bluuebunny/crossref_metadata_2025 huggingface dataset:
Fatal Disseminated
Trichoderma longibrachiatum
Infection in an Adult Bone Marrow Transplant Patient: Species Identification and Review of the Literature
Authors: Richter S. , Cormican M. G. , Pfaller M. A. , Lee C. K. , Gingrich R. , Rinaldi M. G. , Sutton D. A.
Published: December 2019

Trichoderma longibrachiatum
was recovered from stool surveillance cultures and a perirectal ulcer biopsy specimen from a 29-year-old male who had received an allogeneic bone marrow transplant for acute lymphoblastic leukemia. The amphotericin B (2.0 μg/ml) and itraconazole (1.0 μg/ml) MICs for the organism were elevated. Therapy with these agents was unsuccessful, and the patient died on day 58 posttransplantation. At autopsy, histologic sections from the lungs, liver, brain, and intestinal wall showed infiltration by branching septate hyphae. Cultures were positive for
Trichoderma longibrachiatum
. While
Trichoderma
species have been recognized to be pathogenic in profoundly immunosuppressed hosts with increasing frequency, this is the first report of probable acquisition through the gastrointestinal tract. Salient features regarding the identification of molds in the
Trichoderma longibrachiatum
species aggregate are presented.
Identification of
Vibrio parahaemolyticus
Strains at the Species Level by PCR Targeted to the
toxR
Gene
Authors: Kim Yung Bu , Okuda Jun , Matsumoto Chiho , Takahashi Naoki , Hashimoto Satoru , Nishibuchi Mitsuaki
Published: December 2019

The DNA colony hybridization test with the polynucleotide probe for
Vibrio parahaemolyticus toxR
gene was performed. All 373 strains of
V. parahaemolyticus
gave positive results, and the strains belonging to four other
Vibrio
species including
Vibrio alginolyticus
gave weakly positive results, suggesting that
toxR
sequence variation may reflect the phylogenetic relationships of
Vibrio
species. We then established a
toxR
-targeted PCR protocol for the specific detection of
V. parahaemolyticus
.
Aspergillus
Meningitis: Diagnosis by Non-Culture-Based Microbiological Methods and Management
Authors: Verweij Paul E. , Brinkman Kees , Kremer Herbert P. H. , Kullberg Bart-Jan , Meis Jacques F. G. M.
Published: December 2019

The performance of antibody detection, antigen detection, and
Aspergillus
genus-specific PCR for diagnosing
Aspergillus
meningitis was investigated with 26 cerebrospinal fluid (CSF) samples obtained from a single patient with proven infection caused by
Aspergillus fumigatus
. Immunoglobulin G antibodies directed against
Aspergillus
were not detected by enzyme-linked immunosorbent assay in CSF or serum. The antigen galactomannan was detected in the CSF 45 days before a culture became positive, and
Aspergillus
DNA was detected 4 days prior to culture. Decline of the galactomannan antigen titer in the CSF during treatment with intravenous and intraventricular amphotericin B and intravenous voriconazole corresponded with the clinical response to treatment.
Identification of a Novel Genotype of Hepatitis G Virus in Southeast Asia
Authors: Naito Hideo , Win Khin Maung , Abe Kenji
Published: December 2019

Hepatitis G virus (HGV) isolates obtained from 20 Myanmarese and 10 Vietnamese subjects were analyzed. A cluster of isolates not belonging to any known genotype of HGV was found in five Myanmarese subjects and three Vietnamese subjects by phylogenetic analysis, and we classified this new genotype as type 4. These results revealed that the HGV genome can be classified into at least four major genotypes.
PCR-Based Restriction Pattern Typing of the
vacA
Gene Provides Evidence for a Homogeneous Group among
Helicobacter pylori
Strains Associated with Peptic Ulcer Disease
Authors: Donati Manuela , Storni Elisa , D’Apote Lucia , Moreno Sandra , Tucci Antonio , Poli Loris , Cevenini Roberto
Published: December 2019

The results of PCR-based molecular typing of
Helicobacter pylori
strains by restriction fragment length polymorphism analysis of a 1,161-bp nucleotide sequence of the midregion of the
vacA
gene are reported. A total of 48
H. pylori
strains isolated from gastric biopsy specimens obtained from 18 patients with peptic ulcer dyspepsia, 15 patients with nonulcer dyspepsia, and 15 asymptomatic
H. pylori
-infected subjects were studied. Highly heterogeneous restriction patterns were obtained by digestion of PCR products with
Sau
II,
Bgl
II, and
Hha
I, whereas
Hae
III digestion resulted in a strictly homogeneous profile for
H. pylori
strains isolated from 14 of 18 (77.7%) patients with peptic ulcer dyspepsia, but a strictly homogeneous profile was found for strains from only 8 of 15 (53.3%) patients with nonulcer dyspepsia (
P
= 0.163) and 5 of 15 (33.3%) asymptomatic
H. pylori
-infected subjects (
P
= 0.014). A potentially important aspect of the results obtained is the clinical relevance, since a single restriction pattern seems to be able to identify the majority of
H. pylori
strains associated with peptic ulcer disease.
Identification of Nonlipophilic Corynebacteria Isolated from Dairy Cows with Mastitis
Authors: Hommez Jozef , Devriese Luc A. , Vaneechoutte Mario , Riegel Philippe , Butaye Patrick , Haesebrouck Freddy
Published: December 2019

Nonlipophilic corynebacteria associated with clinical and subclinical mastitis in dairy cows were found to belong to four species:
Corynebacterium amycolatum
,
Corynebacterium ulcerans
,
Corynebacterium pseudotuberculosis
, and
Corynebacterium minutissimum
. These species may easily be confused. However, clear-cut differences between
C. ulcerans
and
C. pseudotuberculosis
were found in their acid production from maltotriose and ethylene glycol, susceptibility to vibriostatic agent O129, and alkaline phosphatase. Absence of growth at 20°C and lack of α-glucosidase and 4MU-α-
d
-glycoside hydrolysis activity differentiated
C. amycolatum
from
C. pseudotuberculosis
and
C. ulcerans
. The mastitis
C. pseudotuberculosis
strains differed from the biovar equi and ovis reference strains and from caprine field strains in their colony morphologies and in their reduced inhibitory activity on staphylococcal β-hemolysin.
C. amycolatum
was the most frequently isolated nonlipophilic corynebacterium.
Molecular Markers Demonstrate that the First Described Multidrug-Resistant
Mycobacterium bovis
Outbreak Was Due to
Mycobacterium tuberculosis
Authors: Gutiérrez M. C. , Galán J. C. , Blázquez J. , Bouvet E. , Vincent V.
Published: December 2019

We genetically characterized multidrug-resistant
Mycobacterium tuberculosis
complex strains which caused a nosocomial outbreak of tuberculosis affecting six human immunodeficiency virus (HIV)-positive patients and one HIV-negative staff member (E. Bouvet, E. Casalino, G. Mendoza-Sassi, S. Lariven, E. Vallée, M. Pernet, S. Gottot, and F. Vachon, AIDS 7:1453–1460, 1993). The strains showed all the phenotypic characteristics of
Mycobacterium bovis
. They presented a high copy number of IS
6110
, the spacers 40 to 43 in the direct repeat locus, and the
mtp40
fragment. They lacked the G-A mutation at position 285 in the
oxyR
gene and the C-G mutation at position 169 in the
pncA
gene. These genetic characteristics revealed that these were dysgonic, slow-growing
M. tuberculosis
strains mimicking the
M. bovis
phenotype, probably as a consequence of cellular alterations associated with the multidrug resistance. Spoligotyping and IS
6110
restriction fragment length polymorphism (RFLP) analysis confirmed that the outbreak was due to a single strain. However, the IS
6110
RFLP pattern of the strain isolated from the last patient, diagnosed three years after the index case, differed slightly from the patterns of the other six strains. A model of a possible genetic event is presented to explain this divergence. This study stresses the value of using several independent molecular markers to identify multidrug-resistant tubercle bacilli.
Molecular Evidence for Heterogeneity of the Multiple-Drug-Resistant
Mycobacterium tuberculosis
Population in Scotland (1990 to 1997)
Authors: Fang Z. , Doig C. , Rayner A. , Kenna D. T. , Watt B. , Forbes K. J.
Published: December 2019

Multiple-drug-resistant
Mycobacterium tuberculosis
(MDR-MTB) has been well studied in hospitals or health care institutions and in human immunodeficiency virus-infected populations. However, the characteristics of MDR-MTB in the community have not been well investigated. An understanding of its prevalence and circulation within the community will help to estimate the problem and optimize the strategies for control and prevention of its development and transmission. In this study, MDR-MTB isolates from Scotland collected between 1990 and 1997 were characterized, along with non-drug-resistant isolates. The results showed that they were genetically diverse, suggesting they were unrelated to each other and had probably evolved independently. Several new alleles of
rpoB
,
katG
, and
ahpC
were identified:
rpoB
codon 525 (ACC→AAC; Thr525Asn);
katG
codon 128 (CGG→CAG; Arg128Gln) and codon 291 (GCT→CCT; Ala291Pro); and the
ahpC
synonymous substitution at codon 6 (ATT→ATC). One of the MDR-MTB isolates from an Asian patient had an IS
6110
restriction fragment length polymorphism pattern very similar to that of the MDR-MTB W strain and had the same drug resistance-related alleles but did not have any epidemiological connection with the W strains. Additionally, a cluster of
M. tuberculosis
isolates was identified in our collection of 715 clinical isolates; the isolates in this cluster had genetic backgrounds very similar to those of the W strains, one of which had already developed multiple drug resistances. The diverse population of MDR-MTB in Scotland, along with a low incidence of drug-resistant
M. tuberculosis
, has implications for the control of the organism and prevention of its spread.
Age-Dependent Acquisition of Hepatitis G Virus/GB Virus C in a Nonrisk Population: Detection of the Virus by Antibodies
Authors: Feucht Heinz-Hubert , Schröter Matthias , Zöllner Bernhard , Polywka Susanne , Laufs Rainer
Published: December 2019

Until now there have been few seroepidemiological data for hepatitis G virus/GB virus type C (HGV/GBV-C). A four-antigen HGV/GBV-C immunoblot was established to examine 446 serum specimens from healthy individuals without risk factors for parenteral viral transmission. These individuals were divided into seven groups according to age. Seroprevalence rates were low for children and adolescents (5.6%) and increased for the age groups assumed to be the most sexually active (15.3 to 26.8%). Remarkably, none of the 80 individuals who tested positive for HGV/GBV-C antibodies were simultaneously positive for HGV/GBV-C viremia. From our data we conclude that HGV/GBV-C infection is widespread in the general population (16 to 25%). The development of an antibody response is associated with clearance of HGV/GBV-C viremia. Due to the lack of risk factors for HGV/GBV-C infection of blood, other efficient transmission routes must exist. It must be assumed that HGV/GBV-C transmission may be linked to sexual activity.
Multicenter Comparison of PCR Assays for Detection of Human Herpesvirus 8 DNA in Semen
Authors: Pellett Philip E. , Spira Thomas J. , Bagasra Omar , Boshoff Chris , Corey Lawrence , de Lellis Laura , Huang Meei-Li , Lin Jung-Chung , Matthews Steve , Monini Paolo , Rimessi Paola , Sosa Carlos , Wood Charles , Stewart John A.
Published: December 2019

Reported prevalences of human herpesvirus 8 (HHV-8) (Kaposi’s sarcoma-associated herpesvirus) in semen have ranged widely. This is possibly due to differences in assay sensitivity, geographic or population-based differences in the true presence of the virus in semen, and PCR contamination. This study assessed interlaboratory sensitivity and reproducibility in the analysis of blinded experimental panels, each consisting of 48 specimens and being composed of semen specimens from different healthy artificial-insemination donors (
n
= 30) and human immunodeficiency virus (HIV)-infected patients (
n
= 7) plus positive (
n
= 4) and negative (
n
= 7) controls. The experimental panels analyzed in each laboratory were identical except for being independently coded. Of 10 experiments done in five laboratories, 5 experiments from three laboratories had evidence of PCR contamination; all instances of contamination were in the context of nested PCR procedures. In the experiments with no false-positive results, HHV-8 DNA was detected in three (8%) of the 37 semen specimens (two from artificial-insemination donors and one from an HIV-positive patient) but in only 3 (1.6%) of the 184 PCRs in which these specimens were analyzed. This suggests that HHV-8 DNA is present in semen at concentrations that can be too low to allow its consistent detection. This study emphasizes the importance of performing blinded, multi-institution experiments to provide a coherent basis for comparing results and to motivate standardization of methods.
Discrimination of
Burkholderia multivorans
and
Burkholderia vietnamiensis
from
Burkholderia cepacia
Genomovars I, III, and IV by PCR
Authors: Bauernfeind Adolf , Schneider Ines , Jungwirth Renate , Roller Carsten
Published: December 2019

We present a PCR procedure for identification of
Burkholderia cepacia
,
Burkholderia multivorans
, and
Burkholderia vietnamiensis
. 16S and 23S ribosomal DNAs (rDNAs) of
B. multivorans
and
B. vietnamiensis
were sequenced and aligned with published sequences for definition of species-specific 18-mer oligonucleotide primers. Specific antisense 16S rDNA primers (for
B. cepacia
, 5′-AGC ACT CCC RCC TCT CAG-3′; for
B. multivorans
, 5′-AGC ACT CCC GAA TCT CTT-3′) and 23S rDNA primers (for
B. vietnamiensis
, 5′-TCC TAC CAT GCG TGC AA-3′) were paired with a general sense primer of 16S rDNAs (5′-AGR GTT YGA TYM TGG CTC AG-3′) or with a sense primer of 23S rDNA (5′-CCT TTG GGT CAT CCT GGA-3′). PCR with these primers under optimized conditions is appropriate to specifically and rapidly identify
B. multivorans
,
B. vietnamiensis
, and
B. cepacia
(genomovars I, III, and IV are not discriminated). In comparison with the polyphasic taxonomic analyses presently necessary for species and genomovar identification within the
B. cepacia
complex, our procedure is more rapid and easier to perform and may contribute to clarifying the clinical significance of individual members of the complex in cystic fibrosis.
High Aspartyl Proteinase Production and Vaginitis in Human Immunodeficiency Virus-Infected Women
Authors: de Bernardis F. , Mondello F. , Scaravelli G. , Pachì A. , Girolamo A. , Agatensi L. , Cassone A.
Published: December 2019

Vaginal isolates of
Candida albicans
from human immunodeficiency virus-positive (HIV
+
) and HIV
−
women with or without candidal vaginitis were examined for secretory aspartyl proteinase (Sap) production in vitro and in vivo and for the possible correlation of Sap production with pathology and antimycotic susceptibility in vitro. HIV
+
women with candidal vaginitis were infected by strains of
C. albicans
showing significantly higher levels of Sap, a virulence enzyme, than strains isolated from HIV
+
,
C. albicans
carrier subjects and HIV
−
subjects with vaginitis. The greater production of Sap in vitro was paralleled by greater amounts of Sap in the vaginal fluids of infected subjects. In an estrogen-dependent, rat vaginitis model, a strain of
C. albicans
producing a high level of Sap that was isolated from an HIV
+
woman with vaginitis was more pathogenic than a strain of
C. albicans
that was isolated primarily from an HIV
−
,
Candida
carrier. In the same model, pepstatin A, a strong Sap inhibitor, exerted a strong curative effect on experimental vaginitis. No correlation was found between Sap production and antimycotic susceptibility, as most of the isolates were fully susceptible to fluconazole, itraconazole, and other antimycotics, regardless of their source (subjects infected with strains producing high or low levels of Sap, subjects with vaginitis or carrier subjects, or subjects with or without HIV). Thus, high Sap production is associated with virulence of
C. albicans
but not with fungal resistance to fluconazole in HIV-infected subjects, and Sap is a potentially new therapeutic target in candidal vaginitis.
Use of a Novel Hepatitis C Virus (HCV) Major-Epitope Chimeric Polypeptide for Diagnosis of HCV Infection
Authors: Chien David Y. , Arcangel Phillip , Medina-Selby Angelica , Coit Doris , Baumeister Mark , Nguyen Steve , George-Nascimento Carlos , Gyenes Alexander , Kuo George , Valenzuela Pablo
Published: December 2019

The genome of hepatitis C virus (HCV) consists of seven functional regions: the core, E1, E2/NS1, NS2, NS3, NS4, and NS5 regions. The U.S. Food and Drug Administration-licensed 2.0G immunoassay for the detection of anti-HCV uses proteins from the core, NS3, and NS4 regions (McHutchinson et al., Hepatology 15:19–25, 1992). The 3.0G enzyme-linked immunosorbent assay includes the protein from the NS5 region (Uyttendaele et al., Vox Sang. 66:122–129, 1994). The necessity of detecting antibodies to viral envelope proteins (E1 and E2) and to different genotype samples has been demonstrated previously (Chien et al., Lancet 342:933, 1993; Lok et al., Hepatology 18:497–502, 1993). In this study we have attempted to improve the sensitivity of the anti-HCV assay by developing a single multiple-epitope fusion antigen (MEFA; MEFA-6) which incorporates all of the major immunodominant epitopes from the seven functional regions of the HCV genome. A nucleic acid sequence consisting of proteins from the viral core, E1, E2, NS3, NS4, and NS5 regions and different subtype-specific regions of the NS4 region was constructed, cloned, and expressed in yeast. The epitopes present on this antigen can be detected by epitope-specific monoclonal and polyclonal antibodies. In a competition assay, the MEFA-6 protein competed with 83 to 96% of genotype-specific antibodies from HCV genotype-specific peptides. This recombinant antigen was subsequently used to design an anti-HCV chemiluminescent immunoassay. We designed our assay using a monoclonal anti-human immunoglobulin G antibody bound to the solid phase. Because MEFA-6 is fused with human superoxide dismutase (h-SOD), we used an anti-human superoxide dismutase, dimethyl acridinium ester-labeled monoclonal antibody for detection. Our results indicate that MEFA-6 exposes all of the major immunogenic epitopes. Its excellent sensitivity and specificity for the detection of clinical seroconversion are demonstrated by this assay.
Postsurgical
Candida albicans
Infections Associated with an Extrinsically Contaminated Intravenous Anesthetic Agent
Authors: McNeil Michael M. , Lasker Brent A. , Lott Timothy J. , Jarvis William R.
Published: December 2019

From 16 to 30 April 1990, four of 364 (1%) postsurgical patients at one hospital developed
Candida albicans
fungemia or endophthalmitis. The case patients’ surgeries were clustered on two days. To identify risk factors for
C. albicans
infections, we conducted a cohort study comparing these 4 patients with 67 control patients who had surgeries on the same days but did not acquire
C. albicans
infections. The participation of anesthesiologist 9 (relative risk [RR], undefined;
P < 0.001)
and receipt of intravenous propofol, an anesthetic agent without preservative, which was administered by an infusion pump (RR, 8.8;
P = 0.048)
were identified as risk factors for
C. albicans
infections. The anesthetic had been recently introduced in the hospital. Hand cultures of 8 of 14 (57%) anesthesiologists were positive for
Candida
species; one yielded
C. albicans
. Anesthesiologist 9 was the only one to use stored syringes of propofol in the infusion pump and to reuse propofol syringes. DNA fingerprinting with a digoxigenin-labeled
C. albicans
repetitive element 2 probe and electrophoretic karyotyping showed two distinct banding patterns among patient isolates. We hypothesize that extrinsic contamination of propofol by anesthesiologist 9 likely resulted in
C. albicans
infections. These data suggest that strict aseptic techniques must be used when preparing and administering propofol.
Multicenter Quality Assessment of PCR Methods for Detection of Enteroviruses
Authors: Muir Peter , Ras Albert , Klapper Paul E. , Cleator Graham M. , Korn Klaus , Aepinus Christian , Fomsgaard Anders , Palmer Pierre , Samuelsson Agneta , Tenorio Antonio , Weissbrich Benedikt , van Loon A. M.
Published: December 2019

We conducted a multicenter evaluation of commercial and in-house PCR methods for the detection of enteroviruses. Three coded panels of test and control RNA samples, artificial clinical specimens, and representative enterovirus serotypes were used to assess amplification methods, RNA extraction methods, and reactivities with different enterovirus serotypes. Despite several differences between PCR methods, there was good agreement, although some variation in sensitivity was observed. Most PCR methods were able to detect enterovirus RNA derived from 0.01 50% tissue culture infective dose (TCID
50
) and were able to detect at least 1 TCID
50
of enterovirus in cerebrospinal fluid, stool, or throat swab specimens. Most were also able to detect a wide range of enterovirus serotypes, although serotypic identification was not possible. Some laboratories experienced false-positive results due to PCR contamination, which appeared to result mainly from cross-contamination of specimens during RNA extraction. Provided that this problem is overcome, these PCR methods will prove to be a sensitive and rapid alternative to cell culture for the diagnosis of enterovirus infection.
Clinical and Financial Benefits of Rapid Bacterial Identification and Antimicrobial Susceptibility Testing
Authors: Barenfanger Joan , Drake Cheryl , Kacich Gail
Published: December 2019

To assess the expected clinical and financial benefits of rapid reporting of microbiology results, we compared patients whose cultured samples were processed in the normal manner to patients whose samples were processed more rapidly due to a minor change in work flow. For the samples tested in the rapid-reporting time period, the vast majority of bacterial identification and antimicrobial susceptibility testing (AST) results were verified with the Vitek system on the same day that they were available. This time period was called rapid AST (RAST). For RAST, a technologist on the evening shift verified the data that became available during that shift. For the control time period, cultures were processed in the normal manner (normal AST [NAST]), which did not include evening-shift verification. For NAST, the results for approximately half of the cultures were verified on the first day that the result was available. The average turnaround time for the reporting of AST results was 39.2 h for RAST and 44.4 h for NAST (5.2 h faster for RAST [
P
= 0.001]). Subsequently, physicians were able to initiate appropriate antimicrobial therapy sooner for patients whose samples were tested as part of RAST (
P
= 0.006). The mortality rates were 7.9 and 9.6% for patients whose samples were tested as part of RAST and NAST, respectively (
P
= 0.45). The average length of stay was 10.7 days per patient for RAST and 12.6 days for NAST, a difference of 2.0 days less for RAST (
P
= 0.006). The average variable cost was $4,927 per patient for RAST and $6,677 for NAST, a difference of $1,750 less per patient for RAST (
P
= 0.001). This results in over $4 million in savings in variable costs per year in our hospital.
Clinical Evaluation of the Enhanced Gen-Probe Amplified Mycobacterium Tuberculosis Direct Test for Rapid Diagnosis of Tuberculosis in Prison Inmates
Authors: Bergmann John S. , Yuoh Gbo , Fish Geoffrey , Woods Gail L.
Published: December 2019

The reliability of the enhanced Amplified Mycobacterium Tuberculosis Direct Test (E-MTD; Gen-Probe, Inc., San Diego, Calif.) for rapid diagnosis of pulmonary tuberculosis (TB) was evaluated by testing 1,004 respiratory specimens from 489 Texas prison inmates. Results were compared to those of mycobacterial culture (BACTEC TB 460 and Middlebrook 7H11 biplates), smear for acid-fast bacilli (AFB; auramine O), and clinical course. After chart review, three patients (nine specimens) who were on antituberculosis therapy before the study began were excluded from final analysis. Of the remaining 995 specimens, 21 were AFB smear positive: 13 grew
Mycobacterium tuberculosis
complex (MTBC), 6 grew nontuberculous mycobacteria, and 2 (from two patients diagnosed with TB and started on therapy after the study began) were culture negative. Twenty-eight specimens (20 patients) were positive for MTBC by culture and E-MTD. Seven specimens (seven patients) were positive by culture alone; three were from patients who had other E-MTD-positive specimens, two were false-positive cultures, and two were false-negative E-MTD results. Eight specimens were positive by E-MTD only; four specimens (four patients) were false-positive E-MTD results, and four specimens were from two patients with earlier E-MTD-positive specimens that grew MTBC. Thus, there were 22 patients with TB (10 smear positive and 12 smear negative). The sensitivity and specificity of the AFB smear for diagnosis of TB, by patient, were 45.5 and 98.9%, respectively. After resolving discrepancies, these same values for E-MTD were 90.9 and 99.1% overall, 100 and 100% for the smear-positive patients, and 83.3 and 99.1% for the smear-negative patients. Excluding the one smear-negative patient whose E-MTD-negative, MTBC culture-positive specimen contained inhibitory substances, the sensitivity of E-MTD was 95.2% overall and 90.9% in smear-negative patients. The specificity and positive predictive value of E-MTD can be improved, without altering other performance characteristics, by modifying the equivocal zone recommended by the manufacturer. These data suggest that E-MTD is a reliable method for rapid diagnosis of pulmonary TB, irrespective of the AFB smear result. Guidelines for the most appropriate use of E-MTD with smear-negative patients are needed.
Coaggregation of
Candida dubliniensis
with
Fusobacterium nucleatum
Authors: Jabra-Rizk Mary Ann , Falkler William A. , Merz William G. , Kelley Jacqueline I. , Baqui A. A. M. A. , Meiller Timothy F.
Published: December 2019

The binding of microorganisms to each other and oral surfaces contributes to the progression of microbial infections in the oral cavity.
Candida dubliniensis
, a newly characterized species, has been identified in human immunodeficiency virus-seropositive patients and other immunocompromised individuals.
C. dubliniensis
phenotypically resembles
Candida albicans
in many respects yet can be identified and differentiated as a unique
Candida
species by phenotypic and genetic profiles. The purpose of this study was to determine oral coaggregation (CoAg) partners of
C. dubliniensis
and to compare these findings with CoAg of
C. albicans
under the same environmental conditions. Fifteen isolates of
C. dubliniensis
and 40 isolates of
C. albicans
were tested for their ability to coaggregate with strains of
Fusobacterium nucleatum
,
Peptostreptococcus micros
,
Peptostreptococcus magnus
,
Peptostreptococcus anaerobius
,
Porphyromonas gingivalis
, and
Prevotella intermedia
. When
C. dubliniensis
and
C. albicans
strains were grown at 37°C on Sabouraud dextrose agar, only
C. dubliniensis
strains coaggregated with
F. nucleatum
ATCC 49256 and no
C. albicans
strains showed CoAg. However, when the
C. dubliniensis
and
C. albicans
strains were grown at 25 or 45°C, both
C. dubliniensis
and
C. albicans
strains demonstrated CoAg with
F. nucleatum
. Heating the
C. albicans
strains (grown at 37°C) at 85°C for 30 min or treating them with dithiothreitol allowed the
C. albicans
strains grown at 37°C to coaggregate with
F. nucleatum
. CoAg at all growth temperatures was inhibited by mannose and α-methyl mannoside but not by EDTA or arginine. The CoAg reaction between
F. nucleatum
and the
Candida
species involved a heat-labile component on
F. nucleatum
and a mannan-containing heat-stable receptor on the
Candida
species. The CoAg reactions between
F. nucleatum
and the
Candida
species may be important in the colonization of the yeast in the oral cavity, and the CoAg of
C. dubliniensis
by
F. nucleatum
when grown at 37°C provides a rapid, specific, and inexpensive means to differentiate
C. dubliniensis
from
C. albicans
isolates in the clinical laboratory.
Detection of Unculturable Bacteria in Periodontal Health and Disease by PCR
Authors: Harper-Owen R. , Dymock D. , Booth V. , Weightman A. J. , Wade W. G.
Published: December 2019

Recently developed molecular methods have made it possible to characterize mixed microflora in their entirety, including the substantial numbers of bacteria which do not grow on artificial culture media. In a previous study, molecular analysis of the microflora associated with acute oral infections resulted in the identification of three phylotypes, PUS3.42, PUS9.170, and PUS9.180, representing as-yet-uncultured organisms. The aim of this study was to design and validate specific PCR primers for these phylotypes and to determine their incidences in samples collected from healthy and diseased periodontal tissues. Two specific reverse primers were devised for each phylotype, and these were used in duplex PCRs with universal forward and reverse primers. All three phylotypes were detected in periodontal sites; PUS9.170, related to oral asaccharolytic
Eubacterium
spp., was significantly associated with disease. This study demonstrates the possibility of using unculturable, and therefore uncharacterized, organisms as markers of disease.
Western Immunoblot Analysis of the Antigens ofHaemobartonella feliswith Sera from Experimentally Infected Cats
Authors: Alleman A. Rick , Pate Melanie G. , Harvey John W. , Gaskin Jack M. , Barbet Anthony F.
Published: December 2019

Cats were experimentally infected with a Florida isolate ofHaemobartonella felisin order to collect organisms and evaluate the immune response toH. felis. Cryopreserved organisms were thawed and injected intravenously into nonsplenectomized and splenectomized cats. Splenectomized animals were given 10 mg of methylprednisolone per ml at the time of inoculation. Blood films were evaluated daily for 1 week prior to infection and for up to 60 days postinfection (p.i.). Blood forH. felispurification was repeatedly collected from splenectomized animals at periods of peak parasitemias. Organisms were purified from infected blood by differential centrifugation, separated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, and transferred to nitrocellulose membranes for immunoblot analysis. Serum was collected from nonsplenectomized animals prior to and for up to 60 days p.i. and was used on immunoblots to identify antigens. The combination of splenectomy and corticosteroid treatment resulted in marked, cyclic parasitemias without concurrent severe anemia, providing an opportunity to harvest organisms in a manner that was not lethal to the animals. Several antigens (150, 52, 47, 45, and 14 kDa) were identified. An antigen with a molecular mass of approximately 14 kDa appeared to be one of the most immunodominant and was consistently recognized by immune sera collected at various times during the course of infection. These data suggest that one or more of these antigens might be useful for the serologic diagnosis ofH. felisinfections in cats.
Membrane Filtration Test for Rapid Presumptive Differentiation of Four
Candida
Species
Authors: Bauters T. G. , Peleman R. , Moerman M. , Vermeersch H. , de Looze D. , Noens L. , Nelis H. J.
Published: December 2019

A rapid enzymatic two-step test for the presumptive differentiation of four
Candida
species commonly occurring in various clinical samples is described. The technique involves membrane filtration of a liquid sample, followed by preincubation of the membrane filter on Sabouraud glucose agar supplemented with ticarcillin-clavulanic acid to yield microcolonies. In a separate assay step, parts of the filter are placed on absorbent pads impregnated with fluorogenic 4-methylumbelliferyl (4-MU) enzyme substrates (4-MU-
N
-acetyl-β-
d
-galactosaminide, 4-MU-phosphate, 4-MU-pyrophosphate, and 4-MU-β-
d
-galactoside) in combination with 0.1% digitonin acting as a membrane permeabilizer. The membrane filter in contact with the assay medium is incubated to allow cleavage of the enzyme substrate, resulting in fluorescent microcolonies under long-wavelength UV light. This approach, tested on 301 clinical samples, is able to presumptively differentiate
C. albicans
,
C. glabrata
,
C. krusei
, and
C. tropicalis
and to distinguish them from other
Candida
spp. in about 9 to 11 h. Overall agreement with the conventional methods of 94.4% (one
Candida
species present in the sample) to 83.8% (multiple
Candida
spp. present) was obtained. The false-negative rates with reference to identification by traditional methods were 1.3% (single species) and 3.8% (multiple species).
Evaluation of Recombinant Antigens for Serodiagnosis of Chagas’ Disease in South and Central America
Authors: Umezawa Eufrosina S. , Bastos Sueli F. , Camargo Mario E. , Yamauchi Luci M. , Santos Márcia R. , Gonzalez Antonio , Zingales Bianca , Levin Mariano J. , Sousa Octavio , Rangel-Aldao Rafael , da Silveira José Franco
Published: December 2019

The commercially available diagnostic tests for Chagas’ disease employ whole extracts or semipurified fractions of
Trypanosoma cruzi
epimastigotes. Considerable variation in the reproducibility and reliability of these tests has been reported by different research laboratories, mainly due to cross-reactivity with other pathogens and standardization of the reagents. The use of recombinant antigens for the serodiagnosis of Chagas’ disease is recommended to increase the sensitivity and specificity of serological tests. Expressed in
Escherichia coli
, as fusion products with glutathione
S
-transferase, six
T. cruzi
recombinant antigens (H49, JL7, A13, B13, JL8, and 1F8) were evaluated in an enzyme-linked immunosorbent assay for Chagas’ disease. The study was carried out with a panel of 541 serum samples of chagasic and nonchagasic patients from nine countries of Latin America (Argentina, Bolivia, Brazil, Chile, Colombia, El Salvador, Guatemala, Honduras, and Venezuela). The optimal concentration of each recombinant antigen for coating of plates was determined with the help of
125
I-labelled recombinant proteins. While the specificity of the epimastigote antigen was 84% because of false positives from leishmaniasis cases, for the recombinant antigens it varied from 96.2 to 99.6%. Recombinant antigens reacted with 79 to 100% of serum samples from chronic chagasic patients. In this way, it is proposed that a mixture of a few
T. cruzi
recombinant antigens should be employed in a diagnostic kit to minimize individual variation and promote high sensitivity in the diagnosis of Chagas’ disease.
Visualization of Hydatid Elements: Comparison of Several Techniques
Authors: Clavel A. , Varea M. , Doiz O. , López L. , Quílez J. , Castillo F. J. , Rubio C. , Gómez-Lus R.
Published: December 2019

Some techniques available at our laboratory were tested for their ability to aid in the morphological diagnosis of hydatid elements (
Echinococcus granulosus
[“
Taenia echinococcus
”]) isolated from cysts in humans and sheep. Unstained, methanol-fixed hooklets were fluorescent, most starkly so under violet light (excitation filter wavelength, 405 nm; long-pass filter wavelength, 495 nm). Auramine-rhodamine and Gram procedures failed to stain hooklets. Ziehl-Neelsen stain yielded indifferent results when organisms were viewed under transmitted light but resulted in a surprisingly intense red fluorescence when organisms were viewed under green light (excitation, 546 nm; long pass, 590 nm). Wheatley trichrome stain gave better and more uniform results than fuchsin. Ryan trichrome blue stain was the best under transmitted light; hooklets stained uniformly and intensely and were easily distinguishable from the background. Very satisfactory results were also obtained with a much simpler procedure (modified Baxby technique: no fixation, steaming hot 1% safranin for 2 min, and malachite green for 30 s). Therefore, Ryan and modified Baxby stains are recommended for the examination of
E. granulosus
under transmitted light. For fluorescence microscopy, Ziehl-Neelsen stain under green excitation light, or violet light with no staining, is also very useful. Epifluorescence microscopy is especially convenient for examining samples concentrated by filtration, as it renders the filter pores inconspicuous.
Virulence Factors and Phenotypical Traits of Verotoxin-Producing
Escherichia coli
Strains Isolated from Asymptomatic Human Carriers
Authors: Stephan R. , Untermann F.
Published: December 2019

Fourteen verotoxin-producing
Escherichia coli
strains isolated from stool samples of 14 different asymptomatic human carriers were further characterized. A variety of serotypes was found, but none of the strains belonged to serogroup O157. Only one isolate carried most of the virulence genes that are associated with increased pathogenicity.
Microbiological Characteristics of Yeasts Isolated from Urinary Tracts of Intensive Care Unit Patients Undergoing Urinary Catheterization
Authors: Febré N. , Silva V. , Medeiros E. A. S. , Wey S. B. , Colombo A. L. , Fischman O.
Published: December 2019

We studied 70 intensive care unit patients to determine the incidence of nosocomial candiduria associated with indwelling urinary catheters and to assess microbiological characteristics of the yeasts. The yeasts were isolated, 13 of 17 in urine cultures and 4 of 17 in blood cultures, and colonization had occurred 3 days after the insertion of indwelling urinary catheters. For four strains the MICs of the antifungal drugs were high.
Comparison of Human Immunodeficiency Virus Type 1 (HIV-1) Protease Mutations in HIV-1 Genomes Detected in Plasma and in Peripheral Blood Mononuclear Cells from Patients Receiving Combination Drug Therapy
Authors: Koch Nathalie , Yahi Nouara , Ariasi Franck , Fantini Jacques , Tamalet Catherine
Published: December 2019

Detections of mutations in the protease gene of human immunodeficiency virus type 1 in plasma and peripheral blood mononuclear cells (PBMC) were sought in two matched populations of 23 individuals receiving combination drug therapy with or without protease inhibitors. In the control group (23 patients not receiving protease inhibitors), no primary resistance mutations were found. In contrast, primary resistance mutations (especially at codons M46, V82, and L90) were found in 16 of 23 patients (70%) treated with protease inhibitors. In 30% of the cases, these mutations were detected in plasma but not in PBMC.
Detection of Caprine Herpesvirus 1 in Sacral Ganglia of Latently Infected Goats by PCR
Authors: Tempesta Maria , Pratelli Annamaria , Greco Grazia , Martella Vito , Buonavoglia Canio
Published: December 2019

A study of the latency of caprine herpesvirus 1 (CpHV.1) was carried out with four latently infected goats. Three goats were treated with dexamethasone and euthanized after 4 and 6 days. PCR and virus isolation allowed us to detect CpHV.1 only in the third and fourth sacral ganglia of the two animals euthanized 6 days after the start of treatment.
Evaluation of the MRL Diagnostics Dengue Fever Virus IgM Capture ELISA and the PanBio Rapid Immunochromatographic Test for Diagnosis of Dengue Fever in Jamaica
Authors: Palmer Carol J. , King S. Dorothy , Cuadrado Raul R. , Perez Eddy , Baum Mariana , Ager Arba L.
Published: December 2019

We evaluated two new commercial dengue diagnostic tests, the MRL Diagnostics Dengue Fever Virus IgM Capture ELISA and the PanBio Rapid Immunochromatographic Test, on serum samples collected during a dengue epidemic in Jamaica. The MRL ELISA method correctly identified 98% (78 of 80) of the samples as dengue positive, while the PanBio test identified 100% (80 of 80). Both tests were 100% (20 samples of 20) specific.
Genetic Diversity amongEscherichia coliIsolates Carryingf18Genes from Pigs with Porcine Postweaning Diarrhea and Edema Disease
Authors: Nagy Béla , Wilson Richard A. , Whittam Thomas S.
Published: December 2019

Multilocus enzyme electrophoresis was applied to detect allelic variation and multilocus genotypes (electrophoretic types [ETs]) among 43Escherichia coliisolates from weaned pigs suffering from edema disease or from diarrhea. ETs were analyzed in relation to O serogroups and virulence genes (sta,stb,lt,stx2, andf18) by DNA hybridization. Genomic diversity was the lowest in serogroup O138, while virulence genes (stx2andf18) were the most uniform in serogroup O139. In general, the serogroups or toxin and F18 fimbria types were not related to selected ETs, suggesting that the toxin andf18fimbria genes inE. coliisolates from pigs with postweaning diarrhea or edema disease occur in a variety of chromosomal backgrounds.
Detection of Poliovirus Circulation by Environmental Surveillance in the Absence of Clinical Cases in Israel and the Palestinian Authority
Authors: Manor Y. , Handsher R. , Halmut T. , Neuman M. , Bobrov A. , Rudich H. , Vonsover A. , Shulman L. , Kew O. , Mendelson E.
Published: December 2019

The global eradication of poliomyelitis, believed to be achievable around the year 2000, relies on strategies which include high routine immunization coverage and mass vaccination campaigns, along with continuous monitoring of wild-type virus circulation by using the laboratory-based acute flaccid paralysis (AFP) surveillance. Israel and the Palestinian Authority are located in a geographical region in which poliovirus is still endemic but have been free of poliomyelitis since 1988 as a result of intensive immunization programs and mass vaccination campaigns. To monitor the wild-type virus circulation, environmental surveillance of sewage samples collected monthly from 25 to 30 sites across the country was implemented in 1989 and AFP surveillance began in 1994. The sewage samples were processed in the laboratory with a double-selective tissue culture system, which enabled economical processing of large number of samples. Between 1989 and 1997, 2,294 samples were processed, and wild-type poliovirus was isolated from 17 of them in four clusters, termed “silent outbreaks,” in September 1990 (type 3), between May and September 1991 (type 1), between October 1994 and June 1995 (type 1), and in December 1996 (type 1). Fifteen of the 17 positive samples were collected in the Gaza Strip, 1 was collected in the West Bank, and 1 was collected in the Israeli city of Ashdod, located close to the Gaza Strip. The AFP surveillance system failed to detect the circulating wild-type viruses. These findings further emphasize the important role that environmental surveillance can play in monitoring the eradication of polioviruses.
Epidemiology of Visceral Mycoses: Analysis of Data in
Annual of the Pathological Autopsy Cases in Japan
Authors: Yamazaki Toshikazu , Kume Hikaru , Murase Setsuko , Yamashita Eriko , Arisawa Mikio
Published: December 2019

The data on visceral mycoses that had been reported in the
Annual of the Pathological Autopsy Cases in Japan
from 1969 to 1994 by the Japanese Society of Pathology were analyzed epidemiologically. The frequency of visceral mycoses among the annual total number of pathological autopsy cases increased noticeably from 1.60% in 1969 to a peak of 4.66% in 1990. Among them, the incidences of candidiasis and aspergillosis increased the most. After 1990, however, the frequency of visceral mycoses decreased gradually. Until 1989, the predominant causative agent was
Candida
, followed in order by
Aspergillus
and
Cryptococcus
. Although the rate of candidiasis decreased by degrees from 1990, the rate of aspergillosis increased up to and then surpassed that of candidiasis in 1991. Leukemia was the major disease underlying the visceral mycoses, followed by solid cancers and other blood and hematopoietic system diseases. Severe mycotic infection has increased over the reported 25-year period, from 6.6% of the total visceral mycosis cases in 1969 to 71% in 1994. The reasons for this decrease of candidiasis combined with an increase of aspergillosis or of severe mycotic infection might be that (i) nonsevere (not disseminated) infections were excluded from the case totals, since they have become controllable by antifungal drugs such as fluconazole, but (ii) the available antifungal drugs were not efficacious against severe infections such as pulmonary aspergillosis, and (iii) the number of patients living longer in an immunocompromised state had increased because of developments in chemotherapy and progress in medical care.
Density and Molecular Epidemiology of
Aspergillus
in Air and Relationship to Outbreaks of
Aspergillus
Infection
Authors: Leenders Alexander C. A. P. , van Belkum Alex , Behrendt Myra , Luijendijk Ad , Verbrugh Henri A.
Published: December 2019

After five patients were diagnosed with nosocomial invasive aspergillosis caused by
Aspergillus fumigatus
and
A. flavus
, a 14-month surveillance program for pathogenic and nonpathogenic fungal conidia in the air within and outside the University Hospital in Rotterdam (The Netherlands) was begun.
A. fumigatus
isolates obtained from the Department of Hematology were studied for genetic relatedness by randomly amplified polymorphic DNA (RAPD) analysis. This was repeated with
A. fumigatus
isolates contaminating culture media in the microbiology laboratory. The density of the conidia of nonpathogenic fungi in the outside air showed a seasonal variation: higher densities were measured during the summer, while lower densities were determined during the fall and winter. Hardly any variation was found in the numbers of
Aspergillus
conidia. We found decreasing numbers of conidia when comparing air from outside the hospital to that inside the hospital and when comparing open areas within the hospital to the closed department of hematology. The increase in the number of patients with invasive aspergillosis could not be explained by an increase in the number of
Aspergillus
conidia in the outside air. The short-term presence of
A. flavus
can only be explained by the presence of a point source, which was probably patient related. Genotyping
A. fumigatus
isolates from the department of hematology showed that clonally related isolates were persistently present for more than 1 year. Clinical isolates of
A. fumigatus
obtained during the outbreak period were different from these persistent clones.
A. fumigatus
isolates contaminating culture media were all genotypically identical, indicating a causative point source. Knowledge of the epidemiology of
Aspergillus
species is necessary for the development of strategies to prevent invasive aspergillosis. RAPD fingerprinting of
Aspergillus
isolates can help to determine the cause of an outbreak of invasive aspergillosis.
Survey of Extended-Spectrum β-Lactamases in Clinical Isolates of
Escherichia coli
and
Klebsiella pneumoniae
: Prevalence of TEM-52 in Korea
Authors: Pai Hyunjoo , Lyu Sen , Lee Ji Hyang , Kim Jungmin , Kwon Youngmi , Kim Jong-Won , Choe Kang Won
Published: December 2019

Two hundred ninety isolates of
Escherichia coli
were investigated for the production of extended-spectrum β-lactamases (ESBLs). Fourteen (4.8%) of the 290 strains were found to produce ESBLs. Each of the 14 strains produced one or two ESBLs, as follows: 10 strains produced TEM-52, 1 strain produced SHV-2a, 1 strain produced SHV-12, 1 strain produced a CMY-1-like enzyme, and 1 strain expressed SHV-2a and a CMY-1-like enzyme. Another two strains for which the MICs of ceftazidime and cefoxitin were high, were probable AmpC enzyme hyperproducers. Because of the high prevalence of TEM-52 in
E. coli
isolates, we further investigated the TEM-type ESBLs produced by
Klebsiella pneumoniae
in order to observe the distribution of TEM-52 enzymes among
Enterobacteriaceae
in Korea. All TEM enzymes produced by 12 strains of
K. pneumoniae
were identified as TEM-52. To evaluate the genetic relatedness among the organisms, ribotyping of TEM-52-producing
E. coli
and
K. pneumoniae
was performed. The ribotyping profiles of the organisms showed similar but clearly different patterns. In conclusion, TEM-52 is the most prevalent TEM-type ESBL in Korea.