bluuebunny/crossref_metadata_2025_split huggingface dataset:
Identification of Enteroviruses by Using Monoclonal Antibodies against a Putative Common Epitope
Authors: Shin Soo-Youn , Kim Ki-Soon , Lee Yoon-Sung , Chung Yoon-Seok , Park Kwi-Sung , Cheon Doo-Sung , Na Byoung-Kuk , Kang Yoonsung , Cheong Hyang-Min , Moon Youngjoon , Choi Jee-Hye , Cho Hang-Eui , Min Na-Young , Son Jin-Sook , Park Young-Hoon , Jee Youngmee , Yoon Jae-Deuk , Song Chul-Yong , Lee Kwang-Ho 
Published: July 2003

ABSTRACT
A common epitope region of enteroviruses was identified by sequence-independent single-primer amplification (SISPA), followed by immunoscreening of 11 cDNA libraries from two Korean enterovirus isolates (echoviruses 7 and 30) and a coxsackievirus B3 (ATCC-VR 30). The putative common epitope region was localized in the N terminus of VP1 when the displayed recombinant proteins from the phages were chased by the convalescent-phase sera. The genomic region encoding the common epitope region was amplified and then expressed by using the vector pGEX-5X-1. The antigenicity of the expressed recombinant protein was identified by Western blotting with guinea pig antisera for six different serotypes of enteroviruses. After successive immunization of mice with the recombinant common epitope protein, splenocytes were extracted and hybridized with P3X63-Ag8-653 cells. A total of 24 hybridomas that produced monoclonal antibodies (MAbs) against the putative common epitope of enteroviruses were selected. Four of these were immunoglobulin G1 isotypes with a kappa light chain. These MAbs recognized 15 Korean endemic serotypes and prototypes of enteroviruses in an indirect immunofluorescence assay. These results suggest that the expressed protein might be a useful antigen for producing group common antibodies and that the use of the MAbs against the putative common epitope of enteroviruses might be a valuable diagnostic tool for rapidly identifying a broad range of enteroviruses.
Optimization of Computer Software Settings Improves Accuracy of Pulsed-Field Gel Electrophoresis Macrorestriction Fragment Pattern Analysis
Authors: Duck William M. , Steward Christine D. , Banerjee Shailen N. , McGowan, John E. , Tenover Fred C. 
Published: July 2003

ABSTRACT

Computer-assisted analysis of pulsed-field gel electrophoresis (PFGE) libraries can facilitate comparisons of fragment patterns present on multiple gels. We evaluated the ability of the Advanced Analysis (version 4.01) and Database (version 1.12) modules of the Phoretix gel analysis software package (Nonlinear USA, Inc., Durham, N.C.) to accurately match DNA fragment patterns. Two gels containing 38 lanes of
Sma
I-digested
Enterococcus faecalis
OG1RF DNA were analyzed to assess the impact of (i) varying the lane position of the standards, (ii) using gel plugs made at different times, and (iii) normalizing the fragment patterns by using molecular weight (MW) algorithms versus retardation factor (
R

f

) algorithms. Two sets of PFGE libraries (one containing
Sma
I restriction patterns from 62
Enterococcus faecium
isolates and the other containing
Sma
I restriction patterns of 89
Staphylococcus aureus
isolates) were analyzed to assess the impact of varying the matching tolerance algorithm (designated as the vector box setting [VBS]) in the Phoretix software. Varying the lane position of standards on a gel and using gel plugs made on different days resulted in different VBSs, although it was not possible to judge whether those differences were statistically significant. Normalization of
E. faecalis
OG1RF fragment patterns by
R

f

and MW methodology yielded no statistically significant differences in variability between the same fragment on different lanes. Suboptimal VBSs decreased the specificity with which related isolates were grouped together in dendrograms. The optimal VBS for analysis of PFGE fragment patterns from
E. faecalis
isolates differed from that for
S. aureus
isolates and sometimes was not that recommended by the manufacturer. Thus, computer-assisted analysis of PFGE patterns seemed to compensate for the intra- and intergel variation evaluated in the present study, and optimizing the software for the species to be tested was a critical preliminary step before further PFGE library analysis.
High Prevalence of Human Metapneumovirus Infection in Young Children and Genetic Heterogeneity of the Viral Isolates
Authors: Viazov S. , Ratjen F. , Scheidhauer R. , Fiedler M. , Roggendorf M. 
Published: July 2003

ABSTRACT

RNA of the newly identified human metapneumovirus (HMPV) was detected in nasopharyngeal aspirates of 11 of 63 (17.5%) young children with respiratory tract disease. Markers of infection caused by another member of the
Pneumovirinae
subfamily of the family
Paramyxoviridae
, respiratory syncytial virus (RSV), were identified in 15 of these patients (23.8%). Three patients were simultaneously infected with HMPV and RSV. Studies of the clinical characteristics of HMPV-infected children did not reveal any difference between HMPV-infected patients and a control population of RSV-infected patients with regard to disease severity, but the duration of symptoms was significantly shorter for HMPV-infected patients. Phylogenetic analysis of the amplified viral genome fragments confirmed the existence and simultaneous circulation within one epidemic season of HMPV isolates belonging to two genetic lineages.
Quantification of Hepatitis C Virus (HCV) in Liver Specimens and Sera from Patients with Human Immunodeficiency Virus Coinfection by Using the Versant HCV RNA 3.0 (Branched DNA-Based) DNA Assay
Authors: Tedeschi Rosamaria , Pivetta Eliana , Zanussi Stefania , Bidoli Ettore , Ros Mirna , di Gennaro Giampiero , Nasti Guglielmo , De Paoli Paolo 
Published: July 2003

ABSTRACT

The new generation assay Versant HCV RNA 3.0v (Bayer Diagnostics) was evaluated to quantify hepatitis C virus (HCV) RNA levels in liver biopsy specimens from patients with HCV and human immunodeficiency virus (HIV) coinfection. A total of 25 liver biopsies and sera collected at the time of liver biopsy were used. The efficiency of HCV RNA recovery from spiked samples was between 38.6 and 50.7%, and reproducible measurements of viral load were observed (the intra- and interrun coefficients of variation were 0.5 to 13% and 3.5 to 24.7%, respectively), with good specificity and sensitivity. Linearity was evaluated in the range of 96,154 to 769 IU/μg by using a serially diluted high-titer sample. Coinfected patients had high HCV RNA viral loads in serum and liver (498,471 IU/ml and 231,495 IU/μg, respectively), and both levels were correlated (
r
= 0.63;
P
< 0.01). The amount of hepatic HCV RNA was significantly higher among patients with genotype 1 than among patients with genotype 3 (
P
< 0.01). The virological end-of-treatment response in the serum was associated with a lower pretreatment intrahepatic HCV viral load (
P
= 0.03). The new version of b-DNA is a sensitive, specific, and reproducible method for quantitating HCV RNA in the liver. Given its positive analytical performance, the assay will be used to evaluate the HCV RNA levels in the serum and liver during follow-up of patients treated with an anti-HCV therapeutic regimen.
Major Outbreak of Toxic Shock-Like Syndrome Caused byStreptococcus mitis
Authors: Lu Hong-Zhou , Weng Xin-Hua , Zhu Bai , Li Haijing , Yin You-Kuan , Zhang Yong-Xin , Haas David W. , Tang Yi-Wei 
Published: July 2003

ABSTRACTSevere illness caused by viridans streptococci rarely occurs in immunocompetent hosts. Between December 1990 and May 1991, thousands of patients in the YangZi River Delta area of Jiangsu Province, China, suffered from scarlet fever-like pharyngitis. Fewer cases occurred in subsequent years with the same seasonality. Approximately half of the cases developed complications characteristic of streptococcal toxic shock-like syndrome (TSLS). Throat cultures yielded predominant growth of alpha-hemolytic streptococci. All cases admitted to Haian People's Hospital were investigated. Clinical specimens were collected, medical records were reviewed, and bacterial isolates were identified phenotypically and analyzed by 16S rRNA gene sequencing and pulsed-field gel electrophoresis (PFGE). Proteins were purified from culture supernatants by extraction, ammonium sulfate precipitation, and fast-protein liquid chromatography. Biological activities of protein components were determined by subcutaneous inoculation into rabbits. A total of 178 cases of non-beta-hemolytic streptococcal scarlet fever-like pharyngitis were studied. In 88 (79.3%) of 111 patients, oropharyngeal swab cultures grew morphologically identical alpha-hemolytic streptococci. A protein in culture supernatants was pyrogenic in rabbits, was mitogenic for splenocytes, and enhanced rabbit susceptibility to endotoxin challenge. The N-terminal amino acid sequence of this 34-kDa protein showed no homology with knownStreptococcuspyrogenic exotoxins. The organism was identified asStreptococcus mitisbased on biochemical and 16S rRNA sequence analyses. Representative outbreak isolates from 1990 to 1995 displayed identical PFGE patterns. This TSLS outbreak in southeastern China was caused by a toxigenic clone ofS. mitis. An apparently novel toxin may explain the unusual virulence of this organism.
Field Evaluation of the
gag
-Based Heteroduplex Mobility Assay for Genetic Subtyping of Circulating Recombinant Forms of Human Immunodeficiency Virus Type 1 in Abidjan, Côte d'Ivoire
Authors: Sawadogo Souleymane , Adjé-Touré Christiane , Bilé Celestin E. , Ekpini Rene E. A. , Chorba Terence , Nkengasong John N. 
Published: July 2003

ABSTRACT

The
gag
-based heteroduplex mobility assay (
gag
-HMA) was evaluated for its ease and reliability in subtyping circulating recombinant forms (CRFs) of human immunodeficiency virus type 1 (HIV-1) in Côte d'Ivoire. One hundred thirty-two plasma samples were analyzed blindly for HIV-1 subtypes by sequencing the
pol
gene and by
gag
-HMA. DNA sequencing was used as the “gold standard.” Of the 132 samples sequenced, 108 (82%) were CRF02_AG, 14 (11%) were pure subtype A, 5 (4%) were subtype G, 3 (2%) were subtype D, 1 was CRF01_AE, and 1 was subtype H. The
gag
-HMA correctly classified 126 (95.5%) of the samples. Of the 108 samples that were classified as CRF02_AG by DNA sequencing, 107 (99%) were correctly identified by
gag
-HMA, resulting in a positive predictive value of 96.4%. The
gag
-HMA seems to be a valuable tool for understanding the molecular epidemiology of HIV-1 CRF02_AG in Côte d'Ivoire and West Africa, which could be important for developing and evaluating AIDS vaccines, although DNA sequencing remains necessary for accurate molecular epidemiology.
Comparison of Two Commercial Enzyme-Linked Immunosorbent Assays with an Immunofluorescence Assay for Detection ofLegionella pneumophilaTypes 1 to 6
Authors: Malan Annette K. , Martins Thomas B. , Jaskowski Troy D. , Hill Harry R. , Litwin Christine M. 
Published: July 2003

ABSTRACTMembers of the genusLegionellaare characterized as gram-negative, motile, freshwater-dwelling bacteria that were responsible for a pneumonia outbreak among American Legion members in 1976. Because clinicians routinely order serologic testing forLegionella pneumophilaserogroups 1 to 6 as a screen for possibleL. pneumophilainfections, we evaluated the Wampole LaboratoriesL. pneumophilatype 1 to 6 immunoglobulin G (IgG) and IgM combined enzyme-linked immunosorbent assay (ELISA) and the Zeus ScientificL. pneumophilatype 1 to 6 IgG-IgM-IgA multispecific combined ELISA systems and compared them to an IgG-specific immunofluorescence assay (IFA) forL. pneumophilaserogroups 1 to 6. The Centers for Disease Control and Prevention recommends that the positive titer cutoff for an IFA be 1:256. Regardless of where the positive IFA cutoff titer is placed, however, the sensitivity of both commercial assays was below what would be acceptable for a screening assay. With a 1:256 IFA titer as the positive cutoff, the agreement, sensitivity, and specificity of the Wampole ELISA were 74.6, 21.4, and 98.4%, respectively. The agreement, sensitivity, and specificity of the Zeus ELISA were 72.6, 10.5, and 100.0%, respectively. We recommend that any laboratories attempting to replace an IFA type 1 to 6 screen with an alternative ELISA carefully investigate the sensitivity of the replacement assay.
Invasive Disease Due to Nontypeable
Haemophilus influenzae
among Children in Arkansas
Authors: O'Neill Joshua M. , St. Geme Joseph W. , Cutter David , Adderson Elisabeth E. , Anyanwu Juliana , Jacobs Richard F. , Schutze Gordon E. 
Published: July 2003

ABSTRACT

In this study, we reviewed cases of invasive disease due to nontypeable
Haemophilus influenzae
among children hospitalized at Arkansas Children's Hospital from 1993 to 2001. A total of 28 cases were examined, including 21 associated with bacteremia and 4 associated with meningitis. Of the patients examined, 86% were ≤4 years of age, and 68% had underlying medical conditions. Characterization of the bacterial isolates by multilocus sequence type genotyping revealed significant overall genetic diversity, similar to the diversity in the general population structure for nontypeable
H. influenzae
. However, four separate pairs of isolates were closely related genetically, a relationship confirmed by pulsed-field gel electrophoresis and Southern hybridization studies using probes for the major
H. influenzae
adhesin genes. These results suggest that selected strains of nontypeable
H. influenzae
may have more invasive potential, especially in young children and patients with underlying medical conditions. At this point, the specific factors that contribute to enhanced virulence remain unclear.
Mycobacterium bovis
subsp.
caprae
Caused One-Third of Human
M. bovis
-Associated Tuberculosis Cases Reported in Germany between 1999 and 2001
Authors: Kubica Tanja , Rüsch-Gerdes Sabine , Niemann Stefan 
Published: July 2003

ABSTRACT

The prevalence of the
Mycobacterium bovis
subsp.
caprae
and
M. bovis
subsp.
bovis
among German tuberculosis cases caused by the bovine tubercle bacillus from 1999 to 2001 was determined. Isolates from 166 patients living in Germany and 10 animals were analyzed by conventional laboratory procedures, spoligotyping, and partly by PCR-restriction fragment length polymorphism analysis of the
gyrB
gene. By spoligotyping, 55 of 176 isolates (31%) could be identified as
M. bovis
subsp.
caprae
, and 121 (69%) were confirmed as
M. bovis
subsp.
bovis.
In general, a low variability of spoligotypes with 59 distinct patterns and a cluster rate of 77% (136 isolates/19 clusters) was determined. About half of all isolates were grouped in the three main clusters with 29, 30, and 35 isolates, respectively. Differences in age and gender between the patient groups infected with
M. bovis
subsp.
bovis
and
M. bovis
subsp.
caprae
did not reach statistical significance. However, marked differences in the geographical prevalence of
M. bovis
subsp.
caprae
were observed, ranging from fewer than 10% of all
M. bovis
isolates in the north up to more than 80% of isolates in the south of Germany. In conclusion,
M. bovis
subsp.
caprae
accounts for a high ratio of human
M. bovis
-associated tuberculosis cases in Germany and was more frequently found in the southern part.
Molecular Epidemiology of
Shigella
in aTaiwan Township during 1996 to2000
Authors: Chen Jiann-Hwa , Chiou Chien-Shun , Chen Pei-Chun , Liao Tung-Liang , Liao Tsai-Ling , Li Jen-Ming , Hsu Wen-Bin 
Published: July 2003

ABSTRACT

A
previously identified
Shigella flexneri
serotype 2a strain was
responsible for an outbreak of shigellosis in a Taiwan township in
August 1996. In order to find the relationship between this outbreak
strain and subsequent
Shigella
infections in the
area, 59, 47, 35, and 20
Shigella
isolates recovered in 1997,
1998, 1999, and 2000, respectively, were collected and typed by
serological and pulsed-field gel electrophoresis (PFGE) techniques. Of
these 161 isolates, 139 isolates were
S. flexneri
serotype 2a,
and one-third of them (47 isolates) exhibited the outbreak pattern. The
remaining 92
S. flexneri
serotype 2a isolates displayed 49
different
Not
I-PFGE patterns. Forty-five patterns were closely
related to the outbreak pattern, with deletions of three specific
Not
I fragments occurring with high frequency. While the
outbreak strain remained the main cause of shigellosis after the
outbreak, the continuous emergence of closely related though poorly
transmissible strains from the outbreak strain contributed to the
observed annual decrease of shigellosis in the
area.
Reverse Transcription-PCR Assays for Detection of Bovine Enteric Caliciviruses (BEC) and Analysis of the Genetic Relationships among BEC and Human Caliciviruses
Authors: Smiley J. R. , Hoet A. E. , Tråvén M. , Tsunemitsu H. , Saif L. J. 
Published: July 2003

ABSTRACT
Two genetically distinct bovine enteric caliciviruses (BECs) have been identified: the norovirus (NLV) Jena and Newbury Agent-2 (NA-2) BECs, which are genetically related to human noroviruses, and the Nebraska (NB) BECs, which is related to sapoviruses and lagoviruses but may also represent a new calicivirus genus. The prevalence of these two BEC genotypes in cattle is unknown. Although reverse transcription-PCR (RT-PCR) primers for human NLV recognize NLV-BECs, the genetic relationships between NLV from humans and the NLV-BECs commonly circulating in cattle is undefined. In the present study, veal calf fecal samples were assayed for enteric caliciviruses by using six RT-PCR primer sets designed for the detection of human NLVs or BECs. Caliciviruses genetically related to the NLV-BEC Jena and NA-2 strains or to the recently characterized NB BEC strain were identified in three of four and four of four sampled veal herds, respectively. Extended 3′-terminal genome sequences of two NLV-BECs, designated CV95-OH and CV186-OH, encoding the RNA-dependent RNA polymerase (RdRp; open reading frame 1 [ORF-1]), VP1 (ORF-2), and VP2 (ORF-3) genes were determined. Phylogenetic and sequence identity analyses of each genome region demonstrated these viruses to be most closely related to the NLV-BEC Jena and NA-2 strains. In initial testing, the human P289-P290 (P289/290) primer set was found to be the most sensitive for calicivirus detection. However, its failure to identify all positive fecal pools (as determined by other assays) led us to design two new primer sets, CBECU-F/R and NBU-F/R, for the sensitive and specific detection of NLV-BEC (NLV-BEC Jena and NA-2) and BEC-NB-like viruses, respectively. The RT-PCR assays with the new primers were compared against other primer sets, including P289/290. Composite results of the tests completed by using the new assays identified 72% (54 of 75) of veal calf fecal samples as positive, with 21 of 21 sequenced reaction products specific for the target RdRp gene. The same design strategy used for the new BEC assays may also be applicable to the design of similar assays for the detection of human caliciviruses (HuCVs). Our data support the genetic relationship between NLV-BECs and NLV-HuCVs but with the NLV-BECs comprising two clusters within a third NLV genogroup.
Characterization of Serotype G9 Rotavirus Strains Isolated in the United States and India from 1993 to 2001
Authors: Laird A. R. , Gentsch J. R. , Nakagomi T. , Nakagomi O. , Glass R. I. 
Published: July 2003

ABSTRACT
The emergence of rotavirus serotype G9 as a possible fifth globally common serotype in the last decade, together with its increasing detection in association with various genome constellations, raises questions about the origins and epidemiological importance of recent G9 isolates. We examined a collection of 40 G9 strains isolated in the United States from 1996 to 2001 and in India since 1993 to determine their VP7 gene sequences, P types, E types, subgroup specificities, and RNA-RNA hybridization profiles. With the exception of two U.S. strains, all of the study strains shared high VP7 gene sequence homology (<2.5% sequence divergence on both the nucleotide and amino acid levels) and were more closely related to other recent isolates than to the first G9 strains isolated in the 1980s. The VP7 gene sequence and RNA-RNA hybridization profiles of the long-E-type strains showed greater variation than the short-E-type strains, suggesting that the latter strains are the result of a relatively recent reassortment event of the G9 VP7 gene into a short-E-type lineage. No evidence for reassortment of genes other than VP4 and VP7 between major human rotavirus genogroups was observed. Except for Om46 and Om67, which formed a distinct clade, phylogenetic analysis showed that most of the study strains grouped together, with some subgroups forming according to genetic constellation, geographic location, and date of isolation. The high potential of G9 strains to generate different P and G serotype combinations through reassortment suggests that it will be important to determine if current vaccines provide heterotypic protection against these strains and underscores the need for continued surveillance for G9 and other unusual or emerging rotavirus strains.
Identification ofcagATyrosine Phosphorylation DNA Motifs inHelicobacter pyloriIsolates from Peptic Ulcer Patients by Novel PCR-Restriction Fragment Length Polymorphism and Real-Time Fluorescence PCR Assays
Authors: Owen Robert J. , Sharp Sally I. , Chisholm Stephanie A. , Rijpkema Sjoerd 
Published: July 2003

ABSTRACTCag pathogenicity island-containingHelicobacter pylori(type I) induces signal transduction pathways resulting in tyrosine phosphorylation of proteins adjacent to the site of bacterial adhesion on host gastric epithelial cells. Conventional block PCR-restriction fragment length polymorphism (RFLP) and real-time LightCycler (LC) PCR hybridization assays, validated by direct sequencing, were designed to test for the presence of three nucleotide sequences corresponding to tyrosine phosphorylation motifs (TPMs) A, B, and C in 84 isolates ofH. pyloritype I from patients in England. Overall, the PCR assays demonstrated that one or more TPMs were present in 62 strains (75%). Motif A was common (71% of strains), whereas motifs B and C were rarer (8% of strains). Strains lacking a TPM were typically vacuolating cytotoxin genotypevacAm2. Motif A was widely distributed in relation to disease severity and was more commonly (but not significantly [P= 0.071]) associated with gastric ulcer than with duodenal ulcer (86 versus 56%). The LC hybridization assay provided a rapid means of detecting all three motifs, but RFLP analysis was more specific for TPM-A. TPMs provide novel additional strain markers for definingcagAvariation, including identification of RFLP types within TPM-A. The presence of a particular TPM was not of direct diagnostic value, either singly or in combination, but the higher proportion of TPM-A strains in gastric ulcer patients merits further investigation.