Gallbladder carriage generates genetic variation and genome degradation in Salmonella Typhi

Abstract

Despite recent advances in typhoid fever control, asymptomatic carriage of Salmonella Typhi in the gallbladder remains poorly understood. Aiming to understand if S . Typhi becomes genetically adapted for long-term colonisation in the gallbladder, we performed whole genome sequencing on a collection of S . Typhi isolated from the gallbladders of typhoid carriers. These sequences were compared to contemporaneously sampled sequences from organisms isolated from the blood of acute patients within the same population. We found that S . Typhi carriage was not restricted to any particular genotype or conformation of antimicrobial resistance genes, but was largely reflective of S . Typhi circulating in the general population. However, gallbladder isolates showed a higher genetic variability than acute isolates, with median pairwise SNP distances of 21 and 13 SNPs ( p = 2.8x10 -9 ), respectively. Within gallbladder isolates of the predominant H58 genotype, variation was associated with a higher prevalence of nonsense mutations. Notably, gallbladder isolates displayed a higher frequency of non-synonymous mutations in genes encoding hypothetical proteins, membrane lipoproteins, transport/binding proteins, surface antigens, and carbohydrate degradation. Specifically, we identified several gallbladder-specific non-synonymous mutations involved in LPS synthesis and modification, with some isolates lacking the Vi capsular polysaccharide vaccine target due to the 134Kb deletion of SPI-7. S . Typhi is under strong selective pressure in the human gallbladder, which may be reflected phylogenetically by long terminal branches that may distinguish organisms from chronic and acute infections. Our work shows that selective pressures asserted by the hostile environment of the human gallbladder generate new antigenic variants and raises questions regarding the role of carriage in the epidemiology of typhoid fever.

Multiple introductions of multidrug-resistant typhoid associated with acute infection and asymptomatic carriage, Kenya

“…This translates to a rate of 1.9x10 -7 genome-wide substitutions per site per year (95% HPD = 1.5x10 -7 -2.2x10 -7 ). The novel EA1 isolates from this study (accounting for 35% of cases and 37% of controls) were intermingled with those sequenced previously from Kenya and were genetically diverse (median pairwise distance ~16 SNPs, interquartile range [12][13][14][15][16][17][18][19][20][21][22][23][24][25][26][27].…”

“…Pairwise SNP distances were calculated from alignments using the dist.dna() function in the R package ape v5.3 17 . Non-synonymous mutations were detected using SNPPar (V0.4.2dev) 18 and grouped by function based on the gene in which they were found according to the S. Typhi functional classification scheme developed at the Sanger Institute (www.sanger.ac.uk) using the genome annotation of CT18 1,19 .…”

“…Recent reports of epidemics of typhoid fever in SSA suggest that the disease may be becoming more widespread in the region 1,15,[17][18][19][20] . In Kenya, the rapid growth of population has led to a huge rural-to-urban migration with people increasingly living in informal settlements where clean water and good sanitation are a major challenge 21,22 .…”

Multiple introductions of multidrug-resistant typhoid associated with acute infection and asymptomatic carriage, Kenya

“…Terminal branch lengths were extracted from phylogenies using R package ggtree (v2.2.4) 44 . Non-synonymous mutations occurring in terminal branches were detected using SNPPar (v0.4.2dev) 45 and grouped by function based on the gene in which they were found, according to the functional classification scheme in the genome annotation of S. Typhi CT18 8,36 .…”

“…Control of typhoid is impeded by asymptomatic carriage, which historically was estimated to account for 2-5% of individuals infected [6][7][8] . However, there is a paucity of recent data on the frequency of carriers in different settings including sub-Saharan Africa (SSA) as well as the extent to which they contribute to disease transmission 9 .…”