bluuebunny/crossref_metadata_2025_split huggingface dataset:
Survival of Aerobic and Anaerobic Bacteria in Purulent Clinical Specimens Maintained in the Copan Venturi Transystem and Becton Dickinson Port-a-Cul Transport Systems
Authors: Citron Diane M. , Warren Yumi A. , Hudspeth Marie K. , Goldstein Ellie J. C. 
Published: December 2019

ABSTRACT
Recovery of aerobic and anaerobic bacteria from clinical specimens maintained in the Copan Venturi Transystem and the Becton Dickinson Port-a-Cul transport was assessed. Of 54 anaerobes, 53 were recovered after 4 h, and 52 were recovered after 24 h, from both systems. After 48 h, 45 and 50 were recovered from the two systems, respectively.
Epidemiology and Microbiology of Surgical Wound Infections
Authors: Giacometti A. , Cirioni O. , Schimizzi A. M. , Del Prete M. S. , Barchiesi F. , D'Errico M. M. , Petrelli E. , Scalise G. 
Published: December 2019

ABSTRACT

This study included 676 surgery patients with signs and symptoms indicative of wound infections, who presented over the course of 6 years. Bacterial pathogens were isolated from 614 individuals. A single etiologic agent was identified in 271 patients, multiple agents were found in 343, and no agent was identified in 62. A high preponderance of aerobic bacteria was observed. Among the common pathogens were
Staphylococcus aureus
(191 patients, 28.2%),
Pseudomonas aeruginosa
(170 patients, 25.2%),
Escherichia coli
(53 patients, 7.8%),
Staphylococcus epidermidis
(48 patients, 7.1%), and
Enterococcus faecalis
(38 patients, 5.6%).
Identification of
Helicobacter pylori
and Other
Helicobacter
Species by PCR, Hybridization, and Partial DNA Sequencing in Human Liver Samples from Patients with Primary Sclerosing Cholangitis or Primary Biliary Cirrhosis
Authors: Nilsson Hans-Olof , Taneera Jalal , Castedal Maria , Glatz Elisabeth , Olsson Rolf , Wadström Torkel 
Published: December 2019

ABSTRACT

Helicobacter pylori
was identified in human liver tissue by PCR, hybridization, and partial DNA sequencing. Liver biopsies were obtained from patients with primary sclerosing cholangitis (
n
= 12), primary biliary cirrhosis (
n
= 12), and noncholestatic liver cirrhosis (
n
= 13) and (as controls) normal livers (
n
= 10). PCR analyses were carried out using primers for the
Helicobacter
genus,
Helicobacter pylori
(the gene encoding a species-specific 26-kDa protein and the 16S rRNA),
Helicobacter bilis
,
Helicobacter pullorum
, and
Helicobacter hepaticus
. Samples from patients with primary biliary cirrhosis and primary sclerosing cholangitis (11 and 9 samples, respectively) were positive by PCR with
Helicobacter
genus-specific primers. Of these 20 samples, 8 were positive with the 16S rRNA primer and 9 were positive with the 26-kDa protein primer of
H. pylori
. These nine latter samples were also positive by Southern blot hybridization for the amplified 26-kDa fragment, and four of those were verified to be
H. pylori
by partial 16S rDNA sequencing. None of the samples reacted with primers for
H. bilis
,
H. pullorum
, or
H. hepaticus
. None of the normal livers had positive results in the
Helicobacter
genus PCR assay, and only one patient in the noncholestatic liver cirrhosis group, a young boy who at reexamination showed histological features suggesting primary sclerosing cholangitis, had a positive result in the same assay.
Helicobacter
positivity was thus significantly more common in patients with cholestatic diseases (20 of 24) than in patients with noncholestatic diseases and normal controls (1 of 23) (
P
= <0.00001). Patients positive for
Helicobacter
genus had significantly higher values of alkaline phosphatases and prothrombin complex than
Helicobacter
-negative patients (
P
= 0.0001 and
P
= 0.0003, respectively). Among primary sclerosing cholangitis patients,
Helicobacter
genus PCR positivity was weakly associated with ulcerative colitis (
P
= 0.05). Significant differences related to blood group or HLA status were not found.
Touchdown Enzyme Time Release-PCR for Detection and Identification of
Chlamydia trachomatis
,
C. pneumoniae
, and
C. psittaci
Using the 16S and 16S-23S Spacer rRNA Genes
Authors: Madico Guillermo , Quinn Thomas C. , Boman Jens , Gaydos Charlotte A. 
Published: December 2019

ABSTRACT

Three touchdown enzyme time release (TETR)-PCR assays were used to amplify different DNA sequences in the variable regions of the 16S and 16S-23S spacer rRNA genes specific for
Chlamydia trachomatis
,
Chlamydia pneumoniae
, and
Chlamydia psittaci
as improved tests for sensitive diagnosis and rapid species differentiation. The TETR-PCR protocol used 60 cycles of amplification, which provided improved analytical sensitivity (0.004 to 0.063 inclusion-forming unit of
Chlamydia
species per PCR). The sensitivity of TETR-PCR with primer set CTR 70-CTR 71 was 96.7%, and the specificity was 99.6%, compared to those of the AMPLICOR PCR for the detection of
C. trachomatis
in vaginal swab samples. TETR-PCR for
C. pneumoniae
with primer set CPN 90-CPN 91 was 90% sensitive and 93.3% specific compared with a nested PCR with primer set CP1/2-CPC/D for clinical respiratory samples. TETR-PCR for
C. psittaci
with primer set CPS 100-CPS 101 showed substantial agreement with cell culturing (κ, 0.78) for animal tissue samples. Primer sets were then combined into a single multiplex TETR-PCR test. The respective 315-, 195-, and 111-bp DNA target products were precisely amplified when DNA from each of the respective
Chlamydia
species or combinations of them was used. Multiplex chlamydia TETR-PCR correctly identified one strain of each of the 15 serovars of
C. trachomatis
, 22 isolates of
C. pneumoniae
, and 20 isolates of
C. psittaci
. The primer sets were specific for each species. No target products were amplified when DNA from
C. pecorum
or a variety of other microorganisms was tested for specificity. TETR-PCR with primers selected for specific sequences in the 16S and 16S-23S spacer rRNA genes is a valuable test that could be used either with individual primers or in a multiplex assay for the identification and differentiation of
Chlamydia
species from culture isolates or for the detection of chlamydiae in clinical samples.
Single-Tube Balanced Heminested PCR for Detecting
Mycobacterium tuberculosis
in Smear-Negative Samples
Authors: García-Quintanilla Albert , Garcia Lourdes , Tudó Griselda , Navarro Maria , González Julià , Jiménez de Anta Maria T. 
Published: December 2019

ABSTRACT

In order to achieve more sensitive and specific results for the rapid diagnosis of tuberculosis, we have developed a new method, named balanced heminested PCR, which avoids the inconvenience of asymmetric amplification and has the advantages of single-tube heminested PCR. This was achieved by replacing the outer primer that participates in both rounds of amplification in the standard heminested technique by another primer containing the sequence of the inner primer attached at its 5′ end. When both techniques were tested for the IS
6110
target of
Mycobacterium tuberculosis
complex in 80 smear-negative culture-positive sputum samples and 60 control samples, the results showed 100% specificity for both techniques and sensitivities of 60 and 75% for heminested PCR and balanced heminested PCR, respectively (
P
= 0.02). In conclusion, the balanced heminested technique shows a higher sensitivity than that of the standard heminested, and it could be applied to any PCR by attaching the inner primer at the 5′ end of the opposite outer primer. Thus, the balanced heminested technique provides a target for the inner primer in both strands, avoiding asymmetric amplification and thereby resulting in a more efficient amplification, and, in practice, a higher sensitivity without loss of specificity and with a minimum risk of cross-contamination.
Isolation and Molecular Characterization of
Clostridium difficile
Strains from Patients and the Hospital Environment in Belarus
Authors: Titov Leonid , Lebedkova Natalia , Shabanov Alexander , Tang Yajarayma J. , Cohen Stuart H. , Silva Joseph 
Published: December 2019

ABSTRACT

Toxigenic
Clostridium difficile
is the most common etiologic agent of hospital-acquired diarrhea in developed countries. The role of this pathogen in nosocomial diarrhea in Eastern Europe has not been clearly established. The goal of this study was to determine the prevalence of
C. difficile
in patients and the hospital environment in Belarus and to characterize these isolates as to the presence of toxin genes and their molecular type.
C. difficile
was isolated from 9 of 509 (1.8%) patients analyzed and recovered from 28 of 1,300 (2.1%) environmental sites cultured. A multiplex PCR assay was used to analyze the pathogenicity locus (PaLoc) of all isolates, and strain identity was determined by an arbitrarily primed PCR (AP-PCR). The targeted sequences for all the genes in the PaLoc were amplified in all
C. difficile
strains examined. A predominantly homogenous group of strains was found among these isolates, with five major AP-PCR groups being identified. Eighty-three percent of environmental isolates were classified into two groups, while patient isolates grouped into three AP-PCR types, two of which were also found in the hospital environment. Although no data on the role of
C. difficile
infection or epidemiology of
C. difficile
-associated diarrhea (CDAD) in this country exist, the isolation of toxigenic
C. difficile
from the hospital environment suggests that this pathogen may be responsible for cases of diarrhea of undiagnosed origin and validates our effort to further investigate the significance of CDAD in Eastern Europe.
Diagnostic and Public Health Dilemma of Lactose-Fermenting
Salmonella enterica
Serotype Typhimurium in Cattle in the Northeastern United States
Authors: McDonough Patrick L. , Shin Sang J. , Lein Donald H. 
Published: December 2019

ABSTRACT

The presence of lactose-fermenting
Salmonella
strains in clinical case materials presented to microbiology laboratories presents problems in detection and identification. Failure to detect these strains also presents a public health problem. The laboratory methods used in detecting lactose-fermenting
Salmonella enterica
serotype Typhimurium from six outbreaks of salmonellosis in veal calves are described. Each outbreak was caused by a multiply-resistant and lactose-fermenting strain of
S. enterica
serotype Typhimurium. The use of Levine eosin-methylene blue agar in combination with screening of suspect colonies for C8 esterase enzyme and inoculation of colonies into sulfide-indole-motility medium for hydrogen sulfide production was particularly effective for their detection. A hypothesis for the creation of lactose-fermenting salmonellae in the environment is presented. It is proposed that the environment and husbandry practices of veal-raising barns provide a unique niche in which lactose-fermenting salmonellae may arise.
Molecular Identification and Epidemiological Tracing of
Pasteurella multocida
Meningitis in a Baby
Authors: Boerlin Patrick , Siegrist Hans H. , Burnens André P. , Kuhnert Peter , Mendez Purita , Prétat Gérard , Lienhard Reto , Nicolet Jacques 
Published: December 2019

ABSTRACT

We report a case of
Pasteurella multocida
meningitis in a 1-month-old baby exposed to close contact with two dogs and a cat but without any known history of injury by these animals. 16S rRNA gene sequencing of the isolate from the baby allowed identification at the subspecies level and pointed to the cat as a possible source of infection. Molecular typing of
Pasteurella
isolates from the animals, from the baby, and from unrelated animals clearly confirmed that the cat harbored the same
P. multocida
subsp.
septica
strain on its tonsils as the one isolated from the cerebrospinal fluid of the baby. This case stresses the necessity of informing susceptible hosts at risk of contracting zoonotic agents about some basic hygiene rules when keeping pets. In addition, this study illustrates the usefulness of molecular methods for identification and epidemiological tracing of
Pasteurella
isolates.
Simple Latex Agglutination Assay for Rapid Serodiagnosis of Human Leptospirosis
Authors: Smits Henk L. , van der Hoorn Menno A. W. G. , Goris Marga G. A. , Gussenhoven George C. , Yersin Claude , Sasaki David M. , Terpstra Wiepko J. , Hartskeerl Rudy A. 
Published: December 2019

ABSTRACT

A newly developed latex agglutination assay for the detection of genus-specific
Leptospira
antibodies in human sera was evaluated. The assay is performed by mixing, on an agglutination card, serum with equal volumes of stabilized antigen-coated, dyed test and control latex beads and is read within 2 min. The latex agglutination test was evaluated with groups of serum samples from patients with leptospirosis and control patients from Hawaii, the Seychelles, Thailand, and The Netherlands. The mean overall sensitivity was 82.3%, and the mean overall specificity was 94.6%. The assay is easy to perform and does not require special skills or equipment. The reagents have a long shelf life, even at tropical temperatures. Together, these factors make the assay suitable for use even at the peripheral level of a health care system as a rapid screening test for leptospirosis.
Genetic and Antigenic Evidence Supports the Separation ofHepatozoon canisandHepatozoon americanumat the Species Level
Authors: Baneth Gad , Barta John R. , Shkap Varda , Martin Donald S. , Macintire Douglass K. , Vincent-Johnson Nancy 
Published: December 2019

ABSTRACTRecognition ofHepatozoon canisandHepatozoon americanumas distinct species was supported by the results of Western immunoblotting of canine anti-H. canisand anti-H. americanumsera againstH. canisgamonts. Sequence analysis of 368 bases near the 3′ end of the 18S rRNA gene from each species revealed a pairwise difference of 13.59%.
Identification by 16S rRNA Gene Analyses of a Potential Novel Mycobacterial Species as an Etiological Agent of Canine Leproid Granuloma Syndrome
Authors: Hughes M. S. , James G. , Ball N. , Scally M. , Malik R. , Wigney D. I. , Martin P. , Chen S. , Mitchell D. , Love D. N. 
Published: December 2019

ABSTRACT

PCR amplifications of the 16S rRNA gene were performed on 46 specimens obtained from 43 dogs with canine leproid granuloma syndrome to help determine its etiology. Sequence capture PCR was applied to 37 paraffin-embedded specimens from 37 dogs, and nested PCR was attempted on DNA from 9 fresh tissue specimens derived from 3 of the 37 aforementioned dogs and from an additional 6 dogs. Molecular analyses of the paraffin-embedded tissues and fresh tissue specimen analyses were performed at separate institutions. PCR products with identical sequences over a 350-bp region encompassing variable regions 2 and 3 of the 16S rRNA gene were obtained from 4 of 37 paraffin-embedded specimens and from all 9 specimens of fresh tissue originating from 12 of the 43 dogs. Identical sequences were determined from amplicons obtained from paraffin-embedded and fresh specimens from one dog. The consensus DNA sequence, amplified from paraffin-embedded tissue and represented by GenBank accession no.
AF144747
, shared highest nucleotide identity (99.4% over 519 bp) with mycobacterial strain IWGMT 90413 but did not correspond exactly to any EMBL or GenBank database sequence. With a probe derived from the V2 region of the novel canine sequence, reverse cross blot hybridization identified an additional four paraffin-embedded specimens containing the same novel sequence. In total, molecular methodologies identified the proposed novel mycobacterial sequence in 16 of 43 dogs with canine leproid granuloma syndrome, indicating that the species represented by this sequence may be the principal etiological agent of canine leproid granuloma syndrome.
Comparison of Protein A Gene Sequencing with Pulsed-Field Gel Electrophoresis and Epidemiologic Data for Molecular Typing of Methicillin-Resistant
Staphylococcus aureus
Authors: Tang Yi-Wei , Waddington Michael G. , Smith Douglas H. , Manahan Janice M. , Kohner Peggy C. , Highsmith Leanne M. , Li Haijing , Cockerill Franklin R. , Thompson Rodney L. , Montgomery Stacy O. , Persing David H. 
Published: December 2019

ABSTRACT

The epidemiologic relatedness of methicillin-resistant
Staphylococcus aureus
(MRSA) isolates is currently determined by analysis of chromosomal DNA restriction patterns by pulsed-field gel electrophoresis (PFGE). We have evaluated an alternative typing system (MicroSeq StaphTrack Kit; Perkin-Elmer Biosystems) based on the sequence analysis of the chromosomally encoded polymorphic repeat X region of the
S. aureus
protein A (
spa
) gene. A total of 69 clinical MRSA isolates were divided into 18 groups according to the number and nucleotide sequences of the
spa
repeats. Molecular typing results obtained both by
spa
sequencing and from the PFGE patterns were concordant except for one group, which contained 20 isolates recovered over a 2-year period from hospitalized patients at the Mayo Clinic. Although the
spa
typing patterns were indistinguishable for those isolates, PFGE analysis yielded seven related but distinguishable patterns. Further coagulase gene sequence analysis subtyped those 20 strains into four groups which followed distinct temporal and geographic distributions. During a 2-year epidemic period there were up to 7 fragment changes in PFGE patterns among epidemiologically related isolates, suggesting that PFGE may be unsuitable for long-term typing of strains involved in epidemics. Although more limited than PFGE in discriminatory power,
spa
sequencing analysis could be used as a screening method for typing of MRSA strains because of the shorter turnaround time, ease of use, and the inherent advantages of sequence analysis, storage, and sharing of information.
Molecular Evolution in a Multidrug-Resistant Lineage of
Streptococcus pneumoniae
: Emergence of Strains Belonging to the Serotype 6B Icelandic Clone That Lost Antibiotic Resistance Traits
Authors: Vilhelmsson Sigurdur E. , Tomasz Alexander , Kristinsson Karl G. 
Published: December 2019

ABSTRACT

Since their first detection in 1988, penicillin-resistant
Streptococcus pneumoniae
isolates have rapidly spread in Iceland to account for close to 20% of all pneumococcal disease in that country by 1993. The major component (70%) of the resistant pneumococci identified from 1989 to 1992 was the progeny of a single multidrug-resistant clone (Icelandic clone) with a homogeneous chromosomal macrorestriction profile and identical multilocus enzyme type expressing serotype 6B and resistance to penicillin, tetracycline, chloramphenicol, erythromycin, and trimethoprim-sulfamethoxazole. The rest of the non-penicillin-susceptible isolates included bacteria with serotype 6A and serogroups 19 and 23. The unique geographic and epidemiological setting and the availability of a complete collection of all non-penicillin-susceptible isolates of
S. pneumoniae
in Iceland prompted us to carry out a molecular epidemiological study to monitor the fate of the Icelandic clone between 1989 and 1996; in addition, we wished to extend the characterization to representative groups of all non-penicillin-susceptible serotype 6B pneumococci which showed variations in antibiotype and which were recovered in Iceland between late 1989 and the end of 1996. Also included in the study were non-penicillin-susceptible isolates of serogroup 23. Pulsed-field gel electrophoresis of
Sma
I-restricted chromosomal DNA and Southern hybridization with the
lytA
DNA probe and probes specific for antibiotic resistance genes were used to characterize pneumococcal isolates. The results show that (i) the Icelandic clone remained the predominant type among penicillin-resistant
S. pneumoniae
through 1996; (ii) the emergence of variants of the Icelandic clone which had lost one or more of the antibiotic resistance phenotypes and/or resistant genes, singly or in combination, was documented during the surveillance period; and (iii) isolates belonging to the internationally spread multidrug-resistant serotype 23F clone were present in the Icelandic collection since late 1989 but did not increase in number during the subsequent years.
Quantitative Analysis of Human Herpesvirus 8 Viral Load Using a Real-Time PCR Assay
Authors: Lallemand Francis , Desire Nathalie , Rozenbaum Willy , Nicolas Jean-Claude , Marechal Vincent 
Published: December 2019

ABSTRACT

We have developed a quantitative real-time PCR (TaqMan) assay aimed at measuring the cellular human herpesvirus 8 (HHV-8) DNA load in various clinical samples. Standard curves were obtained by serial dilutions of a control plasmid containing both HHV-8 (ORF73 gene) and the cellular target (human albumin gene). The assay appeared to be very sensitive (100% detection rate for at least 10 copies per well) and specific and was easily reproducible (less than 3% intra-assay variability, 5% interassay variability). This method allowed us to quantify precisely the average HHV-8 copy number per cell in various persistently HHV-8-infected cell lines (BBG-1 cells,
n
= 200; BC-1 cells,
n
= 59; BCBL-1 cells,
n
= 70). A retrospective study was also conducted to assess the HHV-8 DNA load in 12 human immunodeficiency virus-infected patients with either Kaposi's sarcoma (KS; seven patients monitored over a 3-month period) or multicentric Castleman's disease (MCD; five patients). The HHV-8 DNA load ranged from 0 to 9,171 copies/10
6
cells in low-risk KS patients (T0, I0, S0 according to the classification of the AIDS Clinical Trials group). We also measured the viral loads in MCD patients either during symptomatic periods or during remission. The results are in agreement with previously published data, with high viral loads correlating with clinical symptoms (1.3 × 10
6
copies/10
6
cells) and low viral loads correlating with asymptomatic periods (less than 5,000 copies/10
6
cells).
Characterization of Strains of
Mycoplasma mycoides
subsp.
mycoides
Small Colony Type Isolated from Recent Outbreaks of Contagious Bovine Pleuropneumonia in Botswana and Tanzania: Evidence for a New Biotype
Authors: March John B. , Clark Jason , Brodlie Malcolm 
Published: December 2019

ABSTRACT

Four strains of
Mycoplasma mycoides
subsp.
mycoides
small colony type (MmmSC) isolated from recent outbreaks of contagious bovine pleuropneumonia (CBPP) in Africa have been investigated. One Botswanan strain, M375, displayed numerous and significant phenotypic differences from both contemporary field isolates and older field and vaccine strains (African, Australian, and European strains dating back to 1936). Differences include altered morphology, reduced capsular polysaccharide production, high sensitivity to MmmSC rabbit hyperimmune antisera in vitro, and unique polymorphisms following immunoblotting. While insertion sequence analysis using IS
1634
clearly indicates a close evolutionary relationship to west African strains, hybridization with IS
1296
shows the absence of a band present in all other strains of MmmSC examined. The data suggest that a deletion has occurred in strain M375, which may explain its altered phenotype, including poor growth in vitro and a relative inability to cause septicemia in mice. These characteristics are also exhibited by
Mycoplasma capricolum
subsp.
capripneumoniae
(causal agent of contagious caprine pleuropneumonia [CCPP]), against which M375 antiserum exhibited some activity in vitro (unique among the various MmmSC antisera tested). These findings may have evolutionary implications, since CCPP is believed to be lung specific and without a septicemic phase (unlike CBPP). Since M375 was isolated from a clinical case of CBPP, this novel biotype may be fairly widespread but not normally isolated due to difficulty of culture and/or a potentially altered disease syndrome. Bovine convalescent antisera (obtained from contemporary naturally infected cattle in Botswana) were active against strain M375 in an in vitro growth inhibition test but not against any other strains of MmmSC tested. There exists the possibility therefore, that strain M375 may possess a set of protective antigens different from those of other strains of MmmSC (including vaccine strains). These findings have implications for the control of the current CBPP epidemic in Africa.