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PubMed2005

Clonal analysis of invasive pneumococcal isolates in Scotland and coverage of serotypes by the licensed conjugate polysaccharide pneumococcal vaccine: possible implications for UK vaccine policy.

A 7-valent pneumococcal conjugate vaccine (PCV7) has gained licensure and has proven successful in the USA for preventing pneumococcal disease and reducing the incidence of antibiotic-resistant pneumococcal strains. The ability, therefore, to accurately monitor the likely effect of the introduction of PCV7 vaccine on invasive pneumococcal disease in the UK is essential. Serotyping and multilocus sequence typing was performed on invasive isolates of Streptococcus pneumoniae (n=645) from Scotland during 2003. The information gained from this was used to evaluate serotype coverage by the vaccine and the relationship between serotypes. In the present study, invasive pneumococcal disease in Scotland was caused by 33 different serotypes, consisting of 150 sequence types. Overall, 48.4% of the isolates were of serotypes included in the PCV7. Pneumococci were most frequently associated with sequence types 9, 124, and 162. PCV7 would provide protection in 71.8% of infants under 5 years of age against the serotypes in the vaccine. There was limited evidence of the potential for capsule switch among currently circulating invasive pneumococci. The successful implementation of a suitable vaccination programme should lead to a reduction in invasive pneumococcal disease in the UK as well as a reduction in antibiotic resistance of pneumococcal strains.

Child, Preschool↗
PubMed1987

Comparison of the amino acid sequences of the major neutralization protein of four human rotavirus serotypes.

We sequenced the gene coding for the major neutralizing protein (VP7) from eight human rotavirus strains representing serotype 1, 2, 3, or 4. In addition, the corresponding gene of the rhesus rotavirus vaccine strain MMU 18006 (serotype 3) was sequenced. Comparative analyses of their deduced amino acid sequences revealed an overall 15-29% divergence in the VP7 proteins that define four different rotavirus serotypes and confirmed the presence of six discrete regions of clustered sequence divergence (amino acids 39-50, 87-101, 120-130, 143-152, 208-221, and 233-242). When the same regions were compared among rotaviruses belonging to the same serotype, a high degree of homology (91-99%) was detected. These observations indicate that differences in the serotype specificity among rotaviruses are the result of a high degree of sequence divergence in several discrete regions of the VP7 gene and that these regions are highly conserved within a given serotype.

Amino Acid Sequence↗
PubMed1992

A bovine rotavirus serotype 1: serologic characterization of the virus and nucleotide sequence determination of the structural glycoprotein VP7 gene.

Bovine rotavirus T449 was isolated from feces of a calf with diarrhea. Serological characterization by serotype-specific monoclonal antibodies showed that the T449 virus belonged to serotype 1. This is the first report of a bovine rotavirus that does not belong to serotype 6, 8, or 10. The serotype 1 designation was confirmed by using an immunoperoxidase focus neutralization assay. The gene encoding the major neutralization antigen (VP7) was cloned and its nucleotide sequence was determined. The sequence obtained was 1062 bp in length and contained an open reading frame corresponding to 326 amino acid residues. Comparative analysis of the deduced amino acid sequence with the corresponding sequence of the human serotype 1 rotavirus strain, Wa, revealed a 90% identity. When compared to the predicted amino acid sequence of VP7 protein of the other serotypes an overall divergence of 20 to 25% was detected. These data show that the serological typing agrees with the result of the genetic analysis.

Amino Acid Sequence↗
PubMed1988

Genetic relationships of bluetongue virus serotypes isolated from different parts of the world.

Full-length DNA clones representing the 10 double-stranded RNA segments of US bluetongue virus serotype 10 (BTV-10) have been used in a study to determine the genetic relationships among 20 different BTV serotypes. The study was undertaken using Northern blot hybridization techniques involving 32P-labelled DNA probes and total RNA species extracted from BHK-21 cells infected with 20 different BTV serotypes. The results obtained indicate that all the genes representing the nonstructural proteins of BTV (NS1, NS2 and NS3) as well as most of the inner capsid polypeptides are highly conserved (e.g., VP1, VP3, VP4), while VP6 and VP7, the remaining two inner capsid components, are less conserved. The genes representing the two outer capsid polypeptides, VP2 and VP5, vary significantly. When complete DNA clones of RNA segment 2 (representing the VP2 neutralization gene) of 4 other US serotypes (BTV-2, -11, -13 and -17) and one Australian serotype (BTV-1) were used in similar hybridization studies, the data obtained showed that despite geographical distances, a certain BTV serotype exhibits similarities. Some hybridization signals were detected with several of the inner capsid genes and the corresponding RNA segments of epizootic hemorrhagic disease virus (EHDV), a distantly related orbivirus, although none of the BTV outer capsid genes, nor any of the nonstructural genes hybridized with either EHDV-1 or EHDV-2 RNA species.

Animals↗
PubMed1989

Naturally occurring serotype 2/subgroup II rotavirus reassortants in northern Brazil.

Nine serotype 2 human rotavirus strains were isolated in a community-based longitudinal study in Northern Brazil. Five of these strains had a 'long' RNA electrophoretic pattern and all five strains were determined to belong to subgroup II by ELISA assay, in contrast to properties common to serotype 2 human rotaviruses previously characterized. Hybridization studies of one of these unusual strains with 32P-labelled mRNAs derived from the prototype human strains Wa (serotype 1, subgroup II) and S2 (serotype 2, subgroup I) suggested that it was generated by a reassortment event in nature, in which a subgroup II, 'long' electropherotype rotavirus exchanged its serotype-specific gene and gene number 10 for the equivalent genes from a serotype 2, 'short' electropherotype virus.

Brazil↗
PubMed1994

Phylogenetic relationships of the VP2 protein of a virulent isolate of bluetongue virus (BTV-23) compared to those of 6 other BTV serotypes.

To determine the genetic relationship of the virulent Australian bluetongue virus serotype 23 with that of other serotypes and to identify the extent and nature of the antigenic variation among seven serotypes of bluetongue virus (BTV), the complete nucleotide sequence was determined for cDNA clones representing the L2 dsRNA of BTV-23, the gene that codes for the outer capsid neutralization antigen (VP2). The predicted amino acid sequence of the protein was compared with the VP2 sequences of the five USA serotypes (BTV-2, -10, -11, -13 and -17) as well as with an Australian isolate of BTV-1. The comparisons revealed that the VP2 of BTV-23 is most closely related to that of BTV-1, sharing 52% identical and 72% similar sequences, also that the VP2 of the two Australian serotypes are more closely related to that of BTV-2 than to the other four USA serotypes. Only 22% identical sequences are shared by all seven VP2 molecules; however, when homologous substitutions are considered the similarity index was as high as 48%. In addition, the conserved regions that have been identified previously for other VP2 molecules are also conserved in BTV-23.

Amino Acid Sequence↗
PubMed1991

Immunogenicity and safety of rhesus-human rotavirus reassortant vaccines with serotype 1 or 2 VP7 specificity.

Rhesus-human rotavirus reassortants incorporating the gene expressing the VP7 surface protein of human rotavirus serotypes 1 or 2, and the remaining ten genes from rhesus rotavirus (RRV) were evaluated as candidate oral vaccines in 2-4-month-old infants. A single dose of the serotype 1 reassortant vaccine which had a titre of 10(4) plaque-forming units (p.f.u.) induced a fourfold or greater antibody response in 81% of the recipients by a combination of ELISA and neutralization assays; 51% of the vaccinees developed a neutralizing antibody response to the vaccine strain. A single dose of the serotype 2 vaccine (10(4) p.f.u.) induced a seroresponse in all vaccinees by the combination of assays whereas 67% developed neutralizing antibodies to the vaccine strain. A combination of these two vaccines (0.5 x 10(4) p.f.u. of each) induced an overall seroresponse in 95% of the recipients but only 48% and 24% response in neutralizing antibodies to serotypes 1 and 2, respectively. A trivalent combination which included the two reassortants and RRV (0.33 x 10(4) p.f.u. of each strain) induced an overall response in 82% of the vaccinees, but only 30%, 20% and 65% developed a neutralizing antibody response to serotype 1, serotype 2, and RRV, respectively. Febrile reactions on days 2-5 after vaccination were seen in 23-45% of the infants receiving the various vaccines and combinations and in 5% of the placebo group. It is concluded that rhesus-human reassortant rotaviruses may be combined with each other and with RRV as a polyvalent vaccine, but the VP7-specific neutralizing antibody responses are likely to be lower after combined vaccination than following vaccination with a single reassortant rotavirus.

Animals↗
PubMed1994

Efficacy of rhesus rotavirus vaccine MMU-18006 against gastroenteritis due to serotype 1 rotavirus.

We conducted a clinical trial of rhesus rotavirus vaccine MMU-18006 (RRV, serotype 3) to assess the immunogenicity, transmissibility and booster effect of this vaccine in a welfare nursery in Sapporo, from September 1986 to October 1988. After the trial, in March 1989, an outbreak of gastroenteritis due to a wild strain of serotype 1 rotavirus (RV-1) occurred in the study population. Infants were divided into three groups based on vaccination history: five booster vaccinees, 18 one-dose vaccinees and 18 control infants who did not receive vaccine. There was a significant relationship between asymptomatic infection and higher levels of preoutbreak antibody titres against KU (serotype 1) but not RRV. Significant protection from rotavirus illness was observed both in the booster vaccine group and in the one-dose vaccine group but not in the control group. Rotavirus-specific serum IgA immune response was considered to be one of the indicators of recent rotavirus infection, and did not correlate with resistance to rotavirus illness. Our results revealed that protection from rotavirus illness was serotype-specific and that previous rotavirus infection, including vaccination, was important to induce the heterotypic immune response, and that ageing or booster inoculation of RRV might play a role in the protection against serotype 1 rotavirus infection. From our findings, a booster administration was thought to be important to induce effective heterotypic immunity and should be included in a future rotavirus vaccine trial to obtain sufficient protection against four major serotypes of rotavirus.

Antibodies, Viral↗
PubMed1990

Development of specific nucleic acid probes for the differentiation of porcine rotavirus serotypes.

A dot blot hybridization assay is described for the detection and differentiation of porcine rotavirus serotypes. Recombinant complementary DNA (cDNA) representing gene 9 (the gene encoding the neutralization antigens in VP7 glycoprotein) from OSU (porcine rotavirus serotype 1) and Gottfried (porcine rotavirus serotype 2) strains were used to determine the optimal hybridization conditions which allow specific detection of group A porcine rotaviruses. Probes were prepared by excision of the inserts from the recombinant plasmids and radiolabeling of cDNA with 32P by the random primer extension method. Probes were hybridized at various stringencies with viral RNA from different rotavirus serotypes bound to nylon membranes. Hybridization at low stringency (26% base pair mismatch for stable hybrid formation) had high sensitivity but low specificity. Hybridization at high stringency (16% base pair mismatch for stable hybrid formation) produced high specificity but decreased the sensitivity observed at low stringency. Probes were specific for rotavirus at both stringencies and did not hybridize with nucleic acids from other porcine viruses. Subgenomic gene 9 fragments were then tested to provide more specific probes. A 322 bp fragment from OSU gene 9 between nucleotides 382 and 704 and a 266 bp fragment from Gottfried gene 9 between nucleotides 230 and 496 were found to be specific as hybridization probes. These studies demonstrated the feasibility of the dot blot hybridization assay using subgenomic fragments of gene 9 to detect and differentiate serotypes of porcine rotavirus. Additional studies are warranted to further evaluate the sensitivity and the capability of these probes to detect porcine field isolates of the same serotype.

Animals↗
PubMed1991

Virulence and serum-resistance in different clonal groups and serotypes of Yersinia ruckeri.

The virulence in rainbow trout (Oncorhynchus mykiss) of 32 isolates of Yersinia ruckeri, representing a range of biotypes, serotypes, and OMP-types, was examined. Virulence was assayed in fish of average weight 7.7 g by bath challenge for 1 h with approximately 5 x 10(7) cells per ml. Two of the six serotype O1 clonal groups of Y. ruckeri, clones 2 and 5, were virulent, whereas the other four clonal groups, clones 1, 3, 4 and 6, as well as all serotype O2, O5, O6 and O7 isolates examined, were avirulent. Analysis of susceptibility to the bactericidal effect of non-immune rainbow trout serum demonstrated an association between virulence and serum resistance. The virulent serotype O1 clonal groups were serum resistant, whereas the avirulent serotype O1 clonal groups and other serotypes were, with some exceptions, serum sensitive. The fact that some serum resistant isolates were avirulent suggested that other factors may be required for the full expression of virulence. The study also demonstrated that rainbow trout and brook trout (Salvelinus fontinalis) differ in their susceptibility to Y. ruckeri.

Animals↗
PubMed1994

Evaluation of a saline boiled extract, capsular polysaccharides and long-chain lipopolysaccharides of Actinobacillus pleuropneumoniae serotype 1 as antigens for the serodiagnosis of swine pleuropneumonia.

A saline boiled extract (SBE), capsular polysaccharides (CPS) and long-chain lipopolysaccharides (LC-LPS) of Actinobacillus pleuropneumoniae serotype 1 have been evaluated in ELISA for the serodiagnosis of swine pleuropneumonia caused by this serotype. Mean optical densities (ODs) obtained with the three antigens using sera from negative herds as well as from animals experimentally and naturally exposed to A. pleuropneumoniae serotypes 1, 9 or 11 were not significantly different. The positive ELISA reaction with anti-serotypes 9 and 11 was unexpected with the CPS, which are supposed to be serotype-specific; LPS, and to a lesser extent proteins, were present in the CPS and appeared to be responsible for this reaction. In addition, sera from animals exposed to a field strain of A. pleuropneumoniae serotype 3 and to Actinobacillus suis presented a significantly lower mean OD (P < 0.001) when LC-LPS were used. Cross-reacting antigens consisted mainly of LPS core-lipid A present in the SBE and CPS. The specificity and the sensitivity of the ELISA were evaluated using three different cut-off values (the OD plus two, three and four times the standard deviation or SD) obtained with 667 negative sera. The diagnostic sensitivity was of 81% with the three antigens and the different thresholds. The diagnostic specificity was of 84, 86 and 88% for the mean plus two, three and four times the SD respectively using the SBE and the CPS, while that obtained with the LC-LPS was of 96, 98 and 99% using the same thresholds. In conclusion, LC-LPS make an easily obtainable antigen and seem to retain the best specificity while minimizing losses of sensitivity.

Actinobacillus Infections↗
PubMed2006

Multidrug resistance among different serotypes of clinical Salmonella isolates in Taiwan.

Of the 798 clinical Salmonella isolates collected from multiple hospitals in Taiwan, resistance to ampicillin (48.5%), chloramphenicol (55.3%), streptomycin (59.0%), sulfamethoxazole (68.0%), and tetracycline (67.8%) was high, whereas resistance to all 5 antimicrobials (ACSSuT R-type) comprised 327 (41%) and was highly prevalent in Salmonella enterica serotype Typhimurium (72.7%, 176/242), the most common serotype. Additional resistance to trimethoprim was present in 155 (19.4% overall) of the ACSSuT R-type isolates from several serotypes. Reduced susceptibility to fluoroquinolone (FQ) (ciprofloxacin MIC >0.125-1 microg/mL and nalidixic acid-resistant) was detected in 223 (27.9%) isolates including 117 (14.7% overall) that were also ACSSuT-resistant. Full resistance to FQ was detected in Salmonella Choleraesuis (35.5%, 6/17) and Salmonella Schwarzengrund (16.7%, 10/60); both serotypes were also multiresistant to other antimicrobials. Studies are needed to determine the sources of different multidrug-resistant serotypes. Continued national surveillance is underway to monitor changes in resistance trends and to detect further emergence of resistant Salmonella serotypes in Taiwan.

Anti-Bacterial Agents↗
PubMed2004

Colony blot assay: a useful method to detect multiple pneumococcal serotypes within clinical specimens.

The efficacy of pneumococal conjugate vaccines in young children may be complicated by serotype replacement. We developed a colony blot assay which enables the identification of re-colonization with novel serotypes (replacement), overgrowth by minor co-colonizing serotypes or suppression of previously predominant vaccine serotype strains as a result of vaccination. This method allows the identification of multiple serotypes in a single specimen in a ratio of 1:1000. In order to demonstrate the potential of our method, we investigated the consecutive nasopharyngeal samples of 26 children who had shown a shift in pneumococcal colonization after conjugate vaccination. Mixed colonization was found once in 15 pre-vaccination samples and four times in 26 post-vaccination samples. In the remaining children 'true replacement' had presumably occurred. Hence, we conclude that the colony blot assay is an easy to apply method, which allows the identification of different pneumococcal serotypes within single clinical specimens.

Carrier State↗
PubMed2004

The effect of broth media on pneumococcal growth and the latex serotyping result.

The aim was to test Todd-Hewitt broths (TH-broths) for the ability to propagate pneumococci and thereafter to evaluate the serotyping result obtained by the Pneumotest-Latex kit (SSI). TH-broths from four different producers (Oxoid, Sigma, Difco, and SSI) were tested and compared separately and with Serum broth (SSI). Twenty-three pneumococcal strains (different serotypes) were inoculated into the broths with start inoculums of 10(1), 10(3), and 10(6) CFU/ml. After incubation, overnight viable bacterial counts and visible growth were recorded. All pneumococci were serotyped with the Pneumotest-Latex kit. After incubation, the bacterial counts in all TH-broths were within the range of log 4.65-log 7.76 CFU/ml, while Serum broth showed an average growth ranging from log 8.05-log 8.90 CFU/ml. Comparing the growth of the four TH-broths showed no significant differences. In general, Serum broth had a more pronounced visual growth than each of the four TH-broths. Serotyping with Serum broth showed in general positive and correct latex typing results for all serotypes and initial inoculum, while the outcome of the TH-broths showed some false negative results depending on inoculum and serotype. Overall the Serum broth was found to be superior to the four TH-broths tested both with regard to CFU/ml and when used with the Pneumotest-Latex kit. However, if the Pneumotest-latex kit is only used on broths with visible growth as stated in the instruction manual, then the differences between the performances of the broths from the different producers was not significant and all broths could be used for Pneumotest-Latex typing.

Colony Count, Microbial↗
PubMed2006

Infectious bronchitis virus serotypes in poultry flocks in Jordan.

Infectious bronchitis virus (IBV) causes respiratory disease in chickens all over the world. IBV has many serotypes that do not confer cross protection against each other. Hemagglutination inhibition (HI) test has been used to determine the serotypes of IBV as a substitute to the more laborious virus neutralization test and the more sophisticated restriction endonuclease digestion or sequencing of the S1 gene. In Jordan, no previous studies have been carried out to determine the involvement of IBV in respiratory disease in chickens, or the serotypes of IBV that possibly exist. In this study, serum from different chicken flocks (n=20) that suffered from respiratory disease were tested for IBV antibodies using commercial IBV antibody ELISA at time of the initial signs of the respiratory disease and repeated on serum samples from the same flocks 10-14 days later. ELISA titer for IBV increased in 14 out of 20 flocks (70%) after 10-14 days of the initial signs of the respiratory disease and this indicates a recent exposure to IBV. The second serum samples from these 14 flocks were further examined against a panel of five IBV antigens (Ark, Conn, DE-072, JMK, and Mass) by HI test to determine the serotype(s) of IBV they have been exposed to. The HI test results indicated that the exposure of some of these flocks were to Ark, DE-072, and Mass like serotypes. However, the HI titers against the antigens used in this study were relatively similar in 10 out of the 14 flocks (71%) and the serotype of IBV that these flocks were exposed to could not be determined and the possible causes of this are discussed.

Animals↗
PubMed2004

Serotypes and virulence markers of Shiga toxin-producing Escherichia coli (STEC) isolated from dairy cattle in São Paulo State, Brazil.

In order to determine the occurrence, serotypes and virulence markers of Shiga toxin-producing Escherichia coli (STEC) strains, 153 fecal samples of cattle randomly selected from six dairy farms in Sao Paulo State, Brazil, were examined for Shiga toxin (Stx) production by the Vero cell assay. Feces were directly streaked onto MacConkey Sorbitol Agar and incubated at 37 degrees C overnight. Sorbitol-negative colonies (maximum 20) and up to 10 sorbitol-positive colonies from each plate were subcultured onto presumptive diagnostic medium IAL. Sorbitol-negative isolates were screened with O157 antiserum for identification of O157:H7 E. coli. Isolates presenting cytotoxic activity were submitted to colony hybridization assays with specific DNA probes for stx1, stx2, eae, Ehly and astA genes. The isolation rate of STEC ranged from 3.8 to 84.6% depending on the farm analysed. STEC was identified in 25.5% of the animals, and most of them (64.1%) carried a single STEC serotype. A total of 202 STEC isolates were recovered from the animals, and except for the 2 O157:H7 isolates all the others expressed cytotoxic activity. The great majority of the STEC isolates carried both stx1 and stx2 genes (114/202, 56.4%) or stx2 (82/202, 40.6%); and whereas the Ehly sequence occurred in most of them (88%) eae was only observed in O157:H7 and O111:HNM isolates. Serotypes O113:H21, O178:H19 and O79:H14 were the most frequent STEC serotypes identified and widely distributed among animals from different farms, while others such as O77:H18, O88:H25 and O98:H17 occurred only in particular farms. This is the first report on the occurrence of STEC in dairy cattle in Sao Paulo State, and the results point to substantial differences in rate of isolation, serotypes and genetic profile of STEC that has been previously described among beef cattle in our community. Moreover, to our knowledge O79:H14 and O98:H17 represent new STEC serotypes, while O178:H19 has only been recently reported in Spain.

Adhesins, Bacterial↗
PubMed2005

Comparison of the indirect haemagglutination and gel diffusion test for serotyping Haemophilus parasuis.

The aim of this study was to compare the use of indirect haemagglutination (IHA) and gel diffusion (GD) tests for serotyping Haemophilus parasuis by the Kielstein-Rapp-Gabrielson (KRG) scheme. All 15 serovar reference strains, 72 Australian field isolates, nine Chinese field isolates, and seven isolates from seven experimentally infected pigs were evaluated with both tests. With the IHA test, 14 of the 15 reference strains were correctly serotyped-with serovar 10 failing to give a titre with serovar 10 antiserum. In the GD test, 13 reference strains were correctly serotyped-with antigen from serovars 7 and 8 failing to react with any antiserum. The IHA methodology serotyped a total of 45 of 81 field isolates while the GD methodology serotyped a total of 48 isolates. For 29 isolates, the GD and IHA methods gave discordant results. It was concluded that the IHA is a good additional test for the serotyping of H. parasuis by the KRG scheme if the GD methodology fails to provide a result or shows unusual cross-reactions.

Animals↗
PubMed2005

Biochemical analysis, cpn60 and 16S rDNA sequence data indicate that Streptococcus suis serotypes 32 and 34, isolated from pigs, are Streptococcus orisratti.

Streptococcus suis serotypes have traditionally been identified by morphology, biochemical profiling and serotyping. Analysis of the sequences of 16S rRNA and cpn60 genes of the 35 characterized serotypes of S. suis led to the observation that two serotypes 32 and 34, are significantly distinct from other S. suis serotypes and may represent a distinct species. Here we present DNA sequence data and biochemical profiles which indicate that S. suis serotypes 32 and 34, isolated from pigs, are clustered with Streptococcus orisratti, a Voges-Proskauer negative, alpha-haemolytic, aesculin-hydrolytic, Lancefield group A streptococcus isolated from the teeth of rats.

Animals↗