Respiratory carriage of hypervirulent Klebsiella pneumoniae by indigenous populations of Malaysia.

Klebsiella pneumoniae is a Gram-negative Enterobacteriaceae that is classified by the World Health Organisation (WHO) as a Priority One ESKAPE pathogen. South and Southeast Asian countries are regions where both healthcare associated infections (HAI) and community acquired infections (CAI) due to extended-spectrum ß-lactamase (ESBL)-producing and carbapenem-resistant K. pneumoniae (CRKp) are of concern. As K. pneumoniae can also exist as a harmless commensal, the spread of resistance genotypes requires epidemiological vigilance. However there has been no significant study of carriage isolates from healthy individuals, particularly in Southeast Asia, and specially Malaysia. Here we describe the genomic analysis of respiratory isolates of K. pneumoniae obtained from Orang Ulu and Orang Asli communities in Malaysian Borneo and Peninsular Malaysia respectively. The majority of isolates were K. pneumoniae species complex (KpSC) 1 K. pneumoniae (n = 53, 89.8%). Four Klebsiella variicola subsp. variicola (KpSC3) and two Klebsiella quasipneumoniae subsp. similipneumoniae (KpSC4) were also found. It was discovered that 30.2% (n = 16) of the KpSC1 isolates were ST23, 11.3% (n = 6) were of ST65, 7.5% (n = 4) were ST13, and 13.2% (n = 7) were ST86. Only eight of the KpSC1 isolates encoded ESBL, but importantly not carbapenemase. Thirteen of the KpSC1 isolates carried yersiniabactin, colibactin and aerobactin, all of which harboured the rmpADC locus and are therefore characterised as hypervirulent. Co-carriage of multiple strains was minimal. In conclusion, most isolates were KpSC1, ST23, one of the most common sequence types and previously found in cases of K. pneumoniae infection. A proportion were hypervirulent (hvKp) however antibiotic resistance was low.

The characterization of Moraxella catarrhalis carried in the general population.

Moraxella catarrhalis is a common cause of respiratory tract infection, particularly otitis media in children, whilst it is also associated with the onset of exacerbation in chronic obstructive pulmonary disease in adults. Despite the need for an efficacious vaccine against M. catarrhalis, no candidates have progressed to clinical trial. This study, therefore, aimed to characterize the diversity of M. catarrhalis isolated from the upper respiratory tract of healthy children and adults, to gain a better understanding of the epidemiology of M. catarrhalis and the distribution of genes associated with virulence factors, to aid vaccine efforts. Isolates were sequenced and the presence of target genes reported. Contrary to prevailing data, this study found that lipooligosaccharide (LOS) B serotypes are not exclusively associated with 16S type 1. In addition, a particularly low prevalence of LOS B and high prevalence of LOS C serotypes was observed. M. catarrhalis isolates showed low prevalence of antimicrobial resistance and a high gene prevalence for a number of the target genes investigated: ompB2 (also known as copB), ompCD, ompE, ompG1a, ompG1b, mid (also known as hag), mcaP, m35, tbpA, lbpA, tbpB, lbpB, msp22, msp75 and msp78, afeA, pilA, pilQ, pilT, mod, oppA, sbp2, mcmA and mclS.

Carriage of upper respiratory tract pathogens in rural communities of Sarawak, Malaysian Borneo.

INTRODUCTION: Pneumonia is a leading cause of death in Malaysia. Whilst many studies have reported the aetiology of pneumonia in Western countries, the epidemiology of pneumonia in Malaysia remains poorly understood. As carriage is a prerequisite for disease, we sought to improve our understanding of the carriage and antimicrobial resistance (AMR) of respiratory tract pathogens in Malaysia. The rural communities of Sarawak are an understudied part of the Malaysian population and were the focus of this study, allowing us to gain a better understanding of bacterial epidemiology in this population. METHODS: A population-based survey of bacterial carriage was undertaken in participants of all ages from rural communities in Sarawak, Malaysia. Nasopharyngeal, nasal, mouth and oropharyngeal swabs were taken. Bacteria were isolated from each swab and identified by culture-based methods and antimicrobial susceptibility testing conducted by disk diffusion or E test. RESULTS: 140 participants were recruited from five rural communities. Klebsiella pneumoniae was most commonly isolated from participants (30.0%), followed by Staphylococcus aureus (20.7%), Streptococcus pneumoniae (10.7%), Haemophilus influenzae (9.3%), Moraxella catarrhalis (6.4%), Pseudomonas aeruginosa (6.4%) and Neisseria meningitidis (5.0%). Of the 21 S. pneumoniae isolated, 33.3 and 14.3% were serotypes included in the 13 valent PCV (PCV13) and 10 valent PCV (PCV10) respectively. 33.8% of all species were resistant to at least one antibiotic, however all bacterial species except S. pneumoniae were susceptible to at least one type of antibiotic. CONCLUSION: To our knowledge, this is the first bacterial carriage study undertaken in East Malaysia. We provide valuable and timely data regarding the epidemiology and AMR of respiratory pathogens commonly associated with pneumonia. Further surveillance in Malaysia is necessary to monitor changes in the carriage prevalence of upper respiratory tract pathogens and the emergence of AMR, particularly as PCV is added to the National Immunisation Programme (NIP).

The upper respiratory tract microbiome of indigenous Orang Asli in north-eastern Peninsular Malaysia.

Much microbiome research has focused on populations that are predominantly of European descent, and from narrow demographics that do not capture the socio-economic and lifestyle differences which impact human health. Here we examined the airway microbiomes of the Orang Asli, the indigenous peoples of Malaysia. A total of 130 participants were recruited from two sites in the north-eastern state of Terengganu in Peninsular Malaysia. Using 16S rRNA sequencing, the nasal microbiome was significantly more diverse in those aged 5-17 years compared to 50+ years (p = 0.023) and clustered by age (PERMANOVA analysis of the Bray-Curtis distance, p = 0.001). Hierarchical clustering of Bray-Curtis dissimilarity scores revealed six microbiome clusters. The largest cluster (n = 28; 35.4%) had a marked abundance of Corynebacterium. In the oral microbiomes Streptococcus, Neisseria and Haemophilus were dominant. Using conventional microbiology, high levels of Staphylococcus aureus carriage were observed, particularly in the 18-65 age group (n = 17/36; 47.2% 95% CI: 30.9-63.5). The highest carriage of pneumococci was in the <5 and 5 to 17 year olds, with 57.1% (4/7) and 49.2% (30/61), respectively. Sixteen pneumococcal serotypes were identified, the most common being the nonvaccine-type 23A (14.6%) and the vaccine-type 6B (9.8%). The prevalence of pneumococcal serotypes covered by pneumococcal conjugate vaccines support introduction into a Malaysian national immunisation schedule. In addition, the dominance of Corynebacterium in the airway microbiomes is intriguing given their role as a potentially protective commensal with respect to acute infection and respiratory health.

Pneumococcal vaccine impacts on the population genomics of non-typeable Haemophilus influenzae.

The implementation of pneumococcal conjugate vaccines (PCVs) has led to a decline in vaccine-type disease. However, there is evidence that the epidemiology of non-typeable Haemophilus influenzae (NTHi) carriage and disease can be altered as a consequence of PCV introduction. We explored the epidemiological shifts in NTHi carriage using whole genome sequencing over a 5-year period that included PCV13 replacement of PCV7 in the UK's National Immunization Programme in 2010. Between 2008/09 and 2012/13 (October to March), nasopharyngeal swabs were taken from children <5 years of age. Significantly increased carriage post-PCV13 was observed and lineage-specific associations with Streptococcus pneumoniae were seen before but not after PCV13 introduction. NTHi were characterized into 11 discrete, temporally stable lineages, congruent with current knowledge regarding the clonality of NTHi. The increased carriage could not be linked to the expansion of a particular clone and different co-carriage dynamics were seen before PCV13 implementation when NTHi co-carried with vaccine serotype pneumococci. In summary, PCV13 introduction has been shown to have an indirect effect on NTHi epidemiology and there exists both negative and positive, distinct associations between pneumococci and NTHi. This should be considered when evaluating the impacts of pneumococcal vaccine design and policy.

Ecology and diversity in upper respiratory tract microbial population structures from a cross-sectional community swabbing study.

PURPOSE: Respiratory tract infections (RTIs) are responsible for over 2.8 million deaths per year worldwide with pathobiont carriage a required precursor to infection. We sought to determine carriage epidemiology for both bacterial and viral respiratory pathogens as part of a large population-based cross-sectional carriage study. METHODOLOGY: Nose self-swab samples were collected in two separate time-points, May to August 2012 (late spring/summer) and February to April 2013 (winter/early spring). The presence of six bacterial species: S. pneumoniae, H. influenzae, M. catarrhalis, S. aureus, P. aeruginosa and N. meningitidis in addition to respiratory syncytial virus, influenza viruses A and B, rhinovirus/enterovirus, coronavirus, parainfluenza viruses 1-3 and adenovirus was determined using culture and PCR methods.Results/Key findings. Carriage was shown to vary with age, recent RTI and the presence of other species. Spatial structures of microbial communities were more disordered in the 0-4 age group and those with recent RTI. Species frequency distributions were flatter than random expectation in young individuals (X2=20.42, P=0.002), indicating spatial clumping of species consistent with facilitative relationships. Deviations from a neutral model of ecological niches were observed in summer samples and from older individuals but not in the winter or younger individuals (0-4 years), suggesting the presence of seasonal and age-dependent niche processes in respiratory community assembly. CONCLUSION: The application of epidemiological methods and ecological theory to respiratory tract samples has yielded novel insights into the factors that drive microbial community composition.

Secondary Bacterial Infections Associated with Influenza Pandemics.

Lower and upper respiratory infections are the fourth highest cause of global mortality (Lozano et al., 2012). Epidemic and pandemic outbreaks of respiratory infection are a major medical concern, often causing considerable disease and a high death toll, typically over a relatively short period of time. Influenza is a major cause of epidemic and pandemic infection. Bacterial co/secondary infection further increases morbidity and mortality of influenza infection, with Streptococcus pneumoniae, Haemophilus influenzae, and Staphylococcus aureus reported as the most common causes. With increased antibiotic resistance and vaccine evasion it is important to monitor the epidemiology of pathogens in circulation to inform clinical treatment and development, particularly in the setting of an influenza epidemic/pandemic.

The rise and fall of pneumococcal serotypes carried in the PCV era.

Streptococcus pneumoniae is a major cause of meningitis, sepsis and pneumonia worldwide. Vaccination using pneumococcal conjugate vaccines (PCV) has therefore been part of the UK's childhood immunisation programme since 2006. Here we describe pneumococcal carriage rates in children under five years of age attending the paediatric department of a large UK hospital in response to vaccine implementation over seven winter seasons from 2006 to 2013. S. pneumoniae (n=696) were isolated from nasopharyngeal swabs (n=2267) collected during seven consecutive winters, October to March, 2006/7 to 2012/13. This includes the period immediately following the introduction of the seven-valent pneumococcal conjugate vaccine (PCV7) in 2006 in addition to pre- and post-PCV13 introduction in 2010. We show a decrease in PCV13 vaccine serotypes (VT) in the three years following PCV13 vaccine implementation (2010/11 to 2012/13). Serotype 6A represented the only observed VT following PCV13 implementation with all others (including PCV7 serotypes) absent from carriage. Overall pneumococcal carriage, attributable to non-VT (NVT), was consistent across all sampling years with a mean of 31·1%. The ten most frequently isolated NVTs were 6C, 11A, 15B, 23B, 15A, 21, 22F, 35F, 23A and 15C. Fluctuations in the prevalence of each were however noted. Comparing prevalence at 2006/07 with 2012/13 only 15A was shown to have increased significantly (p value of 0·003) during the course of PCV implementation. These data support the increasing evidence that the primary effect of PCVs is due to population immunity by reducing or eliminating the carriage of invasive VT serotypes. With IPD being increasingly attributed to non-vaccine serotypes, surveillance of carriage data continues to act as an early warning system for vaccine design and public health policy that require continual data of both carried pneumococcal serotypes and IPD attributed serotype data.

Vaccination in Southeast Asia--reducing meningitis, sepsis and pneumonia with new and existing vaccines.

Streptococcus pneumoniae, Haemophilus influenzae type b and Neisseria meningitidis are leading causes of vaccine-preventable diseases such as meningitis, sepsis and pneumonia. Although there has been much progress in the introduction of vaccines against these pathogens, access to vaccines remains elusive in some countries. This review highlights the current S. pneumoniae, H. influenzae type b, and N. meningitidis immunization schedules in the 10 countries belonging to the Association of Southeast Asian Nations (ASEAN). Epidemiologic studies may be useful for informing vaccine policy in these countries, particularly when determining the cost-effectiveness of introducing new vaccines.