Rama Kandasamy, Australia

University of New South Wales Women's and Children's Health

Presenter of 3 Presentations

PCV10 IMPACT ON PNEUMOCOCCAL LINEAGES ISOLATED FROM HEALTHY NEPALESE CHILDREN. (ID 736)

Abstract

Background

The 10-valent pneumococcal conjugate vaccine (PCV10) was introduced into the Nepalese infant immunisation schedule in August 2015. We aimed to examine how PCV10 introduction in affected pneumococcal lineages.

Methods

DNA from randomly selected nasopharyngeal pneumococcal isolates of healthy community-based Nepalese children in the Kathmandu valley pre- (2009-2014) and post-PCV10 (2017-2018) introduction, underwent whole-genome-sequencing on the Wellcome Sanger Institutes core sequencing pipeline. Isolates were clustered into lineages based on shared sequence and gene content using Population Partitioning Using Nucleotide K-mers (PopPUNK) software.

Results

313 and 284 pre- and post-PCV10 isolates were sequenced. There was a significant reduction in the proportion of PCV10 serotypes when comparing pre 73/313 (23.3%) with post 37/284 (13%) PCV10 samples (p=0.0014). Overall 122 distinct lineages were identified, 98 pre- and 74 post-PCV10. Simpson's index of diversity for the lineages was 0.992 and 0.987 pre- and post-PCV10 respectively. Within the 3 largest PCV10 serotype lineages there were no examples of non-PCV10 serotype isolates pre-vaccination, whereas all 3 lineages contained non-PCV10 serotypes post-vaccination.

Conclusions

PCV10 serotype prevalence significantly declined following PCV10 introduction. However, strain diversity remained high post-PCV10 and there is evidence suggestive of vaccine escape via capsular-switching among lineages possessing predominantly vaccine-covered capsules prior to PCV10 introduction.

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GENOME-WIDE ASSOCIATION STUDY OF PNEUMOCOCCAL CARRIAGE AMONG NEPALESE CHILDREN (ID 16)

Abstract

Background

Determining the host molecular genetic characteristics of pneumococcal colonisation may inform the development of new clinical interventions which could interrupt pneumococcal transmission and establishment of disease. We performed a genome-wide association study to identify the genes associated with pneumococcal carriage.

Methods

DNA collected from healthy Nepalese children were genotyped using Illumina Global Screening Arrays. Array data underwent QC and filtering before undergoing imputation using the HRC R1.1 2016 reference panel. Nasopharyngeal swabs collected from participants were processed for the presence of pneumococci by conventional microbiological techniques. Association analysis was performed using PLINK 1.9.

Results

Following filtering, 1355 carriers (cases) and 766 non-carriers (controls) were analysed. 10 variants within a single region, were associated with pneumococcal carriage (p<10-8). The variant that had the strongest association with pneumococcal carriage (MAF carriers = 0.07 vs MAF non-carriers = 0.13, OR 0.52, 95% CI 0.42-0.64, p=2.3x10-8), is within an intergenic region between PPFIA2 and CCDC59.

Conclusions

We identified host genetic variants associated with pneumococcal carriage. Further studies confirming this association and its biological role in pneumococcal carriage are needed.

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GENOME-WIDE ASSOCIATION STUDY OF COLONISING NASOPHARYNGEAL PNEUMOCOCCI OBTAINED FROM CHILDREN IN NEPAL TO IDENTIFY GENES ASSOCIATED WITH PNEUMONIA. (ID 729)

Abstract

Background

Identifying the molecular characteristics of pneumococci associated with disease may inform development of new clinical interventions. We aimed to perform a bacterial genome-wide association study to identify pneumococcal genes associated with carriage among children with pneumonia.

Methods

DNA from nasopharyngeal pneumococcal isolates obtained from Nepalese children admitted to hospital with pneumonia (cases) and healthy community-based children (controls), underwent whole-genome-sequencing on the Wellcome Sanger Institutes core sequencing pipeline. The association of variants from sequences mapped against the S. pneumoniae ATCC700669 genome, with cluster of orthologous groups using a fixed effects model, was performed using a python based sequence element enrichment analysis.

Results

245 case and 597 control isolates were sequenced. 405461 variants were identified and 31708 tested after filtering. 20 variants from colonising bacteria had a strong association (p<10-8) with pneumonia. 18/20 of these variants were located within the lacE2 gene. The variant with the strongest association, presence of an A allele at position 1066739, was identified in 240/597 (40%) of controls and 150/245 (61%) of cases (p=10-10).

Conclusions

In this study in Nepal the pneumococcal gene lacE2 was associated with colonisation in children with pneumonia. Studies examining the role of lacE2 in the pathogenesis of pneumococcal pneumonia are needed.

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Author Of 6 Presentations

GENOME-WIDE ASSOCIATION STUDY OF COLONISING NASOPHARYNGEAL PNEUMOCOCCI OBTAINED FROM CHILDREN IN NEPAL TO IDENTIFY GENES ASSOCIATED WITH PNEUMONIA. (ID 729)

Abstract

Background

Identifying the molecular characteristics of pneumococci associated with disease may inform development of new clinical interventions. We aimed to perform a bacterial genome-wide association study to identify pneumococcal genes associated with carriage among children with pneumonia.

Methods

DNA from nasopharyngeal pneumococcal isolates obtained from Nepalese children admitted to hospital with pneumonia (cases) and healthy community-based children (controls), underwent whole-genome-sequencing on the Wellcome Sanger Institutes core sequencing pipeline. The association of variants from sequences mapped against the S. pneumoniae ATCC700669 genome, with cluster of orthologous groups using a fixed effects model, was performed using a python based sequence element enrichment analysis.

Results

245 case and 597 control isolates were sequenced. 405461 variants were identified and 31708 tested after filtering. 20 variants from colonising bacteria had a strong association (p<10-8) with pneumonia. 18/20 of these variants were located within the lacE2 gene. The variant with the strongest association, presence of an A allele at position 1066739, was identified in 240/597 (40%) of controls and 150/245 (61%) of cases (p=10-10).

Conclusions

In this study in Nepal the pneumococcal gene lacE2 was associated with colonisation in children with pneumonia. Studies examining the role of lacE2 in the pathogenesis of pneumococcal pneumonia are needed.

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PCV10 IMPACT ON PNEUMOCOCCAL LINEAGES ISOLATED FROM HEALTHY NEPALESE CHILDREN. (ID 736)

Abstract

Background

The 10-valent pneumococcal conjugate vaccine (PCV10) was introduced into the Nepalese infant immunisation schedule in August 2015. We aimed to examine how PCV10 introduction in affected pneumococcal lineages.

Methods

DNA from randomly selected nasopharyngeal pneumococcal isolates of healthy community-based Nepalese children in the Kathmandu valley pre- (2009-2014) and post-PCV10 (2017-2018) introduction, underwent whole-genome-sequencing on the Wellcome Sanger Institutes core sequencing pipeline. Isolates were clustered into lineages based on shared sequence and gene content using Population Partitioning Using Nucleotide K-mers (PopPUNK) software.

Results

313 and 284 pre- and post-PCV10 isolates were sequenced. There was a significant reduction in the proportion of PCV10 serotypes when comparing pre 73/313 (23.3%) with post 37/284 (13%) PCV10 samples (p=0.0014). Overall 122 distinct lineages were identified, 98 pre- and 74 post-PCV10. Simpson's index of diversity for the lineages was 0.992 and 0.987 pre- and post-PCV10 respectively. Within the 3 largest PCV10 serotype lineages there were no examples of non-PCV10 serotype isolates pre-vaccination, whereas all 3 lineages contained non-PCV10 serotypes post-vaccination.

Conclusions

PCV10 serotype prevalence significantly declined following PCV10 introduction. However, strain diversity remained high post-PCV10 and there is evidence suggestive of vaccine escape via capsular-switching among lineages possessing predominantly vaccine-covered capsules prior to PCV10 introduction.

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MONITORING VACCINE IMPACT ON COMMUNITY CARRIAGE IN NEPAL REVEALS CHANGES IN THE CIRCULATING POPULATION OF PNEUMOCOCCAL SEROTYPES AND ANTIMICROBIAL RESISTANCE GENES (ID 977)

Abstract

Background

Community carriage of pneumococcal serotypes in children was assessed pre- and post-PCV10 introduction in Nepal to monitor pneumococcal vaccine impact. Molecular serotyping by microarray enabled detection of multiple-serotype carriage plus non-encapsulated pneumococcal lineages, related Streptococcus species and selected antimicrobial resistance genes.

Methods

Nasopharyngeal swabs were collected from healthy Nepalese children in 2014-15 (pre-PCV10) and 2017-18 (post-PCV10). DNA was extracted from plate sweeps of 1,241 and 1,445 swab cultures for pre- and post-vaccine periods respectively and analysed by Senti-SP molecular serotyping microarray.

Results

Comparing carriage among children pre- and post-PCV10, there was a decrease in PCV10 serotype carriage (37% vs 17%) and an increase in non-vaccine serotype carriage (67% vs 73%). There was no change for non-encapsulated pneumococcal lineages (16% vs 16%), an increase in related Streptococcal species (22% vs 25%) and an increase in detection of antimicrobial resistance genes (65% vs 74%). Multiple pneumococcal serotype carriage decreased (24% vs 16%) and multiple carriage including non-encapsulated pneumococci and related Streptococcal species also decreased (45% vs 41%).

Conclusions

Introduction of PCV10 in Nepal has resulted in a decrease in vaccine type carriage within two years. However, increases in carriage of non-vaccine types as well as antimicrobial resistance genes and related Streptococcal species were observed.

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GENOME-WIDE ASSOCIATION STUDY OF PNEUMOCOCCAL CARRIAGE AMONG NEPALESE CHILDREN (ID 16)

Abstract

Background

Determining the host molecular genetic characteristics of pneumococcal colonisation may inform the development of new clinical interventions which could interrupt pneumococcal transmission and establishment of disease. We performed a genome-wide association study to identify the genes associated with pneumococcal carriage.

Methods

DNA collected from healthy Nepalese children were genotyped using Illumina Global Screening Arrays. Array data underwent QC and filtering before undergoing imputation using the HRC R1.1 2016 reference panel. Nasopharyngeal swabs collected from participants were processed for the presence of pneumococci by conventional microbiological techniques. Association analysis was performed using PLINK 1.9.

Results

Following filtering, 1355 carriers (cases) and 766 non-carriers (controls) were analysed. 10 variants within a single region, were associated with pneumococcal carriage (p<10-8). The variant that had the strongest association with pneumococcal carriage (MAF carriers = 0.07 vs MAF non-carriers = 0.13, OR 0.52, 95% CI 0.42-0.64, p=2.3x10-8), is within an intergenic region between PPFIA2 and CCDC59.

Conclusions

We identified host genetic variants associated with pneumococcal carriage. Further studies confirming this association and its biological role in pneumococcal carriage are needed.

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SEROTYPE-SPECIFIC PNEUMOCOCCAL CARRIAGE DENSITY AMONG HEALTHY NEPALESE CHILDREN POST-PCV10 INTRODUCTION (ID 1047)

Abstract

Background

The density of pneumococcal carriage provides an insight into the dynamics of transmission, colonisation and vaccine effect. We aimed to measure serotype-specific carriage density 2-3 years after 10-valent pneumococcal conjugate vaccine (PCV10) introduction, using qPCR and bacterial DNA microarray.

Methods

Nasopharyngeal swabs were collected from healthy Nepalese children from the Kathmandu Valley between April 2017 and August 2018. DNA was extracted from the swab media and qPCR performed for pneumococcal autolysin (lytA). Swab media were also plated on blood agar, incubated overnight, and plate sweeps collected. DNA was extracted from plate sweeps and molecular serotyped using the Senti-SPv1.5 microarray (BUGS Bioscience, UK).

Results

1264 swabs were collected and analysed by both microarray and qPCR. The mean density of PCV10 serotypes was significantly higher than non-PCV10 serotypes (10^3.9 vs 10^3.4 copies/ml, p=0.004). Serotypes 1 (10^4.6 copies/ml, 95% CI 10^3.4-5.9) and 6B (10^4.6 copies/ml, 95% CI 10^3.9-5.3) had the highest mean density. Serotypes 4 (10^2.8 copies/ml, 95% CI 10^1.1-4.5) and 9N (10^2.9copies/ml, 95% CI 10^2.1-3.7) had the lowest mean carriage density.

Conclusions

Serotype 1, which causes the greatest proportion of invasive pneumococcal disease in this setting, was found to have the highest carriage density. Further evaluation of the PCV10 impact on carriage density is needed.

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THE IMPACT OF PNEUMOCOCCAL CONJUGATE VACCINE INTRODUCTION IN NEPAL: A SIX-YEAR PAEDIATRIC SURVEILLANCE STUDY (ID 516)

Abstract

Background

S. pneumoniae is a major cause of bacterial pneumonia and an important cause of invasive bacterial disease (IBD) in children under-five years of age in Nepal. Pneumococcal conjugate vaccine, PCV10, was introduced in 2015 with a 2+1 schedule.

Methods

We assessed the programmatic impact of PCV10 introduction using surveillance for nasopharyngeal (NP) colonisation, pneumonia and IBD. NP swabs from pneumonia inpatients and from healthy children, blood cultures from inpatients with suspected IBD, and chest x-rays from inpatient pneumonia cases were obtained over a 6-year period (2014-2019).

Results

The proportion of pneumonia cases with radiographic endpoint-consolidation (likely bacterial) was 34% lower (95%CI 19-46%) in 2018 compared with the pre-vaccine period (2014-2015). Vaccine serotype (VT) carriage in children under 2-years of age with pneumonia in 2019 was 78% lower (95%CI 30-93%) than in the pre-vaccine period.

Among healthy 6-23 month old children (urban and rural cohorts), VT-carriage declined 74% (95%CI 43-82%) by 2019. An increase in PCV13-additional-serotype carriage was seen in 2018 among rural-children (prevalence-ratio 1.65, 95%CI 1.17-2.32), but not urban-children.

Serotype 1 remains the dominant serotype detected in cases of invasive pneumococcal disease.

Conclusions

A decrease in prevalence of endpoint-consolidation-pneumonia and a decrease in vaccine-serotype circulation have been observed post PCV introduction in Nepal.

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