Microbial Genomics - Online First
Articles are published here in their final form, ahead of their inclusion in a final online issue. All articles are published as and when they are approved for publication, complete with supplementary material and supporting data. Each article is citable using its assigned Digital Object Identifier (DOI). This final published version of the article has undergone the full production process including copy editing, typesetting and proof correction. Once the article is included in an issue, the final, paginated Version of Record replaces this version.
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Escherichia coli EC93 deploys two plasmid-encoded class I contact-dependent growth inhibition systems for antagonistic bacterial interactions
Available online: 01 March 2021More LessThe phenomenon of contact-dependent growth inhibition (CDI) and the genes required for CDI (cdiBAI) were identified and isolated in 2005 from an Escherichia coli isolate (EC93) from rats. Although the cdiBAI EC93 locus has been the focus of extensive research during the past 15 years, little is known about the EC93 isolate from which it originates. Here we sequenced the EC93 genome and find two complete and functional cdiBAI loci (including the previously identified cdi locus), both carried on a large 127 kb plasmid. These cdiBAI systems are differentially expressed in laboratory media, enabling EC93 to outcompete E. coli cells lacking cognate cdiI immunity genes. The two CDI systems deliver distinct effector peptides that each dissipate the membrane potential of target cells, although the two toxins display different toxic potencies. Despite the differential expression and toxic potencies of these CDI systems, both yielded similar competitive advantages against E. coli cells lacking immunity. This can be explained by the fact that the less expressed cdiBAI system (cdiBAIEC93-2 ) delivers a more potent toxin than the highly expressed cdiBAIEC93-1 system. Moreover, our results indicate that unlike most sequenced CDI+ bacterial isolates, the two cdi loci of E. coli EC93 are located on a plasmid and are expressed in laboratory media.
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Genomic evolution and local epidemiology of Klebsiella pneumoniae from a major hospital in Beijing, China, over a 15 year period: dissemination of known and novel high-risk clones
Available online: 25 February 2021More LessKlebsiella pneumoniae is a frequent cause of nosocomial and severe community-acquired infections. Multidrug-resistant (MDR) and hypervirulent (hv) strains represent major threats, and tracking their emergence, evolution and the emerging convergence of MDR and hv traits is of major importance. We employed whole-genome sequencing (WGS) to study the evolution and epidemiology of a large longitudinal collection of clinical K. pneumoniae isolates from the H301 hospital in Beijing, China. Overall, the population was highly diverse, although some clones were predominant. Strains belonging to clonal group (CG) 258 were dominant, and represented the majority of carbapenemase-producers. While CG258 strains showed high diversity, one clone, ST11-KL47, represented the majority of isolates, and was highly associated with the KPC-2 carbapenemase and several virulence factors, including a virulence plasmid. The second dominant clone was CG23, which is the major hv clone globally. While it is usually susceptible to multiple antibiotics, we found some isolates harbouring MDR plasmids encoding for ESBLs and carbapenemases. We also reported the local emergence of a recently described high-risk clone, ST383. Conversely to strains belonging to CG258, which are usually associated to KPC-2, ST383 strains seem to readily acquire carbapenemases of different types. Moreover, we found several ST383 strains carrying the hypervirulence plasmid. Overall, we detected about 5 % of simultaneous carriage of AMR genes (ESBLs or carbapenemases) and hypervirulence genes. Tracking the emergence and evolution of such strains, causing severe infections with limited treatment options, is fundamental in order to understand their origin and evolution and to limit their spread. This article contains data hosted by Microreact.
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Genomic relatedness and clinical significance of Streptococcus mitis strains isolated from the urogenital tract of sexual partners
Available online: 25 February 2021More LessResearch into the lower urinary tract (LUT) microbiota has primarily focused on its relationship to LUT symptoms (LUTS), taking snapshots of these communities in individuals with and without LUTS. While certain bacterial taxa have been associated with LUTS, or the lack thereof, the temporal dynamics of this community were largely unknown. Recently, we conducted a longitudinal study and found that vaginal intercourse resulted in a shift in species richness and diversity within the LUT microbiota. This is particularly relevant as frequent vaginal intercourse is a major risk factor for urinary tract infection (UTI) in premenopausal women (Aydin et al. Int Urogynecol J 2015;26:795–804). To further investigate the relationship between vaginal intercourse and LUT microbiota, here we present the results of a 3 week study in which daily urogenital specimens were collected from a female participant and her male sexual partner. Consistent with our previous findings, the LUT microbiota changed after vaginal intercourse, most notably a high abundance of Streptococcus mitis was observed post-coitus. We isolated and sequenced S. mitis from both sexual partners finding that: (i) the S. mitis isolates from the female partner’s urogenital tract were genomically similar throughout the duration of the study, and (ii) they were related to one isolate from the male partner’s oral cavity collected at the end of the study, suggesting transmission between the two individuals. We hypothesize that blooms in S. mitis after vaginal intercourse may play a role in coitus-related UTI. We found that a S. mitis isolate, in contrast to a Lactobacillus jensenii isolate displaced after vaginal intercourse, cannot inhibit the growth of uropathogenic Escherichia coli . Thus, this bloom in S. mitis may provide a window of opportunity for a uropathogen to colonize the LUT.
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Comprehensive in silico survey of the Mycolicibacterium mobilome reveals an as yet underexplored diversity
Available online: 23 February 2021More LessThe mobilome plays a crucial role in bacterial adaptation and is therefore a starting point to understand and establish the gene flow occurring in the process of bacterial evolution. This is even more so if we consider that the mobilome of environmental bacteria can be the reservoir of genes that may later appear in the clinic. Recently, new genera have been proposed in the family Mycobacteriaceae , including the genus Mycolicibacterium , which encompasses dozens of species of agricultural, biotechnological, clinical and ecological importance, being ubiquitous in several environments. The current scenario in the Mycobacteriaceae mobilome has some bias because most of the characterized mycobacteriophages were isolated using a single host strain, and the few plasmids reported mainly relate to the genus Mycobacterium . To fill in the gaps in these issues, we performed a systematic in silico study of these mobile elements based on 242 available genomes of the genus Mycolicibacterium . The analyses identified 156 putative plasmids (19 conjugative, 45 mobilizable and 92 non-mobilizable) and 566 prophages in 86 and 229 genomes, respectively. Moreover, a contig was characterized by resembling an actinomycete integrative and conjugative element (AICE). Within this diversity of mobile genetic elements, there is a pool of genes associated with several canonical functions, in addition to adaptive traits, such as virulence and resistance to antibiotics and metals (mercury and arsenic). The type-VII secretion system was a common feature in the predicted plasmids, being associated with genes encoding virulent proteins (EsxA, EsxB, PE and PPE). In addition to the characterization of plasmids and prophages of the family Mycobacteriaceae , this study showed an abundance of these genetic elements in a dozen species of the genus Mycolicibacterium .
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Genomic epidemiology of a Cryptococcus neoformans case cluster in Glasgow, Scotland, 2018
Available online: 23 February 2021More LessIn 2018, a cluster of two cases of cryptococcosis occurred at the Queen Elizabeth University Hospital (QEUH) in Glasgow, Scotland (UK). It was postulated that these cases may have been linked to pigeon droppings found on the hospital site, given there have been previous reports of Cryptococcus neoformans associated with pigeon guano. Although some samples of pigeon guano taken from the site yielded culturable yeast from genera related to Cryptococcus, they have since been classified as Naganishia or Papiliotrema spp., and no isolates of C. neoformans were recovered from either the guano or subsequent widespread air sampling. In an attempt to further elucidate any possible shared source of the clinical isolates, we used whole-genome sequencing and phylogenetic analysis to examine the relationship of the two Cryptococcus isolates from the QEUH cases, along with two isolates from sporadic cases treated at a different Glasgow hospital earlier in 2018. Our work demonstrated that these four clinical isolates were not clonally related; while all isolates were from the VNI global lineage and of the same mating type (MATα), the genotypes of the two QEUH isolates were separated by 1885 base changes and belonged to different sub-lineages, recently described as the intercontinental sub-clades VNIa-93 and VNIa-5. In contrast, one of the two sporadic 2018 clinical isolates was determined to belong to the VNIb sub-lineage and the other classified as a VNIV/VNI hybrid. Our work demonstrated that the two 2018 QEUH isolates and the two prior C. neoformans clinical isolates were all genetically distinct. It was not possible to determine whether the QEUH genotypes stemmed from independent sources or from the same source, i.e. pigeons carrying different genotypes, but it should be noted that whilst members of allied genera within the Tremellomycetes were isolated from the hospital environment, there were no environmental isolations of C. neoformans.
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Comparative genomics of ST5 and ST30 methicillin-resistant Staphylococcus aureus sequential isolates recovered from paediatric patients with cystic fibrosis
Available online: 18 February 2021More LessStaphylococcus aureus chronic airway infection in patients with cystic fibrosis (CF) allows this pathogen to adapt over time in response to different selection pressures. We have previously shown that the main sequence types related to community-acquired methicillin-resistant S. aureus (MRSA) infections in Argentina – ST5 and ST30 – are also frequently isolated from the sputum of patients with CF, but in these patients they usually display multi-drug antimicrobial resistance. In this study, we sequenced the genomes of MRSA from four paediatric CF patients with the goal of identifying mutations among sequential isolates, especially those possibly related to antimicrobial resistance and virulence, which might contribute to the adaptation of the pathogen in the airways of patients with CF. Our results revealed genetic differences in sequential MRSA strains isolated from patients with CF in both their core and accessory genomes. Although the genetic adaptation of S. aureus was distinct in different hosts, we detected independent mutations in thyA, htrA, rpsJ and gyrA – which are known to have crucial roles in S. aureus virulence and antimicrobial resistance – in isolates recovered from multiple patients. Moreover, we identified allelic variants that were detected in all of the isolates recovered after a certain time point; these non-synonymous mutations were in genes associated with antimicrobial resistance, virulence, iron scavenging and oxidative stress resistance. In conclusion, our results provide evidence of genetic variability among sequential MRSA isolates that could be implicated in the adaptation of these strains during chronic CF airway infection.
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Genomic surveillance, characterization and intervention of a polymicrobial multidrug-resistant outbreak in critical care
Available online: 18 February 2021More LessBackground. Infections caused by carbapenem-resistant Acinetobacter baumannii (CR-Ab) have become increasingly prevalent in clinical settings and often result in significant morbidity and mortality due to their multidrug resistance (MDR). Here we present an integrated whole-genome sequencing (WGS) response to a persistent CR-Ab outbreak in a Brisbane hospital between 2016–2018.
Methods. A. baumannii, Klebsiella pneumoniae, Serratia marcescens and Pseudomonas aeruginosa isolates were sequenced using the Illumina platform primarily to establish isolate relationships based on core-genome SNPs, MLST and antimicrobial resistance gene profiles. Representative isolates were selected for PacBio sequencing. Environmental metagenomic sequencing with Illumina was used to detect persistence of the outbreak strain in the hospital.
Results. In response to a suspected polymicrobial outbreak between May to August of 2016, 28 CR-Ab (and 21 other MDR Gram-negative bacilli) were collected from Intensive Care Unit and Burns Unit patients and sent for WGS with a 7 day turn-around time in clinical reporting. All CR-Ab were sequence type (ST)1050 (Pasteur ST2) and within 10 SNPs apart, indicative of an ongoing outbreak, and distinct from historical CR-Ab isolates from the same hospital. Possible transmission routes between patients were identified on the basis of CR-Ab and K. pneumoniae SNP profiles. Continued WGS surveillance between 2016 to 2018 enabled suspected outbreak cases to be refuted, but a resurgence of the outbreak CR-Ab mid-2018 in the Burns Unit prompted additional screening. Environmental metagenomic sequencing identified the hospital plumbing as a potential source. Replacement of the plumbing and routine drain maintenance resulted in rapid resolution of the secondary outbreak and significant risk reduction with no discernable transmission in the Burns Unit since.
Conclusion. We implemented a comprehensive WGS and metagenomics investigation that resolved a persistent CR-Ab outbreak in a critical care setting.
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Emerging carbapenem-resistant Klebsiella pneumoniae sequence type 16 causing multiple outbreaks in a tertiary hospital in southern Vietnam
Available online: 10 February 2021More LessThe emergence of carbapenem resistance in Klebsiella pneumoniae represents a major global public health concern. Nosocomial outbreaks caused by multidrug-resistant K. pneumoniae are commonly reported to result in high morbidity and mortality due to limited treatment options. Between October 2019 and January 2020, two concurrent high-mortality nosocomial outbreaks occurred in a referral hospital in Ho Chi Minh City, Vietnam. We performed genome sequencing and phylogenetic analysis of eight K. pneumoniae isolates from infected patients and two environmental isolates for outbreak investigation. We identified two outbreaks caused by two distinct lineages of the international sequence type (ST) 16 clone, which displayed extensive drug resistance, including resistance to carbapenem and colistin. Carbapenem-resistant ST16 outbreak strains clustered tightly with previously described ST16 K. pneumoniae from other hospitals in Vietnam, suggesting local persistence and transmission of this particular clone in this setting. We found environmental isolates from a hospital bed and blood pressure cuff that were genetically linked to an outbreak case cluster, confirming the potential of high-touch surfaces as sources for nosocomial spread of K. pneumoniae . Further, we found colistin resistance caused by disruption of the mgrB gene by an ISL3-like element, and carbapenem resistance mediated by a transferable IncF/bla OXA-181 plasmid carrying the ISL3-like element. Our study highlights the importance of coordinated efforts between clinical and molecular microbiologists and infection control teams to rapidly identify, investigate and contain nosocomial outbreaks. Routine surveillance with advanced sequencing technology should be implemented to strengthen hospital infection control and prevention measures.
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Genera specific distribution of DEAD-box RNA helicases in cyanobacteria
Available online: 04 February 2021More LessAlthough RNA helicases are essentially ubiquitous and perform roles in all stages of RNA metabolism, phylogenetic analysis of the DEAD (Asp-Glu-Ala-Asp)-box RNA helicase family in a single phylum has not been performed. Here, we performed a phylogenetic analysis on DEAD-box helicases from all currently available cyanobacterial genomes, comprising a total of 362 helicase protein sequences from 280 strains. DEAD-box helicases belonging to three distinct clades were observed. Two clades, the CsdA (cold shock DEAD-box A)-like and RhlE (RNA helicase E)-like helicases, cluster with the homologous proteins from Escherichia coli . The third clade, the CrhR (cyanobacterial RNA helicase Redox)-like helicases, is unique to cyanobacteria and characterized by a conserved sequence motif in the C-terminal extension. Restricted distribution is observed across cyanobacterial diversity with respect to both helicase type and strain. CrhR-like and CsdA-like helicases essentially never occur together, while RhlE always occurs with either a CrhR-like or CsdA-like helicase. CrhR-like and RhlE-like proteins occurred in filamentous cyanobacteria of the orders Nostocales , Oscillatoriales and Synechococcales . Similarly, CsdA- and RhlE-like proteins are restricted to unicellular cyanobacteria of the genera Cyanobium and Synechococcus . In addition, the unexpected occurrence of RhlE in two Synechococcus strains suggests recent acquisition and evolutionary divergence. This study, therefore, raises physiological and evolutionary questions as to why DEAD-box RNA helicases encoded in cyanobacterial lineages display restricted distributions, suggesting niches that require either CrhR or CsdA RNA helicase activity but not both. Extensive conservation of gene synteny surrounding the previously described rimO–crhR operon is also observed, indicating a role in the maintenance of photosynthesis. The analysis provides insights into the evolution, origin and dissemination of sequences within a single gene family to yield divergent functional roles.
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From genotype to phenotype: adaptations of Pseudomonas aeruginosa to the cystic fibrosis environment
Available online: 02 February 2021More LessPseudomonas aeruginosa is one of the main microbial species colonizing the lungs of cystic fibrosis patients and is responsible for the decline in respiratory function. Despite the hostile pulmonary environment, P. aeruginosa is able to establish chronic infections thanks to its strong adaptive capacity. Various longitudinal studies have attempted to compare the strains of early infection with the adapted strains of chronic infection. Thanks to new ‘-omics’ techniques, convergent genetic mutations, as well as transcriptomic and proteomic dysregulations have been identified. As a consequence of this evolution, the adapted strains of P. aeruginosa have particular phenotypes that promote persistent infection.
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Exact mapping of Illumina blind spots in the Mycobacterium tuberculosis genome reveals platform-wide and workflow-specific biases
Available online: 27 January 2021More LessWhole-genome sequencing (WGS) is fundamental to Mycobacterium tuberculosis basic research and many clinical applications. Coverage across Illumina-sequenced M. tuberculosis genomes is known to vary with sequence context, but this bias is poorly characterized. Here, through a novel application of phylogenomics that distinguishes genuine coverage bias from deletions, we discern Illumina ‘blind spots’ in the M. tuberculosis reference genome for seven sequencing workflows. We find blind spots to be widespread, affecting 529 genes, and provide their exact coordinates, enabling salvage of unaffected regions. Fifty-seven pe/ppe genes (the primary families assumed to exhibit Illumina bias) lack blind spots entirely, while the remaining pe/ppe genes account for 55.1 % of blind spots. Surprisingly, we find coverage bias persists in homopolymers as short as 6 bp, shorter tracts than previously reported. While G+C-rich regions challenge all Illumina sequencing workflows, a modified Nextera library preparation that amplifies DNA with a high-fidelity polymerase markedly attenuates coverage bias in G+C-rich and homopolymeric sequences, expanding the ‘Illumina-sequenceable’ genome. Through these findings, and by defining workflow-specific exclusion criteria, we spotlight effective strategies for handling bias in M. tuberculosis Illumina WGS. This empirical analysis framework may be used to systematically evaluate coverage bias in other species using existing sequencing data.
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A high prevalence of bla OXA-48 in Klebsiella (Raoultella) ornithinolytica and related species in hospital wastewater in South West England
Available online: 08 January 2021More LessKlebsiella species occupy a wide range of environmental and animal niches, and occasionally cause opportunistic infections that are resistant to multiple antibiotics. In particular, Klebsiella pneumoniae (Kpne) has gained notoriety as a major nosocomial pathogen, due principally to the rise in non-susceptibility to carbapenems and other beta-lactam antibiotics. Whilst it has been proposed that the urban water cycle facilitates transmission of pathogens between clinical settings and the environment, the level of risk posed by resistant Klebsiella strains in hospital wastewater remains unclear. We used whole genome sequencing (WGS) to compare Klebsiella species in contemporaneous samples of wastewater from an English hospital and influent to the associated wastewater treatment plant (WWTP). As we aimed to characterize representative samples of Klebsiella communities, we did not actively select for antibiotic resistance (other than for ampicillin), nor for specific Klebsiella species. Two species, Kpne and K. ( Raoultella ) ornithinolytica (Korn), were of equal dominance in the hospital wastewater, and four other Klebsiella species were present in low abundance in this sample. In contrast, despite being the species most closely associated with healthcare settings, Kpne was the dominant species within the WWTP influent. In total, 29 % of all isolates harboured the bla OXA-48 gene on a pOXA-48-like plasmid, and these isolates were almost exclusively recovered from the hospital wastewater. This gene was far more common in Korn (68 % of isolates) than in Kpne (3.4 % of isolates). In general plasmid-borne, but not chromosomal, resistance genes were significantly enriched in the hospital wastewater sample. These data implicate hospital wastewater as an important reservoir for antibiotic-resistant Klebsiella , and point to an unsuspected role of species within the Raoultella group in the maintenance and dissemination of plasmid-borne bla OXA-48. This article contains data hosted by Microreact.
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Plasmids encode niche-specific traits in Lactobacillaceae
Available online: 09 November 2020More LessSpecies belonging to the family Lactobacillaceae are found in highly diverse environments and play an important role in fermented foods and probiotic products. Many of these species have been individually reported to harbour plasmids that encode important genes. In this study, we performed comparative genomic analysis of publicly available data for 512 plasmids from 282 strains represented by 51 species of this family and correlated the genomic features of plasmids with the ecological niches in which these species are found. Two-thirds of the species had at least one plasmid-harbouring strain. Plasmid abundance and GC content were significantly lower in vertebrate-adapted species as compared to nomadic and free-living species. Hierarchical clustering highlighted the distinct nature of plasmids from the nomadic and free-living species than those from the vertebrate-adapted species. EggNOG-assisted functional annotation revealed that genes associated with transposition, conjugation, DNA repair and recombination, exopolysaccharide production, metal ion transport, toxin–antitoxin system, and stress tolerance were significantly enriched on the plasmids of the nomadic and in some cases nomadic and free-living species. On the other hand, genes related to anaerobic metabolism, ABC transporters and the major facilitator superfamily were overrepresented on the plasmids of the vertebrate-adapted species. These genomic signatures correlate with the comparatively nutrient-depleted, stressful and dynamic environments of nomadic and free-living species and nutrient-rich and anaerobic environments of vertebrate-adapted species. Thus, these results indicate the contribution of the plasmids in the adaptation of lactobacilli to their respective habitats. This study also underlines the potential application of these plasmids in improving the technological and probiotic properties of lactic acid bacteria.
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Whole-genome sequencing resolves a polyclonal outbreak by extended-spectrum beta-lactam and carbapenem-resistant Klebsiella pneumoniae in a Portuguese tertiary-care hospital
Available online: 01 April 2020More LessKlebsiella pneumoniae has emerged as an important nosocomial pathogen, with whole-genome sequencing (WGS) significantly improving our ability to characterize associated outbreaks. Our study sought to perform a genome-wide analysis of multiclonal K. pneumoniae isolates (n=39; 23 patients) producing extended spectrum beta-lactamases and/or carbapenemases sourced between 2011 and 2016 in a Portuguese tertiary-care hospital. All isolates showed resistance to third-generation cephalosporins and six isolates (five patients) were also carbapenem resistant. Genome-wide-based phylogenetic analysis revealed a topology representing ongoing dissemination of three main sequence-type (ST) clades (ST15, ST147 and ST307) and transmission across different wards, compatible with missing links that can take the form of undetected colonized patients. Two carbapenemase-coding genes were detected: blaKPC-3 , located on a Tn4401d transposon, and blaGES-5 on a novel class 3 integron. Additionally, four genes coding for ESBLs (blaBEL-1 , blaCTX-M-8 , blaCTX-M-15 and blaCTX-M-32 ) were also detected. ESBL horizontal dissemination across five clades is highlighted by the similar genetic environments of blaCTX-M-15 gene upstream of ISEcp1 on a Tn3-like transposon. Overall, this study provides a high-resolution genome-wide perspective on the epidemiology of ESBL and carbapenemase-producing K. pneumoniae in a healthcare setting while contributing for the adoption of appropriate intervention and prevention strategies.
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Enhancing genomics-based outbreak detection of endemic Salmonella enterica serovar Typhimurium using dynamic thresholds
Available online: 04 November 2019More LessSalmonella enterica serovar Typhimurium is the leading cause of salmonellosis in Australia, and the ability to identify outbreaks and their sources is vital to public health. Here, we examined the utility of whole-genome sequencing (WGS), including complete genome sequencing with Oxford Nanopore technologies, in examining 105 isolates from an endemic multi-locus variable number tandem repeat analysis (MLVA) type over 5 years. The MLVA type was very homogeneous, with 90 % of the isolates falling into groups with a five SNP cut-off. We developed a new two-step approach for outbreak detection using WGS. The first clustering at a zero single nucleotide polymorphism (SNP) cut-off was used to detect outbreak clusters that each occurred within a 4 week window and then a second clustering with dynamically increased SNP cut-offs were used to generate outbreak investigation clusters capable of identifying all outbreak cases. This approach offered optimal specificity and sensitivity for outbreak detection and investigation, in particular of those caused by endemic MLVA types or clones with low genetic diversity. We further showed that inclusion of complete genome sequences detected no additional mutational events for genomic outbreak surveillance. Phylogenetic analysis found that the MLVA type was likely to have been derived recently from a single source that persisted over 5 years, and seeded numerous sporadic infections and outbreaks. Our findings suggest that SNP cut-offs for outbreak cluster detection and public-health surveillance should be based on the local diversity of the relevant strains over time. These findings have general applicability to outbreak detection of bacterial pathogens.
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Molecular characterization of respiratory syncytial viruses circulating in a paediatric cohort in Amman, Jordan
Available online: 18 September 2019More LessRespiratory syncytial viruses (RSVs) are an important cause of mortality worldwide and a major cause of respiratory tract infections in children, driving development of vaccine candidates. However, there are large gaps in our knowledge of the local evolutionary and transmission dynamics of RSVs, particularly in understudied regions such as the Middle East. To address this gap, we sequenced the complete genomes of 58 RSVA and 27 RSVB samples collected in a paediatric cohort in Amman, Jordan, between 2010 and 2013. RSVA and RSVB co-circulated during each winter epidemic of RSV in Amman, and each epidemic comprised multiple independent viral introductions of RSVA and RSVB. However, RSVA and RSVB alternated in dominance across years, potential evidence of immunological interactions. Children infected with RSVA tended to be older than RSVB-infected children [30 months versus 22.4 months, respectively (P value = 0.02)], and tended to developed bronchopneumonia less frequently than those with RSVB, although the difference was not statistically significant (P value = 0.06). Differences in spatial patterns were investigated, and RSVA lineages were often identified in multiple regions in Amman, whereas RSVB introductions did not spread beyond a single region of the city, although these findings were based on small sample sizes. Multiple RSVA genotypes were identified in Amman, including GA2 viruses as well as three viruses from the ON1 sub-genotype that emerged in 2009 and are now the dominant genotype circulating worldwide. As vaccine development advances, further sequencing of RSV is needed to understand viral ecology and transmission, particularly in under-studied locations.
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