bluuebunny/crossref_metadata_2025_split huggingface dataset:
Molecular Typing of
Vibrio parahaemolyticus
Isolates, Obtained from Patients Involved in Food Poisoning Outbreaks in Taiwan, by Random Amplified Polymorphic DNA Analysis
Authors: Wong Hin-Chung , Liu Chi-Chang , Pan Tze-Ming , Wang Tien-Kuei , Lee Chih-Lung , Shih Daniel Yang-Chih 
Published: December 2019

ABSTRACT

Vibrio parahaemolyticus
is one of the most important food-borne pathogens in Taiwan, Japan, and other countries with long coastlines. This paper reports on the development of a new random amplified polymorphic DNA (RAPD) method for the molecular typing of this pathogen. The 10-mer primer 284 (5′-CAG GCG CAC A-3′) was selected to generate polymorphic amplification profiles of the genomic DNA at an annealing temperature of 38°C. A total of 308 clinical isolates of
V. parahaemolyticus
collected during food poisoning outbreaks in Taiwan, mostly occurring between 1993 and 1995, plus 11 environmental and clinical reference strains were analyzed by this RAPD method. A total of 41 polymorphic RAPD patterns were recognized, and these patterns were arbitrarily grouped into 16 types (A to P). Types A, B, C, D, and E were the major types, and subtypes C3, C5, E1, B1, D2, and A2 were the major patterns. The major types were phylogenetically more closely related to each other than to any of the minor types.
Use of PCR in Diagnosis of Human American Tegumentary Leishmaniasis in Rio de Janeiro, Brazil
Authors: Pirmez Claude , da Silva Trajano Valéria , Neto Manoel Paes-Oliveira , da-Cruz Alda Maria , Gonçalves-da-Costa Sylvio Celso , Catanho Marcos , Degrave Wim , Fernandes Octavio 
Published: December 2019

ABSTRACTIn Brazil, the most common etiological agent of American tegumentary leishmaniasis isLeishmania(Viannia)braziliensis. In general, diagnostic techniques envisage the visualization of the parasite, but that technique has a low sensitivity. The main purpose of the present work was to evaluate the PCR as a routine tool for the diagnosis of leishmaniasis. Biopsy specimens from cutaneous or mucosal lesions were taken from 230 individuals from areas whereLeishmaniais endemic: 216 patients who had a clinical picture suggestive ofl leishmaniasis and 14 individuals with cutaneous lesions due to other causes. Each specimen was processed for histopathologic examination, culture, touch preparation, and DNA isolation. Oligonucleotides that amplify the conserved region of the minicircle molecules ofLeishmaniawere used in a hot-start PCR. While at least one conventional technique was positive forLeishmaniafor 62% (134 of 216) of the patients, PCR coupled to hybridization was positive for 94% (203 of 216) of the patients. The 14 patients whose clinical picture was not suggestive of leishmaniasis had negative results by all techniques. The impact of the PCR was striking in mucosal disease. While the disease in only 17% (4 of 24) of the patients could be diagnosed by conventional techniques, PCR was positive for 71% (17 of 24) of the patients. Hybridization showed that all cases of disease were caused by parasites belonging to theVianniasubgenus. Altogether, the results indicate that PCR is a valuable tool for the diagnosis of leishmaniasis on a routine basis and is likely to provide valuable epidemiological information about the disease in countries where it is endemic.
Prevalence of Corynebacterial 16S rRNA Sequences in Patients with Bacterial and “Nonbacterial” Prostatitis
Authors: Tanner Michael A. , Shoskes Daniel , Shahed Asha , Pace Norman R. 
Published: December 2019

ABSTRACT

The etiology of chronic prostatitis syndromes in men is controversial, particularly when positive cultures for established uropathogens are lacking. Although identification of bacteria in prostatic fluid has relied on cultivation and microscopy, most microorganisms in the environment, including some human pathogens, are resistant to cultivation. We report here on an rRNA-based molecular phylogenetic approach to the identification of bacteria in prostate fluid from prostatitis patients. Positive bacterial signals were seen for 65% of patients with chronic prostatitis overall. Seven of 11 patients with bacterial signals but none of 6 patients without bacterial signals were cured with antibiotic-based therapy. Results indicate the occurrence in the prostate fluid of a wide spectrum of bacterial species representing several genera. Most rRNA genes were closely related to those of species belonging to the genera
Corynebacterium
,
Staphylococcus
,
Peptostreptococcus
,
Streptococcus
, and
Escherichia
. Unexpectedly, a wide diversity of
Corynebacterium
species was found in high proportion compared to the proportions of other bacterial species found. A subset of these 16S rRNA sequences represent those of undescribed species on the basis of their positions in phylogenetic trees. These uncharacterized organisms were not detected in control samples, suggesting that the organisms have a role in the disease or are the consequence of the disease. These studies show that microorganisms associated with prostatitis generally occur as complex microbial communities that differ between patients. The results also indicate that microbial communities distinct from those associated with prostatitis may occur at low levels in normal prostatic fluid.
Highly Reproducible Bactericidal Activity Test Results by Using a Modified National Committee for Clinical Laboratory Standards Broth Macrodilution Technique
Authors: Hacek Donna M. , Dressel Dana C. , Peterson Lance R. 
Published: December 2019

ABSTRACT
Bactericidal testing historically has exhibited variable reproducibility, even when prior standardized methods were employed. Several modifications to the National Committee for Clinical Laboratory Standards (NCCLS) broth macrodilution method are proposed to improve reproducibility. Recommended changes from the approved NCCLS guidelines (M21-A and M26-A) include omitting serum supplementation of Mueller-Hinton broth, incubating tubes at 35°C for 24 h with no agitation until they are sampled, running all tests in duplicate with six dilutions instead of nine, reincubating the test for an additional 24 h to resolve discrepant bactericidal activity test results, using a single 0.1-ml sample from each clear tube for subculture, and adopting an alternate method for calculating endpoint determination. In order to test these recommendations in a clinical laboratory setting, we used the modified methodology on 224 separate tests for bactericidal activity. There were 102 serum bactericidal titer (SBT) and 122 minimum bactericidal concentration (MBC) assays performed. By defining reproducibility as agreement between duplicate tests ± 1 dilution, we found 207 of 224 tests (92%) were reproducible at the 24-h subculture point (94% for the SBT assay and 91% for the MBC assay). When the 17 assays with discrepant results were incubated an additional 24 h for a second subculture, only 1 of 224 tests (0.4%) remained discrepant. The method used is practical for a clinical laboratory that chooses to perform bactericidal activity testing and assures a high level of reproducibility between duplicate assays. The total cost of a test was approximately $25.00.
Quantitative Detection of
Borrelia burgdorferi
by Real-Time PCR
Authors: Pahl Andreas , Kühlbrandt Uta , Brune Kay , Röllinghoff Martin , Gessner André 
Published: December 2019

ABSTRACT

Currently, no easy and reliable methods allowing for the quantification of
Borrelia burgdorferi
in tissues of infected humans or animals are available. Due to the lack of suitable assays to detect
B. burgdorferi
CFU and the qualitative nature of the currently performed PCR assays, we decided to exploit the recently developed real-time PCR. This technology measures the release of fluorescent oligonucleotides during the PCR. Flagellin of
B. burgdorferi
was chosen as the target sequence. A linear quantitative detection range of 5 logs with a calculated detection limit of one to three spirochetes per assay reaction mixture was observed. The fact that no signals were obtained with closely related organisms such as
Borrelia hermsii
argues for a high specificity of this newly developed method. A similar method was developed to quantify mouse actin genomic sequences to allow for the standardization of spirochete load. The specificity and sensitivity of the
B. burgdorferi
and the actin real-time PCR were not altered when samples were spiked with mouse cells or spirochetes, respectively. To evaluate the applicability of the real-time PCR, we used the mouse model of Lyme disease. The fate of
B. burgdorferi
was monitored in different tissues from inbred mice and from mice treated with antibiotics. Susceptible C3H/HeJ mice had markedly higher burdens of bacterial DNA than resistant BALB/c mice, and penicillin G treatment significantly reduced the numbers of spirochetes. Since these results show a close correlation between clinical symptoms and bacterial burden of tissues, we are currently analyzing human biopsy specimens to evaluate the real-time PCR in a diagnostic setting.
Rapid and Reliable Identification of
Streptococcus pneumoniae
Isolates by Pneumolysin-Mediated Agglutination
Authors: Cima-Cabal M. D. , Vázquez F. , de los Toyos J. R. , Méndez F. J. 
Published: December 2019

ABSTRACT
A pneumolysin-based agglutination test which allows an easy, rapid, cost-effective, and accurate (100% specific and 95% sensitive) discrimination between pneumococci and other related human and animal pathogenic bacterial strains has been assayed.
Comparison of Three Commercial Systems for Identification of Yeasts Commonly Isolated in the Clinical Microbiology Laboratory
Authors: Wadlin Jill K. , Hanko Gayle , Stewart Rebecca , Pape John , Nachamkin Irving 
Published: December 2019

ABSTRACT

We evaluated three commercial systems (RapID Yeast Plus System; Innovative Diagnostic Systems, Norcross, Ga.; API 20C Aux; bioMerieux-Vitek, Hazelwood, Mo.; and Vitek Yeast Biochemical Card, bioMerieux-Vitek) against an auxinographic and microscopic morphologic reference method for the ability to identify yeasts commonly isolated in our clinical microbiology laboratory. Two-hundred one yeast isolates were compared in the study. The RapID Yeast Plus System was significantly better than either API 20C Aux (193 versus 167 correct identifications;
P
< 0.0001) or the Vitek Yeast Biochemical Card (193 versus 173 correct identifications;
P
= 0.003) for obtaining correct identifications to the species level without additional testing. There was no significant difference between results obtained with API 20C Aux and the Vitek Yeast Biochemical Card system (
P
= 0.39). The API 20C Aux system did not correctly identify any of the
Candida krusei
isolates (
n
= 23) without supplemental testing and accounted for the major differences between the API 20C Aux and RapID Yeast Plus systems. Overall, the RapID Yeast Plus System was easy to use and is a good system for the routine identification of clinically relevant yeasts.
PCR Detection of
Yersinia pestis
in Fleas: Comparison with Mouse Inoculation
Authors: Engelthaler David M. , Gage Kenneth L. , Montenieri John A. , Chu May , Carter Leon G. 
Published: December 2019

ABSTRACT

The “gold standard” for identifying
Yersinia pestis
-infected fleas has been inoculation of mice with pooled flea material. Inoculated mice are monitored for 21 days, and those that die are further analyzed for
Y. pestis
infection by fluorescent-antibody assay and/or culture. PCR may provide a more rapid and sensitive alternative for identifying
Y. pestis
in fleas. To compare these assays, samples were prepared from 381 field-collected fleas. Each flea was analyzed individually by both PCR and mouse inoculation. Sixty of the 381 flea samples were positive for
Y. pestis
by PCR; 48 of these PCR-positive samples caused death in mice (80.0% agreement). None of the 321 PCR-negative samples caused death. Among the 12 mice that survived inoculation with PCR-positive samples, 10 were later demonstrated by serology or culture to have been infected with
Y. pestis
. This suggests that death of inoculated mice is less reliable than PCR as an indicator of the presence of
Y. pestis
in flea samples. Mouse inoculation assays produce results that are comparable to PCR only when surviving as well as dead mice are analyzed for infection. The rapidity and sensitivity (10 to 100 CFU of
Y. pestis
) of PCR suggest that it could serve as a useful alternative to mouse inoculation for routine plague surveillance and outbreak investigations.
Two Cases of
Chromobacterium violaceum
Infection after Injury in a Subtropical Region
Authors: Lee J. , Kim J. S. , Nahm C. H. , Choi J. W. , Kim J. , Pai S. H. , Moon K. H. , Lee K. , Chong Y. 
Published: December 2019

ABSTRACT

Chromobacterium violaceum
is a gram-negative rod and is isolated from soil and water in tropical and subtropical regions. The species have pigmented and nonpigmented colony types. Infections caused by nonpigmented strains are rare. We report on two cases of infection caused by both pigmented and nonpigmented strains of
C. violaceum
. Two 24-year-old Korea Airline stewardesses were admitted to Inha University Hospital, Inchon, South Korea, on 9 August 1997, 3 days after an airplane accident in Guam. Both had multiple lacerations on exposed parts of their bodies. There was swelling, tenderness, and pus discharge. The wounds contained many small fragments of stones and weeds. A pigmented strain was isolated from the left hand and a nonpigmented strain was isolated from the left knee of one patient. For the other patient only a nonpigmented strain was isolated from a foot wound. The nonpigmented colonies from the left-knee and the left-foot wounds did not produce any pigment even after an extended period of incubation. The biochemical characteristics were the same for each strain except for oxidase and indole reactions. The pigmented strain was oxidase negative and indole positive, whereas the nonpigmented strains were oxidase positive and indole negative. The patients were successfully treated by debridement and with appropriate antibiotics.
Borrelia burgdorferiin Tick Cell Culture Modulates Expression of Outer Surface Proteins A and C in Response to Temperature
Authors: Obonyo Marygorret , Munderloh Ulrike G. , Fingerle Volker , Wilske Bettina , Kurtti Timothy J. 
Published: December 2019

ABSTRACTThe Lyme disease spirocheteBorrelia burgdorferisensu stricto downregulates outer surface protein A (OspA) and upregulates outer surface protein C (OspC) during tick feeding. The switching of these proteins correlates with increased spirochetal infectivity for the mammal. We examined the effect of temperature on differential expression of OspA and OspC byB. burgdorfericocultivated with a cell line isolated from the vector tickIxodes scapularis. The effect of incubation at 31, 34, or 37°C on expression of OspA and OspC byB. burgdorferiJMNT and N40 was analyzed by indirect fluorescent-antibody microscopy, polyacrylamide gel electrophoresis, and immunoblotting. The amount of OspA relative to the amount of flagellin was highest in spirochetes cocultivated with tick cells at 31°C and declined with increasing temperature in both strains. OspC production was enhanced in spirochetes cocultivated with tick cells at 37°C. Spirochetes grown axenically in BSK-H medium also produced more OspC at 37°C, but OspA content was not appreciably affected by temperature. Our findings indicate that temperature, along with cultivation in a tick cell culture system, plays a role in the differential expression of OspA and enhances differential expression of OspC by spirochetes.
Detection and Identification ofEhrlichia,Borrelia burgdorferiSensu Lato, andBartonellaSpecies in DutchIxodes ricinusTicks
Authors: Schouls Leo M. , Van De Pol Ingrid , Rijpkema Sjoerd G. T. , Schot Corrie S. 
Published: December 2019

ABSTRACTA sensitive and specific PCR hybridization assay was developed for the simultaneous detection and identification ofEhrlichiaandBorrelia burgdorferisensu lato. In separate assays the 16S rRNA gene ofEhrlichiaspecies and the 23S-5S rRNA spacer region ofB. burgdorferisensu lato were amplified and labeled by PCR. These PCR products were used in a reverse line blot hybridization assay in which oligonucleotide probes are covalently linked to a membrane in parallel lines. Hybridization of the samples with the oligonucleotide probes on this membrane enabled the simultaneous detection and identification ofEhrlichia,B. burgdorferi, andBartonellaspecies in 40 different samples. The application of the assay to DNA extracts from 121Ixodes ricinusticks collected from roe deer demonstrated that 45% of these ticks carriedEhrlichiaDNA. More than half of these positive ticks carried species with 16S rRNA gene sequences closely related to those ofE. phagocytophilaand the human granulocytic ehrlichiosis agent. The majority of the other positive ticks were infected with a newly identifiedEhrlichia-like species. In addition, 13% of the ticks were infected with one or moreB. burgdorferigenospecies. In more than 70% of the ticks 16S rRNA gene sequences forBartonellaspecies or other species closely related toBartonellawere found. In five of the ticks bothEhrlichiaandB. burgdorferispecies were detected.
Relationship between
Helicobacter pylori iceA
,
cagA
, and
vacA
Status and Clinical Outcome: Studies in Four Different Countries
Authors: Yamaoka Yoshio , Kodama Tadashi , Gutierrez Oscar , Kim Jong G. , Kashima Kei , Graham David Y. 
Published: December 2019

ABSTRACT

There is continuing interest in identifying
Helicobacter pylori
virulence factors that might predict the risk for symptomatic clinical outcomes. It has been proposed that
iceA
and
cagA
genes are such markers and can identify patients with peptic ulcers. We compared
H. pylori
isolates from four countries, looking at the
cagA
and
vacA
genotypes,
iceA
alleles, and presentation of the infection. We used PCR to examine
iceA
,
vacA
, and
cagA
status of 424
H. pylori
isolates obtained from patients with different clinical presentations (peptic ulcer, gastric cancer, and atrophic gastritis). The
H. pylori
isolates examined included 107 strains from Bogota, Colombia, 70 from Houston, Tex., 135 from Seoul, Korea, and 112 from Kyoto, Japan. The predominant genotype differed among countries: the
cagA
-positive
iceA1 vacA
s1c-m1 genotype was predominant in Japan and Korea, the
cagA
-positive
iceA2 vacA
s1b-m1 genotype was predominant in the United States, and the
cagA
-positive
iceA2 vacA
s1a-m1 genotype was predominant in Colombia. There was no association between the
iceA
,
vacA
, or
cagA
status and clinical outcome in patients in the countries studied.
iceA
status shows considerable geographic differences, and neither
iceA
nor combinations of
iceA
,
vacA
, and
cagA
were helpful in predicting the clinical presentation of an
H. pylori
infection.
Evaluation of Mycology Laboratory Proficiency Testing
Authors: Reilly Andrew A. , Salkin Ira F. , McGinnis Michael R. , Gromadzki Sally , Pasarell Lester , Kemna Maggi , Higgins Nancy , Salfinger Max 
Published: December 2019

ABSTRACT

Changes over the last decade in overt proficiency testing (OPT) regulations have been ostensibly directed at improving laboratory performance on patient samples. However, the overt (unblinded) format of the tests and regulatory penalties associated with incorrect values allow and encourage laboratorians to take extra precautions with OPT analytes. As a result OPT may measure optimal laboratory performance instead of the intended target of typical performance attained during routine patient testing. This study addresses this issue by evaluating medical mycology OPT and comparing its fungal specimen identification error rates to those obtained in a covert (blinded) proficiency testing (CPT) program. Identifications from 188 laboratories participating in the New York State mycology OPT from 1982 to 1994 were compared with the identifications of the same fungi recovered from patient specimens in 1989 and 1994 as part of the routine procedures of 88 of these laboratories. The consistency in the identification of OPT specimens was sufficient to make accurate predictions of OPT error rates. However, while the error rates in OPT and CPT were similar for
Candida albicans
, significantly higher error rates were found in CPT for
Candida tropicalis
,
Candida glabrata
, and other common pathogenic fungi. These differences may, in part, be due to OPT’s use of ideal organism representatives cultured under optimum growth conditions. This difference, as well as the organism-dependent error rate differences, reflects the limitations of OPT as a means of assessing the quality of routine laboratory performance in medical mycology.
Evaluation of Statens Serum Institut Enteric Medium for Detection of Enteric Pathogens
Authors: Blom Marianne , Meyer Aase , Gerner-Smidt Peter , Gaarslev Knud , Espersen Frank 
Published: December 2019

ABSTRACT

The efficacy of the Statens Serum Institut (SSI) enteric medium for isolation and direct identification of enteric pathogens was evaluated. Six different biochemical reactions can be read by using the SSI enteric medium, allowing direct identification of a range of enteric pathogens. All 248 gram-negative bacterial species that were tested grew on the SSI enteric medium. Only 10 of 248 bacteria (4%) showed discrepant results in the biochemical reactions, and none of these were enteric pathogens. Forty-three of 47 enteric pathogens (92%) produced identical rates of semiquantitative growth on the SSI enteric medium and 5% blood agar, whereas three
Vibrio
spp. and one
Aeromonas
spp. showed reduced growth. Gram-positive bacteria did not grow on the SSI enteric medium. Most enteric pathogens had a detection limit of 50 bacteria per ml of feces, but higher numbers of
Vibrio
spp. and some
Shigella
spp. were required for detection. The growth rates of 125 enteric pathogens and 12
Yersinia
spp. on the SSI enteric medium, xylose lysine deoxycholate (XLD), Hektoen enteric (HE),
Salmonella-Shigella
(SS), and cefsulodin-irgasan-novobiocin (CIN) agar were compared. Detection rates after application of 200 CFU were 99% for SSI enteric medium, 92% for XLD, 88% for HE, and 82% for SS agar. The 12
Yersinia
spp. grew excellently on both the SSI enteric medium and CIN agar. We conclude that the performance of the SSI enteric medium compares favorably to those of other media tested. Its ability to detect
Yersinia
spp. may limit the number of media needed in the typical laboratory. The direct identification of enteric pathogens on the medium may also provide a more rapid diagnosis.