Serotype 19A Research Articles

Characteristics of Clonal Complex Changes and Quinolone Resistance-Determining Region Mutations of Levofloxacin-Resistant Streptococcus pneumoniae in South Korea.

Streptococcus pneumoniae is the most common causative agent of community-acquired pneumonia and invasive pneumococcal diseases with high mortality rates. The aims of this study were to evaluate clonal complex (CC) changes of levofloxacin-resistant S. pneumoniae (LRSP) strains and to investigate the relationship between levofloxacin resistance and pneumococcal serotypes. We analyzed the antimicrobial susceptibility of 145 LRSP strains to 18 antimicrobial agents and the quinolone resistance-determining region mutation. Multilocus sequence typing was performed to investigate the genetic relatedness among LRSP strains. Most LRSP strains (96.6%) were multidrug resistant and had simultaneous mutations in gyrA, parC, and parE (91.7%). The serotypes 11A (44.1%) and 13 (14.5%) accounted for 58.6% of LRSP strains, and 32.0% were nonvaccine serotypes. Most LRSP strains were grouped as CC8279 (N = 83; 57.2%), CC189 (N = 10; 6.9%), or CC320 (N = 5; 3.4%). CC8279 was commonly combined with serotypes 11A and 13. There were numerous changes of serotype and CC accompanying the emergence and spread of LRSP. Continuous monitoring of changes in the serotype and sequence type of LRSP is required to follow the spread of LRSP for public health monitoring.

Extensive/Multidrug-Resistant Pneumococci Detected in Clinical Respiratory Tract Samples in Southern Sweden Are Closely Related to International Multidrug-Resistant Lineages

Background/ObjectiveThe frequencies of non-susceptibility against common antibiotics among pneumococci vary greatly across the globe. When compared to other European countries antibiotic resistance against penicillin and macrolides has been uncommon in Sweden in recent years. Multidrug resistance (MDR) is, however, of high importance since relevant treatment options are scarce. The purpose of this study was to characterize the molecular epidemiology, presence of resistance genes and selected virulence genes of extensively drug-resistant (XDR) (n=15) and MDR (n=10) Streptococcus pneumoniae detected in clinical respiratory tract samples isolated from patients in a southern Swedish county 2016-2018. With the aim of relating them to global MDR pneumococci.MethodsWhole genome sequencing (WGS) was performed to determine molecular epidemiology, resistance genes and presence of selected virulence factors. Antimicrobial susceptibility profiles were determined using broth microdilution testing. Further analyses were performed on isolates from the study and from the European nucleotide archive belonging to global pneumococcal sequence cluster (GPSC) 1 (n=86), GPSC9 (n=55) and GPSC10 (n=57). Bacteria were analyzed regarding selected virulence determinants (pilus islet 1, pilus islet 2 and Zinc metalloproteinase C) and resistance genes.ResultsNineteen of 25 isolates were related to dominant global MDR lineages. Seventeen belonged to GPSC1, GPSC9 or GPSC10 with MDR non-PCV serotypes in GPSC9 (serotype 15A and 15C) as well as GPSC10 (serotype 7B, 15B and serogroup 24). Pilus islet-1 and pilus islet-2 were present in most sequence types belonging to GPSC1 and in two isolates within GPSC9 but were not detected in isolates belonging to GPSC10. Zinc metalloproteinase C was well conserved within all analyzed isolates belonging to GPSC9 but were not found in isolates from GPSC1 or GPSC10.ConclusionsAlthough MDR S. pneumoniae is relatively uncommon in Sweden compared to other countries, virulent non-PCV serotypes that are MDR may become an increasing problem, particularly from clusters GPSC9 and GPSC10. Since the incidence of certain serotypes (3, 15A, and 19A) found among our MDR Swedish study isolates are persistent or increasing in invasive pneumococcal disease further surveillance is warranted.

Recalibrated estimates of non-bacteremic and bacteremic pneumococcal community acquired pneumonia in hospitalized Canadian adults from 2010 to 2017 with addition of an extended spectrum serotype-specific urine antigen detection assay

Objective(s)In the context of age- and risk-based pneumococcal vaccine recommendations in Canada, this study presents updated data from active surveillance of pneumococcal community acquired pneumonia (pCAP) and invasive pneumococcal disease (IPD) in hospitalized adults from 2010 to 2017. MethodsS. pneumoniae was detected using culture (blood and sputum), and urine antigen detection (UAD). Serotyping was performed with Quellung, PCR, or using the PCV13- and PPV23 (non-PCV13)-specific UADs. Laboratory results, demographic, and outcome data were categorized by age (16–49, 50–64, and 65 + ) and by disease [non-bacteremic pCAP, bacteremic pCAP, and IPD(non-CAP)]. Results11,129 CAP cases and 216 cases of IPD (non-CAP) were identified. Laboratory testing for S. pneumoniae was performed in 8912 CAP cases, identifying 1264 (14.2%) as pCAP. Of pCAP cases, 811 (64.1%) were non-bacteremic and 455 (35.9%) were bacteremic. Adults 65 + years represented 54.5% of non-bacteremic pCAP, 41.4% of bacteremic pCAP, and 48.6% of IPD cases. Adults 50–64 years contributed 30.3%, 33.1%, and 29.9%, respectively. In pCAP, PCV13 serotypes declined between 2010 and 2014 due to declines in serotypes 7F and 19A, then plateaued from 2015 to 2017 with persistence of serotype 3. In later study years, non-bacteremic pCAP was predominant, and PPV23 (non-PCV13) serotypes increased from 2015 to 2017, with serotypes 22F, 11A, and 9 N being most frequently identified. Compared to non-pCAP, pCAP cases were more likely to be admitted to intensive care units and require mechanical ventilation. These outcomes and mortality were more common in bacteremic pCAP and IPD, versus non-bacteremic pCAP. Conclusion(s)Along with IPD, pCAP surveillance (bacteremic and non-bacteremic) is important as their trends may differ over time. With insufficient herd protection from PCV13 childhood immunization, or use of PPV23 in adults, this study supports direct adult immunization with PCV13 or higher valency conjugate vaccines to reduce the residual burden of pCAP and IPD.

Persistent serotype 3 and 19A invasive pneumococcal diseases in adults in vaccine era: Serotype-dependent difference in ceftriaxone susceptibility

BackgroundInvasive pneumococcal disease (IPD) is associated with substantial morbidity and mortality in children and elderly populations. Serotype distribution and antibiotic susceptibility of IPD isolates are changing with the implementation of pneumococcal vaccination and increasing antibiotic use worldwide. We aimed to determine serotype distribution, antibiogram, and molecular epidemiology of pneumococci in the late stage of PCV13 era. MethodsProspective multicenter IPD surveillance study was conducted for adults aged ≥ 19 years from July 2019 to June 2021. Clinical and epidemiologic data were collected. In addition, antibiotic susceptibility test, serotype identification and multi-locus sequence typing (MLST) was taken for pneumococcal isolates. ResultsA total of 160 IPD cases were collected with mean age of 65.1 years (male, 72.5%). Serotyping was taken for 116 available pneumococcal isolates. PCV13 and PPSV23 serotypes were 32.8% (n = 38) and 56.0% (n = 65), respectively. Serotype 3 (13.8%) and 19A (9.5%) were the most common causative agents of IPD, followed by serogroup 11 (6.9%), 23A (6.9%), 10A (4.3%), and 15B (4.3%). Notably, 32.5% of invasive pneumococcal isolates were non-susceptible to ceftriaxone. Serotypes 11A, 11E and 19A pneumococci showed high ceftriaxone non-susceptible rate (80%, 100% and 81.8% respectively), and they were related to sequence type (ST) 166 and ST320. In comparison, most serotype 3 isolates were ceftriaxone susceptible and related to ST180. ConclusionsPCV serotypes, especially 3 and 19A, are still prevalent in adult IPDs, suggesting that individual PCV13 immunization would be necessary for the elderly people and chronically ill patients. Ceftriaxone non-susceptible rate was remarkably high in invasive pneumococcal isolates.

Molecular Characterization of Penicillin-Binding Protein2x, 2b and 1a of Streptococcus pneumoniae Causing Invasive Pneumococcal Diseases in China: A Multicenter Study.

Streptococcus pneumoniae is a common human pathogen that can cause severe invasive pneumococcal diseases (IPDs). Penicillin-binding proteins (PBPs) are the targets for β-lactam antibiotics (BLAs), which are the common empirical drugs for treatment of pneumococcal infection. This study investigated the serotype distribution and antibiotic resistance patterns of S. pneumoniae strains causing IPD in China, including exploring the association between penicillin (PEN) susceptibility and PBPs variations. A total of 300 invasive S. pneumoniae isolates were collected from 27 teaching hospitals in China (2010-2015). Serotypes were determined by Quellung reaction. Serotypes 23F and 19F were the commonest serotypes in isolates from cerebrospinal fluid (CSF), whilst serotypes 19A and 23F were most commonly seen in non-CSF specimens. Among the 300 invasive S. pneumoniae strains, only one strain (serotype 6A, MIC = 0.25 μg/ml) with PEN MIC value ≤ 0.25 μg/ml did not have any substitutions in the PBPs active sites. All the strains with PEN MIC value ≥ 0.5 μg/ml had different substitutions within PBPs active sites. Substitutions in PBP2b and PBP2x active sites were common in low-level penicillin-resistant S. pneumoniae (PRSP) strains (MIC = 0.5 μg/ml), with or without PBP1a substitution, while all strains with PEN MIC ≥ 1 μg/ml had substitutions in PBP1a active sites, accompanied by PBP2b and PBP2x active site substitutions. Based on the three PBPs substitution combinations, a high degree of diversity was observed amongst the isolates. This study provides some new insights for understanding the serology and antibiotic resistance dynamics of S. pneumoniae causing IPD in China. However, further genomic studies are needed to facilitate a comprehensive understanding of antibiotic resistance mechanisms of S. pneumoniae.

Pneumococcal meningitis in adults in 2014\u20132018 after introduction of pediatric 13-valent pneumococcal conjugate vaccine in Japan

We assessed the impact of the pediatric 13-valent pneumococcal conjugate vaccine (PCV13) on pneumococcal meningitis in adults in Japan in 2014–2018 by comparing epidemiological characteristics of adults with invasive pneumococcal disease with (n = 222) and without (n = 1258) meningitis. The annual incidence of pneumococcal meningitis in 2016–2018 was 0.20–0.26 cases/100,000 population. Age (p < 0.001) and case fatality rate (p = 0.003) were significantly lower in patients with meningitis than in those without meningitis. The odds of developing meningitis were higher in asplenic/hyposplenic or splenectomized patients (adjusted odds ratio [aOR] 2.29, 95% CI 1.27–4.14), for serotypes 10A (aOR 3.26, 95% CI 2.10–5.06) or 23A (aOR 3.91, 95% CI 2.47–6.19), but lower for those aged ≥ 65 years (aOR 0.59, 95% CI 0.44–0.81). PCV13 had an indirect effect on nonmeningitis, but its impact on meningitis was limited because of an increase in non-PCV13 serotypes. Of meningitis isolates, 78 (35.1%) and 3 (1.4%) were penicillin G- or ceftriaxone-resistant, respectively. We also confirmed an association of the pbp1bA641C mutation with meningitis (aOR 2.92, 95% CI 1.51–5.65).

Virulence genes expression profiling of different Shigella flexneri serotypes in response to sub-inhibitory concentrations of azithromycin and ciprofloxacin

BackgroundShigellosis is a self-limiting disease that antibiotic therapy could decrease its complications and duration. However, sublethal levels of antibiotics, may lead to alteration in disease state, besides its role in the emergence of resistant variants. To understand this link, we investigated diversity of Shigella serogroups in children with diarrhea, diversity of S. flexneri serotypes, cytotoxic potential, resistance patterns to antibiotics, and alteration in transcriptional expression of main virulence genes in response to sub-inhibitory concentrations of azithromycin and ciprofloxacin.ResultsThe most frequently isolated serogroups were S. sonnei (70.3%), followed by S. flexneri (29.1%) and S. boydii (0.6%). Ten serotypes were characterized among the S. flexneri isolates, including 2b, 1b, 2a, 1c, 4a, 3a, 3b, 6 and X and/or Xv. Antimicrobial susceptibility testing showed low frequency of multi-drug resistance phenotype among S. flexneri isolates with minimum inhibitory concentrations (MIC) of 0.5–64 and 0.25–8 µg/mL for azithromycin and ciprofloxacin, respectively. Gene expression analysis showed upregulation of icsA in serotype 4a after exposure with azithromycin, whereas other genes in the VirF pathway were downregulated, and downregulation of virB in serotypes 2a and 3a after exposure with ciprofloxacin, while upregulation of noted genes was detected.ConclusionsAlteration in transcription of key virulence genes of S. flexneri serotypes was shown in response to sublethal concentration of antibiotics. The detected incongruency in the extent of gene transcription proposed that diverse regulatory pathways are possibly mediating response to sub-MIC concentrations of antibiotics in S. flexneri.

Synthetic Library of Oligosaccharides Derived from the Capsular Polysaccharide of Streptococcus pneumoniae Serotypes 6A and 6B and Their Immunological Studies.

Streptococcus pneumoniae serotypes 6A and 6B are two of the common causes of invasive pneumococcal diseases. Although capsular polysaccharide conjugates of these two serotypes are included in the leading 13-valent pneumococcal conjugate vaccine, its low immunogenicity and high threshold for manufacturing technology indicated the need for vaccine improvement. Structurally defined synthetic immunogens have potential in dealing with these problems. To this end, we built a library of capsular polysaccharide fragments through convergent chemical synthesis in [2 + 2], [4 + 4], [4 + 3], [4 + 2], and [4 + 1] coupling manners. The library is comprised of 18 glycan antigens from trisaccharides to pseudo-octasaccharides, derived from the capsular repeating phosphorylated pseudo-tetrasaccharide with or without phosphate. Eight of them were selected for mouse immunization and further immunological studies. Four pseudo-tetrasaccharides with terminal or bridging phosphate elicited opsonic antibodies, which exhibited bactericidal activities and moderate cross-reactivities.

Clonal Distribution, Antimicrobial Resistance, and Pilus Islets in S. pneumoniae Isolates from PCV10-Vaccinated Children with Suppurative AOM in Bulgaria (2015‒2020).

Streptococcus pneumoniae is still a leading bacterial pathogen of acute otitis media (AOM), despite the availability of pneumococcal conjugate vaccines (PCVs). We conducted a study on the population structure, antibiotic nonsusceptibility, serotype distribution, and presence of pilus in middle ear fluids ‒ S. pneumoniae isolates recovered from PCV10-vaccinated children with suppurative АОМ in Bulgaria. Non-susceptibility was observed in 68.75% (n = 33) of the isolates, and multidrug resistance (MDR) was detected in 60.4% of the patients. The dual macrolide resistance mechanism was predominant. The most common serotypes were non-PCV10 serotypes 3 (27.1%, n = 13), 19A (25.0%, n = 12), and VT 19F (23.0%, n = 11). Overall, 64.6% were non-PCV10-serotypes. The presence of Pilus type I was observed mostly in the PCV10-serotypes. We found a strong association between clonal complexes (CCs), serotypes, and antimicrobial resistance. Multilocus sequence typing revealed the presence of four CCs: CC320 (39.6%), CC505 (12.5%), CC1377 8.3%), and CC230 (8.3%). The most abundant CC320 comprised MDR 19A and 19F isolates. CC230 clustered MDR isolates from serotypes 19A, 6C, and 14. CC505 and CC1377 were serotype 3 susceptible isolates. The vaccine-induced changes and trends in antimicrobial resistance and clonality must be the focus of systematic investigations.

Serotype-specific effectiveness against pneumococcal carriage and serotype replacement after ten-valent Pneumococcal Conjugate Vaccine (PCV10) introduction in Pakistan.

ObjectivePakistan was one of the first South-Asian countries to introduce the ten-valent pneumococcal conjugate vaccine (PCV10) at the national level, using a 3+0 schedule without catchup, in 2013.MethodsFrom 2014–18, fifteen children <2 years old were recruited every week in Matiari, Sindh, and nasopharyngeal swabs were collected. The samples were cultured, and pneumococcus was further serotyped through multiplex PCR at the Aga Khan University Hospital as per the method described by the Centers for Disease Control and Prevention, USA.ResultsPneumococcus was detected in 2370/3140 (75%) children. Vaccine type (VT) and non-vaccine type (NVT) serotypes were carried by 379 and 1990 children. There was a significant decline in VT carriage (by 40.3%, p-value <0.001), whereas overall NVT carriage remained the same. The prevalence of VT serotypes 6B, 9V/9A, and 19F showed a significant decline by 58.8%, 79.3%, and 56%, respectively. The prevalence of NVT serotypes 19A, 21, and 10A increased by 70%, 33.3%, and 65.6%, respectively, whereas serotypes 13 and 9N/9L decreased by 53.4% and 51.8%, respectively. Serotype-specific vaccine effectiveness estimates that reached statistical significance were for 9V/9A (VE = 65.0, 95% CI 26.0–83.5%), 19F (VE = 55.3, 95% CI 15.5–76.4%) and for the vaccine related serotype 6A (VE = 28.4, 95% CI 0.9–48.2%).ConclusionThe emergence of NVT serotypes, primarily 19A replacing VT serotypes in this rural community, necessitates continuous monitoring of serotypes in the carriage and invasive disease to evaluate the utility of existing vaccine formulations.

Antibiotic Resistance and Molecular Biological Characteristics of Non-13-Valent-Pneumococcal Conjugate Vaccine Serogroup 15 Streptococcus pneumoniae Isolated From Children in China.

Background: The isolation rate of serogroup 15 Streptococcus pneumoniae has been increasing since developing countries began administering the 13-valent pneumococcal conjugate vaccine.Methods: We detected the antibiotic resistance and molecular characteristics of 126 serogroup 15 S. pneumoniae strains isolated from children in China. Serotypes were determined via the Quellung reaction. Antibiotic resistance was tested using the E-test or disc diffusion method. Sequence types were assigned via multilocus sequence typing. Data were analyzed using WHONET 5.6 software.Results: The frequencies of S. pneumoniae serotypes 15A, 15B, 15C, and 15F were 29.37, 40.48, 28.57, and 1.59%, respectively. Continuous-monitoring data from Beijing showed that the annual isolation rates of serogroup 15 S. pneumoniae were 7.64, 7.17, 2.58, 4.35, 3.85, 7.41, and 10.53%, respectively, from 2013 to 2019. All 126 serogroup 15 strains were susceptible to vancomycin and ceftriaxone. The non-susceptibility rate to penicillin was 78.57%. All strains were resistant to erythromycin with high minimum inhibitory concentrations (MICs). The multidrug resistance rate was 78.57%. The most common clonal complexes were CC3397, CC6011, CC10088, CC9785, and ST8589.Conclusion: Serogroup 15 S. pneumoniae is common among children in China, and these strains should be continuously monitored.

A Novel Pneumococcal Surface Protein K of Nonencapsulated Streptococcus pneumoniae Promotes Transmission among Littermates in an Infant Mouse Model with Influenza A Virus Coinfection.

We established an infant mouse model for colonization and transmission by nonencapsulated Streptococcus pneumoniae (NESp) strains to gain important information about its virulence among children. Invasive pneumococcal diseases have decreased dramatically since the worldwide introduction of pneumococcal capsular polysaccharide vaccines. Increasing prevalence of nonvaccine serotypes, including NESp, has been highlighted as a challenge in treatment strategy, but the virulence of NESp is not well understood. Protective strategies against NESp colonization and transmission between children require particularly urgent evaluation. NESp lacks capsules, a major virulence factor of pneumococci, but can cause a variety of infections in children and older people. PspK, a specific surface protein of NESp, is a key factor in establishing nasal colonization. In our infant mouse model for colonization and transmission by NESp strains, NESp could establish stable nasal colonization at the same level as encapsulated serotype 6A in infant mice and could be transmitted between littermates. Transmission was promoted by NESp surface virulence factor PspK and influenza virus coinfection. However, PspK deletion mutants lost the ability to colonize and transmit to new hosts. Promotion of NESp transmission by influenza was due to increased susceptibility of the new hosts. PspK was a key factor not only in establishment of nasal colonization but also in transmission to new hosts. PspK may be targeted as a new candidate vaccine for NESp infection in children.

Response of Reproductive Hormones in Vaccinated and Non-Vaccinated Pneumonic Female Goats Via Experimental Infection of Mannheimia haemolytica Serotype A2 Under Rainy and Hot Seasons

Response of Reproductive Hormones in Vaccinated and Non-Vaccinated Pneumonic Female Goats Via Experimental Infection of Mannheimia haemolytica Serotype A2 Under Rainy and Hot Seasons

Epidemiology of Streptococcus pneumoniae serotypes in children before and after pneumococcal vaccination

Aim. To investigate how the pneumococcal vaccination affects the distribution of Streptococcus pneumoniae serotypes.Materials and Methods. In 2011-2019, 1,852 healthy children (1,354 aged ≤ 5 years and 480 aged from 6 to 17 years) were examined for the nasopharyngeal pneumococcal carriage. Of them, 539 children were tested before the start of pneumococcal vaccination (2011-2014), while 1,313 were tested during the vaccine campaign (2015-2019). Pneumococcal strains were serotyped using multiplex polymerase chain reaction.Results. Streptococcus pneumoniae serotype distribution considerably differed between children ≤ 5 and 6-17 years of age. Serotypes 23F, 19F, 19A, 6AB, and 15BC were prevalent in children ≤ 5 years of age while the older children were characterised by a high prevalence of capsular serotypes (3 and 33AF/37), serogroup 9 (9AV and 9LN), non-typeable streptococci, as well as 19F, 6AB and 6CD serotypes. Vaccination was associated with a significantly decreased prevalence of Streptococcus pneumoniae carriage (from 41.5% to 19.2%) among children ≤ 5 years of age, while this reduction was less pronounced (from 13.5 to 9.0%) in older children. Vaccination led to the shift in the distribution of pneumococcal serotypes towards an increased prevalence of non-vaccine serotypes that was particularly prominent in children ≤ 5 years of age. In particular, vaccination reduced the prevalence of 23F and 19A pneumococcal serotypes but heightened prevalence of 11AD serotype and to the appearance of previously undetected serotypes such as 8, 10A, 17F, 22F, 24ABF, 34, and 39.Conclusion. Pneumococcal vaccination decreased prevalence of pneumococcal carriage, yet causing a serotype replacement effect requiring improved microbiological monitoring in children of all age groups.

Antimicrobial Resistance in Pneumococcal Carriage Isolates from Children under 2 Years of Age in Rural Pakistan.

ABSTRACTAntimicrobial resistance is an emerging public health concern. Ten-valent pneumococcal vaccine (PCV10) was introduced in Pakistan’s Expanded Program on Immunization (EPI) in 2012 as a 3 + 0 schedule without catchup. From 2014 to 2018, children <2 years were randomly selected in two rural union councils of Matiari, Pakistan. Nasopharyngeal swabs were collected using standard WHO guidelines by trained staff and processed at Infectious Disease Research Laboratory at The Aga Khan University, Karachi using culture on sheep blood agar and Multiplex PCR methods described by CDC, USA. Pneumococcal isolates were identified by optochin sensitivity and bile solubility tests. Isolates were then tested for antimicrobial susceptibility by standard Kirby-Bauer disk-diffusion method on Mueller-Hinton Agar (MHA) with 5% sheep blood agar as per Clinical & Laboratory Standards Institute (CLSI) recommendations. Of 3140 children enrolled, pneumococcal isolates were detected in 2370 (75%). Vaccine coverage improved from 41% to 68.4%. Out of the 2370 isolates, 88.4%, 37.6% and 25% were resistant to cotrimoxazole, tetracycline and erythromycin, respectively. There was no resistance to penicillin, ceftriaxone, and vancomycin. For erythromycin, resistance increased from 20% in 2014/15 to 30.8% in 2017/18 and for tetracycline it increased from 34.9% to 41.8% both of which were explained by an increase in prevalence of serotype 19A. Pneumococcal isolates were susceptible to penicillin, ceftriaxone, and vancomycin. They were largely resistant to cotrimoxazole and tetracycline. There was an increase in erythromycin and tetracycline resistance attributed to increasing prevalence of serotype 19A. Pneumococcal isolates from carriage and invasive disease should be closely monitored for antimicrobial susceptibility.IMPORTANCE Antimicrobial resistance is an emerging public health concern particularly in low- and middle-income countries where there is poor regulation and easy availability of antibiotics. This is the first study from Pakistan to report antimicrobial resistance patterns of pneumococcus after vaccine introduction in the community. Pakistan was the first South-Asian country to introduce PCV10 in its Expanded Program on Immunization (EPI) in 2012 as a 3 + 0 schedule without catchup. In this study, we describe the PCV10 impact on antimicrobial resistance patterns of pneumococcal nasopharyngeal carriage in children younger than 2 years of age in a rural district in Pakistan after the introduction of the vaccine.

Whole-Genome Analysis-Based Phylogeographic Investigation of Streptococcus pneumoniae Serotype 19A Sequence Type 320 Isolates in Japan.

ABSTRACTAfter the introduction of the seven-valent pneumococcal conjugate vaccine, the global spread of multidrug-resistant serotype 19A-sequence type 320 (ST320) strains of Streptococcus pneumoniae became a public health concern. In Japan, the main genotype of serotype 19A was ST3111, and the identification rate of ST320 was low. Although the isolates were sporadically detected in both adults and children, their origin remains unknown. Thus, by combining pneumococcal isolates collected in three nationwide pneumococcal surveillance studies conducted in Japan between 2008 and 2020, we analyzed 56 serotype 19A-ST320 isolates along with 931 global isolates, using whole-genome sequencing to uncover the transmission route of the globally distributed clone in Japan. The clone was frequently detected in Okinawa Prefecture, where the United States returned to Japan in 1972. Phylogenetic analysis demonstrated that the isolates from Japan were genetically related to those from the United States; therefore, the common ancestor may have originated in the United States. In addition, Bayesian analysis suggested that the time to the most recent common ancestor of the isolates from Japan and the U.S. was approximately the 1990s to 2000, suggesting the possibility that the common ancestor could have already spread in the United States before the Taiwan 19F-14 isolate was first identified in a Taiwanese hospital in 1997. The phylogeographical analysis supported the transmission of the clone from the United States to Japan, but the analysis could be influenced by sampling bias. These results suggested the possibility that the serotype 19A-ST320 clone had already spread in the United States before being imported into Japan.

Relationship Between Serotypes and Antimicrobial Nonsusceptibility of Streptococcus pneumoniae Clinical Isolates in Tunisia.

Streptococcus pneumoniae remains a significant cause of morbidity and mortality worldwide despite the overall success of the vaccine programs. In Tunisia, pneumococcal conjugate vaccines (PCV)10 was introduced in the national immunization program in April 2019. We sought to determine the relationship between serotypes and antimicrobial nonsusceptibility of S. pneumoniae isolates recovered from clinical samples in the prevaccination period in the south of Tunisia. A total of 504 nonduplicate S. pneumoniae isolates collected between 2012 and 2018 were tested for antimicrobial susceptibility, among them 439 (87.1%) were serotyped. The most common serotypes were 19F (17.8%), 14 (15.3%), 3 (9.1%), 19A (8.2%), and 23F (7.3%). The proportions of isolates with serotypes covered by PCV7, PCV10, and PCV13 were 55.4%, 56.3%, and 77.9%, respectively. Three-quarters (74.4%) of pneumococcal isolates were nonsusceptible to penicillin, and about half (54.8%) were multidrug resistant. Penicillin nonsusceptibility was observed for all 19A and 23F isolates, and was significantly associated with serotypes 19F (odds ratio [OR]: 33.7) and 14 (OR: 8.7). A significant association with multidrug resistance was noted for serotypes 19A (OR: 10), 19F (OR: 9.4), 23F (OR: 8.6), and 6B (OR: 5.2). The alarming rates of pneumococcal antimicrobial nonsusceptibility and the strong association with the most prevalent serotypes compel microbiologists to monitor the impact of the PCV10 introduced recently in our national immunization program.

1181. Serotype Distribution by Age of Remaining Invasive Pneumococcal Disease After Long-Term PCV10/13 Use: The PSERENADE Project

BackgroundPneumococcal conjugate vaccines (PCV) have reduced invasive pneumococcal disease (IPD) (see other PSERENADE abstract), of which > 70% was vaccine-type pre-PCV. We described the serotype (ST) distribution of remaining IPD in countries with mature infant PCV10/13 programs.MethodsIPD ST distribution data were obtained directly from surveillance sites, supplemented with published literature. Mature programs were defined as exclusive use of PCV10 or PCV13 for at least 5-7 years (depended on if prior PCV7 use and/or PCV10/13 catch-up) with primary series uptake > 70%. The distribution was estimated using a multinomial Dirichlet regression, stratified by PCV product and age (< 5 years, ≥ 50 years).ResultsSerotyped IPD cases from 42 PCV13- (n=78,912) and 12 PCV10-using sites (n=8,429) in 41 countries were analyzed. Most sites were from high-income countries (67%) and used a booster dose schedule (81%). For low- and middle-income countries, only 5 and 7 sites had more than 20 eligible cases for children and adults, respectively. In PCV10 sites, 10.0% (95% CI: 6.3-12.9%) and 15.5% (95% CI: 13.4-19.3%) of the remaining IPD during the mature period was PCV10-type among children and adults, respectively (Figure 1). For PCV13 sites, PCV13-type was 26.4% (95% CI: 21.3-30.0%) among children and 29.5% (95% CI: 27.5-33.0%) among adults. PCV20-, PCV24-, and PPV23-type cases ranged from 62-72% across all age and PCV-use groups. ST 19A was the leading ST at PCV10 sites, though more so for children (30.6%, 95% CI: 18.2-43.1%) than adults (14.8%, 95% CI: 11.9-17.8%; Figure 2). ST 3 was a top ST in both PCV10 and PCV13 sites, causing about 9% of cases in children and 14% in adults. ST 6C was the third most common ST in PCV10 sites, causing 6% of cases in both age groups. Some top non-PCV13 STs are included in higher-valent investigational PCVs (15BC, 12F, 22F, 8, 9N) but others are not (24F, 23B, 23A, 15A). Figure 1. Percentage of IPD cases in the mature PCV10/13 period due to serotypes included in current and upcoming products. Serotype (ST) 3 is illustrated separately in lighter purple in the bars corresponding to products that include ST3 due to the uncertain effectiveness against ST3 in current products. ST6C is illustrated in grey above the bars where ST6A is included. Although ST6C is not included in PCV10 or PCV13, PCV13 offers cross-protection through ST6A. ST6A also benefits from cross-protection with ST6B, included in both PCV10 and PCV13. Therefore, ST6A causes a very small fraction of disease in both settings and age groups, and it is not shown. Confidence intervals do not include ST6C, as this serotype is not included in PCV10/13. PCV13 is Pfizer’s Prevnar13/Prevenar13; PCV10 is GSK’s Synflorix.Figure 2. Serotype-specific distribution of IPD in the mature PCV10/13 period. Serotypes are colored by the lowest valency PCV product they are included in. The “x” in the PCV legend represents the extra serotypes included in that product relative to the next lower product (i.e., PCV13x includes serotypes 3, 6A, and 19A not in PCV10). Serotype (ST) 6C is colored separately because, although it is not included in any product, it is covered through cross-protection with PCV13-type serotype 6A. PCV13 is Pfizer’s Prevnar13/Prevenar13; PCV10 is GSK’s Synflorix.ConclusionIPD due to vaccine STs was low for both children and adults in countries with mature PCV programs. ST distribution of remaining IPD differed between PCV10 and PCV13 sites and between age groups. Higher-valency PCVs under evaluation target over half of remaining IPD cases, but some prevalent STs are not included in known investigational products.Disclosures Maria Deloria Knoll, PhD, Merck (Research Grant or Support)Pfizer (Research Grant or Support)

Molecular surveillance of pneumococcal carriage following completion of immunization with the 13-valent pneumococcal conjugate vaccine administered in a 3\u2009+\u20091 schedule

In a cross-sectional study, with the use of molecular methods, we aimed to gain insight into oropharyngeal pneumococcal colonization over time in 1212 Greek children recruited in general pediatric settings throughout the country; they were fully vaccinated with PCV13 (3 + 1 schedule). A single sample was obtained from each child at a time interval of 26 days to 70 months after administration of the 4th (booster) PCV13 dose; sampling time was divided into six time intervals. Carriage of Streptococcus pneumoniae was detected by real-time PCR targeting the lytA gene and isolates were serotyped by singleplex real-time PCR assays. Multiple control procedures to avoid false-positive results were applied. We showed an overall S. pneumoniae carriage rate of 48.6%. Serotyping identified typeable isolates in 82% of the total lytA-positive samples. Non-PCV13 serotypes represented 83.8% of total isolates when excluding serogroups with mixed PCV13 and non-PCV13 serotypes. In multivariate analysis daycare/school attendance emerged as the main contributing factor. Notably, serotypes 19A and 3 were the only two PCV13 serotypes the colonization rate of which increased over time (χ2 for trend P < 0.001 and P = 0.012, respectively). The application of the SP2020 gene on lytA-positive serotyped samples showed pneumococcal colonization in 97% of cases, and the overall colonization profile over time closely resembled that of the lytA gene. With the provisions of the methodological approach and age group of our study, the use of the oropharynx emerges as a reliable alternative to the nasopharynx in estimating pneumococcal carriage in epidemiological studies.

High-throughput nanofluidic real-time PCR to discriminate Pneumococcal Conjugate Vaccine (PCV)-associated serogroups 6, 18, and 22 to serotypes using modified oligonucleotides

Current real-time high-throughput Polymerase Chain Reaction (qPCR) methods do not distinguish serotypes 6A from 6B, 18C from 18A/B and 22F from 22A. We established a nanofluidic real-time PCR (Fluidigm) for serotyping that included Dual-Priming-Oligonucleotides (DPO), a Locked-Nucleic-Acid (LNA) probe and TaqMan assay-sets for high-throughput serotyping. The designed assay-sets target capsular gene wciP in serogroup 6, wciX and wxcM in serogroup 18, and wcwA in serogroup 22. An algorithm combining results from published assay-sets (6A/B/C/D; 6C/D; 18A/B/C; 22A/F) and designed assay-sets for 6A/C; 18B/C/F; 18C/F, 18F and 22F was validated through blind analysis of 1973 archived clinical samples collected from South African children ≤ 5-years-old (2009–2011), previously serotyped with the culture-based Quellung method. All assay-sets were efficient (92–101%), had low variation between replicates (R2 > 0.98), and were able to detect targets at a limit of detection (LOD) of < 100 Colony-Forming-Units (CFU)/mL of sample. There was high concordance (Kappa = 0.73–0.92); sensitivity (85–100%) and specificity (96–100%) for Fluidigm compared with Quellung for serotyping 6A; 6B; 6C; 18C and 22F. Fluidigm distinguishes vaccine-serotypes 6A, 6B, 18C, next-generation PCV-serotype 22F and non-vaccine-serotypes 6C, 6D, 18A, 18B, 18F and 22A. Discriminating single serotypes is important for assessing serotype replacement and the impact of PCVs on vaccine- and non-vaccine serotypes.