Genetic Admixture and Host-Pathogen Interactions in Helicobacter pylori Infection

Lead Research Organisation: University of Leicester
Department Name: Genetics

Abstract

Helicobacter pylori infection is one of the most common bacterial infections in the world: infection rates range from 25-50% in developed countries to 70-90% in the third world. It is well recognised that H. pylori infection causes upper gastrointestinal diseases; however, understanding how the bacteria causes disease has been challenging, because the disease risk likely depends on complex interactions between both human host human and bacterial genes, as well as dietary and other environmental factors. Until recently, most studies on host-H. pylori interactions were performed in developed countries and have focused on host or bacterial genetic factors in isolation. These studies have failed to explain the high variability in the likelihood to suffer from gastrointestinal disease seen in different individuals and populations following H. pylori infection.

H. pylori has been evolving in concert with humans since their early origins in Africa and migrations out-of Africa. As a consequence, the genomes of the bacteria are genetically differentiated in a similar way as the host genomes among human populations. This has limited past studies to specific host-pathogen combinations. The ideal experiment to understand interactions between human hosts and bacteria would be to expose individuals to random strains of H. pylori and observe disease outcomes.

Recently admixed populations, i.e. populations that derive their genetic ancestry from multiple continents, offer unique opportunities to perform such type of studies, since the mixing process most likely brought together assorted human and H. pylori genomes which are not present in the parental non-admixed populations. The main aim of this project is to characterise the complex host-pathogen-environment interactions that underlie susceptibility to H. pylori infection in the admixed population of Cape Verde. This previously uninhabited archipelago, located 450 km off the coast of Senegal, was discovered by the Portuguese in the mid-15th century and was initially populated with European immigrants and slaves from the neighbouring African mainland. As a consequence of this settlement pattern, extensive genetic admixture occurred between the African and European ancestral populations during the last few centuries. Due to the high admixture in Cape Verde and cross-infection between unrelated individuals, we expect to find not only some correlation in the European/African individual ancestry proportions between the human hosts and the bacteria, as well as individuals who have high African ancestry but are infected by H. pylori that is largely European in origin and vice versa. Specific host-bacteria combinations that are over-represented among the cases offer clues to identify host and pathogen genetic regions that underlie susceptibility to H. pylori infection.

To accomplish our main goal, we will: a) screen H. pylori infection in Cape Verde volunteers, collect DNA samples from 700 individuals and their associated bacteria, and obtain measurements of the degree of the individuals' gastric mucosa damage caused by infection. Our rationale for recruiting volunteers is that a large proportion of the affected individuals may be asymptomatic or un-diagnosed; recruiting volunteers from the general population allows unbiased assessment of both the infection rate and pattern of genetic diversities. b) To examine the association between the genetic diversity and individual ancestry of human host and H. pylori and disease outcomes.

This project establishes a unique and valuable resource for elucidating host-pathogen interactions in gastrointestinal diseases. The study also has significant public health impact, representing a first effort in assessing the infection rate and the correlation between combinations of human and H. pylori genomes and disease outcomes in Cape Verde.

Technical Summary

The main goal of this research is to assess whether individual genomic ancestry and genomic variation of both humans and associated H. pylori affect susceptibility to infection in the admixed population of Cape Verde. To this end, we propose to screen volunteers, of both sexes and aged >35 years, and to collect blood and gastric mucus samples from 700 infected individuals in the two main hospitals of Cape Verde. The blood samples will be used for human DNA extraction, determination of H. pylori infection status through detection of Immunoglobulin G against H. pylori, and for indirect evaluation of the severity of gastric mucosa damage through measurement of the plasma biomarkers pepsinogen I and pepsinogen II. The gastric mucus samples will be used for H. pylori culture, and will be collected on an adhesive nylon string that the participants swallow and that is retrieved an hour later (gastric string). We will then perform high-throughput genotyping of human genomes and next-generation sequencing of complete bacterial genomes. This will allow us to characterise the genomic variation, the admixture structure and the correlation between the individual genomic ancestries (IA) of hosts and H. pylori in Cape Verde. We will also be able to examine the correlation between IA's and the degree of gastric mucosa damage. A significant correlation will indicate the presence of human and bacterial genetic factors that are responsible, at least in part, for the phenotypic variation in disease risk to H. pylori infection observed within and between the ancestral populations to Cape Verde. The relative contribution of these genetic factors can then be quantitated. In addition, we will be able to perform association scans between human and bacterial genotypes or between human and bacterial local ancestry (admixture mapping) and human phenotypes to gain a preliminary characterisation of the genetic architecture underlying susceptibility to H. pylori infection in Cape Verde.

Planned Impact

The anticipated main scientific outcome of the project is a better understanding of the co-evolution between human hosts and H. pylori and of the large heterogeneity in disease outcomes underlying infection observed both within and among populations. We will identify human-H. pylori genomic combinations that are at greater risk for developing disease, and will potentially pinpoint genetic variation that confers susceptibility in humans and pathogenicity in bacteria. Such results have the potential to benefit a wide community, as well as scientists in the field.

In the mid-to-long term, this knowledge will help clinicians to identify those infected individuals that are at greater risk of developing gastric disease and to whom treatment should be targeted (i.e. personalized medicine). Identification of cellular pathways in the human host that might be disrupted in disease can lead to the identification of novel molecular targets for therapeutic drugs, potentially benefiting pharmaceutical industry researchers. This is in support of the MRC's strategic aim to translate research, since it has the potential to bring the health impacts of fundamental research to society more quickly.

There are also beneficiaries within the wider public, apart from those who will benefit from the development of better diagnostic strategies. Specifically, this project sits within MRC's strategic aim to support global research that addresses inequalities in health, which arise particularly in developing countries. Firstly, due to the design of this research programme, there will be a direct and immediate benefit to the Cape Verdean individuals that participate in the study. Infected participants showing signs of gastric disease will be immediately referred for medical consultation, examination and treatment. In the short-run, the population will also benefit from the first screen of H. pylori infection rate and its consequences in Cape Verde. This knowledge will have a large impact in terms of public health policy. Professionals in the epidemiology field will also benefit from knowing how the infection rate and associated morbidity in this particular developing country compares to the global epidemics, and how this is related with the genetic background of the individuals.

Through our communication programmes, A-level and university students in the UK, and high school and university students in Cape Verde, will have contact with the project and with subjects in the main scientific areas of the project. Such experience can direct career decisions and inspire creativity in the student's own areas of research. They will also benefit from a better understanding of how different disciplines such as evolutionary and computational biology can work in concert with medical and bacterial genetics to provide insights into a subject of clinical relevance.

This project also presents the perfect opportunity for masters and PhD students (both in UK and in Cape Verde), and PDRAs that work on the project to get excellent training in the latest computational and statistical approaches in genomics and in methodology related with microbiology and bacterial genetics. These are valuable skills that will help them in their future careers.

Publications

10 25 50

publication icon
Archampong TN (2019) GASTRO-DUODENAL DISEASE IN AFRICA in World Journal of Gatroenterology

 
Description We have collected data from the general population, not solely from individuals that resort to gastroenterology clinics. Epidemiological data from the general population shows an unbiased infection rate of 84%. Genomic data obtained in 534 isolates of Helicobacter pylori from 178 hosts show bacterial within-the-host and population structure. There are African and European strains in the islands, which are clustered into four main population subgroups. In contrast to what was observed in Colombia, where only symptomatic individuals were studied, there was no correlation between the genetic ancestry of the strains and of the host in Cape Verde. Notably, there is a highly frequent and differentiated bacterial cluster in the mainly asymptomatic human population with the particular phenotypic features of absence of genes associated with gastric cancer. This cluster shows signatures of selective advantage in the population of Cape Verde. This interesting finding never reported before is due to the uniqueness of the data collection, and has implications for the assessment of human-H. pylori interactions. Our observations underscore the need to understand the molecular interactions between the bacteria and their host, in order to identify infected individuals that have a greater risk to develop disease and to whom treatment should be targeted. Targeted treatment will also contribute to the prevention of widespread antimicrobial drug resistance. This study demonstrates Cape Verde to be a natural model to study susceptibility to H. pylori infection.
Exploitation Route In the academic setting, both the genomics dataset and the computational approaches we employed constitute a valuable resource that we will share with other population, bacterial and evolutionary geneticists. In addition, within our aim of building research-capacity, four Cape Verdean undergraduate and two UK-based PhD students worked on this project, receiving training in microbiology, molecular genetics, statistical analyses, and bacterial and human genomic and population genetics approaches. Finally, as Leicester is a research-led university, the main concepts and findings of this project and related studies were taught in a third-year Medical Genetics module by the PI.

Regarding implications for Healthcare, we provided the first estimate of an unbiased H. pylori infection rate (84%) to Cape Verde. This was reported to the Ministry of Health, who will now have to set frequent diagnosis of this infection in the clinics. We intend to continue working with them in order to assess associated gastric disease incidence and the genomic variation of Human-H pylori pairs associated with disease risk. We are also evaluating the levels of Antimicrobial drug resistance (AMR) of the strains we collected - we observe high levels of resistance and cases of multi-drug resistance. This has led to the establishment of a science/clinical collaboration to define the impact of the absence of effective infectious disease monitoring and empirical antibiotic prescribing practice on disease burden and in the development of AMR in Cape Verde.
Sectors Education,Healthcare

 
Description Firstly, due to the design of this research programme, there was a direct and immediate benefit to the Cape Verdean individuals that participated in the study - Infected participants showing signs of gastric disease were referred for medical consultation, examination and treatment. Secondly, we provided the first estimate of an unbiased H. pylori infection rate (84%). This was reported to the Ministry of Health, who will now have to set frequent diagnosis of this infection in the clinics. We intend to continue working with them in order to assess associated gastric disease incidence and evaluate the genomic variation of Human-H. pylori pairs associated with disease risk. We have also reported the about the main theme and results of this project to the Cape Verde community both by a conference in the capital of the country and in a radio programme. We are raising awareness regarding this infection in Cape Verde. In this sense, this project sits within ODA strategic aim to support global research that addresses inequalities in health, which arise particularly in developing countries. We are also evaluating the levels of Antimicrobial drug resistance (AMR) of the strains we collected - we observe high levels of resistance and cases of multi-drug resistance. This has led to the expansion of our basic research collaboration into the establishment of a science/clinical collaboration between the University of Leicester and associated hospital, the University of Cape Verde and the Cape Verde Health Ministry, to define the impact of the absence of effective infectious disease monitoring and empirical antibiotic prescribing practice on disease burden and in the development of AMR Cape. The research programme consists in a) To evaluate the natural history and molecular evolution of AMR in CV and how are these related with a restricted panel of antibiotics used locally and to globe-wide patterns of resistance; b) to perform a clinical and health economic study of the impact of the development of resources for accurate AMR testing (clinical microbiology laboratory) on antibiotic prescribing practice and antibiotic resistance in the country. Within this programme, the established network surveyed the clinical microbiology labs of the two main hospitals in Cape Verde and identified high levels of resistance developing across the years for Staphylococcus aureus. We also found gaps in the collection, analysis and reporting of the clinical microbiology labs and we are going to report this to the Health authorities in the country soon.
First Year Of Impact 2018
Sector Education,Healthcare
Impact Types Societal,Policy & public services

 
Description GCRF networking grants (GCRFNG\100338)
Amount £24,600 (GBP)
Organisation Research Councils UK (RCUK) 
Sector Public
Country United Kingdom
Start 04/2018 
End 03/2019
 
Title Protocols for Helicobacter pylori isolation from healthy and diseased individuals and culture under restricted conditions in Cape Verde 
Description We have develop protocols to call for volunteers from the general population, to collect samples of gastric mucus for Helicobacter pylori isolation from healthy and diseased individuals in a minimally invasive way, and to culture Helicobacter pylori under restricted conditions settings 
Type Of Material Biological samples 
Year Produced 2017 
Provided To Others? No  
Impact We developed protocols that can be applied to the culture of other microbial organisms and developed technician and research capacity for Microbiology testing in Cape Verde. 
 
Title Biobank of 743 Helicobacter pylori isolates belonging to 183 Cape Verde individuals 
Description We collected more than one , up to 26 clones of Helicobacter pylori from gastric mucus samples of 183 individuals. We generated whole genome sequences for the 743 clones and will obtain high-throughput genotypes of the respective hosts 
Type Of Material Database/Collection of data 
Year Produced 2018 
Provided To Others? No  
Impact We have develop a protocol to isolate efficiently Helocibacter pylori strains in restricted laboratory conditions in Africa and the transport of samples from African to the UK. 
 
Title Cape Verde dataset on Helicobacter pylori infection and clinical metadata 
Description This database contains clinical metadata (age sex, place of birth, socio-economic variables, smoking habits, history of Helicobacter pylori infection, frequency of antibiotic intake, family history regarding cancer, symptoms of dyspepsia, infection with parasites), ELISA data for infection with Helicobacter pylori, ELISA data on biomarkers of the severity of gastric damage in the blood, information about h pylori isolation with gastric string test, for 938 participants from the general population of Cape Verde. 
Type Of Material Database/Collection of data 
Year Produced 2016 
Provided To Others? No  
Impact This database allow for an evaluation of the prevalence of Helicobacter infection and of its consequences in the Population of Cape Verde. We determined a prevalence of 84%. This will have important public health impact, since currently the Hospitals do not evaluate Helicobacter pylori infection, and gastric cancer is the main cause of death by cancer in the islands. 
 
Description ANCoVer Network - Establishing an multidisciplinary network for antimicrobial drug resistance surveillance in Cabo Verde 
Organisation University Hospitals of Leicester NHS Trust
Country United Kingdom 
Sector Academic/University 
PI Contribution Under this new MRC-funded project we have built a new microbiology lab, utilizing new resources funded by the project and resources available at the university of Cape Verde. We then have trained four students in Microbiology techniques, and specifically in Helicobacter pylori culturing, and in ELISA and basic molecular biology methods. We have determined the rate of Infection by determining the plasma levels of IgG against H. pylori in approximately 900 individuals and Isolated H. pylori strains from approximately 200 gastric mucus samples (~20% of the samples/individuals had more than one strain). We have also determined the resistance status of the bacterial strains to four main antibiotics used in the eradication of infection: Metrodinazole; Amoxicilin; Clarithromycin and Tetracycline. Results from our work determined that the prevalence is 84%, and suggest that the level of antimicrobial resistance (AMR) is significant (from 7.8 % resistance to Clarithromycin to 46% resistance to Metrodinazone, but only one 0.1% resistance to Tetracycline; examples of multi-drug resistance) . This prompted us to establish a multidisciplinary network on AMR in Cape Verde. Our primary objective is to establish a basic science/clinical collaboration to define the impact of the absence of effective infectious disease monitoring and empirical antibiotic prescribing practice on disease burden and in the development of antimicrobial drug resistance (AMR) in the island nation of Cabo Verde.
Collaborator Contribution Reciprocal visits (a two-week visit to Cape Verde and a one-week visit to Leicester) under our long-term collaboration by Leicester and Cape Verde scientists and the director of health in Cape Verde have made this application possible. In these meetings we have discussed the health priorities in Cape Verde, where the need to evaluate the health and economic impact of the absence of a clinical microbiology lab in the diagnosis of infections and definition of antibiotic prescription practices was established and where a feasibility study of a clinical trial was developed. Within this project, we have performed a five-year retrospective study concerning antimicrobial drug resistance (AMR) in Cape Verde main hospitals, and have found an alarming increase of Methicilin-resistant Staphylococcus aureus (MRSA) over the past years, with 80% of hospital isolates being methicillin resistant in 2019. This poses a very high threat to empirical antibiotic therapy in a setting with low availability of diagnostic facilities. Further analyses of the genomes of over 100 S. aureus isolates led to the finding of a lineage with a unique pattern of antibiotic resistance. These two findings were submitted for peer-review. We now intend to expand this work and develop a research programme with two research main goals: A. To evaluate the natural history and molecular evolution of AMR in CV and how are these related with a restricted panel of antibiotics used locally and to global-wide patterns of resistance. Starting with the data collected in S. aureus (we have more samples for genomic sequencing), we will perform an improved characterisation of AMR patterns in S. aureus, including MRSA, and of the colonisation and transmission of resistant bacteria in Cape Verde. B. To perform a clinical and health economic study (an interrupted time series design) of the impact of a health intervention aimed at decreasing human carriage by MRSA in Cabo Verde. We have obtained funding from the University of Leicester for a series of meetings with the local hospitals and the ministry of health in Cape Verde to discuss the feasibility of applying for funding to perform an evaluation of such an intervention in Cabo Verde in collaboration with Cabo Verdean colleagues.
Impact The feasibility study of the clinical trial will start in April 2018. Submitted manuscripts: Monteiro T, M Wysocka, E Tellez, O Monteiro, L Spencer, E Veiga, S Monteiro, C de Pina, D Gonçalves, S de Pina, L Correia, J Moreno, T Conceicao, M Aires De Sousa, H De Lencastre, L Gray, M Pareek, DR Jenkins, the ANCoVer project, S Beleza, MR Oggioni, II Araujo. A five-year retrospective study shows increasing rates of antimicrobial drug resistance in Cabo Verde for both Staphylococcus aureus and Escherichia coli. Submitted. Wysocka M, T Conceicao, Monteiro T, C de Pina, D Gonçalves, S de Pina, L Correia, J Moreno, L Gray, M Pareek, DR Jenkins, M Aires De Sousa, H De Lencastre, S Beleza, II Araujo, MR Oggioni. Genome sequencing shows frequent loss of beta-lactamase in Staphylococcus aureus leading to oxacillin-susceptible MRSA in Cabo Verde. In preparation.
Start Year 2018
 
Description ANCoVer Network - Establishing an multidisciplinary network for antimicrobial drug resistance surveillance in Cabo Verde 
Organisation University of Cape Verde
Country Cape Verde 
Sector Academic/University 
PI Contribution Under this new MRC-funded project we have built a new microbiology lab, utilizing new resources funded by the project and resources available at the university of Cape Verde. We then have trained four students in Microbiology techniques, and specifically in Helicobacter pylori culturing, and in ELISA and basic molecular biology methods. We have determined the rate of Infection by determining the plasma levels of IgG against H. pylori in approximately 900 individuals and Isolated H. pylori strains from approximately 200 gastric mucus samples (~20% of the samples/individuals had more than one strain). We have also determined the resistance status of the bacterial strains to four main antibiotics used in the eradication of infection: Metrodinazole; Amoxicilin; Clarithromycin and Tetracycline. Results from our work determined that the prevalence is 84%, and suggest that the level of antimicrobial resistance (AMR) is significant (from 7.8 % resistance to Clarithromycin to 46% resistance to Metrodinazone, but only one 0.1% resistance to Tetracycline; examples of multi-drug resistance) . This prompted us to establish a multidisciplinary network on AMR in Cape Verde. Our primary objective is to establish a basic science/clinical collaboration to define the impact of the absence of effective infectious disease monitoring and empirical antibiotic prescribing practice on disease burden and in the development of antimicrobial drug resistance (AMR) in the island nation of Cabo Verde.
Collaborator Contribution Reciprocal visits (a two-week visit to Cape Verde and a one-week visit to Leicester) under our long-term collaboration by Leicester and Cape Verde scientists and the director of health in Cape Verde have made this application possible. In these meetings we have discussed the health priorities in Cape Verde, where the need to evaluate the health and economic impact of the absence of a clinical microbiology lab in the diagnosis of infections and definition of antibiotic prescription practices was established and where a feasibility study of a clinical trial was developed. Within this project, we have performed a five-year retrospective study concerning antimicrobial drug resistance (AMR) in Cape Verde main hospitals, and have found an alarming increase of Methicilin-resistant Staphylococcus aureus (MRSA) over the past years, with 80% of hospital isolates being methicillin resistant in 2019. This poses a very high threat to empirical antibiotic therapy in a setting with low availability of diagnostic facilities. Further analyses of the genomes of over 100 S. aureus isolates led to the finding of a lineage with a unique pattern of antibiotic resistance. These two findings were submitted for peer-review. We now intend to expand this work and develop a research programme with two research main goals: A. To evaluate the natural history and molecular evolution of AMR in CV and how are these related with a restricted panel of antibiotics used locally and to global-wide patterns of resistance. Starting with the data collected in S. aureus (we have more samples for genomic sequencing), we will perform an improved characterisation of AMR patterns in S. aureus, including MRSA, and of the colonisation and transmission of resistant bacteria in Cape Verde. B. To perform a clinical and health economic study (an interrupted time series design) of the impact of a health intervention aimed at decreasing human carriage by MRSA in Cabo Verde. We have obtained funding from the University of Leicester for a series of meetings with the local hospitals and the ministry of health in Cape Verde to discuss the feasibility of applying for funding to perform an evaluation of such an intervention in Cabo Verde in collaboration with Cabo Verdean colleagues.
Impact The feasibility study of the clinical trial will start in April 2018. Submitted manuscripts: Monteiro T, M Wysocka, E Tellez, O Monteiro, L Spencer, E Veiga, S Monteiro, C de Pina, D Gonçalves, S de Pina, L Correia, J Moreno, T Conceicao, M Aires De Sousa, H De Lencastre, L Gray, M Pareek, DR Jenkins, the ANCoVer project, S Beleza, MR Oggioni, II Araujo. A five-year retrospective study shows increasing rates of antimicrobial drug resistance in Cabo Verde for both Staphylococcus aureus and Escherichia coli. Submitted. Wysocka M, T Conceicao, Monteiro T, C de Pina, D Gonçalves, S de Pina, L Correia, J Moreno, L Gray, M Pareek, DR Jenkins, M Aires De Sousa, H De Lencastre, S Beleza, II Araujo, MR Oggioni. Genome sequencing shows frequent loss of beta-lactamase in Staphylococcus aureus leading to oxacillin-susceptible MRSA in Cabo Verde. In preparation.
Start Year 2018
 
Description Collaboration with University of Accra, Ghana 
Organisation University of Accra
Country Ghana 
Sector Academic/University 
PI Contribution I am supervising Mr Tim Archampong in his MD thesis. Dr Achampong is studying the genetic diversity in Helicobacter pylori in asymptomatic individuals versus patients, analysing strains from Cape Verde, Ghana and worldwide samples included in public databases.
Collaborator Contribution Tim Archampong is collecting samples of Helicobacter pylori strains (for his project) that will be used as reference samples in the MRC-funded Project
Impact 10.1186/s13104-017-2542-8
Start Year 2016
 
Description Contribution for science TV programme in Cape Verde 
Form Of Engagement Activity A broadcast e.g. TV/radio/film/podcast (other than news/press)
Part Of Official Scheme? No
Geographic Reach National
Primary Audience Public/other audiences
Results and Impact Science TV programe on the Genetic structure of Cape Verde and its use for the study of susceptibility to Helicobacter pylori infection (http://rtc.cv/tcv/index.php?paginas=47&id_cod=47020 ; program in portuguese). This program was intended to both report the main results obtained during the longstanding collaboration with the university of Cape Verde about the genetic structure of Cape Verde , the use of the population as a model to study complex disease that differ between European and African populations, and what were the main findings that led us to develop a new project on Host-pathogen interactions in Helicobacter pylori. We took the opportunity to explain the concepts and goals of the new project and publicize the sampling step to the general Cape Verde public.
Year(s) Of Engagement Activity 2017
URL http://rtc.cv/tcv/index.php?paginas=47&id_cod=47020
 
Description Epidemiology of gastric disease in Cape Verde 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach Local
Primary Audience Study participants or study members
Results and Impact An afternoon with two talks and a Q&A session. The first talk was general about Helicobater pylori infection, worlwide epidemiology and association with gastric disease. The second concerned the presentation of the main results of the project: Epidemiological study in Cape Verde, genetic variation of H. pylori in the asymptomatic host population and Antimicrobial drug resistance levels to 4 antibiotics used in clinics. We finished with a session of questions and answers done the attendees to a panel composed by the main researchers of the project (the PI, Dr Sandra Beleza, of the University of Leicester, and Dr Isabel Araújo of the University of Cape Verde) and the gastroenterologist consultant in the study.
Year(s) Of Engagement Activity 2018