bluuebunny/crossref_metadata_2025_split huggingface dataset:
Phenotypic Diversity of Enterotoxigenic
Escherichia coli
Strains from a Community-Based Study of Pediatric Diarrhea in Periurban Egypt
Authors: Peruski Leonard F. , Kay Bradford A. , El-Yazeed Remon Abu , El-Etr Sahar H. , Cravioto Alejandro , Wierzba Thomas F. , Rao Malla , El-Ghorab Nemat , Shaheen Hind , Khalil Sami B. , Kamal Karim , Wasfy Momtaz O. , Svennerholm Ann-Mari , Clemens John D. , Savarino Stephen J. 
Published: December 2019

ABSTRACT

No past studies of diarrhea in children of the Middle East have examined in detail the phenotypes of enterotoxigenic
Escherichia coli
(ETEC) strains, which are important pathogens in this setting. During a prospective study conducted from November 1993 to September 1995 with 242 children under 3 years of age with diarrhea living near Alexandria, Egypt, 125 episodes of diarrhea were positive for ETEC. ETEC strains were available for 98 of these episodes, from which 100 ETEC strains were selected and characterized on the basis of enterotoxins, colonization factors (CFs), and O:H serotypes. Of these representative isolates, 57 produced heat-stable toxin (ST) only, 34 produced heat-labile toxin (LT) only, and 9 produced both LT and ST. Twenty-three ETEC strains expressed a CF, with the specific factors being CF antigen IV (CFA/IV; 10 of 23; 43%), CFA/II (5 of 23; 22%), CFA/I (3 of 23; 13%), PCFO166 (3 of 23; 13%), and CS7 (2 of 23; 9%). No ETEC strains appeared to express CFA/III, CS17, or PCFO159. Among the 100 ETEC strains, 47 O groups and 20 H groups were represented, with 59 O:H serotypes. The most common O serogroups were O159 (13 strains) and O43 (10 strains). O148 and O21 were each detected in five individual strains, O7 and O56 were each detected in four individual strains, O73, O20, O86, and O114 were each detected in three individual strains, and O23, O78, O91, O103, O128, and O132 were each detected in two individual strains. The most common H serogroups were H4 (16 strains), 12 of which were of serogroup O159; H2 (9 strains), all of which were O43; H18 (6 strains); H30 (6 strains); and H28 (5 strains); strains of the last three H serogroups were all O148. Cumulatively, our results suggest a high degree of clonal diversity of disease-associated ETEC strains in this region. As a low percentage of these strains expressed a CF, it remains possible that other adhesins for which we either did not assay or that are as yet undiscovered are prevalent in this region. Our findings point out some potential barriers to effective immunization against ETEC diarrhea in this population and emphasize the need to identify additional protective antigens commonly expressed by ETEC for inclusion in future vaccine candidates.
Simple and Accurate PCR-Based System for Typing Vacuolating Cytotoxin Alleles of
Helicobacter pylori
Authors: Atherton J. C. , Cover T. L. , Twells R. J. , Morales M. R. , Hawkey C. J. , Blaser M. J. 
Published: December 2019

ABSTRACT

Alleles of the vacuolating cytotoxin gene (
vacA
) of
Helicobacter pylori
vary between strains, particularly in the region encoding the signal sequence (which may be type s1 or s2) and the midregion (which may be type m1 or m2). Using a PCR-based typing system developed in the United States, we showed that 36 strains from Asia and South America were all
vacA
signal sequence type s1; 3 were midregion type m1 and 11 were m2, but 22 could not be typed for the
vacA
midregion. All strains possessed
cagA
(cytotoxin-associated gene A), another virulence marker.
vacA
nucleotide sequence analysis showed that midregion typing failure was due to base substitutions at the primer annealing sites. Using the new sequence data, we developed two new PCR-based
vacA
midregion typing systems, both of which correctly typed 41 U.S. strains previously typed by the old system and successfully typed all 36 of the non-U.S. strains. All previously untypeable strains were
vacA
m1, other than one m1/m2 hybrid. In summary, we describe and validate a simple PCR-based system for typing vacuolating cytotoxin (
vacA
) alleles of
H. pylori
and show that this system correctly identifies the signal and midregion types of
vacA
in 77 strains from Asia and North and South America.
Colonization of Mexican Patients by Multiple
Helicobacter pylori
Strains with Different
vacA
and
cagA
Genotypes
Authors: Morales-Espinosa Rosario , Castillo-Rojas Gonzalo , Gonzalez-Valencia Gerardo , Ponce de León Sergio , Cravioto Alejandro , Atherton John C. , López-Vidal Yolanda 
Published: December 2019

ABSTRACT

Helicobacter pylori
virulence determinants have not previously been studied in detail in Latin Americans with
H. pylori
infections. We characterized the
vacA
(vacuolating cytotoxin gene A) and
cagA
(cytotoxin-associated gene A) types of more than 400 single-colony isolates from 20 patients in Mexico City. For 17 patients
H. pylori
strains of two or more different
vacA
genotypes were isolated from gastric biopsy specimens, indicating infection with two or more strains of
H. pylori
. The most frequent
vacA
genotype was s1b/m1.
vacA
diversity was more marked than that described previously, in that isolates from seven patients had untypeable
vacA
midregions and isolates from nine patients had type s1 signal sequence coding regions which could not be further subtyped. Previously undescribed
vacA
type s2/m1 strains were found in five patients. All patients were infected with
cagA
-positive strains, but occasionally, these coexisted with small numbers of
cagA
-negative strains. In conclusion, coinfection with multiple
H. pylori
strains is common in Mexico, and
vacA
in these strains is genetically more diverse than has been described in other populations.
Evaluation of Two Rapid Assays for Detection of
Clostridium difficile
Toxin A in Stool Specimens
Authors: Fedorko Daniel P. , Engler Howard D. , O’Shaughnessy Elizabeth M. , Williams Esther C. , Reichelderfer Cynthia J. , Smith William I. 
Published: December 2019

ABSTRACT

Rapid laboratory diagnosis of
Clostridium difficile
-associated diarrhea (CDAD) is highly desirable in the setting of hospital cost containment. We tested 654 stool specimens to compare the performance of two assays for rapid detection of toxin A, the Immunocard Toxin A test (Meridian Diagnostics, Inc.) and the Culturette Brand Toxin CD enzyme immunoassay (EIA) (Becton Dickinson Microbiology Systems), with a cytotoxin assay (Cytotoxi Test; Advanced Clinical Diagnostics) and culture on cycloserine-cefoxitin-fructose agar followed by determination of the production of toxins A and B. A chart review was performed for patients whose stool specimens provided positive results on one to three of the assays. With the “gold standard” of all four assays positive or chart review evidence of CDAD, 97 (14.8%) stool specimens were positive by one or more assays and 557 (85.2%) were negative by all methods. Total agreement for all assays was 90.5% (592 of 654). The sensitivity, specificity, positive predictive value, and negative predictive value for toxigenic culture were 94.7, 98.6, 87.1, and 99.5%, respectively, for toxigenic culture; 87.7, 98.6, 86.2, and 98.8%, respectively, for the cytotoxin assay; 71.9, 99.3, 91.1, and 97.3%, respectively, for the Immunocard; and 68.4, 99.1, 88.6, and 96.9%, respectively, for the Culturette EIA. While easy to perform and highly specific, these rapid assays do not appear to be sufficient for accurate diagnosis of CDAD.
Prevalence of Group A Rotavirus, Human Calicivirus, Astrovirus, and Adenovirus Type 40 and 41 Infections among Children with Acute Gastroenteritis in Dijon, France
Authors: Bon F. , Fascia P. , Dauvergne M. , Tenenbaum D. , Planson H. , Petion A. M. , Pothier P. , Kohli E. 
Published: December 2019

ABSTRACT
Group A rotaviruses, human caliciviruses, astroviruses, and adenovirus types 40 and 41 were detected by enzyme immunoassay or reverse transcription-PCR in 61, 14, 6, and 3% of stool specimens from 414 children consulting for gastroenteritis between 1995 and 1998. These data highlight the importance of caliciviruses in infantile gastroenteritis. Among these, Norwalk-like viruses belonging to genogroup II were predominant.
Multicenter Evaluation of AMPLICOR and Automated COBAS AMPLICOR CT/NG Tests for
Neisseria gonorrhoeae
Authors: Martin David H. , Cammarata Cathy , Van Der Pol Barbara , Jones Robert B. , Quinn Thomas C. , Gaydos Charlotte A. , Crotchfelt Kimberly , Schachter Julius , Moncada Jeanne , Jungkind D. , Turner Buffy , Peyton Cynthia 
Published: December 2019

ABSTRACT

The fully automated COBAS AMPLICOR CT/NG and semiautomated AMPLICOR CT/NG tests were evaluated in a multicenter trial for their ability to detect
Neisseria gonorrhoeae
infections. Test performance compared to that of culturing was evaluated for 2,192 matched endocervical swab and urine specimens obtained from women and for 1,981 matched urethral swab and urine specimens obtained from men. Culture-negative, PCR-positive specimens that tested positive in a confirmatory PCR test for an alternative target sequence within the
N. gonorrhoeae
16S rRNA gene were considered to be true positives. The overall prevalences of gonorrhea were 6.6% in women and 20.1% in men. The COBAS AMPLICOR and AMPLICOR formats yielded concordant results for 98.8% of the specimens and exhibited virtually identical sensitivities and specificities. The results that follow are for the COBAS AMPLICOR format. With the infected patient as the reference standard, the resolved sensitivities of PCR were 92.4% for endocervical swab specimens and 64.8% for female urine specimens. There were no significant differences in these rates between women with and without symptoms. Among symptomatic men, COBAS AMPLICOR sensitivities were 94.1% for urine and 98.1% for urethral swabs; for asymptomatic men, the results were 42.3 and 73.1%, respectively. In comparison, the sensitivities of culturing were 84.8% for endocervical specimens, 92.7% for symptomatic male urethral specimens, and only 46.2% for urethral specimens obtained from asymptomatic men. When PCR results were analyzed as if only a single test had been performed on a single specimen type, the resolved sensitivity was always higher. The resolved specificities of PCR were 99.5% for endocervical swab specimens, 99.8% for female urine specimens, 98.9% for male urethral swab specimens, and 99.9% for male urine specimens. The internal control revealed that 2.1% of specimens were inhibitory when initially tested. Nevertheless, valid results were obtained for 99.2% of specimens because 60.0% of the inhibitory specimens were not inhibitory when a second aliquot was tested. The COBAS AMPLICOR CT/NG test for
N. gonorrhoeae
exhibited high sensitivity and specificity with urethral swab and urine specimens from men and endocervical swab specimens from women and thus is well suited for diagnosing and screening for
N. gonorrhoeae
infection.
Serodiagnosis of Louse-Borne Relapsing Fever with Glycerophosphodiester Phosphodiesterase (GlpQ) from
Borrelia recurrentis
Authors: Porcella Stephen F. , Raffel Sandra J. , Schrumpf Merry E. , Schriefer Martin E. , Dennis David T. , Schwan Tom G. 
Published: December 2019

ABSTRACT

Human louse-borne relapsing fever occurs in sporadic outbreaks in central and eastern Africa that are characterized by significant morbidity and mortality. Isolates of the causative agent,
Borrelia recurrentis
, were obtained from the blood of four patients during a recent epidemic of the disease in southern Sudan. The
glpQ
gene, encoding glycerophosphodiester phosphodiesterase, from these isolates was sequenced and compared with the
glpQ
sequences obtained from other relapsing-fever spirochetes. Previously we showed that GlpQ of
Borrelia hermsii
is an immunogenic protein with utility as a serological test antigen for discriminating tick-borne relapsing fever from Lyme disease. In the present work, we cloned and expressed the
glpQ
gene from
B. recurrentis
and used recombinant GlpQ in serological tests. Acute- and convalescent-phase serum samples obtained from 42 patients with louse-borne relapsing fever were tested with an indirect immunofluorescence assay (IFA) and an enzyme-linked immunosorbent assay (ELISA) that used whole cells of
B. recurrentis
and with immunoblotting to whole-cell lysates of the spirochete and
Escherichia coli
producing recombinant GlpQ. The geometric mean titers of the acute- and convalescent-phase serum samples measured by IFA were 1:83 and 1:575, respectively. The immunoblot analysis identified a high level of reactivity and seroconversion to GlpQ, and the assay was more sensitive than the whole-cell IFA and ELISA using purified, recombinant histidine-tagged GlpQ. Serum antibodies to GlpQ and other antigens persisted for 27 years in one patient. We conclude that assessment of anti-GlpQ antibodies will allow serological confirmation of louse-borne relapsing fever and determination of disease prevalence.
Spoligotyping and Polymorphic GC-Rich Repetitive Sequence Fingerprinting of
Mycobacterium tuberculosis
Strains Having Few Copies of IS
6110
Authors: Yang Z. H. , Ijaz K. , Bates J. H. , Eisenach K. D. , Cave M. D. 
Published: December 2019

ABSTRACT

Several genetic loci have been utilized to genotype isolates of
Mycobacterium tuberculosis
. A shortcoming of the most commonly used method, IS
6110
fingerprinting, is that it does not adequately discriminate between isolates having few copies of IS
6110
. This study was undertaken to compare pTBN12 fingerprinting of polymorphic GC-rich repetitive sequence genes and spoligotyping of the direct repeat locus as secondary typing procedures for
M. tuberculosis
isolates having fewer than six copies of IS
6110
. A total of 88 isolates (100% of the isolates with fewer than six copies of IS
6110
isolated in Arkansas during 1996 and 1997) were included in this study. Among the 88 isolates, 34 different IS
6110
patterns were observed, 10 of which were shared by more than 1 isolate, involving a total of 64 isolates. The 64 isolates were subdivided into 13 clusters (containing 37 isolates) and 27 unique isolates based on a combination of IS
6110
and pTBN12 fingerprinting and into 11 clusters (containing 51 isolates) and 13 unique isolates based on a combination of IS
6110
fingerprinting and spoligotyping. Identical spoligotypes were found among isolates having different IS
6110
patterns, as well as among isolates showing different pTBN12 patterns. In contrast, all isolates that had different IS
6110
patterns were found to be unique by pTBN12 typing. The clustering rate was 73, 58, and 42%, respectively, for IS
6110
fingerprinting alone, IS
6110
fingerprinting and spoligotyping combined, and IS
6110
and pTBN12 combined fingerprinting. The data indicate that the pTBN12 method has greater discriminating power among low-copy-number isolates than does spoligotyping.
Use of a Repetitive DNA Probe To Type Clinical and Environmental Isolates of
Aspergillus flavus
from a Cluster of Cutaneous Infections in a Neonatal Intensive Care Unit
Authors: James Michael J. , Lasker Brent A. , McNeil Michael M. , Shelton Mark , Warnock David W. , Reiss Errol 
Published: December 2019

ABSTRACT

Aspergillus flavus
is second to
A. fumigatus
as a cause of invasive aspergillosis, but no standard method exists for molecular typing of strains from human sources. A repetitive DNA sequence cloned from
A. flavus
and subcloned into a pUC19 vector, pAF28, was used to type 18 isolates from diverse clinical, environmental, and geographic sources. The restriction fragment length polymorphisms generated with
Eco
RI- or
Pst
I-digested genomic DNA and probed with digoxigenin-labeled pAF28 revealed complete concordance between patterns. Eighteen distinct fingerprints were observed. The probe was used to investigate two cases of cutaneous
A. flavus
infection in low-birth-weight infants in a neonatal intensive care unit (NICU). Both infants were transported by the same ambulance and crew to the NICU on the same day.
A. flavus
strains of the same genotype were isolated from both infants, from a roll of tape used to fasten their umbilical catheters, from a canvas bag used to store the tape in the ambulance, and from the tape tray in the ambulance isolette. These cases highlight the need to consider exposures in critically ill neonates that might occur during their transport to the NICU and for stringent infection control practices. The hybridization profiles of strains from a second cluster of invasive
A. flavus
infections in two pediatric hematology-oncology patients revealed a genotype common to strains from a definite case patient and a health care worker. A probable case patient was infected with a strain with a genotype different from that of the strain from the definite case patient but highly related to that of an environmental isolate. The high degree of discrimination and reproducibility obtained with the pAF28 probe underscores its utility for typing clinical and environmental isolates of
A. flavus
.
16S Ribosomal DNA Sequence Analysis of a Large Collection of Environmental and Clinical Unidentifiable Bacterial Isolates
Authors: Drancourt Michel , Bollet Claude , Carlioz Antoine , Martelin Rolland , Gayral Jean-Pierre , Raoult Didier 
Published: December 2019

ABSTRACT

Some bacteria are difficult to identify with phenotypic identification schemes commonly used outside reference laboratories. 16S ribosomal DNA (rDNA)-based identification of bacteria potentially offers a useful alternative when phenotypic characterization methods fail. However, as yet, the usefulness of 16S rDNA sequence analysis in the identification of conventionally unidentifiable isolates has not been evaluated with a large collection of isolates. In this study, we evaluated the utility of 16S rDNA sequencing as a means to identify a collection of 177 such isolates obtained from environmental, veterinary, and clinical sources. For 159 isolates (89.8%) there was at least one sequence in GenBank that yielded a similarity score of ≥97%, and for 139 isolates (78.5%) there was at least one sequence in GenBank that yielded a similarity score of ≥99%. These similarity score values were used to defined identification at the genus and species levels, respectively. For isolates identified to the species level, conventional identification failed to produce accurate results because of inappropriate biochemical profile determination in 76 isolates (58.7%), Gram staining in 16 isolates (11.6%), oxidase and catalase activity determination in 5 isolates (3.6%) and growth requirement determination in 2 isolates (1.5%). Eighteen isolates (10.2%) remained unidentifiable by 16S rDNA sequence analysis but were probably prototype isolates of new species. These isolates originated mainly from environmental sources (
P
= 0.07). The 16S rDNA approach failed to identify
Enterobacter
and
Pantoea
isolates to the species level (
P
= 0.04; odds ratio = 0.32 [95% confidence interval, 0.10 to 1.14]). Elsewhere, the usefulness of 16S rDNA sequencing was compromised by the presence of 16S rDNA sequences with >1% undetermined positions in the databases. Unlike phenotypic identification, which can be modified by the variability of expression of characters, 16S rDNA sequencing provides unambiguous data even for rare isolates, which are reproducible in and between laboratories. The increase in accurate new 16S rDNA sequences and the development of alternative genes for molecular identification of certain taxa should further improve the usefulness of molecular identification of bacteria.
Molecular Surveillance of European Quinolone-Resistant Clinical Isolates ofPseudomonas aeruginosaandAcinetobacterspp. Using Automated Ribotyping
Authors: Brisse Sylvain , Milatovic Dana , Fluit A. C. , Kusters Karlijn , Toelstra Annet , Verhoef Jan , Schmitz Franz-Josef 
Published: December 2019

ABSTRACTNosocomial isolates ofPseudomonas aeruginosaandAcinetobacterspp. exhibit high rates of resistance to antibiotics and are often multidrug resistant. In a previous study (D. Milatovic, A. Fluit, S. Brisse, J. Verhoef, and F. J. Schmitz, Antimicrob. Agents Chemother. 44:1102–1107, 2000), isolates of these species that were resistant to sitafloxacin, a new advanced-generation fluoroquinolone with a high potency and a broad spectrum of antimicrobial activity, were found in high proportion in 23 European hospitals. Here, we investigate the clonal diversity of the 155P. aeruginosaand 145Acinetobacterspp. sitafloxacin-resistant isolates from that study by automated ribotyping. Numerous ribogroups (sets of isolates with indistinguishable ribotypes) were found among isolates ofP. aeruginosa(n= 34) andAcinetobacterspp. (n= 16), but the majority of the isolates belonged to a limited number of major ribogroups. Sitafloxacin-resistant isolates (MICs > 2 mg/liter, used as a provisional breakpoint) showed increased concomitant resistance to piperacillin, piperacillin-tazobactam, ceftriaxone, ceftazidime, amikacin, gentamicin, and imipenem. The major ribogroups were repeatedly found in isolates from several European hospitals; these isolates showed higher levels of resistance to gentamicin and imipenem, and some of them appeared to correspond to previously described multidrug-resistant international clones ofP. aeruginosa(serotype O:12) andAcinetobacter baumannii(clones I and II). Automated ribotyping, when used in combination with more discriminatory typing methods, may be a convenient library typing system for monitoring future epidemiological dynamics of geographically widespread multidrug-resistant bacterial clones.
Rapid Quantification and Differentiation of Human Polyomavirus DNA in Undiluted Urine from Patients after Bone Marrow Transplantation
Authors: Biel Stefan S. , Held Thomas K. , Landt Olfert , Niedrig Matthias , Gelderblom Hans R. , Siegert Wolfgang , Nitsche Andreas 
Published: December 2019

ABSTRACTA combined PCR assay was developed for the detection and typing of human polyomavirus (huPoV) in clinical samples, consisting of (i) a qualitative seminested PCR assay (snPCR) to discriminate between huPoV BK and JC and (ii) a high-throughput, quantitative TaqMan PCR assay (TM-PCR) for the general detection of huPoV. The TM-PCR detects huPoV DNA in a linear range from 107to 101copies per assay. In reproducibility runs, the inter- and intra-assay variabilities were ≤60 and ≤50%, respectively. The snPCR assay uses a set of four primers for the same region of the BK and JC viral genomes. In the first round of amplification, two general primers were used; in the second round, one of these general primers and two additional, BK- or JC-specific primers were used simultaneously to produce amplicons of different sizes specific for BK virus (246 bp) and JC virus (199 bp), respectively. We tested different urine dilutions in order to determine the inhibitory effects of urine on PCR amplification. Furthermore, we compared the use of native urine with DNA purified by different preparation procedures. Our results show, that a 1:10 dilution of the urine led to complete reduction of the amplification inhibition found with 6% of undiluted urine samples. In a clinical study including 600 urine specimens, our assay turned out to be fast, cheap, and reliable in both qualitative and quantitative aspects.