Bacterial shedding and serologic responses following an outbreak of Salmonella Typhi in an endemic cohort.

BACKGROUND: Salmonella enterica serovar Typhi (Salmonella Typhi) is the cause of typhoid fever. Salmonella Typhi may be transmitted through shedding in the stool, which can continue after recovery from acute illness. Shedding is detected by culturing stool, which is challenging to co-ordinate at scale. We hypothesised that sero-surveillance would direct us to those shedding Salmonella Typhi in stool following a typhoid outbreak. METHODS: In 2016 a typhoid outbreak affected one in four residents of a Nursing School in Malosa, Malawi. The Department of Health asked for assistance to identify nursing students that might spread the outbreak to other health facilities. We measured IgG antibody titres against Vi capsular polysaccharide (anti-Vi IgG) and IgM / IgG antibodies against H:d flagellin (anti-H:d) three and six months after the outbreak. We selected participants in the highest and lowest deciles for anti-Vi IgG titre (measured at visit one) and obtained stool for Salmonella culture and PCR. All participants reported whether they had experienced fever persisting for three days or more during the outbreak (in keeping with the WHO definitions of 'suspected typhoid'). We tested for salmonellae in the Nursing School environment. RESULTS: We obtained 320 paired serum samples from 407 residents. We cultured stool from 25 residents with high anti-Vi IgG titres and 24 residents with low titres. We did not recover Salmonella Typhi from stool; four stool samples yielded non-typhoidal salmonellae; one sample produced a positive PCR amplification for a Salmonella Typhi target. Median anti-Vi and anti-H:d IgG titres fell among participants who reported persistent fever. There was a smaller fall in anti-H:d IgG titres among participants who did not report persistent fever. Non-typhoidal salmonellae were identified in water sampled at source and from a kitchen tap. CONCLUSION: High titres of anti-Vi IgG did not identify culture-confirmed shedding of Salmonella Typhi. There was a clear serologic signal of recent typhoid exposure in the cohort, represented by waning IgG antibody titres over time. The presence of non-typhoidal salmonellae in drinking water indicates sub-optimal sanitation. Developing methods to detect and treat shedding remains an important priority to complement typhoid conjugate vaccination in efforts to achieve typhoid elimination.

Case-control investigation of invasive Salmonella disease in Malawi reveals no evidence of environmental or animal transmission of invasive strains, and supports human to human transmission.

BACKGROUND: Invasive Salmonella infections cause significant morbidity and mortality in Sub-Saharan Africa. However, the routes of transmission are uncertain. We conducted a case-control study of index-case and geographically-matched control households in Blantyre, Malawi, sampling Salmonella isolates from index cases, healthy people, animals, and the household environment. METHODOLOGY: Sixty index cases of human invasive Salmonella infection were recruited (March 2015-Oct 2016). Twenty-eight invasive Non-Typhoidal Salmonella (iNTS) disease and 32 typhoid patients consented to household sampling. Each index-case household was geographically matched to a control household. Extensive microbiological sampling included stool sampling from healthy household members, stool or rectal swabs from household-associated animals and boot-sock sampling of the household environment. FINDINGS: 1203 samples from 120 households, yielded 43 non-Typhoidal Salmonella (NTS) isolates from 25 households (overall sample positivity 3.6%). In the 28 iNTS patients, disease was caused by 3 STs of Salmonella Typhimurium, mainly ST313. In contrast, the isolates from households spanned 15 sequence types (STs). Two S. Typhimurium isolates from index cases closely matched isolates from their respective asymptomatic household members (2 and 3 SNP differences respectively). Despite the recovery of a diverse range of NTS, there was no overlap between the STs causing iNTS disease with any environmental or animal isolates. CONCLUSIONS: The finding of NTS strains from index cases that matched household members, coupled with lack of related animal or environmental isolates, supports a hypothesis of human to human transmission of iNTS infections in the household. The breadth of NTS strains found in animals and the household environment demonstrated the robustness of NTS sampling and culture methodology, and suggests a diverse ecology of Salmonella in this setting. Healthy typhoid (S. Typhi) carrier state was not detected. The lack of S. Typhi isolates from the household environment suggests that further methodological development is needed to culture S. Typhi from the environment.

Large-scale sequencing of SARS-CoV-2 genomes from one region allows detailed epidemiology and enables local outbreak management.

The COVID-19 pandemic has spread rapidly throughout the world. In the UK, the initial peak was in April 2020; in the county of Norfolk (UK) and surrounding areas, which has a stable, low-density population, over 3200 cases were reported between March and August 2020. As part of the activities of the national COVID-19 Genomics Consortium (COG-UK) we undertook whole genome sequencing of the SARS-CoV-2 genomes present in positive clinical samples from the Norfolk region. These samples were collected by four major hospitals, multiple minor hospitals, care facilities and community organizations within Norfolk and surrounding areas. We combined clinical metadata with the sequencing data from regional SARS-CoV-2 genomes to understand the origins, genetic variation, transmission and expansion (spread) of the virus within the region and provide context nationally. Data were fed back into the national effort for pandemic management, whilst simultaneously being used to assist local outbreak analyses. Overall, 1565 positive samples (172 per 100 000 population) from 1376 cases were evaluated; for 140 cases between two and six samples were available providing longitudinal data. This represented 42.6 % of all positive samples identified by hospital testing in the region and encompassed those with clinical need, and health and care workers and their families. In total, 1035 cases had genome sequences of sufficient quality to provide phylogenetic lineages. These genomes belonged to 26 distinct global lineages, indicating that there were multiple separate introductions into the region. Furthermore, 100 genetically distinct UK lineages were detected demonstrating local evolution, at a rate of ~2 SNPs per month, and multiple co-occurring lineages as the pandemic progressed. Our analysis: identified a discrete sublineage associated with six care facilities; found no evidence of reinfection in longitudinal samples; ruled out a nosocomial outbreak; identified 16 lineages in key workers which were not in patients, indicating infection control measures were effective; and found the D614G spike protein mutation which is linked to increased transmissibility dominates the samples and rapidly confirmed relatedness of cases in an outbreak at a food processing facility. The large-scale genome sequencing of SARS-CoV-2-positive samples has provided valuable additional data for public health epidemiology in the Norfolk region, and will continue to help identify and untangle hidden transmission chains as the pandemic evolves.

CoronaHiT: high-throughput sequencing of SARS-CoV-2 genomes.

We present CoronaHiT, a platform and throughput flexible method for sequencing SARS-CoV-2 genomes (≤ 96 on MinION or > 96 on Illumina NextSeq) depending on changing requirements experienced during the pandemic. CoronaHiT uses transposase-based library preparation of ARTIC PCR products. Method performance was demonstrated by sequencing 2 plates containing 95 and 59 SARS-CoV-2 genomes on nanopore and Illumina platforms and comparing to the ARTIC LoCost nanopore method. Of the 154 samples sequenced using all 3 methods, ≥ 90% genome coverage was obtained for 64.3% using ARTIC LoCost, 71.4% using CoronaHiT-ONT and 76.6% using CoronaHiT-Illumina, with almost identical clustering on a maximum likelihood tree. This protocol will aid the rapid expansion of SARS-CoV-2 genome sequencing globally.

Stepwise evolution of Salmonella Typhimurium ST313 causing bloodstream infection in Africa.

Bloodstream infections caused by nontyphoidal Salmonella are a major public health concern in Africa, causing ~49,600 deaths every year. The most common Salmonella enterica pathovariant associated with invasive nontyphoidal Salmonella disease is Salmonella Typhimurium sequence type (ST)313. It has been proposed that antimicrobial resistance and genome degradation has contributed to the success of ST313 lineages in Africa, but the evolutionary trajectory of such changes was unclear. Here, to define the evolutionary dynamics of ST313, we sub-sampled from two comprehensive collections of Salmonella isolates from African patients with bloodstream infections, spanning 1966 to 2018. The resulting 680 genome sequences led to the discovery of a pan-susceptible ST313 lineage (ST313 L3), which emerged in Malawi in 2016 and is closely related to ST313 variants that cause gastrointestinal disease in the United Kingdom and Brazil. Genomic analysis revealed degradation events in important virulence genes in ST313 L3, which had not occurred in other ST313 lineages. Despite arising only recently in the clinic, ST313 L3 is a phylogenetic intermediate between ST313 L1 and L2, with a characteristic accessory genome. Our in-depth genotypic and phenotypic characterization identifies the crucial loss-of-function genetic events that occurred during the stepwise evolution of invasive S. Typhimurium across Africa.