bluuebunny/crossref_metadata_2025_split huggingface dataset:
Sequence-Based Identification of
Mycobacterium
Species Using the MicroSeq 500 16S rDNA Bacterial Identification System
Authors: Patel Jean Baldus , Leonard Debra G. B. , Pan Xai , Musser James M. , Berman Richard E. , Nachamkin Irving 
Published: May 2021

ABSTRACT

We evaluated the MicroSeq 500 16S rDNA Bacterial Sequencing Kit (PE Applied Biosystems), a 500-bp sequence-based identification system, for its ability to identify clinical
Mycobacterium
isolates. The organism identity was determined by comparing the 16S rDNA sequence to the MicroSeq database, which consists primarily of type strain sequences. A total of 113 isolates (18 different species), previously recovered and identified by routine methods from two clinical laboratories, were analyzed by the MicroSeq method. Isolates with discordant results were analyzed by
hsp65
gene sequence analysis and in some cases repeat phenotypic identification, AccuProbe rRNA hybridization (Gen-Probe, Inc., San Diego, Calif.), or high-performance liquid chromatography of mycolic acids. For 93 (82%) isolates, the MicroSeq identity was concordant with the previously reported identity. For 18 (16%) isolates, the original identification was discordant with the MicroSeq identification. Of the 18 discrepant isolates, 7 (six unique sequences) were originally misidentified by phenotypic analysis or the AccuProbe assay but were correctly identified by the MicroSeq assay. Of the 18 discrepant isolates, 11 (seven unique sequences) were unusual species that were difficult to identify by phenotypic methods and, in all but one case, by molecular methods. The remaining two isolates (2%) failed definitive phenotypic identification, but the MicroSeq assay was able to definitively identify one of these isolates. The MicroSeq identification system is an accurate and rapid method for the identification of
Mycobacterium
spp.
VP7 and VP4 Genotyping of Human Group A Rotavirus in Buenos Aires, Argentina
Authors: Argüelles M. H. , Villegas G. A. , Castello A. , Abrami A. , Ghiringhelli P. D. , Semorile L. , Glikmann G. 
Published: May 2021

ABSTRACTSpecific and sensitive tests for the detection and typing of group A rotavirus strains are needed for a more comprehensive knowledge of the epidemiology of rotaviral infection. In this study 500 stool specimens taken from 1996 to 1998 from children with acute diarrhea in Buenos Aires were examined. Group A rotavirus was unequivocally demonstrated in 62% of the samples tested by enzyme-linked immunosorbent assay (ELISA) for detection of VP6 antigen, polyacrylamide gel electrophoresis of double-stranded RNA, and reverse transcription-PCR (RT-PCR) for amplification of the VP7:G (1,062 bp) and VP4:P (876 bp) genes. Only five positive specimens were found by RT-PCR but not by ELISA. G and P typing was carried out by nested amplification of variable sequences of the VP7 and the VP4 genes with six G- and five P-type-specific primers (multiplex PCR). Results obtained by this method showed the prevalence of the following G and P types: G1, 39%; G2, 43%; G4, 4%; P[8], 16%; P[4], 71%. Unexpectedly, the G-P type combination most frequently found was G2P[4] (43%) rather than G1P[8] (12%), which is the most commonly found worldwide. Unusual strains of the type G1P[4] accounted for 14% of the total, while mixed infections with more than one type were found in 10% of the samples. Detection of fecal rotavirus-specific immunoglobulin M (IgM) and IgA antibodies in consecutive samples of two patients taken at daily intervals demonstrated that high levels of IgM and IgA antibodies were detected on day 1 after the onset of disease and that the samples remained positive for about 10 days, after which virus shedding was no longer observed. Multiplex PCR offers a sensitive and specific alternative to determine the prevalence of group A rotavirus G and P types and to identify the emergence of uncommon strains, whereas detection of fecal IgM and IgA antibodies represents a useful supplement to virus detection for the diagnosis of current or recently acquired infections.
Diagnosis of Bubonic Plague by PCR in Madagascar under Field Conditions
Authors: Rahalison L. , Vololonirina E. , Ratsitorahina M. , Chanteau S. 
Published: May 2021

ABSTRACT

The diagnostic value of a PCR assay that amplifies a 501-bp fragment of the
Yersinia pestis caf1
gene has been determined in a reference laboratory with 218 bubo aspirates collected from patients with clinically suspected plague managed in a regional hospital in Madagascar. The culture of
Y. pestis
and the detection of the F1 antigen (Ag) by enzyme-linked immunosorbent assay (ELISA) were used as reference diagnostic methods. The sensitivity of PCR was 89% (57 of 64) for the
Y. pestis
-positive patients, and 80.7% (63 of 78) for the F1 Ag-positive patients. The specificity of PCR for the culture-, F1 Ag-, and antibody-negative patients (
n
= 105) was 100%. Because in Madagascar most patients with plague are managed and their clinical samples are collected in remote villages, the usefulness of PCR was evaluated for routine diagnostic use in the operational conditions of the control program. The sensitivity of PCR was 50% (25 of 50) relative to the results of culture and 35.2% (19 of 54) relative to the results of the F1 Ag immunocapture ELISA. The specificity of PCR under these conditions was 96%. In conclusion, the PCR method was found to be very specific but not as sensitive as culture or the F1 Ag detection method. The limitation in sensitivity may have been due to suboptimal field conditions and the small volumes of samples used for DNA extraction. This technique is not recommended as a routine diagnostic test for plague in Madagascar.
Genotype Profiles of Rotavirus Strains from Children in a Suburban Community in Guinea-Bissau, Western Africa
Authors: Fischer Thea Kølsen , Steinsland Hans , Mølbak Kåre , Ca Rui , Gentsch Jon R. , Valentiner-Branth Palle , Aaby Peter , Sommerfelt Halvor 
Published: May 2021

ABSTRACT
The P (VP4) and G (VP7) genotypes of 167 group A rotavirus strains obtained during the period 1996 to 1998 from 149 children living in a suburban community in Guinea-Bissau, western Africa, were determined by the reverse transcription-PCR technique. A total of nine combinations including five different P types and five different G types were identified. The globally common genotype pairs P[8], G1; P[4], G2; P[8], G3 and P[8], G4 were underrepresented in this study area. We found a substantial year-to-year variation in the occurrence of the genotype combinations. In 1996 and 1997, P[6], G2 was the most frequent, whereas P[8], G1 was more common in 1998. The unusual type P[9], G3 and a few mixed infections were detected. Sixteen percent of the rotavirus-positive samples were nontypeable.
Etiology of Genital Ulcer Disease in Dakar, Senegal, and Comparison of PCR and Serologic Assays for Detection of
Haemophilus ducreyi
Authors: Totten Patricia A. , Kuypers Jane M. , Chen Cheng-Yen , Alfa Michelle J. , Parsons Linda M. , Dutro Susan M. , Morse Stephen A. , Kiviat Nancy B. 
Published: May 2021

ABSTRACT

We used PCR assays to determine the etiology of genital ulcers in patients presenting to a sexually transmitted disease clinic in Dakar, Senegal, and evaluated the ability of two PCR tests (
groEL
and
recD
) and two serological tests (adsorption enzyme immunoassay [EIA] and lipooligosaccharide [LOS] EIA) to detect current
Haemophilus ducreyi
infection. We found that in this population,
H. ducreyi
,
T. pallidum
, and herpes simplex virus HSV DNA were detected in 56, 15, and 13% of 39 genital ulcer specimens, respectively, and
H. ducreyi
DNA was detected in 60% (3 of 5) of samples from ulcerated bubos. Among 40 consecutive patients with genital ulcer disease and with sufficient sample for both PCR assays, the
recD
and
groEL H. ducreyi
PCR assays were 83% concordant, with the
recD
PCR assay detecting six (15%) additional positive specimens and the
groEL
assay detecting one (3%) additional positive specimen. Compared to PCR, the adsorption EIA and LOS EIA tests had sensitivities of 71 and 59% and specificities of 57 and 90%, respectively, for the diagnosis of current
H. ducreyi
infection. While these differences in specificity could be due either to previous infection with
H. ducreyi
or to the detection of cross-reacting antibodies, only 6% of patients from a nearby family planning clinic gave a positive reaction in both the adsorption EIA and LOS EIA assays, indicating that cross-reacting antibodies are not prevalent among clinic attendees in this city. Our studies indicate that the adsorption EIA detects both current and past infection, while the LOS EIA assay is more specific for current infection with
H. ducreyi
in this population.
Prevalence of Toxin Types and Colonization Factors in Enterotoxigenic
Escherichia coli
Isolated during a 2-Year Period from Diarrheal Patients in Bangladesh
Authors: Qadri Firdausi , Das Swadesh Kumar , Faruque A. S. G. , Fuchs George J. , Albert M. John , Sack R. Bradley , Svennerholm Ann-Mari 
Published: May 2021

ABSTRACT

The prevalence of toxin types and colonization factors (CFs) of enterotoxigenic
Escherichia coli
(ETEC) was prospectively studied with fresh samples (
n
= 4,662) obtained from a 2% routine surveillance of diarrheal stool samples over 2 years, from September 1996 to August 1998. Stool samples were tested by enzyme-linked immunoassay techniques and with specific monoclonal antibodies for the toxins and CFs. The prevalence of ETEC was 14% (
n
= 662), with over 70% of the strains isolated from children 0 to 5 years of age, of whom 93% were in the 0- to 3-year-old age range. Of the total ETEC isolates, 49.4% were positive for the heat-stable toxin (ST), 25.4% were positive for the heat-labile toxin (LT) only, and 25.2% were positive for both LT and ST. The rate of ETEC isolation peaked in the hot summer months of May to September and decreased in winter. About 56% of the samples were positive for 1 or more of the 12 CFs that were screened for. The coli surface antigens CS4, CS5, and/or CS6 of the colonization factor antigen (CFA)/IV complex were most prevalent (incidence, 31%), followed by CFA/I (23.5%) and coli surface antigens CS1, CS2, and CS3 of CFA/II (21%). In addition, other CFs detected in decreasing order were CS7 (8%), CS14 (PCFO166) (7%), CS12 (PCFO159) (4%), CS17 (3%), and CS8 (CFA/III) (2.7%). The ST- or LT- and ST-positive ETEC isolates expressed the CFs known to be the most prevalent (i.e., CFA/I, CFA/II, and CFA/IV), while the strains positive for LT only did not. Among children who were infected with ETEC as the single pathogen, a trend of relatively more severe disease in children infected with ST-positive (
P
< 0.001) or LT- and ST-positive (
P
< 0.001) ETEC isolates compared to the severity of the disease in children infected with LT only-positive ETEC isolates was seen. This study supports the fact that ETEC is still a major cause of childhood diarrhea in Bangladesh, especially in children up to 3 years of age, and that measures to prevent such infections are needed in developing countries.
Importance of Inoculum Size and Sampling Effect in Rapid Antigen Detection for Diagnosis of
Streptococcus pyogenes
Pharyngitis
Authors: Kurtz Bradley , Kurtz Michael , Roe Martha , Todd James 
Published: May 2021

ABSTRACT

Current recommendations suggest that negative rapid
Streptococcus pyogenes
antigen tests be backed up with a culture, reflecting evidence that culture may have a higher sensitivity and also that testing of a second swab may yield a different (i.e., a positive) result because of variation in sample size or distribution. If the latter is common, the sensitivities of current antigen detection tests might be improved by simply increasing the amount of sample tested. The present study assessed the effect of antigen testing of two swabs extracted together compared to independent testing of each swab extracted separately for children with clinical pharyngitis.
S. pyogenes
grew from one or both swabs for 198 (37%) of 537 children. The combined culture was significantly (
P
< 0.05) more sensitive than culture of either swab alone. Compared to combined culture, antigen testing of two swabs extracted and tested together was significantly more sensitive than two single swab extractions (94.1 versus 80%;
P
= 0.03); however, the specificity was decreased (81.5 versus 89.8 to 92.7%;
P
< 0.05). This study suggests that sample size and/or uneven sample distribution may have influenced the apparent sensitivities of prior studies that compared antigen tests to a single plate culture. A strategy, such as the one used in the present study, that increases the sample size available for antigen testing (i.e., extraction of samples from both swabs) may improve detection rates to a level that will better approximate true disease status and obviate the need for backup cultures if specificity can be improved.
Prevalence of
Campylobacter
,
Arcobacter
,
Helicobacter
, and
Sutterella
spp. in Human Fecal Samples as Estimated by a Reevaluation of Isolation Methods for Campylobacters
Authors: Engberg Jørgen , On Stephen L. W. , Harrington Clare S. , Gerner-Smidt Peter 
Published: May 2021

ABSTRACT

The aims of this study were to investigate the prevalence of campylobacteria including
Campylobacter jejuni
subsp.
jejuni
(
C. jejuni
) and
Campylobacter coli
in human clinical samples and in samples from healthy individuals and to reevaluate the efficacies of conventional selective methods for isolation of
Campylobacter
spp. Two charcoal-based selective media, modified charcoal cefoperazone deoxycholate agar (mCCDA) and cefoperazone-amphotericin-teicoplanin (CAT) agar, were compared with Skirrow's blood-based medium and with a filter method (filter) applied to a yeast-enriched blood agar. A total of 1,376 specimens were tested on all four media, and the percentages of thermophilic
Campylobacter
-positive specimens isolated on Skirrow's medium, filters, CAT agar, and mCCDA were 82, 83, 85, and 95%, respectively. When additional samples were processed with the three selective media, mCCDA recovered significantly more thermophilic
Campylobacter
spp. than Skirrow's medium (
P
= 0.0034). No significant difference between Skirrow's medium and CAT agar was observed in this study. Another six taxa were identified, namely,
Campylobacter concisus
,
Campylobacter curvus
-like bacteria,
Arcobacter butzleri
,
Arcobacter cryaerophilus
,
Helicobacter cinaedi
, and
Sutterella wadsworthensis
. Most of these strains were isolated after 5 to 6 days of incubation by use of the filter technique. This paper provides evidence for the existence of
S. wadsworthensis
in human feces from clinical cases of gastrointestinal disorders and in feces from a healthy individual. Furthermore,
C. concisus
was isolated from a large number of diarrheal cases, particularly those at the extremes of age, but was additionally isolated from the feces of healthy people. Further investigations to establish the role of
C. concisus
and
S. wadsworthensis
in enteric disease is needed. We conclude that a range of campylobacteria may cause infections in Denmark.
Molecular Methods for the Epidemiological Typing of
Salmonella enterica
Serotype Typhi from Hong Kong and Vietnam
Authors: Ling J. M. , Lo N. W. S. , Ho Y. M. , Kam K. M. , Hoa Nguyen Thi Tuyet , Phi Le Thi , Cheng A. F. 
Published: May 2021

ABSTRACT

A total of 217 and 73 strains of
Salmonella enterica
serotype Typhi isolated from 1985 to 1997 in Hong Kong and in 2 months of 1989 and 1990 in Vietnam, respectively, were studied. These isolates were typed by plasmid profile analysis, plasmid fingerprinting, ribotyping with
Pst
I, and total DNA fingerprinting with
Nar
I. There appeared to be no major outbreak of typhoid fever in Hong Kong during the study period since there was considerable heterogeneity among the isolates. Isolates from Hong Kong were different from those from Vietnam. Thirty-seven percent of Vietnamese isolates belonged to two predominant clones, with the rest being heterogeneous in nature. Total DNA fingerprinting supplemented with ribotyping could be a reliable and rapid method for epidemiological typing of
S. enterica
serotype Typhi.
Differentiation of Phylogenetically Related Slowly Growing Mycobacteria by Their
gyrB
Sequences
Authors: Kasai Hiroaki , Ezaki Takayuki , Harayama Shigeaki 
Published: May 2021

ABSTRACT

The conventional methods for identifying mycobacterial species are based on their phenotypic characterization. Since some problematic species are slow growers, their taxonomy takes several weeks or months to identify. The ribosomal DNA (rDNA) sequence-based identification strategy has been adopted to solve this problem. More recently, the
gyrB
sequences have been shown to be useful phylogenetic markers for the identification of species. We determined the
gyrB
sequences of 43 slowly growing strains belonging to 15 species in the genus
Mycobacterium
. The frequencies of base substitutions in the
gyrB
sequences were comparable to those in the 16S-23S rDNA internal transcribed spacer (ITS) sequences. The ITS sequences of four species belonging to the
M. tuberculosis
complex (
M. tuberculosis
,
M. bovis
,
M. africanum
, and
M. microti
) were 100% identical, while four synonymous substitutions were found in the
gyrB
sequences of these strains. Based on the differences found in the
gyrB
sequences, we developed PCR and PCR-restriction fragment length polymorphism methods to discriminate these species.
Evaluation of Whole-Cell and OspC Enzyme-Linked Immunosorbent Assays for Discrimination of Early Lyme Borreliosis from OspA Vaccination
Authors: Wieneke Chad A. , Lovrich Steven D. , Callister Steven M. , Jobe Dean A. , Marks Jennifer A. , Schell Ronald F. 
Published: May 2021

ABSTRACT

A recombinant Lyme borreliosis vaccine consisting of outer surface protein A (OspA) is commercially available for vaccination of humans against infection with
Borrelia burgdorferi
. Vaccination with OspA induces an antibody response that makes serologic interpretation of infection with
B. burgdorferi
difficult, especially by screening tests based on whole-cell preparations of
B. burgdorferi
. We show that an enzyme-linked immunosorbent assay with
B. burgdorferi
sensu stricto 50772, which lacks the plasmid encoding OspA and OspB, or a full-length recombinant OspC protein can identify patients infected with
B. burgdorferi
. We found that 69 and 65% of serum samples from patients with case-defined early Lyme borreliosis had anti-
B. burgdorferi
sensu stricto 50772 and anti-OspC reactivities, respectively. In addition, little or no reactivity was detected with sera obtained from individuals vaccinated with OspA. Unfortunately, 51 and 33% of sera from healthy patients and sera from patients with other illnesses were also reactive against
B. burgdorferi
sensu stricto 50772 and OspC, respectively. Although these assays can discriminate
B. burgdorferi
infection from vaccination with OspA, their lack of specificity highlights the necessity for confirming equivocal or positive reactivities with more specific serodiagnostic tests.
Prevalence of
Escherichia coli
O157:H7 from Cull Dairy Cows in New York State and Comparison of Culture Methods Used during Preharvest Food Safety Investigations
Authors: McDonough Patrick L. , Rossiter Christine A. , Rebhun Robert B. , Stehman Susan M. , Lein Donald H. , Shin Sang J. 
Published: May 2021

ABSTRACT

A number of protocols for the cultural detection of
Escherichia coli
O157:H7 in clinical fecal specimens have been proposed. In the present study direct plating of cattle feces was compared to three different broth enrichment protocols, i.e., a protocol with modified
E. coli
broth with novobiocin, a protocol with Trypticase soy broth with cefixime and vancomycin, and a protocol with Gram-Negative Broth with novobiocin, for their relative abilities to detect
E. coli
O157:H7 in feces. In all enrichment protocols, dilutions of the enrichment broths onto 150-mm sorbitol-MacConkey agar plates to which cefixime and tellurite were added were used along with reading of agar plates at both 24 and 48 h. Fecal samples came from a preharvest food safety project in which feces from New York cull dairy cattle from a northeastern packing plant along with experimentally inoculated adult dairy cow feces were tested. The performances of the broth enrichments were comparable to each other, but the broth enrichments were superior to direct plating in their ability to detect
E. coli
O157:H7. Regardless of the culture protocol used, recovery of
E. coli
O157:H7 is more likely from fresh fecal specimens than from frozen samples. An overall prevalence of
E. coli
O157:H7 fecal shedding by New York cull dairy cattle of 1.3% was found in specimens just before processing at the packing plant.
Stability of Plasma Human Immunodeficiency Virus Load in VACUTAINER PPT Plasma Preparation Tubes during Overnight Shipment
Authors: Holodniy Mark , Rainen Lynne , Herman Steve , Yen-Lieberman Belinda 
Published: May 2021

ABSTRACT
VACUTAINER PPT plasma preparation tubes were evaluated to determine the effects of various handling and shipping conditions on plasma human immunodeficiency virus (HIV) load determinations. Plasmas obtained from PPT tubes stored and shipped under nine different conditions were compared to conventional EDTA tube plasmas stored at −70°C within 2 h after phlebotomy. Compared to viral loads in frozen EDTA plasma, those in PPT tube plasma that was frozen immediately and either separated or shipped in situ were not significantly different. Viral loads in PPT tube plasma after storage for 6 h at either room temperature or 4°C, followed by shipment at ambient temperature or on wet or dry ice, were not significantly different from baseline viral loads in EDTA or PPT plasma. The results of this study indicate that the HIV load in PPT tube plasma is equivalent to that in standard EDTA plasma. Plasma viral load is not affected by storage or shipment temperature when plasma is collected in PPT tubes. Furthermore, plasmas can be shipped in spun PPT tubes, and the tubes provide a safer and more convenient method for sample collection and transport than regular EDTA tubes.
Comparison ofPasteurellaspp. Simultaneously Isolated from Nasal and Transtracheal Swabs from Cattle with Clinical Signs of Bovine Respiratory Disease
Authors: DeRosa D. C. , Mechor G. D. , Staats J. J. , Chengappa M. M. , Shryock T. R. 
Published: May 2021

ABSTRACTTwenty-four matched pairs of isolates ofPasteurella haemolyticaand three matched pairs of isolates ofPasteurella multocidawere isolated by using a nasal swab and a transtracheal swab from individual calves with clinical signs of bovine respiratory disease. The identity of each matched pair was confirmed biochemically and serologically. The similarity of the isolates obtained from a nasal swab and from a transtracheal swab was compared by using ribotyping and antibiotic susceptibility analyses. Although the calves were sampled only once with a nasal and a transtracheal swab, when both samples were bacteriologically positive the nasal swab identified the same bacterial species as the transtracheal swab 96% of the time. The nasal swab isolate was genetically identical to the transtracheal isolate in 70% of the matched pairs. Six different ribotypes were observed for theP. haemolyticaisolates, while only one ribotype was observed for the limited number ofP. multocidaisolates. Of the sixP. haemolyticaribotypes, two ribotypes predominated. All the paired isolates displayed similar susceptibility to ceftiofur, erythromycin, tilmicosin, trimethoprim-sulfamethoxazole, and florfenicol, with some minor variations for ampicillin and spectinomycin. These results suggest that a nasal swab culture can be predictive of the bacterial pathogen within the lung when the isolates are from an acutely ill animal and can be used to determine antibiotic susceptibility.