Candida parapsilosis, an Emerging Fungal Pathogen

Candida parapsilosis is an emerging major human pathogen that has dramatically increased in significance and prevalence over the past 2 decades, such that C. parapsilosis is now one of the leading causes of invasive candidal disease. Individuals at the highest risk for severe infection include neonates and patients in intensive care units. C. parapsilosis infections are especially associated with hyperalimentation solutions, prosthetic devices, and indwelling catheters, as well as the nosocomial spread of disease through the hands of health care workers. Factors involved in disease pathogenesis include the secretion of hydrolytic enzymes, adhesion to prosthetics, and biofilm formation. New molecular genetic tools are providing additional and much-needed information regarding C. parapsilosis virulence. The emerging information will provide a deeper understanding of C. parapsilosis pathogenesis and facilitate the development of new therapeutic approaches for treating C. parapsilosis infections.

Related papers:

title: Epidemiology of Invasive Candidiasis: a Persistent Public Health Problem
abstract: Invasive candidiasis (IC) is a leading cause of mycosis-associated mortality in the United States. We examined data from the National Center for Health Statistics and reviewed recent literature in order to update the epidemiology of IC. IC-associated mortality has remained stable, at approximately 0.4 deaths per 100,000 population, since 1997, while mortality associated with invasive aspergillosis has continued to decline. Candida albicans remains the predominant cause of IC, accounting for over half of all cases, but Candida glabrata has emerged as the second most common cause of IC in the United States, and several less common Candida species may be emerging, some of which can exhibit resistance to triazoles and/or amphotericin B. Crude and attributable rates of mortality due to IC remain unacceptably high and unchanged for the past 2 decades. Nonpharmacologic preventive strategies should be emphasized, including hand hygiene; appropriate use, placement, and care of central venous catheters; and prudent use of antimicrobial therapy. Given that delays in appropriate antifungal therapy are associated with increased mortality, improved use of early empirical, preemptive, and prophylactic therapies should also help reduce IC-associated mortality. Several studies have now identified important variables that can be used to predict risk of IC and to help guide preventive strategies such as antifungal prophylaxis and early empirical therapy. However, improved non-culture-based diagnostics are needed to expand the potential for preemptive (or early directed) therapy. Further research to improve diagnostic, preventive, and therapeutic strategies is necessary to reduce the considerable morbidity and mortality associated with IC.

title: Nosocomial bloodstream infections in US hospitals: analysis of 24,179 cases from a prospective nationwide surveillance study.
abstract: Nosocomial bloodstream infections (BSIs) are important causes of morbidity and mortality in the United States.



Data from a nationwide, concurrent surveillance study (Surveillance and Control of Pathogens of Epidemiological Importance [SCOPE]) were used to examine the secular trends in the epidemiology and microbiology of nosocomial BSIs.



Our study detected 24,179 cases of nosocomial BSI in 49 US hospitals over a 7-year period from March 1995 through September 2002 (60 cases per 10,000 hospital admissions). Eighty-seven percent of BSIs were monomicrobial. Gram-positive organisms caused 65% of these BSIs, gram-negative organisms caused 25%, and fungi caused 9.5%. The crude mortality rate was 27%. The most-common organisms causing BSIs were coagulase-negative staphylococci (CoNS) (31% of isolates), Staphylococcus aureus (20%), enterococci (9%), and Candida species (9%). The mean interval between admission and infection was 13 days for infection with Escherichia coli, 16 days for S. aureus, 22 days for Candida species and Klebsiella species, 23 days for enterococci, and 26 days for Acinetobacter species. CoNS, Pseudomonas species, Enterobacter species, Serratia species, and Acinetobacter species were more likely to cause infections in patients in intensive care units (P<.001). In neutropenic patients, infections with Candida species, enterococci, and viridans group streptococci were significantly more common. The proportion of S. aureus isolates with methicillin resistance increased from 22% in 1995 to 57% in 2001 (P<.001, trend analysis). Vancomycin resistance was seen in 2% of Enterococcus faecalis isolates and in 60% of Enterococcus faecium isolates.



In this study, one of the largest multicenter studies performed to date, we found that the proportion of nosocomial BSIs due to antibiotic-resistant organisms is increasing in US hospitals.

title: Clinical and microbiological aspects of candidemia due to Candida parapsilosis in Brazilian tertiary care hospitals.
abstract: In order to characterize the epidemiology, microbiology and outcome of candidemia due to Candida parapsilosis, we examined a database of 282 episodes of candidemia prospectively collected from four tertiary care hospitals in São Paulo, Brazil between March 2002 and February 2003, and compared the characteristics of patients with candidemia due to C. parapsilosis (n=64) with those caused by Candida albicans (n=107). C. parapsilosis candidemia was associated with neutropenia (p=0.005), tunneled central venous catheter (p=0.005) and cancer chemotherapy (p=0.03). By multivariate analysis, candidemia due to C. parapsilosis was associated with the presence of a tunneled central venous catheter (relative risk 3.71, 95% confidence interval 1.28-10.70). Except for a single isolate of C. parapsilosis that exhibited MIC >1 microg/ml to amphotericin B, no resistance was observed in 166 isolates tested against fluconazole, itraconazole, 5-flucytosine and amphotericin B. The caspofungin MIC values of C. parapsilosis isolates were significantly higher than those exhibited by C. albicans isolates (p<0.001). The overall mortality of patients with candidemia due to C. parapsilosis was significantly lower (45% vs. 62%, p=0.03). The association between C. parapsilosis candidemia and a tunneled central venous catheter supports the idea that the main mode of acquisition of C. parapsilosis is from an external source.

title: Prospective Observational Study of Candidemia in São Paulo, Brazil: Incidence Rate, Epidemiology, and Predictors of Mortality
abstract: Studies conducted in tertiary care hospitals of different European countries and the United States have shown incidence rates of candidemia ranging from 0.17 to 0.76 and 0.28 to 0.96 per 1,000 admissions, respectively. So far, only 1 study has evaluated the incidence rates of candidemia in tertiary care hospitals in Latin American countries.  To evaluate the epidemiology of candidemia in 4 tertiary care hospitals in São Paulo, Brazil.  Multicenter, laboratory-based surveillance of candidemia.  A total of 7,038 episodes of bloodstream infection were identified, and Candida species accounted for 282 cases (4%). The incidence rate of candidemia was 1.66 candidemic episodes per 1,000 hospital admissions. Candida albicans was the most frequently isolated Candida species in all hospitals, but Candida species other than C. albicans accounted for 62% of isolates, including predominantly Candida parapsilosis and Candida tropicalis. Azole resistance was restricted to only 2% of all Candida isolates (1 isolate of Candida glabrata and 4 isolates of Candida rugosa). Candidemia was mostly documented in surgical patients with long durations of hospital stay. The crude mortality rate was 61%, and advanced age and high Acute Physiology and Chronic Health Evaluation II score were both conditions independently associated with risk of death.  We observed in our series a higher incidence rate of candidemia than that reported in European countries and the United States. Advanced age and a high Acute Physiology and Chronic Health Evaluation II score were factors associated with a higher probability of death in candidemic patients. Fluconazole-resistant Candida strains are still a rare finding in our case-based study of candidemia.

title: Epidemiology of Candidemia in Brazil: a Nationwide Sentinel Surveillance of Candidemia in Eleven Medical Centers
abstract: Candidemia studies have documented geographic differences in rates and epidemiology, underscoring the need for surveillance to monitor trends. We conducted prospective candidemia surveillance in Brazil to assess the incidence, species distribution, frequency of antifungal resistance, and risk factors for fluconazole-resistant Candida species. Prospective laboratory-based surveillance was conducted from March 2003 to December 2004 in 11 medical centers located in 9 major Brazilian cities. A case of candidemia was defined as the isolation of Candida spp. from a blood culture. Incidence rates were calculated per 1,000 admissions and 1,000 patient-days. Antifungal susceptibility tests were performed by using the broth microdilution assay, according to the Clinical and Laboratory Standards Institute guidelines. We detected 712 cases, for an overall incidence of 2.49 cases per 1,000 admissions and 0.37 cases per 1,000 patient-days. The 30-day crude mortality was 54%. C. albicans was the most common species (40.9%), followed by C. tropicalis (20.9%) and C. parapsilosis (20.5%). Overall, decreased susceptibility to fluconazole occurred in 33 (5%) of incident isolates, 6 (1%) of which were resistant. There was a linear correlation between fluconazole and voriconazole MICs (r = 0.54 and P < 0.001 [Spearman's rho]). This is the largest multicenter candidemia study conducted in Latin America and shows the substantial morbidity and mortality of candidemia in Brazil. Antifungal resistance was rare, but correlation between fluconazole and voriconazole MICs suggests cross-resistance may occur.

title: A first Portuguese epidemiological survey of fungaemia in a university hospital
abstract: A prospective, observational study was conducted at the biggest Portuguese hospital, aiming to evaluate the epidemiology of bloodstream fungal infection. During a period of 12 months (2004), all yeasts isolated from the blood cultures of patients with fungaemia admitted at a university hospital of Porto were collected. Demographic and clinical data, as well as haematological and biochemical profiles, were registered. Antifungal susceptibility was evaluated. The incidence of fungaemia and nosocomial fungaemia were 2.7 and 2 per 1,000 hospital admissions, respectively. Blood strains from 117 patients were identified. Thirty-five percent of yeast isolates were Candida albicans, followed by C. parapsilosis (25.6%). The mortality rate associated with fungaemia was 39.3%; the highest values were found in patients with C. glabrata and C. tropicalis infection. Seventy-five percent of the fungaemia episodes were nosocomial, with 48% mortality; the main predisposing factors were parenteral nutrition, gastric protection with omeprazole, surgical drainage and the presence of central venous catheters (CVCs). Thrombocytopaenia, urinary catheter, gastrointestinal pathology and nosocomial fungaemia were independently associated with a poor outcome. Antifungal susceptibility testing showed high fluconazole resistance (15%), mostly in C. tropicalis. We observed a high incidence of nosocomial fungaemia with high mortality rates. Important predisposing factors were identified, deserving further investigation. Local surveillance is warranted to monitor the incidence of in vitro antifungal resistance.

title: Changing Incidence of Candida Bloodstream Infections Among NICU Patients in the United States: 1995–2004
abstract: Recent reports suggest that candidemia caused by fluconazole-resistant strains is increasing in certain adult populations. We evaluated the annual incidence of neonatal candidemia and the frequency of disease caused by different species of Candida among neonates in the United States.  The study included neonates admitted to 128 NICUs participating in the National Nosocomial Infections Surveillance system from January 1, 1995, to December 31, 2004 (study period).  Reports of bloodstream infection (BSI) with Candida spp.; Candida BSIs, patient admissions, patient-days, and central venous catheter days were pooled by birth weight category. The number of Candida BSIs per 100 patients (attack rate) and per 1000 patient-days (incidence density) was determined. Both overall and species-specific rates were calculated; data were pooled over time to determine the differences by birth weight category and by year to determine trends over time.  From the 130523 patients admitted to NICUs during the study period, there were 1997 Candida spp. BSIs reported. Overall, 1472 occurred in the <1000-g birth weight group. Candida albicans BSIs were most common, followed by Candida parapsilosis, Candida tropicalis, Candida lusitaniae, Candida glabrata, and only 3 Candida krusei. Among neonates <1000 g, incidence per 1000 patient-days decreased from 3.51 during 1995–1999 to 2.68 during 2000–2004 but remained stable among heavier neonates. No increase in infections by species that tend to demonstrate resistance to fluconazole (C glabrata or C krusei) was observed.  Although Candida BSI is a serous problem among neonates <1000 g, incidence has declined over the past decade, and disease with species commonly resistant to azoles was extremely rare.

title: Fungemia in Neonates: Report of 80 Cases From Seven University Hospitals
abstract: Candidemia is an emerging infection in neonatology, due to multiple risk factors for fungal infections in neonates, eg, central venous catheter (CVC) insertion, total parenteral nutrition (TPN), corticosteroid and antibiotic therapy, very low birth weight (VLBW) and arteriovenous support (AV).1–3 

Within 10 years, from 1989 to 1998, a prospective nation-wide survey of fungemia in 7 university children's clinics in Slovakia was performed. A total of 310 fungemias developed; 145 appeared in children (48.5%) and 80 (55.3% of all children and 23.3% of all cases) appeared in neonates. The most common risk factors (Table 1) for fungemia in neonates were prior antibiotic therapy, CVC insertion, and TPN, which appeared in …

title: International Surveillance of Candida spp. and Aspergillus spp.: Report from the SENTRY Antimicrobial Surveillance Program (2003)
abstract: 2003, a total of 1,397 Candida isolates, 73 Aspergillus isolates, 53 Cryptococcus neoformans isolates, and 25 other fungal isolates from infected, normally sterile, body sites in patients hospitalized in North America, Europe, and Latin America were studied as a component of the longitudinal SENTRY Antimicrobial Surveillance Program. The MICs for seven antifungal agents were determined in a central laboratory (JMI Laboratories, North Liberty, IA) using testing methods promulgated by the Clinical and Laboratory Standards Institute (formerly the National Committee for Clinical Laboratory Standards). The rank order of Candida spp. occurrence was as follows: C. albicans (48.7%), C. parapsilosis (17.3%), C. glabrata (17.2%), C. tropicalis (10.9%), C. krusei (1.9%), and other Candida spp. (4.0%). C. albicans accounted for 51.5, 47.8, and 36.5% of candidal infections in North America, Europe, and Latin America, respectively. Ravuconazole, voriconazole, and fluconazole were highly active against C. albicans, C. parapsilosis, and C. tropicalis, with both former agents being more potent (MIC at which 90% of the isolates tested are inhibited [MIC90] of ≤0.008 to 0.12 μg/ml) than fluconazole (MIC90 of 0.5 to 2 μg/ml). C. glabrata isolates were less susceptible to these agents, with MIC90s of 1, 1, and 64 μg/ml, respectively. Ravuconazole and voriconazole were the most active agents tested against C. krusei (MIC90 of 0.5 μg/ml). Among Aspergillus spp., A. fumigatus was the most commonly (71.2% of isolates) recovered species; 96.2, 96.2, 84.6, and 11.5% of strains were inhibited by ≤1 μg/ml of ravuconazole, voriconazole, itraconazole, and amphotericin B, respectively. Of the antifungal agents tested, ravuconazole and voriconazole displayed the greatest spectrum of activity against pathogenic Candida and Aspergillus spp., regardless of geographic origin. These results extend upon previous findings from SENTRY Program reports (1997 to 2000), further characterizing species composition as seen in local clinical practice and demonstrating the potent activity of selected, newer triazole antifungal agents.

title: International Surveillance of Bloodstream Infections Due toCandida Species: Frequency of Occurrence and In Vitro Susceptibilities to Fluconazole, Ravuconazole, and Voriconazole of Isolates Collected from 1997 through 1999 in the SENTRY Antimicrobial Surveillance Program
abstract: A surveillance program (SENTRY) of bloodstream infections (BSI) in the United States, Canada, Latin America, and Europe from 1997 through 1999 detected 1,184 episodes of candidemia in 71 medical centers (32 in the United States, 23 in Europe, 9 in Latin America, and 7 in Canada). Overall, 55% of the yeast BSIs were due to Candida albicans, followed by Candida glabrata andCandida parapsilosis (15%), Candida tropicalis(9%), and miscellaneous Candida spp. (6%). In the United States, 45% of candidemias were due to non-C. albicansspecies. C. glabrata (21%) was the most common non-C. albicans species in the United States, and the proportion of non-C. albicans BSIs was highest in Latin America (55%). C. albicans accounted for 60% of BSI in Canada and 58% in Europe. C. parapsilosiswas the most common non-C. albicans species in Latin America (25%), Canada (16%), and Europe (17%). Isolates ofC. albicans, C. parapsilosis, and C. tropicaliswere all highly susceptible to fluconazole (97 to 100% at ≤8 μg/ml). Likewise, 97 to 100% of these species were inhibited by ≤1 μg/ml of ravuconazole (concentration at which 50% were inhibited [MIC50], 0.007 to 0.03 μg/ml) or voriconazole (MIC50, 0.007 to 0.06 μg/ml). Both ravuconazole and voriconazole were significantly more active than fluconazole against C. glabrata(MIC90s of 0.5 to 1.0 μg/ml versus 16 to 32 μg/ml, respectively). A trend of increased susceptibility of C. glabrata to fluconazole was noted over the three-year period. The percentage of C. glabrataisolates susceptible to fluconazole increased from 48% in 1997 to 84% in 1999, and MIC50s decreased from 16 to 4 μg/ml. A similar trend was documented in both the Americas (57 to 84% susceptible) and Europe (22 to 80% susceptible). Some geographic differences in susceptibility to triazole were observed with Canadian isolates generally more susceptible than isolates from the United States and Europe. These observations suggest susceptibility patterns and trends among yeast isolates from BSI and raise additional questions that can be answered only by continued surveillance and clinical investigations of the type reported here (SENTRY Program).

title: International Surveillance of Bloodstream Infections Due to Candida Species: Frequency of Occurrence and Antifungal Susceptibilities of Isolates Collected in 1997 in the United States, Canada, and South America for the SENTRY Program
abstract: An international program of surveillance of bloodstream infections (BSIs) in the United States, Canada, and South America between January and December 1997 detected 306 episodes of candidemia in 34 medical centers (22 in the United States, 6 in Canada, and 6 in South America). Eighty percent of the BSIs were nosocomial and 50% occurred in patients hospitalized in an intensive care unit. Overall, 53.3% of the BSIs were due to Candida albicans, 15.7% were due toC. parapsilosis, 15.0% were due to C. glabrata, 7.8% were due to C. tropicalis, 2.0% were due to C. krusei, 0.7% were due to C. guilliermondii, and 5.8% were due to Candida spp. However, the distribution of species varied markedly by country. In the United States, 43.8% of BSIs were due to non-C. albicansspecies. C. glabrata was the most common non-C. albicans species in the United States. The proportion of non-C. albicans BSIs was slightly higher in Canada (47.5%), where C. parapsilosis, not C. glabrata, was the most common non-C. albicansspecies. C. albicans accounted for 40.5% of all BSIs in South America, followed by C. parapsilosis (38.1%) andC. tropicalis (11.9%). Only one BSI due to C. glabrata was observed in South American hospitals. Among the different species of Candida, resistance to fluconazole (MIC, ≥64 μg/ml) and itraconazole (MIC, ≥1.0 μg/ml) was observed with C. glabrata and C. krusei and was observed more rarely among other species. Isolates of C. albicans,C. parapsilosis, C. tropicalis, and C. guilliermondii were all highly susceptible to both fluconazole (99.4 to 100% susceptibility) and itraconazole (95.8 to 100% susceptibility). In contrast, 8.7% of C. glabrata isolates (MIC at which 90% of isolates are inhibited [MIC90], 32 μg/ml) and 100% of C. krusei isolates were resistant to fluconazole, and 36.9% of C. glabrata isolates (MIC90, 2.0 μg/ml) and 66.6% of C. kruseiisolates were resistant to itraconazole. Within each species there were no geographic differences in susceptibility to fluconazole or itraconazole.

title: [Multicenter study of fungemia due to yeasts in Argentina].
abstract: The incidence of candidemia has increased approximately 500% in high-complexity hospitals. A change in the spectrum of Candida infections due to species other than Candida albicans has also been detected. Between April 1999 and April 2000 a multicenter study was performed in order to determine the species distribution associated to candidemias in Argentina and the susceptibility profile of the isolates to the current antifungal drugs. Thirty six institutions have participated. All the 265 yeast strains isolated from blood cultures were identified by morphological, physiological, and biochemical tests. The antifungal susceptibility testing of isolates was performed based on the reference NCCLS procedure. The distribution of species was: Candida albicans (40.75%), Candida parapsilosis (28.67%), Candida tropicalis (15.84%), Candida famata (3.77%), Cryptococcus neoformans (3.77%), Candida glabrata (2.64%), and others (4.53%). Most of the isolates were susceptible to amphotericin B, fluconazole and itraconazole. Mortality associated to the fungemia by yeasts episodes (n=265) was 30%, lower than results previously determined (33-54%). The mortality percentage in patients who received antifungal therapy versus patients without treatment was 26.3% and 47%, respectively.

title: Epidemiological study of Candida infections in blood: susceptibilities of Candida spp. to antifungal agents, and clinical features associated with the candidemia
abstract: The purpose of this study was to evaluate the susceptibility to antifungal agents of Candida spp. isolated from blood samples from patients in our hospital, located in Osaka, Japan. We also examined the clinical background of these patients. We analyzed fungi isolated from clinical blood samples obtained in our hospital over a period of 10 years (1993 to 2002). Antifungal susceptibility testing was carried out for six agents, using the National Committee of Clinical Laboratory Standards (NCCLS) M-27-A2 method. The clinical backgrounds were reviewed using the medical records of 125 patients who were diagnosed as having candidemia. The major fungi isolated were Candida parapsilosis (39.2%) and C. albicans (30.1%), and both were sensitive to fluconazole. One strain of C. glabrata and six strains of C. krusei were resistant to fluconazole, and they constituted 4.4% of all Candida spp. isolated. With the exception of C. parapsilosis, most fungi were susceptible to micafungin, although there is no universally agreed breakpoint for this drug. Analysis of the patients' clinical backgrounds revealed that the major underlying disease was cancer (46.4% excluding hematological malignancies). C. krusei was detected almost exclusively in patients with hematological malignancies. Indwelling venous catheters had been responsible for infection in 93.6% of the infected patients. The clinical outcomes of the 125 patients were favorable in 52% and poor in 48%, and subsequent removal of the indwelling catheters was effective in about half of the patients in whom this was done, with good prognosis. To prevent mycosis and its complications, indwelling catheters should be avoided as much as possible. Attention must be paid to the possibility that resistant isolates of Candida spp. can be selected as a result of the use of antifungal agents.

title: Systemic Candida infection in University Hospital 1997–1999: the distribution of Candida biotypes and antifungal susceptibility patterns
abstract: A total of 102 Candida species were isolated from blood cultures from January 1997 to October 1999. Using assimilation of carbohydrate test, 52 (51.0%) of the Candida sp. were identified as C. parapsilosis, 25.5% (26) were C. tropicalis. C. albicans made up 11.8% (12), 6.9% (7) were C. rugosa, 3.8% (4) C. glabrata and 1% (1) C. guilliermondii. No C. dubliniensis was found in the study. In vitro antifungal susceptibility tests showed that all Candida species were sensitive to nystatin, amphotericin B and ketoconazole. Although all isolates remained sensitive to fluconazole, intermediate susceptibility was found in 3 C. rugosa isolates. Antifungal agents with high frequency of resistance were econazole, clotrimazole, miconazole and 5-fluorocytosine. Candida species found to have resistance to these antifungal agents were non-C. albicans.

title: Candida parapsilosis: epidemiology, pathogenicity, clinical manifestations, and antimicrobial susceptibility.
abstract: Early reports associated Candida parapsilosis with endocarditis in intravenous narcotic addicts. More recently, this species has emerged as an important nosocomial pathogen, with clinical manifestations including fungemia, endocarditis, endophthalmitis, septic arthritis, and peritonitis, all of which usually occur in association with invasive procedures or prosthetic devices. Outbreaks of C. parapsilosis infections have been caused by contamination of hyperalimentation solutions, intravascular pressure monitoring devices, and ophthalmic irrigating solution. Experimental studies have generally shown that C. parapsilosis is less virulent than Candida albicans or Candida tropicalis. However, characteristics of C. parapsilosis that may relate to its increasing occurrence in nosocomial settings include frequent colonization of the skin, particularly the subungual space, and an ability to proliferate in glucose-containing solutions, with a resultant increase in adherence to synthetic materials. Recently developed molecular techniques may facilitate the continued exploration of the epidemiology and pathogenesis of C. parapsilosis infections.

title: SUBACUTE ENDOCARDITIS AND SYSTEMIC MYCOSIS (MONILIA)
abstract: As far as could be ascertained, no case of subacute bacterial endocarditis with systemic mycosis due to Monilia recognized during life has ever been reported in the literature. Dr. Emanuel Libman, who saw this case with us, referred us to a recent report by Friedman and Donaldson. 1 In their case nests of yeastlike cells were isolated from the aortic vegetations on postmortem examination of a patient who died of subacute bacterial endocarditis. However, the authors were unable to isolate the organism clinically or identify it on postmortem examination. Because of the rarity of this condition and the circumstances under which the infection occurred in our case, it was considered worthy of publication. REPORT OF CASE History.— F. M., a white man aged 48, American born, married, a bookkeeper, was admitted to the surgical service of the Cumberland Hospital on April 24, 1939, because of midabdominal pain of two

title: Candida orthopsilosis and Candida metapsilosis spp. nov. To Replace Candida parapsilosis Groups II and III
abstract: Two new species, Candida orthopsilosis and C. metapsilosis, are proposed to replace the existing designations of C. parapsilosis groups II and III, respectively. The species C. parapsilosis is retained for group I isolates. Attempts to construct a multilocus sequence typing scheme to differentiate individual strains of C. parapsilosis instead revealed fixed DNA sequence differences between pairs of subgroups in four genes: COX3, L1A1, SADH, and SYA1. PCR amplicons for sequencing were obtained for these four plus a further seven genes from 21 group I isolates. For nine group II isolates, PCR products were obtained from only 5 of the 11 genes, and for two group III isolates PCR products were obtained from a different set of 5 genes. Three of the PCR products from group II and III isolates differed in size from the group I products. Cluster analysis of sequence polymorphisms from COX3, SADH, and SYA1, which were common to the three groups, consistently separated the isolates into three distinct sets. All of these differences, together with DNA sequence similarities <90% in the ITS1 sequence, suggest the subgroups should be afforded species status. The near absence of DNA sequence variability among isolates of C. parapsilosis and relatively high levels of sequence variability among isolates of C. orthopsilosis suggest that the former species may have evolved very recently from the latter.

title: Phenotype switching affects biofilm formation by Candida parapsilosis.
abstract: Generation of biofilms by the pathogenic yeast Candida parapsilosis is correlated closely with disease. The phenomenon of phenotype switching in 20 isolates of C. parapsilosis was examined and the relationship with biofilm development was investigated. Four stable and heritable phenotypes were identified--crepe, concentric, smooth and crater. Cells from crepe and concentric phenotypes are almost entirely pseudohyphal, whilst cells from smooth and crater phenotypes are mostly yeast-like. The pseudohyphae from concentric phenotypes are approximately 45 % wider than those from crepe cells. The cell size of the smooth phenotype is smaller than those of the other three phenotypes. On polystyrene surfaces, the concentric phenotype generates up to twofold more biofilm than the crepe and crater phenotypes. Smooth phenotypes generate the least biofilm. Concentric phenotypes also invade agar surfaces more than the crepe and crater phenotypes, whilst smooth phenotypes do not invade at all. The smooth phenotype, however, grows significantly faster than the others. The quorum-sensing molecule farnesol inhibits formation of biofilms by the crepe, concentric and crater phenotypes.

title: Amino acids mediate colony and cell differentiation in the fungal pathogen Candida parapsilosis.
abstract: Candida parapsilosis is responsible for severe cases of non-albicans systemic candidiasis and is one of the leading causes of mortality in neonates. The molecular mechanisms underlying this organism's virulence remain unknown. Unlike C. albicans, which can exist in several morphogenetic forms, C. parapsilosis exists in either the yeast or pseudohyphal forms. The environmental signals that trigger pseudohyphal differentiation and the signalling pathways that transduce these signals are unknown. This paper provides evidence for the role of amino acids in morphogenesis in C. parapsilosis. The cell and colony morphologies, pseudohyphal differentiation and invasive growth of five C. parapsilosis isolates were characterized in ammonium-rich minimal media lacking or supplemented with naturally occurring amino acids. C. parapsilosis underwent dramatic changes in cellular and colony morphology and formed pseudohyphae in response to a specific subset of amino acids. Transport studies showed that these amino acid inducers activate the transport of some, but not all, unrelated amino acids. Interestingly, citrulline, an amino acid that is not transported in the presence of ammonium, strongly induced pseudohyphal morphogenesis in C. parapsilosis under these conditions. Together the data suggest that amino acids are important morphogens in C. parapsilosis and that amino-acid-mediated morphogenesis in this organism does not require transport of the ligand across the plasma membrane.

title: Epidemiologic and Molecular Characterization of an Outbreak of Candida parapsilosis Bloodstream Infections in a Community Hospital
abstract: Candida parapsilosis is an important cause of bloodstream infections in the health care setting. We investigated a large C. parapsilosis outbreak occurring in a community hospital and conducted a case-control study to determine the risk factors for infection. We identified 22 cases of bloodstream infection with C. parapsilosis: 15 confirmed and 7 possible. The factors associated with an increased risk of infection included hospitalization in the intensive care unit (adjusted odds ratio, 16.4; 95% confidence interval, 1.8 to 148.1) and receipt of total parenteral nutrition (adjusted odds ratio, 9.2; 95% confidence interval, 0.9 to 98.1). Samples for surveillance cultures were obtained from health care worker hands, central venous catheter insertion sites, and medical devices. Twenty-six percent of the health care workers surveyed demonstrated hand colonization with C. parapsilosis, and one hand isolate was highly related to all case-patient isolates by tests with the DNA probe Cp3-13. Outbreak strain isolates also demonstrated reduced susceptibilities to fluconazole and voriconazole. This largest known reported outbreak of C. parapsilosis bloodstream infections in adults resulted from an interplay of host, environment, and pathogen factors. Recommendations for control measures focused on improving hand hygiene compliance.

title: Candida parapsilosis infection in very low birthweight infants
abstract: In a UK national surveillance study, we found that Candida parapsilosis accounted for one quarter of all cases of invasive fungal infection in very low birthweight infants. C parapsilosis was associated with fewer deep-seated infections than C albicans, but mortality was similar. Ongoing surveillance is needed to monitor the epidemiology of invasive fungal infection in very low birthweight infants.

title: Mortality Following Blood Culture in Premature Infants: Increased with Gram-negative Bacteremia and Candidemia, but Not Gram-positive Bacteremia
abstract: To describe survival following nosocomial bloodstream infections and quantify excess mortality associated with positive blood culture. Multicenter cohort study of premature infants. First blood culture was negative for 4648/5497 (78%) of the neonates — 390/4648 (8%) died prior to discharge. Mortality prior to discharge was 19% in the 161 infants with Gram-negative rod (GNR) bacteremia, 8% in the 854 neonates with coagulase negative staphylococcus (CONS), 6% in the 169 infants infected with other Gram-positive bacteria (GP-o), and 26% in the 115 neonates with candidemia. The excess 7-day mortality was 0% for Gram-positive organisms and 83% for GNR bacteremia and candidemia. Using negative blood culture as referent, GNR [hazard ratio (HR)=2.61] and candidemia (HR=2.27) were associated with increased mortality; CONS (HR=1.08) and GP-o (HR=0.97) were not. Nosocomial GNR bacteremia and candidemia were associated with increased mortality but Gram-positive bacteremia was not.

title: Systematics of yeast species in the Candida parapsilosis group
abstract: The systematics ofCandida species that may or may not ferment carbohydrates and which assimilate trehalose, xylose, and mannitol, but do not assimilate lactose, melibiose, raffinose, inositol, dulcitol, or KNO3 were examined. Included were:C. albicans, C. albicans var.stellatoidea, C. diddensii, C. bogoriensis, C. claussenii, C. lusilaniae, C. natalensis, C. obtusa, C. oregonensis, C. parapsilosis, C. parapsilosis var.querci, C. polymorpha, C. pulcherrima. C. reukaufii, C. solani, C. tropicalis, C. tropicalis var.lambica andC. viswanathii. The species were delimited and a basis for separation established.

title: Candidemia before and during the fluconazole era: prevalence, type of species and approach to treatment in a tertiary care community hospital.
abstract: The characteristics of candidemia before and after the introduction of fluconazole were compared at our institution. In the pre-fluconazole era (1986-89), the incidence of candidemia rose from 0.02% to 13% over the course of 4 y. Candida albicans accounted for 44/66 isolates (67%) and C. glabrata and C. parapsilosis were the predominant non-albicans species (9% each). In one-third of cases antifungal therapy was not given and the overall mortality rate was 55%. During the fluconazole era (1994-97), the incidence of candidemia remained constant between 1994 and 1996 at 0.09-0.11% and dropped to 0.06% in 1997. Bloodstream infection associated with neutropenia increased significantly but only accounted for 13% of cases; antifungal prophylaxis was not employed. The isolation of C. albicans decreased (n = 49; 50%) whereas isolation of both C. parapsilosis (n = 23; 24%) and C. tropicalis (n = 16; 16%) increased. The vast majority of patients received antifungal therapy and the overall mortality rate was 39%. These findings show that the incidence of candidemia rose steadily prior to the fluconazole era and then stabilized in spite of a shift towards non-albicans species. When candidemia was encountered in the fluconazole era, it was rare not to give antifungal treatment.

title: Candidemia in a Tertiary Care Hospital: Epidemiology and Factors Influencing Mortality
abstract: The present study was conducted in order to assess the epidemiology and clinical course of candidemia and to identify the risk factors associated with mortality. A total of 143 episodes of nosocomial candidemia were identified during a 5-year period, and these were included in the study. The majority of candidemic episodes were due to Candida albicans (63, 44%), followed by Candida parapsilosis (32, 22%). The overall mortality was 45%. The following independent prognostic factors for mortality were identified: bacterial sepsis, rapidly fatal illness, chronic obstructive lung disease, presence of a central venous catheter, candidemia due to Candida albicans, and lack of antifungal therapy.

title: Results from the ARTEMIS DISK Global Antifungal Surveillance Study, 1997 to 2005: an 8.5-Year Analysis of Susceptibilities of Candida Species and Other Yeast Species to Fluconazole and Voriconazole Determined by CLSI Standardized Disk Diffusion Testing
abstract: Fluconazole in vitro susceptibility test results for 205,329 yeasts were collected from 134 study sites in 40 countries from June 1997 through December 2005. Data were collected for 147,776 yeast isolates tested with voriconazole from 2001 through 2005. All investigators tested clinical yeast isolates by the CLSI M44-A disk diffusion method. Test plates were automatically read and results recorded with a BIOMIC image analysis system. Species, drug, zone diameter, susceptibility category, and quality control results were collected quarterly. Duplicate (same patient, same species, and same susceptible-resistant biotype profile during any 7-day period) and uncontrolled test results were not analyzed. Overall, 90.1% of all Candida isolates tested were susceptible (S) to fluconazole; however, 10 of the 22 species identified exhibited decreased susceptibility (<75% S) on the order of that seen with the resistant (R) species C. glabrata and C. krusei. Among 137,487 isolates of Candida spp. tested against voriconazole, 94.8% were S and 3.1% were R. Less than 30% of fluconazole-resistant isolates of C. albicans, C. glabrata, C. tropicalis, and C. rugosa remained S to voriconazole. The non-Candida yeasts (8,821 isolates) were generally less susceptible to fluconazole than Candida spp. but, aside from Rhodotorula spp., remained susceptible to voriconazole. This survey demonstrates the broad spectrum of these azoles against the most common opportunistic yeast pathogens but identifies several less common yeast species with decreased susceptibility to antifungal agents. These organisms may pose a future threat to optimal antifungal therapy and emphasize the importance of prompt and accurate species identification.

title: In Vitro Susceptibility of Invasive Isolates of Candida spp. to Anidulafungin, Caspofungin, and Micafungin: Six Years of Global Surveillance
abstract: The echinocandins are being used increasingly as therapy for invasive candidiasis. Prospective sentinel surveillance for the emergence of in vitro resistance to the echinocandins among invasive Candida sp. isolates is indicated. We determined the in vitro activities of anidulafungin, caspofungin, and micafungin against 5,346 invasive (bloodstream or sterile-site) isolates of Candida spp. collected from over 90 medical centers worldwide from 1 January 2001 to 31 December 2006. We performed susceptibility testing according to the CLSI M27-A2 method and used RPMI 1640 broth, 24-h incubation, and a prominent inhibition endpoint for determination of the MICs. Of 5,346 invasive Candida sp. isolates, species distribution was 54% C. albicans, 14% C. parapsilosis, 14% C. glabrata, 12% C. tropicalis, 3% C. krusei, 1% C. guilliermondii, and 2% other Candida spp. Overall, all three echinocandins were very active against Candida: anidulafungin (MIC50, 0.06 μg/ml; MIC90, 2 μg/ml), caspofungin (MIC50, 0.03 μg/ml; MIC90, 0.25 μg/ml), micafungin (MIC50, 0.015 μg/ml; MIC90, 1 μg/ml). More than 99% of isolates were inhibited by ≤2 μg/ml of all three agents. Results by species (expressed as the percentages of isolates inhibited by ≤2 μg/ml of anidulafungin, caspofungin, and micafungin, respectively) were as follows: for C. albicans, 99.6%, 100%, and 100%; for C. parapsilosis, 92.5%, 99.9%, and 100%; for C. glabrata, 99.9%, 99.9%, and 100%; for C. tropicalis, 100%, 99.8%, and 100%; for C. krusei, 100%, 100%, and 100%; and for C. guilliermondii, 90.2%, 95.1%, and 100%. There was no significant change in the activities of the three echinocandins over the 6-year study period and no difference in activity by geographic region. All three echinocandins have excellent in vitro activities against invasive strains of Candida isolated from centers worldwide. Our prospective sentinel surveillance reveals no evidence of emerging echinocandin resistance among invasive clinical isolates of Candida spp.

title: Comparison of adherence of Candida albicans and Candida parapsilosis to silicone catheters in vitro and in vivo.
abstract: Although Candida parapsilosis has been associated with device-related infections in the clinical settings, factors that contribute to this association have not been previously examined. The objectives of this study were to compare in vitro and in vivo the adherence to silicone catheters of: (1) Candida albicans vs. C. parapsilosis, and (2) invasive vs. colonizing isolates of C. albicans and C. parapsilosis.



The records of 840 patients who had had Candida species isolated at three teaching hospitals during a three-month period were reviewed. A total of 20 clinical isolates of each of C. parapsilosis and C. albicans were examined for their adherence to silicone catheters in vitro and in a rabbit model of percutaneously placed catheters. For each Candida species, ten invasive isolates that had caused clinical device-related infection and 10 colonizing isolates that had caused only device colonization were studied.



Candida parapsilosis accounted for <5% of yeast isolates from all sites, while three-quarters were C. albicans. Candida parapsilosis was isolated proportionately more often from blood and/or devices than C. albicans (34.3% vs. 8.5%, respectively, P < 0.0001). There were no significant differences in the degrees of adherence in vitro and in vivo between C. albicans and C. parapsilosis or between invasive and colonizing Candida.



Although C. parapsilosis was isolated proportionately more often from blood and/or devices than C. albicans in our studied population, there was no significant difference in the adherence of the two Candida species to silicone, nor between invasive and colonizing Candida in our in vitro and in vivo models. Factors other than microbial adherence may help explain the observed association of C. parapsilosis with device-related infections.

title: Fungemia in Patients with Leukemia
abstract: A nine-year retrospective study on fungemia in patients with leukemia was conducted. A total of 79 episodes of fungemia in 77 patients with leukemia were documented. Candida parapsilosis fungemia was associated more frequently with the presence of a central venous line and to the use of parenteral nutrition than the other fungal species (p = 0.00026 and p = 0.01, respectively). The same fungus was isolated from both blood and surveillance cultures in 95% of Candida albicans and in 89% of Candida tropicalis fungemia (p < 0.01 and p = 0.02, respectively). The neutropenia and fungus colonization that resulted was associated significantly with the presence of invasive disease (p = 0.0024 and p = 0.0028, respectively). Conversely, central venous catheterization and parenteral nutrition appeared to be associated with episodes without deep tissue invasion (p = 0.000037 and p = 0.001, respectively). Invasive mycosis due to the fungus isolated from blood was documented in 51 patients with a mortality rate of 6970, whereas in 20 patients without invasive mycosis, mortality rate was 21% (p = 0.000059). In patients with fungemia, related or unrelated to the presence of a central venous catheter, mortality was 24% and 6470, respectively (p = 0.00042). Mortality was highest with C. tropicalis (p = 0.0017) and lowest with C. parapsilosis (p = 0.057). Severe neutropenia (polymorpho-nuclears < 100/mmc) appeared associated with a higher mortality rate (p = 0.012), whereas the recovery of neutropenia was related adversely to a fatal outcome (p < 0.01). With antifungal therapy, there was no statistically significant difference whether antifungal therapy was given or not. This study confirms the major roles of neutropenia, previous colonization, and the central venous catheter as predisposing factors in the development of fungemia in patients with leukemia. The prognosis of patients with fungemia depends on the presence of a deep infection, on the recovery of neutropenia, and on the species of fungus involved.

title: High-dose fluconazole therapy for cancer patients with solid tumors and candidemia: an observational, noncomparative retrospective study
abstract: BackgroundResponse rates for candidemia treated with standard-dose fluconazole (400 mg/day) are approximately 70%. Higher doses of fluconazole have been recommended for susceptible dose-dependent Candida isolates. Herein, we describe the outcome of 20 patients with solid tumors and candidemia treated with high-dose fluconazole (HDF) at The University of Texas M.D. Anderson Cancer Center (1998–2002).Patients and methodsPatients were identified either by searching the microbiology laboratory database or through direct referral from primary oncology services to the Infectious Diseases Consultative Services. A retrospective review of cases was performed. HDF was defined as ≥600 mg/day.ResultsFive patients were treated with 600 mg/day, whereas 15 patients received 800 mg/day. Only one patient was neutropenic. The median APACHE II score at the onset of candidemia was 12 (range 6–24). The most common species identified were Candida albicans (eight patients, 40%) and Candida parapsilosis (seven patients, 35%). Of 19 patients whose quantitative data were available, eight (42%) had high-grade candidemia [≥200 colony forming units (CFU)/ml]. Fifteen (83%) of 18 isolates were fluconazole susceptible, and two (both Candida glabrata) were fluconazole resistant (MIC 64 each) in vitro. Nineteen patients (95%) responded to HDF therapy. The only HDF failure occurred in a patient with C. glabrata (MIC 64.0) infection. The other patient with C. glabrata (MIC 64.0) infection responded to HDF. Central venous catheters were removed from all patients with ≥10 CFU/ml candidemias. All patients with high-grade candidemias responded to HDF. The median duration of HDF therapy was 16 (range 6–42) days. No significant toxicity occurred.ConclusionsAlthough our data are limited, HDF appears to be well tolerated and may be associated with higher response rates than standard-dose fluconazole in a selected group of patients with solid tumors and candidemia caused by species that are susceptible to this triazole.

title: Importance of Candida species other than C. albicans as pathogens in oncology patients.
abstract: A number of surveys have documented increased rates of candida infection over the past several decades. In this assessment of the frequency and distribution of non-albicans Candida species among patients with cancer, 37 reports that were published between 1952 and 1992 and that described 1,591 cases of systemic candida infection were reviewed. Species other than Candida albicans accounted for 46% of all systemic candida infections in patients with cancer; specifically, Candida tropicalis accounted for 25%, Candida glabrata for 8%, Candida parapsilosis for 7%, and Candida krusei for 4%. Other species were uncommon. C. tropicalis was the predominant pathogenic Candida species in five reports, C. glabrata in two, C. krusei in two, and Candida stellatoidea in one. The perception that, over time, a greater proportion of candida infections have been caused by non-albicans species was not borne out. The wide variability in reported findings was striking and was due in part to differences in the underlying disease affecting the patients described. For example, patients with leukemia were more likely to be infected by C. albicans or C. tropicalis but less likely to be infected by C. glabrata than patients with other types of cancer. The recent increase in the rate of bone marrow transplantation may also have contributed to discrepancies among reports. Bone marrow transplant recipients were more likely to be infected by C. krusei or C. lusitaniae. The other factors partially responsible for the variability among reports included common-source contamination and the pressures imposed by antimicrobial measures.

title: Death from inappropriate therapy for Lyme disease.
abstract: A 30-year-old woman died as a result of a large Candida parapsilosis septic thrombus located on the tip of a Groshong catheter. The catheter had been in place for 28 months for administration of a 27 month course of intravenous cefotaxime for an unsubstantiated diagnosis of chronic Lyme disease.

title: Rising incidence of Candida parapsilosis fungemia in patients with hematologic malignancies: clinical aspects, predisposing factors, and differential pathogenicity of the causative strains.
abstract: Over the years 1983-1994, Candida parapsilosis causes 35 or 138 fungemic episodes (24 of 69 candidemias in the last quadriennium) in patients with hematologic malignancies who were being treated at a large university hospital in Italy. The central venous catheter was usually the source of bloodstream invasion; in most cases, the resolution of fungemia in patients receiving antifungal therapy required catheter removal. In seven cases, C. parapsilosis fungemia evolved to five proven (two cases with endocarditis) and two probable deep-seated infections; three of these seven patients died of deep-seated infections. Deep seated infection was associated with the detection of a circulating mannoprotein antigen of C. parapsilosis but not with in vitro resistance to antifungal agents. Almost all fungal isolates produced slime in vitro, but only 34% were pathogenic in a model of bloodstream infection in neutropenic mice. The four isolates associated with endocarditis or persistent fungemia with multiorgan failure were among the most virulent in the model of infection. Overall, our findings highlight the role of C. parapsilosis as an agent of fungemia in patients with malignant hemopathies.

title: Candida parapsilosis fungemia associated with implantable and semi-implantable central venous catheters and the hands of healthcare workers.
abstract: A cluster of six cases of fungemia among hematology, bone marrow transplant, and oncology patients was investigated in a case-control study (18 controls). The use of implantable and semi-implantable central venous catheters was significantly associated with cases (p = 0.016). The hands of three healthcare workers (HCWs) were positive for Candida parapsilosis. Electrophoretic karyotyping showed two profiles among patients and HCWs, and five among six unrelated strains. The profiles of two HCWs matched the ones of the patients they had handled. The patients' strains were moderate or strong slime producers, whereas none of the HCWs' were strong producers. In conclusion, our results indicated the occurrence of an outbreak C. parapsilosis fungemia related to long-term central venous catheters in which the hands of HCWs were implicated. The amount of slime production might be associated with the pathogenicity of the strains.

title: Risk factors for candidemia in a children's hospital.
abstract: Candida species are increasingly important nosocomial pathogens in critically ill children. A 2.3-fold increase in the rate of nosocomial candidemia at our 200-bed tertiary care children's hospital prompted a study to identify risk factors for this infection. Twenty-six cases were identified between 1992 and 1993, representing 21% of all nosocomial bloodstream infections. Candida albicans was the most frequent isolate (58%), followed by Candida parapsilosis (27%). A case-control study revealed that there was a statistically significant association between the occurrence of candidemia and placement of a central venous catheter in the femoral vein (P = .03), the use of a tunneled central venous catheter (P = .05), and prolonged hyperalimentation (P = .04). Patients with candidemia also were noted to have candiduria more often than controls (P = .003) and were more likely to have had topical antifungal agents prescribed (P = .04). Multivariate analysis showed that hyperalimentation was an independent risk factor for the development of candidemia. We conclude that measures must be taken to reduce these risk factors whenever possible.

title: An outbreak of Candida parapsilosis bloodstream infections in patients receiving parenteral nutrition.
abstract: From July to September 1981, five patients at one hospital had bloodstream infections or colonization of an intravascular cannula with Candida parapsilosis. All five cases, but none of 34 controls, were receiving parenteral nutrition at the onset of infection or colonization (P less than 0.01; Fisher's exact test, one-tailed). Epidemiologic investigation showed that human serum albumin was more frequently added to parenteral nutrition solutions during the epidemic period than during a comparable period a year earlier. The increase in human serum albumin use coincided with the more frequent use of an electrically powered vacuum pump to assist in compounding parenteral nutrition solutions. Cultures from the vacuum pump showed heavy growth of C parapsilosis from multiple sites. Laboratory investigation demonstrated that sterile solutions could be contaminated by use of the vacuum pump. Use of the vacuum pump was stopped, and no further cases occurred.

title: Nosocomial acquisition of Candida parapsilosis: an epidemiologic study.
abstract: The purpose of this study was to determine aspects of the epidemiology of nosocomial infection due to Candida parapsilosis. Candida species are important nosocomial pathogens; however, little epidemiologic information is available.



We prospectively cultured specimens from 98 patients admitted to the bone marrow transplant unit and a medicine intensive care unit (ICU) of a tertiary care hospital. Specimens from hands of personnel and environmental surfaces were also cultured. Environmental cultures were done before patients were admitted to a studied unit. Restriction enzyme analysis (REA) of chromosomal DNA was used as a typing system to determine the relatedness of strains.



C. parapsilosis was identified from five patients, six hand cultures from four hospital staff, and two environmental surfaces. All five patients had negative initial cultures and acquired C. parapsilosis after admission to the study unit. There were no significant differences between patients and control subjects in age, underlying disease, immunosuppressive therapy, and instrumentation. The duration of antibiotic therapy (median: 32.8 versus 11.8 days, p = 0.05) and the duration in the unit (means: 30.1 versus 16.1 days, p = 0.048) was longer in patients than in controls. No common source was identified. REA revealed three strain types; however, one strain type was identical in four patients, three staff members, and two environmental surfaces.



These results suggest exogenous acquisition of C. parapsilosis. Based upon isolation of identical patient strains of C. parapsilosis from inanimate surfaces before patients were admitted to a study unit, there is evidence that the organism may have been acquired from the hospital environment. The principal mechanism of transmission was probably indirect contact via the hands of hospital personnel.

title: Neonatal Candida parapsilosis outbreak with a high case fatality rate.
abstract: A Candida parapsilosis outbreak of 58 cases in a neonatal intensive care unit lasted for 55 months. Patients infected by or colonized with C. parapsilosis were mainly very low birth weight infants (birth weight < 1500 g). Their mean birth weight was 817 g and their mean gestational age was 28 weeks. Statistical analysis including logistic regression confirmed that prematurity was the main risk factor. The analysis also suggested that C. parapsilosis infection (or colonization) was associated with a poor prognosis. In infants with gestational age < 29 weeks the risk for death in C. parapsilosis-infected patients was 16-fold greater than in those with no C. parapsilosis infection. The case fatality rate of C. parapsilosis patients was higher than that of the controls (9 of 23 vs. 1 of 40; P < 0.0001). The outbreak was most likely a result of cross-infection because C. parapsilosis could be isolated only from the patients and from the hands of four nurses immediately after they had cared for a colonized patient. Cessation of the outbreak was temporally associated with long term parenteral fluconazole (6 mg/kg/day) prophylaxis.

title: Candida parapsilosis fungemia associated with parenteral nutrition and contaminated blood pressure transducers
abstract: During the period September 1983 through May 1985, Candida parapsilosis was isolated from intravascular sites (blood or vascular catheter tips) in 12 patients at a pediatric hospital. Of 205 patients with cultures of any site positive for Candida species, 32 (16%) had cultures positive for C. parapsilosis. In contrast, of 23 patients with intravascular cultures positive for Candida species, 12 (51%) had cultures positive for C. parapsilosis (P less than 0.001, Fisher's exact test). The 12 patients with intravascular cultures positive for C. parapsilosis were more likely to have received central venous nutrition therapy (10 of 12 versus 7 of 23; P less than 0.01, Mantel-Haenzel chi-square test) and had a longer duration of exposure to blood pressure transducers (P less than 0.08, paired t test) than the 23 ward- and age-matched controls. C. parapsilosis was isolated from 11 (32%) of 34 in-use and stored blood pressure transducers. After ethylene oxide sterilization of blood pressure transducers was begun, in-use pressure transducers showed no growth of C. parapsilosis. This study emphasizes the role of C. parapsilosis as a nosocomial pathogen associated with invasive devices and parenteral nutrition; it also emphasizes the importance of adhering to recommended procedures for sterilizing blood pressure transducers.

title: Candida parapsilosis Characterization in an Outbreak Setting
abstract: Candida parapsilosis is an important non-albicans species which infects hospitalized patients. No studies have correlated outbreak infections of C. parapsilosis with multiple virulence factors. We used DNA fingerprinting to determine genetic variability among isolates from a C. parapsilosis outbreak and from our clinical database. We compared phenotypic markers of pathogenesis, including adherence, biofilm formation, and protein secretion (secretory aspartic protease [SAP] and phospholipase). Adherence was measured as colony counts on silicone elastomer disks immersed in agar. Biofilms formed on disks were quantified by dry weight. SAP expression was measured by hydrolysis of bovine albumin; a colorimetric assay was used to quantitate phospholipase. DNA fingerprinting indicated that the outbreak isolates were clonal and genetically distinct from our database. Biofilm expression by the outbreak clone was greater than that of sporadic isolates (p < 0.0005). Adherence and protein secretion did not correlate with strain pathogenicity. These results suggest that biofilm production plays a role in C. parapsilosis outbreaks.

title: When to Suspect Fungal Infection in Neonates: A Clinical Comparison of Candida albicans and Candida parapsilosis Fungemia With Coagulase-Negative Staphylococcal Bacteremia
abstract: To determine the epidemiology of candidemia in our neonatal intensive care unit; to compare risk factors, clinical presentation, and outcomes for neonates infected with Candida albicans, Candida parapsilosis,and coagulase-negative staphylococcus (CoNS); and to suggest a rational approach to empiric antifungal therapy of neonates at risk for nosocomial infection.  Retrospective chart review of all neonatal intensive care unit patients with systemic candidiasis or CoNS infection between January 1, 1995 and July 31, 1998 at Duke University Medical Center.  Fifty-one patients were reviewed. Nine of 19 patients infected with C parapsilosis and 5 of 15 patients infected with C albicans died of fungemia. Seventeen neonates had >2 positive cultures for CoNS obtained within 96 hours and 1 died. There was no statistically significant difference in birth weight, gestational age, or age at diagnosis between patient groups; however, candidemic patients had a sevenfold higher mortality rate. Before diagnosis, candidemic patients had greater exposure to systemic steroids, antibiotics, and catecholamine infusions. Of the 51 patients, 32 received third-generation cephalosporins in the 2 weeks before diagnosis and 19 did not. Twenty-nine of the 32 who were treated with third-generation cephalosporins subsequently developed candidemia, while candidemia occurred in only 5 of 19 patients who were not treated with cephalosporins. At the time of diagnosis, candidemic patients were more likely to have required mechanical ventilation and were less likely to be tolerating enteral feeding. Multivariate clustered logistic regression analysis revealed that candidemic patients had more exposure to third-generation cephalosporins. Once the clinician was notified of a positive blood culture for Candida, patients infected with C parapsilosis retained their central catheters longer than patients infected with C albicans.  In this retrospective review, we were able to identify aspects of the clinical presentation and medication history that may be helpful in differentiating between candidemia and CoNS bacteremia. Those key features may be used by clinicians to initiate empiric amphotericin B therapy in premature neonates at risk for nosocomial infections. Prolonged use of third-generation cephalosporins was strongly associated with candidemia. There was no statistically significant difference in the morbidity and mortality between patients infected with C parapsilosis and those infected withC albicans. Observed delays in removal of the central venous catheter may have contributed to finding a mortality rate fromC parapsilosis that was higher than was previously reported.

title: Incidence of Candida parapsilosis colonization in an intensive care nursery population and its association with invasive fungal disease
abstract: &NA; We have studied 82 consecutive intensive care nursery admissions to determine rates of colonization and incidence of fungal sepsis. Cultures were obtained from stool, gastric aspirate and skin at three different times. Infants studied ranged in gestational age from 23 to 38 weeks (mean ± SEM 29 ± 0.4 weeks). Nineteen percent of all infants were colonized with Candida sp.; stools were more frequently culture‐positive than skin or gastric aspirates. Colonized infants began enteral feeds at a later time compared with noncolonized neonates. Five of the study infants developed fungal sepsis. One had congenital Candida albicans sepsis and died at 10 days of age; the other four had Candida parapsilosis sepsis and survived. The development of C. parapsilosis sepsis was significantly associated with gastrointestinal colonization. Our results suggest that early initiation of enteral feeds decreases gastrointestinal colonization with C. parapsilosis. Gastrointestinal colonization was strongly associated with the subsequent development of C. parapsilosis sepsis in this group of high risk neonates.

title: Colonization and epithelial adhesion in the pathogenesis of neonatal candidiasis.
abstract: Candida species are important nosocomial pathogens in the newborn population, particularly among the premature very-low-birth-weight infants in neonatal intensive care units. Candida colonization of the neonatal skin and gastrointestinal tract is an important first step in the pathogenesis of invasive disease. C albicans is the most commonly isolated species in colonized or infected infants. Over the past decade the incidence of both colonization and infection with other Candida species, particularly C parapsilosis, has risen dramatically. Colonization of the infant occurs early in life and is affected by a variety of common practices in neonatal intensive care. Microbial factors also augment colonization, including the ability of Candida to adhere to human epithelium. A better understanding of the complex interactions between host risk factors and virulence traits of colonizing yeast may allow the risk of systemic spread to be reduced in the population of premature infants.

title: Vertical and horizontal transmission of unique Candida species to premature newborns.
abstract: The number of nosocomial bloodstream infections due to Candida species in critically ill newborns is increasing. This pathogen may be vertically transmitted from the mother or nosocomially acquired in the nursery. The goal of this study was to identify the route of transmission of unique Candida species and strains from mothers to their preterm offspring. Specimens from mothers for fungal cultures were obtained before delivery, and specimens from infants for sequential fungal cultures were obtained at defined intervals. Candida species were identified by standard methods and were typed by electrophoretic karyotyping (EK) and restriction endonuclease analysis of genomic DNA (REAG) with pulsed-field gel electrophoresis. Antifungal susceptibility testing was performed on all isolates. Fungal cultures were positive for Candida species in 12 (63%) of 19 mothers' specimens and in seven (33%) of 21 infants' specimens. EK and REAG revealed that both the mother and the infant in three (14%) of 21 mother-infant pairs were colonized with the identical strain of Candida albicans. C. albicans was most commonly transmitted vertically. Candida parapsilosis colonized other infants and could not be accounted for by a maternal reservoir.

title: High frequency of Candida parapsilosis on the hands of healthy hosts.
abstract: The presence of yeasts on the hands of 86 healthy hosts (62 hospital workers and 24 healthy members of the community with no hospital exposure) was investigated. A high rate of colonization was found (59.3%). Candida parapsilosis was the most frequently isolated species (51%), independent of the origin of the samples. The potential virulence and resistance to antifungals of the 26 C. parapsilosis isolates were determined. All were proteinase producers and formed biofilms. The haemolytic activity was variable, with a predominance of total haemolysis of sheep erythrocytes. All isolates were susceptible to amphotericin B but two showed reduced susceptibility to fluconazole. Healthy people may be colonized by a species of yeast with a high capacity for adhesion to plastic surfaces, providing an infection risk to susceptible individuals.

title: Candida parapsilosis fungemia in neonates: genotyping results suggest healthcare workers hands as source,
and review of published studies
abstract: An outbreak of Candida parapsilosis fungemia involving 17 neonatal intensive care unit (NICU) patients was studied. There were 14 blood culture and nine colonizing isolates from other sites available. The hands of NICU healthcare workers (HCW) yielded eight isolates. Screening of the isolates by random amplified polymorphic DNA (RAPD) method showed only three profiles. Typing by restriction fragment length polymorphism (RFLP) revealed all blood isolates were RFLP subtype VII-1. Among the nine infant colonizing isolates, there were four different RFLP subtypes; four of the isolates were subtype VII-1. Seven of the eight isolates from HCW were RFLP subtype VII-1. The majority of infant colonizers were not found in the blood, suggesting a possible direct spread of the epidemic subtype VII-1 strain from HCW hands to infant blood. The source of the infant colonizing strains is unclear, but non-VII-1 strains may be largely of maternal origin and VII-1 strains from HCW. These findings reinforce prior studies that have implicated HCW hands as the source of nosocomial, including neonatal, fungemia.

title: Horizontal Transmission of Candida parapsilosis Candidemia in a Neonatal Intensive Care Unit
abstract: This report describes the nosocomial acquisition of Candida parapsilosis candidemia by one of the six premature newborns housed in the same room of a neonatal intensive care unit at the Ospedale Santa Chiara, Pisa, Italy. The infant had progeria, a disorder characterized by retarded physical development and progressive senile degeneration. The infant, who was not found to harbor C. parapsilosis at the time of his admission to the intensive care unit, had exhibited symptomatic conjunctivitis before the onset of a severe bloodstream infection. In order to evaluate the source of infection and the route of transmission, two independent molecular typing methods were used to determine the genetic relatedness among the isolates recovered from the newborn, the inanimate hospital environment, hospital personnel, topically and intravenously administered medicaments, and indwelling catheters. Among the isolates collected, only those recovered from the hands of two nurses attending the newborns and from both the conjunctiva and the blood of the infected infant were genetically indistinguishable. Since C. parapsilosis was never recovered from indwelling catheters or from any of the drugs administered to the newborn, we concluded that (i) horizontal transmission of C. parapsilosis occurred through direct interaction between nurses and the newborn and (ii) the conjunctiva was the site through which C. parapsilosis entered the bloodstream. This finding highlights the possibility that a previous C. parapsilosis colonization and/or infection of other body sites may be a predisposing condition for subsequent C. parapsilosis hematogenous dissemination in severely ill newborns.

title: Outbreak ofCandida parapsilosis fungemia in neonatal intensive care units: Clinical implications and genotyping analysis
abstract: SummaryDuring a 5-month period, 17 infants hospitalized in neonatal intensive care units of a medical center and a branch hospital developed 18 episodes ofCandida parapsilosis fungemia. The mean age at onset was 35 days. Prior to fungemia, all the infants had received hyperalimentation and antibiotics, and 15 infants had had central venous catheters. The presenting symptoms were variable but only vague in 40% of the episodes. Despite administration of antifungal agents, subsequent eradication of fungemia was achieved in only two-thirds of the episodes. None of the environmental samples was positive forC. parapsilosis, while 20% of hand-washing samples of staff working in both units yielded this microorganism. Four genotypes with two main types were identified from 14 outbreak strains and eight genotypes from 14 hand-washing strains, with one type predominant. The results suggest thatC. parapsilosis fungemia increases the morbidity and mortality of neonates but does not cause acute lethal events. The outbreak was caused by two main genotypes, possibly via cross-infection by the hands of health care workers.

title: Epidemiological trends in nosocomial candidemia in intensive care
abstract: BackgroundInfection represents a frequent complication among patients in Intensive Care Units (ICUs) and mortality is high. In particular, the incidence of fungal infections, especially due to Candida spp., has been increasing during the last years.MethodsIn a retrospective study we studied the etiology of candidemia in critically ill patients over a five-year period (1999–2003) in the ICU of the San Martino University Hospital in Genoa, Italy.ResultsIn total, 182 episodes of candidaemia were identified, with an average incidence of 2.22 episodes/10 000 patient-days/year (range 1.25–3.06 episodes). Incidence of candidemia increased during the study period from 1.25 in 1999 to 3.06/10 000 patient-days/year in 2003. Overall, 40% of the fungemia episodes (74/182) were due to C.albicans, followed by C. parapsilosis(23%), C.glabrata (15%), C.tropicalis (9%) and other species (13%). Candidemia due to non-albicans species increased and this was apparently correlated with an increasing use of azoles for prophylaxis or empirical treatment.ConclusionThe study demonstrates a shift in the species of Candida causing fungemia in a medical and surgical ICU population during a 5 year period. The knowledge of the local epidemiological trends in Candida species isolated in blood cultures is important to guide therapeutic choices.

title: Candidemia in a Brazilian hospital: the importance of Candida parapsilosis.
abstract: The aim of this study was to perform a retrospective analysis of cases of candidemia in a Brazilian hospital in the city of Fortaleza, Ceará. A total of 50 blood cultures were analyzed from 40 candidemic patients. The mycological diagnosis was based on the phenotypical analysis and the patients' data were recorded in appropriate files. The most frequent species were Candida parapsilosis (n = 18), followed by C. albicans (n = 14), C. tropicalis (n = 8), C. guillermondii (n = 6), C. glabrata (n = 2), and Candida spp. (n = 2). A detailed descriptive study was undertaken with 21 patients whose medical records were complete. The candidemia episodes occurred in eight male patients and 13 female patients. The most representative risk factors implicated in candidemia were prior antibiotic therapy, central venous catheters, parenteral nutrition, gastric probes and mechanical ventilation. Death occurred in 13 of the 21-candidemic patients. This study demonstrated the emergence of candidemia caused by C. parapsilosis in a Brazilian hospital in the city of Fortaleza, Ceará.

title: Candidemia in a neonatal intensive care unit: trends during fifteen years and clinical features of 111 cases.
abstract: To determine changes in the incidence of candidemia in a neonatal intensive care unit (NICU) during a 15-year period (1981 to 1995) and to compare the prevalence and case fatality rates of Candida albicans and Candida parapsilosis infections.



A retrospective study was conducted of candidemia occurring in infants in a NICU between January 1, 1981, and December 31, 1995. Cases were identified through computerized searching of a microbiology blood culture database. Candidemia was considered contributory to mortality if death occurred within 3 days of positive blood cultures or if there was autopsy evidence of disseminated candidiasis.



One hundred eleven cases of candidemia occurred in 107 infants, representing 1% of all NICU patients during the study period. The rate of candidemia in the NICU increased from 2.5 cases per 1000 admissions in 1981 to 1985, to 4.6 per 1000 admissions in 1986 to 1990 and to 28.5 per 1000 in 1991 to 1995 (P = 0.001). C. albicans was the predominant cause of candidemia between 1981 and 1990. C. parapsilosis was the most prevalent species between 1991 and 1995, causing 53 of 89 cases (60%). The mortality from C. albicans, 13 of 50 cases (26%), was significantly higher than the mortality from C. parapsilosis, 2 of 54 (4%) (P = 0.002; relative risk, 7; 95% confidence interval, 1.7 to 30).



The rate of candidemia in our neonatal intensive care unit increased >11-fold in the 15 years from 1981 to 1995; the prevalent Candida species shifted from C. albicans to C. parapsilosis; and candidemia associated with C. albicans has significantly higher mortality than with C. parapsilosis.

title: Nosocomial outbreak of Candida parapsilosis fungemia related to intravenous infusions.
abstract: Candida parapsilosis is rarely isolated from blood cultures. Our hospital surveillance detected an increased rate of isolation of C parapsilosis during a fourmonth period. Fourteen postoperative patients receiving intravenous (IV) hyperalimentation and eight burn patients receiving IV albumin were involved. Hectic fever, the major clinical manifestation, was seen in 61% of cases. Therapy in the postoperative patients consisted merely of discontinuing IV catheters and hyperalimentation, while amphotericin B was needed in five of eight burn patients to control persistent fungemia. Epidemiologic analysis identified a source of the organism in the IV-additive preparation room, where C parapsilosis was found contaminating a vacuum system. Organisms apparently refluxed into IV bottles when aliquots were removed to accommodate additives. Of 103 patients who received fluids prepared with the contaminated system, 21% became infected with C parapsilosis . Infection surveillance was instrumental in detection and control of the outbreak. Routine guidelines should be established to insure the sterility of IV fluids containing additives.

title: Nosocomial fungemia in neonates associated with intravascular pressure‐monitoring devices
abstract: In the period from January, 1982, to March, 1983, eight infants in the neonatal intensive care unit at one hospital had blood cultures positive for Candida parapsilosis; six cases had occurred after December, 1982. Epidemiologic investigation included a case-control study comparing the 8 cases with 29 birth weight-matched controls. Logistic regression analysis indicated that the model that best fit the observed data included the following risk factors for fungemia: duration of umbilical artery catheterization; duration of receipt of parenteral nutrition; and estimated gestational age. Parenteral nutrition therapy was often administered through the umbilical artery catheters, which were also used for monitoring arterial pressure; transducer domes thus contained parenteral nutrition fluid. Transducers were usually disinfected with alcohol. Laboratory investigation showed that the heads of 6 of 11 in-use blood pressure transducers and 1 of 4 transducers in storage after cleaning were culture-positive for C. parapsilosis. After control measures were instituted no further cases occurred.

title: Candida parapsilosis bloodstream infections in neonatal intensive care unit patients: epidemiologic and laboratory confirmation of a common source outbreak.
abstract: Candida parapsilosis is a common cause of sporadic and epidemic infections in neonatal intensive care units (NICUs). When a cluster of C. parapsilosis bloodstream infections occurred in NICU patients in a hospital in Louisiana, it provided us with the opportunity to conduct an epidemiologic investigation and to apply newly developed molecular typing techniques.



A case-patient was defined as any NICU patient at Louisiana State University Medical Center, University Hospital, with a blood culture positive for C. parapsilosis during July 20 to 27, 1991. To identify risk factors for C. parapsilosis bloodstream infection, a cohort study of all NICU infants admitted during July 17 to 27, 1991, was performed. Electrophoretic karyotyping was used to assess the relatedness of C. parapsilosis isolates.



The receipt of liquid glycerin given as a suppository was identified as a risk factor (relative risk, 31.2; 95% confidence intervals, 4.3 to 226.8). Glycerin was supplied to the NICU in a 16-oz multidose bottle. Bottles used at the time of the outbreak were not available for culture. All six available isolates from four case-patients had identical chromosomal banding patterns; six University Hospital non-outbreak isolates had different banding patterns.



This study demonstrates the utility of combined epidemiologic and laboratory techniques in identifying a novel common source for a C. parapsilosis bloodstream infection outbreak and illustrates that extreme caution should be exercised when using multidose medications in more than one patient.

title: Secular Trends of Candidemia in a Large Tertiary-Care Hospital From 1988 to 2000: Emergence of Candida parapsilosis
abstract: To analyze the secular trends of candidemia in a large tertiary-care hospital to determine the overall incidence, as well as the incidence by ward and by species, and to detect the occurrence of outbreaks.  Retrospective descriptive analysis. Secular trends were calculated using the Mantel-Haenszel test.  A large tertiary-care referral center in Spain with a pediatric intensive care unit (ICU) to which more than 500 children with congenital cardiac disease are admitted annually.  All patients with candidemia occurring from 1988 to 2000 were included. Cases were identified from laboratory records of blood cultures.  There were 331 episodes of candidemia. The overall incidence of nosocomial candidemia was 0.6 episode per 1,000 admissions and remained stable throughout the study period (P = .925). The species most frequently isolated was Candida albicans, but the incidence of C. parapsilosis candidemia increased (P = .035). In the pediatric ICU, the incidence of C. parapsilosis was 5.6 episodes per 1,000 admissions and it was the predominant species. Outbreaks occurred occasionally in the pediatric ICU, suggesting nosocomial transmission.  During this 13-year period, the incidence of candidemia remained stable in this hospital, but C. parapsilosis increased in frequency. Occasional outbreaks of candidemia suggested nosocomial transmission of Candida species (Infect Control Hosp Epidemiol 2005;26:548-552).

title: Candida parapsilosis endocarditis: a comparative review of the literature
abstract: Fungal endocarditis (FE) is an uncommon disease, and while accounting for only 1.3–6% of all cases of infectious endocarditis, it carries a high mortality risk. Although Candida albicans represents the main etiology of FE, C. parapsilosis is the most common non-albicans species. We report the case of a 32-year-old man with a history of prior intravenous drug (IVD) use hospitalized with endocarditis due to C. parapsilosis and review all 71 additional cases documented in the literature. A retrospective analysis of the 72 C. parapsilosis cases compared to 52 recently reviewed cases of C. albicans endocarditis was conducted to identify organism-specific clinical peculiarities. The most common predisposing factor for C. parapsilosis endocarditis (41/72; 57.4%) involved prosthetic valves followed by IVD use (12/72; 20%). Peripheral embolic and/or hemorrhagic events occurred in 28/64 (43.8%) patients, mostly in cerebral and lower limb territories. Overall mortality was 41.7%. Combined surgical and clinical treatment was associated with a lower mortality. Few patients received the newer antifungal agents, and it would appear that more experience is required for their use in the treatment of C. parapsilosis endocarditis.

title: [A case report of Candida prosthetic endocarditis: an autopsy review].
abstract: Most of the increasing numbers of cases of Candida endocarditis have developed after cardiac surgery. We report a case of Candida parapsilosis endocarditis after mitral valve replacement in a patient with no predisposing factors. In this report we mainly examined the pathological findings in this patient with postmortem examination. A woman, age 55, underwent valve replacement with a Björk-Shiley prosthesis for mitral stenosis. The spike fever occurred on the 22nd postoperative day and Candida parapsilosis was cultured from the blood. On the 76th day, a conscious disturbance with the right side hemiplegia was experienced. A CT scan revealed massive hemorrhagic infarction. Despite the intensive care the patient died of cerebral damage. Pathologic findings showed that the vegetation in the major orifice of the Björk-Shiley valve was composed of both necrotic debris and tissue, with Candida organisms being scattered throughout the interstices. Additionally there were also multiple myocardial abscesses with necrotic tissue.

title: Isolated native tricuspid valve Candida endocarditis in a non-drug-addicted patient: case report and review of the literature.
abstract: A case is reported of isolated native tricuspid calve Candida parapsilosis endocarditis (INTVCE) in a male patient with no history of drug abuse or heart disease. The patient had received hyperalimentation and antibiotics for four months via a central venous catheter after abdominal surgery. He underwent successful treatment with tricuspid valve debridement, liposomal amphotericin (AmBisome) and fluconazole, and remained without relapse during an eight-year follow up. A literature review of 12 similar cases (including the present patient) without history of drug abuse or heart disease, dating from 1970, is included.

title: Isolated tricuspid valve endocarditis due to Candida parapsilosis associated with long-term central venous catheter implantation.
abstract: A 72-year-old man was treated for fungal tricuspid valve endocarditis (TVE) with significant tricuspid valvular regurgitation and severe congestive heart failure caused by Candida parapsilosis. The patient had received hyperalimentation and antibiotic therapy for three months through a central venous catheter after the surgical treatment of ileus. The patient was treated medically with amphotericin B and fluconazole because of high surgical risk due to severe pulmonary emphysema, and he responded well. Although TVE caused by C. parapsilosis is rare, we should consider this possibility in patients receiving long-term hyperalimentation and antibiotic therapy using a central venous catheter.

title: [Candida parapsilosis endocarditis after prolonged antibiotic therapy].
abstract: Infective endocarditis is still a very serious disease whose prognosis has improved in recent years, even though advanced technologies have brought a greater number of cases attributable to increased growth of fungi in more virulent forms. We describe a case of endocarditis caused by Candida parapsilosis on a previously healthy aortic valve in a man who had received prolonged intravenous antibiotic treatment a few months earlier. The initial presentation included acute arterial ischaemia of the lower limbs. The diagnosis of endocarditis was then confirmed by echocardiography and microbiology. Specific antifungal therapy was started, and embolectomy and valve replacement were performed; however, the infection could not be controlled and the patient died a few days later.

title: Native valve endocarditis due to Candida parapsilosis: a late complication after bone marrow transplantation-related fungemia.
abstract: A case of Candida parapsilosis endocarditis observed 16 months after BMT is reported. The patient, a 35-year-old female with CML, suffered from Candida parapsilosis fungemia on day +22 after BMT. In spite of treatment with amphotericin B, fluconazole and catheter withdrawal, the same yeast was isolated > 1 year later from a vegetation on an old rheumatic mitral valve. Although the patient remained in complete cytogenetical and hematological remission, in vitro tests showed reduced phagocytic and chemotactic capacity of neutrophils and monocytes. This case stresses the need of prolonged therapy for patients with candidemia after BMT.

title: A case of recurrent Candida parapsilosis prosthetic valve endocarditis: cure by medical treatment alone.
abstract: A patient with recurrent fungal endocarditis on prosthetic mitral valve is presented. Candida parapsilosis was the causative agent. The patient was treated medically with conventional amphotericin during the first episode. When the disease recurred conventional amphotericin B was used again, but had to be stopped because of severe side effects. Treatment was continued with amphotericin B colloidal dispersion, followed by fluconazole for 8 months. The patient is healthy 16 months after discontinuation of fluconazole. Medical treatment of fungal endocarditis on prosthetic valves can be successful in selected cases.

title: Long-term suppressive therapy for Candida parapsilosis-induced prosthetic valve endocarditis.
abstract: Prompt valve replacement is advocated in patients in whom candidal prosthetic valve endocarditis develops. Unfortunately, some patients with this condition are considered nonsurgical candidates, and they are unable to tolerate long-term administration of amphotericin B with or without flucytosine. Herein we describe a patient with Candida parapsilosis-induced prosthetic valve endocarditis in whom oral administration of fluconazole during an 11-month period successfully suppressed the fungal infection. Three previously published cases indicate that long-term noncurative suppressive therapy for C. parapsilosis-induced prosthetic valve endocarditis may allow prolonged symptom-free survival for such patients.

title: Medical treatment of recurrent candidemia in a patient with probable Candida parapsilosis prosthetic valve endocarditis.
abstract: Fungal endocarditis is considered an absolute indication for valve replacement surgery. We describe the successful medical treatment of recurrent Candida parapsilosis candidemia with sequential treatment with amphotericin B and fluconazole in a patient with probable prosthetic valve endocarditis. Because of the presumed effectiveness of amphotericin B and fluconazole in the treatment of this patient, medical therapy should be considered as potentially useful in the treatment of recurrent C parapsilosis fungemia or endocarditis or both.

title: Epidemiology of central nervous system mycoses.
abstract: Fungal infections of the central nervous system (CNS) were considered rare until the 1970s. This is no longer true in recent years due to widespread use of corticosteroids, cytotoxic drugs and antibiotics. Immunocompromised patients with underlying malignancy organ transplantations and acquired immune deficiency syndrome are all candidates for acquiring fungal infections either in meninges or brain. A considerable number of cases of CNS fungal infections even in immunocompetent hosts have been reported. A vast array of fungi may cause infection in the CNS, but barring a few, most of them are anecdotal case reports. Cryptococcus neoformans, Candida albicans, Coccidioides immitis. Histoplasma capsulatum are common causes of fungal meningitis; Aspergillus spp, Candida spp, Zygomycetes and some of the melanized fungi are known to cause mass lesions in brain. Few fungi like C. neoformans, Cladophialophora bantiana, Exophiala dermatitidis, Ramichloridium mackenzie, Ochroconis gallopava are considered as true neurotropic fungi. Most of the fungi causing CNS infection are saprobes with worldwide distribution; a few are geographically restricted like Coccidioides immitis. The infections reach the CNS either by the hematogenous route or by direct extension from colonized sinuses or ear canal or by direct inoculation during neurosurgical procedures.

title: Candidal infection in the central nervous system.
abstract: Candida has become the most prevalent cerebral mycosis at autopsy, indicating a significant incidence coupled with inadequate eradication. Of 29 patients with systemic candidiasis, 48 percent (14 of 29) also had central nervous system involvement. Of these patients, however, only 21 percent (three of 14) had antemortem diagnosis, and only one of these three patients remains alive; the two patients with antemortem diagnosis who died had a meningeal form that, although easier to document on the basis of cerebrospinal fluid examination, is now distinctly rarer than other forms of the disease in adults. The lone surviving patient was treated with amphotericin B for endocarditis and mycotic aneurysms of the cerebral vessels. One clue to central nervous system candidal infection was the striking correlation between cardiac and cerebral involvement; 80 percent of patients with myocardial or valve infection also had central nervous system candidiasis. Most forms of immunosuppression represent a risk factor for both the systemic and cerebral mycoses. Neuropathologically, there is a spectrum of disease entities associated with Candida, including two previously unrecognized lesions reported herein: fungus balls of both white and gray matter and mycotic aneurysms secondary to Candida parapsilosis. Other parenchymal presentations include thrombosis, vasculitis, abscess, hemorrhage, and demyelination. For drug therapy such as amphotericin B to be more effective, earlier diagnosis of these parenchymal infections must be sought.

title: Systemic Candida infections in infants in intensive care nurseries: high incidence of central nervous system involvement.
abstract: The clinical courses in 27 infants with culture or autopsy evidence of systemic candidiasis were reviewed. Twenty-two infants (group 1) had persistent signs of sepsis and clinical deterioration or died before institution of antifungal therapy. Five infants (group 2) improved markedly before culture results were reported, and recovered without systemic antifungal therapy. Fourteen infants in group 1 (64%) had central nervous system infection. Of four patients in whom CNS involvement was diagnosed only postmortem, antemortem cerebrospinal fluid from three was abnormal despite sterile cultures; no antemortem CSF was obtained in the other. In meningitis caused by susceptible organisms addition of flucytosine sterilized CSF within 5 days, although prior amphotericin monotherapy had been unsuccessful. Of 14 patients in group 1 who received systemic antifungal therapy, only one died with Candida infection. Toxicity from antifungal agents occurred in 11 of 13 successfully treated infants, but was reversible in every case except one by modifying the dosage. Our data indicate that (1) CNS infection is very common in infants with systemic candidiasis, (2) combined flucytosine-amphotericin therapy may facilitate treatment of CNS infection and should be the initial therapy for systemic candidiasis in infants, (3) Gram stains of CSF and urine enhance early diagnosis, (4) isolation of Candida from normally sterile body fluids in high-risk infants should be considered pathogenic and therapy initiated unless the clinical course strongly suggests otherwise, and (5) toxicity from antifungal agents is common but usually reversible.

title: Candida peritonitis. Report of 22 cases and review of the English literature.
abstract: Thirty-one patients with Candida isolated from peritoneal fluid were examined. Twenty-two were considered to have Candida peritonitis. The data on these 22 patients, plus 12 additional patients described in the literature, were reviewed. This infection was observed as a complication of peritoneal dialysis, gastrointestinal surgery or perforation of an abdominal viscus. Recent antibiotic administration seemed to be an important predisposing factor. The disease usually remained localized intra-abdominally, although disseminated candidiasis was also noted in three cases. Clinically significant infection could be differentiated from peritoneal contamination with Candida by the presence and persistence of fever, peritoneal signs, peripheral leukocytosis, positive peritoneal cultures for Candida, abnormal films of the abdomen and purulent ascitic fluid. Surgical interventions and removal of infected peritoneal fluid were the cornerstones of therapy. Short-term, low-dose systemic and/or intraperitoneally administered amphotericin B appeared promising in the treatment of unremitting infection. Mortality in treated patients was low and was comparable to that in patients with bacterial peritonitis.

title: Candida meningitis in newborn infants: a review and report of combined amphotericin B--flucytosine therapy.
abstract: Meningitis due to Candida albicans was successfully treated in a 1.1 kg premature infant using combined antifungal therapy of amphotericin B for three weeks and 5-fluorocytosine for four months. Hydrocephalus and profound psychomotor retardation were present one year later. Psychomotor retardation, aqueductal stenosis and hydrocephalus were found to be common in a review of 16 previously reported cases of central nervous system (CNS) candidiasis in newborn infants. The diagnosis and institution of therapy were frequently delayed, and the mortality rate was 29% in the 17 patients reviewed here. The subacute course, lack of clinical findings, variable cerebrospinal fluid (CSF) findings, negative CSF cultures due to low concentrations of organisms, slow in vitro growth of C. albicans and misinterpretation of positive cultures as contaminants are factors frequently leading to delayed diagnoses. Using combination therapy, it should be possible to use lower doses and shorter courses of amphotericin B therapy for C. albicans meningitis in the newborn infant.

title: Chronic candidal meningitis: an uncommon manifestation of candidiasis.
abstract: Chronic meningitis is an uncommon manifestation of candidiasis. We present the case of an elderly woman who had symptoms such as headache, malaise, and fever for 8 months and was found to have Candida albicans meningitis, and we review 17 similar cases. An underlying illness or risk factor for candidiasis was present in only 13 (72%) of the 18 patients. Headache, fever, and nuchal rigidity were the predominant clinical findings. Analysis of CSF showed either mononuclear or neutrophilic pleocytosis, an elevated protein level, and a decreased level of glucose. Only 17% of CSF smears were positive, and only 44% of initial CSF cultures yielded Candida species. In four cases, Candida species grew only after special techniques were used; in three cases, CSF cultures remained negative. The overall mortality associated with candidal meningitis was 53%, but among 12 patients who were treated and followed, the rate was 33%. In addition to acute meningitis seen with disseminated infection, Candida species can cause chronic meningitis that mimics tuberculosis and the more common fungal meningitides, such as cryptococcosis.

title: Candidal meningitis in neonates: a 10-year review.
abstract: Candidal meningitis may complicate systemic candidiasis in the premature neonate. We conducted a 10-year retrospective review of 106 cases of systemic candidiasis in neonates to define the incidence, clinical features, laboratory findings, treatment, and outcome of candidal meningitis. Twenty-three of the 106 neonates had candidal meningitis (0.4% of admissions to the neonatal intensive care unit). The median gestational age was 26.2 weeks, the median birth weight was 820 g, and the median age at the onset of illness was 8 days. Clinical disease was severe and commonly was manifested by respiratory decompensation. Findings of cerebrospinal fluid (CSF) analyses varied: pleocytosis was inconsistent, hypoglycorrhachia was common, gram staining was uniformly negative, and Candida was isolated from 17 neonates (74%). Each infant was treated with amphotericin B (median cumulative dose, 30 mg/kg); 5 also received flucytosine therapy. In conclusion, initial clinical features of candidal meningitis are indistinguishable from those of other causes of systemic infection in premature neonates, and normal CSF parameters do not exclude meningitis. Timely initiation of amphotericin B monotherapy was associated with an excellent outcome.

title: Candida species isolated from cerebrospinal fluid
abstract: In the period from November 1998 to June 2001 13 cases of nosocomial meningitis were reported.Candida albicans was isolated from 54% of the patients (7);C. parapsilosis from 23% (3);C. tropicalis from 15% (2) andC. krusei from 8% (1).C. albicans was isolated from the cerebrospinal fluid (CSF) of five children with the following diagnoses: nonspecified tumor of the central nervous system, Hodgkin's disease, meningitis, suspect neuroinfection, and sepsis. Examination of CSF allowed us to detect 2 strains ofC. albicans from adult patients, one after neurosurgery because of a brain tumor and one with a vascular disease of the brain.C. parapsilosis was found in CSF from two premature children and one child with epilepsy. Two isolates ofC. tropicalis were obtained from both blood and the CSF of a child from the neonatal intensive care unit and from a child from pediatric oncology with multiple malignant neoplasms. Only one strain ofC. krusei was found in the oral cavity and CSF of a patient after neurosurgery performed after head trauma.

title: [Severe infection caused by Candida parapsilosis in an infant].
abstract: Three cases of serious infections due to Candida parapsilosis in infants are reported. Two patients are children exposed to abdominal surgery, parenteral nutrition and antibiotic treatment, who developed catheter-associated sepsis with successful response to change of catheter and intravenous amphotericin B, despite one of the patients showed a relapse some weeks after. Third patient, a hydrocephalic child with external ventricular shunt, developed cerebrospinal fluid infection without response to antifungal treatment and died. Sepsis and meningitis due to Candida parapsilosis can be observed in infants with some risk factors, as use of intravascular or intraventricular catheters. Change of the catheter and treatment with amphotericin B are indicated in these infections.

title: Factors predicting outcome of fungal peritonitis in peritoneal dialysis: analysis of a 9-year experience of fungal peritonitis in a single center.
abstract: Fungal peritonitis causes significant morbidity and mortality for patients undergoing continuous ambulatory peritoneal dialysis (CAPD). We retrospectively reviewed 70 episodes of fungal peritonitis in a single center over the last 9 years in 896 CAPD patients. Seventy percent of the episodes of fungal peritonitis were caused by Candida species, among which 50% were Candida parapsilosis. As a result of fungal peritonitis, 44% of the patients died, whereas further peritoneal dialysis failed in 14%, requiring a change to long-term hemodialysis. Only 37% managed to continue CAPD. The remaining 5% either underwent transplantation or were lost to follow-up. We identified the factors associated with poor outcome, namely mortality and technique failure. The presence of abdominal pain, bowel obstruction, and a catheter remaining in situ were significantly associated with greater mortality. Abdominal pain, antibiotic use within 3 months before fungal peritonitis, and complication by bowel obstruction were associated with greater technique failure. In choosing antifungal agents with catheter removal, oral fluconazole alone appears equally as effective as combined oral fluconazole with 5-flucytosine for peritonitis caused by Candida species. For peritonitis caused by species other than Candida, the choice of antifungal therapy needs to be individualized, based on fungal species and sensitivities.

title: Fungal peritonitis in peritoneal dialysis: critical review of six cases.
abstract: Fungal peritonitis (FP) is uncommon in patients on peritoneal dialysis (PD); it is difficult to treat and has a high mortality rate. We report 6 cases of fungal peritonitis observed between 1980 and 1992 in our center. The etiologic agents were: Candida spp., C. guilliermondi, C. parapsilosis, C. albicans, and Verticillium spp. All 6 patients had suffered at least one episode of bacterial peritonitis in the two months before the fungal infection appeared and were all treated by intraperitoneal administration of antibiotics. The catheter was removed early in 3 patients followed by antimycotic therapy, while the remaining 3 patients received antimycotic therapy, with removal of the catheter in a later stage. The result in the first group was that they all switched permanently to hemodialysis, while in the second group there were 2 deaths and 1 transfer to hemodialysis. In the light of these 6 cases, we analyzed 22 published reports to assess risk factors, therapy, and outcome of this pathology. The major predisposing factors were intraperitoneal antibiotics and bacterial peritonitis, and the best results were obtained by continuing PD plus intraperitoneal and systemic antifungal agents.

title: A Retrospective Study of Seven Cases of Candida Parapsilosis Peritonitis in CAPD Patients: The Therapeutic Implications
abstract: Candida peritonitis accounts for the majority of fungal peritonitis in continuous ambulatory peritoneal dialysis (CAPD), but the Candida species were not routinely subtyped in previous studies. The clinical course and the outcome of Candida parapsilosis peritonitis remain unclear.  To study the clinical course and outcome of C. parapsilosis peritonitis in CAPD patients.  Peritoneal dialysis unit in a regional hospital.  A retrospective study on seven cases of C. parapsilosis peritonitis occurring in a single center over 3 years.  The 7 patients included 4 males and 3 females. Their mean age was 62 ± 11.5 years. Two (29%) were diabetic. Three (43%) had a history of preceding peritonitis and 5 (71%) had received broad spectrum antibiotic within the previous 1 month. All presented with cloudy dialysate, abdominal pain, and fever. The mean dialysate white cell count was 300 ± 168/mm3 with a predominance of neutrophils (81.4% ± 13.1%). The mean time from onset of symptoms to diagnosis was 5.7 ± 3.1 days. All had been treated with immediate catheter removal within 24 hours of diagnosis and antifungal therapy, including oral fluconazole, intravenous (IV) amphotericin, or their sequential combination. Environmental samplings were negative for C. parapsilosis. The overall complication rate was exceptionally high (71%), with three (43%) complicated by abscess formation requiring surgical drainage, one peritoneal adhesion (14%), and one mortality (14%). In the end, only two (29%) could resume CAPD.  The outcome of this study group appeared worse than those previously described in the literature, and the optimal treatment for this group of patients remains unclear.

title: Candida Parapsilosis peritonitis in patients on CAPD
abstract: Twenty-four episodes of C. parapsilosis peritonitis in 23 patients on continuous ambulatory peritoneal dialysis (CAPD) over 6 years were reviewed. Clinical manifestations and laboratory findings were similar to those of other pathogens. All started treatment with intravenous amphotericin B. In six cases it was attempted to maintain a peritoneal catheter in situ, but removal became essential to relieve fungal peritonitis. Of the patients who developed peritonitis, 15 episodes (62.5%) continued the CAPD program. Nine cases could not resume CAPD because of death in 4, patient preference in 2, and abdominal adhesion in 3. Antifungal treatment alone was ineffective in most cases. It was found that peritonitis developing after gram negative bacterial peritonitis and the use of fluconazole after catheter removal were associated with CAPD discontinuation. It was suggested that C. parapsilosis peritonitis in CAPD patients should be treated with rapid catheter removal, particularly those with fungal peritonitis who had prior gram negative peritonitis.

title: In vitro preclinical evaluation studies with the echinocandin antifungal MK-0991 (L-743,872)
abstract: The echinocandin MK-0991, formerly L-743,872, is a water-soluble lipopeptide that has been demonstrated in preclinical studies to have potent activity against Candida spp., Aspergillus fumigatus, and Pneumocystis carinii. An extensive in vitro biological evaluation of MK-0991 was performed to better define the potential activities of this novel compound. Susceptibility testing with MK-0991 against approximately 200 clinical isolates of Candida, Cryptococcus neoformans, and Aspergillus isolates was conducted to determine MICs and minimum fungicidal concentrations MF(s). The MFC at which 90% of isolates are inhibited for 40 C. albicans clinical isolates was 0.5 microg/ml. Susceptibility testing with panels of antifungal agent-resistant species of Candida and C. neoformans isolates indicated that the MK-0991 MFCs for these isolates are comparable to those obtained for susceptible isolates. Growth kinetic studies of MK-0991 against Candida albicans and Candida tropicalis isolates showed that the compound exhibited fungicidal activity (i.e., a 99% reduction in viability) within 3 to 7 h at concentrations ranging from 0.06 to 1 microg/ml (0.25 to 4 times the MIC). Drug combination studies with MK-0991 plus amphotericin B found that this combination was not antagonistic against C. albicans, C. neoformans, or A. fumigatus in vitro. Studies with 0 to 50% pooled human or mouse serum established that fungal susceptibility to MK-0991 was not significantly influenced by the presence of human or mouse serum. Results from resistance induction studies suggested that the susceptibility of C. albicans was not altered by repeated exposure (40 passages) to MK-0991. Erythrocyte hemolysis studies with MK-0991 with washed and unwashed human or mouse erythrocytes indicated minimal hemolytic potential with this compound. These favorable results of preclinical studies support further studies with MK-0991 with humans.

title: Comparison of Peritoneal Fluid Culture Results from Adults and Children Undergoing Capd
abstract: Peritonitis is a common complication in patients with end-stage renal disease treated by continuous ambulatory peritoneal dialysis (CAPD). Empirical treatment is based on the organisms that are most frequently isolated and their susceptibilities.  To analyze and then compare peritoneal fluid culture results from adult and pediatric patients on CAPD, with respect to micro-organisms and antimicrobial susceptibilities.  Three-year retrospective review of peritoneal fluid cultures from adults and children on CAPD.  We isolated 481 organisms from 378 peritoneal fluid specimens, collected from 135 patients (45 children, 90 adults). There were 191 episodes of peritonitis in children (mean 4.2 ± 3.5, range 1 – 15) compared to 187 in adults (2.1 ± 1.9, range 1 – 10) (p < 0.001). Two or more episodes occurred in 30 of 45 children (67%) compared to 33 of 90 adults (37%) (p < 0.001). The number of different organisms/patient as well as the total number of isolates/patient were significantly greater in children (respectively, 2.8 ± 2.3, range 1 – 12; and 5.3 ± 5.2, range 1 – 27) than in adults (2.0 ± 1.3, range 1 – 6; and 2.7 ± 2.4, range 1 – 10) (p < 0.005). After Staphylococcus epidermidis, S. aureus was the most frequently isolated organism, occurring in 18% of episodes in adults and 12% of episodes in children (p < 0.01). Twenty-two of 33 fungal isolates (67%) in children were Candida parapsilosis compared to 3 of 24 (12%) in adults (p < 0.001). Subanalysis of multiple episodes revealed that Pseudomonas and Candida occurred significantly more often in children (p < 0.01), whereas S. aureus occurred more often in adults (p < 0.001). In polymicrobial episodes S. epidermidis occurred more often in adults (p < 0.05). Significant differences in susceptibilities to ampicillin, ceftriaxone, chloramphenicol, and gentamicin were found between children and adults (p < 0.05 – 0.001).  CAPD-associated peritonitis occurs significantly more often in children than adults. Significant differences in microbial etiology and susceptibilities were found between pediatric and adult patients. Each dialysis unit should periodically analyze peritoneal fluid culture results from its CAPD patients. These data can then be used for optimization of empirical antimicrobial therapy of peritonitis.

title: Fungal peritonitis in peritoneal dialysis patients: effect of fluconazole treatment and use of the twin-bag disconnect system.
abstract: Fungal peritonitis is an uncommon but potentially life-threatening complication for patients undergoing continuous ambulatory peritoneal dialysis. This retrospective study evaluated the efficacy of fluconazole in fungal peritonitis treatment and the incidence of fungal peritonitis in different peritoneal dialysis disconnect systems. Fungal peritonitis was caused by Candida species in 67% of episodes. The most common pathogen in this series was Candida parapsilosis (29%), followed by Candida albicans (14%). One patient (5%) died within 1 month after admission for treatment of fungal peritonitis. Only 1 patient (5%) in this series could resume peritoneal dialysis. Treatment with fluconazole alone has an effect comparable to intraperitoneal (IP) amphotericin B alone or IP amphotericin B combined with oral or intravenous fluconazole. The incidence of fungal peritonitis in patients who used the spike, Y-set, and UV antiseptic systems was 5.69, 6.20, and 2.93 times, respectively, as frequent as that of fungal peritonitis in patients who used the twin-bag disconnect system.

title: Candida parapsilosis Peritonitis Has More Complications Than Other Candida Peritonitis in Peritoneal Dialysis Patients
abstract: Candida parapsilosis is the most prevalent pathogen of fungal peritonitis in peritoneal dialysis (PD). The difference between C. parapsilosis peritonitis and other C. species for clinical outcomes and treatment responses to fungal peritonitis remains unclear. This retrospective study of fungal peritonitis attempts to answer that question. A total 22 patients with fungal peritonitis in 762 PD patients were enrolled in this study. The mean age of the 22 patients, 9 males and 13 females, was 54.7 ± 12.5 years with a mean PD duration of 39.7 ± 33.4 months. Candida species accounted for 86% (19 cases) of fungal peritonitis and 41% (9 cases) were C. parapsilosis. Thirteen (59%) patients received fluconazole as monotherapy; others received either amphotericin B alone or in combination with fluconazole. Catheters were removed for all patients. The mean duration from peritonitis onset to catheter removal was 5.8 ± 4.1 days. Eleven (50%) patients developed severe complications, with abscess formation or persistent peritonitis after catheter removal. C. parapsilosis peritonitis had a higher complication rate than other Candida species (78% versus 20%, p = 0.012). In patients who received fluconazole as monotherapy, the rate of severe complications of C. parapsilosis peritonitis was statistically higher than those of other Candida species (100% versus 29%, p = 0.013). Because of different severity and prognosis, C. parapsilosis peritonitis in PD patients should be treated more aggressively than other Candida species.

title: Pigeons and peritonitis?
abstract: We report an outbreak of fungal peritonitis due to Candida parapsilosis in 12 patients undergoing chronic ambulatory peritoneal dialysis (CAPD). All 12 patients were treated by removal of the CAPD catheter together with systemic antifungal therapy. There were no peritonitis-related deaths. Four patients were successfully returned to CAPD at a later date. Microbiological investigation during the outbreak demonstrated colonization of various areas of the CAPD Unit and medical ward with the organism. C. parapsilosis was also isolated from pigeon guano obtained from window-sills. The number of cases of peritonitis due to this organism decreased markedly after bird-proof netting was installed. We believe that this is the first report of an outbreak of CAPD peritonitis due to faecal carriage of C. parapsilosis by pigeons.

title: Candida arthritis in adult patients who are not intravenous drug addicts: report of three cases and review of the literature.
abstract: Candida species are an uncommon cause of infectious arthritis, although the frequency has increased during recent years. Three cases of septic arthritis caused by Candida species are reported, and the literature is reviewed. The first patient developed a popliteal cyst infected by Candida albicans after multiple intravenous antibiotic treatments. The second patient had acute myelogenous leukemia and experienced knee arthritis after chemotherapy, and the third suffered oligoarthritis after a second heart transplantation. All patients received treatment with a standard dose of intravenous amphotericin B. Responses were achieved only in the first two cases; the third patient died despite therapy. Thirty-six previously reported Candida arthritis cases are reviewed, and epidemiologic, diagnostic, therapeutic, and prognostic features are analyzed. Cases are divided into two categories: direct inoculation of fungus and hematogenously disseminated disease; these two categories are compared in terms of sex, age, pathogen species, treatment, and survival. Arthritis induced through direct inoculation of fungus is seen in older individuals, is more frequently produced by species other than C albicans (Candida parapsilosis especially), and has a better prognosis than arthritis caused by disseminated candidiasis. Arthritis can be resolved even in the persistence of the systemic disease. It is believed that the first case of Baker's cyst infected by C albicans and the first case of Candida arthritis in a heart transplant patient are reported here.

title: Successful treatment of Candida parapsilosis (fluconazole-resistant) osteomyelitis with caspofungin in a HIV patient
abstract: Treating Candida arthritis is challenging. We report a case of Candida parapsilosis arthritis successfully treated with caspofungin. We illustrate the likelihood of severe infections due fluconazole resistant C. parapsilosis after extensive fluconazole use and discuss the role of newer antifungal agents in the treatment of arthritis due to Candida spp.

title: Candidemia in children with central venous catheters: role of catheter removal and amphotericin B therapy
abstract: We reviewed retrospectively 31 cases of candidemia in children with central venous catheters. Infection rate was significantly higher in 1− to 4-year-old children than in other age groups (8.4% vs. 2.2%; P < 0.05). Serious sequelae occurred in 11 (35%) cases and included fatal outcome (5 instances), Candida endocarditis (2), renal abscesses, meningitis, arthritis and osteomyelitis (1 each). Complications were significantly more common in infants than in older children (P < 0.05) and appeared 3 to 52 days after the first positive blood culture (mean, 16 days). In fatal cases catheters were left in place a significantly greater number of days than in nonfatal cases (P < 0.05). A literature review identified 43 additional cases of catheter-related candidemia described in 11 series. The rate of Candida infection in the group as a whole was 2.7%. Patients treated with catheter removal plus amphotericin B had a significantly higher cure rate then patients treated with catheter retention plus amphoteriein B (P = 0.009). Prompt catheter removal remains crucial in the treatment of catheter-related candidemia.

title: Candida tropicalis arthritis - assessment of amphotericin B therapy.
abstract: A 28 year old male heroin addict developed Candida tropicalis infection of the knee joint in association with candidemia. Assessment of amphotericin B therapy was facilitated by the determination of serum and synovial fluid amphotericin B concentrations using a radiometric bio-assay method. The results indicate that adequate synovial fluid drug levels were achieved with intravenous systemic therapy.

title: Candidal arthritis after complete treatment of systemic candidiasis.
abstract: Over the last few decades, the incidence of invasive candidal infections in neonatal intensive care units has increased dramatically. Various complications, such as arthritis, endocarditis, meningitis, and endophthalmitis, have been reviewed. We present the case of a premature infant with systemic candidemia. Arthritis was discovered 6 months after completion of amphotericin B therapy, and was successfully treated with oral fluconazole for 6 weeks. We conclude that long-term follow-up is particularly important in patients with treated candidemia. To prevent complications, prolonged treatment with high-dose amphotericin B is suggested for systemic fungal infection, and oral fluconazole is an effective alternative for candidal arthritis.

title: Candida prosthetic arthritis: report of a case treated with fluconazole and review of the literature.
abstract: Prosthetic arthritis due to Candida species is uncommon, with only 15 cases reported in the literature. We recently cared for a human immunodeficiency virus-infected patient who developed Candida parapsilosis prosthetic arthritis unresponsive to resection arthroplasty, intravenous amphotericin B, and suppressive ketoconazole therapy. Treatment with fluconazole led to mycologic cure and symptom improvement, although he subsequently underwent above-the-knee amputation due to continued joint instability. Fluconazole may be useful follow-up therapy after a course of amphotericin B combined with resection arthroplasty or when removal of the prosthesis cannot be accomplished.

title: Synovial fluid levels of fluconazole in a patient with Candida parapsilosis prosthetic joint infection who had an excellent clinical response.
abstract: A 73-year-old woman presenting with persistent swelling of the right knee 30 months after total joint arthroplasty was found to have a Candida parapsilosis infection. Treatment with fluconazole resulted in full recovery.

title: Successful salvage of a primary total knee arthroplasty infected with Candida parapsilosis.
abstract: Fungal infections of total joint arthroplasties are extremely rare with only 21 previous reported cases in the literature. In 19 of these cases, the offending organism has been a member of the candida species. In all of these cases, the patients had no clinical evidence of disseminated fungal infection. All previously reported cases of total joint fungal infections required removal of the primary prosthesis to eradicate the infection. There has also been a great reluctance to reimplant these patients. In fact, reimplantation has been successful in only one reported case. We report the first case of successful salvage of an arthroplasty infected with candida.

title: Staged reimplantation of total knee arthroplasty after Candida infection.
abstract: Prosthetic joint infection with Candida is uncommon. Only 28 cases have been reported in the English literature. Successful reimplantation after eradication of Candida infection has been reported in 3 hip joints and only 1 knee. We present the case of a 68-year-old woman with chronic Candida parapsilosis infection of a prosthetic knee joint. Removal of the prosthesis, thorough débridement, and antifungal therapy treated the infection successfully. Antifungal therapy included 6 weeks of parenteral administration of fluconazole followed by 4 weeks of oral fluconazole. The involved knee joint was reimplanted 3 months after initial treatment. The prosthetic joint was pain free and functioned satisfactorily during the ensuing 4 years. No recurrence of infection was noted. The principle in treating Candida prosthetic infection generally has been the same as that of bacterial prosthetic infection. In chronic cases, removal of implants, thorough débridement, and effective antifungal therapy are mandatory for the eradication of infection. Reimplantation of the prosthesis can be performed successfully in a staged surgical procedure with the interval between the 2 stages shortened to 3 months.

title: Fungal arthritis of the knee caused by Candida parapsilosis in a kidney transplant recipient.
abstract: Fungal arthritis is a rare infection usually caused by Candida species. Candida parapsilosis has emerged as a new pathogen and has been implicated in several cases of fungal arthritis, mostly associated with a joint prosthesis or after arthrocentesis. We report an unusual case of fungal arthritis of the knee caused by this organism in a 38-year-old female kidney transplant recipient, who had no previous invasive procedures on her knee. Systemic and intraarticular antifungal therapy combined with surgical debridement proved to be an effective treatment of this clinically difficult condition. To our knowledge, this case is the first report of fungal arthritis caused by C. parapsilosis in a transplant recipient. The multimodality approach used in the treatment of this patient should be considered in similar forms of fungal arthritis in immunocompromised patients.

title: Successful treatment of postoperative Candida endophthalmitis in an eye with an intraocular lens implant.
abstract: Candida endophthalmitis developed in the left eye of a 71-year-old man two months after he underwent extracapsular cataract extraction and intraocular lens implantation. The symptoms included decreased vision and redness but no pain. The discovery of the cause of the endophthalmitis was delayed because it was initially treated as a sterile postoperative inflammation. Vitrectomy with intracameral amphotericin B and treatment with topical and systemic amphotericin B and flucytosine led to resolution of the infection and a final visual acuity of 20/80. It was not necessary to remove the intraocular lens.

title: POSTOPERATIVE ENDOPHTHALMITIS SECONDARY TO CANDIDA PARAPSILOSIS: A Case Treated by Vitrectomy and Intravitreous Therapy
abstract: A 79-year-old woman developed Candida parapsilosis endophthalmitis nine weeks after an uneventful cataract extraction. The diagnosis was suspected on clinical examination and documented by culture of the vitreous biopsy obtained during vitrectomy. The endophthalmitis was treated by pars plana vitrectomy and intravitreous injection of amphoterlcin B and dexamethasone. The patient was also treated with systemic flucytosine, to which the operative isolate proved resistant. This was therefore immediately discontinued; however, the intraocular infection cleared rapidly after the initial vitrectomy and intravitreous injection. The patient's visual acuity ultimately returned to 20/20. The combination of these two newer methods of treatment for fungal endophthalmitis achieved a favorable outcome in a disease process that has often previously wrought disaster.

title: Outbreak of Candida parapsilosis endophthalmitis after cataract extraction and intraocular lens implantation
abstract: Between November 1983 and January 1984, 13 cases of Candida parapsilosis endophthalmitis occurred in Florida, Georgia, and Tennessee in patients who had had an intraocular lens implantation (IOLI) or cataract extraction with an IOLI. This outbreak followed the introduction in July 1983 of a new brand of balanced salt solution (BSS) used as an intraoperative ophthalmic irrigation solution. This product was subsequently recalled because of intrinsic fungal contamination. A retrospective cohort study including 704 ophthalmology patients at risk for exposure to this brand of BSS revealed that definite exposure to that product was a significant risk factor for C. parapsilosis endophthalmitis (P less than 0.001, Fisher exact test). A retrospective case control study including 203 control patients with definite exposure to BSS suggested that exposure to systemic steroids (P = 0.007, Fisher exact test) was an additional risk factor for C. parapsilosis endophthalmitis. Treatment modalities among the 13 patients included topical, intraocular, or systemic antifungal therapy (or a combination of these modalities) in 13 patients and vitrectomy in 10 patients. No patients had systemic symptoms or complete visual loss. Laboratory investigations showed a 6.7% overall contamination of the product with C. parapsilosis. After recall of the product by the manufacturer, no patients having a cataract extraction or IOLI at the institutions studied are known to have developed C. parapsilosis endophthalmitis.

title: Epidemic postsurgical Candida parapsilosis endophthalmitis. Clinical findings and management of 15 consecutive cases.
abstract: Fifteen cases of postoperative Candida parapsilosis endophthalmitis occurring secondary to a contaminated lot of an irrigating solution were studied. All patients underwent a vitreous tap or diagnostic and therapeutic vitrectomy. Eleven of the 15 specimens were positive for the organism. Fourteen patients were treated with pars plana vitrectomy surgery. All patients were treated with intravitreal amphotericin B and systemic amphotericin B and 5-fluorocytosine. Two clinical recurrences were successfully treated with intravitreal amphotericin B, removal of the pseudophakos, and oral ketoconazole. The intraocular lens was retained in 11 of the 14 pseudophakic patients. Final visual acuities ranged from 20/25 to no light perception with eight of 15 patients having 20/60 or better visual acuities. Measurable levels of intraocular amphotericin B were found after systemic amphotericin B administration. Two patients with totals of 20 and 30 micrograms of intravitreal amphotericin B over 48 and 96 hours, respectively, had near normal ERGs one year later. Posterior capsulotomy and vitrectomy appear to decrease amphotericin B toxicity and allow sequential intraocular injection of this drug within a short time period.

title: Endophthalmitis in patients with disseminated fungal disease.
abstract: Fungal endophthalmitis caused by dissemination from extraocular fungal infections has been reported to vary between 9% and 45%. However, recent clinical experience disagrees with that. This study is an investigation of patients in an inner city teaching hospital, the risks associated with endogenous fungal endophthalmitis, and this incidence.



All ophthalmology consultations between February 1995 and August 2000 that might be associated with disseminated fungal infection were examined in a prospective manner. Patients were excluded if there was no evidence of a positive fungal culture from any site at any time. Visual symptoms were recorded along with ophthalmologic and systemic examination features. Information was gathered, including the identity of cultured organisms, the sites from which the organisms were obtained, and the patients' disposition.



During this interval, 170 consultation requests contained the words "endophthalmitis" or "retinitis" and/or indicated concern about disseminated fungal infections. Extraocular fungal infections were found in 114 patients, but only 82 of them had evidence of systemic dissemination. Some patients had more than one organism. The following are listed in decreasing frequency of occurrence: Candida albicans, Torulopsis glabrata, Candida tropicalis, Candida parapsilosis, Candida krusei, Aspergillus niger, and others. Only two patients had evidence of chorioretinitis and progressed to fungal endophthalmitis.



Endophthalmitis was rare among these patients with known fungal infections. Less than 2% had any related ophthalmic manifestations. Nevertheless, since treatment can save vision, evidence of intraocular infection should be sought as eagerly as before.

title: Yeast-like microorganisms in eye infections
abstract: The proportion of yeast species involved in eye infections in 11 patients was examined. The presence of yeast organisms as causative agents of endophthalmitis was found in corneal smears (n=4), conjunctival swabs (4), and vitreous fluid (3). Altogether 5 strains ofCandida albicans, 2 strains ofC. krusei and one strain each ofC. guilliermondii, C. parapsilosis, C. tropicalis andCryptococcus neoformans were isolated from the clinical material. The hematogenic origin of endophthalmitis was proved in 7 cases on the basis of positive blood samples and in 2 cases by the isolation of yeasts from the tip of an intravenous catheter. Endophthalmitis-supporting risk factors such as indwelling intravenous catheters, prolonged use of broad-spectrum antibiotics and chemotherapy, surgical intervention, diabetes mellitus, and malignancy were observed in the patients.

title: Fungal Colonization and Infection in Boston Keratoprosthesis
abstract: To determine the incidence of and risk factors for fungal keratitis and endophthalmitis in patients with a Boston keratoprosthesis (KPro) and to determine whether surveillance cultures were helpful in predicting fungal infection.  A retrospective chart review was performed of 182 patients (202 eyes) who received a type 1 (through cornea) or type 2 (through cornea and lid) KPro between March 1, 1990, and December 31, 2004, and who were followed for at least 1 month (range, 1 month to 13 years; mean, 2.84 years). There were 148 eyes with type 1 and 54 eyes with type 2. Beginning in late 1999, many eyes were given a prophylactic topical regimen containing vancomycin, and many eyes with type 1 KPro were given therapeutic contact lenses. Cases of fungal keratitis or endophthalmitis were analyzed. To determine the fungal colonization rate, 70 surveillance cultures of the ocular or lid surface around the KPro optic were obtained of 36 uninfected KPro eyes (35 patients) at random time-points over 1 year (August 2002 to July 2003).  There were 4 definite and 1 probable fungal infections in 6893 patient-months of follow-up, or 0.009 fungal infections per patient-year. These included 3 cases of definite or probable Candida endophthalmitis (C. parapsilosis, C. glabrata, and C. albicans) and 2 cases of mold keratitis (Alternaria, Fusarium). The rate was higher in eyes receiving a vancomycin-containing topical prophylactic regimen than those with on a non-vancomycin regimen (5 cases/2774 person-months vs. 0 cases/4119 person-months; P = 0.011). In eyes with type 1 KPro, the rate was higher with therapeutic contact lens wear than without (4/1682 vs. 0/3115 person-months; P = 0.015). Surveillance cultures did not predict fungal infection, and none of the 6 surveillance eyes colonized with fungi (all Candida) developed a fungal infection. The prevalence of fungal colonization in KPro eyes had not changed since our 1996 surveillance study (11% vs. 10%, P > 0.05).  Fungal infections in KPro eyes have appeared since we introduced broad-spectrum antibiotic prophylaxis and therapeutic contact lenses 5 years ago, but the infection rate remains very low in our mostly New England-based patient population. Cleaning or replacing the contact lens on a regular basis and prescribing a short course of topical amphotericin at the first visible signs of fungal colonization may prevent these infections.

title: Pseudophakic Candida parapsilosis Endophthalmitis with a Consecutive Keratitis
abstract: We describe a case of Candida parapsilosis endophthalmitis with a consecutive keratitis after phacoemulsification and posterior chamber intraocular lens implantation in an otherwise healthy eye. Despite aggressive medical and surgical management during a 2-year period, multiple episodes recurred, with the development of an intracapsular plaque and an infectious nidus on the corneal endothelium 6 months after the initial presentation. After subtotal removal of the culture-positive capsule, intravitreal and topical amphotericin B, and oral fiuconazole, the inflammation improved. However, the corneal endothelial plaque persisted with recurrent inflammation 2 months later, prompting debridement of the culture-positive plaque and further removal of the culture-negative capsular remnants and lens implant. The infection was quiescent for the subsequent 12-month period until recurrent intraocular inflammation developed with enlargement of the endothelial plaque. Culture of this plaque was again positive for C. parapsilosis. After debridement and intraocular and topical amphotericin B, the eye has now been quiescent for 13 months. This case demonstrates the development of a secondary keratitis in an eye affected by pseudophakic C. parapsilosis endophthalmitis, with the posterior cornea serving as a sanctuary site for the fungus despite aggressive management leading to recurrent infection 1 year after the clinical disease appeared to be quiescent.

title: Candida endophthalmitis associated with intraocular lens implantation: efficacy of fluconazole therapy
abstract: Four patients with Candida parapsilosis endophthalmitis following intraocular lens implantation were treated with oral fluconazole. All had failed prior therapy with intravitreal amphotericin B. The only patient who had the lens implant removed was cured after treatment with fluconazole for one year. Three patients who did not have the lens implant removed had resolution of symptoms and return of visual acuity towards normal during the year they were on fluconazole. However, five months after therapy had been stopped, all three had decreasing visual acuity, and in two, culture of vitreous fluid yielded C. parapsilosis. Fluconazole may be effective for treatment of endophthalmitis associated with an intraocular lens implant, but only when the implant is removed also.

title: Bilateral Candida parapsilosis endophthalmitis.
abstract: We describe a patient with bilateral, delayed endophthalmitis who underwent bilateral pars plana vitrectomy, total capsulectomy, intraocular lens exchange, intravitreal injection of amphotericin B, and oral fluconazole therapy. The long-term inflammation resolved, and vitreous cultures from both eyes yielded Candida parapsilosis. Histopathologic examination revealed sequestered yeast forms in the capsular bags.

title: The changing spectrum of fungal keratitis in south Florida.
abstract: To review the clinical experience with fungal keratitis in south Florida over a 10-year period.



One hundred twenty-five cases of fungal keratitis were identified in the microbiology laboratory records between January 1982 and January 1992. The medical record of each patient was reviewed.



The most commonly associated risk factor was trauma (44%). Fungal keratitis developed in five patients using extended wear contact lenses and one patient wearing a therapeutic bandage contact lens. Clinical features included irregular, feathery margins (62%), a dry, rough texture (47%), and satellite lesions (41%). An initial positive culture was obtained in 90% of patients, with a majority of cultures becoming positive within 48 hours. The Fusarium sp accounted for 62% of the isolates, with Fusarium oxysporum being the most commonly isolated organism. New fungal isolates include Candida parapsilosis, Aspergillus terreus, Candida tropicalis, and Trichosporon beigellii. Natamycin 5% suspension was the initial antifungal agent used for 91% of the patients, with an average duration of treatment of 38 days. Twenty-five patients were treated with oral ketoconazole for a median duration of 2 weeks, in addition to topical antifungal therapy. Thirty-four patients (27%) required a penetrating keratoplasty. Six patients had recurrence of fungal keratitis after penetrating keratoplasty.



Trauma, including contact lens wear, is the most commonly associated risk factor. The fungal organisms can be readily identified in culture. F. oxysporum is the most common organism, with new isolates identified. The mainstay of therapy is topical natamycin with the increasing use of imidazoles.

title: Late Microbial Keratitis After Corneal Transplantation
abstract: Infectious keratitis after penetrating keratoplasty can be devastating to the survival of the graft and its visual outcome. From November 1989 to October 1994, we treated 41 episodes of late microbial keratitis among 354 consecutive penetrating keratoplasty patients and reviewed their medical records retrospectively. The time interval between the corneal transplantation and the onset of graft infection was averaged 10.4 ± 10.9 months (range: 1–52 months). The precipitating factors of keratitis included epithelial defect (49%), suture-related problems (41%), use of contact lenses (17%), trichiasis (17%), dry eye (12%), and lid abnormalities (10%). Gram-positive cocci and gram-negative bacilli were associated with 51 and 40%, respectively, of the infectious keratitis, with Streptococcus being the most common species. Despite fortified antibiotic treatments, major complications such as graft failure and wound dehiscence could result. The overall result was that clarity was retained in only 43% of our grafts. We conclude that to prevent infectious keratitis there is a need to implement appropriate preventive measures as well as close monitoring of the graft after operation.

title: Candida Parapsilosis Keratitis Following Treatment of Epithelial Ingrowth After Laser In Situ Keratomileusis
abstract: To present a case of Candida parapsilosis following laser in situ keratomileusis (LASIK).  Retrospective chart review.  A case report of a 51-year-old woman who underwent bilateral LASIK is presented. Two weeks after the procedure, the patient presented with epithelial ingrowth OD. The ingrowth was treated with flap lifting and scraping, followed by postoperative antibiotics. Four weeks later, the patient presented with numerous interface infiltrates. Smears were positive for yeast forms and cultures grew Candida parapsilosis. Administration of topical and systemic antifungal therapy resulted in clearing of the infection with partial visual recovery.  To our knowledge, this represents the first reported case of a post-LASIK Candida parapsilosis keratitis. A high degree of suspicion coupled with rapid and appropriate treatment can result in visual recovery.

title: Mycological and histological investigations in humans with middle ear infections *
abstract: The aim of our investigations was to characterize fungal colonization of the ear in immunocompetent patients. From 1993 to 2000, 128 patients supposed to suffer from otomycosis were included. Mycological examination conducted by direct microscopy and fungal cultures was performed on 139 specimens. Among these, 115 patients suffered from chronic otitis media with persisting tympanum perforation and otorrhea. A further 13 patients had clinical signs of an otitis externa only. Out of 139 samples, fungi were identified in the auditory canal (n = 54), on the tympanic membrane (n = 5), and in the middle ear (n = 5). Two‐thirds were as moulds and one‐third yeasts. The dominating species were Aspergillus niger and Candida parapsilosis. Samples from 15 patients supposed to have mastoiditis or cholesteatoma were examined histologically. Fungal hyphae were observed in the middle ear cavity and/or between horny lamellae of cholesteatoma in four patients. In the middle ear of immunocompetent patients chronic‐hyperplastic (polypoid) inflammation was detected with increased production of mucus, which probably promotes colonization by pathogenic fungi in the middle ear as well as in the auditory canal. In vasive fungal growth into the subepithelial connective tissue was not observed.

title: Fungal causes of otitis externa and tympanostomy tube otorrhea.
abstract: To describe the occurrence of fungal organisms in the setting of otitis externa and tympanostomy tube otorrhea, review the treatment course, timing of diagnosis, organism identified and time to resolution with fungal infections.



Retrospective review.



Pediatric otolaryngology clinic within a tertiary care hospital.



One hundred and sixty-six patients (ages 16 days to 18 years) with fungal organisms on ear culture.



Number of prior therapies, number of office visits, time to resolution and anti-fungal therapy.



Ear cultures positive for fungal organisms were found in 166 patients seen between 1 January 1996 and 30 September 2003 from a total of 1242 patients undergoing ear culture. Comparing the 3-year period (1996-1998) prior to the availability of fluoroquinolone ototopical drops to the 3-year period after (1999-2001), there is a statistically significant increase in the incidence of positive fungal culture (p<0.001). Otitis media was diagnosed in 72% of these children, with otitis externa comprising 25%. Approximately 3% carried a diagnosis of both otitis externa and otitis media. Candida albicans was identified in 43% of fungal organism-positive cultures. Candida parapsilosis was found in 24% of and Aspergillus fumigatus in 13%. The remainder of the cultures yielded three other Candida and three other Aspergillus species, each at less than 5%. Time to resolution ranged from 1 week to 9 months, with a median of 3.8 weeks for symptom resolution. Patients were treated with an average of 1.7 oral antibiotics and 1.1 ototopical agent before a culture was taken.



Otorrhea due to fungal organisms occurs in the setting of refractory infection and is often discovered after multiple oral and ototopical antibacterial medications. Due to the extended treatment period required to clear fungal organism, timely diagnosis with culture for bacteria and fungus is required in patients with persistent otorrhea. An increase in incidence of fungal infections of the ear was found in the period after widespread use of ofloxacin began.

title: [Mycotic infection of the ear: a harmless saprophyte or pathognomonic risk factor?].
abstract: The mycotic infection of the auditory canal and middle ear in patients with special risks (transplantation, extensive ear surgery, immunosuppression) and long term antibiotic treatment is discussed as a possible pathogenetic factor for inner ear lesions and generalization. This problem was also studied by experimental investigations with artificially infected guinea pigs. Further on we report about a clinical study with a new antimycotic substance which was used for local therapy.

title: Fungal otitis externa--its association with fluoroquinolone eardrops.
abstract: The value of ototopical antimicrobials in the treatment of otorrhea has been debated for  Several recent articles have suggested that fluoroquinolone eardrops have the correct antimicrobial profile to treat posttympanostomy tube otorrhea and otitis externa in  Studies showing a lack of ototoxicity and efficacy similar to oral amoxicillin-clavulanate led the Food and Drug Administration to approve ofloxacin for treatment of otitis externa and tube otorrhea in children. Several authors have advocated the use of fluoroquinolone drops as first-line therapy for these indications,3 …

title: The frequency of Candida parapsilosis in onychomycosis. An epidemiological survey in Israel
abstract: Candida albicans is regarded as the major pathogen in yeast‐induced onychomycosis. Based on our impression of an increasing prevalence of Candida parapsilosis in this disease, we examined the data of two mycology laboratories in the same geographic location, from 1994 to 1996 in one (centre A) and for 1995 (6 months) in the other (centre B). A total of 954 and 230 toenails and 621 and 190 fingernails, respectively, underwent KOH microscopy and culture studies in each centre. Positive findings were noted in 45 and 65% of the toenails and 44 and 72% of the fingernails, respectively. In the toenails, Candida spp. were found in 22 and 15%, respectively, and in the fingernails, in 77 and 63%, respectively. The most frequent Candida species was C. parapsilosis (39.5% in toenails, 36.7% in fingernails), followed by C. albicans (19.5% in toenails, and 34.4% in fingernails). These results demonstrate a higher frequency of isolation of C. parapsilosis compared with C. albicans in onychomycosis. This might have important therapeutic implications.

title: Onychomycosis in a Premature Infant Caused by Candida parapsilosis
abstract: We report onychomycosis caused by Candida parapsilosis in a 35‐day‐old premature infant. To our knowledge, this represents the youngest instance of an isolated lesion of the nail plate without involvement of glabrous skin, caused by C. parapsilosis in this age group to be reported in the literature.

title: Candida onychomycosis—an evaluation of the role of Candida species in nail disease
abstract: Nail infections caused by Candida species are normally associated with chronic paronychia or chronic mucocutaneous candidiasis (CMCC). However, the role of Candida in the pathogenesis of other primary nail dystrophies has been questioned in view of their response to antifungal therapy alone. In the present study of 86 patients with primary nail dystrophies from which Candida was isolated, three patterns of nail involvement were found. Nineteen patients, of whom 17 had CMCC, had total dystrophic onychomycosis of at least two nails. The second group consisted of 27 patients with paronychia and lateral onycholysis. In a further 40 patients, who did not have paronychia, Candida was isolated from nails showing primary distal and lateral onycholysis. These changes were mainly seen in patients with peripheral vascular disease, particularly Raynaud's disease, or Cushing's syndrome. Nail biopsies from patients in the latter two groups confirmed the presence of yeasts and mycelium in the nail plate and 17 (46%) of those receiving antifungal therapy with ketoconazole or itraconazole showed complete clearance of the nail dystrophy. Good responses to therapy were seen more frequently in patients with peripheral vascular disease or Cushing's syndrome of whom 15 (72%) recovered; distal erosion of the nail plate, mycelium in the nail plate on biopsy or direct microscopy together with the isolation of C. albicans were all associated with good responses to antifungals. In addition to patients with CMCC or paronychia, therefore, Candida appears to be a significant pathogen in some patients with primary onycholysis of the finger nails, particularly where there is underlying peripheral vascular disease or Cushing's syndrome.

title: Clinical and mycological diagnostic aspects of onychomycoses and dermatomycoses
abstract: The most common fungal disease of man is tinea pedis, and it has been estimated that 10-15% of the adult population in developed countries are affected at any one time. This prevalence may be even higher among at-risk groups such as swimmers and coal miners. There is no published data on the extent of onychomycoses in the general population but, extrapolating from the above figures and assuming that one-third of subjects will have some form of nail involvement, Evans' postulated that about 5% of the general population could have fungal disease. In view of the dimensions of these clinical conditions and because of their relatively poor response to treatment, it is important to summarize the natural history and diagnosis of nail and foot infections.

title: Causative agents of onychomycosis — a retrospective study
abstract: Dermatophytes, yeasts and moulds all are potential causative agents of onychomycosis.The aim of this study was to determine the percentage of cases of onychomycoses caused by each group. In addition, the responsible genus and species was identified for each nail infection.

title: Vaginal Candida parapsilosis: pathogen or bystander?
abstract: Candida parapsilosis is an infrequent isolate on vaginal cultures; its role as a vaginal pathogen remains unstudied. This retrospective study of women with positive culture for C. parapsilosis sought to characterize the significance of this finding and its response to antifungal therapy.



From February 2001 to August 2002, we identified all individuals with positive fungal isolates among a population of women with chronic vulvovaginal symptoms. Charts of women with C. parapsilosis cultures were reviewed with regard to patient demographics, clinical presentation and therapeutic response. Mycological cure, defined as a negative fungal culture at the next office visit, and clinical cure, i.e. symptom resolution, were determined for each subject.



A total of 582 women had positive vaginal cultures for 635 isolates, of which 54 (8.5%) were C. parapsilosis. The charts of 51 subjects with C. parapsilosis were available for review and follow-up cultures and clinical information were available for 39 (76.5%). Microscopy was positive in 9 (17.6%). Antifungal treatment resulted in mycological cure in 17/19 patients with fluconazole, 7/7 with butoconazole, 6/6 with boric acid, 1/1 with miconazole and occurred spontaneously in 6/7: 24/37 (64.9%) patients with a mycological cure experienced clinical cure.



Although C. parapsilosis is often a cause of vaginal symptoms, it seems to respond to a variety of antifungal agents and may even be a transient vaginal colonizer.

title: Antifungal Susceptibilities of Candida Species Causing Vulvovaginitis and Epidemiology of Recurrent Cases
abstract: There are limited data regarding the antifungal susceptibility of yeast causing vulvovaginal candidiasis, since cultures are rarely performed. Susceptibility testing was performed on vaginal yeast isolates collected from January 1998 to March 2001 from 429 patients with suspected vulvovaginal candidiasis. The charts of 84 patients with multiple positive cultures were reviewed. The 593 yeast isolates were Candida albicans (n = 420), Candida glabrata (n = 112), Candida parapsilosis (n = 30), Candida krusei (n = 12), Saccharomyces cerevisiae ( n = 9), Candida tropicalis (n = 8), Candida lusitaniae (n = 1), and Trichosporon sp. (n = 1). Multiple species suggesting mixed infection were isolated from 27 cultures. Resistance to fluconazole and flucytosine was observed infrequently (3.7% and 3.0%); 16.2% of isolates were resistant to itraconazole (MIC ≥ 1 μg/ml). The four imidazoles (econazole, clotrimazole, miconazole, and ketoconazole) were active: 94.3 to 98.5% were susceptible at ≤1 μg/ml. Among different species, elevated fluconazole MICs (≥16 μg/ml) were only observed in C. glabrata (15.2% resistant [R], 51.8% susceptible-dose dependent [S-DD]), C. parapsilosis (3.3% S-DD), S. cerevisiae (11.1% S-DD), and C. krusei (50% S-DD, 41.7% R, considered intrinsically fluconazole resistant). Resistance to itraconazole was observed among C. glabrata (74.1%), C. krusei (58.3%), S. cerevisiae (55.6%), and C. parapsilosis (3.4%). Among 84 patients with recurrent episodes, non-albicans species were more common (42% versus 20%). A ≥4-fold rise in fluconazole MIC was observed in only one patient with C. parapsilosis. These results support the use of azoles for empirical therapy of uncomplicated candidal vulvovaginitis. Recurrent episodes are more often caused by non-albicans species, for which azole agents are less likely to be effective.

title: Symptomatic vulvovaginitis due to fluconazole-resistant Candida albicans in a female who was not infected with human immunodeficiency virus.
abstract: pancreatitis: correlation between the severity of the disease and the antienterobacterial common antigen antibody titre. Gut 1984;25:1065-70. 4. Runkel NS, Rodriguez LF, Moody FG. Mechanisms of sepsis in acute pancreatitis in opossums.  1987;317:940-5. 6. Michie HR, Manogue KR, Spriggs DR, et al. Detection of circulating tumor

title: Chronic fungal vaginitis: the value of cultures.
abstract: Our purpose was to examine the importance of fungal cultures in evaluating patients with symptoms of chronic vaginitis by assessing the relative contribution of various yeast species and by comparing infections caused by Candida albicans with those caused by other species.



A prospective observational study of patients referred with chronic vaginal symptoms was undertaken. In addition to a standard evaluation of symptoms, cultures for yeast were performed on modified Sabouraud agar plates.



Seventy-seven isolates were obtained from 74 patients. A total of 68% were Candida albicans; 32% were other species. The clinical syndromes caused by non-Candida albicans isolates were indistinguishable from Candida albicans infections. Fluconazole gave a short-term mycologic cure in all Candida albicans but only 25% of non-Candida albicans cases (p < 0.001). In non-Candida albicans infections, boric acid suppositories achieved the best mycologic cure rate (85%).



Because non-Candida albicans species are responsible for a significant number of chronic fungal vaginal infections and are more resistant to therapy with fluconazole, fungal cultures are a valuable aid in confirming the diagnosis and selecting appropriate therapy.

title: Prevalence of vulvovaginal candidiasis and susceptibility to fluconazole in women.
abstract: The purpose of this study was to determine the prevalence of vaginal colonization by Candida with the use of a rapid detection method, to examine the determinants of vaginal candidiasis, and to evaluate susceptibility for fluconazole.



Vaginal swabs were collected from unselected women at the outpatient clinic of the Department of Obstetrics and Gynecology. A differentiation was made between patients with a positive and a negative potassium hydroxide examination.



Six hundred twelve women were recruited, of whom 39 women (6.3%) had clinical candidiasis. The overall rate of yeast colonization was 20.1%. Candida albicans was isolated most frequently(68.3%), followed by C glabrata (16.3%) and C parapsilosis (8.9%). Clinical candidiasis was related positively with the state of estrogen impregnation. In vitro susceptibility testing by the National Committee for Clinical Laboratory Standards method for fluconazole revealed that 21.1% of the isolates were resistant.



More than 20% of the unselected women were colonized with Candida species. Hyperestrogenemia was associated with an increased vulvovaginal colonization by Candida. Surprisingly, 21% of the isolates was resistant to fluconazole, according to the National Committee for Clinical Laboratory Standards method.

title: Vulvovaginal candidiasis in female sex workers
abstract: Vulvovaginal candidiasis is a frequent inflammatory process in women but it has not been widely studied in female sex workers FSWs . To estimate the frequency of Candida species infection in FSWs and to identify related risk factors and clinical findings, we carried out a retrospective study of 1923 FSWs over 11 years. We also performed a prospective study of 163 consecutive FSWs with a history of candidiasis during a 4 year period. Candida species were isolated in 1967 samples 18.5 of the total . Candida albicans 89.3 was the most frequent species, followed by Candida glabrata 2.7 , Candida parapsilosis 1.2 and Saccharomyces cerevisiae 0.4 . In the prospective study of 163 patients, we found vaginal discharge in 76.1 of cases, soreness in 52.1 and vulval pruritus in 32.5 . We identified 12 patients 7.4 with recurrent vulvovaginal candidiasis. No statistical difference was found between recurrent vulvovaginitis and the use of oral contraceptives, oral sex, tight fitting clothing and synthetic underwear. FSWs have the same prevalence of candidiasis as other groups of women described in published literature. The proportion of albicans and non albicans species does not differ between women with recurrent and non recurrent vulvovaginal candidiasis VVC .

title: Identification of Candida strains isolated from Tanzanian pregnant women with vaginal candidiasis.
abstract: To identify Candida strains isolated from Tanzanian women (13 to 45 years) with vaginal candidiasis.



A cross-sectional study.



Antenatal clinic in Ilala district hospital in Dar es Salaam, Tanzania from March 1998 to December 2000.



The identities of the 272 isolates tested with API Candida were: Candida albicans 180(66.2%), Candida tropicalis 13(4.7%), Candida glabrata 20(7.35%), Candida famata 6(2.2%), Candida parapsilosis 6 (2.2%) and Candida lusitaniae one (0.37%). API Candida could not speciate 43 (15.8%) isolates of these; two (0.7%) fell between C. albicans and C. tropicalis, 17(6.25%) C. lusitaniael, C. guilliermondii/C. famata, 14(5.15%) C. krusei, C. inconspicua, and C. norvegensis and nine (3.3%) either C. parapsilosis, C. krusei, C. incospicua or Geotrichum spp. Four (1.5%) isolates had an assimilation pattern of Trichosporo spp, but were all germ tube positive and had morphological features on cornemeal agar that were consistent with C. albicans. API 20C AUX was used for testing 29 isolates and results showed: 11/29 (37.9%) C. albicans, 1/29 (3.4%) C. tropicalis, 4/29 (13.8%) C. glabrata, 1/29 (3.4%) C. parapsilosis, 1/29 (3.4%) C. famata, 1/29 (3.4%) C. lusitaniae, 1/29 (3.4%) C. colliculosa/C. magnoliae, 5/29(17.2%) C. albicans/ C. tropicals 2/29 (6.8%) C. norvegensis/C. parapsilosis, and 2/29(6.8%) C. kruseil/C. inconspicua. Results of 20 isolates identified by Randomly Amplified Polymorphic DNA (RAPID) technique showed a 95% agreement with API Candida and a 100% agreement with API 20C AUX.



Although most (66.3%) of the species isolated from Dar es Salaam women with vaginal candidiasis were C. albicans, a considerable percentage (33.7%) were non-albicans, mainly C. glabrata, C. krusei and C. tropicalis. The high prevalence of non-albicans Candida spp observed in this population may have therapeutic implications.

title: Isolation, acid proteinase secretion, and experimental pathogenicity of Candida parapsilosis from outpatients with vaginitis
abstract: Candida parapsilosis was isolated from the vaginas of several nonpregnant, nondiabetic, mostly premenopausal outpatients who presented the characteristic signs and symptoms of a frank vulvovaginal candidiasis (heavy discharge with cottage cheese appearance and intense itching, with or without vulvar erythema and dyspareunia). All isolates conformed morphologically, biochemically, and serologically to the standard description of the species. They showed high acid proteinase-secretory activity in vitro and were appreciably pathogenic for cyclophosphamide-immunodepressed mice. Some isolates were also tested for their vaginopathic potential in ovariectomized rats under estradiol administration. In all cases, the rat vagina was colonized by C. parapsilosis to an extent and duration not different from those caused by a vaginopathic isolate of Candida albicans. Periodic acid-Schiff-stained vaginal smears taken at intervals during rat experimental infection showed C. parapsilosis yeasts adhering to exfoliated epithelial cells of rat vagina. Overall, these results emphasize the proteolytic and pathogenic potential of C. parapsilosis and suggest that this fungus may be an agent of clinical vaginitis.

title: Experimental rat vaginal infection with Candida parapsilosis.
abstract: The experimental vaginopathic potential of Candida parapsilosis was determined in ovariectomized rats maintained under pseudoestrus by estrogen administrations. Of the 3 strains of C. parapsilosis tested, that isolated from the vagina of a woman affected by vulvovaginal candidosis gave a prolonged and sustained experimental vaginitis, not different in extent and duration from that caused by a vaginal isolate of C. albicans from a vaginitis patient. The other two isolates of C. parapsilosis (one from the vagina of an asymptomatic subject and another from soil) were unable to infect rat vagina. Microscopic observations of PAS-stained vaginal smears from rats infected with the vaginopathic isolate of C. parapsilosis showed pronounced adherence of yeasts to exfoliated cells. In addition, this isolate of C. parapsilosis produced an elevated quantity of acid proteinase in vitro.

title: Candiduria in critically ill patients admitted to intensive care medical units
abstract: BackgroundThe purpose of this study was to determine the incidence of candiduria in critically ill patients admitted to intensive care medical units (ICUs), to identify risk factors for candiduria and to assess the frequency distribution of different Candida spp.Subjects and methodsThis was a prospective cohort observational and multicenter study. A total of 1,765 patients older than 18 years of age who were admitted for at least 7 days to 73 medical-surgical ICUs of 70 Spanish hospitals were included in the study. Urine cultures were performed once a week.ResultsIn 389 patients (22%), Candida spp. in one or more urine samples were isolated. In the multivariate analysis, independent risk factors for candiduria included: age >65 years, female sex, length of hospital stay before ICU admission, diabetes mellitus, total parenteral nutrition, mechanical ventilation and previous use of antimicrobials. Candida albicans was recovered in 266 cases (68.4%), followed by C. glabrata (32 cases, 8.2%) and C. tropicalis (14 cases, 36%). Previous use of antifungal agents was the only risk factor for the selection of Candida non-albicans candiduria (OR 2.64, 95% CI 1.35–5.14, P =0.004). In-hospital mortality was 48.8% in patients with candiduria compared to 36.6% in those without candiduria ( P <0.001). Significant differences were also found for ICU mortality (38.% vs. 28.1%, P <0.001).ConclusionsTwenty-two percent of critically ill patients admitted for more than 7 days in the ICU developed candiduria. C. albicans was the most frequent causative pathogen. Previous use of antifungals was the only risk factor for the selection of Candida non-albicans.

title: Candiduria in a public hospital of São Paulo (1999-2004): characteristics of the yeast isolates.
abstract: The study involved 100 yeast isolates, obtained from urine samples provided by a Public Pediatric Hospital of São Paulo, Brazil, from 1999 to 2004. The most frequent species was Candida albicans, followed by C. tropicalis, C. glabrata and C. parapsilosis. In regard to virulence, 97% of the isolates showed index 3 for proteinase and 63% index 2 for phospholipase. The most frequent killer biotypes were 511 and 888.

title: Correlation between candiduria and departmental antibiotic use.
abstract: The incidence of candiduria is increasing in teaching hospitals. We examined the hypothesis that this trend was correlated with the amount of departmental antibiotic consumption. In the setting of a large teaching hospital in Israel, the correlation coefficient between departmental intravenous antibiotic consumption (expressed as daily defined dose (DDD)/1000 patient-days) and the incidence of candiduria per 1000 patient-days was 0.47 (P=0.03). For broad-spectrum antibiotics, the corresponding correlation coefficient was 0.66 (P=0.001). The strongest correlation with candiduria was shown for the use of meropenem (r=0.79, P<0.001) and ceftazidime (r=0.66, P=0.001). This is the first time that departmental habits of antibiotic use have been shown to be strongly correlated with the incidence of candiduria in hospitalized patients. These results add an important new dimension to the strategy of restricting broad-spectrum antibiotics.

title: Candiduria in Hospital Patients: A Study Prospective
abstract: The presence of Candida species in the urine is frequent among hospitalized patients. We studied sample urine of 205 hospitalized patients during a 1-year period to determine the incidence of nosocomial candiduria. The yeasts were isolated in 22% (45/205) urine cultures and risk factors in these patients were analyzed. Candida albicans was isolated in 35.6% and C. tropicalis (22%) was the second most frequent species isolated. Most patients were women (57.8%) with a mean age of 48.7 years. The principal risk factors that were observed in patients with candiduria included antibiotics therapy (100%), urinary catheterization (84.4%), surgical procedure (66.7%), female sex and extended hospitalization. The efficacy of fluconazole therapy to eradicate Candida from urine was demonstrated (p = 0.05). Of the 23 individuals who received antifungal therapy, candiduria persisted in 9 (39.2%) and of 22 patients who received no antifungal therapy, the candiduria persisted in 15 (68.2%).

title: Investigation of the sequence of colonization and candidemia in nonneutropenic patients
abstract: Among neutropenic patients with hematologic malignancies, candidemia has been shown to arise typically from autoinfection after colonization. In patients without neutropenia, we examined the similarities of strains colonizing or infecting various body sites and those subsequently causing Candida bloodstream infections. Strain similarity was examined by karyotyping and restriction endonuclease analysis of genomic DNA (REAG) by using two restriction enzymes (SfiI and BssHII). The banding patterns of 42 isolates from 19 patients were independently evaluated in a blinded fashion by three observers. The interobserver reliability measured with a generalized kappa statistic was 0.59 for karyotyping, 0.84 for REAG with SfiI, and 0.88 for REAG with BssHII (P < 0.001 for each). REAG classified the initial colonizing or infecting isolate and subsequent blood isolates as identical in 16 patients (84%). The mean duration of colonization or infection prior to a positive blood culture was 5 and 23 days in patients infected with related and unrelated isolates, respectively (P = 0.14; 95% confidence interval = -14.5 to 50.5). Karyotyping results matched the REAG results for isolates from 14 of the 19 patients (74%). In patients infected with identical isolates, the initial isolate was most frequently recovered from the urine (n = 5) or vascular catheter tips (n = 4). In the five subjects with organisms showing disparate results between the methods, karyotyping revealed different banding patterns, whereas REAG suggested that the isolates were identical. Candida colonization or infection with an identical strain frequently precedes bloodstream infection in nonneutropenic patients. Future studies should evaluate whether patients at high risk for candidemia and who have vascular catheter or urine samples that are positive for a Candida on culture should be treated empirically.

title: Incidence and significance of candiduria.
abstract: The 100,000 colony count has been used without careful evaluation for the diagnosis of renal candidiasis. Therefore, a prospective study was done on 1,004 urine samples from patients without signs of candidiasis to determine the incidence of candiduria and Candida colony counts. These were compared with colony counts from histologically proved cases of renal candidiasis. The incidence of candiduria varied from a low of 4% in men to a high of 39% in girls during their second week of antibiotic therapy. The overall mean colony count was 1,292 +/- 1,500. The mean colony count in clean-catch urine specimens from six proved cases of renal candidiasis was 23,750 +/- 12,311. The difference in colony counts from proved and unproved cases is statistically significant. Counts greater than 10,000 Candida organisms per milliliter require further investigation.

title: Adhesion of Candida parapsilosis to epithelial and acrylic surfaces correlates with cell surface hydrophobicity
abstract: We investigated in vitro adherence of 24 isolates of Candida parapsilosis and 12 isolates of Candida albicans with regard to their relative cell‐surface hydrophobicity (CSH), adherence to human buccal epithelial cells (BEC) and acrylic surfaces. There was no significant interspecies difference in the relative adherence of C. parapsilosis and C. albicans isolates to BEC, although the former demonstrated a tendency for increased adherence. However, a significant intra‐species variation in adherence among isolates of C. parapsilosis (P < 0.0001) to BEC, but not of C. albicans was noted. The superficial isolates of C. parapsilosis demonstrated a higher avidity (33%) to BEC than the systemic isolates. On regression analysis a significant positive correlation between C. parapsilosis adherence to BEC and denture acrylic surfaces was noted (r = 0.45, P = 0.02). Similarly, buccal cell adherence correlated strongly with CSH of C. parapsilosis (r = 0.63, P = 0.0008). These results shed further light on the intimate relationship between adherence and CSH in candidal colonization and imply that both C. parapsilosis and C. albicans are equally potent in colonizing mucosal surfaces with respect to the attributes investigated.

title: Genotypic variation and slime production among blood and catheter isolates of Candida parapsilosis
abstract: Candida parapsilosis is an important nosocomial pathogen that can proliferate in high concentrations of glucose and form biofilms on prosthetic materials. We investigated the genotypic diversity and slime production among 31 isolates of C. parapsilosis from individual patients with bloodstream or catheter infections. DNA subtyping was performed by using electrophoretic karyotyping plus restriction endonuclease analysis with BssHII followed by pulsed-field gel electrophoresis. Slime production was evaluated by growing organisms in Sabouraud broth with 8% glucose and examining the walls of the tubes for the presence of an adherent slime layer. Overall there were 14 DNA subtypes among the 31 isolates. Eighty percent of the isolates produced slime; 67% of the isolates were moderately to strongly positive, 13% were weakly positive, and 20% were not slime producers. The ability of isolates of a given DNA type to produce slime under these conditions was variable. The results of these studies indicate moderate genotypic variation among clinical isolates of C. parapsilosis. The propensity of these isolates to form slime in glucose-containing solutions suggests that this phenotypic characteristic may contribute to the ability of C. parapsilosis to adhere to plastic catheters and cause infections.

title: A comparison of phospholipase activity, cellular adherence and pathogenicity of yeasts.
abstract: Phospholipase A and lysophospholipase activities were measured in the culture fluid and in the blastospores of Candida albicans. When phospholipase activity was measured in six yeasts (four strains of C. albicans and a single strain each of Candida parapsilosis and Saccharomyces cerevisiae) a correlation was found between this activity and two potential parameters of pathogenicity. The C. albicans isolates which adhered most strongly to buccal epithelial cells and were most pathogenic in mice had the highest phospholipase activities. Non-pathogenic yeasts, including C. albicans isolates which did not adhere and did not kill mice, had lower phospholipase activities.

title: Adherence of Candida albicans and other Candida species to mucosal epithelial cells
abstract: To study the possible involvement of candidal adherence in mucosal colonization, we examined the in vitro adherence capabilities of seven Candida species. Adherence was evaluated by direct microscopic examination and by a quantitative radiometric adherence test. The results indicate that C. albicans adheres to vaginal and buccal epithelial cells to a significantly greater degree (P less than 0.01) than the other species tested. C. tropicalis and C. stellatoidea demonstrated moderate adherence capabilities, while C. parapsilosis adhered only to a slight degree. Other species failed to interact with isolated mucosal cells. These findings suggest that there is a relationship between the adherence capabilities of the Candida species and their abilities to colonize mucosal surfaces, since those species which adhere are those which most frequently colonize mucosal surfaces. C. albicans was found to be adherent under a variety of environmental conditions. Stationary-phase blastospores of C. albicans were found to be more adherent than logarithmic-phase yeasts, and larger blastospore cell-to-epithelial cell ratios resulted in greater adherence values. The actual number of adherent yeasts varied considerably when epithelial cells were obtained from different donors.

title: Biotype diversity of Candida parapsilosis and its relationship to the clinical source and experimental pathogenicity.
abstract: Environmental, vaginal, and blood isolates of Candida parapsilosis were biotyped by karyotype analysis in pulsed-field gel electrophoresis. Morphotype and resistotype were also determined as was aspartyl proteinase secretion and pathogenicity in a systemic mouse infection model. Overall, the karyotype patterns consisted of 6-9 chromosome bands (> 3.0-0.6 Mb) with limited clustering, since most isolates had unique chromosome profiles. Major clusters C. parapsilosis, differing by source of isolation and in experimental pathogenicity, could be discriminated by morphoresistotyping. The morphotypes of isolates from subjects with candidemia ranged from one that caused elevated mortality in the normal mouse to those that were totally avirulent in the neutropenic animal. Among clinical isolates, secretion of aspartyl proteinase was higher in vaginitis than in candidemia isolates and did not correlate with the experimental pathogenicity. These results emphasize the biotype diversity of C. parapsilosis and have potentially important epidemiologic and pathologic implications.

title: Evidence that members of the secretory aspartyl proteinase gene family, in particular SAP2, are virulence factors for Candida vaginitis.
abstract: Virulence of Candida albicans strains with targeted disruption of secretory aspartyl proteinase genes (SAP1 to SAP6) was assessed in an estrogen-dependent rat vaginitis model. Null sap1 to sap3 but not sap4 to sap6 mutants lost most of the virulence of their parental strain SC5314. In particular, the sap2 mutant was almost avirulent in this model. Reinsertion of the SAP2 gene into this latter mutant led to the to recovery of the vaginopathic potential. The vaginal fluids of the animals infected by the wild type strain or by the sap1 or sap3 mutants expressed a pepstatin-sensitive proteinase activity in vitro. No traces of this activity were found in the vaginal fluid of rats challenged by the sap2 mutant. All strains were capable of developing true hyphae during infection. Thus, members of SAP family, in particular SAP2, play a clear pathogenic role in vaginitis and may constitute a novel target for chemoimmunotherapy of this infection.

title: Antifungal Susceptibility of Candida Biofilms: Unique Efficacy of Amphotericin B Lipid Formulations and Echinocandins
abstract: Biofilms, likely the predominant mode of device-related microbial infection, exhibit resistance to antimicrobial agents. Evidence suggests that Candida biofilms have dramatically reduced susceptibility to antifungal drugs. We examined antifungal susceptibilities of Candida albicans and Candida parapsilosis biofilms grown on a bioprosthetic model. In addition to conventional agents, we determined if new antifungal agents (triazoles, amphotericin B lipid formulations, and echinocandins) have activities against Candida biofilms. We also explored effects of preincubation of C. albicans cells with subinhibitory concentrations (sub-MICs) of drugs to see if they could modify subsequent biofilm formation. Finally, we used confocal scanning laser microscopy (CSLM) to image planktonic- and biofilm-exposed blastospores to examine drug effects on cell structure. Candida biofilms were formed on silicone elastomer and quantified by tetrazolium and dry weight (DW) assays. Susceptibility testing of fluconazole, nystatin, chlorhexidine, terbenafine, amphotericin B (AMB), and the triazoles voriconazole (VRC) and ravuconazole revealed resistance in all Candida isolates examined when grown as biofilms, compared to planktonic forms. In contrast, lipid formulations of AMB (liposomal AMB and AMB lipid complex [ABLC]) and echinocandins (caspofungin [Casp] and micafungin) showed activity against Candida biofilms. Preincubation of C. albicans cells with sub-MIC levels of antifungals decreased the ability of cells to subsequently form biofilm (measured by DW; P < 0.0005). CSLM analysis of planktonic and biofilm-associated blastospores showed treatment with VRC, Casp, and ABLC resulted in morphological alterations, which differed with each agent. In conclusion, our data show that Candida biofilms show unique susceptibilities to echinocandins and AMB lipid formulations.

title: Role of dimorphism in the development of Candida albicans biofilms.
abstract: Two model biofilm systems, involving growth on disks of catheter material or on cylindrical cellulose filters, were used to investigate the structure of Candida albicans biofilms. To assess the importance of dimorphism in biofilm development, biofilms produced by two wild-type strains were compared with those formed by two morphological mutants, incapable of yeast and hyphal growth, respectively. Scanning electron microscopy and thin sections of biofilms examined by light microscopy revealed that biofilms of the wild-type strains formed on catheter disks consisted of two distinct layers: a thin, basal yeast layer and a thicker, but more open, hyphal layer. The hypha- mutant produced only the basal layer, whereas the yeast- mutant formed a thicker, hyphal biofilm equivalent to the outer zone of the wild-type structures. Biofilms of the yeast- mutant were more easily detached from the catheter surface than the others, suggesting that the basal yeast layer has an important role in anchoring the biofilm to the surface. Biofilms formed on cylindrical cellulose filters were quite different in appearance. The hypha- mutant and both wild types produced exclusively yeast-form biofilms whereas the yeast- mutant generated a dense hyphal mat on the top of the filter. All these biofilms, irrespective of morphological form, were resistant to the antifungal agent, amphotericin B. Overall, these results indicate that the structure of a C. albicans biofilm depends on the nature of the contact surface, but that some surfaces produce biofilms with a layered architecture resembling to that described for bacterial systems.

title: Biofilm formation by Candida species on the surface of catheter materials in vitro
abstract: A model system for studying Candida biofilms growing on the surface of small discs of catheter material is described. Biofilm formation was determined quantitatively by a colorimetric assay involving reduction of a tetrazolium salt or by [3H]leucine incorporation; both methods gave excellent correlation with biofilm dry weight (r = 0.997 and 0.945, respectively). Growth of Candida albicans biofilms in medium containing 500 mM galactose or 50 mM glucose reached a maximum after 48 h and then declined; however, the cell yield was lower in low-glucose medium. Comparison of biofilm formation by 15 different isolates of C. albicans failed to reveal any correlation with pathogenicity within this group, but there was some correlation with pathogenicity when different Candida species were tested. Isolates of C. parapsilosis (Glasgow), C. pseudotropicalis, and C. glabrata all gave significantly less biofilm growth (P < 0.001) than the more pathogenic C. albicans. Evaluation of various catheter materials showed that biofilm formation by C. albicans was slightly increased on latex or silicone elastomer (P < 0.05), compared with polyvinyl chloride, but substantially decreased on polyurethane or 100% silicone (P < 0.001). Scanning electron microscopy demonstrated that after 48 h, C. albicans biofilms consisted of a dense network of yeasts, germ tubes, pseudohyphae, and hyphae; extracellular polymeric material was visible on the surfaces of some of these morphological forms. Our model system is a simple and convenient method for studying Candida biofilms and could be used for testing the efficacy of antifungal agents against biofilm cells.

title: Comparison of Biofilms Formed by Candidaalbicans and Candidaparapsilosis on Bioprosthetic Surfaces
abstract: Little is known about fungal biofilms, which may cause infection and antibiotic resistance. In this study, biofilm formation by different Candida species, particularly Candidaalbicans and C. parapsilosis, was evaluated by using a clinically relevant model of Candida biofilm on medical devices. Candida biofilms were allowed to form on silicone elastomer and were quantified by tetrazolium (XTT) and dry weight (DW) assays. Formed biofilm was visualized by using fluorescence microscopy and confocal scanning laser microscopy with Calcofluor White (Sigma Chemical Co., St. Louis, Mo.), concanavalin A-Alexafluor 488 (Molecular Probes, Eugene, Oreg.), and FUN-1 (Molecular Probes) dyes. Although minimal variations in biofilm production among invasive C. albicans isolates were seen, significant differences between invasive and noninvasive isolates (P < 0.001) were noted. C. albicans isolates produced more biofilm than C. parapsilosis, C. glabrata, and C. tropicalis isolates, as determined by DW assays (P was <0.001 for all comparisons) and microscopy. Interestingly, noninvasive isolates demonstrated a higher level of XTT activity than invasive isolates. On microscopy, C. albicans biofilms had a morphology different from that of other species, consisting of a basal blastospore layer with a dense overlying matrix composed of exopolysaccharides and hyphae. In contrast, C. parapsilosis biofilms had less volume than C. albicans biofilms and were comprised exclusively of clumped blastospores. Unlike planktonically grown cells, Candida biofilms rapidly (within 6 h) developed fluconazole resistance (MIC, >128 μg/ml). Importantly, XTT and FUN-1 activity showed biofilm cells to be metabolically active. In conclusion, our data show that C. albicans produces quantitatively larger and qualitatively more complex biofilms than other species, in particular, C. parapsilosis.

title: Candida biofilms on implanted biomaterials: a clinically significant problem.
abstract: In recent years there has been an increasing appreciation that microbial biofilms are ubiquitous, which has resulted in a number of studies on infectious diseases from a biofilm perspective. Biofilms are defined as structured microbial communities that are attached to a surface and encased in a matrix of exopolymeric material. A wide range of biomaterials used in clinical practice have been shown to support colonization and biofilm formation by Candida spp., and the increase in Candida infections in the last decades has almost paralleled the increase and widespread use of a broad range of medical implant devices, mainly in populations with impaired host defenses. Formation of Candida biofilms has important clinical repercussions because of their increased resistance to antifungal therapy and the ability of cells within biofilms to withstand host immune defenses. Further recognition and understanding of the role of Candida biofilms in human infection should help in the clinical management of these recalcitrant infections.

title: Biofilm Production by Candida Species and Inadequate Antifungal Therapy as Predictors of Mortality for Patients with Candidemia
abstract: Nosocomial Candida bloodstream infections rank among infections with highest mortality rates. A retrospective cohort analysis was conducted at Catholic University Hospital to estimate the risk factors for mortality of patients with candidemia. We reviewed records for patients with a Candida bloodstream infection over a 5-year period (January 2000 through December 2004). Two hundred ninety-four patients (42.1% male; mean age ± standard deviation, 65 ± 12 years) were studied. Patients most commonly were admitted with a surgical diagnosis (162 patients [55.1%]), had a central venous catheter (213 [72.4%]), cancer (118 [40.1%]), or diabetes (58 [19.7%]). One hundred fifty-four (52.3%) patients died within 30 days. Of 294 patients, 168 (57.1%) were infected by Candida albicans, 64 (21.7%) by Candida parapsilosis, 28 (9.5%) by Candida tropicalis, and 26 (8.8%) by Candida glabrata. When fungal isolates were tested for biofilm formation capacity, biofilm production was most commonly observed for isolates of C. tropicalis (20 of 28 patients [71.4%]), followed by C. glabrata (6 of 26 [23.1%]), C. albicans (38 of 168 [22.6%]), and C. parapsilosis (14 of 64 [21.8%]). Multivariable analysis identified inadequate antifungal therapy (odds ratio [OR], 2.35; 95% confidence interval [95% CI], 1.09 to 5.10; P = 0.03), infection with overall biofilm-forming Candida species (OR, 2.33; 95% CI, 1.26 to 4.30; P = 0.007), and Acute Physiology and Chronic Health Evaluation III scores (OR, 1.03; 95% CI, 1.01 to 1.15; P < 0.001) as independent predictors of mortality. Notably, if mortality was analyzed according to the different biofilm-forming Candida species studied, only infections caused by C. albicans (P < 0.001) and C. parapsilosis (P = 0.003) correlated with increased mortality. Together with well-established factors, Candida biofilm production was therefore shown to be associated with greater mortality of patients with candidemia, probably by preventing complete organism eradication from the blood.

title: Biofilm Production by Isolates of Candida Species Recovered from Nonneutropenic Patients: Comparison of Bloodstream Isolates with Isolates from Other Sources
abstract: Biofilm production has been implicated as a potential virulence factor of some Candida species responsible for catheter-related fungemia in patients receiving parenteral nutrition. We therefore compared clinical bloodstream isolates representing seven different Candida species to each other and to those from other anatomical sites for the capacity to form biofilms in glucose-containing medium. Potential associations between the capacity to form biofilms and the clinical characteristics of fungemia were also analyzed. Isolates included the following from nonneutropenic patients: 101 bloodstream isolates (35 C. parapsilosis, 30 C. albicans, 18 C. tropicalis, 8 C. glabrata, and 10 other Candida species isolates) and 259 clinical isolates from other body sites (116 C. albicans, 53 C. glabrata, 43 C. tropicalis, 17 C. parapsilosis, and 30 other Candida species isolates). Organisms were grown in Sabouraud dextrose broth (SDB) containing a final concentration of 8% glucose to induce biofilm formation, as published previously. Biofilm production was determined by both visual and spectrophotometric methods. In this medium, biofilm production by C. albicans isolates was significantly less frequent (8%) than that by non-C. albicans Candida species (61%; P < 0.0001). The overall proportion of non-C. albicans Candida species isolates from the blood that produced biofilms was significantly higher than that of non-C. albicans Candida isolates obtained from other sites (79% versus 52%; P = 0.0001). Bloodstream isolates of C. parapsilosis alone were significantly more likely to be biofilm positive than were C. parapsilosis isolates from other sites (86% versus 47%; P = 0.0032). Non-C. albicans Candida species, including C. parapsilosis, were more likely to be biofilm positive if isolates were derived from patients whose candidemia was central venous catheter (CVC) related (95%; P < 0.0001) and was associated with the use of total parenteral nutrition (TPN) (94%; P < 0.005). These data suggest that the capacity of Candida species isolates to produce biofilms in vitro in glucose-containing SDB may be a reflection of the pathogenic potential of these isolates to cause CVC-related fungemia in patients receiving TPN.

title: Importance of biofilm inCandida parapsilosis and evaluation of its susceptibility to antifungal agents by colorimetric method
abstract: The ability ofC. parapsilosis (an important cause of nosocomial infections) to produce biofilm was evaluated in 32 bloodstream isolates and 85 strains isolated from skin. The biofilm formation was found in 19 (59%) blood isolates and only in 33 (39%) isolates from skin. The antifungal susceptibility was assessed for amphotericin B, itraconazole and voriconazole in planktonic and biofilm form of the 19 biofilm-positive bloodstream strains by broth microdilution method according toNCCLS standards. The method was modified by the use of resazurin as a colorimetric indicator of the metabolically active cells which makes the determination of the effect of antifungal agents easier. Biofilm forms of all strains were more resistant than their planktonic form.

title: Targeted gene deletion in Candida parapsilosis demonstrates the role of secreted lipase in virulence.
abstract: Candida parapsilosis is a major cause of human disease, yet little is known about the pathogen's virulence. We have developed an efficient gene deletion system for C. parapsilosis based on the repeated use of the dominant nourseothricin resistance marker (caSAT1) and its subsequent deletion by FLP-mediated, site-specific recombination. Using this technique, we deleted the lipase locus in the C. parapsilosis genome consisting of adjacent genes CpLIP1 and CpLIP2. Additionally we reconstructed the CpLIP2 gene, which restored lipase activity. Lipolytic activity was absent in the null mutants, whereas the WT, heterozygous, and reconstructed mutants showed similar lipase production. Biofilm formation was inhibited with lipase-negative mutants and their growth was significantly reduced in lipid-rich media. The knockout mutants were more efficiently ingested and killed by J774.16 and RAW 264.7 macrophage-like cells. Additionally, the lipase-negative mutants were significantly less virulent in infection models that involve inoculation of reconstituted human oral epithelium or murine intraperitoneal challenge. These studies represent what we believe to be the first targeted disruption of a gene in C. parapsilosis and show that C. parapsilosis-secreted lipase is involved in disease pathogenesis. This efficient system for targeted gene deletion holds great promise for rapidly enhancing our knowledge of the biology and virulence of this increasingly common invasive fungal pathogen.

title: Development of a Gene Knockout System in Candida parapsilosis Reveals a Conserved Role for BCR1 in Biofilm Formation
abstract: Candida parapsilosis is an important cause of candidiasis, yet few molecular tools are available. We adapted a recyclable nourseothricin resistance marker gene (SAT1) originally developed for use with C. albicans in order to generate gene knockouts from C. parapsilosis. We first replaced the promoters driving expression of the FLP recombinase and the SAT1 genes with the equivalent sequences from C. parapsilosis. We then used the cassette to generate a homozygous knockout of C. parapsilosis URA3. The ura3 knockouts have altered colony morphologies. We also knocked out both alleles of an ortholog of BCR1, a gene encoding a transcription factor known to be required for biofilm development in C. albicans. We show that C. parapsilosis BCR1 is necessary for biofilm development in C. parapsilosis and for expression of the cell wall protein encoded by RBT1. Our results suggest that there are significant similarities in the regulation of biofilms between the two species, despite the fact that C. parapsilosis does not generate true hyphae and that BCR1 regulates the expression of many hypha-specific adhesins in C. albicans.

title: Differential Activities of Newer Antifungal Agents against Candida albicans and Candida parapsilosis Biofilms
abstract: The activities of voriconazole, posaconazole, caspofungin, and anidulafungin against Candida albicans and Candida parapsilosis biofilms were evaluated. In contrast to planktonic cells, the MICs for voriconazole and posaconazole for the biofilms of the two species were high (≥256 and >64 mg/liter, respectively) but relatively low for the echinocandins caspofungin and anidulafungin (≤1 and ≤2 mg/liter, respectively).

title: Anti-metabolic activity of caspofungin against Candida albicans and Candida parapsilosis biofilms.
abstract: Candidiasis can be associated with the formation of biofilms on bioprosthetic surfaces and the intrinsic resistance of Candida albicans biofilms to the most commonly used antifungal agents has been demonstrated. In this study, we report on the antifungal activity of caspofungin at two different concentrations, on C. albicans and Candida parapsilosis biofilms with different ages of maturation.



Fifteen strains of C. albicans (10 strains susceptible to fluconazole in vitro and five strains resistant to this antifungal agent) and six strains of C. parapsilosis (all were susceptible to fluconazole in vitro) were studied. The antifungal activity of caspofungin was assessed by looking for a significant inhibition of the metabolic activity of yeasts within biofilms. Biofilms of Candida were produced in vitro, on silicone catheters.



Caspofungin used at MIC did not modify the metabolic activity of C. albicans, whatever the maturation age of the biofilms. The same concentration of caspofungin significantly reduced the metabolism (P<or=0.001) of 25% (biofilms of 48 h) to 50% (biofilms of 2 h) of the C. parapsilosis yeasts. The use of a therapeutic concentration of caspofungin (2 mg/L) significantly decreased (P<or=0.001) the metabolism of all the strains of C. albicans and C. parapsilosis tested, independently of the biofilm maturation age. This potent antifungal activity of caspofungin on C. albicans biofilms was observed independently of the yeast susceptibility to fluconazole.



This study demonstrated that caspofungin used at MIC was not sufficient to reduce C. albicans biofilms, but it suggested an activity on C. parapsilosis biofilms depending on their maturation age. This study also indicated that caspofungin used at 2 mg/L could be a good candidate in the prevention of candidiasis associated with silicone medical devices. Our results also suggested that fluconazole resistance of yeasts did not affect caspofungin activity.

title: Quorum Sensing in the Dimorphic FungusCandida albicans Is Mediated by Farnesol
abstract: The inoculum size effect in the dimorphic fungus Candida albicans results from production of an extracellular quorum-sensing molecule (QSM). This molecule prevents mycelial development in both a growth morphology assay and a differentiation assay using three chemically distinct triggers for germ tube formation (GTF): l-proline, N-acetylglucosamine, and serum (either pig or fetal bovine). In all cases, the presence of QSM prevents the yeast-to-mycelium conversion, resulting in actively budding yeasts without influencing cellular growth rates. QSM exhibits general cross-reactivity within C. albicans in that supernatants from strain A72 are active on five other strains ofC. albicans and vice versa. The QSM excreted by C. albicans is farnesol (C15H26O; molecular weight, 222.37). QSM is extracellular, and is produced continuously during growth and over a temperature range from 23 to 43°C, in amounts roughly proportional to the CFU/milliliter. Production is not dependent on the type of carbon source nor nitrogen source or on the chemical nature of the growth medium. Both commercial mixed isomer and (E,E)-farnesol exhibited QSM activity (the ability to prevent GTF) at a level sufficient to account for all the QSM activity present in C. albicans supernatants, i.e., 50% GTF at ca. 30 to 35 μM. Nerolidol was ca. two times less active than farnesol. Neither geraniol (C10), geranylgeraniol (C20), nor farnesyl pyrophosphate had any QSM activity.

title: Inhibition of Candida albicans Biofilm Formation by Farnesol, a Quorum-Sensing Molecule
abstract: Farnesol is a quorum-sensing molecule that inhibits filamentation in Candida albicans. Both filamentation and quorum sensing are deemed to be important factors in C. albicans biofilm development. Here we examined the effect of farnesol on C. albicans biofilm formation. C. albicans adherent cell populations (after 0, 1, 2, and 4 h of adherence) and preformed biofilms (24 h) were treated with various concentrations of farnesol (0, 3, 30, and 300 μM) and incubated at 37°C for 24 h. The extent and characteristics of biofilm formation were then assessed microscopically and with a semiquantitative colorimetric technique based on the use of 2,3-bis(2-methoxy-4-nitro-5-sulfo-phenyl)-2H-tetrazolium-5-carboxanilide. The results indicated that the effect of farnesol was dependent on the concentration of this compound and the initial adherence time, and preincubation with 300 μM farnesol completely inhibited biofilm formation. Supernatant media recovered from mature biofilms inhibited the ability of planktonic C. albicans to form filaments, indicating that a morphogenetic autoregulatory compound is produced in situ in biofilms. Northern blot analysis of RNA extracted from cells in biofilms indicated that the levels of expression of HWP1, encoding a hypha-specific wall protein, were decreased in farnesol-treated biofilms compared to the levels in controls. Our results indicate that farnesol acts as a naturally occurring quorum-sensing molecule which inhibits biofilm formation, and we discuss its potential for further development and use as a novel therapeutic agent.

title: Guidelines for the management of intravascular catheter-related infections.
abstract: These guidelines from the Infectious Diseases Society of America (IDSA), the American College of Critical Care Medicine (for the Society of Critical Care Medicine), and the Society for Healthcare Epidemiology of America contain recommendations for the management of adults and children with, and diagnosis of infections related to, peripheral and nontunneled central venous catheters (CVCs), pulmonary artery catheters, tunneled central catheters, and implantable devices. The guidelines, written for clinicians, contain IDSA evidence-based recommendations for assessment of the quality and strength of the data. Recommendations are presented according to the type of catheter, the infecting organism, and the associated complications. Intravascular catheter-related infections are a major cause of morbidity and mortality in the United States. Coagulase-negative staphylococci, Staphylococcus aureus, aerobic gram-negative bacilli, and Candida albicans most commonly cause catheter-related bloodstream infection. Management of catheter-related infection varies according to the type of catheter involved. After appropriate cultures of blood and catheter samples are done, empirical i.v. antimicrobial therapy should be initiated on the basis of clinical clues, the severity of the patient's acute illness, underlying disease, and the potential pathogen(s) involved. In most cases of nontunneled CVC-related bacteremia and fungemia, the CVC should be removed. For management of bacteremia and fungemia from a tunneled catheter or implantable device, such as a port, the decision to remove the catheter or device should be based on the severity of the patient's illness, documentation that the vascular-access device is infected, assessment of the specific pathogen involved, and presence of complications, such as endocarditis, septic thrombosis, tunnel infection, or metastatic seeding. When a catheter-related infection is documented and a specific pathogen is identified, systemic antimicrobial therapy should be narrowed and consideration given for antibiotic lock therapy, if the CVC or implantable device is not removed. These guidelines address the issues related to the management of catheter-related bacteremia and associated complications. Separate guidelines will address specific issues related to the prevention of catheter-related infections. Performance indicators for the management of catheter-related infection are included at the end of the document. Because the pathogenesis of catheter-related infections is complicated, the virulence of the pathogens is variable, and the host factors have not been well defined, there is a notable absence of compelling clinical data to make firm recommendations for an individual patient. Therefore, the recommendations in these guidelines are intended to support, and not replace, good clinical judgment. Also, a section on selected, unresolved clinical issues that require further study and research has been included. There is an urgent need for large, well-designed clinical studies to delineate management strategies more effectively, which will improve clinical outcomes and save precious health care resources.

title: Therapeutic approaches in patients with candidemia. Evaluation in a multicenter, prospective, observational study.
abstract: To evaluate the morbidity and mortality of Candida fungemia and to assess the efficacy of low- vs high-dose amphotericin B and fluconazole vs amphotericin B in patients with candidemia.



Multicenter, prospective, observational study of 427 consecutive patients with candidemia.



The mortality rate for patients with candidemia was 34%. The mortality rate for patients with catheter-related candidemia in whom the catheters were retained was significantly higher than that of patients in whom the catheters were removed (41% vs 21%, P < .001). We found no overall difference in mortality in patients treated with low-dose (total amphotericin B dose of < or = 500 mg) (13%) vs high-dose amphotericin B (total amphotericin B dose of > 500 mg) (15%), but the group treated with a low dose had fewer side effects (40%) than those treated with a high dose (55%) (P = .03). Fluconazole was as efficacious as amphotericin B in the therapy of candidemia, even when stratified by risk factors for mortality. Fewer side effects were seen with fluconazole (12%) compared with amphotericin B (44%) (P < .001).



In selected patients with candidemia, low-dose amphotericin B was as efficacious as high-dose amphotericin B. Based on other studies and ours, fluconazole seems to be an alternative therapeutic option to amphotericin B in selected patients.

title: Practice Guidelines for the Treatment of Candidiasis
abstract: Infections due to Candida species are the most common of the fungal infections. Candida species produce a broad range of infections, ranging from nonlife-threatening mucocutaneous illnesses to invasive process that may involve virtually any organ. Such a broad range of infections requires an equally broad range of diagnostic and therapeutic strategies. This document summarizes current knowledge about treatment of multiple forms of candidiasis and is the guideline of the Infectious Diseases Society of America (IDSA) for the treatment of candidiasis. Throughout this document, treatment recommendations are scored according to the standard scoring scheme used in other IDSA guidelines to illustrate the strength of the underlying data. The document covers 4 major topical areas. The role of the microbiology laboratory. To a greater extent than for other fungi, treatment of candidiasis can now be guided by in vitro susceptibility testing. The guidelines review the available information supporting current testing procedures and interpretive breakpoints and place these data into clinical context. Susceptibility testing is most helpful in dealing with infection due to non-albicans species of Candida. In this setting, especially if the patient has been treated previously with an azole antifungal agent, the possibility of microbiological resistance must be considered. Treatment of invasive candidiasis. In addition to acute hematogenous candidiasis, the guidelines review strategies for treatment of 15 other forms of invasive candidiasis. Extensive data from randomized trials are really available only for therapy of acute hematogenous candidiasis in the nonneutropenic adult. Choice of therapy for other forms of candidiasis is based on case series and anecdotal reports. In general, both amphotericin B and the azoles have a role to play in treatment. Choice of therapy is guided by weighing the greater activity of amphotericin B for some non-albicans species (e.g., Candida krusei) against the lesser toxicity and ease of administration of the azole antifungal agents. Flucytosine has activity against many isolates of Candida but is not often used. Treatment of mucocutaneous candidiasis. Therapy for mucosal infections is dominated by the azole antifungal agents. These drugs may be used topically or systemically and have been proven safe and efficacious. A significant problem with mucosal disease is the propensity for a small proportion of patients to suffer repeated relapses. In some situations, the explanation for such a relapse is obvious (e.g., relapsing oropharyngeal candidiasis in an individual with advanced and uncontrolled HIV infection), but in other patients the cause is cryptic (e.g., relapsing vaginitis in a healthy woman). Rational strategies for these situations are discussed in the guidelines and must consider the possibility of induction of resistance over time. Prevention of invasive candidiasis. Prophylactic strategies are useful if the risk of a target disease is sharply elevated in a readily identified group of patients. Selected patient groups undergoing therapy that produces prolonged neutropenia (e.g., some bone-marrow transplant recipients) or who receive a solid-organ transplant (e.g., some liver transplant recipients) have a sufficient risk of invasive candidiasis to warrant prophylaxis.

title: Genetic evidence for role of extracellular proteinase in virulence of Candida albicans
abstract: The relationship between extracellular proteinase and the virulence for mice in Candida albicans was studied by using a set of three isolates. The set included a proteinase-producing parent (C9), a proteinase-deficient mutant derived from C9 by nitrous acid treatment (C9M1), and a spontaneous revertant (C9M1M) obtained by mouse passage of C9M1. The morphological markers and the carbon assimilation pattern were identical in these isolates. Isolate C9 produced a high level of proteinase in vitro and caused fatal infection (100%) within 21 days. The mutant produced no detectable enzymes in vitro, and all mice survived until day 22. Only 30% of the mice infected with C9M1 died between day 23 and 30. The isolates recovered from the dead mice were found to be proteinase sufficient, indicating that the mice died after the organism in tissue had reverted. The C9M1M isolate produced proteinase in vitro at 44% the level of C9 and induced fatal infection in 90% of the mice within 30 days. The number of CFU recovered from the kidneys correlated with the level of proteinase produced in vitro and, in turn, the rate of fatal infection produced by the isolates. These results support a previous observation indicating that proteinase activity is one of the virulence factors associated with C. albicans.

title: Secreted Aspartic Proteases as Virulence Factors of Candida Species
abstract: Candida infections have emerged as a significant medical problem during the last few decades. Among the different virulence traits of C. albicans, secreted proteolytic activity has been intensively investigated. Pathogenesis of the various forms of candidiasis was shown to be associated with the differential and temporal regulation of the expression of genes coding for secreted aspartic proteases (Sap). These enzymes act as cytolysins in macrophages after phagocytosis of Candida, are present in tissue penetration and are also involved in adherence to epithelial cells. Since the introduction of new antiretroviral therapeutics such as HIV protease inhibitors, oropharyngeal candidiasis is less often observed in AIDS patients. Different HIV aspartic protease inhibitors were able to inhibit the C. albicans Saps involved in adherence. The lower rates of oropharyngeal candidiasis observed in individuals receiving antiretroviral combination therapy could reflect not only an improvement in the immune system but also direct inhibition of Candida Saps by HIV protease inhibitors. Therefore, the development of specific aspartic protease inhibitors might be of interest for the inhibition of candidiasis.

title: In Vivo Analysis of Secreted Aspartyl Proteinase Expression in Human Oral Candidiasis
abstract: Secreted aspartyl proteinases are putative virulence factors inCandida infections. Candida albicans possesses at least nine members of a SAP gene family, all of which have been sequenced. Although the expression of the SAPgenes has been extensively characterized under laboratory growth conditions, no studies have analyzed in detail the in vivo expression of these proteinases in human oral colonization and infection. We have developed a reliable and sensitive procedure to detect C. albicans mRNA from whole saliva of patients with oral C. albicans infection and those with asymptomaticCandida carriage. The reverse transcription-PCR protocol was used to determine which of the SAP1 to SAP7genes are expressed by C. albicans during colonization and infection of the oral cavity. SAP2 and the SAP4to SAP6 subfamily were the predominant proteinase genes expressed in the oral cavities of both Candida carriers and patients with oral candidiasis; SAP4, SAP5, orSAP6 mRNA was detected in all subjects. SAP1and SAP3 transcripts were observed only in patients with oral candidiasis. SAP7 mRNA expression, which has never been demonstrated under laboratory conditions, was detected in several of the patient samples. All seven SAP genes were simultaneously expressed in some patients with oral candidiasis. This is the first detailed study showing that the SAP gene family is expressed by C. albicans during colonization and infection in humans and that C. albicans infection is associated with the differential expression of individualSAP genes which may be involved in the pathogenesis of oral candidiasis.

title: Transcriptional Regulators Cph1p and Efg1p Mediate Activation of the Candida albicans Virulence Gene SAP5 during Infection
abstract: The opportunistic fungal pathogen Candida albicans can cause superficial as well as systemic infections. Successful adaptation to the different host niches encountered during infection requires coordinated expression of various virulence traits, including the switch between yeast and hyphal growth forms and secretion of aspartic proteinases. Using an in vivo expression technology that is based on genetic recombination as a reporter of gene activation during experimental candidiasis in mice, we investigated whether two signal transduction pathways controlling hyphal growth, a mitogen-activated protein kinase cascade ending in the transcriptional activator Cph1p and a cyclic AMP-dependent regulatory pathway that involves the transcription factor Efg1p, also control expression of the SAP5 gene, which encodes one of the secreted aspartic proteinases and is induced by host signals soon after infection. Our results show that both transcriptional regulators are important for SAP5 activation in vivo. SAP5 expression was reduced in a cph1 mutant, although filamentous growth in infected tissue was not detectably impaired. SAP5 expression was also reduced, but not eliminated, in an efg1 null mutant, although this strain grew exclusively in the yeast form in infected tissue, demonstrating that in contrast to in vitro conditions, SAP5 activation during infection does not depend on growth of C. albicans in the hyphal form. In a cph1 efg1 double mutant, however, SAP5 expression in infected mice was almost completely eliminated, suggesting that the two signal transduction pathways are important for SAP5 expression in vivo. The avirulence of the cph1 efg1 mutant seemed to be caused not only by the inability to form hyphae but also by a loss of expression of additional virulence genes in the host.

title: Profile of Candida albicans-Secreted Aspartic Proteinase Elicited during Vaginal Infection
abstract: Vaginal infections caused by the opportunistic yeast Candida albicans are a significant problem in women of child-bearing age. Several factors are recognized as playing a crucial role in the pathogenesis of superficial candidiasis; these factors include hyphal formation, phenotypic switching, and the expression of virulence factors, including a 10-member family of secreted aspartic proteinases. In the present investigation, we analyzed the secreted aspartic proteinase gene (SAP) expression profile of C. albicans that is elicited in the course of vaginal infection in mice and how this in vivo expression profile is associated with hyphal formation. We utilized two different genetic reporter systems that allowed us to observe SAP expression on a single-cell basis, a recombination-based in vivo expression technology and green fluorescent protein-expressing Candida reporter strains. Of the six SAP genes that were analyzed (SAP1 to SAP6), only SAP4 and SAP5 were detectably induced during infection in this model. Expression of both of these genes was associated with hyphal growth, although not all hyphal cells detectably expressed SAP4 and SAP5. SAP5 expression was induced soon after infection, whereas SAP4 was expressed at later times and in fewer cells compared with SAP5. These findings point to a link between morphogenetic development and expression of virulence genes during Candida vaginitis in mice, where host signals induce both hyphal formation and expression of SAP4 and SAP5, but temporal gene expression patterns are ultimately controlled by other factors.

title: Secreted aspartic proteases of Candida albicans, Candida tropicalis, Candida parapsilosis and Candida lusitaniae. Inhibition with peptidomimetic inhibitors.
abstract: The frequency of Candida infections has increased in recent years and it has been accompanied by a significant rise in morbidity and mortality. The secretion of aspartic proteases by Candida spp. was demonstrated to be one of the virulence determinants. Candida albicans is classified as the major human pathogen in the genus Candida. However, other species of this genus have been found to cause an increasing number of candidiases. We isolated secreted aspartic proteases (Saps) of C. albicans (Sap2p), C. tropicalis (Sapt1p), C. parapsilosis (Sapp1p), and C. lusitaniae (Saplp) from culture media. All the isolated proteases were N-terminally sequenced. Their specific proteolytic activities and sensitivity to series of peptidomimetic inhibitors modified in the type of scissile bond replacement as well as in the N- and C-termini were analyzed. The most divergent substrate specificity was observed for the Sap of C. tropicalis. The specificity of Sap of C. lusitaniae is most closely related to that of Sap of C. parapsilosis. We designed and prepared an inhibitor containing phenylstatine isoster that was equipotent towards all four proteases within the range of 10-10-10-9 M. The HIV-1 protease inhibitors ritonavir, saquinavir, indinavir, and nelfinavir were also tested for the inhibition of four Saps. Only ritonavir and saquinavir inhibited Sap2p, Sapt1p, Sapp1p, and Saplp in micromolar concentrations.

title: Candida infections: an overview.
abstract: The range of human infections caused by the yeast Candida albicans and a handful of related species is considerable. They range from relatively trivial conditions such as oral and genital thrush to fatal, systemic superinfections in patients who are already seriously ill with other diseases. Interest in Candida infections, and in C. albicans in particular, has become huge in recent years as fatal infections have become more prevalent and new Candida-based pathologies have been recognized. There is now even a large pseudoscientific cult based on the notion that chronic allergy to Candida can cause all sorts of common illnesses. The medical importance of Candida infections and the scientific value of C. albicans as a model for fungal cellular development have stimulated enormous advances in our understanding of the epidemiology of candidosis, pathogenesis of the disease, and the genetics and biochemistry of C. albicans.

title: Characterization of a secretory proteinase of Candida parapsilosis and evidence for the absence of the enzyme during infection in vitro
abstract: The opportunistic yeastlike fungi of the genus Candida comprise three species which are proteolytic in vitro. Among them, C. albicans and C. tropicalis are of foremost medical importance. However, a strict correlation between extracellular proteolytic activity and virulence is opposed by the low virulence of the third proteolytic species, C. parapsilosis. We purified the secretory acid proteinase of C. parapsilosis (clinical isolate 265). The enzyme is a carboxyl proteinase (EC 3.4.23) like all other secretory Candida proteinases handled so far. Proteinase 265 is distinguished by a lower molecular weight (approximately 33,000); it has increased hydrophobicity, which accounts for inhibition of the enzyme by hemin, and required the presence of nonionic detergent in the initial steps of purification. The enzyme already undergoes alkaline denaturation at neutrality. Its activity is thus confined to the acid microenvironment of the fungal cell wall. Within this range, the enzyme may degrade immunoglobulins like immunoglobulin A1 (IgA1), IgA2, and secretory IgA. No indication was found for glycosylation of proteinase 265 and the related enzyme of C. albicans CBS 2730. However, the comparable proteinase of C. tropicalis 293 was identified as a manno protein. Antiserum against proteinase 265 cross-reacted strongly with corresponding enzymes from other Candida species. Antisera against proteinases of C. albicans and C. tropicalis reacted only weakly with proteinase 265. Thus, secretory Candida proteinases are likely to possess common and species-specific antigenic sites. In contrast to C. albicans, infection of phagocytes by C. parapsilosis 265 was not accompanied by secretion of fungal proteinase. This lack of induction of the enzyme under conditions of infection may account for the low virulence of most isolates of C. parapsilosis.

title: The precursor of secreted aspartic proteinase Sapp1p from Candida parapsilosis can be activated both autocatalytically and by a membrane-bound processing proteinase
abstract: Opportunistic pathogens of the genus Candida produce secreted aspartic proteinases (Saps) that play an important role in virulence. Saps are synthesized as zymogens, but cell-free culture supernatants of Candida spp. contain only mature Saps. To study the zymogen conversion, the gene encoding a precursor of C. parapsilosis proteinase Sapp1p was cloned, expressed in E. coli and the product was purified. When placed in acidic conditions, the precursor was autocatalytically processed, yielding an active proteinase. The self-activation proceeded through an intermediate product and the resulting enzyme was one amino acid shorter than the authentic enzyme. This truncation did not cause changes in proteinase activity or secondary structure compared to the authentic Sapp1p. Accurate cleavage of the pro-mature junction, however, required a processing proteinase. A crude membrane fraction prepared from C. parapsilosis cells contained an enzyme with Kex2-like activity, which processed the Sapp1p precursor at the expected site. The pro-segment appeared to be indispensable for Sapp1p to attain an appropriate structure. When expressed without the pro-segment, the Sapp1p mature domain was not active and had a lower content of α-helical conformation, as measured by circular dichroism. A similar effect was observed when a His6-tag was linked to the C-terminus of Sapp1p or its precursor.

title: Extracellular aspartic proteinases from Candida albicans, Candida tropicalis, and Candida parapsilosis yeasts differ substantially in their specificities.
abstract: Extracellular aspartic proteinases have been implicated for some time as virulence factors associated with Candida opportunistic fungal infections. Our present knowledge of the enzymatic properties of these proteinases is rather limited. Information on their substrate specificity is important for understanding their roles in invasive Candida infections. We have isolated aspartic proteinases from each of the three Candida yeasts, Candida albicans, Candida tropicalis, and Candida parapsilosis, and investigated the specificities of these proteinases using a library of synthetic substrates and testing inhibition by pepstatin A. The specificities of these aspartic proteinases are different from those of major human proteinases, including gastric pepsins, renal renin, and cathepsin D. For the peptide substrate, Lys-Pro-Ala-Leu-Phe*Phe(p-NO2)-Arg-Leu, the values of kcat/Km were 2.95 x 10(6) M-1 s-1 for cleavage by Candida albicans proteinase, 1.60 x 10(6) M-1 s-1 for cleavage by Candida tropicalis proteinase, and 0.59 x 10(6) M-1 s-1 for Candida parapsilosis proteinase. Substantial differences in specificity among the Candida yeast proteinases were identified. For example, Candida tropicalis shows large changes in the kcat/Km value depending on the acidobasic character of the residue occupying the P2 position (1.6 x 10(6) M-1 s-1 for Leu, 0.47 x 10(6) M-1 s-1 for Lys, and 0.05 x 10(6) M-1 s-1 for Asp at P2, respectively). Candida parapsilosis by comparison is tolerant of these substitutions at P2 and is highly restrictive at position P4.(ABSTRACT TRUNCATED AT 250 WORDS)

title: Candida parapsilosis expresses and secretes two aspartic proteinases
abstract: We have isolated and characterized a second aspartic proteinase secreted by the CHUV E‐18 strain of Candida parapsilosis. This proteinase is produced at a level corresponding to approximately 25% of the production of the main proteinase described earlier . This minor proteinase has similar molecular weight and pH optimum but differs in the isoelectric point and in the specificity when compared with the major secreted form. The determination of the amino terminal amino acid sequence identified this minor form of Candida parapsilosis aspartic proteinase as a protein which corresponds to the sequence deduced from genomic DNA originally reported as a pseudogene . We conclude that strain CHUV E‐18 of Candida parapsilosis expresses and secretes two different aspartic proteinases.

title: Acid proteinase, phospholipase and adherence properties of Candida parapsilosis strains isolated from clinical specimens of hospitalised patients
abstract: In this study, we investigated the presence of virulence factors in a total of 33 Candida parapsilosis strains, including adherence, acid proteinase and phospholipase production capabilities. Of the 33 strains, 19 were isolated from blood cultures of hospitalised fungaemic patients (study group 2). We also investigated the importance of other factors present in the development of fungaemia. The mean adherence values of 19 blood isolates and 14 non‐blood isolates were found to be 52.63 and 57.96, respectively; these values did not differ significantly (P > 0.05). Nine non‐blood strains (64.29%) of study group 2 and five blood strains (26.31%) of study group 1 were found to be positive for acid proteinase production; this difference was statistically significant (P < 0.05). Phospholipase production was found in five strains (26.31%) of the 19 blood isolates. None of the 14 non‐blood strains exhibited phospholipase activity; this difference between the two groups was statistically significant (P < 0.05). In this study, correlations between adherence‐phospholipase, and adherence‐acid proteinase properties of strains were detected as well. In the present study, it has been observed that phospholipase production can be an important virulence factor in bloodstream infections caused by C. parapsilosis.

title: Biotyping and Virulence Properties of Skin Isolates ofCandida parapsilosis
abstract: The biotype and virulence of skin isolates of Candida parapsilosis were compared with blood isolates of the same fungus. Morphotype, resistotype, and electrophoretic karyotype determinations did not reveal any special cluster with a unique or dominant pathogenic feature among all of the isolates, regardless of their source. However, all cutaneous isolates had uniformly elevated secretory aspartyl-protease (Sap) activity, more than four times higher than the enzyme activity of the blood isolates. They were also highly vaginopathic in a rat vaginitis model, being significantly more virulent than blood isolates in this infection model. In contrast, skin isolates were nonpathogenic in systemic infection of cyclophosphamide-immunodepressed mice, while some blood isolates were, in this model, highly pathogenic (median survival time, 2 days, with internal organ invasion at autopsy). Finally, skin isolates did not differ, as a whole, from blood isolates in their adherence to plastic. This property was associated with a morphotype, as defined by a colony with continuous fringe, which was present among both skin and blood isolates. While confirming the genetic heterogenicity of C. parapsilosis, our data strongly suggest that the potential of this fungus to cause mucosal disease is associated with Sap production and is substantially distinct from that of systemic invasion.

title: Purification and Characterization of Secretory Proteinase of Candida albicans
abstract: Proteolytic activity of medically important yeasts was tested in both YCB‐BSA agar and medium. All of 134 strains of Candida albicans, 13 of 18 strains of Candida tropicalis and 11 of 18 strains of Candida parapsilosis had this activity, while none of 52 Candida glabrata strains or of 11 Cryptococcus neoformans strains tested had proteolytic activity. Strains of C. albicans fell into five groups based on the level and time‐course of in vitro proteinase productivity. Five strains randomly selected from each group were tested for pathogenicity in mice. The strain possessing the strongest pathogenicity was used to purify proteinase. The molecular weight of the proteinase was approximately 44,000 daltons and its isoelectric point was pH 4.2. Optimal pH of the proteinase was 3.2 and the enzyme was stable below pH 7.0 and lost its activity above pH 8.0 at 37 C in a 60‐min incubation. The 23 amino acid sequence of the proteinase N‐terminus was determined.

title: Exposure to Therapeutic Concentrations of Ritonavir, but Not Saquinavir, Reduces Secreted Aspartyl Proteinase of Candida parapsilosis
abstract: The effect of ritonavir and saquinavir, HIV proteinase inhibitors, on the secreted aspartyl proteinase (Sap) activity of Candida parapsilosis was studied. In a proteinase-inducing medium (yeast carbon base-bovine serum albumin), Sap activity in all clinical isolates of C. parapsilosis (n = 20) was observed at 37°C but not at 22°C. The presence of ritonavir at a concentration of 8 µg/ml produced an inhibition close to 50% albumin consumption and also delayed yeast growth; however, saquinavir did not have any effect on growth or on Sap activity. In Sabouraud broth, which does not induce Sap production, no effect was shown on yeast growth by either of the two HIV proteinase inhibitors studied.

title: Virulence of Candida parapsilosis, Candida orthopsilosis, and Candida metapsilosis in reconstituted human tissue models.
abstract: Candida parapsilosis is an increasingly important human pathogen. To study the interactions of C. parapsilosis with human tissues, we evaluated the effects of the CBS 604 type strain and three different clinical isolates on reconstituted human oral epithelial and epidermal tissues. The newly described species Candida orthopsilosis and Candida metapsilosis were also examined in these models. Microscopy of reconstituted tissues infected with yeast cells revealed severe attenuation, morphological changes and cellular damage. C. orthopsilosis caused damage similar to C. parapsilosis isolates, whereas C. metapsilosis was less virulent. To further quantitate tissue damage, we measured lactate dehydrogenase (LDH) in the culture supernatant. The relative LDH measurements correlated with our histopathological observations. We also examined the effect of the lipase inhibitor Ebelactone B and proteinase inhibitor Pepstatin A, to establish the utility of this model for studying factors of C. parapsilosis virulence. Both Ebelactone B and Pepstatin A reduced the destruction of epidermal and epithelial tissues. Our data show that reconstituted human tissues are extremely useful for modeling host interactions with C. parapsilosis and for studying fungal virulence factors.

title: Potential role of phospholipases in virulence and fungal pathogenesis.
abstract: Microbial pathogens use a number of genetic strategies to invade the host and cause infection. These common themes are found throughout microbial systems. Secretion of enzymes, such as phospholipase, has been proposed as one of these themes that are used by bacteria, parasites, and pathogenic fungi. The role of extracellular phospholipase as a potential virulence factor in pathogenic fungi, including Candida albicans, Cryptococcus neoformans, and Aspergillus, has gained credence recently. In this review, data implicating phospholipase as a virulence factor in C. albicans, Candida glabrata, C. neoformans, and A. fumigatus are presented. A detailed description of the molecular and biochemical approaches used to more definitively delineate the role of phospholipase in the virulence of C. albicans is also covered. These approaches resulted in cloning of three genes encoding candidal phospholipases (caPLP1, caPLB2, and PLD). By using targeted gene disruption, C. albicans null mutants that failed to secrete phospholipase B, encoded by caPLB1, were constructed. When these isogenic strain pairs were tested in two clinically relevant murine models of candidiasis, deletion of caPLB1 was shown to lead to attenuation of candidal virulence. Importantly, immunogold electron microscopy studies showed that C. albicans secretes this enzyme during the infectious process. These data indicate that phospholipase B is essential for candidal virulence. Although the mechanism(s) through which phospholipase modulates fungal virulence is still under investigations, early data suggest that direct host cell damage and lysis are the main mechanisms contributing to fungal virulence. Since the importance of phospholipases in fungal virulence is already known, the next challenge will be to utilize these lytic enzymes as therapeutic and diagnostic targets.

title: The relationship between the acid and alkaline phosphatase activity and the adherence of clinical isolates of Candida parapsilosis to human buccal epithelial cells
abstract: Candida parapsilosis is an emerging fungal pathogen implicated in many diseases, especially in compromised hosts. Candidal colonization and infection depends on the initial ability to adhere to host surfaces, which in turn depends upon the cell wall components and the allied structures of both the host and the fungus. Examination of a miscellaneous collection of 24 C. parapsilosis isolates, from both superficial and deep infections, for their potential pathogenic traits displayed a relationship between the phosphatase activity measured with p‐nitrophenol phosphate and adhesion of the yeasts to human buccal epithelial cells (BECs). Significant intraspecies differences were seen in both the alkaline and acid phosphatase activity as well as in their adhesion to BECs (p<0.0001). The acid phosphatase activity of the superficial isolates was significantly greater (152%) than that of the systemic isolates (p=0.0352). A highly significant positive correlation was also established between the yeast adhesion to BECs and both the acid (r=0.88, p<0.0001) and alkaline (r=0.9, p<0.0001) phosphatase activity. These relationships, described here for the first time, imply that phosphatases of Candida species may play a crucial role in potentiating their virulence.

title: The cytochemical localization of phospholipase in Candida albicans infecting the chick chorio-allantoic membrane.
abstract: The cytochemical localization of phospholipase A and lysophospholipase in blastospores and hyphae of Candida albicans is described. The possible role of these fungal enzymes in the invasion of the chick embryonic membrane is discussed.

title: An ultrastructural and a cytochemical study of candidal invasion of reconstituted human oral epithelium.
abstract: Opportunistic yeast, Candida albicans causes superficial and systemic mycoses in compromised patients. Adhesion to host tissues, morphogenesis and extracellular phospholipases (PL) are thought to contribute to its virulence. The nature of numerous host-parasite interactions at the invasive phase of oral candidiasis is not fully understood. Hence in this study, we explore the ultrastructural features of oral candidiasis using a tissue culture model based on reconstituted human oral epithelium (RHOE).



Reconstituted human oral epithelium (Skinethic Laboratory, Nice, France) was inoculated with C. albicans SC5314 and incubated up to 48 h. The infected tissue was harvested at 12, 24 and 48 h and examined using light, scanning (SEM) and transmission electron microscopy (TEM). Localized activity of PLs of C. albicans during tissue invasion was also examined using a cytochemical method.



Over a period of 48 h C. albicans invaded the RHOE, and histological examination revealed characteristic hallmarks of pathological tissue invasion. Hyphal penetration into the superficial epithelium, particularly at cell junctions, together with features of cellular internalization of yeasts was noted. Phospholipase activity was visible at the tips of hyphae and initial sites of bud formation. Further, SEM studies revealed cavitations on the surface epithelial cells particularly pronounced at the sites of hyphal invasion. Hyphal invasion was seen both at cell surfaces and intercellular cell junctions of the epithelium, the latter resembling thigmotropic behaviour.



Our findings confirm that multiple cellular interactions such as internalization, thigmotropism and extracellular PLs contribute to invasive candidiasis. The RHOE model, described here, appears to be a satisfactory model for the investigation of ultrastructural and histochemical features of invasive candidiasis in humans.

title: Candida albicans stimulates endothelial cell eicosanoid production.
abstract: The response of human umbilical vein endothelial cells to invasion by Candida species was examined in vitro. Live Candida albicans caused significant endothelial release of eicosanoids, mainly prostaglandins. Since prostaglandin I2 (PGI2) is an important prostaglandin produced by endothelial cells, factors influencing its release were studied. The ability of different strains and species of Candida to induce endothelial PGI2 release was closely related to their capacity to injure the endothelium (r = .99). C. albicans was the only species tested that either stimulated PGI2 release or damaged the endothelial cells; only this organism possessed detectable phospholipase activity. Candida glabrata and Candida tropicalis had no phospholipase activity and neither increased PGI2 release nor caused significant endothelial damage. Close proximity with germinated C. albicans was required for endothelial injury and PGI2 release. The ability of C. albicans to stimulate endothelial cells may have important implications in regulating neutrophil activities against organisms that interact with endothelial cells.

title: Hydrolytic enzymes as virulence factors of Candida albicans
abstract: Candida albicans is a facultative pathogenic micro‐organism that has developed several virulence traits enabling invasion of host tissues and avoidance of host defence mechanisms. Virulence factors that contribute to this process are the hydrolytic enzymes. Most of them are extracellularly secreted by the fungus. The most discussed hydrolytic enzymes produced by C. albicans are secreted aspartic proteinases (Saps). The role of these Saps for C. albicans infections was carefully evaluated in numerous studies, whereas only little is known about the physiological role of the secreted phospholipases (PL) and almost nothing about the involvement of lipases (Lip) in virulence. They may play an important role in the pathogenicity of candidosis and their hydrolytic activity probably has a number of possible functions in addition to the simple role of digesting molecules for nutrition. Saps as the best‐studied member of this group of hydrolytic enzymes contribute to host tissue invasion by digesting or destroying cell membranes and by degrading host surface molecules. There is also some evidence that hydrolytic enzymes are able to attack cells and molecules of the host immune system to avoid or resist antimicrobial activity. High hydrolytic activity with broad substrate specificity has been found in several Candida species, most notably in C. albicans. This activity is attributed to multigene families with at least 10 members for Saps and Lips and several members for PL B. Distinct members of these gene families are differentially regulated in various Candida infections. In future, prevention and control of Candida infections might be achieved by pharmacological or immunological tools specifically modulated to inhibit virulence factors, e.g. the family of Saps.

title: Adherence of Candida species to fibrin clots in vitro
abstract: The adherence of six Candida species to fibrin clots was studied using a simple, in vitro technique. Yeast suspensions were incubated with fibrin clots and the number of adherent organisms quantified as follows: after washing, the clots were subjected to vortex mixing and the number of CPU's which subsequently grew on Sabourauds medium were counted. Adhesion was directly proportional to the concentration of Candida species in the suspension (r=0.99 p<0.001). C. albicans and C. tropicalis exhibited marked adherence whereas C. krusei, C. gulliermondi and C. glabrata adhered less readily. C. parapsilosis was intermediate in its ability to adhere.

title: Studies on hyaluronidase, chondroitin sulphatase, proteinase and phospholipase secreted by Candida species
abstract: We studied the ability of different Candida species to produce, at the same time, hyaluronidase, chondroitin sulphatase, proteinase, and phospholipase to assess whether they could be related to Candida pathogenicity. Only C. albicans was able to produce the four enzymes tested (73%) and was highly virulent to mice. Strains, that lack the capacity to produce one or more of the enzymes assayed, seemed less virulent or avirulent, similarly to the spontaneous hyaluronidase, chondroitin sulphatase, phospholipase and proteinase‐deficient C. albicans strain FCF 14,1 which was non‐pathogenic to mice. Among the other Candida species tested, none of them produced the four enzymes simultaneously, being less virulent in intravenously inoculated mice.

title: Model of interaction of polar lipids, cholesterol, and proteins in biological membranes
abstract: Membranes are proposed to consist of a hydrophobic core, two hydrogen belts, and two polar zones. The hydrogen belts consist of hydrogen bond acceptors, i.e. the carbonyl groups of phospholipids and sphingolipids, and hydrogen bond donors, i.e. the labile hydrogens of cholesterol, sphingosine, proteins, and water. The density of anhydrous hydrogen bonding and the impermeability of the membrane increase with increasing concentrations of cholesterol, sphingolipids, α-hydroxy acyl residues, plasmalogens, and ether phospholipids. Cholesterol owes its membrane-closing properties to its rigid longitudinal orientation in the membrane combined with the latitudinal orientation of the O−H bond. It is suggested that the intrinsic proteins of membranes are held in position by hydrogen bonding, as well as by hydrophobic and electrostatic forces, and that hydrogen bonding also mediates the penetration of membranes by proteins.

title: Expression analysis of the Candida albicans lipase gene family during experimental infections and in patient samples.
abstract: Secreted lipases of Candida albicans are encoded by a gene family with at least 10 members (LIP1-LIP10). The expression pattern of this multigene family was investigated using reverse transcription polymerase chain reaction in experimental infections and in samples of patients suffering from oral candidosis. The findings illustrate that individual lipase genes are differentially regulated in a mouse model of systemic candidosis with some members showing sustained expression and others being transiently expressed or even silent. The lipase gene expression profile depended on the stage of infection rather than on the organ localization. This temporal regulation of lipase gene expression was also detected in an experimental model of oral candidosis. Furthermore, the expression of candidal lipase genes in human specimens is shown for the first time.

title: Induction of human neutrophils chemotaxis by staphylococcal lipase.
abstract: Influence of highly purified staphylococcal lipase on chemotactic activity of human polymorphonuclear leukocytes (PMN), has been studied. Staphylococcal lipase exhibited both chemotactic and chemokinetic properties. Chemotactic response was stimulated at concentration as low as 10(-9) M and was dose-dependent. Pretreatment of PMN with lipase, strongly enhanced chemotactic response toward casein. At the same time, spontaneous migration of these PMN was inhibited. It has been demonstrated that staphylococcal lipase binds to the leukocyte surface. Several possible mechanisms of lipase-induced chemotactic response of human PMN are discussed.

title: Propionibacterium acnes, a resident of lipid-rich human skin, produces a 33 kDa extracellular lipase encoded by gehA.
abstract: Five independent clones of the Propionibacterium acnes P-37 lipase gene (gehA) were obtained in Escherichia coli, and the gene was localized to a 2.75 kb Xhol fragment by subcloning. The five clones were shown to contain the same gene by Southern blotting with a DIG-labelled probe to gehA. The nucleotide sequence of gehA was determined, and shown to contain a single ORF of 1017 kb, encoding a protein of 339 amino acids. The predicted molecular mass was 36 kDa. A 33 kDa (PAGE) radiolabelled polypeptide was detected from E. coli minicell preparations harbouring gehA, which could correspond to GehA after cleavage of the putative 26 amino acid residue signal peptide. gehA was overexpressed in E. coli under the control of the bacteriophage T7 promoter, and the corresponding polypeptide was found to be present in insoluble aggregates. Active lipase was produced when the overexpressing strain was incubated at a reduced temperature in the presence of sucrose. Purification of lipase from P. acnes culture supernatant fluids confirmed the production of a 33 kDa (PAGE) lipase.

title: Extracellular lipase from Pseudomonas aeruginosa is an amphiphilic protein.
abstract: Lipase (triacylglycerol acylhydrolase, EC 3.1.1.3) secreted by Pseudomonas aeruginosa PAC1R was purified from cell-free growth medium by preparative isoelectric focusing. After blotting the N-terminal amino acid sequence and the amino acid composition were determined and compared to P. fragi and P. cepacia lipases yielding significant homology between all three species. Additionally, a consensus sequence K-Y-P-i-v-l-V-H-G was identified residing at the N-terminus of Pseudomonas lipases and in the central part of Staphylococcus lipases. Treatment of lipase with the serine-specific inhibitor diethyl p-nitrophenyl phosphate caused a rapid and complete inhibition of enzyme activity indicating the presence of a serine at the catalytic site as expected from lipase consensus sequences. Upon charge-shift electrophoresis the electrophoretic mobility of purified lipase was shifted either anodally or cathodally in the presence of sodium deoxycholate and cetyltrimethylammoniumbromide, respectively. This result demonstrates that extracellular lipase of P. aeruginosa exhibits an amphiphilic character like intrinsic membrane proteins.

title: Lipase of Malassezia furfur: some properties and their relationship to cell growth.
abstract: Lipase activity of Malassezia furfur, detected with alpha-naphthyl palmitate as a substrate, appeared to be associated with the insoluble fraction of the organism. Profiles of M. furfur lipase were similar to those of Candida cylindracea lipase. The pH optimum of the lipase was acidic, pH 5.0, which is very similar to skin surface pH. The enzyme activity was strongly activated by a lipase activator, sodium taurocholate (STC). Addition of STC to the culture medium activated cell growth in a dose-dependent manner and induced hyphae production. These results suggest that M. furfur lipase plays an important role in cell growth.

title: Cell‐surface hydrophobicity and lipolysis as essential factors in human tinea nigra
abstract: Hydrophobic adhesion of cells of the black yeast Hortaea werneckii (Horta) Nishimura & Miyaji, causative agent of human tinea nigra, was established by microbial adhesion to hexadecane (MATH) and adhesion to polystyrene, and compared with adhesion of other species of black yeasts. Additional ecophysiological tests were performed. Hortaea werneckii cells proved to have a high degree of hydrophobicity (98.5% MATH). The species is unable to degrade keratin, but shows significant lipolytic activity. It is concluded that H. werneckii is a commensal, that shows lipophilic adhesion to human skin and survives by the assimilation of excretion products.

title: Secreted lipases of Candida albicans: cloning, characterisation and expression analysis of a new gene family with at least ten members
abstract: Extracellular lipolytic activity enabled the human pathogen Candida albicans to grow on lipids as the sole source of carbon. Nine new members of a lipase gene family (LIP2–LIP10) with high similarities to the recently cloned lipase gene LIP1 were cloned and characterised. The ORFs of all ten lipase genes are between 1281 and 1416 bp long and encode highly similar proteins with up to 80% identical amino acid sequences. Each deduced lipase sequence has conserved lipase motifs, four conserved cysteine residues, conserved putative N-glycosylation sites and similar hydrophobicity profiles. All LIP genes, except LIP7, also encode an N-terminal signal sequence. LIP3–LIP6 were expressed in all media and at all time points of growth tested as shown by Northern blot and RT-PCR analyses. LIP1, LIP3, LIP4, LIP5, LIP6 and LIP8 were expressed in medium with Tween 40 as a sole source of carbon. However, the same genes were also expressed in media without lipids. Two other genes, LIP2 and LIP9, were only expressed in media lacking lipids. Transcripts of most lipase genes were detected during the yeast-to-hyphal transition. Furthermore, LIP5, LIP6, LIP8 and LIP9 were found to be expressed during experimental infection of mice. These data indicate lipid-independent, highly flexible in vitro and in vivo expression of a large number of LIP genes, possibly reflecting broad lipolytic activity, which may contribute to the persistence and virulence of C. albicans in human tissue.

title: Lipase 8 Affects the Pathogenesis of Candida albicans
abstract: The production of lipases can affect microbial fitness and virulence. We examined the role of the lipase 8 (LIP8) gene in the virulence of Candida albicans by constructing Δlip8 strains by the URA-blaster disruption method. Reverse transcription-PCR experiments demonstrated the absence of LIP8 expression in the homozygous knockout mutants. Reconstituted strains and overexpression mutants were generated by introducing a LIP8 open reading frame under control of a constitutive actin promoter. Knockout mutants produced more mycelium, particularly at higher temperatures and pH ≥7. Diminished LIP8 expression resulted in reduced growth in lipid-containing media. Mutants deficient in the LIP8 gene were significantly less virulent in a murine intravenous infection model. The results clearly indicate that Lip8p is an important virulence factor of C. albicans.

title: The lipase/acyltransferase from Candida parapsilosis: molecular cloning and characterization of purified recombinant enzymes.
abstract: Candida parapsilosis has been previously shown to produce a lipase (i.e. able to catalyze efficiently the hydrolysis of insoluble lipid esters such as triacylglycerols) that preferentially catalyses transfer reactions such as alcoholysis in the presence of suitable nucleophiles other than water, even in aqueous media with high (> 0.9) water thermodynamic activity. The present work describes the cloning and the overexpression of the gene coding for this enzyme. Two ORFs (CpLIP1 and CpLIP2) were isolated. The deduced 465-amino-acid protein sequences contained the consensus motif (G-X-S-X-G) which is conserved among lipolytic enzymes. Only one of the two deduced proteins (CpLIP2) contained peptide sequences obtained from the purified lipase/acyltransferase. Homology investigations showed that CpLIP2 has similarities principally with 11 lipases produced by C. albicans (42-61%) and the lipase A from Candida antarctica (31%) but not with the other lipases sequenced so far. Both CpLIP1 and CpLIP2 were expressed in Saccharomyces cerevisiae, but only CpLIP2 coded for an active protein. The substrate specificity and the catalytic behavior of purified recombinant CpLIP2, with or without a C-terminal histidine tag, were not changed compared to those of the native lipase.

title: Echinocandins--an advance in the primary treatment of invasive candidiasis.
abstract: Candida species are now the fourth leading cause of nosocomial bloodstream infection in the United States, exceeded in some series only by Staphylococcus aureus, coagulase-negative staphylococcus, and enterococcus species.1,2 Wey and colleagues estimate the attributable mortality of candidemia at 38 percent.3 Candidemia is a surrogate marker for the presence of deeply invasive candidiasis, particularly in immunocompromised  In patients with untreated or inadequately treated candidemia, the rate of clinically overt complications, including endophthalmitis, endocarditis, septic arthritis, and renal candidiasis, may be as high as 15 percent. Candidemia may be complicated by chronic disseminated candidiasis in neutropenic hosts and by . . .

title: Multiechinocandin- and Multiazole-Resistant Candida parapsilosis Isolates Serially Obtained during Therapy for Prosthetic Valve Endocarditis
abstract: Echinocandins are approved for the treatment of candidal infections. In vitro they have been shown to be less potent against strains of Candida parapsilosis than against other Candida spp. This is the first case report describing the development of a secondary multidrug (echinocandin-azole)-resistant Candida strain during therapy.

title: Pharmacokinetics, outcome of treatment, and toxic effects of amphotericin B and 5-fluorocytosine in neonates.
abstract: To determine the pharmacokinetics of amphotericin B and 5-fluorocytosine in neonates, we measured serum concentrations at first dose and after 5 days of therapy by high-performance liquid chromatography in 13 neonates (mean birth weight 1.2 +/- 0.8 kg). The dose of amphotericin B was serially increased from 0.1 to 0.5 mg/kg/day in 10 infants but was decreased from 0.8 to 1.0 to 0.5 mg/kg/day in three infants. Amphotericin B concentrations were not detectable in infants receiving 0.1 mg/kg/day. Amphotericin B cerebrospinal fluid concentrations were 40% to 90% of serum values obtained simultaneously. Serum concentrations after oral administration of 5-fluorocytosine (dose 25 to 100 mg/kg/day) were detectable in all infants. We found extreme interindividual variability for the half-life, volume of distribution, and clearance for both drugs. Four infants had minimal elimination for both drugs between doses, a finding that correlates with rises in serum creatinine (greater than 0.4 mg/dl, 40 mumol/L) and blood urea nitrogen (greater than 10 mg/dl, 3.6 mmol/L). We recommend that the dose of amphotericin B given on the first day of treatment be greater than the usual testing dose of 0.1 mg/kg/day. We also recommend an initial 24-hour dosing interval for amphotericin B and 5-fluorocytosine. Serum drug concentrations may need to be monitored in high-risk, low birth weight infants.

title: Amphotericin B lipid complex for neonatal invasive candidiasis
abstract: This study describes the safety and efficacy of amphotericin B lipid complex (ABLC) in 11 neonates with systemicCandida infections. Nine of the 11 improved clinically, and eight of nine evaluable patients had a mycological cure with ABLC. Creatinine levels improved or did not significantly change in eight of the 11 patients.

title: Treatment of candidaemia in premature infants: comparison of three amphotericin B preparations.
abstract: The objective of this study was to compare the effectiveness and tolerability of three antifungal preparations, amphotericin B, liposomal amphotericin B (LamB) and amphotericin B colloidal dispersion (ABCD), in the treatment of neonatal Candida bloodstream infection (CBSI).



All patients hospitalized in the neonatal intensive care unit from 1996 to 2000 with CBSI were enrolled. Patients with a serum creatinine concentration of <1.2 mg/dL received amphotericin B, and those with serum creatinine > or =1.2 mg/dL received LamB or ABCD. Complete blood counts, and renal and hepatic function tests were obtained before, during and after treatment; blood cultures were performed daily until three consecutive cultures were negative. If cultures were positive for more than 10 days with clinical signs of fungal infection and/or persistent thrombocytopenia, a second antifungal drug was added.



Fifty-six infants met the study criteria: four term and 52 preterm, including 36 extremely low birth weight infants. Amphotericin B was the initial treatment for 34, LamB for 6 and ABCD for 16 infants. No differences in mortality were found between the three groups. Sterilization of the blood was achieved with amphotericin B in 67.6% of patients, LamB in 83.3% and ABCD in 57.1%, when used as monotherapy; with the addition of a second antifungal agent, success rates were 100%, 83.3% and 92.8%, respectively. There were no differences between the groups in the time to resolution of fungaemia. No patients had immediate local or systemic adverse events and none showed deterioration in renal function.



ABCD and LamB appear to be effective, safe and well tolerated in premature infants with CBSI and renal dysfunction. Larger trials are needed before routine use can be recommended.

title: The safety and efficacy of amphotericin B colloidal dispersion in the treatment of invasive mycoses.
abstract: Amphotericin B colloidal dispersion (ABCD), a novel formulation of amphotericin B and cholesteryl sulfate in a 1:1 ratio, was developed to reduce the toxicity of amphotericin B yet retain its antifungal efficacy. In an open-label trial, ABCD at dosages as high as 6 mg/(kg.d) was administered to 168 patients with documented or presumed systemic mycoses. All patients had responded incompletely to at least 7 days' treatment with conventional amphotericin B (CAB), had experienced CAB-induced nephrotoxic effects, had preexisting renal impairment, or had experienced other CAB-related, treatment-limiting toxic effects. The clinical response to ABCD could be evaluated in 97 patients. Complete clinical response or improvement was noted in 48 (49%) of them after a mean treatment duration of 18.5 days. All 168 enrolled patients were evaluated with regard to safety of the treatment. Even at daily doses as high as 6 mg/kg, and mean and median cumulative doses of 4.0 g and 2.4 g, respectively, ABCD had little renal toxicity: the mean change in serum level of creatinine from baseline to final value was -0.02 mg/dL. Hypokalemia developed in eight patients (5%). This study provides preliminary evidence that ABCD is effective in treating invasive mycoses and lacks the dose-limiting nephrotoxicity of CAB.

title: Successful caspofungin treatment of multidrug resistant Candida parapsilosis septicaemia in an extremely low birth weight neonate
abstract: Candida septicaemia with multidrug resistance is an uncommon event in preterm neonates. We present an extremely low birth weight infant (gestational age of 27 weeks, birth weight of 980 g) who developed congenital Candida parapsilosis septicaemia. Because the fungus was resistant both to amphotericin B and fluconazole, caspofungin was chosen for therapy. The fungus was successfully eradicated without any clinical or laboratory adverse effects.

title: Antifungal Susceptibility Survey of 2,000 Bloodstream Candida Isolates in the United States
abstract: Candida bloodstream isolates (n = 2,000) from two multicenter clinical trials carried out by the National Institute of Allergy and Infectious Diseases Mycoses Study Group between 1995 and 1999 were tested against amphotericin B (AMB), flucytosine (5FC), fluconazole (FLU), itraconazole (ITR), voriconazole (VOR), posaconazole (POS), caspofungin (CFG), micafungin (MFG), and anidulafungin (AFG) using the NCCLS M27-A2 microdilution method. All drugs were tested in the NCCLS-specified RPMI 1640 medium except for AMB, which was tested in antibiotic medium 3. A sample of isolates was also tested in RPMI 1640 supplemented to 2% glucose and by using the diluent polyethylene glycol (PEG) in lieu of dimethyl sulfoxide for those drugs insoluble in water. Glucose supplementation tended to elevate the MIC, whereas using PEG tended to decrease the MIC. Trailing growth occurred frequently with azoles. Isolates were generally susceptible to AMB, 5FC, and FLU. Rates of resistance to ITR approached 20%. Although no established interpretative breakpoints are available for the candins (CFG, MFG, and AFG) and the new azoles (VOR and POS), they all exhibited excellent antifungal activity, even for those strains resistant to the other aforementioned agents.

title: In vitro susceptibility of Candida species to five antifungal agents in a German university hospital assessed by the reference broth microdilution method and Etest.
abstract: The aim of this study is to evaluate the susceptibilities of Candida spp. to the common antifungal agents in a German university hospital. Since quick results of in vitro testing are desirable, Etest and the CLSI broth microdilution (BMD) method (reference method) were compared, focusing on the validity of early readings.



A total of 512 Candida spp. isolates, including 174 from primarily sterile sites, were collected in the clinical routine. The yeasts were differentiated by CHROMagar and verified by API 20C AUX if necessary. In vitro susceptibilities to amphotericin B, flucytosine, fluconazole, voriconazole and caspofungin were determined using the BMD method described in the CLSI (formerly NCCLS) M27-A2 document and Etest. MICs were noted after 24 and 48 h of incubation.



The most frequently isolated species was Candida albicans. Among the non-albicans species, Candida glabrata was the most prevalent, followed by Candida tropicalis, Candida parapsilosis and Candida krusei. MICs (mg/L) at which 90% of the strains were inhibited were 1 for amphotericin B, 32 for flucytosine, 8 for fluconazole, 0.25 for voriconazole and 1 for caspofungin. Susceptibility to fluconazole was 85.0% for C. glabrata and 5.3% for C. krusei, almost all other isolates were susceptible in over 90% except very rare species. The 48 h MIC values of Etest and BMD were in agreement (no more than 2 log(2) dilutions) in 88.7% to 98.1% with categorical agreement rates of 91.6% to 98.2%, depending on the antifungal agent. Comparison of the 24 h MICs of both BMD and Etest with the 48 h MICs of the reference method showed categorical agreement in 94.9% to 99.2%. For caspofungin, however, a comparison of the categorical agreement was not possible due to the lack of interpretive breakpoints. The order of frequency and the resistance patterns of the isolates from primarily sterile sites and those of isolates from non-sterile sites did not differ.



No alarming resistances against the agents tested were found; however, owing to the relatively high frequency of C. glabrata with elevated fluconazole MICs, this species and, to a certain extent, C. krusei must be taken into consideration when choosing antifungal agents for calculated therapy. Etest is a reliable method for the susceptibility testing of Candida spp. and the 24 h readings of both Etest and BMD can serve as helpful preliminary results in most cases.

title: Trends in frequency and in vitro susceptibilities to antifungal agents, including voriconazole and anidulafungin, of Candida bloodstream isolates. Results from a six-year study (1996-2001).
abstract: The frequency of isolation and antifungal susceptibility patterns to established and two new antifungal agents were determined for 218 Candida spp isolates causing bloodstream infection from 1996 to 2001. Overall, 41.7% of the candidemias were due to C. albicans, followed by C. parapsilosis (22%), C. tropicalis (16.1%), C. glabrata (11.9%), C. krusei (6%) and miscellaneous Candida spp (2.3%). Isolates of C. albicans C. parapsilosis and C. tropicalis (80% of isolates) were highly susceptible to fluconazole (94 to 100% at </= 8 microg/ml) and voriconazole (97 to 100% at </= 1 microg/ml). By comparison with the newer agents itraconazole was less active (77 to 97% at </=0.12 microg/ml). Only 77% and 15% of C. glabrata isolates were inhibited by fluconazole at </= 8 microg/ml and itraconazole at </=0.12 microg/ml, respectively. Voriconazole showed a remarkable in vitro potency against C. glabrata as well as C. krusei isolates (100% at </= 1 microg/ml). Anidulafungin was very active against Candida spp isolates (MIC90: </= 0.5 microg/ml), except C. parapsilosis (MIC90: 4 microg/ml) and two C. guilliermondii isolates (MIC: >/= 32 microg/ml).

title: Amphotericin B: spectrum and resistance.
abstract: Amphotericin B is a polyene macrolide antibiotic derived from the actinomycete Streptomyces nodosus. Of the 200 known polyene agents, amphotericin B is the only one with toxicities that are sufficiently limited to permit intravenous administration. All polyenes have a common mechanism of action in that they preferentially bind to ergosterol, the primary sterol in the fungal cell membrane. The consequence of this binding includes disruption of the osmotic integrity of the membrane, with leakage of intracellular potassium and magnesium, and also the disruption of oxidative enzymes in target cells. Amphotericin B has a relatively broad spectrum of action and is useful in treating cases of candidosis, cryptococcosis, histoplasmosis, blastomycosis, paracoccidioidomycosis, coccidioidomycosis, aspergillosis, extracutaneous sporotrichosis and mucormycosis, and some cases of hyalohyphomycosis and phaeohyphomycosis. Resistance (MIC > 2 mg/L) tends to be species-dependent and emerges uncommonly and slowly in isolates from patients treated with amphotericin B. These include some individual strains of Candida albicans, Candida tropicalis, Candida parapsilosis and Candida lusitaniae, which may acquire resistance during treatment. Some isolates of Scedosporium apiospermum, Fusarium spp. and Sporothrix schenckii also show primary resistance, whereas all strains of Scedosporium prolificans demonstrate resistance. The main problems associated with the use of conventional amphotericin B have always been due to its poor aqueous solubility and toxicity rather than antifungal resistance.

title: In Vitro Susceptibilities of CandidaBloodstream Isolates to the New Triazole Antifungal Agents BMS-207147, Sch 56592, and Voriconazole
abstract: Sch 56592, and voriconazole are three new investigational triazoles with broad-spectrum antifungal activity. The in vitro activities of these three agents were compared with those of itraconazole and fluconazole against 1,300 bloodstream isolates ofCandida species obtained from over 50 different medical centers in the United States. The MICs of all of the antifungal drugs were determined by broth microdilution tests performed according to the National Committee for Clinical Laboratory Standards method using RPMI 1640 as a test medium. BMS-207147, Sch 56592, and voriconazole were all quite active against all Candida sp. isolates (MICs for 90% of the isolates tested [MIC90s], 0.5, 1.0, and 0.5 μg/ml, respectively). Candida albicans was the most susceptible species (MIC90s, 0.03, 0.06, and 0.06 μg/ml, respectively), and C. glabrata was the least susceptible (MIC90s, 4.0, 4.0, and 2.0 μg/ml, respectively). BMS-207147, Sch 56592, and voriconazole were all more active than itraconazole and fluconazole against C. albicans, C. parapsilosis, C. tropicalis, and C. krusei. There existed a clear rank order of in vitro activity of the five azoles examined in this study when they were tested versusC. glabrata: voriconazole > BMS-207147 = Sch 56592 = itraconazole > fluconazole (MIC90s, 2.0, 4.0, 4.0, 4.0, and 64 μg/ml, respectively). For isolates ofCandida spp. with decreased susceptibility to both itraconazole and fluconazole, the MICs of BMS-207147, Sch 56592, and voriconazole were also elevated. These results suggest that BMS-207147, Sch 56592, and voriconazole all possess promising antifungal activity and that further in vitro and in vivo investigations are warranted to establish the clinical value of this improved potency.

title: The European Confederation of Medical Mycology (ECMM) survey of candidaemia in Italy: antifungal susceptibility patterns of 261 non-albicans Candida isolates from blood.
abstract: To analyse the in vitro antifungal susceptibility of 261 non-albicans Candida bloodstream strains isolated during the European Confederation of Medical Mycology survey of candidaemia performed in Lombardia, Italy (September 1997-December 1999).



In vitro susceptibility to flucytosine, fluconazole, itraconazole, posaconazole and voriconazole was determined using the broth microdilution method described in the NCCLS M27-A guidelines. Etest strips were used to assess susceptibility to amphotericin B. In vitro findings were correlated with the patient's underlying condition and previous antifungal treatment.



MICs (mg/L) at which 90% of the strains were inhibited were, respectively, 2 for flucytosine, 8 for fluconazole, 0.5 for itraconazole, 0.25 for voriconazole and 0.25 for posaconazole. Amphotericin B MIC endpoints were <0.50 mg/L in all the isolates tested. Flucytosine resistance was detected in 19 isolates (7%), mainly among Candida tropicalis strains (30%). Innate or secondary fluconazole resistance was detected in 13 strains (5%). Among the 13 patients with fluconazole-resistant Candida bloodstream infection, three were HIV positive, including one treated with fluconazole for oral candidosis; the four who were HIV negative had received the azole during the 2 weeks preceding the candidaemia. Cross-resistance among fluconazole and other azoles was a rare event.



Resistance is still uncommon in non-albicans Candida species recovered from blood cultures. However, in fungaemias caused by C. tropicalis, Candida glabrata and Candida krusei, there is a high prevalence of resistance to fluconazole and flucytosine. Fluconazole resistance should be suspected in patients treated previously with azoles, mainly those with advanced HIV infection.

title: Resistance of Candida species to antifungal agents: molecular mechanisms and clinical consequences.
abstract: Candida albicans and related species pathogenic for man become resistant to antifungal agents, in particular triazole compounds, by expression of efflux pumps that reduce drug accumulation, alteration of the structure or concentration of antifungal target proteins, and alteration of membrane sterol composition. The clinical consequences of antifungal resistance can be seen in treatment failures in patients and in changes in the prevalences of Candida species causing disease. These effects were seen unequivocally in HIV-infected patients with oropharyngeal candida infections, but their incidence has decreased dramatically with the introduction of highly active antiretroviral therapy. The evidence for similar emergence of antifungal-resistant yeast strains and species in other types of candida infections is confounded by non-standardised susceptibility testing methods and definitions of a resistant fungal isolate. Recent large-scale surveys of yeasts isolated from blood cultures, based on standardised methodology and resistance definitions, do not support the view that antifungal resistance in pathogenic yeasts constitutes a significant or growing therapeutic problem.

title: Emergence of Fluconazole Resistance in a Candida parapsilosis Strain That Caused Infections in a Neonatal Intensive Care Unit
abstract: Candida parapsilosis is an increasing cause of bloodstream infections (BSIs) in neonatal intensive care units (NICUs). It has been a persistent problem in the NICU of Hospital for Children and Adolescents, Helsinki University Central Hospital, Helsinki, Finland, since 1987. Fluconazole prophylaxis has been used to control the problem. The number of new infections has, however, increased markedly since September 2000. We assessed fluconazole consumption and occurrence of all Candida species in the NICU from 1991 to 2002. C. parapsilosis bloodstream isolates obtained in the NICU from 1990 to 2002 (n = 26) were genotyped and their fluconazole susceptibility was defined. A low rate of C. parapsilosis BSIs was correlated with high rates of consumption of fluconazole. No emergence of Candida species with primary resistance to fluconazole was detected. However, genotyping with a complex DNA fingerprinting probe revealed that a single strain of C. parapsilosis with decreasing susceptibility to fluconazole was responsible for cross-infections that caused BSIs in the NICU over a 12-year period. The emergence of fluconazole resistance in that strain was observed after more than 10 years of fluconazole prophylaxis.

title: Caspofungin Activity against Clinical Isolates of Fluconazole-Resistant Candida
abstract: A total of 7,837 clinical isolates of Candida were tested against fluconazole, and 351 resistant (fluconazole MIC ≥ 64 μg/ml) isolates were identified (4% of the total tested). All fluconazole-resistant isolates were inhibited by caspofungin at concentrations that can be exceeded by standard doses (MIC at which 90% of the isolates were inhibited, 1 μg/ml; 99% of the MICs were ≤2 μg/ml).

title: Fluconazole prophylaxis against fungal colonization and infection in preterm infants.
abstract: Invasive fungal infection is associated with substantial morbidity and mortality in preterm infants. We evaluated the efficacy of prophylactic fluconazole in preventing fungal colonization and invasive infection in extremely-low-birth-weight infants.



We conducted a prospective, randomized, double-blind clinical trial over a 30-month period in 100 preterm infants with birth weights of less than 1000 g. The infants were randomly assigned during the first five days of life to receive either intravenous fluconazole or placebo for six weeks. We obtained weekly surveillance cultures from all patients.



The 50 infants randomly assigned to fluconazole and the 50 control infants were similar in terms of birth weight, gestational age at birth, and base-line risk factors for fungal infection. During the six-week treatment period, fungal colonization was documented in 30 infants in the placebo group (60 percent) and 11 infants in the fluconazole group (22 percent; difference in risk, 0.38; 95 percent confidence interval, 0.18 to 0.56; P=0.002). Invasive fungal infection with positive growth of fungal isolates from the blood, urine, or cerebrospinal fluid developed in 10 infants in the placebo group (20 percent) and none of the infants in the fluconazole group (difference in risk, 0.20; 95 percent confidence interval, 0.04 to 0.36; P=0.008). The sensitivities of the fungal isolates to fluconazole did not change during the study, and no adverse effects of the fluconazole therapy were documented.



Prophylactic administration of fluconazole during the first six weeks of life is effective in preventing fungal colonization and invasive fungal infection in infants with birth weights of less than 1000 g.

title: Targeted Short-Term Fluconazole Prophylaxis Among Very Low Birth Weight and Extremely Low Birth Weight Infants
abstract: To assess whether targeted short-term fluconazole prophylaxis reduces late-onset (>3 days of age) invasive fungal infection (IFI) among very low birth weight infants and extremely low birth weight (ELBW) infants and to assess mortality rates, toxicity, and costs associated with this intervention.  An observational study of 2 subsequent epochs of inborn infants with birth weight of <1500 g or gestational age of <32 weeks, 1 before (control) and 1 after (fluconazole) initiation of routine targeted fluconazole prophylaxis in March 2003, was performed. Targeted fluconazole (3 mg/kg) prophylaxis was administered to infants for whom a decision was made to administer broad-spectrum antibiotics for >3 days.  IFI was observed for 13 (6.3%) of 206 infants in the control epoch and 2 (1.1%) of 178 in the fluconazole epoch, with a common odds ratio of 0.166. Logistic regression analysis taking into account all published factors (except for fungal colonization) showed that the fluconazole epoch was associated significantly with lower IFI rates. We observed no change in late (>3 days) mortality rates (11 of 206 infants in the control epoch vs 8 of 178 infants in the prophylaxis epoch). The mortality rate for ELBW infants with IFI was low (15%) in our study. Fluconazole was administered to 81% of ELBW infants, who received a median of 8 doses, and 41% of larger infants, who received a median of 5 doses. The intervention was cost-effective, and the effective number needed to treat to prevent 1 IFI was 10.  This study suggests that targeted short-course fluconazole prophylaxis in very low birth weight and ELBW infants may be efficacious and cost effective.

title: Fluconazole prophylaxis: can we eliminate invasive Candida infections in the neonatal ICU?
abstract: Owing to the high mortality, risk of neurodevelopmental impairment and end-organ involvement with fungal infections in the neonate, prevention of invasive Candida infections in extremely preterm infants should be a priority for each neonatal ICU.  Even with prompt or empiric treatment, mortality and neurodevelopmental impairment is high (57%) in infants <1000 g. Multiple studies have been performed with fluconazole prophylaxis, including a recent multicenter randomized controlled trial. All of the studies have demonstrated efficacy and safety with no increase or emergence of fungal resistance. Analysis of these studies demonstrates that fluconazole prophylaxis decreased the incidence of invasive Candida infections in high-risk infants <1000 g by 91% (P = 0.0004) and all infants <1500 g by 85% (P < 0.0001). The mortality rate from all causes was 25% lower (P = 0.029). Furthermore, studies have demonstrated that all Candida-related mortality can be eliminated in an entire neonatal ICU by targeting fluconazole prophylaxis in infants <1000 g.  Targeting fluconazole prophylaxis to infants who are either <1000 g or ≤27 weeks is highly effective, safe and inexpensive, and can eliminate these infections as a cause of neurodevelopmental impairment and mortality.

title: In vitro susceptibility testing of Candida and Aspergillus spp. to voriconazole and other antifungal agents using Etest: results of a French multicentre study.
abstract: Minimum inhibitory concentrations (MICs) of the antifungal agent voriconazole were determined using the Etest and compared with those of amphotericin B, itraconazole and fluconazole using 1986 clinical isolates of Candida spp. Voriconazole MICs were also compared with those of amphotericin B and itraconazole using 391 clinical isolates of Aspergillus spp. Voriconazole was found to have more potent activity and lower MIC values than amphotericin B, itraconazole and fluconazole against C. albicans, C. tropicalis, C. parapsilosis and C. kefyr. Against C. glabrata and C. krusei, voriconazole was more active than either of the other two azole antifungals but had similar activity to amphotericin B. For species of Aspergillus, MIC values of voriconazole were lower than those of amphotericin B and itraconazole against A. fumigatus and A. flavus, and were similar to those of amphotericin B against A. niger. Against A. terreus, MIC values for voriconazole and itraconazole were similar. A. terreus is known to be resistant to amphotericin B, and this was reflected in higher MIC values compared with those of voriconazole and itraconazole. Voriconazole therefore compares very favourably with other antifungal agents against a large number of clinical isolates of Candida and Aspergillus spp.

title: Activities of Fluconazole and Voriconazole against 1,586 Recent Clinical Isolates of Candida Species Determined by Broth Microdilution, Disk Diffusion, and Etest Methods: Report from The ARTEMIS Global Antifungal Susceptibility Program, 2001
abstract: The ARTEMIS Global Antifungal Susceptibility Program (ARTEMIS Program) was initiated in 2001 to provide focused surveillance of the activities of fluconazole and voriconazole against Candida spp. isolated from blood and other normally sterile sites. A total of 1,586 episodes of infection were detected at 61 international study sites. Overall, 57.7% of the infections were due to Candida albicans, followed by C. glabrata (14.8%), C. parapsilosis (12.5%), C. tropicalis (9.4%), C. krusei (2.7%), and C. lusitaniae (1.5%). Isolates of C. albicans, C. parapsilosis, and C. tropicalis were all highly susceptible to fluconazole (for 99% of the isolates the MICs were ≤8 μg/ml). Likewise, 99 to 100% of these species were inhibited by ≤1 μg of voriconazole per ml. Voriconazole was also active against C. glabrata (93% of the isolates were susceptible [MICs ≤ 1 μg/ml]) and C. krusei (100% of the isolates were susceptible). The agar-based Etest and disk diffusion methods performed well for the testing of both fluconazole and voriconazole compared to the broth microdilution MIC reference method. These observations establish the continued importance of C. albicans as a pathogen and the sustained activity of fluconazole and the broad spectrum of activity of voriconazole and will serve as the first-year benchmark for the ARTEMIS Program. Continued surveillance and refinement of broth- and agar-based test methods will help to identify susceptibility trends and improve the laboratory capability for antifungal susceptibility testing.

title: Development of resistance to 5-fluorocytosine in Candida parapsilosis during therapy.
abstract: After an initially favorable response, infective endocarditis caused by Candida parapsilosis recurred on day 37 of treatment with 5-fluorocytosine. Development of resistance of the organism to 5-fluorocytosine during therapy was documented by in vitro tests (MIC, 6.5 [tg/ml initially, > 129 ptg/ml after relapse). Although strains isolated before treatment and after relapse were able to incorporate exogenous uracil into RNA and were susceptible to 5-fluorouracil, the isolate found after relapse differed from the one isolated before treatment in being unable to incorporate exogenous cytosine and resistant to 5-fluorocytosine. The enzymatic basis of this resistance was identified as a virtual lack of cytosine deaminase activity in the strain isolated after relapse.

title: Comparison of caspofungin and amphotericin B for invasive candidiasis.
abstract: Caspofungin is an echinocandin agent with fungicidal activity against candida species. We performed a double-blind trial to compare caspofungin with amphotericin B deoxycholate for the primary treatment of invasive candidiasis.



We enrolled patients who had clinical evidence of infection and a positive culture for candida species from blood or another site. Patients were stratified according to the severity of disease, as indicated by the Acute Physiology and Chronic Health Evaluation (APACHE II) score, and the presence or absence of neutropenia and were randomly assigned to receive either caspofungin or amphotericin B. The study was designed to compare the efficacy of caspofungin with that of amphotericin B in patients with invasive candidiasis and in a subgroup with candidemia.



Of the 239 patients enrolled, 224 were included in the modified intention-to-treat analysis. Base-line characteristics, including the percentage of patients with neutropenia and the mean APACHE II score, were similar in the two treatment groups. A modified intention-to-treat analysis showed that the efficacy of caspofungin was similar to that of amphotericin B, with successful outcomes in 73.4 percent of the patients treated with caspofungin and in 61.7 percent of those treated with amphotericin B (difference after adjustment for APACHE II score and neutropenic status, 12.7 percentage points; 95.6 percent confidence interval, -0.7 to 26.0). An analysis of patients who met prespecified criteria for evaluation showed that caspofungin was superior, with a favorable response in 80.7 percent of patients, as compared with 64.9 percent of those who received amphotericin B (difference, 15.4 percentage points; 95.6 percent confidence interval, 1.1 to 29.7). Caspofungin was as effective as amphotericin B in patients who had candidemia, with a favorable response in 71.7 percent and 62.8 percent of patients, respectively (difference, 10.0 percentage points; 95.0 percent confidence interval, -4.5 to 24.5). There were significantly fewer drug-related adverse events in the caspofungin group than in the amphotericin B group.



Caspofungin is at least as effective as amphotericin B for the treatment of invasive candidiasis and, more specifically, candidemia.

title: Current treatment strategies for disseminated candidiasis.
abstract: The incidence of disseminated candidiasis has increased dramatically over the past several decades. Fortunately, in recent years, a variety of new antifungal agents have become available to treat these infections. On the basis of efficacy, safety, and cost considerations, fluconazole is the agent of choice for the empirical treatment of disseminated candidiasis in nonneutropenic, hemodynamically stable patients, unless a patient is suspected to be infected with an azole-resistant species (i.e., Candida glabrata or Candida krusei). For hemodynamically unstable or neutropenic patients, agents with broader species coverage, such as polyenes, echinocandins, or, possibly, voriconazole, are preferred for empirical treatment of candidemia. Modification of the initial, empirical regimen depends on the response to therapy and the subsequent identification of the species of the offending pathogen. Echinocandins or high-dose polyenes are preferred for the treatment of infections with C. glabrata or C. krusei. Central venous catheters should be removed from all patients who have disseminated candidiasis, if feasible, and antifungal therapy should be administered to all patients who have candidemia or proven candidiasis.

title: Effects of Caspofungin against Candida guilliermondii and Candida parapsilosis
abstract: The in vitro activity of caspofungin (CAS) was investigated against 28 yeast isolates belonging to Candida albicans (n = 5), Candida guilliermondii (n = 10), and Candida parapsilosis (n = 13). CAS MICs obtained by broth dilution and Etest methods clearly showed a rank order of susceptibility to the echinocandin compound with C. albicans > C. parapsilosis > C. guilliermondii. Similarly, time-kill assays performed on selected isolates showed that CAS was fungistatic against C. albicans and C. parapsilosis, while it did not exert any activity against C. guilliermondii. In a murine model of systemic candidiasis, CAS given at doses as low as 1 mg/kg of body weight/day was effective at reducing the kidney burden of mice infected with either C. albicans or C. guilliermondii isolates. Depending on the isolate tested, mice infected with C. parapsilosis responded to CAS given at 1 and/or 5 mg/kg/day. However, the overall CFU reduction for C. guilliermondii and C. parapsilosis was approximately 100-fold less than that for C. albicans. Our study shows that CAS was active in experimental systemic candidiasis due to C. guilliermondii and C. parapsilosis, but this activity required relatively high drug dosages.

title: Caspofungin in Combination with Amphotericin B against Candida parapsilosis
abstract: Candida parapsilosis has emerged as an important nosocomial pathogen. In the present study, a checkerboard broth microdilution method was performed to investigate the in vitro activities of caspofungin (CAS) in combination with amphotericin B (AMB) against three clinical isolates of C. parapsilosis. Although there was a significant reduction of the MIC of one or both drugs used in combination, an indifferent interaction (fractional inhibitory concentration index greater than 0.50 and less than or equal to 4.0) was observed in 100% of cases. This finding was confirmed by killing curve studies. By a disk diffusion assay, the halo diameters produced by antifungal agents in combination were often significantly greater than those produced by each drug alone. Antagonism was never observed. In a murine model of systemic candidiasis, CAS at either 0.25 or 1 mg/kg/day combined with AMB at 1 mg/kg/day was significantly more effective than each single drug at reducing the colony counts in kidneys. Higher doses of the echinocandin (i.e., 5 and 10 mg/kg/day) combined with the polyene did not show any advantage over CAS alone. Overall, our study showed a positive interaction of CAS and AMB against C. parapsilosis.

title: In Vitro Activities of Anidulafungin against More than 2,500 Clinical Isolates of Candida spp., Including 315 Isolates Resistant to Fluconazole
abstract: Anidulafungin is an echinocandin antifungal agent with potent activity against Candida spp. We assessed the in vitro activity of anidulafungin against 2,235 clinical isolates of Candida spp. using the CLSI broth microdilution method. Anidulafungin was very active against Candida spp. (the MIC at which 90% of strains are inhibited [MIC90] was 2 μg/ml when MIC endpoint criteria of partial inhibition [MIC-2] were used). Candida albicans, C. glabrata, C. tropicalis, C. krusei, and C. kefyr were the most susceptible species of Candida (MIC90, 0.06 to 0.12 μg/ml), and C. parapsilosis, C. lusitaniae, and C. guilliermondii were the least susceptible (MIC90, 0.5 to 2 μg/ml). In addition, 315 fluconazole-resistant isolates were tested, and 99% were inhibited by ≤1 μg/ml of anidulafungin. These results provide further evidence for the spectrum and potency of anidulafungin activity against a large and geographically diverse collection of clinically important isolates of Candida spp.

title: Evaluation of the echinocandin antifungal MK-0991 (L-743,872): efficacies in mouse models of disseminated aspergillosis, candidiasis, and cryptococcosis
abstract: The in vivo activity of the Merck antifungal echinocandin drug candidate MK-0991 (L-743,872) was evaluated in mouse models of disseminated candidiasis, aspergillosis, and cryptococcosis. The echinocandins are potent inhibitors of 1,3-beta-D-glucan synthase. Two models of disseminated candidiasis were used. In a Candida albicans mouse survival model with both DBA/2N and CD-1 mice, estimates of the 50% effective doses (ED50s) of MK-0991 were 0.04 and 0.10 mg/kg of body weight/dose at 21 days after challenge, respectively. In a C. albicans target organ assay (TOA) with DBA/2N mice, MK-0991 at levels of > or =0.09 mg/kg/dose significantly reduced the numbers of C. albicans CFU/g of kidneys compared to the numbers in the kidneys of control mice from 1 to 28 days after challenge. Even when given as a single intraperitoneal dose either 30 min or 24 h after challenge, MK-0991 was effective and significantly reduced the numbers of C. albicans CFU/g of kidney compared to those in the controls. MK-0991 was >300-fold less active when it was administered orally than when it was administered parenterally. MK-0991 was efficacious in mouse TOAs against other C. albicans strains and Candida species including Candida tropicalis, Candida (Torulopsis) glabrata, Candida lusitaniae, Candida parapsilosis, and Candida krusei. MK-0991 was ineffective against disseminated Cryptococcus neoformans infections. In the model of disseminated aspergillosis in mice, MK-0991 at doses of > or =0.02 mg/kg/dose significantly prolonged the survival of DBA/2N mice, with estimates of the ED50 and ED90 of MK-0991 being 0.03 and 0.12 mg/kg/dose, respectively, at 28 days after challenge. MK-0991 is a potent, parenterally administered therapeutic agent against disseminated candidiasis and aspergillosis that warrants further investigation in human clinical trials.

title: Echinocandin antifungal drugs
abstract: The echinocandins are large lipopeptide molecules that are inhibitors of beta-(1,3)-glucan synthesis, an action that damages fungal cell walls. In vitro and in vivo, the echinocandins are rapidly fungicidal against most Candida spp and fungistatic against Aspergillus spp. They are not active at clinically relevant concentrations against Zygomycetes, Cryptococcus neoformans, or Fusarium spp. No drug target is present in mammalian cells. The first of the class to be licensed was caspofungin, for refractory invasive aspergillosis (about 40% response rate) and the second was micafungin. Adverse events are generally mild, including (for caspofungin) local phlebitis, fever, abnormal liver function tests, and mild haemolysis. Poor absorption after oral administration limits use to the intravenous route. Dosing is once daily and drug interactions are few. The echinocandins are widely distributed in the body, and are metabolised by the liver. Results of studies of caspofungin in candidaemia and invasive candidiasis suggest equivalent efficacy to amphotericin B, with substantially fewer toxic effects. Absence of antagonism in combination with other antifungal drugs suggests that combination antifungal therapy could become a general feature of the echinocandins, particularly for invasive aspergillosis.

title: Candida parapsilosis fungaemia treated unsuccessfully with amphotericin B and fluconazole but eliminated with caspofungin: a case report.
abstract: A case of Candida parapsilosis Hickman catheter infection with fungaemia presenting with severe hypoxia, lactic acidosis and renal failure is described. Four months previously, the Hickman catheter had become infected with Candida parapsilosis (sensitive to fluconazole and amphotericin B) with blood cultures remaining positive despite an intravenous course of fluconazole 200 mg 12-hourly increasing to 400 mg-12-hourly. Subsequently, treatment with an amphotericin B infusion (30 mg over 24 hours) for five days was associated with a progressive increase in serum creatinine from 0.042 mmol/L to 0.278 mmol/L with blood cultures remaining positive. While arrangements were being made to remove and replace the catheter via a thoracotomy, the patient developed severe hypoxia, lactic acidosis and renal failure and was admitted to the intensive care unit for further management. An intravenous course of caspofungin (70 mg over 1 hour followed by 50 mg over 1 hour, daily) was associated with negative blood cultures and a normal plasma C-reactive protein for the first time in four months. The hypoxia was managed with spontaneous ventilation and high inspired oxygen concentrations, and the anaemia was treated with red blood cell transfusions. A rapid reversal of the respiratory failure and lactic acidosis occurred and the patient was discharged home from the intensive care unit after 10 days to continue the caspofungin infusions for a total of 6 weeks. She has subsequently remained well with negative blood cultures two months later.

title: [Successful treatment with micafungin in a case of candidemia associated with pneumonia].
abstract: A 63-year-old woman was admitted to our hospital with fever and cough. Candidemia was diagnosed by blood culture and culture of IVH catheter. Although, the patient was treated with fluconazole, clinical symptoms and chest radiographic findings worsened. After micafungin was replaced with fluconazole, her symptoms, chest radiographic findings improved and stabilized. It is suggested that micafungin is useful for the treatment of candidemia associated with Candida parapsilosis.

title: Caspofungin Therapy of Neonates With Invasive Candidiasis
abstract: Invasive candidiasis is an increasing problem in neonatal intensive care units worldwide and is an important cause of morbidity, mortality and prolongation of hospital stay. Despite administration of amphotericin B, invasive candidiasis in neonates is sometimes complicated by persistent fungemia and refractory invasive candidiasis. The problem has been augmented by the increasing prevalence of non-albicans species that often are resistant to fluconazole and to amphotericin B.  The population consisted of 1 term and 9 premature neonates with invasive candidiasis caused by Candida albicans (n = 4), Candida parapsilosis (n = 3), Candida tropicalis (n = 2) and Candida glabrata (n = 1). Despite initial therapy with deoxycholate amphotericin B, blood cultures remained positive in all patients for 13–49 days. Invasive candidiasis progressed to meningitis and enlarging renal Candida bezoars in the kidney of one patient and an enlarging atrial vegetation in another. Another patient developed severe hypokalemia refractory to potassium supplementation. Two of the C. albicans and all of the non-albicans Candida isolates were resistant to fluconazole; the C. glabrata isolate was resistant to amphotericin B. Amphotericin B was discontinued and caspofungin initiated in all patients in a dosage of 1 mg/kg/d for 2 days followed by 2 mg/kg/d.  All positive blood cultures cleared between 3 and 7 days after initiation of caspofungin, the atrial vegetation resolved and the renal Candida bezoars disappeared. Renal and hepatic function tests did not show any values above normal throughout caspofungin therapy. There were no attributable clinical adverse events during the administration of caspofungin in any of the patients.  Caspofungin was effective, safe and well-tolerated as an alternative therapy for persistent and progressive candidiasis in those neonates who were unresponsive to or intolerant of deoxycholate amphotericin B.

title: Caspofungin Susceptibility Testing of Isolates from Patients with Esophageal Candidiasis or Invasive Candidiasis: Relationship of MIC to Treatment Outcome
abstract: The caspofungin clinical trial database offers an opportunity to assess susceptibility results for Candida pathogens obtained from patients with candidiasis and allows for correlations between efficacy outcomes and MICs. Candida isolates have been identified from patients enrolled in four studies of esophageal candidiasis and two studies of invasive candidiasis. The MICs of caspofungin for all baseline isolates were measured at a central laboratory using NCCLS criteria (document M-27A); MICs for caspofungin were defined as the lowest concentration inhibiting prominent growth at 24 h. MICs were then compared to clinical and microbiological outcomes across the two diseases. Susceptibility testing for caspofungin was performed on 515 unique baseline isolates of Candida spp. obtained from patients with esophageal candidiasis. MICs for caspofungin ranged from 0.008 to 4 μg/ml; the MIC50 and MIC90 were 0.5 and 1.0 μg/ml, respectively. Susceptibility testing was also performed on 231 unique baseline isolates of Candida spp. from patients with invasive candidiasis. The majority (∼96%) of MICs were between 0.125 and 2 μg/ml, with MIC50 and MIC90 for caspofungin being 0.5 and 2.0 μg/ml, respectively. Overall, caspofungin demonstrated potent in vitro activity against clinical isolates of Candida species. A relationship between MIC for caspofungin and treatment outcome was not seen for patients with either esophageal candidiasis or invasive candidiasis. Patients with isolates for which the MICs were highest (>2 μg/ml) had better outcomes than patients with isolates for which the MICs were lower (<1 μg/ml). Additionally, no correlation between MIC and outcome was identified for specific Candida species.

title: Interlaboratory Comparison of Results of Susceptibility Testing with Caspofungin against Candida and Aspergillus Species
abstract: Seventeen laboratories participated in a study of interlaboratory reproducibility with caspofungin microdilution susceptibility testing against panels comprising 30 isolates of Candida spp. and 20 isolates of Aspergillus spp. The laboratories used materials supplied from a single source to determine the influence of growth medium (RPMI 1640 with or without glucose additions and antibiotic medium 3 [AM3]), the same incubation times (24 h and 48 h), and the same end point definition (partial or complete inhibition of growth) for the MIC of caspofungin. All tests were run in duplicate, and end points were determined both spectrophotometrically and visually. The results from almost all of the laboratories for quality control and reference Candida and Aspergillus isolates tested with fluconazole and itraconazole matched the NCCLS published values. However, considerable interlaboratory variability was seen in the results of the caspofungin tests. For Candida spp. the most consistent MIC data were generated with visual “prominent growth reduction” (MIC2) end points measured at 24 h in RPMI 1640, where 73.3% of results for the 30 isolates tested fell within a mode ± one dilution range across all 17 laboratories. MIC2 at 24 h in RPMI 1640 or AM3 also gave the best interlaboratory separation of Candida isolates of known high and low susceptibility to caspofungin. Reproducibility of MIC data was problematic for caspofungin tests with Aspergillus spp. under all conditions, but the minimal effective concentration end point, defined as the lowest caspofungin concentration yielding conspicuously aberrant hyphal growth, gave excellent reproducibility for data from 14 of the 17 participating laboratories.

title: Early Clinical Experience with Anidulafungin at a Large Tertiary Care Medical Center
abstract: To evaluate early clinical experience with anidulafungin.

title: Paradoxical Effect of Echinocandins across Candida Species In Vitro: Evidence for Echinocandin-Specific and Candida Species-Related Differences
abstract: Paradoxical growth of some Candida isolates occurs at concentrations above the MIC for echinocandins. In 60 Candida bloodstream isolates from cancer patients (20 C. albicans isolates and 10 isolates each of C. parapsilosis, C. tropicalis, C. krusei, and C. glabrata), paradoxical growth was more frequent with caspofungin than micafungin or anidulafungin, was unrelated to MIC, and was strikingly absent in C. glabrata isolates.

title: Increasing incidence of Candida parapsilosis candidemia with caspofungin usage.
abstract: To determine the impact on the change in epidemiology of Candida species at our institution since the introduction of caspofungin.



A 5-year retrospective review of all candidemia at a major tertiary care center. Only one episode of candidemia per patient per admission was counted. All antifungal defined daily doses were also collected in this same time period. Regression analysis was performed on the data and correlation statistics among antifungal use and Candida species were assessed using a Pearson correlation analysis.



There were 469 individual episodes of candidemia between fiscal year 2002 and 2006 with the rate increasing every year. On regression analysis there was a significant increase in Candida parapsilosis candidemia (R(2)=0.90, p=0.02) and significant increase in caspofungin usage (R(2)=0.80, p<0.01), with a correspondingly significant decline in conventional (R(2)=-0.77, p<0.01) and lipid amphotericin B (R(2)=-0.95, p<0.05) usage. We found correlations between increased caspofungin usage (R(2)=0.94, p=0.017) and increased C. parapsilosis candidemia and decreased Candida tropicalis candidemia (R(2)=0.92, p<0.05) and a trend towards decreased Candida glabrata (R(2)=0.64, p=0.1).



We showed significant correlations between increased caspofungin usage and an increased incidence of C. parapsilosis candidemia and reduction in C. tropicalis candidemia, with a trend towards less C. glabrata candidemia.

title: Combination Treatment of Invasive Fungal Infections
abstract: The persistence of high morbidity and mortality from systemic fungal infections despite the availability of novel antifungals points to the need for effective treatment strategies. Treatment of invasive fungal infections is often hampered by drug toxicity, tolerability, and specificity issues, and added complications often arise due to the lack of diagnostic tests and to treatment complexities. Combination therapy has been suggested as a possible approach to improve treatment outcome. In this article, we undertake a historical review of studies of combination therapy and also focus on recent studies involving newly approved antifungal agents. The limitations surrounding antifungal combinations include nonuniform interpretation criteria, inability to predict the likelihood of clinical success, strain variability, and variations in pharmacodynamic/pharmacokinetic properties of antifungals used in combination. The issue of antagonism between polyenes and azoles is beginning to be addressed, but data regarding other drug combinations are not adequate for us to draw definite conclusions. However, recent data have identified potentially useful combinations. Standardization of assay methods and adoption of common interpretive criteria are essential to avoid discrepancies between different in vitro studies. Larger clinical trials are needed to assess whether combination therapy improves survival and treatment outcome in the most seriously debilitated patients afflicted with life-threatening fungal infections.

title: Successful treatment of Candida parapsilosis mural endocarditis with combined caspofungin and voriconazole
abstract: BackgroundFungal mural endocarditis is a rare entity in which the antemortem diagnosis is seldom made. Seven cases of mural endocarditis caused by Candida spp. have been collected from literature and six of these patients died after treatment with amphotericin B.Case presentationWe report a case of mural endocarditis diagnosed by transesophageal echocardiogram and positive blood cultures to Candida parapsilosis. Because blood cultures continued to yield C. parapsilosis despite caspofungin monotherapy, treatment with voriconazole was added.ConclusionThis is the first description of successful treatment of C. parapsilosis mural endocarditis with caspofungin and voriconazole.

title: Three distinct genotypes within Candida parapsilosis from clinical sources
abstract: Three genetically distinct groups of Candida parapsilosis were detected among clinical isolates. These were distinguishable on the basis of isoenzyme profiles and DNA sequences of internally transcribed spacer (ITS) sequences flanking the 5.8S RNA gene. In an investigation of 45 strains, including 32 clinical isolates from Texas, C. parapsilosis group I composed the majority of the common clinical isolates. The type strain of C. parapsilosis was a member of this group. The 10 group II isolates were indistinguishable from group I strains when tested with the API 20C kit. The two group III isolates differed from those in groups I and II by being D-xylitol positive by the API 20C kit; however, isolates in all groups assimilated D-xylitol from broth. Isoenzyme profiles excluded the close relationship of any of these groups to Lodderomyces elongisporus, which is a teleomorphic yeast that has a physiological profile similar to that of C. parapsilosis. Although there were insignificant differences in the ITS2 rDNA sequences, comparisons of the ITS1 sequences revealed several differences. A sequence analysis of ITS1 in which missing bases were counted as mismatches showed the following similarities: group I versus group II, 87.7%; group I versus group III, 82.1%; group II versus group III, 84.5%. Also, the activity of secreted proteinase showed differences among the three groups, with many group I isolates having moderate to high activity. The degree of susceptibility to antifungal agents, amphotericin B, ketoconazole, and 5-fluorocytosine, could not be used to determine an isolate's group.

title: Genomic heterogeneity in the yeast Candida parapsilosis
abstract: Candida parapsilosis shows a wide intraspecies variation in chromosome/homolog size distribution. As a prerequisite for delineating modes of transmission, we have undertaken an analysis of genetic variation at different levels. In the present study we have observed that a majority of isolates display similar electrophoretic karyotype patterns consistent for the species, with variations in the smaller group of chromosomes. In two strains we observed phenotypic “switching”; one of these also exhibited a mixed karyotypic subpopulation. In contrast, a few isolates displayed a greater degree of chromosome/ homolog size variation. We also observed, through randomly amplified polymorphic DNA (RAPD) analysis, results consistent with those of pulsed-field electrophoresis. Isolates displaying a high degree of chromosome/homolog variation also displayed a high degree of variation in genomic “fingerprints”. Polymorphisms, although present, were much reduced in the majority of isolates. These parallel observations suggest a common underlying mechanism. Our results are consistent with the hypothesis that chromosome-sized variations in C. parapsilosis are due to random genetic events. A similar mechanism has been hypothesized for the taxonomically related yeast Candida albicans.

title: Identification of clinically important ascomycetous yeasts based on nucleotide divergence in the 5' end of the large-subunit (26S) ribosomal DNA gene
abstract: Clinically important species of Candida and related organisms were compared for extent of nucleotide divergence in the 5' end of the large-subunit (26S) ribosomal DNA (rDNA) gene. This rDNA region is sufficiently variable to allow reliable separation of all known clinically significant yeast species. Of the 204 described species examined, 21 appeared to be synonyms of previously described organisms. Phylogenetic relationships among the species are presented.

title: Confirmation of the Distinct Genotype Groups within the Form Species Candida parapsilosis
abstract: The DNA relatedness and restriction fragment length polymorphism patterns of whole-cell DNA and the general phenotypic properties of 14 isolates of the form species Candida parapsilosis representing three diverse genetic groups were compared. The data confirm the unrelatedness of the three groups at the species level.

title: Phylogenetic relationship and mode of evolution of yeast DNA topoisomerase II gene in the pathogenic Candida species.
abstract: We have determined the nucleotide sequences of about 55% of the region of the DNA topoisomerase II gene (approximately 2.3 kb) isolated from the pathogenic Candida species, C. dubliniensis, C. parapsilosis, C. tropicalis, C. krusei, C. kefyr, C. guilliermondii and C. lusitaniae. Evolutionary relationships among nine Candida species including those of C. albicans and C. glabrata were studied based on the DNA topoisomerase II gene. The nucleotide sequences of 2192 bp, which covered two catalytic domains, ATPase and cutting/resealing, were subjected to phylogenetic analysis. Sequence comparison and evolutionary analysis have revealed that the Candida species tested here are not monophyletic, and the two strains within the species C. tropicalis and C. parapsilosis are too diverse to be in a single species. A wide variety of divergence was observed among the functional domains of DNA topoisomerase II, suggesting that Candida species were in different evolutionary paths at least as regarding the DNA topoisomerase II gene. Sequence information and the observation on the species-specific manner of molecular evolution of DNA topoisomerase II in Candida will be applied to develop a method of identification and characterization of the Candida species in both natural and clinical isolates.

title: Variations in DNA subtype, antifungal susceptibility, and slime production among clinical isolates of Candida parapsilosis.
abstract: Candida parapsilosis is an important nosocomial pathogen that can proliferate in high concentrations of glucose and form biofilms on prosthetic materials. We investigated the genotypic diversity, slime production, and antifungal susceptibility among 60 isolates of C. parapsilosis from 44 patients and 10 patient care providers from five different medical centers. Molecular typing was performed using macrorestriction digest profiles with BssHII followed by pulsed-field gel electrophoresis (REAG) and by electrophoretic karyotyping (EK). Slime production was evaluated by growing the organisms in Sabouraud broth with 8% glucose and examining the walls of the tubes for the presence of an adherent slime layer. Antifungal susceptibility to amphotericin B, 5-fluorocytosine, fluconazole, and itraconazole was determined using National Committee for Clinical Laboratory Standards proposed standard methods. Overall 28 different DNA types were identified by REAG and EK methods. MIC90 values ranged from 0.12 microgram/ml for itraconazole to 1.0 microgram/ml for fluconazole and amphotericin B. Sixty-five percent of the isolates produced slime: 37% were moderately to strongly positive, 28% were weakly positive, and 35% were negative. Overall, 83% of blood and catheter isolates were slime positive versus 53% of isolates from all other sites (P < 0.05). These data underscore the genetic diversity and susceptibility of C. parapsilosis to antifungal agents. Slime production may be important in enabling C. parapsilosis to cause catheter-related bloodstream infections.

title: Linear versus circular mitochondrial genomes: intraspecies variability of mitochondrial genome architecture in Candida parapsilosis.
abstract: The yeast species Candida parapsilosis, an opportunistic pathogen, exhibits genetic and genomic heterogeneity. To assess the polymorphism at the level of mitochondrial DNA (mtDNA), the organization of the mitochondrial genome in strains belonging to the three variant groups of this species was investigated. Although these analyses revealed a group-specific restriction fragment pattern of mtDNA, strains belonging to different groups appear to have similar genes in the same gene order. An extensive survey of C. parapsilosis isolates uncovered surprising alterations in the molecular architecture of their mitochondrial genome. A screening strategy for strains harbouring mtDNA with rearranged architecture showed that nearly all strains from groups I and III possess linear mtDNA molecules terminating with arrays of tandem repeat units, while most of the group II strains have a circular mitochondrial genome. In addition, it was found that linear genophores in mitochondria of strains from different groups differ in the sequence of the mitochondrial telomeric repeat unit. The occurrence of altered forms of mtDNA among C. parapsilosis strains opens up the unique possibility to address questions concerning the evolutionary origin and replication strategy of linear and circular genomes in mitochondria.

title: A yeast with linear molecules of mitochondrial DNA
abstract: SummaryMitochondrial DNA from the yeast strain SR23, tentatively allocated to the species, Candida rhagii, consists of linear molecules 30 kb long. This has been demonstrated by restriction analysis and selective radioactive labelling of terminal restriction fragments. Preliminary sequence analysis indicated that the two ends of the molecule are formed by inverted repeats. The arrangement of several genes in the mitochondrial genome of C. rhagii SR23 was established by specific hybridisation with probes prepared from mitochondrial DNA of Saccharomyces cerevisiae. The arrangement is unique, with genes coding for the two ribosomal RNAs placed widely apart. Intron(s) may be present in the gene coding for cytochrome b.

title: Linear mitochondrial DNAs from yeasts: telomeres with large tandem repetitions
abstract: The terminal structure of the linear mitochondrial DNA (mtDNA) from the yeast Candida parapsilosis was investigated. This mtDNA, 30 kb long, has symmetrical ends forming inverted terminal repeats. These repeats are made up of a variable number of tandemly repeating units of 738 by each; the terminal nucleotide corresponds to a precise position within the last repeat unit sequence. The ends had an open structure accessible to enzymes, with a 5′ single-stranded extension of about 110 nucleotides. No circular forms were detected in the DNA preparations. Two other unrelated species, Pichia philodendra and Candida salmanticensis also appear to have a linear mtDNA of similar organization. These linear DNAs (which we name Type 2 linear mtDNAs) are distinct from the previously described linear mtDNAs of yeasts whose termini are formed by a closed hairpin loop (Type 1 linear mtDNA). The terminal structure of C. parapsilosis mtDNA is reminiscent of the linear mitochondrial genomes of the ciliate Tetrahymena although, in the latter, the telomeric tandem repeat unit is considerably shorter.

title: Mitochondrial Telomeres as Molecular Markers for Identification of the Opportunistic Yeast Pathogen Candida parapsilosis
abstract: Recent studies have demonstrated that a large number of organisms carry linear mitochondrial DNA molecules possessing specialized telomeric structures at their ends. Based on this specific structural feature of linear mitochondrial genomes, we have developed an approach for identification of the opportunistic yeast pathogen Candida parapsilosis. The strategy for identification of C. parapsilosis strains is based on PCR amplification of specific DNA sequences derived from the mitochondrial telomere region. This assay is complemented by immunodetection of a protein component of mitochondrial telomeres. The results demonstrate that mitochondrial telomeres represent specific molecular markers with potential applications in yeast diagnostics and taxonomy.

title: Complete DNA sequence of the linear mitochondrial genome of the pathogenic yeast Candida parapsilosis
abstract: The complete sequence of the mitochondrial DNA of the opportunistic yeast pathogen Candida parapsilosis was determined. The mitochondrial genome is represented by linear DNA molecules terminating with tandem repeats of a 738-bp unit. The number of repeats varies, thus generating a population of linear DNA molecules that are heterogeneous in size. The length of the shortest molecules is 30,922 bp, whereas the longer molecules have expanded terminal tandem arrays (n×738 bp). The mitochondrial genome is highly compact, with less than 8% of the sequence corresponding to non-coding intergenic spacers. In silico analysis predicted genes encoding fourteen protein subunits of complexes of the respiratory chain and ATP synthase, rRNAs of the large and small subunits of the mitochondrial ribosome, and twenty-four transfer RNAs. These genes are organized into two transcription units. In addition, six intronic ORFs coding for homologues of RNA maturase, reverse transcriptase and DNA endonucleases were identified. In contrast to its overall molecular architecture, the coding sequences of the linear mitochondrial DNA of C. parapsilosis are highly similar to their counterparts in the circular mitochondrial genome of its close relative C. albicans. The complete sequence has implications for both mitochondrial DNA replication and the evolution of linear DNA genomes.

title: A Genome Sequence Survey Shows that the Pathogenic Yeast Candida parapsilosis Has a Defective MTLa1 Allele at Its Mating Type Locus
abstract: Candida parapsilosis is responsible for ca. 15% of Candida infections and is of particular concern in neonates and surgical intensive care patients. The related species Candida albicans has recently been shown to possess a functional mating pathway. To analyze the analogous pathway in C. parapsilosis, we carried out a genome sequence survey of the type strain. We identified ca. 3,900 genes, with an average amino acid identity of 59% with C. albicans. Of these, 23 are predicted to be predominantly involved in mating. We identified a genomic locus homologous to the MTLa mating type locus of C. albicans, but the C. parapsilosis type strain has at least two internal stop codons in the MTLa1 open reading frame, and two predicted introns are not spliced. These stop codons were present in MTLa1 of all eight C. parapsilosis isolates tested. Furthermore, we found that all isolates of C. parapsilosis tested appear to contain only the MTLa idiomorph at the presumptive mating locus, unlike C. albicans and C. dubliniensis. MTLα sequences are present but at a different chromosomal location. It is therefore likely that all (or at least the majority) of C. parapsilosis isolates have a mating pathway that is either defective or substantially different from that of C. albicans.

title: Multilocus Genotypes and DNA Fingerprints Do Not Predict Variation in Azole Resistance among Clinical Isolates ofCandida albicans
abstract: If variation in azole resistance is due to inherent differences in strains of Candida albicans, as a predominantly clonal organism, then correlation between multilocus genotypes and drug resistance would be expected. A sample of 81 clinical isolates from patients infected with human immunodeficiency virus in Toronto, Canada, plus 3 reference isolates were genotyped at 16 loci, distributed on all linkage groups, by means of oligonucleotide hybridizations specific for each of the alleles at each locus. These multilocus genotypes were significantly correlated with DNA fingerprints obtained with the species-specific probe 27A, indicating widespread linkage disequilibrium in the genome. There were 64 multilocus diploid genotypes and 77 DNA fingerprint types delineated in this sample. Neither the multilocus genotyping nor DNA fingerprinting alone identified all of the 81 types identified by the combination of these two methods. Multilocus genotypes were not predictive of fluconazole resistance, suggesting that resistance is gained or lost too quickly to be predicted by linkage with neutral markers.

title: Optimization and Validation of Multilocus Sequence Typing for Candida albicans
abstract: Multilocus sequence typing (MLST) was applied to 75 Candida albicans isolates, including 2 that were expected to be identical, 48 that came from diverse geographical and clinical sources, and 15 that were sequential isolates from two patients. DNA fragments (≈500 bp) of eight genes encoding housekeeping functions were sequenced, including four that have been described before for C. albicans MLST, and four new gene fragments, AAT1a, AAT1b, MPI, and ZWF1. In total, 87 polymorphic sites were found among 50 notionally different isolates, giving 46 unique sequence types, underlining the power of MLST to differentiate isolates for epidemiological studies. Additional typing information was obtained by detecting variations in size at the transcribed spacer region of the 25S rRNA gene and tests for homozygosity at the mating type-like (MTL) locus. The stability of MLST was confirmed in two sets of consecutive isolates from two patients. In each set the isolates were identical or varied by a single nucleotide. Reference strain SC5314 and a derived mutant, CAF2, gave identical MLST types. Heterozygous polymorphisms were found in at least one isolate for all but 16 (18.4%) of the variable nucleotides, and 35 (41%) of the 87 individual sequence changes generated nonsynonymous amino acids. Cloning and restriction digestion of a gene fragment containing heterozygous polymorphisms indicated that the heterozygosity was genuine and not the result of sequencing errors. Our data validate and extend previous MLST results for C. albicans, and we propose an optimized system based on sequencing eight gene fragments for routine MLST with this species.

title: Evidence for aneuploidy and recombination in the human commensal yeast Candida parapsilosis.
abstract: Isolates of Candida parapsilosis, including representatives of the three major sub-species groups, were screened for single nucleotide polymorphisms (SNPs) by sequencing five independent loci totaling 4kb per isolate. Group I isolates were highly conserved and in some cases, group I alleles were found in group II and III strains. Unique alleles were also associated with groups II and III, consistent with earlier observations of intergroup divergence. There was no heterozygosity in any strain, and a FACS analysis demonstrated that for all three groups nuclei are variant in size, ranging from 0.5 to 1.0 x the size of other diploid yeast genomes. This suggests that natural isolates of C. parapsilosis are aneuploid, with some isolates being essentially haploid. Taken collectively with the observation of group I alleles within group II and III strains, we propose that some form of recombination is occurring between groups.

title: Occurrence and genetic variability of Candida parapsilosis sensu lato in Hungary.
abstract: The occurrence and genetic variability of Candida parapsilosis isolates in two Hungarian hospitals, located in Debrecen and Pécs, were examined. Among the 209 Candida isolates examined, 20 were found to belong to C. parapsilosis sensu lato, based on morphological, physiological and molecular data. The frequency of occurrence of C. parapsilosis isolates (9.6%) was lower than that observed in Europe but higher than that observed previously in Hungary. The genetic variability of C. parapsilosis sensu lato isolates was also examined using random amplified polymorphic DNA (RAPD) analysis and sequence analysis of the intergenic transcribed spacer (ITS) region of the rRNA gene cluster. The genetic variability of the isolates was relatively high, as revealed by RAPD analysis. Two isolates were found to belong to the recently described Candida metapsilosis species (C. parapsilosis group III), based on ITS sequence data, RAPD analysis and phenotypic data. These two isolates could also be distinguished from C. parapsilosis sensu stricto isolates using a primer pair developed for the detection of C. parapsilosis group I isolates. To the best of the authors' knowledge, this is the first report on the identification of C. metapsilosis from bloodstream infection.

title: Genetic manipulation of the pathogenic yeast Candida parapsilosis
abstract: Candida parapsilosis is an important human pathogen, responsible for severe cases of systemic can-didiasis and one of the leading causes of mortality in neonates. In this report, we describe the first system for genetic manipulation of C. parapsilosis. We isolated and subsequently determined DNA sequences of genes encoding galactokinase (CpGALl) and orotidine-5′-phos-phate decarboxylase (CpURA3) from a genomic DNA library of C. parapsilosis by functional complementation of corresponding mutations in Saccharomyces cerevisiae. The predicted protein products, Gallp and Ura3p, displayed a high degree of homology with corresponding sequences of C. albicans and S. cerevisiae, respectively. A collection of galactokinase-deficient (gall) strains of C. parapsilosis was prepared using direct selection of mutagenized cells on media containing 2-deoxy-galac-tose. Additionally, we constructed a plasmid vector carrying CpGALl as a selection marker and a genomic DNA fragment with an autonomously replicating sequence activity that transforms the C. parapsilosis gall mutant strain with high efficiency. This system for genetic transformation of C. parapsilosis may significantly advance the study of this human pathogen, greatly improving our understanding of its biology and virulence, with implications for drug development.

title: The SAT1 flipper, an optimized tool for gene disruption in Candida albicans.
abstract: The construction of Candida albicans mutants by targeted gene disruption usually depends on the use of nutritional markers for the selection of prototrophic transformants from auxotrophic host strains, but it is becoming increasingly evident that this strategy may cause difficulties in the interpretation of mutant phenotypes. Here, we describe a new method for inactivating both alleles of a target gene in C. albicans wild-type strains to obtain homozygous null mutants. The SAT1 flipping method relies on the use of a cassette that contains a dominant nourseothricin resistance marker (caSAT1) for the selection of integrative transformants and a C. albicans-adapted FLP gene that allows the subsequent excision of the cassette, which is flanked by FLP target sequences, from the genome. Two rounds of integration/excision generate homozygous mutants that differ from the wild-type parent strain only by the absence of the target gene, and reintegration of an intact gene copy for complementation of mutant phenotypes is performed in the same way. Transformants are obtained after only 1 day of growth on a selective medium, and integration into the target locus occurs with high specificity after adding homologous flanking sequences on both sides of the cassette. FLP-mediated excision of the SAT1 flipper cassette is achieved by simply growing the transformants for a few hours in medium without selective pressure, and nourseothricin-sensitive (NouS) derivatives can easily be identified by their slower growth on indicator plates containing a low concentration of nourseothricin. We demonstrate the use of the system by deleting the OPT1 gene, which encodes an oligopeptide transporter, in the C. albicans model strain SC5314. The null mutants became resistant to the toxic peptide KLLEth, and reintroduction of an intact OPT1 copy restored susceptibility. The SAT1 flipping method provides a highly efficient method for gene disruption in C. albicans wild-type strains, which eliminates currently encountered problems in the genetic analysis of this important human fungal pathogen.