bluuebunny/crossref_metadata_2025 huggingface dataset:
Reverse transcription-PCR detection of LaCrosse virus in mosquitoes and comparison with enzyme immunoassay and virus isolation
Authors: Wasieloski L P , Rayms-Keller A , Curtis L A , Blair C D , Beaty B J
Published: January 2020

A reverse transcription-PCR (RT-PCR) assay was developed and compared with enzyme immunoassay (EIA) and virus isolation for detecting LaCrosse virus (LAC) in mosquito pools. All three techniques were able to detect a single LAC-infected mosquito in a pool of 99 negative mosquitoes. Virus isolation was the most sensitive of the three techniques; it was possible to isolate virus immediately following intrathoracic inoculation of mosquitoes. RT-PCR was second in sensitivity; LAC RNA was detected 1 day postinfection. EIA detected LAC antigen 2 days postinfection. Additionally, RT-PCR and EIA were able to detect LAC RNA and protein, respectively, from mosquito samples which were subjected to seven freeze-thaw cycles, and RT-PCR was able to detect LAC RNA from mosquito samples which remained at room temperature for up to 7 days.
Detection of Ehrlichia risticii from feces of infected horses by immunomagnetic separation and PCR
Authors: Biswas B , Vemulapalli R , Dutta S K
Published: January 2020

Potomac horse fever, caused by Ehrlichia risticii, is an important disease of equines. The major features of the disease are fever, leukopenia, and diarrhea. The organism has been detected from the blood mononuclear cells of infected horses, but its presence in the feces has not been known. A method for immunomagnetic separation of E. risticii from the feces of infected horses was developed, and the separated organisms were detected by PCR. Coating immunomagnetic beads (Dynabeads) with a 1:5 dilution of rabbit anti-E. risticii serum and incubating the Dynabeads with fecal samples for 25 min at room temperature gave optimum results. E. risticii was detected from the feces during the course of diarrhea from two experimentally infected horses. In horse 1, watery diarrhea occurred from days 11 to 16 postinfection (p.i.), after which the feces became soft on day 17 p.i. and then returned to normal. The organisms were first detected from the feces on day 11 p.i., peaked on day 13 p.i., and then gradually decreased until day 16 p.i., after which they became undetectable. In horse 2, first, on day 12 p.i., there was soft feces which continued and progressed to diarrhea on day 17 p.i. The feces became normal after day 18 p.i. The organisms in the feces of this horse were first detected on day 12 p.i. and peaked on day 14 p.i., after which they declined until day 16 p.i. and then became undetectable. In both horses, the number of organisms in the mononuclear cells peaked on days 10 and 11 p.i., respectively, 3 days prior to the respective peaks in the feces. E. risticii was not detected from the plasma samples obtained from these horses. There was a drastic reduction in PCR amplification of E. risticii DNA for fecal samples stored frozen at -20 degrees C in comparison with those stored at 4 degrees C. The presence of the organism in the feces only during the soft- or diarrheal-feces phase supports the previous hypothesis that the diarrhea is caused by the organisms replicating in cells lining the intestines. This rapid simple method of detection of the organisms from the feces will be helpful in diagnostic and epidemiologic studies of Potomac horse fever.
Simultaneous culture for adenovirus, cytomegalovirus, and herpes simplex virus in same shell vial by using three-color fluorescence
Authors: Brumback B G , Wade C D
Published: January 2020

A simultaneous triple culture for adenovirus, cytomegalovirus, and herpes simplex virus, using three different fluorochromes for virus detection and differentiation in the same shell vial, was developed. In evaluations the triple culture performed comparably to separate cultures, required less specimen volume, and was less expensive to perform.
Detection of human parvovirus B19 DNA PCR products by RNA probe hybridization enzyme immunoassay
Authors: Erdman D D , Durigon E L , Holloway B P
Published: January 2020

We have developed an RNA probe hybridization enzyme immunoassay for detection of human parvovirus B19 PCR-amplified DNA. The assay is easy to perform and increases assay sensitivity without the added inconvenience and risk of false-positive results associated with nested PCR.
First report of involvement of Nodulisporium species in human disease
Authors: Cox G M , Schell W A , Scher R L , Perfect J R
Published: January 2020

Allergic fungal sinusitis is a common disease that results from a hypersensitivity reaction mounted by the host against fungi living in the paranasal sinuses. We have recently treated a patient with allergic fungal sinusitis due to a Nodulisporium species. This is the first description of a Nodulisporium species involved in human disease. The genus Nodulisporium contains both dematiaceous and nondematiaceous members. These fungi occur worldwide in nature, often as accompanying conidial anamorphs of certain wood decay ascomycetes. Clinical mycology laboratories may encounter this new agent of phaeohyphomycosis.
Effect of potential interference factors on performance of enzyme immunoassay and latex agglutination assay for cryptococcal antigen
Authors: Engler H D , Shea Y R
Published: January 2020

The PREMIER Cryptococcal Antigen Enzyme Immunoassay (Meridian Diagnostics) did not give discrepant results with rheumatoid factor, syneresis fluid, or serum macroglobulins from systemic lupus erythematosis patients. The Cryptococcal Antigen Latex Agglutination System (Meridian Diagnostics) did cross-react with syneresis fluid but not with the other serum factors tested.
Hafnia alvei in stool specimens from patients with diarrhea and healthy controls
Authors: Ridell J , Siitonen A , Paulin L , Mattila L , Korkeala H , Albert M J
Published: January 2020

We found an epidemiological association of Hafnia alvei with diarrhea, because the organism was isolated from 12 of 77 (16%) adult Finnish tourists to Morocco who developed diarrhea and from 0 of 321 tourists without diarrhea (P < 0.001). From another group of 112 adult Finnish diarrheal patients, only 2 (2%) yielded H. alvei. In contrast to some Bangladeshi strains of H. alvei, the Finnish strains were negative for the attachment-effacement lesion by an in vitro fluorescent acting staining test and also did not show homology to the Escherichia coli attachment-effacement gene (eaeA) by PCR. These results suggest that a mechanism or mechanisms other than the attachment-effacement lesion may also be involved in the association of H. alvei with diarrhea.
Susceptibility to levofloxacin predicted from in vitro susceptibility testing results obtained with ciprofloxacin and with ofloxacin
Authors: Cormican M G , Jones R N
Published: January 2020

A test battery of bacterial strains with a high incidence of resistance to fluoroquinolones was studied to determine the extent to which susceptibility to levofloxacin could be predicted from susceptibility tests performed with ciprofloxacin or ofloxacin as reagents. Isolates susceptible or intermediately susceptible to ofloxacin (MICs < or = 4 micrograms/ml) may be regarded as susceptible to levofloxacin, with the exception of Enterococcus faecium. Ciprofloxacin-susceptible isolates (MICs < or = 1 micrograms/ml) were also completely susceptible to levofloxacin. Clinical laboratories could use either of the currently used fluoroquinolones (ciprofloxacin or ofloxacin) to predict levofloxacin activity and spectrum with little risk of false-susceptible errors (0.0 to 1.3%). Results obtained with ofloxacin are superior in maximizing recognition of levofloxacin-susceptible clinical isolates.
Evaluation of an immunoassay for direct detection of Escherichia coli O157 in stool specimens
Authors: Dylla B L , Vetter E A , Hughes J G , Cockerill F R
Published: January 2020

An enzyme-linked immunosorbent assay (ELISA) produced by LMD Laboratories, Inc., Carlsbad, Calif., was compared with culture for the detection of Escherichia coli O157. Nine of 185 stool specimens evaluated had positive results by the LMD E. coli O157 ELISA and grew E. coli O157 on culture; 174 had negative by LMD E. coli O157 ELISA results and did not grow E. coli O157 on culture. Of 174 specimens negative by LMD E. coli O157 ELISA, 117 specimens grew other enteric pathogens: Campylobacter spp. (46 isolates), Salmonella spp. (43 isolates), Yersinia spp. (20 isolates), and Shigella spp. (8 isolates). There were two indeterminant results by the LMD E. coli O157 ELISA. One stool specimen did not grow other enteric pathogens on culture, and one grew a Campylobacter sp. on culture. Both had negative LMD E. coli O157 ELISA results upon repeat testing. The LMD E. coli O157 ELISA is an accurate, easy-to-read screening method for the detection of E. coli O157 in fecal specimens.
Genetic homogeneity of Taylorella equigenitalis from Norwegian trotting horses revealed by chromosomal DNA fingerprinting
Authors: Thoresen S I , Jenkins A , Ask E
Published: January 2020

Chromosomal DNA fingerprinting indicated that Norwegian Taylorella equigenitalis strains are genetically homogeneous and similar to some Swedish isolates but different from other European strains. As contagious equine metritis is rarely a serious disease in Norwegian horses, we conclude that the dominant T. equigenitalis strain in Norway is a genetically homogeneous clone of low virulence.
Meningococcal infections in the Province of Québec, Canada, during the period 1991 to 1992
Authors: Ringuette L , Lorange M , Ryan A , Ashton F
Published: January 2020

A total of 234 strains of Neisseria meningitidis obtained from hospitalized patients living in the province of Québec during the period 1991 to 1992 were characterized according to their serogroup, serotype, subtype, electrophoretic type, and antimicrobial susceptibility. All these strains were recovered from sterile body fluids, except for one strain that was isolated postmortem from a cutaneous lesion. For both years, serogroup C was the most prevalent (69.7%), followed by serogroup B (27.4%). Serotype 2a represented 80.3% of serogroup C isolates, and P1.2 was the most common subtype associated with this serotype. Clone ET 15 accounted for 76.5% of serogroup C isolates and 90.0% of serotype 2a strains. Although meningococcal disease occurred mostly in children under the age of 5 (9.7 cases per 100,000 children), with a peak incidence for children under 1 (20.3 cases per 100,000 children), most fatalities occurred among teenagers (12 to 19 years old). The total fatality rate was 11.5%, and serogroup C strains were responsible for 88.9% of these fatalities. Thirteen strains had a reduced susceptibility to penicillin G, and 28 strains were resistant to sulfadiazine. One strain was resistant to both rifampin and sulfadiazine and showed a reduced susceptibility to penicillin G.
Comparison of characteristics of Q beta replicase-amplified assay with competitive PCR assay for Chlamydia trachomatis
Authors: An Q , Liu J , O'Brien W , Radcliffe G , Buxton D , Popoff S , King W , Vera-Garcia M , Lu L , Shah J
Published: January 2020

In order to study infections due to Chlamydia trachomatis, we have compared semiquantitative PCR and Q beta replicase-amplified assays for detection of this organism. The PCR assay was directed against the C. trachomatis 16S rRNA gene. Quantitation was accomplished by adding known amounts of a plasmid containing a truncated segment of the 16S rRNA gene target to chlamydia-containing samples and then amplifying with a common primer set. The Q beta replicase assay consisted of reversible target capture of C. trachomatis 16S rRNA, which was followed by amplification of an RNA detector probe in the presence of the enzyme Q beta replicase. In a clinical matrix, the lower limit of detection of both the PCR and Q beta replicase assays was five elementary bodies. The Q beta replicase and PCR assays were quantitative over 10,000- and 1,000-fold ranges of organisms, respectively. Analysis of the effects of endocervical matrix on amplification was accomplished by examining 94 endocervical specimens by each technique. Both assays detected five of six culture-confirmed specimens as well as three culture-negative specimens. PCR inhibitors were detected in 13 specimens. The Q beta replicase assay, in contrast, showed no evidence of sample inhibition. The Q beta replicase and PCR assays should allow quantitative investigation of infections due to C. trachomatis. In addition, because it targets highly labile RNA, the Q beta replicase assay may facilitate investigations into the role of active persisting infection in culture-negative inflammatory conditions.
Factors which influence levels of selected organisms in saliva of older individuals
Authors: Loesche W J , Schork A , Terpenning M S , Chen Y M , Stoll J
Published: January 2020

The most commonly measured bacterial parameters in saliva are the levels of the mutans group streptococci and lactobacilli, which have diagnostic implications for the incidence of dental decay. Diagnostic guidelines which are applicable to children and young adults in whom most, if not all, teeth are present and in whom the rate of stimulated saliva is almost always greater than 0.5 ml/min have been developed. Dental decay is a potential health problem of considerable magnitude among elderly individuals. In elderly individuals, missing teeth, the presence of dentures, and a reduced salivary flow could confound the interpretation of salivary levels of cariogenic bacteria. In the present study, in which saliva was collected from more than 560 elderly individuals (average age, 70 +/- 8 years), there was a significant positive relationship between the salivary levels of Streptococcus mutans and increased numbers of teeth. There was a positive association between the salivary levels of S. mutans and decay when the data were stratified for the presence of a complaint of xerostomia and the presence of dentures. However, a similar analysis indicated that lactobacilli and yeasts were more likely to be associated with decay. The various variables which could influence the bacterial counts per milliliter of saliva, e.g., independent or dependent living status, complaint of xerostomia, stimulated salivary flow, salivary pH, the presence of dentures, number of teeth, and decay, were analyzed simultaneously by using a multivariable linear model. In that analysis the number of decayed teeth was significantly associated with the presence of lactobacilli (P = 0.0001) and yeasts (P = 0.025) but not with the presence of S. mutans. Our findings indicate that salivary levels of lactobacilli and yeasts, as well as the salivary levels of S. mutans, should be monitored when seeking microbial indicators that might predict the incidence of caries in elderly individuals.
Evaluation of Amplicor PCR for direct detection of Mycobacterium tuberculosis from sputum specimens
Authors: Beavis K G , Lichty M B , Jungkind D L , Giger O
Published: January 2020

We evaluated the Amplicor PCR assay (Roche Molecular Systems, Branchburg, N.J.) for direct detection of Mycobacterium tuberculosis in sputum. A total of 532 specimens from 270 patients were decontaminated and stored at 4 or -75 degrees C until assayed by PCR. This assay used three-step sample preparation, biotinylated primer pairs, AmpErase, and a microtiter format for amplicon capture and detection. Amplicor PCR results were compared with clinical history, culture from a Lowenstein-Jensen slant, and results from the BACTEC TB-460 system. Eighty-seven cultures from 15 patients grew M. tuberculosis; of these, 83 (95%) were positive with the Amplicor PCR test. The false negatives were most likely due to sample variation and inhibitors. Of the 445 specimens from which M. tuberculosis was not isolated, 428 (96%) were negative with the Amplicor PCR test. Of the 17 M. tuberculosis culture-negative, Amplicor-positive specimens, 15 were reclassified as true positives because previous cultures grew M. tuberculosis. Of the 445 specimens which did not grow M. tuberculosis, Mycobacterium spp. other than M. tuberculosis were isolated from 150 specimens. Three of these 150 specimens were Amplicor positive; two were from a patient with a history of tuberculosis, and one specimen gave a false-positive result. We do not feel that this represents cross-reactivity, because repeated Amplicor testing of the isolate gave negative results. The microtiter plate has 96 wells. Allowing for six controls, 90 decontaminated specimens can be tested by one technologist in 7.5 h. This PCR assay took 7.5 h to complete and is a sensitive and specific, rapid method for the direct detection of M. tuberculosis from sputum.
Inhibition of PCR by aqueous and vitreous fluids
Authors: Wiedbrauk D L , Werner J C , Drevon A M
Published: January 2020

The detection of viral nucleic acids in intraocular fluids and tissues by PCR has become increasingly important in clinical ophthalmology. While much attention has been directed toward minimizing false-positive reactions resulting from specimen contamination or amplicor carryover, relatively little attention has been given to the causes of false-negative PCRs. This report describes a PCR inhibitor in normal aqueous and vitreous fluids that can produce false-negative PCR results. As little as 0.5 microliter of vitreous fluid and 20 microliters of aqueous fluid can completely inhibit DNA amplification in a 100-microliters PCR mixture. This inhibition was not primer specific, nor was it due to chelation of Mg2+ ions or DNase activity in the ocular fluid. The inhibitor was completely resistant to boiling for 15 min. However, the inhibitory effects were completely removed by a single chloroform-isoamyl alcohol (24:1) extraction. The extent of PCR inhibition depended upon the type of thermostable DNA polymerase used in the reaction. Taq DNA polymerase was very sensitive to the inhibitor, while thermostable DNA polymerases from Thermus thermophilus HB-8 (Tth) and Thermus flavus (Tfl) were completely resistant. Thus, the inhibitory effects of intraocular fluids on PCRs can be removed by diluting the specimen, by chloroform extraction, or by using Tth or Tfl DNA polymerases.
Comparative evaluation of macrodilution and alamar colorimetric microdilution broth methods for antifungal susceptibility testing of yeast isolates
Authors: To W K , Fothergill A W , Rinaldi M G
Published: January 2020

A comparative evaluation of the macrodilution method and the Alamar colorimetric method for the susceptibility testing of amphotericin B, fluconazole, and flucytosine was conducted with 134 pathogenic yeasts. The clinical isolates included 28 Candida albicans, 17 Candida tropicalis, 15 Candida parapsilosis, 12 Candida krusei, 10 Candida lusitaniae, 9 Candida guilliermondii, 18 Torulopsis glabrata, and 25 Cryptococcus neoformans isolates. The macrodilution method was performed and interpreted according to the recommendations of the National Committee for Clinical Laboratory Standards (document M27-P), and the Alamar colorimetric method was performed according to the manufacturer's instructions. For the Alamar colorimetric method, MICs were determined at 24 and 48 h of incubation for Candida species and T. glabrata and at 48 and 72 h of incubation for C. neoformans. The overall agreement within +/- 1 dilution for Candida species and T. glabrata against the three antifungal agents was generally good, with the values for amphotericin B, fluconazole, and flucytosine being 85.3, 77.9, and 86.2%, respectively, at the 24-h readings and 69.3, 65.2, and 97.2%, respectively, at the 48-h readings. Most disagreement was noted with fluconazole against C. tropicalis and T. glabrata. Our studies indicate that determination of MICs at 24 h by the Alamar colorimetric method is a valid alternate method for testing amphotericin B, fluconazole, and flucytosine against Candida species but not for testing fluconazole against C. tropicalis and T. glabrata. For flucytosine, much better agreement can be demonstrated against Candida species and T. glabrata at the 48-h readings by the Alamar method. Excellent agreement within +/- dilution can also be observed for amphotericin B, fluconazole, and flucytosine (80, 96, and 96%, respectively) against c. neoformans when the MICs were determined at 72 h by the Alamar method.
Fatal cerebral mycoses caused by the ascomycete Chaetomium strumarium
Authors: Abbott S P , Sigler L , McAleer R , McGough D A , Rinaldi M G , Mizell G
Published: January 2020

Three cases of fatal cerebral mycosis in males with prior histories of intravenous drug use from the United States and Australia are reported. Infection in each case was limited to brain abscess; no other sites of infection were observed. The fungus seen by histopathology and isolated from the brain tissue in each case was identified as Chaetomium strumarium. This is the first report of human infection by this species, and C. strumarium is the second species of Chaetomium known to cause primary brain infection. Chaetomium strumarium is unusual among members of the genus Chaetomium in forming ascocarps covered with pale, thin-walled, flexuous hairs, a feature leading to its original placement in the genus Achaetomium. Presence of pinkish exudate droplets and/or crystals associated with hyphae or ascocarps, sometimes accompanied by a pinkish diffusible pigment; good growth at 42 degrees C; and production of small conidia further distinguish this species. The brain abscess isolates were compared with isolates from prior cases of cerebral infection which had been identified as either Chaetomium atrobrunneum or Chaetomium globosum. With reidentification of one isolate originally identified as C. globosum to C. atrobrunneum, only C. strumarium and C. atrobrunneum have been confirmed to cause infection involving the brain.
Hemocytic rickettsia-like organisms in ticks: serologic reactivity with antisera to Ehrlichiae and detection of DNA of agent of human granulocytic ehrlichiosis by PCR
Authors: Magnarelli L A , Stafford K C , Mather T N , Yeh M T , Horn K D , Dumler J S
Published: January 2020

Ixodid ticks were collected from Connecticut, Massachusetts, Missouri, Pennsylvania, Rhode Island, and British Columbia (Canada) during 1991 to 1994 to determine the prevalence of infection with hemocytic (blood cell), rickettsia-like organisms. Hemolymph obtained from these ticks was analyzed by direct and indirect fluorescent antibody (FA) staining methods with dog, horse, or human sera containing antibodies to Ehrlichia canis, Ehrlichia equi, or Rickettsia rickettsii. Of the 693 nymphal and adult Amblyomma americanum, Dermacentor variabilis, Ixodes scapularis, and Ixodes pacificus ticks tested with dog anti-E. canis antiserum, 209 (32.5%) contained hemocytic bacteria. The prevalence of infected ticks varied greatly with species and locale. In parallel tests of duplicate hemolymph preparations from adult I. scapularis ticks, the hemocytic organisms reacted positively with E. canis and/or E. equi antisera, including sera from persons who had granulocytic ehrlichiosis. In separate PCR analyses, DNA of the agent of human granulocytic ehrlichiosis was detected in 59 (50.0%) of 118 adult and in 1 of 2 nymphal I. scapularis ticks tested from Connecticut. There was no evidence of Ehrlichia chaffeensis DNA in these ticks. In indirect FA tests of hemolymph for spotted fever group rickettsiae, the overall prevalence of infection was less than 4%. Specificity tests of antigens and antisera used in these studies revealed no cross-reactivity between E. canis and E. equi or between any of the ehrlichial reagents and those of R. rickettsii. The geographic distribution of hemocytic microorganisms with shared antigens to Ehrlichia species or spotted fever group rickettsiae is widespread.
Characterization of Vibrio cgolerae non-O1 serogroups obtained from an outbreak of diarrhea in Lima, Peru
Authors: Dalsgaard A , Albert M J , Taylor D N , Shimada T , Meza R , Serichantalergs O , Echeverria P
Published: January 2020

In February 1994, an outbreak of diarrhea caused by non-O1 Vibrio cholerae occurred among volunteers in a vaccine trial study area in Lima, Peru. Clinically, 95% of the patients presented with liquid diarrhea with either no or mild dehydration. Serogrouping of 58 isolates recovered from diarrheal patients affected in the outbreak revealed seven different serogroups, with serogroups O10 (21%) and O12 (65%) being predominant. Most of these isolates were susceptible to a variety of antimicrobial agents. None of the 58 isolates hybridized with a DNA probe previously used to detect the gene encoding the heat-stable enterotoxin NAG-ST or produced cholera toxin as assessed by GM1 ganglioside enzyme-linked immunosorbent assay. Ribotyping exhibited 10 different BglI ribotype patterns among the 58 V. cholera non-O1 strains studied. However, ribotyping showed that all isolates belonging to serogroup O12 exhibited identical ribotypes and that 83% of the serogroup O10 isolates belonged to another identical ribotype, thus showing excellent correlation between ribotypes and serogroups. Among a group of O10 and O12 isolates selected for virulence studies, none produced enterotoxin whereas the majority produced a cytotoxin, as assessed in Y1 and HeLa cells. These isolates were also negative for the gene encoding zonula occludens toxin (Zot) as assessed by a PCR assay. The isolates tested showed strong adherence and some degree of invasion in the HEp-2 cell assay, whereas none of the isolates was positive in the PCR assay for the gene encoding the toxin coregulated pilus subunit A antigen (tcpA). In the removable intestinal tie adult rabbit diarrhea model, O10 and O12 serogroup isolates produced severe diarrhea and occasionally death when rabbits were challenged with 10(10) bacterial cells. Fluid accumulation was shown in the rabbit intestinal loop test when whole cultures were injected. No significant difference in virulence was shown between serogroup O10 and O12 isolates. This study provides further evidence that V. chlorae non-O1 non-O139 strains have diarrhegenic potential for humans through a yet-undefined mechanism(s) and that such strains can cause outbreaks.
Fluconazole- and itraconazole-resistant Candida albicans strains from AIDS patients: multilocus enzyme electrophoresis analysis and antifungal susceptibilities
Authors: Le Guennec R , Reynes J , Mallié M , Pujol C , Janbon F , Bastide J M
Published: January 2020

Multilocus enzyme electrophoresis and in vitro susceptibility testing with a broth microdilution method were used to analyze Candida albicans strain diversity in four AIDS patients with recurrent oropharyngeal candidiasis who successively developed clinical resistance to fluconazole (FCZ) and itraconazole (ITZ). One to ten colonies per sample were randomly chosen from oral washings collected before the initial FCZ treatment and just before every other antifungal treatment; a total of 98 isolates were analyzed. Multilocus enzyme electrophoresis analysis revealed 14 different electrophoretic types (ETs). Statistical analysis of genetic distances showed that C. albicans isolates clustered into five subpopulations (I to V). In each subpopulation, isolates are closely related, and genetic distances between subpopulations I to IV are short. In contrast, subpopulation V, which contained isolates typed as ET8 and ET14, is strongly divergent from the others; these isolates may represent atypical C. albicans isolates. Only one patient was infected with a single strain during the course of azole therapy; for the three remaining patients, variants of the same strain and different strains were concurrently isolated. Clinical FCZ resistance was clearly correlated with in vitro data for three patients. Moreover, MICs of ITZ increased during FCZ therapy, and MICs of ITZ which were > or = 1.56 micrograms/ml were found when clinical ITZ resistance occurred; isolates from subpopulation V showed the highest MICs of ITZ. Because of the emergence of clinical ITZ resistance after clinical FCZ resistance, the feasibility of long-term azole therapy for mucosal candidiasis in AIDS patients is questioned.
Six newly recognized types of Streptococcus pneumoniae
Authors: Henrichsen J
Published: January 2020

The serological properties of six new pneumococcal capsular types are described. A table listing all 90 pneumococcal types and their cross-reactions is included.
Improved PCR detection of Chlamydia trachomatis by using an altered method of specimen transport and high-quality endocervical specimens
Authors: Kellogg J A , Seiple J W , Klinedinst J L , Stroll E S , Cavanaugh S H
Published: January 2020

Duplicate endocervical swabs were collected for detection of Chlamydia trachomatis by PCR (Roche Diagnostics). One swab was swirled in Specimen Transport Medium (Roche) for PCR testing and discarded. A saline aliquot from the other specimen, sent as a dry swab to the laboratory, was Papanicolaou stained to determine specimen adequacy, and the remainder was PCR tested. Significantly more (24%) PCR-positive results (118 versus 95; P < 0.001) were obtained with the dry specimens than with the swirled specimens when first tested. In addition, PCR-positive results were obtained with 107 (10.6%) of 1,007 microscopically adequate specimens but with only 3 (0.9%) of 341 inadequate specimens (P < 0.001).
Nocardia species as an etiologic agent in Parkinson's disease: serological testing in a case-control study
Authors: Hubble J P , Cao T , Kjelstrom J A , Koller W C , Beaman B L
Published: January 2020

To test the hypothesis that Nocardia spp. may be an etiologic factor in Parkinson's disease (PD), we used a serodiagnostic panel to determine if PD patients had antibodies specific for Nocardia spp. To validate the serological test panel, sera from healthy volunteers and from patients with culture-proven nocardiosis (n = 307) were compared in part 1 of the study. The sensitivity of the panel was 88% for detection of culture-proven nocardial infections, and specificity was 85% (excluding cross-reactive leprosy cases). In part 2, no difference in seropositivity was found when PD patients were compared with their age- and gender-matched controls (n = 140). We found a high exposure rate of humans to nocardial antigens, especially among men and older individuals. Our results offer no support to the hypothesis that Nocardia spp. are causative in PD; however, it is possible that serological testing may not be optimal for detection of nocardial central nervous system infection.
Nonisotopic hybridization assay for determination of relative amounts of genotypic human immunodeficiency virus type 1 zidovudine resistance
Authors: Eastman P S , Boyer E , Mole L , Kolberg J , Urdea M , Holodniy M
Published: January 2020

A nonisotopic hybridization assay for human immunodeficiency virus genotypic zidovudine resistance determination is described. Biotinylated PCR product was hybridized with enzyme-labeled probes for wild-type or resistant mutant sequences and detected colorimetrically or chemiluminescently in a microplate format. Changes in mutant-to-wild-type ratios allow for the monitoring of longitudinal patient samples.
Recovery of Helicobacter pylori from gastric biopsy specimens is not dependent on the transport medium used
Authors: Roosendaal R , Kuipers E J , Peña A S , de Graaff J
Published: January 2020

The survival of six clinical isolates of Helicobacter pylori at room temperature was investigated after suspension in five different media: brucella broth with 5% lysed horse blood (BBLH), phosphate-buffered saline (PBS), 20% glucose, Stuart medium, and PBS with 10% Fildes enrichment (PBS-F). Only in BBLH and PBS-F no decrease in mean bacterial numbers was observed during the 24-h study period. No H. pylori isolates could be cultured from Stuart medium after 7 h of incubation. In contrast, the recovery rates in PBS-F or Stuart medium of H. pylori isolates from gastric tissue specimens collected from 19 H. pylori-positive patients were not significantly different even after a delay of culture of up to 24 h. Our data show that the medium composition is not critical for the survival of H. pylori within gastric tissue specimens.
Mycobacterium conspicuum sp. nov., a new species isolated from patients with disseminated infections
Authors: Springer B , Tortoli E , Richter I , Grünewald R , Rüsch-Gerdes S , Uschmann K , Suter F , Collins M D , Kroppenstedt R M , Böttger E C
Published: January 2020

A new type of slowly growing, nonphotochromogenic mycobacterium was recovered from two patients with disseminated disease. The growth characteristics, acid fastness, acids were consistent with those for Mycobacterium species. The results of biochemical investigations, lipid analyses, and comparative 16S rRNA sequencing showed that these isolates represent a new slowly growing Mycobacterium species which is named Mycobacterium conspicuum.
Genetic relatedness of Cryptococcus neoformans clinical isolates grouped with the repetitive DNA probe CNRE-1
Authors: Chen F , Currie B P , Chen L C , Spitzer S G , Spitzer E D , Casadevall A
Published: January 2020

Cryptococcus neoformans isolates from eight patients with cryptococcal infection were previously assigned into three groups on the basis of repetitive DNA probe (CNRE-1) restriction fragment length polymorphisms. These groups accounted for a disproportionate number of recent clinical isolates in New York City. To further examine the genetic relatedness of isolates within and across CNRE-1 groups, the DNA sequence of the 779-base URA5 gene from each strain was amplified and sequenced. The number of nucleotide differences occurred in the third codon position or in introns. Pairwise comparisons revealed average nucleotide differences within a CNRE-1 group of 4.8 +/- 2.6 (n = 8) and between CNRE-1 groups of 21.9 +/- 7.0 (n =20) (P <0.001) Analysis of URA5 sequences defined three groups that were congruent with those defined by CNRE-1 restriction fragment length polymorphisms. PCR amplification of an rDNA intergenic spacer revealed conservation of the intergenic spacer length within groups. Electrophoretic karyotyping did not distinguish between two isolates in each of two CNRE-1 groups. DNA from all isolates studied hybridized to an alpha mating type-specific probe. We interpret these results as suggesting a clonal population structure for some pathogenic isolates of C. neoformans in New York City.
Vancomycin-resistant enterococci colonizing the intestinal tracts of hospitalized patients
Authors: Gordts B , Van Landuyt H , Ieven M , Vandamme P , Goossens H
Published: January 2020

A point prevalence culture survey was carried out to investigate the prevalence of fecal carriage of vancomycin-resistant enterococci (VRE) among patients admitted to an 800-bed general hospital where no VRE had been isolated previously. Twenty-two of 636 patients (3.5%) were found to be VRE carriers. Eighteen strains were identified as Enterococcus faecium, three were identified as Enterococcus gallinarum, and one was identified as Enterococcus faecalis. The susceptibilities of the enterococci to ampicillin, vancomycin, and teicoplanin were determined by the disk diffusion and the agar dilution methods. High-level resistance (HLR) to gentamicin and streptomycin was determined by the agar screening method. Eighteen strains (82%) were highly resistant to vancomycin, and four strains (18%) were moderately resistant to vancomycin. Five strains were susceptible to teicoplanin (23%; MICs, < or = 8 micrograms/ml). Only one strain (4.5%, E. faecium) showed HLR to gentamicin, and six strains (27%) showed HLR to streptomycin (one E. faecalis and five E. faecium strains). All 18 E. faecium and 1 E. faecalis strain carried the vanA gene, and 3 E. gallinarum strains carried the vanC gene. An epidemiological investigation revealed several risk factors for VRE colonization: hospitalization and duration of stay in the hematology department and prior vancomycin treatment. The study demonstrates that the patient's gastrointestinal tract is a possible reservoir for VRE, even in hospitals where VRE infections have not yet been observed. Therefore, we conclude that infection control precautions and restriction of glycopeptide usage may be key issues in limiting the emergence and spread of nosocomial VRE infections.
Controlled clinical evaluation of BACTEC Plus Aerobic/F and BacT/Alert Aerobic FAN bottles for detection of bloodstream infections
Authors: Pohlman J K , Kirkley B A , Easley K A , Basille B A , Washington J A
Published: January 2020

A total of 7,190 blood culture sets were obtained from adult patients with a suspected bloodstream infection. A 20-ml sample of blood was distributed equally between the aerobic FAN bottle which was monitored in the BacT/Alert system and a Plus Aerobic/F bottle which was monitored in the BACTEC 9240 system. A total of 988 positive cultures were obtained from 483 patients; however, only 453 positive cultures from 173 patients met the criteria for volume ( > or = ml per bottle) and clinical significance on the basis of concurrent case review required for data analysis. There were 25 and 68 false positives from the FAN and Plus Aerobic/F bottles, respectively. There were no statistically significant differences between systems in the number of positive cultures or septic episodes by species; however, the total number of Enterobacteriaceae and Pseudomonas aeruginosa isolates combined was significantly greater in the FAN bottle (P = 0.04). Detection times did not differ significantly between systems for positive cultures; however, episodes of Staphylococcus aureus bacteremia were detected significantly more rapidly from the FAN bottle (P = 0.005). There was no significant difference between systems in the detection of bloodstream infections in patients receiving antibiotics at the time of blood culture.
Genotypic and phenotypic characterization of Helicobacter cinaedi and Helicobacter fennelliae strains isolated from humans and animals
Authors: Kiehlbauch J A , Brenner D J , Cameron D N , Steigerwalt A G , Makowski J M , Baker C N , Patton C M , Wachsmuth I K
Published: January 2020

By DNA-DNA hybridization, we classified 26 human strains, 4 dog and cat strains, and 4 hamster strains putatively identified as Helicobacter cinaedi as well as 2 human strains and 2 animal strains of Helicobacter fennelliae. All but one human strain belonged to the same hybridization group as the type strain of H. cinaedi. The animal strains also appeared to belong to this hybridization group. Both human strains of H. fennelliae were shown to be H. fennelliae by DNA-DNA hybridization, but both animal strains were less than 15% related to the type strain. All strains were also characterized by plasmid profiles and ribotyping. Plasmids were found in 23% of the human strains, 100% of the hamster strains, and 33% of the dog and cat strains. Human strains were essentially identical by ribotyping, but were clearly differentiated from the hamster and dog and cat strains. Some strains may be difficult to culture on primary isolation; we found that our strains grew well on anaerobic CDC agar, brucella agar, and tryptic soy agar II. Our H. cinaedi and H. fennelliae strains differed from those previously described because some were resistant to cephalothin: some H. cinaedi strains were also resistant to nalidixic acid. All isolates were also characterized by antimicrobial susceptibility testing. We found that human strains of H. cinaedi were more resistant to clindamycin and erythromycin than were animal isolates; 19% of the human strains were resistant to ciprofloxacin. Therefore, we recommend that antimicrobial susceptibility results be obtained before initiating therapy for H. cinaedi and H. fennelliae infections.
Detection and identification of mycobacteria by DNA amplification and oligonucleotide-specific capture plate hybridization
Authors: De Beenhouwer H , Liang Z , De Rijk P , Van Eekeren C , Portaels F
Published: January 2020

We have developed an easy and rapid detection and identification system for the diagnosis of mycobacterial diseases. The system is based on selective amplification by PCR of mycobacteria with primers based on the genes coding for 16S rRNA. During PCR, a label (digoxigenin-11-dUTP) is incorporated with biotinylated species-specific oligonucleotides (oligonucleotide-specific capture plate hybridization [OSCPH]. One oligonucleotide specific for the genus Mycobacterium and seven species-specific (Mycobacterium tuberculosis, M. avium, M. intracellulare, M. scrofulaceum, M. xenopi, M. genavense, and M. chelonae) oligonucleotides were designed as capturing probes. After specific hybridization, an enzyme immunoassay reveals the specifically bound complexes and thus permits identification of the mycobacterium. A total of 70 mycobacterial strains were tested. For 69 strains, results concordant with conventional identification were obtained. One M. chelonae strain was negative with the M. chelonae probe and was later reidentified as M. fortuitum. Moreover, for 15 clinical samples suspected of harboring nontuberculous mycobacteria, OSCPH was able to confirm all culture results and could identify one M. genavense infection for which standard culture results were negative. PCR-OSCPH is easily applicable and much faster than culture. It could become a valuable alternative approach for the diagnosis of mycobacterial infections.
Comparison of PCR and standard cytological staining for detection of Pneumocystis carinii from respiratory specimens from patients with or at high risk for infection by human immunodeficiency virus
Authors: Leibovitz E , Pollack H , Moore T , Papellas J , Gallo L , Krasinski K , Borkowsky W
Published: January 2020

The detection of Pneumocystis carinii DNA by PCR was compared with routine cytologic staining techniques (CYT). A total of 284 clinical respiratory specimens, including 137 bronchoalveolar lavage (BAL), 63 bronchoalveolar washing, 63 sputum, and 21 induced sputum samples, obtained from patients with or at high risk for human immunodeficiency virus infection were evaluated. Eighty specimens were positive by PCR, and 69 were positive by CYT. PCR was able to detect P. carinii in more bronchoalveolar washing specimens (15 versus 11) and in comparable BAL specimens (53 versus 54) compared with CYT. PCR was particularly more sensitive than CYT in detecting P. carinii in expectorated sputum (12 versus 4 samples). Of the 19 patients whose respiratory specimens were positive for P. carinii by PCR but negative by CYT, 5 had P. carinii pneumonia (PCP) confirmed by subsequent BAL and transbronchial or mediastinal lymph node biopsy and 9 had a clinical course highly suggestive of acute PCP. Eleven (58%) of the 19 patients with discordant PCR and CYT results had received prior anti-PCP prophylaxis. In this clinical setting in particular and in the evaluation of sputum specimens, the ability of PCR to detect a low parasitic load suggests that this technique may become an important additional tool, along with current cytological methods, for the detection of P. carinii.
Assessment of Difco ESP 384 blood culture system by terminal subcultures: failure to detect Cryptococcus neoformans in clinical specimens
Authors: Tinghitella T J , Lamagdeleine M D
Published: January 2020

Terminal subcultures were performed on 1,162 5-day negative blood culture sets which had been monitored by the Difco ESP 384, a continuous-monitor blood culture system. Of these, 16 (1.4%) had growth upon terminal subculture. The isolates not detected by the Difco ESP 384 were Cryptococcus neoformans (eight isolates), Candida albicans (one isolate), Staphylococcus aureus (two isolates) coagulase-negative staphylococcus (three isolates), Bacillus sp. (one isolate), and Corynebacterium sp. (one isolate). Acridine orange staining was performed on 200 randomly selected negative blood culture sets from the study group. Of these, two sets were positive and grew out C. neoformans, as did the terminal subculture. A review of patient's medical records indicated that many of these false-negative isolates were clinically insignificant. The Difco ESP 384 failed to detect 1.4% of the isolates in this study, 50% of which were C. neoformans, indicating a deficiency in the detection mechanism of the system. Further studies demonstrated that while these isolates (C. neoformans) grew in the Difco media, the system did not detect this growth when the standard 5-day protocol was used.
Application of PCR for detection of toxigenic Corynebacterium diphtheriae strains isolated during the Russian diphtheria epidemic, 1990 through 1994
Authors: Mikhailovich V M , Melnikov V G , Mazurova I K , Wachsmuth I K , Wenger J D , Wharton M , Nakao H , Popovic T
Published: January 2020

A total of 250 Corynebacterium diphtheriae isolates from clinical cases and carriers in Russia were assayed by PCR directed at the A subunit of the diphtheria toxin gene to distinguish toxigenic from nontoxigenic strains; 170 strains were positive as indicated by the presence of the 248-bp amplicon. The results of this PCR assay were in complete concordance with those of the standard immunoprecipitation assay (Elek), and the PCR assay is a useful tool for rapid identification in clinical laboratories.
Antimicrobial susceptibilities and serotypes of invasive Streptococcus pneumoniae strains in Switzerland
Authors: Wüst J , Huf E , Kayser F H
Published: January 2020

In 1993 and 1994, 10 microbiological laboratories in Switzerland collected 351 strains of Streptococcus pneumoniae from invasive infections. Susceptibilities to the main representatives of the chemical classes were as follows: penicillin, 93%; chloramphenicol, 92%; erythromycin, 94%; sulfamethoxazole-trimethoprim, 86%; tetracycline, 92%; vancomycin, 100%. Forty-three strains showed resistance to one agent, and 35 strains showed resistance to two or more antimicrobial agents simultaneously; i.e., 22% of the strains were resistant to at least one antimicrobial agent. Four strains (1%) were fully resistant to penicillin, whereas 21 strains (6%) showed reduced susceptibility. Of these 25 strains not fully susceptible to penicillin, 10 were resistant to one, 3 were resistant to two, and 8 were resistant to three additional antimicrobial agents. Of the quinolones, sparfloxacin was the most active substance, with an MIC at which 90% of the strains are inhibited of 0.5 mg/liter. The most common serotypes were types 6 (13.6% of isolates), 7 (10.5%), 19 (10.5%), 14 (9.1%), and 1 (8.5%) as well as 3 and 23 (8.0% each). Reduced susceptibility to penicillin was found mainly among serotypes 6, 14, 19, and 23. The currently available 23-valent pneumococcal vaccine covers 320 (91%) of the pneumococci isolated. Regional differences within Switzerland with regard to serotypes and antimicrobial resistance were not observed.