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PubMed1992

Monoclonal antibodies as probes to examine serotype-specific and cross-reactive epitopes of lipopolysaccharides from serotypes O2, O5, and O16 of Pseudomonas aeruginosa.

Serotypes O2, O5, and O16 of Pseudomonas aeruginosa are chemically related, and the O antigens of their lipopolysaccharides share a similar trisaccharide repeat backbone structure. Serotype-specific monoclonal antibodies (MAbs) MF71-3, MF15-4, and MF47-4 against the O2, O5, and O16 serotypes, respectively, were isolated. MAb 18-19, which is cross-reactive with all strains of this chemically related serogroup, was also produced. When column chromatography or sodium dodecyl sulfate-polyacrylamide gel electrophoresis-separated lipopolysaccharide (LPS) samples from each of the serotypes were probed with the MAbs in Western immunoblots, each of the serotype-specific MAbs interacted only with high-molecular-weight bands of the homologous LPS, with a minimum O-antigen chain length of at least 6 to 10 repeats. In contrast, cross-reactive MAb 18-19 was shown to interact in Western immunoblots with the entire LPS banding pattern except the fastest-running band, which lacks O antigen. Chemical modification of P. aeruginosa LPS by alkali treatment and carboxyl reduction abolished reactions between LPS and MAb 18-19, while reactions of modified LPS with serotype-specific MAbs were not affected. Therefore, cross-reactive MAb 18-19 likely recognizes the chemical backbone structure of the O repeat that is common to all three serotypes of the O2-O5-O16 group, while the O-specific MAbs appeared to recognize LPS epitopes that could be presented when 6 to 10 or more O-antigen repeat units are present on the LPS molecule. Thus, the O-specific LPS epitopes likely involve unique chemical structures, glycosidic linkages, and some order of folding of the O side chains.

Antibodies, Bacterial↗
PubMed1995

Relationships between rfb gene clusters required for biosynthesis of identical D-galactose-containing O antigens in Klebsiella pneumoniae serotype O1 and Serratia marcescens serotype O16.

The lipopolysaccharide O antigens of Klebsiella pneumoniae serotype O1 and Serratia marcescens serotype O16 both contain a repeating unit disaccharide of [-->3)-beta-D-Galf-(1-->3)-alpha-D-Galp-(1-->]; the resulting polymer is known as D-galactan I. In K. pneumoniae serotype O1, the genes responsible for the synthesis of D-galactan I are found in the rfb gene cluster (rfbKpO1). We report here the cloning and analysis of the rfb cluster from S. marcescens serotype O16 (rfbSmO16). This is the first rfb gene cluster examined for the genus Serratia. Synthesis of D-galactan I is an rfe-dependent process for both K. pneumoniae serotype O1 and S. marcescens serotype O16. Hybridization experiments with probes derived from each of the six rfbKpO1 genes indicate that the cloned rfbSmO16 cluster contains homologous genes arranged in the same order. However, the degree of homology at the nucleotide sequence level was sufficiently low that hybridization was detected only under low-stringency conditions. rfbABSmO16 genes were subcloned and shown to encode an ABC-2 (ATP-binding cassette) transporter which is functionally identical to the one encoded by the corresponding rfb genes from K. pneumoniae serotype O1. The amino acid sequences of the predicted RfbA and RfbB homologs showed identities of 75.7% (87.9% total similarity) and 78.0% (86.5% total similarity), respectively. The last gene of the rfbKpO1 cluster, rfbFKpO1, encodes a bifunctional galactosyltransferase which initiates the formation of D-galactan I. RfbFKpO1 and RfbFSmO16 are 57.6% identical (with 71.1% total similarity), and both show similarity with RfpB, the galactosyltransferase involved in the synthesis of Shigella dysenteriae type I O-polysaccharide. The G+C contents of the rfbAB genes from each organism are quite similar, and values are lower than those typical for the species. However, the G+C content of rfbFSmO16 (47.6%) was much higher than that of rfbFKpO1 (37.3%), despite the fact that the average for each species (52 to 60%) falls within the same range.

ATP-Binding Cassette Transporters↗
PubMed2004

Dengue virus serotyping based on envelope and membrane and nonstructural protein NS1 serotype-specific capture immunoglobulin M enzyme-linked immunosorbent assays.

Envelope and membrane (E/M) and nonstructural protein NS1 serotype-specific capture Immunoglobulin M (IgM) enzyme-linked immunosorbent assays (ELISAs) were developed to differentiate four dengue virus serotypes. A total of 93 anti-dengue virus IgM-positive serum samples collected between days 5 and 45 of illness from 59 confirmed dengue patients were analyzed. The results showed that positive serotype specificity could be identified for 86.1 and 47.6% of serum samples tested for E/M-specific IgM antibodies versus 83.3 and 42.9% of serum samples tested for NS1-specific IgM antibodies from patients with primary and secondary dengue virus infections, respectively. Dual analyses with both E/M and NS1 serotype-specific capture IgM ELISAs showed that positive serotype specificity could be correctly identified for 98.6 and 61.9% of all of the primary and secondary serum samples tested, respectively. These findings suggested that E/M and NS1 serotype-specific capture IgM ELISAs have the potential to be of use in dengue virus serotyping.

Antibodies, Viral↗
PubMed2005

Clonal association between Streptococcus pneumoniae serotype 23A, circulating within the United States, and an internationally dispersed clone of serotype 23F.

Streptococcus pneumoniae is an important pathogen in the United States and is associated with significant morbidity and mortality. Since the introduction of the seven-valent conjugate vaccine, a significant decline in pneumococcal disease has been reported. However, surveillance for pneumococcal disease remains essential, as the extent of cross protection against vaccine-related serotypes is still unclear. Further, any increase in non-vaccine-related serotypes also needs monitoring. We report on a new clonal association between a vaccine-related serotype, serotype 23A, obtained as part of the Active Bacterial Core surveillance, with an established internationally dispersed Pneumococcal Molecular Epidemiology Network (PMEN) clone, clone Colombia(23F)-26. Sixty-two isolates of serotype 23A collected from sterile sites during a 2-year period (2002 and 2003) were characterized. Twenty-one (34%) isolates were penicillin nonsusceptible, although none were fully resistant. Pulsed-field gel electrophoresis and multilocus sequence typing analysis showed that 24 (39%) of the serotype 23A isolates shared either genetic identity or high genetic relatedness with PMEN clone Colombia(23F)-26. Extensive variability was noted within the sequenced region of pbp2b in two penicillin-nonsusceptible isolates as well as in PMEN clone Colombia(23F)-26, suggesting that these isolates probably acquired penicillin resistance independently. The emergence of such new serotype and genotype associations highlights the dynamic nature of the pneumococcal population, necessitating continuous monitoring in the post-vaccine era.

Adolescent↗
PubMed2005

Fatal meningitis in a previously healthy young adult caused by Streptococcus pneumoniae serotype 38: an emerging serotype?

BACKGROUND: In December 2001, a fatal case of pneumococcal meningitis in a Marine Corps recruit was identified. As pneumococcal vaccine usage in recruit populations is being considered, an investigation was initiated into the causative serotype. CASE PRESENTATION: Traditional and molecular methods were utilized to determine the serotype of the infecting pneumococcus. The pneumococcal isolate was identified as serotype 38 (PS38), a serotype not covered by current vaccine formulations. The global significance of this serotype was explored in the medical literature, and found to be a rare but recognized cause of carriage and invasive disease. CONCLUSION: The potential of PS38 to cause severe disease is documented in this report. Current literature does not support the hypothesis that this serotype is increasing in incidence. However, as we monitor the changing epidemiology of pneumococcal illness in the US in this conjugate era, PS38 might find a more prominent and concerning niche as a replacement serotype.

Adolescent↗
PubMed1993

A comparative evaluation of the safety and immunogenicity of a single dose of unbuffered oral rhesus rotavirus serotype 3, rhesus/human reassortant serotypes 1, 2 and 4 and combined (tetravalent) vaccines in healthy infants.

To assess safety and immunogenicity, 213 healthy infants aged 6 weeks to 4 months were randomized to receive a single dose of placebo, a 10(4) or 10(5) p.f.u. dose of rhesus rotavirus (RRV) serotype 3, human-RRV reassortant (VP-7 serotypes 1, 2 or 4) or a 10(4) or 10(5) p.f.u. dose of tetravalent rotavirus vaccine (containing equal parts of serotype 1, 2, 3 and 4 strains). The infants were fed ad libitum before and after vaccination; no buffer was used. For 7 days after vaccination, potential vaccine side effects were monitored, and no significant differences were noted for any symptom evaluated among the single serotype, tetravalent or placebo groups. Sera, obtained before and 28 days after vaccination, were measured for antibody to rotavirus by IgG, IgA and IgM enzyme-linked immunosorbent assay in all subjects, and by neutralizing antibody to the individual serotypes by plaque reduction in placebo and tetravalent vaccinees. The serological response rates for serotypes 1, 2, 3, 4 and the tetravalent vaccine were 25, 12, 19, 11 and 22%, respectively, at 10(4) p.f.u.; 47, 50, 35, 29 and 61%, respectively, at 10(5) p.f.u.; and 37% for placebo. The tetravalent vaccine was more immunogenic at 10(5) than at 10(4) p.f.u. (p = 0.04). Grouped together, the vaccines at 10(5) p.f.u. (single serotype and tetravalent) were more immunogenic than the vaccines at 10(4) p.f.u. (38 of 85 versus 17 of 94 seroresponders; p < 0.001).(ABSTRACT TRUNCATED AT 250 WORDS)

Antibodies, Viral↗
PubMed2004

[Recovery method of serotypable character in non serotypable pseudomonas aeruginosa strains].

Serotyping is one of the most used techniques for typing Pseudomonas aeruginosa strains. During chronic infections, and especially in cystic fibrosis, the decrease of lipopolysaccharide production is responsible for difficulties in determining O antigens. The possibility of serotyping can be simply restored by using a primary culture broth containing amikacin (1/6 of the strain MIC for this antibiotic); this is due to the ability of this antibiotic to inhibit alginate production. This technique allowed us to determine the serotype of 108 non-serotypable strains of P. aeruginosa isolated in 14 different hospitals. Among these isolates, serotype O:1 and O:13, had a high prevalence; the origin is a deficiency in D-glucose and L-rhamnose, required for the synthesis of lipopolysaccharide. In contrast, these sugars are not present in lipopolysaccharide of O:12, and these strains are always serotypable. The main protein is Alg C; this bifunctional enzyme is required in the exopolysaccharide and lipopolysaccharide production, according stress conditions in the bacterial-cells' environment. Determination of the serotype, as Antibiogram, is essential for genotypic inquiries.

Bacterial Typing Techniques↗
PubMed1992

The restriction endonuclease map of Marek's disease virus (MDV) serotype 2 and collinear relationship among three serotypes of MDV.

A BamHI, EcoRI, and XhoI restriction endonuclease map of Marek's disease virus (MDV) serotype 2 (MDV2) DNA was constructed by double-digest analyses of 28 cloned BamHI and 11 cloned EcoRI fragments of MDV2 DNA, followed by hybridization tests of these cloned BamHI DNA fragments with electrophoretically separated digests of MDV2-infected cell DNA. On this map, MDV2 genome consisted of two segments which have unique regions inserted between two inverted repeat regions as observed in MDV serotype 1 and 3 genomes. Further, the DNA homology among three serotypes of MDV was examined by hybridization under less stringent conditions using cloned BamHI fragments of MDV2 DNA. Most of the MDV2 fragments located within the unique regions hybridized with MDV serotype 1 and 3 DNAs, indicating the presence of the collinear relationship among three serotypes. In addition, MDV2 DNA fragments which hybridized with the DNA fragments encoding MDV1 gp57-65 (or A antigen) or MDV1 gp100, gp60, gp49 (or B antigen) were identified and these fragments of serotypes 1 and 2 found to be collinear.

Animals↗
PubMed2006

Development of a DNA microarray to identify the Streptococcus pneumoniae serotypes contained in the 23-valent pneumococcal polysaccharide vaccine and closely related serotypes.

Streptococcus pneumoniae is a major worldwide human pathogen. This investigation has developed a reliable and accurate DNA microarray method for inter-species differentiation of S. pneumoniae and intra-species differentiation of the 23 groups of S. pneumoniae including serotypes represented in the 23-valent pneumococcal vaccine and the other 20 closely related serotypes. In addition to 16S rDNA probes, serotype- or serogroup-specific probes targeting the capsular polysaccharide synthesis (cps) genes, wzy or capA were generated. We adopted a two-step multiplex PCR to improve the sensitivity of detection to a level of 10(5) cfu/ml in pure culture or 50 ng DNA. A total of 169 isolates (from China, Australia, Canada and New Zealand) including 147 belonging to 23-valent vaccine and closely related serotypes of S. pneumoniae, 11 belonging to other serotypes and 11 of different species commonly isolated from respiratory tract were tested to verify the method. The DNA microarray method developed provides a sensitive means to rapidly identify the members of the most common S. pneumoniae serotypes in patients and to monitor their distribution in different patient groups and geographic locations. Such information is needed for disease surveillance and to monitor vaccine efficacy.

Bacterial Proteins↗
PubMed1976

The stability of the serotypes of Bordetella pertussis with particular reference to serotype 1,2,3,4.

Strains of Bordetella pertussis in which all the organisms contain agglutinogens 1 and 3 or 1,2 and 4 are easy to identify as serotypes 1,0,3,0 and 1,2,0,4 respectively; and similarly, stable strains of serotype 1,0,3,4 are occasionally found. During repeated subcultures, passage in vivo, and lyophilization and preservation for many years, these serotypes do not change. Mixing 1,0,3,0 and 1,2,0,4 serotypes and culturing them together in vivo and in vitro produces cultures from which organisms of the same two serotypes can be isolated. In contrast, strains which type as 1,2,3,4 are often a heterogeneous group. We have attempted to classify these as "stable", "variable" and "mixed" cultures. Some strains comprise organisms all of which contain the four agglutinogens and are as easy to type as the strains described above. These we have called "stable" 1,2,3,4 strains. Other 1,2,3,4 strains are made up of colonies possessing all four agglutinogens, as shown by agglutinin production, but in amounts varying from day to day so that direct typing is inconsistent. These we have called "variable" 1,2,3,4 strains. The last category, "mixed", is made up of organisms most of which give rise to stable 1,2,3,4 cultures; a few of the component organisms, however, have one or two of the four agglutinogens missing. The importance of the "variable" cultures is emphasized for work on apparent change of serotype, e.g. during infection.

Agglutination Tests↗
PubMed1997

Serotyping of Streptococcus pyogenes isolated from common and severe invasive infections in Japan, 1990-5: implication of the T3 serotype strain-expansion in TSLS. The Working Group for Group A Streptococci in Japan.

To clarify the relationship between the epidemics of severe invasive group A streptococcal infections (streptococcal Toxic Shock-Like Syndrome: TSLS) and common group A streptococcal infections in Japan, we examined the T serotypes of S. pyogenes strains (group A streptococci) isolated from clinical specimens of the streptococcal infections (17999 cases) in the period 1990-5, including the severe infections (TSLS) (29 cases) in the period 1992-5. Characteristic points of the analyses were: (1) dominant serotypes of the infections in these periods were T12, T4, T1, T28 and TB3264, which were consistently isolated; (2) isolates of T3 rapidly increased through 1990 to 1994 while T6 decreased in the period 1990-3; (3) when Japanese area was divided into three parts, T3 serotype tended to spread out from the north-eastern to the south-western area; (4) strains of T3 and T1 serotypes were dominant in the TSLS. Dominant-serotype strains of streptococcal infections did not always induce severe infections and dominance of T3 serotype in the TSLS seemed to be correlated with the increase of T3 in streptococcal infections. These results may indicate that certain clones of S. pyogenes are involved in the pathogenesis of the TSLS.

Adolescent↗
PubMed1985

Protection against infection with Neisseria meningitidis group B serotype 2b by passive immunization with serotype-specific monoclonal antibody.

Hybridomas derived from mice immunized with Neisseria meningitidis serogroup B serotype 2b (B,2b) outer membrane preparations produced monoclonal antibodies (MAbs) specific for major outer membrane proteins of classes 1, 2, and 5. The MAbs were examined by enzyme-linked immunosorbent assay against a selected panel of seven strains of N. meningitidis (B,2b) of different sodium dodecyl sulfate-polyacrylamide gel electrophoresis patterns, a serotype 2a, and a nontypable strain. The five MAbs selected were all bactericidal and of different immunoglobulin subclasses. None of the MAbs reacted with other bacterial strains in a dot-enzyme immunoassay. The corresponding antigenic determinant for each MAb was localized on a specific outer membrane protein by immunoblotting of sodium dodecyl sulfate-polyacrylamide gel electrophoresis patterns of major outer membrane proteins. MAbs M5-11 and M5-30 bound to the class 2 protein and were serotype 2b specific. MAb M2-20 bound to the class 1 protein, and MAbs M5-16 and M5-19 bound to the class 5 protein. A mouse model of infection was established whereby a local infection progressed to lethal bacteremia over 3 days, and 50% of the animals were killed with an intraperitoneal injection of 10 meningococci plus 4% mucin and 1.6% hemoglobin. The ability of the MAbs to provide passive protection against experimental infection with N. meningitidis (B,2b) was examined. Both serotype-specific MAbs M5-11 and M5-30 were highly protective even though they were of different immunoglobulin subclasses. The class 5-specific MAb offered no protection, while the class 1-specific MAb gave limited protection. It may therefore be possible to provide protection against serotype 2b infection by using as vaccine the class 2 serotype-specific surface-exposed outer membrane protein epitopes defined by MAb M5-11 or M5-30.

Animals↗
PubMed2001

Clinical isolates of non-O157 Shiga toxin-producing Escherichia coli: serotypes, virulence characteristics, and molecular profiles of strains of the same serotype.

All human Shiga toxin-producing Escherichia coli (STEC) non-O157 strains (n = 56) isolated in Finland from 1990 to August 2000 were characterized for the O:H serotype, stx(1) and stx(2) genes, production of enterohemolysin, and sensitivity to 12 antimicrobial agents. Strains of the same serotype were genotyped by pulsed-field gel electrophoresis (PFGE) after XbaI restriction of total DNA. The 56 non-O157 isolates belonged to 29 serotypes. Two of the serotypes (O102:H7 and OX181:H49) have not previously been described as being associated with STEC infections in humans or isolated from animals. Thirty-four strains (61%) within seven serotypes (O103:H2 [14 isolates], O26:H11 [6 isolates], O145:H28 [4 isolates], O145:HNM [3 isolates], O15:HNM [3 isolates], OX174:H21 [2 isolates], and O Rough:HNM [2 isolates]) were represented by more than one isolate. Of these strains, O103:H2 isolates were divided into seven, O26:H11 isolates were divided into four, and the rest within a serotype were divided into two genotypes in PFGE. In PCR, 31 (55%) of the 56 strains were positive for the stx(2) gene only and 24 strains (43%) were positive for stx(1) only. One strain (O43:H2) carried both stx(1) and stx(2). Forty-two strains (75%) produced enterohemolysin, and 39 strains (70%) possessed the eae gene. Of the latter 39 strains, 36 (92%) were enterohemolytic, whereas only 6 (35%) of the 17 isolates lacking the eae gene were enterohemolytic (P < 0.001). The majority of the strains (44 strains, 79%) were sensitive to all 12 antimicrobials tested. Of the 56 strains, 20 (36%) were associated with small family outbreaks in nine families and 14 (25%) were associated with recent travel abroad.

Bacterial Typing Techniques↗
PubMed2003

Serotyping of Listeria monocytogenes by enzyme-linked immunosorbent assay and identification of mixed-serotype cultures by colony immunoblotting.

Routine analysis of Listeria monocytogenes by serotyping using traditional agglutination methods is limited in use because of the expense and limited availability of commercially prepared antisera and intra- and interlaboratory discrepancies arising from differences in antiserum preparation and visual determination of agglutination. We have adapted a commercially available set of L. monocytogenes antisera to an enzyme-linked immunosorbent assay (ELISA) format for high-throughput, low-cost serotype determination. Rather than subjective visualization of agglutination, positive antigen and antiserum reactions were scored by a quantitative, colorimetric reaction. ELISA serotyping of 89 of 101 L. monocytogenes isolates agreed with slide agglutination serotyping data, and 100 previously uncharacterized isolates were serotyped unambiguously by the ELISA method. In addition, mixed-serotype cultures of L. monocytogenes were identified by a colony immunoblot procedure, in which serogroup 1/2 and serogroup 4 colonies were discriminated by differential staining.

Agglutination↗
PubMed2004

Serotype-specific entry of dengue virus into liver cells: identification of the 37-kilodalton/67-kilodalton high-affinity laminin receptor as a dengue virus serotype 1 receptor.

Dengue virus, the causative agent of dengue fever, dengue shock syndrome, and dengue hemorrhagic fever, infects susceptible cells by initially binding to a receptor(s) located on the host cell surface. Evidence to date suggests that receptor usage may be cell and serotype specific, and this study sought to identify dengue virus serotype 1 binding proteins on the surface of liver cells, a known target organ. By using a virus overlay protein binding assay (VOPBA), in both nondenaturing and denaturing gel systems, a putative dengue virus serotype 1 binding protein of approximately 37 kDa expressed on the surface of liver (HepG2) cells was identified. Mass spectrometry analysis identified a candidate protein, the 37/67-kDa high-affinity laminin receptor. Entry of the dengue virus serotype 1 was significantly inhibited in a dose-dependent manner by both antibodies directed against the 37/67-kDa high-affinity laminin receptor and soluble laminin. No inhibition of virus entry was seen with dengue virus serotypes 2, 3, or 4, demonstrating that the 37/67-kDa high-affinity laminin receptor is a serotype-specific receptor for dengue virus entry into liver cells.

Animals↗
PubMed1987

Serotypic antigens of Plasmodium falciparum recognized by serotype-restricted inhibitory human sera.

Immune sera from some Cambodian refugees contain functional serotypic antibodies that inhibit invasion of erythrocytes by the Camp strain but not by the FCR-3 strain of Plasmodium falciparum. Using a new assay, the "competitive heterologous antigen assay" (CHAA), the serotypic antibodies in a pool of three inhibitory sera were characterized by the antigens they precipitated. In the CHAA, immunoprecipitation of antigens by antibodies to common or cross-reacting antigenic determinants was blocked with excess heterologous unlabeled FCR-3 antigens before 3H-labeled Camp schizont and merozoite antigens were immunoprecipitated. The predominant Camp strain serotypic antigens revealed after electrophoresis and autoradiography were the major 195 Kd glycoprotein surface antigen (gp195) and its processed products at 150, 83, 73, and possibly 45 Kd. Additional serotypic antigens were identified at 180, 130, 65, 50, and 32 Kd. It is likely that one or more of these serotypic antigens is a target for the serotypic antibodies that inhibit invasion.

Animals↗
PubMedDate not supplied

Identification and analysis of the Georgia 98 serotype, a new serotype of infectious bronchitis virus.

Twenty-five field infectious bronchitis viruses (IBVs) similar to, but genetically distinct from, the DE072 serotype were isolated from several states in the United States from 1990 through 1999 and were examined molecularly and antigenically. A 421-bp sequence in the hypervariable region of the S1 gene was examined, and phylogenetic analysis on that region indicated that these viruses are closely related but fall into unique groups. Cross-virus neutralization testing and entire S1 sequence analysis on selected isolates further confirmed that fact, and we divided the viruses into the DE072 serotype and two other unique groups. In a vaccine protection trial, the commercially available DE072 vaccine showed less than 50% protection against viruses in one of the groups. The majority of the recent isolates belong to that group and share very low antigenic relatedness to the DE072 strain as well as other serotypes of IBV. Consequently, we designated this group as a new serotype, Georgia 98. We developed a restriction fragment length polymorphism analysis that can differentiate this new serotype from all other serotypes of IBV.

Animals↗