2022.09.26 iNTS household survey.pptx
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Presentation for ASTMH 2022 on invasive non-typhoidal Salmonella in Blantyre, Malawi
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2022.09.26 iNTS household survey.pptx
- 1. Case-control investigation of invasive Salmonella disease in Blantyre, Malawi reveals no evidence of environmental or animal reservoirs of invasive strains Dr Philip Ashton, Malawi-Liverpool-Wellcome Programme #ASTMH22 Add slideshare link
- 2. Acknowledgments Dr Leonard Koolman Dr Reenesh Prakash Prof Melita Gordon
- 3. Invasive Non-Typhoidal Salmonella (iNTS) introduction Achtman, Wain, Weill, Nair et al., 2012, PLOS Pathogens, e1009040 iNTS Burden • 535,000 (95% CI 409-705k) cases in 2017 • 77,500 (95% CI 46-123k) deaths iNTS Risk factors • HIV • Severe malaria • Malnutrition iNTS associated types • ST313 & ST3257 • Some lineages of S. Enteritidis • Becoming human restricted?
- 4. What is the source of iNTS-causing Salmonella infections? Kariuki et al., 2020 Kenya Kasumba et al., 2021 7 sites in Africa & Asia Phobia at al., 2020 DR Congo Wilson et al., 2020 Kenya and Malawi Crump et al., 2020 Tanzania & Kenya Dekker et al., 2018 Ghana Dione et al., 2011 The Gambia Carroll et al., 2021 South Africa Kariuki et al., 2006 Kenya Kariuki et al., 2002 Kenya Post et al., 2019 Burkina Faso Koolman et al., 2022 Malawi iNTS associated types are rarely found in the domestic environment, animals, or food sources. However, they have been found in either healthy human stool or the stool of people with diarrhoea.
- 5. Study objectives • Identify household shedding and potential household environmental sources of invasive Salmonellae within Blantyre • Compare invasive isolates to healthy human, animal and environmental strains using whole genome sequencing • Generate information to inform control strategies for Salmonella infections in Africa
- 6. Methods Index Case identification at QECH* (Typhi + NTS) 60 cases 60 controls URBAN Visit case household within 2 weeks Select Control household by bottle-spin and 100 yard walk Obtain Consents Conduct Sampling at Homes Family Animals Environment Culture and identification WGS & bioinformatic analysis QECH = Queen Elizabeth Central Hospital, a tertiary referral hospital in Blantyre, Malawi
- 7. Results – Salmonella positivity Total samples No. samples with Salmonella % positive samples No. HH tested No. HH with Salmonella % Positive HH Total samples No. samples with Salmonella % positive samples No. HH tested No. HH with Salmonella % Positive HH Human Stool 116 9 7.8 28 4 14.3 97 0 0.0 28 0 0.0 Animal stool/swab 27 0 0.0 20 0 0.0 38 0 0.0 17 0 0.0 HH environment 137 0 0.0 28 0 0.0 131 6 4.6 28 3 10.7 Total 280 9 3.2 28.0 4 14.3 266 6 2.3 28 3 10.7 Human Stool 157 4 2.5 32 2 6.3 121 3 2.5 32 3 9.4 Animal stool/swab 22 2 9.1 18 2 11.1 23 2 8.7 16 2 12.5 HH environment 168 5 3.0 32 4 12.5 166 12 7.2 32 9 28.1 Total 347 11 3.2 32 7 21.9 310 17 5.5 32 11 34.4 Typhoid Control households (HH) (n=32) iNTS Control Households (HH) (n=28) Typhoid Index Case households (HH) (n=32) iNTS Index Case Households (HH) (n=28) Salmonella positivity Overall sample positivity was 3.6%, with Salmonella isolated from every niche investigated
- 8. Results – Sequence Type presence in different niches The only niche with the same STs as index cases was household member stool
- 9. Results – Whole genome SNP phylogeny of index-case/household pairs In both cases, the isolate from the household family member was very closely related to that from the index case
- 10. Summary & conclusions • We were successful in isolating non-typhoidal Salmonella from the environment. • The only iNTS-associated types found in the environment were from the stool of household members – these were very closely related to the isolates from the index cases • Our results support a hypothesis that healthy humans were the source of iNTS infections.
- 11. Acknowledgements Leonard Koolman Reenesh Prakash Yohane Diness Chisomo Msefula Tonney S. Nyirenda Franziska Olgemoeller Nicholas A. Feasey Melita A. Gordon Core lab staff Jay C. D. Hinton Siân V. Owen Blanca Perez-Sepulveda Paul Wigley
Editor's Notes
- Hello, my name is philip ashton. I'm a post-doc in prof melita gordon's group at the malawi-liverpool-wellcome reserach programme, based here in blantyre, malawi, and i'm going to talk to you today about sources of invasive non-typhoidal salmonella. this presentation is also available from slideshare, in case you want to double check any of the references or anything later.
- first things first, i want to acknowledge the particular contribution of 3 people to this work. leonard koolman and reenesh prakash led the development of all the lab protocols and led the sampling teams, while melita gordon is the PI on this project. we recently posted a pre-print with more details of this work, so please see medrxiv for more details.
- ok, so, salmonella. there are two sepcies of salmonella, s. enterica and s. bongori. within s. enterica there are 6 sub-species, of which 1, s. enterica, subsp enterica causes 99% of human and animal infections. within s. enterica, there are more than 1500 serovars, and these are divided based on the disease they cause into typhoidal salmonella and non-typhoidal salmonella. typhoidal salmonella such as SAlmonella tyhphi and paratyphi cause serious invasive disease, while non-typhoidal salmonella are typcailly associated with self limiting gastroenteritis. however, starting at around the turn of the millenium, there has been an increasing acknowledgement of invasive non-typhoidal salmonella. this is syndromically very different from typhical non-typhoidal salmonella. there is a significant burden of infeciton, particularly in sub-saharan Africa, and a substantial mortality rate. the risk factors are HIV in adults and children, and severe malaria and malnutrition in children. most iNTS is caused by partciular sub-types of non-typhoidal salmonella. the two main causes are Salmonella typhimuiurm ST313 (and it's single locus variant ST3257) and some lineages of S. Enteritidis. there is some evidence from genomics, and comparisons with the typhoidal serovars, the iNTS associated types are becoming human restricted.
- One persistent mystery of iNTS epidemiology is the source of INTS infections. Normally non-typhoidal salmonella are zoonotic infections from animal reservoirs, however from the earliest studies on iNTS, there has not been much evidence of animal reservoirs being important for the sub-types of salmonella that cause iNTS. Here we have a figure summarising the different potential sources of iNTS, such as food, water, animals, the domestic environment, and family members. There are 12 studies that have tried to address the question of sources of iNTS infections, and they used different sampling frames. However, the overall message from all these studies is that iNTS associated types are rarely found in the domestic environment, animals, or food sources. While they have been found in either healthy human stool or the stool of people with diarrhoea.
- so, to outline our study objectives, we want to...
- Briefly, our methods were:
- So, for this slide, we split the households into four groups based on whether the index case was iNTS or typhoid, and whether the household was a case or control household. starting with the iNTS case households, the three rows are results from sampling the stool of household members, animal stools or rectal swabs, and the household envrionment (e.g. bootsocks). then, the columns are the total number of samples taken, the number and percentage of samples that were positive for salmonella. the number of households tested, and the number and percentage of households with salmonella. so, you can see here that from the iNTS case households, 3.2% of samples were positive, all human stools, and 14% of housholds were positive for salmonella. then, in the iNTS control households, 2.3% of samples were positive, just from the household environment, and 11% of households were positive. then, 3.2% of the samples from typhoid case households were positive, with salmonella isolated from each category of niches, and finally from the typhoid control households, 5.5% of samples were positive, and 34% of households.
- this slide shows the sequence types identified in different niches. by sequence type, i mean that the sequences of 7 housekeeping genes were analysed, and if the sequences were identical, then the isolates have the same sequence type.this figure is a minimum spanning tree. sequence types that vary by only one of the 7 housekeeping genes are connected by a line. each circle is a sequence type, with the size approximately proportional to the number of isolates of that type. they are coloured by the source of the isolates. furthermore, whenever isolates of the same sequence type are from the same household, they have a shared border colour. for example, here, in green, there are a pair of ST3263 isolates from the same household (green border), one from animal, and one environmental. also st3263, there were a pair of isolates from the same household, also one animal one environmental. but, the ones that are really interesting to our research question are these two - st313 and st3257. and you can see that one of each ST has a pair of isolates from the same household. one from an index case (human invasive disease) and one from a human, not invasive disease (household member).
- because we have whole genome sequences for these isolates, we can look across the whole genome, not just at 7 housekeeping genes. this is a maximum likelihood phylogeny that shows that there were only 2 SNPs separateing the index case and family member pair in one case, and 3 SNPs separating the index and family member in the other case.