Identification of epidemic strains of Streptococcus suis by genomic fingerprinting.

A natural outbreak of Streptococcus suis meningitis in two closed swine herds was studied. DNA fingerprinting, serotyping, and biochemical profiles were assessed. Multiple serotypes were recovered from these herds. In farm A, 50 S. suis strains were isolated from 330 swabs collected. Eighteen strains belonged to serotype 2, and 32 strains belonged to serotypes 3, 5, 6, 8, 9, and 11. In farm B, 16 S. suis strains were recovered from a total of 70 samples. Eight strains belonged to serotype 7 and eight belonged to serotypes 2, 3, 5, and 8. In each epidemiological situation, a single strain characterized by a distinctive restriction fragment pattern predominated among affected penmates. The epidemic serotype 2 strain was detected in farm A in weaned pigs between the ages of 5 and 7 weeks. In contrast, the pathogenic strain in farm B belonged to serotype 7 and was isolated from pigs up to 3 weeks of age. The results from both farms strongly suggest a lateral spread of these organisms. No vertical transmission could be shown in either herd. It was concluded that genomic fingerprinting is an appropriate method to distinguish outbreak isolates of S. suis from nonoutbreak strains, within the same serotype or from epidemiologically unrelated clusters of strains.     

Clonal analysis of the Actinobacillus pleuropneumoniae population in a geographically restricted area by multilocus enzyme electrophoresis.

The genetic diversity among 250 isolates of Actinobacillus pleuropneumoniae from lungs of pigs with pleuropneumonia and from tonsils of apparently healthy pigs at slaughter was estimated by multilocus enzyme electrophoresis. The Danish strains were derived from both specific-pathogen-free and conventional herds. Sixty-six percent of the isolates belonged to three electrophoretic types (ETs) of a total of 37 ETs detected. While five biotype 2 isolates constituted a separate ET closely related to biotype 1 isolates, the type strain of the species (Shope 4074) belonged to its own ET, with a genetic distance of 0.30 from its nearest neighbor. Isolates of serotypes traditionally considered to have less pathogenic potential (serotypes 6, 10, and 12) from herds with acute outbreaks of pleuropneumonia belonged to the same ETs as isolates from apparently healthy pigs, suggesting that factors such as cross immunity and management may lead to divergent clinical results. Isolates from four herds harboring more than one serotype showed distinct profiles between the serotypes, indicating no or only limited chromosomal recombination among clones. Isolates from tonsils belonged to the same ET as isolates from lungs. The same ET was isolated from widely different parts of the world. Evidence from this study indicates that multilocus enzyme electrophoresis may be a valuable tool for the epidemiological analysis of A. pleuropneumoniae.     

Clonal diversity in Haemophilus pleuropneumoniae.

Genetic diversity among 135 isolates of nine serotypes of Haemophilus pleuropneumoniae recovered from pigs with pleuropneumonia or other invasive diseases in 14 countries was estimated by multilocus enzyme electrophoresis, which detects allelic variation in structural genes. Thirty-two multilocus genotypes (electrophoretic types [ETs]) were distinguished on the basis of allele profiles at 15 enzyme loci, and 36 distinctive combinations of ET and serotype were identified. The recovery of isolates with identical properties in widely separated geographic regions and over a 20-year period indicated that the population structure of H. pleuropneumoniae is clonal. Isolates of the same ET generally shared the same serotype and electrophoretic pattern of the outer membrane proteins, but some ETs were represented by isolates of several different serotypes, outer membrane protein patterns, or both. On average, the genetic diversity among ETs of the same serotype was 56% of the total genetic diversity in the species. Isolates of serotype 1, which are unusually pathogenic, belong to a distinctive group of clones that are closely related to clones marked by serotype 9.     

Characterization of prototype and clinically defined strains of Streptococcus suis by genomic fingerprinting.

A collection of Streptococcus suis strains from animal and human infections was examined for DNA-banding patterns after restriction endonuclease digestion and agarose gel electrophoresis. The endonuclease HaeIII produced the most discriminating restriction profiles among 23 serotypes studied. DNA from serotypes 9, 11, 12, and 16 was resistant to HaeIII cleavage. DNA from serotypes 9 through 16 was cleaved with HindIII and showed substantial genomic differences. We also examined 106 epidemiologically unrelated strains isolated from cases of pig meningitis or pneumonia and 5 strains isolated from cases of human meningitis in order to compare genomic fingerprinting and serotyping as epidemiological tools. Heterogeneity was found among fingerprints of serologically identical isolates, indicating genetic diversity within some serotypes. DNA fingerprints of some serotype 2 strains from different sources appeared identical, suggesting a clonal relationship among strains of this serotype. The data suggest that this technique represents an important tool for examining the natural history of disease caused by S. suis.     

Analysis of genetic diversity of Streptococcus suis clinical isolates from pigs in Spain by pulsed-field gel electrophoresis

Pulsed-field gel electrophoresis (PFGE) was used to investigate the diversity of Streptococcus suis isolates of various serotypes recovered from swine clinical samples in Spain. Capsular types 9 (64.9%) and 2 (14.8%) were the most frequently isolated serotypes followed by serotype 7 (5.9%) and serotype 8 (4.3%). The PFGE results of this study with 60 different pulsotypes indicate a great genetic diversity among the S. suis isolates, which is consistent with the broad distribution of S. suis in the swine population. Forty-five percent of the pulsotypes corresponded to single isolates, no pulsotype was common to all farms, and at least 3 different pulsotypes were isolated in 56% of herds in which more than 3 clinical isolates were analyzed. These results reveal a great diversity both between and within herds throughout the strains of S. suis studied, demonstrating that different strains of S. suis are associated with infection in pigs. Some pulsotypes were more frequently isolated and exhibited a wider distribution over herds than others, and were the unique or predominant strains in several herds, suggesting the existence of a prevalent or a few prevalent clones responsible for a large proportion of clinical cases. Overall, the great genetic heterogeneity of the clinical strains of S. suis, the isolation of different strains within the same herd, and the predominance of particular strains in some herds are evidence that infection by S. suis is a dynamic process and reinforce the idea that the epidemiology of S. suis infection is very complex.     

Relatedness of Streptococcus suis isolates of various serotypes and clinical backgrounds as evaluated by macrorestriction analysis and expression of potential virulence traits

We evaluated the genetic diversity of Streptococcus suis isolates of different serotypes by macrorestriction analysis and elucidated possible relationships between the genetic background, expression of potential virulence traits, and source of isolation. Virulence traits included expression of serotype-specific polysaccharides, muramidase-released protein (MRP), extracellular protein factor (EF), hemolysin activity, and adherence to epithelial cells. Macrorestriction analysis of streptococcal DNA digested with restriction enzymes SmaI and ApaI allowed differentiation of single isolates that could be assigned to four major clusters, named A1, A2, B1, and B2. Comparison of the genotypic and phenotypic features of the isolates with their source of isolation showed that (i) the S. suis population examined, which originated mainly from German pigs, exhibited a genetic diversity and phenotypic patterns comparable to those found for isolates from other European countries; (ii) certain phenotypic features, such as the presence of capsular antigens of serotypes 2, 1, and 9, expression of MRP and EF, and hemolysin activity (and in particular, combinations of these features), were strongly associated with the clinical background of meningitis and septicemia; and (iii) isolates from pigs with meningitis and septicemia showed a significantly higher degree of genetic homogeneity compared to that for isolates from pigs with pneumonia and healthy pigs. Since the former isolates are considered highly virulent, this supports the theory of a clonal relationship among highly virulent strains.     

Development of a multilocus sequence typing scheme for the pig pathogen Streptococcus suis: identification of virulent clones and potential capsular serotype exchange

Streptococcus suis is an important pathogen of pigs and occasionally causes serious human disease. However, little is known about the S. suis population structure, the clonal relationships between strains, the potential of particular clones to cause disease, and the relevance of serotype as a marker for epidemiology. Here we describe a multilocus sequence typing (MLST) scheme for S. suis developed in order to begin to address these issues. Seven housekeeping gene fragments from each of 294 S. suis isolates obtained from various S. suis diseases and from asymptomatic carriage representing 28 serotypes and nine distinct countries of origin were sequenced. Between 32 and 46 alleles per locus were identified, giving the ability to distinguish >1.6 x 10(11) sequence types (STs). However only 92 STs were identified in this study. Of the 92 STs 18 contained multiple isolates, the most common of which, ST1, was identified on 141 occasions from six countries. Assignment of the STs to lineages resulted in 37 being identified as unique and unrelated STs while the remaining 55 were assigned to 10 complexes. ST complexes ST1, ST27, and ST87 dominate the population; while the ST1 complex was strongly associated with isolates from septicemia, meningitis, and arthritis, the ST87 and ST27 complexes were found to contain significantly higher numbers of lung isolates. In agreement with the observed distribution of disease-causing isolates of S. suis, most isolates previously characterized as of high virulence in porcine infection models belong to ST1, while isolates belonging to other STs appear to be less virulent in general. Finally nine STs were found to contain isolates of multiple serotypes, and many isolates belonging to the same serotypes were found to have very disparate genetic backgrounds. As well as highlighting that the serotype can often be a poor indicator of genetic relatedness between S. suis isolates, these findings suggest that capsular genes may be moving horizontally through the S. suis population.     

Characterization of Streptococcus agalactiae strains by multilocus enzyme genotype and serotype: identification of multiple virulent clone families that cause invasive neonatal disease.

The chromosomal genotypes of 277 isolates of 16 serotypes of Streptococcus agalactiae were characterized by analysis of electrophoretically demonstrable allele profiles at 12 metabolic enzyme loci. The collection comprised the type strain and 276 strains recovered from French symptomatic and asymptomatic subjects. Sixty-one distinctive electrophoretic types (ETs), representing multilocus clonal genotypes, were identified. Cluster analysis of the ETs revealed two primary phylogenetic divisions separated by a genetic distance of 0.62, Division I contained 67 isolates which could be assigned to 13 ETs. Twenty-seven of these isolates were from samples of cerebrospinal fluid (CSF) from neonatal meningitis patients. Two ETs, separated by a genetic distance of 0.217, contained 26 of these 27 isolates. Division II contained 210 isolates, of which 27 were isolated from CSF. This division was more polymorphic and included 48 ETs. Spanning a genetic distance of 0.3, three clusters and one ET were identified within this group. Twenty-four of 27 strains isolated from CSF belonged to one cluster, and 19 of them belonged to two adjacent ETs with a genetic distance of 0.083. Fifty-five of the 68 serotype Ia strains and 24 of the 26 serotype Ib strains were each confined to one of the evolutionary lineages, and 85 of the 86 strains which carried protein antigen c belonged to phylogenetic division II. Most of the type III organisms were assigned to two clone families. The characteristics of this French population argue for the existence of particular groups of strains responsible for neonatal meningitis and demonstrate that serotyping can supply information about the genetic distribution of strains.     

Virulent strains of Streptococcus suis serotype 2 and highly virulent strains of Streptococcus suis serotype 1 can be recognized by a unique ribotype profile.

The ribotype profiles of 42 different Streptococcus suis strains were studied. These strains belonged to five serotypes and differed in their virulence for pigs as well as in the expression of the muramidase-released protein and the extracellular protein factor. For the ribotyping, chromosomal DNAs were digested with EcoRI and were hybridized with a 1,066-bp ribosomal DNA probe. The hybridization patterns showed genetic heterogeneity within and between the serotypes. Pathogenic strains of serotype 2 and highly pathogenic strains of serotype 1 could be recognized by their unique ribotype profiles. Nonpathogenic strains showed a high degree of genetic heterogeneity. Moreover, by comparing the 16S ribosomal DNA sequences of a number of S. suis strains, we were able to design two DNA probes which specifically hybridized with S. suis strains.     

Analysis of Erysipelothrix rhusiopathiae and Erysipelothrix tonsillarum by multilocus enzyme electrophoresis.

The genetic diversity of 74 Australian field isolates of Erysipelothrix rhusiopathiae and 22 reference strains for serovars of E. rhusiopathiae or Erysipelothrix tonsillarum was examined by multilocus enzyme electrophoresis. Four serovar reference strains of E. tonsillarum (strains KS 20 A, Wittling, Lengyel-P, and Bano 107 for serovars 25, 3, 10, and 22, respectively) were genetically distinct from E. rhusiopathiae. However, the E. tonsillarum reference strain for serovar 14 (Iszap-4) and the reference strain for serovar 13 (Pecs-56), which has been said to represent a new genomic species, were found to cluster with typical isolates and reference strains of E. rhusiopathiae. Our reference strain for serovar 7 (Rotzunge) was also genetically typical of E. rhusiopathiae, thus indicating that these serotype reactivities cannot be relied upon as a means of identifying isolates as E. tonsillarum. Australian field isolates of E. rhusiopathiae were genetically diverse. Those recovered from sheep or birds were more diverse than those isolated from pigs, and isolates of serovar 1 were more diverse than those of serovar 2. The diversity found among isolates of the same serovar and the presence of isolates of different serovars in the same electrophoretic types (ETs) indicated that serotyping of E. rhusiopathiae was unreliable for use as an epidemiological tool. Some ETs contained isolates recovered from different animal species. ET 41 contained 32.2% of the field isolates and two reference strains, indicating that this clone of E. rhusiopathiae is both widespread and commonly associated with disease in various species of animals.     

Molecular characterization of Escherichia coli strains isolated from pigs with edema disease.

The present study was conducted to investigate the epidemiological relationship of isolates of Escherichia coli causing edema disease. Classical edema disease has not previously been described in Denmark, but between February 1994 and November 1995 cases appeared in 51 pig herds, among which direct or indirect trading contacts were confirmed for 36 of the herds. A total of 213 isolates from pigs with edema disease in Denmark and other countries and 23 E. coli O139 isolates from pigs with diarrhea or healthy pigs were analyzed to characterize their O serogroups, HindIII ribotypes, and pulsed-field gel electrophoresis (PFGE) types, and 183 of the isolates were also analyzed for their plasmid profiles. The resulting PFGE types of the isolates from pigs with edema disease were examined by cluster analysis. Ten isolates from three herds could not be typed with the available O antisera, whereas all other isolates were of serotype O139. However, all isolates from pigs with edema disease belonged to the same HindIII ribotype, which was not observed among the isolates from pigs with diarrhea or healthy pigs. All isolates from Danish pigs with edema disease except for three isolates originating from two herds belonged to the same or closely related XbaI PFGE types; the other three isolates were assigned to possibly related types. Isolates from pigs with edema disease in different countries belonged to different PFGE types. All isolates from Danish pigs with edema disease grouped together in one cluster, in contrast to isolates from other countries, which did not form any clusters. E. coli strains of serogroup O139 from pigs with diarrhea or isolated from the feces of healthy Danish pigs were very different. Plasmid profiles differed largely among isolates. However, among the isolates from Danish pigs with edema disease, one type predominated within herds. The present study indicated that most, if not all, of the observed cases of edema disease in Denmark were part of the same outbreak. The combination of PFGE typing and ribotyping was useful for studying the possible clonal relationship among strains, whereas plasmid profiling was less informative.     

Genetic relationships and clonal population structure of serotype 2 strains of Neisseria meningitidis.

Two hundred and thirty-four strains of Neisseria meningitidis, including 94 serotype 2a, 111 serotype 2b, and 19 serotype 2c isolates, together with 10 isolates that were serotyped as 2 with polyvalent antiserum but did not react with monoclonal antibodies, were characterized by the electrophoretic mobilities of 15 metabolic enzymes. Of these enzymes, 14 were polymorphic, and 56 distinctive combinations of alleles at the enzyme loci (electrophoretic types) were identified, among which the mean genetic diversity per locus was 0.413, or about 75% of that recorded for the species N. meningitidis as a whole. Mean genetic diversity among electrophoretic types of the same serotype (2a, 2b, or 2c) was, however, on average, less than half the total species diversity, and no multilocus genotypes were shared between isolates of the different serotypes, which belong to distinctive clonal lineages. Recent temporal changes in the frequencies of recovery of pathogenic strains of serotypes 2a and 2b in South Africa and North America resulted from clone replacement in these populations rather than evolutionary modification of the serotype protein of the initially dominant clones.     

Genetic relatedness within serotypes of penicillin-susceptible Streptococcus pneumoniae isolates

The molecular epidemiological characteristics of all Streptococcus pneumoniae strains isolated in a nationwide manner from patients with meningitis in The Netherlands in 1994 were investigated. Restriction fragment end labeling analysis demonstrated 52% genetic clustering among these penicillin-susceptible strains, a value substantially lower than the percentage of clustering among Dutch penicillin-nonsusceptible strains. Different serotypes were found within 8 of the 28 genetic clusters, suggesting that horizontal transfer of capsular genes is common among penicillin-susceptible strains. The degree of genetic clustering was much higher among serotype 3, 7F, 9V, and 14 isolates than among isolates of other serotypes, i.e., 6A, 6B, 18C, 19F, and 23F. We further studied the molecular epidemiological characteristics of pneumococci of serotype 3, which is considered the most virulent serotype and which is commonly associated with invasive disease in adults. Fifty epidemiologically unrelated penicillin-susceptible serotype 3 invasive isolates originating from the United States (n = 27), Thailand (n = 9), The Netherlands (n = 8), and Denmark (n = 6) were analyzed. The vast majority of the serotype 3 isolates (74%) belonged to two genetically distinct clades that were observed in the United States, Denmark, and The Netherlands. These data indicate that two serotype 3 clones have been independently disseminated in an international manner. Seven serotype 3 isolates were less than 85% genetically related to the other serotype 3 isolates. Our observations suggest that the latter isolates originated from horizontal transfer of the capsular type 3 gene locus to other pneumococcal genotypes. In conclusion, epidemiologically unrelated serotype 3 isolates were genetically more related than those of other serotypes. This observation suggests that serotype 3 has evolved only recently or has remained unchanged over long periods.     

Serotypes and clonal types of penicillin-susceptible streptococcus pneumoniae causing invasive disease in children in five Latin American countries

We used multilocus sequencing typing (MLST) to determine the genetic backgrounds of 185 recent penicillin susceptible Streptococcus pneumoniae isolates with serotypes that most frequently cause invasive disease in preschool age children in five Latin American countries-Argentina, Brazil, Colombia, Mexico, and Uruguay. Most of the isolates were associated with pneumonia (90/185), meningitis (74/185), and bacteremia (17/185). The collection of strains included seven serotypes-14, 6B, 5, 1, 23 F-which represent the serotypes of S. pneumoniae most frequently associated with sterile site infections in children. Also included were strains expressing serotypes 7F and 3. Comparison of serotype and multilocus sequence type allowed division of the isolates into two groups: strains expressing serotypes 1, 5, 3, and 7 were represented by a relatively few sequence types while strains expressing serotypes 6B, 14, and 23 F showed great genetic diversity. The genetic diversity of serotypes 14, 6B, and 23 F may be related to the capacity of these serotypes to colonize the nasopharynx of healthy carriers during which opportunities for diversification through genetic exchanges can occur. The findings present an interesting contrast with the results of an earlier study in which over 80% of invasive penicillin- resistant serotype 14 and 23 isolates from the same countries were found to belong to as few as two pandemic clones of S. pneumoniae.     

Multiplex PCR assay for detection of Streptococcus suis species and serotypes 2 and 1/2 in tonsils of live and dead pigs

A PCR assay was developed for the detection of Streptococcus suis serotypes 2 and 1/2. This multiplex PCR is based on the amplification of the gene coding for 16S rRNA of S. suis and on the amplification of the cps2J gene coding for the capsule of S. suis serotypes 2 and 1/2. An internal control was constructed and added in this test to monitor the efficiency of amplification in each reaction. To evaluate the specificity of the test, 31 strains of other bacterial species related to S. suis or isolated from pigs and 42 strains of S. suis serotypes 1 and 3 to 34 were analyzed. The detection threshold of the test was 28 S. suis CFU/ml. The specificity and the sensitivity of the multiplex PCR test and the presence of an internal control allowed the analysis of biological samples without a culture step. The PCR assay was then applied to the detection of 14 S. suis serotype 1/2 strains, 88 S. suis serotype 2 strains isolated from pigs, and 25 S. suis serotype 2 strains isolated from humans. This test was also applied to analyze tonsil samples of pigs experimentally infected and carrier pigs without any symptoms.     

Distribution and Genetic Diversity of Suilysin in Streptococcus suis Isolated from Different Diseases of Pigs and Characterization of the Genetic Basis of Suilysin Absence

Streptococcus suis is an economically important pathogen of pigs responsible for a variety of diseases including meningitis, septicemia, arthritis, and pneumonia, although little is known about the mechanisms of pathogenesis or virulence factors associated with this organism. Here, we report on the distribution and genetic diversity of the putative virulence factor suilysin, a member of the thiol-activated toxin family of gram-positive bacteria. On the basis of PCR analysis of over 300 isolates of S. suis, the suilysin-encoding gene, sly, was detected in 69.4% of isolates. However, sly was present in a considerably higher proportion of isolates obtained from cases of meningitis, septicemia, and arthritis (>80%) and isolates obtained from asymptomatic tonsillar carriage (>90%) than lung isolates associated with pneumonia (44%). With the exception of serotypes 1, 14, and 1/14, there was no strong correlation between the presence of suilysin and serotype. Analysis of the genetic diversity of suilysin by restriction fragment length polymorphism and sequence analysis found that the suilysin gene, where present, is highly conserved with a maximum of 1.79% diversity at the nucleotide level seen between sly alleles. Assays of hemolytic activity and hybridization analysis provided no evidence for a second member of the thiol-activated toxin family in S. suis. Inverse PCR was used to characterize regions flanking sly, which in turn allowed the first characterization of the equivalent region in a strain lacking sly. Sequence comparison of these regions from sly-positive (P1/7) and sly-negative (DH5) strains indicated that two alternative arrangements are both flanked by genes with highest similarity to haloacid dehalogenase-like hydrolases (5' end) and putative N-acetylmannosamine-6-phosphate epimerases (3' end). However, sly appears to be completely absent from the alternative arrangement, and a gene of unknown function is located in the equivalent position. Finally, PCR analysis of multiple sly-positive and -negative strains indicated that these two alternative genetic arrangements are conserved among many S. suis isolates.     

Persistence of two invasive Streptococcus pneumoniae clones of serotypes 1 and 5 in comparison to that of multiple clones of serotypes 6B and 23F among children in southern Israel

We conducted a study to examine the clonal distribution of invasive serotype 1 and 5 isolates as representatives of serotypes that are rarely carried by healthy individuals compared to that of invasive serotype 6B and 23F isolates as representatives of serotypes often carried by young children for prolonged periods. All invasive serotype 1, 5, 6B, and 23F isolates recovered from blood cultures during January 1995 to May 1999 were analyzed; these included 66 serotype 1, 30 serotype 5, 11 serotype 6B, and 15 serotype 23F isolates. One hundred thirty-three nasopharyngeal (NP) isolates of the indicated four serotypes from healthy children were also studied. The strains were characterized using serotyping, antimicrobial susceptibility testing, and pulsed-field gel electrophoresis profiling. We found that both invasive and NP serotype 1 and 5 isolates were susceptible to penicillin and that each serotype showed only one clonal type. In contrast, serotype 6B and 23F strains showed different phenotypic characteristics as well as multiple clonal types; 10 clones were identified among 6B isolates, and 11 clones were identified among 23F isolates.     

Phenotypes, genotypes, serotypes and molecular epidemiology of erythromycin-resistant Streptococcus agalactiae in Italy

The purpose of this investigation was to analyse Streptococcus agalactiae (group B Streptococcus, GBS) isolates collected in Italy from vaginal and urine samples in respect to their clonality, distribution of virulence factors and antimicrobial resistance determinants. Three hundred and eighty-eight GBS were recovered from clinical samples. They were analysed for antibiotic resistance profiling. Erythromycin-resistant strains were further characterised by multilocus sequence typing (MLST), serotyping and the detection of alp genes of the alpha-like protein (Alp) family. GBS isolates represented 40 different sequence types (STs), grouped in five clonal complexes (CCs) and belonged to seven serotypes. Most serotype V strains (81%) possessed alp2-3; serotype Ia carried mainly epsilon, while the serotype III mainly rib. All isolates were susceptible to penicillin, whereas resistance to erythromycin was detected in 15% of isolates. Most erythromycin-resistant GBS strains were of serotype V (56.8%) and belonged to the CC-1 group (50%). Macrolide resistance phenotypes were the cMLS(B) (46.5%) and the M phenotypes (46.5%) due to the presence of ermB and mefA/E genes, respectively. These results provide data which establish a baseline for monitoring erythromycin resistance in this region and also provide an insight into the correlation among clonal types, serotypes, surface protein and resistance genes. The increased prevalence of strains that displayed the M phenotype strengthens the importance of the epidemiological surveillance of macrolide resistance in GBS, which may also represent an important reservoir of resistance genes for other species.     

Multiplex PCR assays for simultaneous detection of six major serotypes and two virulence-associated phenotypes of Streptococcus suis in tonsillar specimens from pigs

Multiplex PCR assays for the detection and identification of various Streptococcus suis strains in tonsillar specimens from pigs were developed and evaluated. In two separate reactions, five distinct DNA targets were amplified. Three targets, based on the S. suis capsular polysaccharide (cps) genes specific for serotypes 1 (and 14), 7, and 9, were amplified in multiplex PCR I. Two other targets, based on the serotype 2- (and 1/2-) specific cps gene and the epf gene, encoding the EF proteins of virulent serotype 2 and highly virulent serotype 1 strains, were amplified in multiplex PCR II. To identify false-negative results, firefly luciferase (luc) DNA and primers based on the luc gene were included in the assay. The multiplex PCR assays were evaluated with tonsillar specimens from pigs infected with S. suis strains. The results obtained with the PCR assays were compared with the results obtained with a bacteriological examination. Most (94%) of the results obtained with multiplex PCR assays were confirmed by the bacteriological examination. The PCR method seems to be more sensitive compared to the bacteriological method, since the remaining 6% of the samples were positive by PCR and negative by bacteriological examination. These results indicate that the PCR method is highly specific for the detection of S. suis strains most frequently involved in clinical disease in infected pig herds. The serotypes found by PCR in tonsillar specimens from diseased pigs were compared with the serotypes of the strains isolated from the affected tissues of the same pigs. The results showed that there is significant association between carriership and clinical illness for S. suis serotype 9 and EF-positive serotype 2 strains and not for serotype 7 and EF-negative serotype 2 (or 1/2) strains.     

Phenotypic and genetic diversity of invasive pneumococcal isolates recovered from French children

We investigated the phenotypic and genetic diversity (by ribotyping) of Streptococcus pneumoniae isolates recovered from French children, by blood culture from meningitis-free patients (n = 244) and by cerebrospinal fluid culture from patients with meningitis (n = 154). Isolates belonging to serotypes associated with carriage and penicillin-resistant isolates were significantly more frequent in children under 2 years of age than in older children. The seven-valent pneumococcal conjugate vaccine covered 68% of strains associated with bacteremia and 61% of strains associated with meningitis in children under 2 years. Although some serotypes were recovered more frequently from children with bacteremia than from those with meningitis, no difference in the genetic backgrounds of the two groups of strains was found.