bluuebunny/crossref_metadata_2025_split huggingface dataset:
Development of a Simple Latex Agglutination Assay for Detection of Shiga Toxin-Producing
Escherichia coli
(STEC) by Using Polyclonal Antibody against STEC
Authors: Hajra Tapas K. , Bag Prasanta K. , Das Suresh C. , Mukherjee Souryadeep , Khan Asis , Ramamurthy T. 
Published: March 2007

ABSTRACT

Rabbit antiserum raised against the whole-cell antigen of Shiga toxin-producing
Escherichia coli
(STEC) strain VT3 (
stx
1
+
stx
2
+
eae
+
) was repeatedly adsorbed with heat-killed cells of different non-STEC strains and other enteric bacteria. Thus, the antiserum obtained was designated VT3 antiserum. VT3 antiserum reacted with intimin type γ. We assessed the reactivity of VT3 antiserum to whole-cell lysates of 87 strains of
E. coli
and other enteric bacteria by immunoblotting. The antiserum recognized the 97-kDa protein in whole-cell lysate from strain VT3, and 36 (83.7%) of the 43 STEC strains were positive for the STEC antigen. None of the non-STEC strains or strains of other species examined tested positive by immunoblotting. Based on this result, we developed a latex agglutination assay for the detection of STEC strains. Thirty-five (81.4%) of the 43 STEC strains tested positive for the STEC antigen by the latex agglutination assay. One (3.3%) of the 30 non-STEC strains and none of the strains of the other enteric bacteria included in this study tested positive by the latex agglutination assay. The corresponding specificity of the latex agglutination assay was approximately 98%. Results of this study showed the production of STEC antiserum and the generation of a simple, cost-effective, sensitive, and specific latex agglutination assay for establishing an etiological diagnosis of STEC.
Detection of H5 Avian Influenza Viruses by Antigen-Capture Enzyme-Linked Immunosorbent Assay Using H5-Specific Monoclonal Antibody
Authors: He Qigai , Velumani Sumathy , Du Qingyun , Lim Chee Wee , Ng Fook Kheong , Donis Ruben , Kwang Jimmy 
Published: March 2007

ABSTRACT
The unprecedented spread of highly pathogenic avian influenza virus subtype H5N1 in Asia and Europe is threatening animals and public health systems. Effective diagnosis and control management are needed to control the disease. To this end, we developed a panel of monoclonal antibodies (MAbs) against the H5N1 avian influenza virus (AIV) and implemented an antigen-capture enzyme-linked immunosorbent assay (AC-ELISA) to detect the H5 viral antigen. Mice immunized with denatured hemagglutinin (HA) from A/goose/Guangdong/97 (H5N1) expressed in bacteria or immunized with concentrated H5N2 virus yielded a panel of hybridomas secreting MAbs specific for influenza virus HA. The reactivity of each MAb with several subtypes of influenza virus revealed that hybridomas 3D4 and 8B6 specifically recognized H5 HA. Therefore, purified antibodies from hybridomas 3D4 and 8B6, which secrete immunoglobulin G (IgG) and IgM, respectively, were used as the capture antibodies and pooled hyperimmune guinea pig serum IgG served as the detector antibody. The specificity of the optimized AC-ELISA was evaluated by using AIV subtypes H5 H3, H4, H7, H9, and H10. Specimens containing AIV subtype H5 subtype yielded a specific and strong signal above the background, whereas specimens containing all other subtypes yielded background signals. The detection limits of the AC-ELISA were 62.5 ng of bacterium-expressed H5N1 HA1 protein and 124, 62, and 31 50% tissue culture infective doses of influenza virus subtypes H5N1/PR8, H5N2, and H5N3, respectively. Reconstituted clinical samples consisting of H5 AIVs mixed with pharyngeal-tracheal mucus from healthy chickens also yielded positive signals in the AC-ELISA, and the results were confirmed by reverse transcription-PCR. The tracheal swab samples from H9N2-infected chickens did not give positive signals. Taken together, the newly developed MAb-based AC-ELISA offers an attractive alternative to other diagnostic approaches for the specific detection of H5 AIV.
Maternal Milk Contains Antimicrobial Factors That Protect Young Rabbits from Enteropathogenic
Escherichia coli
Infection
Authors: Gallois Mélanie , Gidenne Thierry , Tasca Christian , Caubet Cécile , Coudert Cécile , Milon Alain , Boullier Séverine 
Published: March 2007

ABSTRACT

Enteropathogenic
Escherichia coli
(EPEC) colibacillosis represents a major cause of lethal diarrhea in young children in developing countries. EPEC strains also infect numerous mammal species and represent a major economical problem in rabbit industry. Protection against this pathogen is a challenging goal both in humans and in other mammal species. Despite a good knowledge of the pathogenicity mechanisms of EPEC, the intrinsic and environmental factors that control the expression of EPEC virulence in mammals remain unknown. For instance, the exacerbated sensitivity of young mammals to EPEC infection is still unexplained. Our goal was to investigate if age or other factors, like milk consumption, could be determinants that trigger the disease. We used rabbits as an animal model to study the role of milk in the sensitivity to an EPEC infection. Weaned and suckling rabbits were orally inoculated with EPEC strain E22 (O103:H2:K−) at 28 days of age, and the evolution of the disease was investigated in the two groups. In addition, in order to better characterize the interactions between milk and EPEC, we determined in vitro bacterial growth and the abilities of EPEC cells to adhere to epithelial cells in the presence of milk. Our results demonstrate a protective role of milk in vivo in association with in vitro antibacterial activity. These effects are independent of the presence of specific anti-EPEC antibodies.
Early Identification of Seronegative Human Immunodeficiency Virus Type 1 Infection with Severe Presentation
Authors: Chin Bum Sik , Lee Sun Hee , Kim Gab Jung , Kee Mee Kyung , Suh Soon Deok , Kim Sung Soon 
Published: March 2007

ABSTRACT
Specific antibodies against human immunodeficiency virus (HIV), usually used for diagnosis, almost invariably become detectable within 3 months of exposure. We report on a patient whose HIV infection was identified early by a combined antigen/antibody test, but seroconversion did not occur for 7 months, until the implementation of antiretroviral therapy.
Variability of Properties of
Vibrio parahaemolyticus
Strains Isolated from Individual Patients
Authors: Bhoopong Phuangthip , Palittapongarnpim Prasit , Pomwised Rattanaruji , Kiatkittipong Arpapak , Kamruzzaman Muhammad , Nakaguchi Yoshitsugu , Nishibuchi Mitsuaki , Ishibashi Masanori , Vuddhakul Varaporn 
Published: March 2007

ABSTRACT

Infections by strains belonging to the O3:K6 pandemic clone of
Vibrio parahaemolyticus
are prevalent in southern Thailand, and serovariants of these strains have also been detected.
V. parahaemolyticus
strains lacking important virulence genes (
tdh
and
trh
) were isolated from 6.5 to 10.9% of clinical specimens during the period from 2000 to 2003. In order to understand whether changes to the characteristics of
V. parahaemolyticus
occur during infection, 10 isolates collected from each of 63 patients who presented with diarrhea at the Hat Yai hospital from 2003 to 2004 were examined for the presence of the
tdh
and
trh
genes, the O:K serotype, and genetic markers for the pandemic clone. A total of 42 patients (66.7%) yielded identical isolates (homogeneous populations), and 21 of the patients (33.3%) yielded isolates that differed in at least one character from the other isolates (heterogeneous populations). The DNA fingerprints (examined by arbitrarily primed PCR and pulsed-field gel electrophoresis) of some, but not all, of the heterogeneous populations from single patients were indistinguishable. The results indicated that some patients were infected with a unique strain and that in vivo changes (
tdh
deletion or serotype conversion) might have occurred in certain individuals. It is therefore important to bear in mind that epidemiological studies based on the analysis of a single colony from a single patient might lead to misleading conclusions. Finally, the present study did not rule out the possibility that isolates lacking
tdh
and
trh
have unknown virulence mechanisms other than the
tdh
and
trh
genes.
Prevalence and Significance of a Negative Extended-Spectrum β-Lactamase (ESBL) Confirmation Test Result after a Positive ESBL Screening Test Result for Isolates of
Escherichia coli
and
Klebsiella pneumoniae
: Results from the SENTRY Asia-Pacific Surveillance Program
Authors: Bell Jan M. , Chitsaz Mohsen , Turnidge John D. , Barton Mary , Walters Luke J. , Jones Ronald N. 
Published: March 2007

ABSTRACT

A negative extended-spectrum β-lactamase (ESBL) confirmation test result obtained after a positive ESBL screening test result using Clinical and Laboratory Standards Institute methods has been a common occurrence among isolates of
Escherichia coli
and
Klebsiella pneumoniae
in the SENTRY Antimicrobial Surveillance Program in the Asia-Pacific region. Among isolates collected between 1998 and 2004 this screen-positive, nonconfirmed profile (failed to show clavulanate synergy) was observed in 8.9% of 4,515
E. coli
isolates and 20.3% of 2,303
K. pneumoniae
isolates. We then selected 52
E. coli
isolates and 68
K. pneumoniae
isolates with a negative ESBL confirmation test, as well as comparable number of isolates with confirmed ESBL-positive tests, and examined them for the presence of TEM, SHV, plasmid-borne
ampC
, and CTX-M genes. We found that 62% of nonconfirming
E. coli
isolates and 75% of nonconfirming
K. pneumoniae
harbored a plasmid-borne AmpC enzyme of the CIT or DHA type. The majority of nonconfirming
E. coli
and
K. pneumoniae
from the Asia-Pacific region harbor important β-lactamases, and a positive screening test alone should be sufficient grounds to report resistance to extended-spectrum cephalosporins in this region.
Comparison of the SPF
10
-LiPA System to the Hybrid Capture 2 Assay for Detection of Carcinogenic Human Papillomavirus Genotypes among 5,683 Young Women in Guanacaste, Costa Rica
Authors: Safaeian Mahboobeh , Herrero Rolando , Hildesheim Allan , Quint Wim , Freer Enrique , Van Doorn Leen-Jan , Porras Carolina , Silva Sandra , González Paula , Bratti M. Concepcion , Rodriguez Ana Cecilia , Castle Philip 
Published: March 2007

ABSTRACT

The objective of this analysis was to compare the performance characteristics of two human papillomavirus (HPV) DNA detections assays, the Hybrid Capture 2 assay (HC2) and the SPF
10
assay, for the detection of carcinogenic HPV. Data are from the enrollment visits of women who participated in the randomized, double-blind, placebo-controlled phase III HPV16/18 Vaccine Trial in Guanacaste, Costa Rica. We compared the results of HC2 and SPF
10
testing of cervical specimens. Since the line probe assay (LiPA) detection system does not distinguish between HPV type 68 (HPV68; which is targeted by HC2) and HPV73 (which is not targeted by HC2), for SPF
10
-LiPA, we defined the carcinogenic HPV types as the 12 HC2-targeted types (types 16, 18, 31, 33, 35, 39, 45, 51, 52, 56, 58, and 59), HPV68/73, and the HC2-cross-reactive, carcinogenic type HPV66. The kappa values and the performance characteristics for the detection of cervical abnormalities were ascertained. Paired observations were available for 5,683 sexually active, young women (median age, 21 years). The prevalence of carcinogenic HPV types was 35% (
n
= 1,962) by HC2 and 35% (
n
= 2,003) by SPF
10
-LiPA. There were no differences in the prevalence of carcinogenic HPV types by HC2 and SPF
10
-LiPA among women with normal, atypical squamous cells of undetermined significance and high-grade squamous intraepithelial lesion cytology. Among women with low-grade squamous intraepithelial lesion cytology, HC2 was more likely to test positive than SPF
10
-LiPA for the carcinogenic HPV types (87% and 79%, respectively;
P
= 0.001) as a result of HC2 cross-reactivity with HPV types 40, 43, 44, 53, 54, 60, 70, and 74. The crude agreement between the two assays was 88%, with a kappa value of 0.75 (95% confidence limits, 0.73 to 0.76). We observed very good agreement between HC2 and SPF
10
-LiPA for carcinogenic HPV type detection.
N-Terminal Mutants of Herpes Simplex Virus Type 2 gH Are Transported without gL but Require gL for Function
Authors: Cairns Tina M. , Friedman Lisa S. , Lou Huan , Whitbeck J. Charles , Shaner Marie S. , Cohen Gary H. , Eisenberg Roselyn J. 
Published: March 2007

ABSTRACT
Glycoprotein H (gH) is conserved among all herpesviruses and is essential for virus entry and cell fusion along with gL, gB, and, in most alphaherpesviruses, gD. Within the gH/gL heterodimer, it is thought that gH accounts for the fusion function and gL acts as a chaperone for the folding and transport of gH. Here, we found that the N terminus of gH2 contains important elements involved in both its folding and its transport. Our conclusions are based on the phenotypes of a series of gH deletion mutants in which the signal sequence (residues 1 to 18) was retained and N-terminal residues were removed up to the number indicated. The first mutant, gH2Δ29 (deletion of residues 19 to 28), like wild-type (WT) gH, required gL for both transport and function. To our surprise, two other mutants (gH2Δ64 and gH2Δ72) were transported to the cell surface independent of gL but were nonfunctional, even when complexed with gL. Importantly, a fourth mutant (gH2Δ48) was transported independent of gL but was functional only when complexed with gL. Using a panel of monoclonal antibodies against gH2, we found that when gH2Δ48 was expressed alone, its antigenic structure differed from that of gH2Δ48/gL or gH2-WT/gL. Mutation of gH2 residue R39, Y41, W42, or D44 allowed gL-independent transport of gH. Our results also show that gL is not merely required for gH transport but is also necessary for the folding and function of the complex. Since gH2Δ64/gL and gH2Δ72/gL were nonfunctional, we hypothesized that residues critical for gH/gL function lie within this deleted region. Additional mutagenesis identified L66 and L72 as important for function. Together, our results highlight several key gH residues: R39, Y41, W42, and D44 for gH transport and L66 and L72 for gH/gL structure and function.
Role of the Foamy Virus Pol Cleavage Site in Viral Replication
Authors: Roy Jacqueline , Linial Maxine L. 
Published: March 2007

ABSTRACT
Foamy virus Pol precursor protein processing by the viral protease occurs at only one site, releasing a protease-reverse transcriptase and an integrase protein. To examine whether the cleavage of the Pol precursor protein is necessary for enzymatic activities and efficient viral replication, several mutations were generated around the cleavage site. All cleavage site mutants synthesize wild-type levels of Pol precursor protein. Mutants containing more than two amino acid substitutions around the cleavage site exhibit no detectable Pol processing. The Pol cleavage site is not required for the production of infectious particles in a single round of infection, but is important for subsequent rounds of viral infection. Mutations around the cleavage site affected the enzymatic activities of the protease and reverse transcriptase and prevented replication after two rounds of infection. Interestingly, Pol encapsidation is significantly reduced in some of the mutants.
Demonstration of Coinfection with and Recombination by Caprine Arthritis-Encephalitis Virus and Maedi-Visna Virus in Naturally Infected Goats
Authors: Pisoni Giuliano , Bertoni Giuseppe , Puricelli Maria , Maccalli Marina , Moroni Paolo 
Published: March 2007

ABSTRACT

Recombination of different strains and subtypes is a hallmark of lentivirus infections, particularly for human immunodeficiency virus, and contributes significantly to viral diversity and evolution both within individual hosts and within populations. Recombinant viruses are generated in individuals coinfected or superinfected with more than one lentiviral strain or subtype. This, however, has never been described in vivo for the prototype lentivirus maedi-visna virus of sheep and its closely related caprine counterpart, the caprine arthritis-encephalitis virus. Cross-species infections occur in animals living under natural conditions, which suggests that dual infections with small-ruminant lentiviruses (SRLVs) are possible. In this paper we describe the first documented case of coinfection and viral recombination in two naturally infected goats. DNA fragments encompassing a variable region of the envelope glycoprotein were obtained from these two animals by end-limiting dilution PCR of peripheral blood mononuclear cells or infected cocultures. Genetic analyses, including nucleotide sequencing and heteroduplex mobility assays, showed that these goats harbored two distinct populations of SRLVs. Phylogenetic analysis permitted us to assign these sequences to the maedi-visna virus group (SRLV group A) or the caprine arthritis-encephalitis virus group (SRLV group B). SimPlot analysis showed clear evidence of A/B recombination within the
env
gene segment of a virus detected in one of the two goats. This case provides conclusive evidence that coinfection by different strains of SRLVs of groups A and B can indeed occur and that these viruses actually recombine in vivo.
Incorporation of Human Immunodeficiency Virus Type 1 Reverse Transcriptase into Virus-Like Particles
Authors: Liao Wei-Hao , Huang Kuo-Jung , Chang Yu-Fen , Wang Shiu-Mei , Tseng Ying-Tzu , Chiang Chien-Cheng , Wang Jaang-Jiun , Wang Chin-Tien 
Published: March 2007

ABSTRACT

We demonstrate that a genetically engineered human immunodeficiency virus type 1 (HIV-1) reverse transcriptase (RT) composed mainly of p66 or p51 subunits can be incorporated into virus-like particles (VLPs) when coexpressed with HIV-1 Pr55

gag

. VLP-associated RT exhibited a detergent-resistant association with immature cores during sucrose gradient equilibrium centrifugation, suggesting that RT is incorporated into VLPs. However, RT that retains downstream integrase (IN) is severely inhibited in terms of incorporation into VLPs. Results from immunofluorescence tests reveal that RT-IN is primarily localized at the perinuclear area and exhibits poor colocalization with Gag. IN removal leads to a redistribution of RT throughout the cytoplasm and improved RT incorporation into VLPs. Similar results were observed for RT-IN in which alanine was substituted for 186-Lys-Arg-Lys-188 residues of the IN putative nuclear localization signal, suggesting that IN karyophilic properties may partly account for the inhibitory effect of IN on RT incorporation. Although the membrane-binding capacity of RT was markedly reduced compared to that of wild-type Gag or Gag-Pol, the correlation of membrane-binding ability with particle incorporation efficiency was incomplete. Furthermore, we observed that membrane-binding-defective myristylation-minus RT can be packaged into VLPs at the same level as its normal myristylated counterpart. This suggests that the incorporation of RT into VLPs is independent of membrane affinity but very dependent on RT-Gag interaction. Results from a genetic analysis suggest that the Gag-interacting regions of RT mainly reside in the thumb subdomain and that the RT-binding domains of Gag are located in the matrix (MA) and p6 regions.
Severe Acute Respiratory Syndrome Coronavirus Accessory Protein 6 Is a Virion-Associated Protein and Is Released from 6 Protein-Expressing Cells
Authors: Huang Cheng , Peters C. J. , Makino Shinji 
Published: March 2007

ABSTRACT
Analysis of severe acute respiratory syndrome coronavirus (SCoV) by either sucrose gradient equilibrium centrifugation or a virus capture assay using an anti-SCoV S protein antibody demonstrated that the SCoV 6 protein, which is one of the accessory proteins of SCoV, was incorporated into virus particles. Coexpression of the SCoV S, M, E, and 6 proteins was sufficient for incorporation of the 6 protein into virus-like particles. Cells transfected with plasmid expressing the 6 protein released SCoV 6 protein; however, infected cells released SCoV 6 protein only in association with SCoV particles.
Biologic, Antigenic, and Full-Length Genomic Characterization of a Bovine-Like Coronavirus Isolated from a Giraffe
Authors: Hasoksuz Mustafa , Alekseev Konstantin , Vlasova Anastasia , Zhang Xinsheng , Spiro David , Halpin Rebecca , Wang Shiliang , Ghedin Elodie , Saif Linda J. 
Published: March 2007

ABSTRACTCoronaviruses (CoVs) possess large RNA genomes and exist as quasispecies, which increases the possibility of adaptive mutations and interspecies transmission. Recently, CoVs were recognized as important pathogens in captive wild ruminants. This is the first report of the isolation and detailed genetic, biologic, and antigenic characterization of a bovine-like CoV from a giraffe (Giraffa camelopardalis) in a wild-animal park in the United States. CoV particles were detected by immune electron microscopy in fecal samples from three giraffes with mild-to-severe diarrhea. From one of the three giraffe samples, a CoV (GiCoV-OH3) was isolated and successfully adapted to serial passage in human rectal tumor 18 cell cultures. Hemagglutination assays, receptor-destroying enzyme activity, hemagglutination inhibition, and fluorescence focus neutralization tests revealed close biological and antigenic relationships between the GiCoV-OH3 isolate and selected respiratory and enteric bovine CoV (BCoV) strains. When orally inoculated into a BCoV-seronegative gnotobiotic calf, GiCoV-OH3 caused severe diarrhea and virus shedding within 2 to 3 days. Sequence comparisons and phylogenetic analyses were performed to assess its genetic relatedness to other CoVs. Molecular characterization confirmed that the new isolate belongs to group 2a of the mammalian CoVs and revealed closer genetic relatedness between GiCoV-OH3 and the enteric BCoVs BCoV-ENT and BCoV-DB2, whereas BCoV-Mebus was more distantly related. Detailed sequence analysis of the GiCoV-OH3 spike gene demonstrated the presence of a deletion in the variable region of the S1 subunit (from amino acid 543 to amino acid 547), which is a region associated with pathogenicity and tissue tropism for other CoVs. The point mutations identified in the structural proteins (by comparing GiCoV-OH3, BCoV-ENT, BCoV-DB2, and BCoV-Mebus) were most conserved among GiCoV-OH3, BCoV-ENT, and BCoV-DB2, whereas most of the point mutations in the nonstructural proteins were unique to GiCoV-OH3. Our results confirm the existence of a bovine-like CoV transmissible to cattle from wild ruminants, namely, giraffes, but with certain genetic properties different from those of BCoVs.