Laboratory and molecular surveillance of paediatric typhoidal Salmonella in Nepal: Antimicrobial resistance and implications for vaccine policy

Abstract

BackgroundChildren are substantially affected by enteric fever in most settings with a high burden of the disease, including Nepal. However pathogen population structure and transmission dynamics are poorly delineated in young children, the proposed target group for immunization programs. Here we present whole genome sequencing and antimicrobial susceptibility data on 198 S. Typhi and 66 S. Paratyphi A isolated from children aged 2 months to 15 years of age during blood culture surveillance at Patan Hospital, Nepal, 2008–2016.Principal findingsS. Typhi was the dominant agent and comprised several distinct genotypes, dominated by 4.3.1 (H58). The heterogeneity of genotypes in children under five was reduced compared to data from 2005–2006, attributable to ongoing clonal expansion of H58. Most isolates (86%) were non-susceptible to fluoroquinolones, associated mainly with S. Typhi H58 lineage II and S. Paratyphi A harbouring mutations in the quinolone resistance-determining region (QRDR); non-susceptible strains from these groups accounted for 50% and 25% of all isolates. Multi-drug resistance (MDR) was rare (3.5% of S. Typhi, 0 S. Paratyphi A) and restricted to chromosomal insertions of resistance genes in H58 lineage I strains. Temporal analyses revealed a shift in dominance from H58 Lineage I to H58 Lineage II, with the latter being significantly more common after 2010. Comparison to global data sets showed the local S. Typhi and S. Paratyphi A strains had close genetic relatives in other South Asian countries, indicating regional strain circulation. Multiple imports from India of ciprofloxacin-resistant H58 lineage II strains were identified, but these were rare and showed no evidence of clonal replacement of local S. Typhi.SignificanceThese data indicate that enteric fever in Nepal continues to be a major public health issue with ongoing inter- and intra-country transmission, and highlights the need for regional coordination of intervention strategies. The absence of a S. Paratyphi A vaccine is cause for concern, given its prevalence as a fluoroquinolone resistant enteric fever agent in this setting.

Population structure and antimicrobial resistance patterns of Salmonella Typhi isolates in Bangladesh from 2004 to 2016

“…For each SNP that passes these criteria in any one isolate, consensus base calls for the SNP locus were extracted from all genomes mapped, with those having phred quality scores under 20 being treated as unknown alleles and represented with a gap character. These SNPs were then used to assign isolates to previously defined genotypes according to an extended S. Typhi genotyping framework using the GenoTyphi python script, available at https://github.com/katholt/genotyphi [6, 21]. Unique SNPs defining novel genotypes and lineages detected in the present study were manually extracted from allele tables output by RedDog using R, with SNPs responsible for non-synonymous mutations in highly conserved genes prioritised for lineage definitions.…”

“…To provide global context, 1,560 additional S. Typhi genomes belonging to the genotypes found in Bangladesh [6, 7, 9, 14, 15, 21, 27] were subjected to both SNP calling and genotyping as described above, resulting in an alignment of 14,852 chromosomal SNPs.…”

“…ISMapper (v2.0) [40] was run with default parameters to screen for insertion sites the transposases IS 1 (accession number J01730) and IS 26 (accession number X00011), relative to the CT18 reference sequence in order to identify the location of these in the chromosome of each Bangladeshi S. Typhi genome. Point mutations in the QRDR of genes gyrA and parC associated with reduced susceptibility to fluoroquinolones [15] were determined using GenoTyphi [6, 21].…”

“…Multi-drug resistant (MDR) S . Typhi, defined as resistance to the first-line antibiotics ampicillin, chloramphenicol and trimethoprim-sulfamethoxazole, was first observed in the 1970s [6–9]. The more recent emergence of MDR S. Typhi with nalidixic acid resistance and reduced susceptibility to fluoroquinolones complicates treatment options.…”

“…H58 S. Typhi encoding nonsynonymous mutations in the quinolone resistance determining region (QRDR) of DNA gyrase genes gyrA and gyrB and DNA topoisomerase IV genes parC and parE have shown reduced susceptibility to fluoroquinolones [7, 15]. Studies on typhoid in Nepal have reported the evolution of a fluoroquinolone resistant H58 lineage II isolates carrying three QRDR mutations ( gyrA-S83F, gyrA-D87N , and parC-S80I ) responsible for the failure of a gatifloxacin treatment trial [6, 15]. More recently, in Bangladesh, a new H58 lineage I triple QRDR mutant carrying three different mutations ( gyrA-S83F, gyrA-D87G , and parC-E84K ) has been observed [14].…”