Prevalence and Phase Variable Expression Status of Two Autotransporters,NalP and MspA,in Carriage and Disease Isolates of Neisseria meningitidis

Neisseria meningitidis is a human nasopharyngeal commensal capable of causing life-threatening septicemia and meningitis. Many meningococcal surface structures, including the autotransporter proteins NalP and MspA, are subject to phase variation (PV) due to the presence of homopolymeric tracts within their coding sequences. The functions of MspA are unknown. NalP proteolytically cleaves several surface-located virulence factors including the 4CMenB antigen NhbA. Therefore, NalP is a phase-variable regulator of the meningococcal outer membrane and secretome whose expression may reduce isolate susceptibility to 4CMenB-induced immune responses. To improve our understanding of the contributions of MspA and NalP to meningococcal-host interactions, their distribution and phase-variable expression status was studied in epidemiologically relevant samples, including 127 carriage and 514 invasive isolates representative of multiple clonal complexes and serogroups. Prevalence estimates of >98% and >88% were obtained for mspA and nalP, respectively, with no significant differences in their frequencies in disease versus carriage isolates. 16% of serogroup B (MenB) invasive isolates, predominately from clonal complexes ST-269 and ST-461, lacked nalP. Deletion of nalP often resulted from recombination events between flanking repetitive elements. PolyC tract lengths ranged from 6–15 bp in nalP and 6–14 bp in mspA. In an examination of PV status, 58.8% of carriage, and 40.1% of invasive nalP-positive MenB isolates were nalP phase ON. The frequency of this phenotype was not significantly different in serogroup Y (MenY) carriage strains, but was significantly higher in invasive MenY strains (86.3%; p<0.0001). Approximately 90% of MenB carriage and invasive isolates were mspA phase ON; significantly more than MenY carriage (32.7%) or invasive (13.7%) isolates. This differential expression resulted from different mode mspA tract lengths between the serogroups. Our data indicates a differential requirement for NalP and MspA expression in MenB and MenY strains and is a step towards understanding the contributions of phase-variable loci to meningococcal biology.     

Antigenic and genetic characterization of serogroup C meningococci isolated from invasive meningococcal disease cases in Canada from 1999 to 2003

Four hundred and forty-two serogroup C Neisseria meningitidis isolates from individual invasive meningococcal disease (IMD) patients in Canada during the period 1999 to 2003 were analyzed. The majority (84%) of the serogroup C meningococci were characterized by the serotype antigen 2a and belonged to the clonal complex of electrophoretic type ET-15. However, after more than a decade of endemic disease as well as a number of outbreaks and many vaccination campaigns, both genetic and antigenic variants of the serogroup C serotype 2a meningococci were noted. Such variants include strains characterized as C:2a:P1.5 and C:2a:P1.7,1 as well as a non-serotypeable phenotype due to a mutational hot spot on the serotype 2a PorB outer-membrane protein. Meningococci characterized by the antigen formula B:2a:P1.5,2 and B:2a:P1.7,1 have also been found, which suggests capsule switching. Besides the clonal group of ET-15/ET-37, small numbers of serogroup C isolates were found to belong to the clonal complexes of ST-8 (Cluster A4), ST-41/44 (Lineage 3), ST-35, and ST-269.     

Diversity of prophage DNA regions of Streptococcus agalactiae clonal lineages from adults and neonates with invasive infectious disease

The phylogenetic position and prophage DNA content of the genomes of 142 S. agalactiae (group-B streptococcus, GBS) isolates responsible for bacteremia and meningitis in adults and neonates were studied and compared. The distribution of the invasive isolates between the various serotypes, sequence types (STs) and clonal complexes (CCs) differed significantly between adult and neonatal isolates. Use of the neighbor-net algorithm with the PHI test revealed evidence for recombination in the population studied (PHI, P = 2.01 × 10(-6)), and the recombination-mutation ratio (R/M) was 6:7. Nevertheless, the estimated R/M ratio differed between CCs. Analysis of the prophage DNA regions of the genomes of the isolates assigned 90% of the isolates to five major prophage DNA groups: A to E. The mean number of prophage DNA fragments amplified per isolate varied from 2.6 for the isolates of prophage DNA group E to 4.0 for the isolates of prophage DNA group C. The isolates from adults and neonates with invasive diseases were distributed differently between the various prophage DNA groups (P < 0.00001). Group C prophage DNA fragments were found in 52% of adult invasive isolates, whereas 74% of neonatal invasive isolates had prophage DNA fragments of groups A and B. Differences in prophage DNA content were also found between serotypes, STs and CCs (P < 0.00001). All the ST-1 and CC1 isolates, mostly of serotype V, belonged to the prophage DNA group C, whereas 84% of the ST-17 and CC17 isolates, all of serotype III, belonged to prophage DNA groups A and B. These data indicate that the transduction mechanisms, i.e., gene transfer from one bacterium to another by a bacteriophage, underlying genetic recombination in S. agalactiae species, are specific to each intraspecies lineage and population of strains responsible for invasive diseases in adults and neonates.     

Genetic structure of Neisseria meningitidis populations in relation to serogroup, serotype, and outer membrane protein pattern.

The genetic structure of populations of Neisseria meningitidis was examined by an analysis of electrophoretically demonstrable allelic variation at 15 genes encoding enzymes in 650 isolates of eight serogroups (A, B, C, W135, X, Y, Z, and 29E) and 38 nonserogroupable isolates. A total of 331 distinctive multilocus genotypes (electrophoretic types, ETs) was identified, among which mean genetic diversity per locus (H = 0.547) was greater than in Escherichia coli and other bacterial species thus far studied. The intercontinental distribution of some ETs and the recovery of organisms of identical genotype over periods of many years strongly suggest that the genetic structure of N. meningitidis is basically clonal as a consequence of low rates of recombination of chromosomal genes. Variation among strains in serogroup, serotype, and the electrophoretic pattern of the major outer membrane proteins has little relationship to the complex structure of populations revealed by enzyme electrophoresis, which involves 14 major lineages of clones diverging from one another at genetic distances greater than 0.50. Genetic diversity among ETs of isolates of the same serogroup was, on average, 84% of that in the total sample. Clones of serogroup A were unusual in being genotypically less heterogeneous than those of other serogroups and in forming a single phylogenetic group. Isolates of the same serotype or outer membrane protein pattern were also highly heterogeneous; on average, 87 and 97%, respectively, of the total species diversity was represented by ETs of the same serotype or outer membrane protein.     

Population Genetic Structure of Streptococcus pneumoniae in Kilifi,Kenya, Prior to the Introduction of Pneumococcal Conjugate Vaccine

Background

The 10-valent pneumococcal conjugate vaccine (PCV10) was introduced in Kenya in 2011. Introduction of any PCV will perturb the existing pneumococcal population structure, thus the aim was to genotype pneumococci collected in Kilifi before PCV10.

Methods and Findings

Using multilocus sequence typing (MLST), we genotyped >1100 invasive and carriage pneumococci from children, the largest collection genotyped from a single resource-poor country and reported to date. Serotype 1 was the most common serotype causing invasive disease and was rarely detected in carriage; all serotype 1 isolates were members of clonal complex (CC) 217. There were temporal fluctuations in the major circulating sequence types (STs); and although 1-3 major serotype 1, 14 or 23F STs co-circulated annually, the two major serotype 5 STs mainly circulated independently. Major STs/CCs also included isolates of serotypes 3, 12F, 18C and 19A and each shared ≤2 MLST alleles with STs that circulate widely elsewhere. Major CCs associated with non-PCV10 serotypes were predominantly represented by carriage isolates, although serotype 19A and 12F CCs were largely invasive and a serotype 10A CC was equally represented by invasive and carriage isolates.

Conclusions

Understanding the pre-PCV10 population genetic structure in Kilifi will allow for the detection of changes in prevalence of the circulating genotypes and evidence for capsular switching post-vaccine implementation.     

Differences in the population structure of invasive Streptococcus suis strains isolated from pigs and from humans in The Netherlands

Streptococcus suis serotype 2 is the main cause of zoonotic S. suis infection despite the fact that other serotypes are frequently isolated from diseased pigs. Studies comparing concurrent invasive human and pig isolates from a single geographical location are lacking. We compared the population structures of invasive S. suis strains isolated between 1986 and 2008 from human patients (N?=?24) and from pigs with invasive disease (N?=?124) in The Netherlands by serotyping and multi locus sequence typing (MLST). Fifty-six percent of pig isolates were of serotype 9 belonging to 15 clonal complexes (CCs) or singleton sequence types (ST). In contrast, all human isolates were of serotype 2 and belonged to two non-overlapping clonal complexes CC1 (58%) and CC20 (42%). The proportion of serotype 2 isolates among S. suis strains isolated from humans was significantly higher than among strains isolated from pigs (24/24 vs. 29/124; P<0.0001). This difference remained significant when only strains within CC1 and CC20 were considered (24/24 vs. 27/37,P?=?0.004). The Simpson diversity index of the S. suis population isolated from humans (0.598) was smaller than of the population isolated from pigs (0.765, P?=?0.05) indicating that the S. suis population isolated from infected pigs was more diverse than the S. suis population isolated from human patients. S. suis serotype 2 strains of CC20 were all negative in a PCR for detection of genes encoding extracellular protein factor (EF) variants. These data indicate that the polysaccharide capsule is an important correlate of human S. suis infection, irrespective of the ST and EF encoding gene type of S. suis strains.     

The establishment of Neisseria meningitidis serogroup W135 of the clonal complex ET-37/ST-11 as an epidemic clone and the persistence of serogroup A isolates in Burkina Faso

We analyzed 48 invasive isolates of Neisseria meningitidis that were isolated from meningitis cases in Burkina Faso (April 2002 to April 2003). Thirty-nine of these isolates had the phenotype (serogroup:serotype:serosubtype) W135:2a:P1.5,2, eight isolates were A:4:P1.9 and one isolate was nongroupable:nonserotypable:nonserosubtypable. Genotyping of meningococcal isolates showed that W135 isolates belonged to the sequence type (ST)-11. The nongroupable isolate was of genogroup W135 and belonged to ST-192. Isolates of serogroup A belonged to ST-2859 (a member of the subgroup III/ST-5 clonal complex). W135 (ST-11) isolates involved in meningitis outbreaks in Burkina Faso differed from those involved in the Hajj-2000 associated outbreak by their pulsed-field gel electrophoresis profile. These data confirm the changing epidemiology of meningococcal infection in Burkina Faso with the establishment and expansion of serogroup W135 N. meningitidis strains of the ET-37/ST-11 clonal complex, as well as the emergence of a new clone within the subgroup III/ST-5 clonal complex.     

A variable region within the genome of Streptococcus pneumoniae contributes to strain-strain variation in virulence

The bacterial factors responsible for the variation in invasive potential between different clones and serotypes of Streptococcus pneumoniae are largely unknown. Therefore, the isolation of rare serotype 1 carriage strains in Indigenous Australian communities provided a unique opportunity to compare the genomes of non-invasive and invasive isolates of the same serotype in order to identify such factors. The human virulence status of non-invasive, intermediately virulent and highly virulent serotype 1 isolates was reflected in mice and showed that whilst both human non-invasive and highly virulent isolates were able to colonize the murine nasopharynx equally, only the human highly virulent isolates were able to invade and survive in the murine lungs and blood. Genomic sequencing comparisons between these isolates identified 8 regions >1 kb in size that were specific to only the highly virulent isolates, and included a version of the pneumococcal pathogenicity island 1 variable region (PPI-1v), phage-associated adherence factors, transporters and metabolic enzymes. In particular, a phage-associated endolysin, a putative iron/lead permease and an operon within PPI-1v exhibited niche-specific changes in expression that suggest important roles for these genes in the lungs and blood. Moreover, in vivo competition between pneumococci carrying PPI-1v derivatives representing the two identified versions of the region showed that the version of PPI-1v in the highly virulent isolates was more competitive than the version from the less virulent isolates in the nasopharyngeal tissue, blood and lungs. This study is the first to perform genomic comparisons between serotype 1 isolates with distinct virulence profiles that correlate between mice and humans, and has highlighted the important role that hypervariable genomic loci, such as PPI-1v, play in pneumococcal disease. The findings of this study have important implications for understanding the processes that drive progression from colonization to invasive disease and will help direct the development of novel therapeutic strategies.     

A new perspective on Listeria monocytogenes evolution

Listeria monocytogenes is a model organism for cellular microbiology and host-pathogen interaction studies and an important food-borne pathogen widespread in the environment, thus representing an attractive model to study the evolution of virulence. The phylogenetic structure of L. monocytogenes was determined by sequencing internal portions of seven housekeeping genes (3,288 nucleotides) in 360 representative isolates. Fifty-eight of the 126 disclosed sequence types were grouped into seven well-demarcated clonal complexes (clones) that comprised almost 75% of clinical isolates. Each clone had a unique or dominant serotype (4b for clones 1, 2 and 4, 1/2b for clones 3 and 5, 1/2a for clone 7, and 1/2c for clone 9), with no association of clones with clinical forms of human listeriosis. Homologous recombination was extremely limited (r/m<1 for nucleotides), implying long-term genetic stability of multilocus genotypes over time. Bayesian analysis based on 438 SNPs recovered the three previously defined lineages, plus one unclassified isolate of mixed ancestry. The phylogenetic distribution of serotypes indicated that serotype 4b evolved once from 1/2b, the likely ancestral serotype of lineage I. Serotype 1/2c derived once from 1/2a, with reference strain EGDe (1/2a) likely representing an intermediate evolutionary state. In contrast to housekeeping genes, the virulence factor internalin (InlA) evolved by localized recombination resulting in a mosaic pattern, with convergent evolution indicative of natural selection towards a truncation of InlA protein. This work provides a reference evolutionary framework for future studies on L. monocytogenes epidemiology, ecology, and virulence.     

Population pattern of pneumococci with lower susceptibility to penicillin and prospects of antipneumococcal vaccination to control antibiotic resistance distribution

Large-scale antipneumococcal vaccination is followed by changes in the serotype composition and level of antibiotic resistance in pneumococci. The aim of the study was to evaluate the serotype composition and population pattern of pneumococci with lower susceptibility to penicillin before large-scale antipneumococcal vaccination. Among 260 Streptococcus pneumoniae strains isolated in the Russian Federation within 2003-2007, serotypes 23F (37.2%) and 19F (13.9%) were the most frequent ones. 19.3% of the isolates belonged to serogroup 6, 3.6% of the isolates each belonged to serotype 3 and serogroup 18, 4.9% of the isolates belonged to serotype 14 and 2.2% of the isolates belonged to serotype 19A. 66.8% of the isolates belonged to serotypes of the 7-valent conjugated pneumococcal vaccine, 67.3 and 82.1% of the isolates belonged to the 10- and 13-valent conjugated pneumococcal vaccines respectively. The isolates with lower susceptibility to penicillin were characterized by significant clonality and 56.9% of them belonged to 4 global clonal complexes (CC81, CC156, CC320 and CC315). Inclusion of the conjugated antipneumococcal vaccine to the National Vaccination Time-Table of the Russian Federation could promote lower levels of antibiotic resistance in pneumococci.     

Influence of Two Vaccination Campaigns on Genetic Diversity of Invasive Neisseria meningitidis Isolates in Northern Spain (1997–2008)

Background

Neisseria meningitidis diversifies rapidly, due to its high recombination rates. The aim of this study was to analyze the possible impact of two vaccination campaigns (a once-off A/C polysaccharide vaccination campaign in people aged 18 months to 20 years old in 1997, and a meningococcal C conjugate vaccination campaign in children aged ≤6 years old from 2000 to 2008) on diversification of the population of invasive isolates obtained between 1997 and 2008. All of the 461 available isolates were included (2, 319, 123, 11 and 6 belonging to serogroups A, B, C, Y and W-135, respectively).

Methodology/Principal Findings

The isolates were analyzed for diversity using multilocus sequence typing, eBURST and the S.T.A.R.T.2 program. One hundred and seven sequence types (ST) and 20 clonal complexes were obtained. Five different STs (ST11, ST8, ST33, ST1163 and ST3496) included 56.4% of the isolates. With the exception of ST11, all other STs were associated with a specific serogroup. Epidemic circulation of serogroup C ST8 isolates was detected in 1997–1998, as well as epidemic circulation of ST11 isolates (serogroups B and C) in 2002–2004. The epidemic behavior of serogroup B ST11 (ST11_B:2a:P1.5) was similar, although with lesser intensity, to that of ST11 of serogroup C. Although clonality increased during epidemic years, the overall diversity of the meningococcal population did not increase throughout the 12 years of the study.

Conclusion

The overall diversity of the meningococcal population, measured by the frequency of STs and clonal complexes, numbers of alleles, polymorphic sites, and index of association, remained relatively constant throughout the study period, contradicting previous findings by other researchers.     

Identification of the High-Virulence Clone of Group B Streptococci by Using a Probe Containing a Putative Aldolase Gene

A high-virulence clone (HVC) was proposed as causing much of the morbidity and mortality when a collection of group B Streptococcus (GBS) isolates was examined by multi-locus enzyme electrophoresis. HVC isolates could be further distinguished by their inability to grow at 40°C, and a temperature-sensitive aldolase was identified as responsible for this characteristic. In the present study, the HVC was sought in a collection of 57 GBS isolates by hybridization with a probe containing a putative aldolase gene on genomic DNA restriction enzyme digests. Isolates were initially classified as HVC or non-HVC by their inability to grow at 40°C. Three serotype III invasive isolates had the HVC control restriction/hybridization pattern. They were also unable to grow at 40°C. The remaining 11 invasive and all carrier isolates showed a pattern identical to that of the non-HVC control. These results provide additional support for the existence of a highly virulent clonal group among serotype III isolates and suggest that hybridization with a probe containing the aldolase gene on DNA restriction enzyme digests can be an alternative method for identifying highly virulent isolates. Received: 2 September 2002 / Accepted: 13 December 2002     

Population Snapshot of Streptococcus pneumoniae Causing Invasive Disease in South Africa Prior to Introduction of Pneumococcal Conjugate Vaccines

We determined the sequence types of isolates that caused invasive pneumococcal disease (IPD) prior to routine use of pneumococcal conjugate vaccines (PCV) in South Africa. PCV-13 serotypes and 6C isolates collected in 2007 (1 461/2 437, 60%) from patients of all ages as part of on-going, national, laboratory-based surveillance for IPD, were selected for genetic characterization. In addition, all 134 non-PCV isolates from children <2 years were selected for characterization. Sequence type diversity by serotype and age category (children <5 years vs. individuals ≥5 years) was assessed for PCV serotypes using Simpson’s index of diversity. Similar genotypes circulated among isolates from children and adults and the majority of serotypes were heterogeneous. While globally disseminated clones were common among some serotypes (e.g., serotype 1 [clonal complex (CC) 217, 98% of all serotype 1] and 14 [CC230, 43%)]), some were represented mainly by clonal complexes rarely reported elsewhere (e.g., serotype 3 [CC458, 60%] and 19A [CC2062, 83%]). In children <2 years, serotype 15B and 8 were the most common serotypes among non-PCV isolates (16% [22/134] and 15% [20/134] isolates, respectively). Sequence type 7052 and 53 were most common among serotypes 15B and 8 isolates and accounted for 58% (7/12) and 64% (9/14) of the isolates, respectively. Serotype 19F, 14, 19A and 15B had the highest proportions of penicillin non-susceptible isolates. Genotypes rarely reported in other parts of the world but common among some of our serotypes highlight the importance of our data as these genotypes may emerge post PCV introduction.     

Pseudomonas aeruginosa displays an epidemic population structure

Bacteria can have population structures ranging from the fully sexual to the highly clonal. Despite numerous studies, the population structure of Pseudomonas aeruginosa is still somewhat contentious. We used a polyphasic approach in order to shed new light on this issue. A data set consisting of three outer membrane (lipo)protein gene sequences (oprI, oprL and oprD), a DNA-based fingerprint (amplified fragment length polymorphism), serotype and pyoverdine type of 73 P. aeruginosa clinical and environmental isolates, collected across the world, was analysed using biological data analysis software. We observed a clear mosaicism in the results, non-congruence between results of different typing methods and a microscale mosaic structure in the oprD gene. Hence, in this network, we also observed some clonal complexes characterized by an almost identical data set. The most recent clones exhibited serotypes O1, 6, 11 and 12. No obvious correlation was observed between these dominant clones and habitat or, with the exception of some recent clones, geographical origin. Our results are consistent with, and even clarify, some seemingly contradictory results in earlier epidemiological studies. Therefore, we suggest an epidemic population structure for P. aeruginosa, comparable with that of Neisseria meningitidis, a superficially clonal structure with frequent recombinations, in which occasionally highly successful epidemic clones arise.     

How clonal is Staphylococcus aureus?

Staphylococcus aureus is an important human pathogen and represents a growing public health burden owing to the emergence and spread of antibiotic-resistant clones, particularly within the hospital environment. Despite this, basic questions about the evolution and population biology of the species, particularly with regard to the extent and impact of homologous recombination, remain unanswered. We address these issues through an analysis of sequence data obtained from the characterization by multilocus sequence typing (MLST) of 334 isolates of S. aureus, recovered from a well-defined population, over a limited time span. We find no significant differences in the distribution of multilocus genotypes between strains isolated from carriers and those from patients with invasive disease; there is, therefore, no evidence from MLST data, which index variation within the stable "core" genome, for the existence of hypervirulent clones of this pathogen. Examination of the sequence changes at MLST loci during clonal diversification shows that point mutations give rise to new alleles at least 15-fold more frequently than does recombination. This contrasts with the naturally transformable species Neisseria meningitidis and Streptococcus pneumoniae, in which alleles change between 5- and 10-fold more frequently by recombination than by mutation. However, phylogenetic analysis suggests that homologous recombination does contribute toward the evolution of this species over the long term. Finally, we note a striking excess of nonsynonymous substitutions in comparisons between isolates belonging to the same clonal complex compared to isolates belonging to different clonal complexes, suggesting that the removal of deleterious mutations by purifying selection may be relatively slow.     

The carriage population of Staphylococcus aureus from Mali is composed of a combination of pandemic clones and the divergent Panton-Valentine leukocidin-positive genotype ST152

Staphylococcus aureus is an important human pathogen, but it appears more commonly in asymptomatic colonization of the nasopharynx than in cases of invasive disease. Evidence concerning the global population structure of S. aureus is limited by the overrepresentation in the multilocus sequence testing database of disease isolates recovered from Western Europe, the Americas, Australia, and Japan. We address this by presenting data from the S. aureus carriage population in Mali, the first detailed characterization of asymptomatic carriage from an African population. These data confirm the pandemic spread of many of the common S. aureus clones in the carriage population. We also note the high frequency (approximately 24%) of a single divergent genotype, sequence type 152 (ST152), which has not previously been recovered from nasal carriage isolates but corresponds to a sporadic Panton-Valentine leukocidin (PVL)-positive, community-acquired methicillin-resistant S. aureus clone noted mostly in Central Europe. We show that 100% of the ST152 isolates recovered from nasal carriage samples in Mali are PVL positive and discuss implications relating to the emergence and spread of this virulent genotype.     

Epidemiology of Streptococcus pneumoniae Serogroup 6 Isolates from IPD in Children and Adults in Germany

This study presents serogroup 6 isolates from invasive pneumococcal disease (IPD) before and after the recommendation for childhood pneumococcal conjugate vaccination in Germany (July 2006). A total of 19,299 (children: 3508, adults: 15,791) isolates were serotyped. Serogroup 6 isolates accounted for 9.5% (children) and 6.7% (adults), respectively. 548 isolates had serotype 6A, 558 had serotype 6B, 285 had serotype 6C, and 4 had serotype 6D. Among children, serotype 6B was most prevalent (7.5% of isolates) before vaccination, followed by 6A and 6C. After the 7-valent pneumococcal conjugate vaccine (PCV7), the prevalence of serotype 6B significantly decreased (p = 0.040), a pattern which continued in the higher-valent PCV period (PCV10, PCV13). Serotype 6A prevalence showed a slight increase directly after the start of PCV7 vaccination, followed by a decrease which continued throughout the PCV10/13 period. Serotype 6C prevalence remained low. Serotype 6D was not found among IPD isolates from children. Among adults, prevalence of both 6A and 6B decreased, with 6B reaching statistical significance (p = 0.045) and 6A showing a small increase in 2011–2012. Serotype 6C prevalence was 1.5% or lower before vaccination, but increased post-vaccination to 3.6% in 2011/12 (p = 0.031). Four serotype 6D isolates were found post-PCV7 childhood vaccination, and two post-PCV10/13. Antibiotic resistance was found mainly in serotype 6B; serotype 6A showed lower resistance rates. Serotype 6C isolates only showed resistance among adults; serotype 6D isolates showed no resistance. Multilocus sequence typing showed that sequence type (ST) 1692 was the most prevalent serotype 6C clone. Thirty-two other STs were found among serotype 6C isolates, of which 12 have not been previously reported. The four serotype 6D isolates had ST 948, ST 2185 and two new STs: 8422 and 8442. Two serogroup 6 isolates could not be assigned to a serotype, but had STs common to serogroup 6.     

Molecular Characterization of Invasive Meningococcal Isolates from Countries in the African Meningitis Belt before Introduction of a Serogroup A Conjugate Vaccine

Background

The serogroup A conjugate meningococcal vaccine, MenAfriVac, was introduced in mass vaccination campaigns in December 2010 in Burkina Faso, Mali and Niger. In the coming years, vaccination will be extended to other African countries at risk of epidemics. To document the molecular characteristics of disease-causing meningococcal strains circulating in the meningitis belt of Africa before vaccine introduction, the World Health Organization Collaborating Centers on Meningococci in Europe and United States established a common strain collection of 773 isolates from cases of invasive meningococcal disease collected between 2004 and 2010 from 13 sub-Saharan countries.

Methodology

All isolates were characterized by multilocus sequence typing, and 487 (62%) were also analyzed for genetic variation in the surface antigens PorA and FetA. Antibiotic susceptibility was tested for part of the collection.

Principal Findings

Only 19 sequence types (STs) belonging to 6 clonal complexes were revealed. ST-5 clonal complex dominated with 578 (74.8%) isolates. All ST-5 complex isolates were remarkably homogeneous in their PorA (P1.20,9) and FetA (F3-1) and characterized the serogroup A strains which have been responsible for most epidemics during this time period. Sixty-eight (8.8%) of the 773 isolates belonged to the ST-11 clonal complex which was mainly represented by serogroup W135, while an additional 38 (4.9%) W135 isolates belonged to the ST-175 complex. Forty-eight (6.2%) serogroup X isolates from West Africa belonged to the ST-181 complex, while serogroup X cases in Kenya and Uganda were caused by an unrelated clone, ST-5403. Serogroup X, ST-181, emerged in Burkina Faso before vaccine introduction.

Conclusions

In the seven years preceding introduction of a new serogroup A conjugate vaccine, serogroup A of the ST-5 clonal complex was identified as the predominant disease-causing strain.     

Spatio-temporal analysis of infra-specific genetic variations among a Pseudomonas aeruginosa water network hospital population: invasion and selection of clonal complexes

Aims: To investigate infra-specific spatio-temporal dynamics of a hospital water network Pseudomonas aeruginosa population. To infer the origin of water network isolates and assess their potential health hazard. Methods and Results: 168 P. aeruginosa strains were isolated from tap waters and swabs of tap nozzle aerators of a hospital unit, over 2 years, and from rectal swabs and nosocomial infections. Genetic diversity among this collection was assessed by pulsed field gel electrophoresis of SpeI restricted genomic DNA. Virulence gene sets, biofilm properties, and hypochlorite resistance were analysed. Exactly 68% of the water samples and 74% of the tap nozzle aerators harboured P. aeruginosa. The strains were divided into 22 clonal lineages, with one dominant clone shown to have been involved in a nosocomial infection. Conclusions: An important turnover among the P. aeruginosa hospital population was observed. Some clonal lineages were found to persist, spread in the unit, and diversify into clonal complexes. Rectal carriage appeared an important source of contamination of the water network. Significance and Impact of the Study: High P. aeruginosa infra-specific population diversity suggested a broad ability in colonizing water networks but persistence analysis indicated a strong selection leading to the emergence of dominant clones.     

Genetic Diversity and Levels of Expression of Factor H Binding Protein among Carriage Isolates of Neisseria meningitidis

The prevention of meningococcal disease may be improved by recombinant vaccines such as 4CMenB and rLP2086 that target the factor H binding protein (fHbp), an immunogenic surface component of Neisseria meningitidis present as one of three variants. Whether such vaccines decrease carriage of invasive isolates and thus induce herd immunity is unknown. We analyzed the genetic diversity and levels of expression of fHbp among 268 carriage strains and compare them to those of 467 invasive strains. fhbp gene sequencing showed higher proportions of variants 2 and 3 among carriage isolates (p<0.0001). Carriage isolates expressed lower levels of fHbp (p<0.01) but that remain high enough to predict targeting by antibodies against fHbp particularly in group B isolates belonging to the frequent hypervirulent clonal complexes in Europe and North America (cc32, cc41/44, cc269). This suggests that fHbp targeting meningococcal vaccines might reduce, at least in part, the acquisition of some hyperinvasive isolates.