This study evaluated detection methods for Salmonella Typhi (S. Typhi) in the environment, to establish a novel pathway from field sampling to isolation of viable organisms and molecular confirmation from complex environmental samples, thus enabling environmental surveillance of typhoid.Methods and Results: Multiple media were assessed using clinical isolates from the Public Health England's (PHE) Culture collection. The culture pathway selected consisted of a primary 2% bile broth and secondary Selenite F broth, followed by modified Chromogenic Agar for Salmonella Esterase (mCASE). A qPCR assay was adapted from a validated S. Typhi PCR panel for confirmation of isolates, with comparison to biochemical and serological tests showing good specificity. Sampling locations in Blantyre, Malawi were used to compare sampling methods. Viable S. Typhi were isolated from a mixture of trap and grab river water samples on six occasions. Conclusions:Culture of viable S. Typhi from environmental samples was possible using effective capture and culture techniques.Significance and impact of study: Whilst several studies have attempted to detect S. Typhi from the environment, this is the first successful attempt to isolate the organism from river water since the 1980s. Supplementing clinical data with environmental screening offers the potential for enhanced surveillance, which might inform interventions and assess vaccination programmes.

BackgroundStudies have shown that households using sludge from human excreta for agriculture are at an increased risk of soil transmitted helminths. However, while use of ecological sanitation (EcoSan) latrines is increasing in most African countries including Malawi, few studies have been done to check whether use of such sludge could potentially increase the prevalence of helminthic infections among household members as a results of exposure to faecal sludge/compared to use of traditional latrines.MethodsA cross sectional study was done targeting households using EcoSan and traditional pit latrines. Samples were collected from both types of latrines in Chikwawa (rural) and Blantyre (urban) districts. These two districts have a high number of EcoSan latrines in southern region of Malawi. 156 latrines were sampled (n = 95 traditional; n = 61 EcoSan), and processed following standard guidelines using modified triple floatation method. Identification of helminth ova (Ascaris lumbricoides, hookworms, Trichuris trichiura, Taenia spp. and Diphyllobothrium latum) was done using standard microscopy methods. The difference between the prevalence and mean concentration of helminths between the two types of latrines was tested using Chi Square and t test respectively.ResultsOf the total latrines tested, 85.9% (n = 134) had at least one species of helminth while 84.6% (n = 132) had at least a STH, with 82.0% (n = 50) in EcoSan and 86.3% (n = 82) in traditional pit latrines. There was no significant difference between the prevalence of helminths in EcoSan and traditional pit latrines [χ2 = 0.43 (1), P = 0.5]. The prevalence of Ascaris lumbricoides was significantly higher in EcoSan than in traditional pit latrines [χ2 = 5.44 (1) p = 0.02] while prevalence of hookworms was significantly higher in traditional pit latrines than in EcoSan latrines [χ2 = 13.98 (1) p < 0.001]. The highest concentration of helminths per gram of faecal sludge was in traditional pit latrines [31.2 (95% CI 19.1–43.2)] than in EcoSan latrines [26.4 (95% CI 16.5–36.3)].ConclusionThere was no significant difference between overall prevalence of helminths between households using EcoSan and those using traditional pit latrines. However, Ascaris lumbricoides was significantly higher in households using EcoSan latrines. EcoSan users need awareness on safe ways of handling faecal sludge in order to reduce chances of reinfection from Ascaris lumbricoides. Further research should be undertaken on household members to identify those infected and potential routes of infection to enable preventive targeting.

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.

Background Invasive Salmonella infections are a major cause of morbidity and mortality in Sub-Saharan Africa (SSA), but the sources and transmission routes are uncertain. We investigated potential sources for cases of invasive disease by sampling healthy people, animals, and the environment in index-case and geographically-matched control households. Methods Sixty index cases of human invasive Salmonella infection were recruited (28 invasive Non-Typhoidal Salmonella (iNTS) disease, and 32 typhoid). 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. Results 1203 samples were taken from 120 households, yielding 43 Salmonella isolates from 25 households (overall sample positivity 3.6%). Isolates from households were all NTS and spanned 15 STs. iNTS disease was caused by 3 STs of Salmonella Typhimurium, mainly ST313. Two S. Typhimurium isolates from index cases closely matched isolates from their respective asymptomatic household members (2 and 3 SNPs different respectively). There was no overlap of STs causing iNTS disease with environmental or animal isolates, despite recovery of diverse NTS. Conclusions The finding of NTS strains from index cases matching household members, coupled with lack of overlap with either animal or environmental isolates, supports a hypothesis that healthy humans are the source of iNTS infections in the household. The breadth of NTS strains found in the household environment across all sites demonstrated the robustness of sampling and methodology to detect NTS, and suggests a diverse ecology of Salmonella in this setting. The lack of S. Typhi isolated from the household environment may suggest a need for further methodological development to culture sources of typhoid.