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Bovine rotavirus serotypes and their significance for immunization.

Neutralization assays on calf fecal rotavirus with antisera to two previously described bovine rotavirus serotypes allowed the isolation of four rotaviruses belonging to a distinct third serotype. In a survey of 85 calf isolates, 80 rotaviruses belonged to serotype 1 (91%), 1 belonged to serotype 2 (1%), and 4 belonged to serotype 3 (5%). Serotypes 1 and 2 were shown to not cross-protect in a passive immunization experiment in gnotobiotic lambs. Ingestion of specific antiserum protected against infection with the homologous, but not heterologous, serotype. Rabbits with no previous exposure to rotavirus responded to sequential vaccination with bovine and human rotavirus serotypes with antibody specific to those serotypes, and the response did not broaden to include serotypes to which they had not been exposed. These factors suggested the need for multivalent rotavirus vaccines. By contrast, 47 adult cows on 11 farms had neutralizing antibodies to two bovine and three human rotavirus serotypes. After vaccination with one bovine rotavirus serotype, these cows produced a significant increase in antibody titers to these same five serotypes but not to two other serotypes to which they had no preexisting antibody. These results were interpreted to indicate that cows will respond heterotypically after monovalent vaccination to all rotavirus serotypes with which they have had experience and, therefore, that single serotype vaccination may be sufficient. This conclusion has practical importance for rotavirus immunization procedures.

Animals↗

Porcine rotaviruses antigenically related to human rotavirus serotypes 1 and 2.

Fecal samples from rotavirus-infected piglets were characterized by a serotyping enzyme-linked immunosorbent assay (ELISA) by using monoclonal antibodies (MAbs) specific to human serotypes 1, 2, 3, and 4 (D. O. Matson, M. K. Estes, J. W. Burns, H. B. Greenberg, K. Taniguchi, and S. Urasawa, submitted for publication). Rotavirus in 19 of 25 specimens tested from two herds of pigs from Buenos Aires province, Argentina, were classified antigenically as follows: one serotype 1, four serotype 2, two serotype 3, and no serotype 4. Six specimens reacted with both serotype 1 and 2 MAbs, and viruses in six specimens probably belonged to other serotypes because they reacted only with a VP7 common epitope MAb. Two porcine rotavirus fecal samples found to contain both serotype 1 and 2 viruses by the MAb-based test and one found to contain a serotype 2 virus were grown in tissue culture. When plaque-purified preparations of these tissue culture-adapted viruses were analyzed in the serotyping ELISA, the C60 and C86 preparations reacted only as serotype 1 viruses, indicating that the original fecal samples, which showed multiple VP7 reactivities, were heterogeneous and apparently contained two types of viruses. Testing of plaque-purified C134 virus confirmed its serotype 2 reactivity. The MAb-based serotype designations of these viruses also were confirmed by using a neutralization immunoperoxidase focus reduction assay. This is the first report of the occurrence of serotype 1 and 2 rotaviruses in animals. The MAbs originally developed to serotype human rotaviruses can be utilized to type animal rotaviruses.

Animals↗

Analysis of loci required for determination of serotype antigenicity in Streptococcus mutans and its clinical utilization.

We recently identified the genes responsible for the serotype c-specific glucose side chain formation of rhamnose-glucose polysaccharide (RGP) in Streptococcus mutans. These genes were located downstream from the rgpA through rgpF locus that is involved in the synthesis of RGP. In the present study, the corresponding chromosomal regions were isolated from serotype e and f strains and characterized. The rgpA through rgpF homologs were well conserved among the three serotypes. By contrast, the regions downstream from the rgpF homolog differed considerably among the three serotypes. Replacement of these regions in the different serotype strains converted their serotypic phenotypes, suggesting that these regions participated in serotype-specific glucose side chain formation in each serotype strain. Based on the differences among the DNA sequences of these regions, a PCR method was developed to determine serotypes. S. mutans was isolated from 198 of 432 preschool children (3 to 4 years old). The serotypes of all but one S. mutans isolate were identified by serotyping PCR. Serotype c predominated (84.8%), serotype e was the next most common (13.3%), and serotype f occured rarely (1.9%) in Japanese preschool children. Caries experience in the group with a mixed infection by multiple serotypes of S. mutans was significantly higher than that in the group with a monoinfection by a single serotype.

Child, Preschool↗

Rapid multiplex assay for serotyping pneumococci with monoclonal and polyclonal antibodies.

We have developed and characterized a rapid semiautomated pneumococcal serotyping system incorporating a pneumococcal lysate preparation protocol and a multiplex serotyping assay. The lysate preparation incorporates a bile solubility test to confirm pneumococcal identification that also enhances assay specificity. The multiplex serotyping assay consists of 24 assays specific for 36 serotypes: serotypes 1, 2, 3, 4, 5, 6A, 6B, 7A/7F, 8, 9L/9N, 9V, 10A/10B/39/(33C), 11A/11D/11F, 12A/12B/12F, 14, 15B/(15C), 17F, 18C, 19A, 19F, 20, 22A/22F, 23F, and 33A/33F. The multiplex assay requires a flow cytometer, two sets of latex particles coated with pneumococcal polysaccharides, and serotype-specific antibodies. Fourteen newly developed monoclonal antibodies specific for common serotypes and a pool of polyclonal rabbit sera for some of the less-common serotypes are used. The two monoclonal antibodies specific for serotypes 18C and 23F recognize serotype-specific epitopes that have not been previously described. These monoclonal antibodies make the identification of the 14 common serotypes invariant. The specificity of the serotyping assay is fully characterized with pneumococci of all known (i.e., 90) serotypes. The assay is sensitive enough to use bacterial lysates diluted 20 fold. Our serotyping system can identify not only all the serotypes in pneumococcal vaccines but also most (>90%) of clinical isolates. This system should be very useful in serotyping clinical isolates for evaluating pneumococcal vaccine efficacy.

Animals↗

Sequential multiplex PCR approach for determining capsular serotypes of Streptococcus pneumoniae isolates.

Accurate serotyping is essential to monitor the changes in the seroepidemiology of Streptococcus pneumoniae. We devised a simple and schematic sequence-based system of seven multiplex PCRs, in a sequence order based upon Active Bacterial Core surveillance (ABCs) serotype distribution during 2002 to 2003, to reliably deduce specific pneumococcal serotypes. A total of 421 isolates from ABCs were randomly chosen to evaluate this system. Two hundred twenty-nine of the isolates (54.3%) were specifically assigned 1 of 17 serotypes by the multiplex PCR system, with the results in complete concordance with conventional serotyping. One hundred seventy-two additional isolates (40.9%) were assigned to 11 specific sets of 2 to 4 serotypes that with one exception (serotypes 6A and 6B) consisted of the single frequently occurring targeted serotype and 1 to 3 additional rare serotypes primarily within the same serogroup as the targeted serotype. Only 20 isolates (4.8%) could not be assigned specific serotypes or serotype sets, since they were either of rare serotypes not included in the assay design or were nonserotypeable. Overall, we found this system to be highly reliable, with the potential to greatly reduce our reliance upon conventional serotyping. Especially important is the capability of this system to give serotype-determining potential to any facility that lacks the expensive typing sera and expertise needed for conventional serotyping yet has the modest equipment necessary for DNA amplification and electrophoresis.

Bacterial Capsules↗

Characteristics and serologic studies of two serotypes of infectious bursal disease virus in turkeys.

Two serotypes of infectious bursal disease virus (IBDV) were distinguished using the virus-neutralization test. An IBDV isolate from chickens, two commercial IBDV vaccine viruses, and an IBDV isolate from turkeys were designated serotype I. Two IBDV isolates from turkeys were designated serotype II. A common antigen shared by serotype I and serotype II viruses was detected using an indirect immunofluorescent assay. The virions of both IBDV serotypes were determined to be nonenveloped icosahedrons with diameters ranging from 58 to 60 nm. The virion densities of one serotype I isolate and two serotype II isolates were similar (1.32 g/ml). Twenty-three turkey breeder flocks and 22 commercial turkey flocks from Ohio, North Carolina, and Indiana were tested for antibodies to the two IBDV serotypes. Seventy percent of the 23 breeder flocks had antibody titers to serotype I, and 75% had antibody titers to serotype II. None of the 22 commercial flocks had antibodies to serotype I, and 77% had antibody titers to serotype II. Antibodies to serotype I were detected only in breeder flocks that were vaccinated using a commercial serotype I IBDV vaccine. Only antibodies to serotype II viruses were naturally widespread in the flocks tested.

Animals↗

Biological and physicochemical characteristics of four serotypes of Salmonella enteritidis.

Four serotypes of Salmonella enteritidis, Anatum ATCC 9270, Newbrunswick ATCC 1608, Oranienburg 200 E, and Pullorum RM, were studied to determine biological, chemical, or physical differences which might explain variations in Salmonella virulence as previously reported by McCullough and Eisele (J. Infect. Dis. 88:278-289, 1951; 89:259-265, 1951). These investigators found that serotype Pullorum was significantly less virulent than serotypes Newport, Derby, Barielly, Meleagridis and Anatum when fed to healthy humans. Results of our own experiments showed that serotype Pullorum RM had a generation time approximately twice that of serotype Anatum 9270. The volume of serotype Pullorum was approximately one-half the volume of the other serotypes used (Anatum 9270, Newbrunswick 1608, Oranienburg 200 E, Cubana 12007, and Meleagridis DR). The number of cells required to yield 1 g dry weight was substantially higher for serotype Pullorum RM than for serotypes Anatum 9270, Newbrunswick 1608, and Oranienburg 200 E. The yield of endotoxin per gram dry weight for serotype Pullorum RM averaged 22 mg/g, whereas yields of endotoxin for serotypes Anatum 9270, Newbrunswick 1608, and Oranienburg 200 E averaged 32 to 35 mg/g. The relative abundance of the four major fatty acids (measured by gas chromatography) also showed distinct differences among the serotypes. Pullorum RM contained less lauric and 3-hydroxymyristic acids and more myristic and palmitic acids than the other three serotypes. The identity of 3-hydroxymyristate was confirmed by mass spectroscopy. Serotype Pullorum RM required 10 times more lipopolysaccharides (endotoxin) to obtain a 50% lethal dose in mice than the other three serotypes. When the lipid part was separated from the polysaccharide and solubilized with bovine serum, the 50% lethal dose of serotype Pullorum RM was equal to that of the other three.

Endotoxins↗

Effects of local immunization with glucosyltransferase fractions from Streptococcus mutans on dental caries in hamsters caused by homologous and heterologous serotypes of Streptococcus mutans.

Seven serotypes of Streptococcus mutans have been identified. The biochemical, genetic, and serological characteristics of these serotypes have indicated that certain serotypes are quite similar, whereas others are quite distinct. The effect of local immunization with glucosyltransferase (GTF) enzymes from serotypes a, c, or g on infection and disease caused by homologous or heterologous cariogenic S. mutans is reported. Organisms with either similar (a and g) or different (c and g) biochemical and serological characteristics were selected for heterologous challenge. NIH white hamsters were injected four times at weekly intervals with GTF prepared by 6 M guanidine-hydrochloride elution from water-insoluble glucan of serotypes a, c, or g, which resulted in enzyme (homologous) inhibitory activity in sera and salivas. After infection of GTF-immunized and sham-immunized groups of hamsters with cariogenic S. mutans of the same serotype as the injected antigen (homologous infection) or with S. mutans of a different serotype from the injected antigen (heterologous infection), the numbers of streptomycin-labeled S. mutans, caries, and lesions were determined. Immunization with GTF preparations from each of the three serotypes resulted in statistically significant reductions in the extent of infection and disease and number of lesions caused by infections with homologous cariogenic S. mutans. Statistically significant reductions in these three parameters were also observed in groups immunized with enzyme from serotype a (strain E49) and challenged with cariogenic serotype g (strain 6715) organisms; or immunized with enzyme from serotype c (strain Ingbritt) and challenged with cariogenic serotype g (strain 6715) organisms; or immunized with enzyme from serotype g (strain 6715) and challenged with cariogenic serotype c (strain Ingbritt) organisms. These studies suggest that soluble antigen preparations containing GTF from one serotype may elicit a protective immune response against infection with cariogenic S. mutans from many or possibly all serotypes.

Animals↗

Identification of a second hemolysin (HlyII) in Actinobacillus pleuropneumoniae serotype 1 and expression of the gene in Escherichia coli.

Hemolysin genes of the reference strains of Actinobacillus pleuropneumoniae serotypes 1 and 2 were identified, cloned, and expressed in Escherichia coli by using polymerase chain reaction amplification with oligonucleotides derived from the DNA sequence of the corresponding appA gene from A. pleuropneumoniae serotype 5. The three genes from serotypes 1, 2, and 5 have identical restriction maps and appear to encode a hemolysin which was previously identified in serotype 2 and designated HlyII. Gene appA is different from hlyIA encoding the major hemolysin type I (HlyI) which was identified earlier in serotype 1. Polymerase chain reaction amplification with oligonucleotides derived from the DNA sequence of hlyIA of serotype 1 showed that the gene encoding HlyI is present in serotype 1 but not in serotype 2, in contrast to the gene encoding HlyII that was present in both serotypes. This was confirmed by Western blot (immunoblot) experiments using monoclonal antibodies specific for either recombinant HlyI or recombinant HlyII, which showed that A. pleuropneumoniae serotype 1 strain 4074 produces both HlyI and HlyII, whereas serotype 2 strain S1536 produces only HlyII. The expression of both hemolysins was investigated in all serotypes by the use of monoclonal antibodies. HlyI was shown to be expressed by the reference strains of serotypes 1, 5a, 5b, 9, 10, and 11, whereas HlyII was shown to be expressed by the reference strains of all 12 serotypes tested except serotype 10. A. pleuropneumoniae serotype 1 strain 4074 is the first bacterium which has been shown to contain two different actively expressed RTX toxin genes. Comparison of our data with those from other groups shows that the originally described strongly hemolytic hemolysin type I (HlyI) corresponds to cytolysin I (ClyI) which was recently described by others, while the weakly hemolytic hemolysin type II (HlyII) seems to be identical to ClyII and AppA.

Actinobacillus pleuropneumoniae↗

Epidemiological study of the G serotype distribution of group A rotaviruses in Kenya from 1991 to 1994.

An epidemiological study on the G serotype distribution of group A rotaviruses (GARV) isolated in Kenya was carried out in one urban hospital in Nairobi and in two rural hospitals in Nanyuki and Kitui to clarify the prevalent G serotypes before future introduction of the ready licensed rotavirus vaccine in Kenya. A total of 1,431 stool specimens were collected from children, who were mainly outpatients, aged from 0 to 6 years old with acute gastroenteritis from August 1991 to July 1994. Samples positive for GARV by conventional ELISA were then analyzed by subgrouping and serotyping ELISA and by PAGE. To ascertain the G serotypes of viruses in samples that were unable to be typed by serotyping ELISA, polymerase chain reaction was also attempted. The prevalence of GARV was 28.4% in the urban hospital, 22.5% in Nanyuki, and 13.7% in Kitui. Among rotavirus-positive samples, subgroup II rotaviruses were detected in 63.1%, and subgroup I rotaviruses were 25.9%. Serotype G4 was most prevalent, accounting for 41.6% followed by 23.3% of serotype G1, 17.0% of serotype G2, and serotype G3 was rarely isolated. Seven strains of serotype G8/P1B rotavirus was detected for the first time in Kenya by RT-PCR. Eleven specimens with an unusual composition of subgroup, serotype, and electropherotype were atypical GARV in which the P-serotype was P1A, P1B, or P2. Although uncommon GARV serotype G8/P1B and atypical GARV were detected, the four major GARV serotypes, G1 through G4, should be targeted at this moment for vaccination to control this diarrheal disease in Kenya. Continuous monitoring of the G- and P-serotype distribution of GARV should provide important information about the impact of rotavirus vaccination in Kenya.

Child↗

Postvaccine genetic structure of Streptococcus pneumoniae serotype 19A from children in the United States.

BACKGROUND: The introduction of the 7-valent conjugate pneumococcal vaccine (PCV7) in children may result in serotype replacement. We estimated the rate of increase of invasive pneumococcal disease (IPD) caused by serotype 19A in children <5 years old and determined the genetic composition of these isolates. METHODS: Cases of IPD between July 1999 and June 2004 were identified through the Active Bacterial Core Surveillance. Serotype 19A isolates obtained from children <5 years old between January 2003 and June 2004 were characterized by serotyping, antibiotic susceptibility testing, and pulsed-field gel electrophoresis (PFGE). Select isolates representing homologous PFGE clusters were subjected to multilocus sequence typing, and eBURST was used to delineate clonal groups. RESULTS: Between July 1999 and June 2004, the overall rate of IPD decreased from 23.3 to 13.1 cases/100,000 population (P<.00001). In children <5 years old, the rate decreased from 88.7 to 22.4 cases/100,000 population (P<.00001), whereas the rate in persons > or =5 years old decreased from 18.4 to 12.4 cases/100,000 population (P<.0001). The rate of serotype 19A IPD in children <5 years old increased significantly from 2.6 cases/100,000 population in 1999-2000 to 6.5 cases/100,000 population in 2003-2004; this was accompanied by significant increases in penicillin nonsusceptibility (P=.008) and multidrug resistance (P=.002) among serotype 19A isolates. As was observed during the pre-PCV7 era, clonal complex (CC) 199 predominated within serotype 19A, representing approximately 70% of invasive serotype 19A isolates from children <5 years old during 2003-2004. New serotype 19A genotypes were observed during 2003-2004, including 6 CCs that were not found among pneumococcal serotype 19A isolates during surveillance in 1999. CONCLUSION: Serotype 19A is, at present, the most important cause of IPD by replacement serotypes, and it is increasingly drug resistant. CC199 is the predominant CC among type 19A serotypes in children <5 years old. Our data suggest that some of the increase in rates of infection with serotype 19A may be due to serotype switching within certain vaccine type strains.

Anti-Bacterial Agents↗

Antibiotic resistance and putative virulence factors of Serratia marcescens with respect to O and K serotypes.

Serratia marcescens serotypes O6:K14, O8:K14 and O28:K28 are common in the natural environment, but rare in hospitals. Serotypes O14:K14 and O27:K14 predominate among clinical strains, but not in the environment, suggesting that the latter serotypes may be more suited for survival in the clinical setting. Consequently, 469 epidemiologically distinct strains of S. marcescens were tested for various putative virulence factors and analysed for associations with serotype. The factors positively associated with serotype O14:K14 were agglutination of five different species of red blood cells and expression of type 1 fimbriae. These were found in 63% and 53% of O14:K14 strains, respectively, compared with 7% and 12% of the three 'environmental serotypes'. Almost a quarter of the collection expressed the mannose-resistant haemagglutinin indicative of type 3 fimbriae, but this was not associated with any serotype. The production of DNAase, haemolysin, lipase, lecithinase, proteases and siderophores was almost universal and showed no serotype correlations. Almost half of the strains (46%) were resistant to serum and serotypes O27:K14 and O6:K14 were strongly associated with this characteristic. Serotype O27:K14 was also associated with higher proportions of antibiotic-resistant strains than other serotypes, but the same was not true of serotype O14:K14. All three 'environmental serotypes' were associated with low frequencies of antibiotic resistance; <12% were resistant to gentamicin, carbenicillin or piperacillin, or any combination of these three, compared with 20-25% of O14:K14 strains and >42-51% of O27:K14 strains. Pigment production was strongly associated with serotype. None of the O14:K14 or O27:K14 strains produced prodigiosin, but frequencies for the three 'environmental serotypes' ranged from 31% of O28:K28 strains to 85% of O6:K14 strains. The results of this study suggest that the adherence capability of S. marcescens strains may play a role in the colonisation of hospital patients, while the production of prodigiosin is a marker of environmental origin.

Animals↗

Distribution of serotypes, IS901 and a 40 kDa protein in Mycobacterium avium complex strains isolated from man and animals in Denmark.

The aim of the present study was to characterize all strains of the Mycobacterium avium complex isolated in Denmark in 1993. A total of 141 M. avium complex strains (86 from man, 38 from animals, and 17 from peat) were analysed by serotyping, ELISA specific for a 40 kDa protein, and IS901-specific PCR. Serotype analysis showed that the most frequent serotypes among human strains were serotype 4 (27%) and serotype 6 (19%), which differs from an earlier survey where serotype 1 was most prevalent. The most frequent serotypes in animals were serotype 2 (53%) and serotype 6 (13%), whereas the most prevalent serotypes among strains isolated from peat were serotype 4 (29%) and serotype 9 (18%). There was a concurrent appearance of IS901 and p40 in all strains. Only M. avium complex strains isolated from animals, and belonging to serotype 1 or serotype 2, contained the IS901/p40 markers. The different distribution of serotypes of M. avium complex strains in animals and man, and the presence of IS901/p40 exclusively in animal strains, suggests that transmission of M. avium from animals to man is not of significance in Denmark.

Animals↗

[Comparison of O-serotype distribution of Escherichia coli isolated from faecal specimens of patients with sporadic diarrhea and healthy persons and regional difference in Japan].

To investigate the isolation frequency of O-serotype of Escherichia coli, a total of 1,563 faecal specimens obtained from patients with sporadic diarrhea in Ishikawa between July 1997 and June 1998, were examined. As a result of O-serotyping of isolated strains using commercially E. coli antisera (43 different types), 247 strains of 29 different O-serotypes were isolated. Isolation rate was 15.8%. Most predominant O-serotype was O1 (128 strains, 52%), followed by O18 (26 strains, 11%), O6 (17 strains, 7%), O111 (16 strains, 6%), and these 4 different O-serotypes took up three quarters of the isolated E. coli. Between August 1996 and May 1997, E. coli isolation from faecal samples of 51,893 healthy persons and O-serotyping of isolated strains using commercial antisera to 6 predominant O-serotypes (O-26, 111, 114, 128, 157 and O1) of VTEC/EHEC were carried out. Among 6 O-serotypes, the most predominant O-serotype was O1 (93% of isolates), followed by O26, 111, 128 (6%) and O114, 157 (1%). These isolation frequencies in patients were 80%, 18%, 2%, respectively, have resembled each other in healthy persons in many points. In a similar way, of these distributions of O-serotype of strains hemolysed on Beutin's blood agar plates, we compared patients with healthy persons. Fifty-six strains (3.6% of the total) of E. coli of different O-serotypes were isolated from 1,563 patients and 57 strains (2.8% of the total) belonging to 11 serotypes from 2,036 healthy persons. As a result of O-serotype frequency, both groups resembled each other. O18 and O6, the most predominant O-serotypes, occupied 64% of the isolated strains in patients and 74% in healthy persons. Next in patients, O1, 26 were 7%--level, O28 ac, 152, 157 were 4%--level, respectively, and in healthy persons. O1 was 5%--level, O28 ac, 55, 146, 152 were 4%--level respectively. In the comparison of O-serotype frequency of E. coli isolated from sporadic diarrhea in other 5 areas (Kanto district, Tokyo, Oita, Aichi and Ishikawa), O1, 6, 8, 18, 25, 26, 55, 86a, 111, 125, 126, 127a, 128, 146, 148, 157 and 166 (17 types) have covered a wide area. On the other hand, O29, 44, 78, 112ac, 115, 136, 143, 152, 168 and 169 (10 types) have a tendency to distribute in local areas, we believe that there are regional differences even in the same Japanese territory.

Diarrhea↗

Prevalence of neutralizing antibodies against different rotavirus serotypes in children with severe rotavirus-induced diarrhea and their mothers.

Neutralizing antibody (NAb) responses to different rotavirus serotypes were compared in 64 convalescent-phase serum samples from hospitalized rotavirus-positive children less than 2 years of age and their mothers. Compared to the child patients, the mothers showed significantly higher NAb positivity to animal rotavirus serotypes G3 simian (96.88%), G6 bovine (85.94%), and G10 bovine (25.0%) and to human rotavirus serotypes G8 (79.69%) and G3 (57.81%) (P < 0.01 for each) but not to human serotypes G1, G2, G4, and G9 (P > 0.05). The overall prevalence of NAb among the child patients was low for human rotavirus serotypes G1 (20.31%) and G3 (21.8%). The comparative NAb response in individual mother-child paired serum samples was analyzed against each rotavirus serotype. A substantial number of child patients showed higher NAb titers than their mothers to serotypes G1, G2, G4, and G9, indicating that these serotypes are the major serotypes causing rotavirus diarrhea among the children of Pune, India. In these cases, the mothers were either negative or had lower titers of NAbs than their children. Correlation was observed between the infecting serotype and child patient serum that showed a homologous NAb response at a higher level than that of the mother. It appears that when the level of NAb to a particular serotype is higher among child patients than among their mothers, that serotype is the infecting serotype, and that low titers of NAb among the mothers predispose the children to infection with that serotype, if the serotype is in circulation.

Adult↗

Structural and genetic analyses of O polysaccharide from Actinobacillus actinomycetemcomitans serotype f.

The oral bacterium Actinobacillus actinomycetemcomitans is implicated as a causative agent of localized juvenile periodontitis (LJP). A. actinomycetemcomitans is classified into five serotypes (a to e) corresponding to five structurally and antigenically distinct O polysaccharide (O-PS) components of their respective lipopolysaccharide molecules. Serotype b has been reported to be the dominant serotype isolated from LJP patients. We determined the lipopolysaccharide O-PS structure from A. actinomycetemcomitans CU1000, a strain isolated from a 13-year-old African-American female with LJP which had previously been classified as serotype b. The O-PS of strain CU1000 consisted of a trisaccharide repeating unit composed of L-rhamnose and 2-acetamido-2-deoxy-D-galactose (molar ratio, 2:1) with the structure -->2)-alpha-L-Rhap-(1-3)-2-O-(beta-D-GalpNAc)-alpha-L-Rhap-(1-->* O-PS from strain CU1000 was structurally and antigenically distinct from the O-PS molecules of the five known A. actinomycetemcomitans serotypes. Strain CU1000 was mutagenized with transposon IS903phikan, and three mutants that were deficient in O-PS synthesis were isolated. All three transposon insertions mapped to a single 1-kb region on the chromosome. The DNA sequence of a 13.1-kb region surrounding these transposon insertions contained a cluster of 14 open reading frames that was homologous to gene clusters responsible for the synthesis of A. actinomycetemcomitans serotype b, c, and e O-PS antigens. The CU1000 gene cluster contained two genes that were not present in serotype-specific O-PS antigen clusters of the other five known A. actinomycetemcomitans serotypes. These data indicate that strain CU1000 should be assigned to a new A. actinomycetemcomitans serotype, designated serotype f. A PCR assay using serotype-specific PCR primers showed that 3 out of 20 LJP patients surveyed (15%) harbored A. actinomycetemcomitans strains carrying the serotype f gene cluster. The finding of an A. actinomycetemcomitans serotype showing serological cross-reactivity with anti-serotype b-specific antiserum suggests that a reevaluation of strains previously classified as serotype b may be warranted.

Actinobacillus Infections↗

Adherence to, invasion by, and cytokine production in response to serotype VIII group B Streptococci.

The adherence to and invasion of the human epithelial cell line A549 by group B streptococcus (GBS) serotype VIII strains were compared with those of serotype III strains by a conventional method and the dynamic in vitro attachment and invasion system. Twenty GBS strains, including nine vaginal isolates and one invasive isolate each of serotypes III and VIII, were used in the conventional attachment and invasion assay. Adherence to and invasion of A549 cells by serotype VIII GBS strains were significantly greater (P < 0.0001) than those by serotype III strains for both the invasive strain and vaginal isolates. Cytokine production by A549 cells following stimulation with GBS serotypes III and VIII or their purified capsular polysaccharides (CPS) was measured. Serotype III strains stimulated significantly greater tumor necrosis factor alpha (TNF-alpha) (P < 0.0001) and interleukin-10 (IL-10) (P < 0.05) production than did serotype VIII strains. IL-8 production in response to serotype VIII was significantly higher (P < 0.001) than that in response to serotype III. TNF-alpha, IL-8, and IL-10 production was greater in A549 cells infected with GBS than in the untreated control cells. TNF-alpha production was significantly greater (P < 0.005) after stimulation with purified GBS serotype III CPS than after stimulation with serotype VIII CPS, a result similar to that after stimulation with whole GBS. IL-12 production by A549 cells was observed only in response to infection with GBS serotype III, resulting in the possibility of a greater TH1 response in serotype III GBS. These results suggest differences in immune responses to infection with GBS serotypes III and VIII.

Bacterial Adhesion↗

Serotypes of Ureaplasma urealyticum isolated from normal pregnant women and patients with pregnancy complications.

To compare the distribution of Ureaplasma urealyticum serotypes 1 to 10 in different patient populations, the serotypes of 240 U. urealyticum strains from 207 patients were determined by the indirect immunofluorescence test by using U. urealyticum antisera 1 to 10. Strains were obtained from the following four patient groups: group 1, 24 couples in which the women had a history of recurrent spontaneous abortion; group 2, 25 patients who had their first spontaneous abortion; group 3, 14 pregnant patients with pregnancy complications (premature delivery, intrauterine death); and group 4, 138 patients with uneventful pregnancies. The serotypes most often found in these 207 patients were as follows: serotype 3, 52.2%; serotype 6, 30.3%; serotype 10, 11.4%; serotype 1, 9.5%; serotype 4, 6.5%; serotype 8, 6.5%. Serotypes 2, 5, 7, and 9 were found in less than 1% of the patients. More than one serotype was found in 16.9% of the patients. The overall distribution of the 10 serotypes in the different groups was similar, except for that of serotype 4. Serotype 4 was isolated from 20.8% of the patients in group 1 and from 5.1% of the patients in group 4 (P less than 0.01). Results of this study indicate that U. urealyticum serotype 4 can be isolated more frequently from patients with a history of recurrent miscarriages than from normal pregnant women.

Abortion, Habitual↗