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Overview Phase 1 – Local AMR & Virulence Mapping of Major Tilapia Pathogens in Burkina Faso to Support Autovaccine Development Scientific Rationale & Importance Antibiotic overuse in tilapia farming drives antimicrobial resistance (AMR), threatening productivity, public health, and food security. Streptococcus agalactiae and Aeromonas hydrophila are major pathogens causing mass mortalities and economic losses. These bacteria often display multidrug resistance (MDR) and biofilm-forming ability, reducing treatment efficacy. This project will generate the first local AMR and virulence dataset from Burkina Faso to support the development of preventive alternatives, including autogenous vaccines and post-biotic adjuvants, aligned with One Health principles. Funding / Facilities / Ethics Budget requested: 5 000 USD Facilities: Veterinary Microbiology Lab, Kırıkkale University, Turkey (PhD host); collaborative access to local lab in Burkina Faso for sampling. Ethics: Non-invasive sampling; farm-level consent; no animal experimentation; BSL-2 compliance. Timeline 12 months (M1–M12): sampling, isolation/identification, antibiograms, virulence testing, data analysis, dissemination. Data Sharing All AMR and virulence data will be deposited in an open-access repository (Zenodo/OSF) within 3 months of completion. Strains will be available for non-commercial research under an MTA. 2. Introduction Theoretical background Antimicrobial resistance (AMR) is a global One-Health emergency. Intensive tilapia culture relies heavily on antibiotics, selecting for multidrug-resistant (MDR) pathogens that spread to humans via food and water. Two taxa drive mortality in African tilapia farms: Streptococcus agalactiae (streptococcosis; ≤80 % mortalities) and Aeromonas hydrophila (ulcerative syndrome, septicaemia). Both form biofilms and carry MDR plasmids, yet no baseline AMR or virulence data exist for Burkina Faso. Key literature Asian and East-African surveys report tetracycline and fluoroquinolone resistance rates >60 % in these species (Chen 2020; Hounmanou 2021). Biofilm-positive isolates show 4–8× higher minimal inhibitory concentrations and persist in pond sediments (Watts 2017). Autogenous vaccines, adjuvanted with low-cost post-biotics, are emerging as sustainable alternatives, but strain selection requires local phenotypic and resistance maps—currently missing for West Africa. Research questions RQ1 Which S. agalactiae and A. hydrophila genotypes dominate Burkinabè tilapia farms? RQ2 What are their AMR profiles against 12 antibiotic classes? RQ3 Do they exhibit high-virulence phenotypes (haemolysis, biofilm, extracellular enzymes) that warrant vaccine inclusion? Numbered hypotheses H1 Local isolates display MDR (≥3 classes), especially to tetracyclines and fluoroquinolones. H2 ≥50 % of isolates are strong biofilm-formers and β-haemolytic. H3 A quantifiable subset combines high prevalence, MDR and virulence, providing optimal autovaccine candidates. Impact statement Regardless of resistance levels, this first national surveillance will: (i) establish an open-access AMR/virulence atlas and strain bank; (ii) guide rational antimicrobial use; (iii) select candidate strains for an immediate PhD-driven autogenous vaccine trial with post-biotic adjuvant; and (iv) create a replicable One-Health surveillance model for West African aquaculture. 3. Methods Participants / Sample Sites: 5-7 tilapia farms in Burkina Faso Samples: gills, kidney, liver of diseased/suspect fish; pond water; equipment swabs Exclusion: fish treated with antibiotics <7 days Target: 60–90 isolates (30–45 per species) Materials & Procedures Isolation & ID: Todd-Hewitt, TSA, MacConkey, blood agar; Gram, catalase, oxidase; biochemical galleries (API or low-cost equivalent); optional species-specific PCR if budget allows. Antibiograms: Kirby-Bauer, EUCAST/CLSI; panel: β-lactams, tetracyclines, fluoroquinolones, sulfonamides/trimethoprim, aminoglycosides, phenicols. Virulence: haemolysis on blood agar, crystal-violet biofilm (96-well), gelatinase/DNase plates. Planned Analyses Resistance rates & MDR index Site comparisons (Chi²/Fisher) Biofilm: ANOVA/Tukey Strain prioritisation: matrix combining prevalence, MDR, and virulence score Contingency: if isolation rate < expected → increase sites/samples; if no PCR → focus on robust phenotyping Ethics & Data Management Farm consent; anonymised data BSL-2 biosafety; national waste regulations Open data: CSV + metadata → Zenodo/OSF (CC-BY 4.0) 4. Pilot Data (Optional) Summarize any pilot data or simulations that demonstrate method feasibility and inform power analysis. Clearly distinguish from main study. 5. Budget Category Cost (USD) Media & reagents (BHI/TSB, blood agar, Gram, antibiotic discs) 1,200 Consumables (plates, swabs, tips, tubes, cryotubes) 900 Transport/field (coolers, ice packs, travel) 600 Targeted confirmations (PCR/16S outsourced – optional) 700 Small equipment (pipettes/calibration, racks, pH meter) 500 Quality & control (reference strains, control discs) 250 Communication & reporting (printing, farm briefings) 200 Contingency 350 Estimated total 4,700 6. References FAO. The State of World Fisheries and Aquaculture. 2022. Cabello FC. Environ Microbiol, 2006. Watts JEM et al. Front Microbiol, 2017. Amal MNA, Zamri-Saad M. Aquaculture, 2011. Austin B, Austin DA. Bacterial Fish Pathogens. Springer, 2016. Chen L et al. Sci Total Environ, 2020. Phu TM et al. Rev Aquac, 2016. Hounmanou YAG et al. Aquaculture Reports, 2021.57:T168f,Proposal Template 1. Overview Phase 1 – Local AMR & Virulence Mapping of Major Tilapia Pathogens in Burkina Faso to Support Autovaccine Development Scientific Rationale & Importance Antibiotic overuse in tilapia farming drives antimicrobial resistance (AMR), threatening productivity, public health, and food security. Streptococcus agalactiae and Aeromonas hydrophila are major pathogens causing mass mortalities and economic losses. These bacteria often display multidrug resistance (MDR) and biofilm-forming ability, reducing treatment efficacy. This project will generate the first local AMR and virulence dataset from Burkina Faso to support the development of preventive alternatives, including autogenous vaccines and post-biotic adjuvants, aligned with One Health principles. Funding / Facilities / Ethics Budget requested: 5 000 USD Facilities: Veterinary Microbiology Lab, Kırıkkale University, Turkey (PhD host); collaborative access to local lab in Burkina Faso for sampling. Ethics: Non-invasive sampling; farm-level consent; no animal experimentation; BSL-2 compliance. Timeline 12 months (M1–M12): sampling, isolation/identification, antibiograms, virulence testing, data analysis, dissemination. Data Sharing All AMR and virulence data will be deposited in an open-access repository (Zenodo/OSF) within 3 months of completion. Strains will be available for non-commercial research under an MTA. 2. Introduction Theoretical background Antimicrobial resistance (AMR) is a global One-Health emergency. Intensive tilapia culture relies heavily on antibiotics, selecting for multidrug-resistant (MDR) pathogens that spread to humans via food and water. Two taxa drive mortality in African tilapia farms: Streptococcus agalactiae (streptococcosis; ≤80 % mortalities) and Aeromonas hydrophila (ulcerative syndrome, septicaemia). Both form biofilms and carry MDR plasmids, yet no baseline AMR or virulence data exist for Burkina Faso. Key literature Asian and East-African surveys report tetracycline and fluoroquinolone resistance rates >60 % in these species (Chen 2020; Hounmanou 2021). Biofilm-positive isolates show 4–8× higher minimal inhibitory concentrations and persist in pond sediments (Watts 2017). Autogenous vaccines, adjuvanted with low-cost post-biotics, are emerging as sustainable alternatives, but strain selection requires local phenotypic and resistance maps—currently missing for West Africa. Research questions RQ1 Which S. agalactiae and A. hydrophila genotypes dominate Burkinabè tilapia farms? RQ2 What are their AMR profiles against 12 antibiotic classes? RQ3 Do they exhibit high-virulence phenotypes (haemolysis, biofilm, extracellular enzymes) that warrant vaccine inclusion? Numbered hypotheses H1 Local isolates display MDR (≥3 classes), especially to tetracyclines and fluoroquinolones. H2 ≥50 % of isolates are strong biofilm-formers and β-haemolytic. H3 A quantifiable subset combines high prevalence, MDR and virulence, providing optimal autovaccine candidates. Impact statement Regardless of resistance levels, this first national surveillance will: (i) establish an open-access AMR/virulence atlas and strain bank; (ii) guide rational antimicrobial use; (iii) select candidate strains for an immediate PhD-driven autogenous vaccine trial with post-biotic adjuvant; and (iv) create a replicable One-Health surveillance model for West African aquaculture. 3. Methods Participants / Sample Sites: 5-7 tilapia farms in Burkina Faso Samples: gills, kidney, liver of diseased/suspect fish; pond water; equipment swabs Exclusion: fish treated with antibiotics <7 days Target: 60–90 isolates (30–45 per species) Materials & Procedures Isolation & ID: Todd-Hewitt, TSA, MacConkey, blood agar; Gram, catalase, oxidase; biochemical galleries (API or low-cost equivalent); optional species-specific PCR if budget allows. Antibiograms: Kirby-Bauer, EUCAST/CLSI; panel: β-lactams, tetracyclines, fluoroquinolones, sulfonamides/trimethoprim, aminoglycosides, phenicols. Virulence: haemolysis on blood agar, crystal-violet biofilm (96-well), gelatinase/DNase plates. Planned Analyses Resistance rates & MDR index Site comparisons (Chi²/Fisher) Biofilm: ANOVA/Tukey Strain prioritisation: matrix combining prevalence, MDR, and virulence score Contingency: if isolation rate < expected → increase sites/samples; if no PCR → focus on robust phenotyping Ethics & Data Management Farm consent; anonymised data BSL-2 biosafety; national waste regulations Open data: CSV + metadata → Zenodo/OSF (CC-BY 4.0) 4. Pilot Data (Optional) Summarize any pilot data or simulations that demonstrate method feasibility and inform power analysis. Clearly distinguish from main study. 5. Budget Category Cost (USD) Media & reagents (BHI/TSB, blood agar, Gram, antibiotic discs) 1,200 Consumables (plates, swabs, tips, tubes, cryotubes) 900 Transport/field (coolers, ice packs, travel) 600 Targeted confirmations (PCR/16S outsourced – optional) 700 Small equipment (pipettes/calibration, racks, pH meter) 500 Quality & control (reference strains, control discs) 250 Communication & reporting (printing, farm briefings) 200 Contingency 350 Estimated total 4,700 6. References FAO. The State of World Fisheries and Aquaculture. 2022. Cabello FC. Environ Microbiol, 2006. Watts JEM et al. Front Microbiol, 2017. Amal MNA, Zamri-Saad M. Aquaculture, 2011. Austin B, Austin DA. Bacterial Fish Pathogens. Springer, 2016. Chen L et al. Sci Total Environ, 2020. Phu TM et al. Rev Aquac, 2016. Hounmanou YAG et al. Aquaculture Reports, 2021. 58:T5ff0,{"type":"doc","content":[{"type":"heading","attrs":{"id":"9c5ba187-6d06-42d0-8252-30a7a00f02bf","data-toc-id":"9c5ba187-6d06-42d0-8252-30a7a00f02bf","textAlign":"left","level":1},"content":[{"type":"text","text":"Proposal Template"}]},{"type":"heading","attrs":{"id":"7dd8f266-aecf-4b9c-8f82-d367f77fd815","data-toc-id":"7dd8f266-aecf-4b9c-8f82-d367f77fd815","textAlign":"left","level":2},"content":[{"type":"text","text":"1. Overview"}]},{"type":"paragraph","attrs":{"id":"271192d8-b908-4663-9f9b-e6e69217f3e3","class":null,"textAlign":"left"},"content":[{"type":"text","marks":[{"type":"bold"}],"text":"Phase 1 – Local AMR & Virulence Mapping of Major Tilapia Pathogens in Burkina Faso to Support Autovaccine Development"},{"type":"hardBreak"}]},{"type":"paragraph","attrs":{"id":"9c0ebf4f-b275-4b2f-84db-cbe0744ab762","class":null,"textAlign":"left"},"content":[{"type":"text","marks":[{"type":"textStyle","attrs":{"fontSize":null,"fontFamily":null,"color":"#121212"}},{"type":"bold"}],"text":"Scientific Rationale & Importance"},{"type":"hardBreak","marks":[{"type":"textStyle","attrs":{"fontSize":null,"fontFamily":null,"color":"#121212"}}]},{"type":"text","marks":[{"type":"textStyle","attrs":{"fontSize":"16px","fontFamily":"-apple-system, BlinkMacSystemFont, \"Segoe UI\", system-ui, -apple-system, \"Segoe UI\", Roboto, Ubuntu, Cantarell, \"Noto Sans\", sans-serif, Arial, \"PingFang SC\", \"Source Han Sans SC\", \"Microsoft YaHei UI\", \"Microsoft YaHei\", \"Noto Sans CJK SC\", sans-serif","color":"#121212"}}],"text":"Antibiotic overuse in tilapia farming drives antimicrobial resistance (AMR), threatening productivity, public health, and food security. "},{"type":"text","marks":[{"type":"textStyle","attrs":{"fontSize":null,"fontFamily":null,"color":"#121212"}},{"type":"italic"}],"text":"Streptococcus agalactiae"},{"type":"text","marks":[{"type":"textStyle","attrs":{"fontSize":"16px","fontFamily":"-apple-system, BlinkMacSystemFont, \"Segoe UI\", system-ui, -apple-system, \"Segoe UI\", Roboto, Ubuntu, Cantarell, \"Noto Sans\", sans-serif, Arial, \"PingFang SC\", \"Source Han Sans SC\", \"Microsoft YaHei UI\", \"Microsoft YaHei\", \"Noto Sans CJK SC\", sans-serif","color":"#121212"}}],"text":" and "},{"type":"text","marks":[{"type":"textStyle","attrs":{"fontSize":null,"fontFamily":null,"color":"#121212"}},{"type":"italic"}],"text":"Aeromonas hydrophila"},{"type":"text","marks":[{"type":"textStyle","attrs":{"fontSize":"16px","fontFamily":"-apple-system, BlinkMacSystemFont, \"Segoe UI\", system-ui, -apple-system, \"Segoe UI\", Roboto, Ubuntu, Cantarell, \"Noto Sans\", sans-serif, Arial, \"PingFang SC\", \"Source Han Sans SC\", \"Microsoft YaHei UI\", \"Microsoft YaHei\", \"Noto Sans CJK SC\", sans-serif","color":"#121212"}}],"text":" are major pathogens causing mass mortalities and economic losses. These bacteria often display multidrug resistance (MDR) and biofilm-forming ability, reducing treatment efficacy. This project will generate the first local AMR and virulence dataset from Burkina Faso to support the development of "},{"type":"text","marks":[{"type":"textStyle","attrs":{"fontSize":null,"fontFamily":null,"color":"#121212"}},{"type":"bold"}],"text":"preventive alternatives"},{"type":"text","marks":[{"type":"textStyle","attrs":{"fontSize":"16px","fontFamily":"-apple-system, BlinkMacSystemFont, \"Segoe UI\", system-ui, -apple-system, \"Segoe UI\", Roboto, Ubuntu, Cantarell, \"Noto Sans\", sans-serif, Arial, \"PingFang SC\", \"Source Han Sans SC\", \"Microsoft YaHei UI\", \"Microsoft YaHei\", \"Noto Sans CJK SC\", sans-serif","color":"#121212"}}],"text":", including "},{"type":"text","marks":[{"type":"textStyle","attrs":{"fontSize":null,"fontFamily":null,"color":"#121212"}},{"type":"bold"}],"text":"autogenous vaccines"},{"type":"text","marks":[{"type":"textStyle","attrs":{"fontSize":"16px","fontFamily":"-apple-system, BlinkMacSystemFont, \"Segoe UI\", system-ui, -apple-system, \"Segoe UI\", Roboto, Ubuntu, Cantarell, \"Noto Sans\", sans-serif, Arial, \"PingFang SC\", \"Source Han Sans SC\", \"Microsoft YaHei UI\", \"Microsoft YaHei\", \"Noto Sans CJK SC\", sans-serif","color":"#121212"}}],"text":" and "},{"type":"text","marks":[{"type":"textStyle","attrs":{"fontSize":null,"fontFamily":null,"color":"#121212"}},{"type":"bold"}],"text":"post-biotic adjuvants"},{"type":"text","marks":[{"type":"textStyle","attrs":{"fontSize":"16px","fontFamily":"-apple-system, BlinkMacSystemFont, \"Segoe UI\", system-ui, -apple-system, \"Segoe UI\", Roboto, Ubuntu, Cantarell, \"Noto Sans\", sans-serif, Arial, \"PingFang SC\", \"Source Han Sans SC\", \"Microsoft YaHei UI\", \"Microsoft YaHei\", \"Noto Sans CJK SC\", sans-serif","color":"#121212"}}],"text":", aligned with "},{"type":"text","marks":[{"type":"textStyle","attrs":{"fontSize":null,"fontFamily":null,"color":"#121212"}},{"type":"bold"}],"text":"One Health"},{"type":"text","marks":[{"type":"textStyle","attrs":{"fontSize":"16px","fontFamily":"-apple-system, BlinkMacSystemFont, \"Segoe UI\", system-ui, -apple-system, \"Segoe UI\", Roboto, Ubuntu, Cantarell, \"Noto Sans\", sans-serif, Arial, \"PingFang SC\", \"Source Han Sans SC\", \"Microsoft YaHei UI\", \"Microsoft YaHei\", \"Noto Sans CJK SC\", sans-serif","color":"#121212"}}],"text":" principles."}]},{"type":"paragraph","attrs":{"id":"f7054684-5b53-4900-9fbd-31b8f9441e1e","class":null,"textAlign":"left"},"content":[{"type":"text","marks":[{"type":"bold"}],"text":"Funding / Facilities / Ethics"}]},{"type":"bulletList","content":[{"type":"listItem","content":[{"type":"paragraph","attrs":{"id":"ce1c0740-dcf8-4e18-be0c-9f6fc1ee8f68","class":null,"textAlign":null},"content":[{"type":"text","marks":[{"type":"bold"}],"text":"Budget requested"},{"type":"text","text":": 5 000 USD"}]}]},{"type":"listItem","content":[{"type":"paragraph","attrs":{"id":"413a7719-3e7e-462a-9572-49efacb6e781","class":null,"textAlign":null},"content":[{"type":"text","marks":[{"type":"bold"}],"text":"Facilities"},{"type":"text","text":": Veterinary Microbiology Lab, Kırıkkale University, Turkey (PhD host); collaborative access to local lab in Burkina Faso for sampling."}]}]},{"type":"listItem","content":[{"type":"paragraph","attrs":{"id":"d1c0a168-5b8d-4f89-ada1-4ba7d8d0eb4a","class":null,"textAlign":null},"content":[{"type":"text","marks":[{"type":"bold"}],"text":"Ethics"},{"type":"text","text":": Non-invasive sampling; farm-level consent; no animal experimentation; BSL-2 compliance."}]}]}]},{"type":"paragraph","attrs":{"id":"a191084f-a363-490b-a2dd-0dc92ae087a8","class":null,"textAlign":null},"content":[{"type":"text","marks":[{"type":"bold"}],"text":"Timeline"},{"type":"hardBreak"},{"type":"text","text":"12 months (M1–M12): sampling, isolation/identification, antibiograms, virulence testing, data analysis, dissemination."}]},{"type":"paragraph","attrs":{"id":"9ef864bf-df7e-44b9-9c4a-b9fb84e2f5bd","class":null,"textAlign":null},"content":[{"type":"text","marks":[{"type":"bold"}],"text":"Data Sharing"},{"type":"hardBreak"},{"type":"text","text":"All AMR and virulence data will be deposited in an "},{"type":"text","marks":[{"type":"bold"}],"text":"open-access repository (Zenodo/OSF)"},{"type":"text","text":" within 3 months of completion. Strains will be available for non-commercial research under an MTA."}]},{"type":"heading","attrs":{"id":"bb3a2b07-742e-45fd-9cd4-ad61f29b58e9","data-toc-id":"bb3a2b07-742e-45fd-9cd4-ad61f29b58e9","textAlign":"left","level":2},"content":[{"type":"text","text":"2. Introduction"}]},{"type":"paragraph","attrs":{"id":"2690e203-a0ba-4d2c-bb8b-a6a7bcf46426","class":null,"textAlign":"left"},"content":[{"type":"text","text":"Theoretical background"},{"type":"hardBreak"},{"type":"text","text":"Antimicrobial resistance (AMR) is a global One-Health emergency. Intensive tilapia culture relies heavily on antibiotics, selecting for multidrug-resistant (MDR) pathogens that spread to humans via food and water. Two taxa drive mortality in African tilapia farms: Streptococcus agalactiae (streptococcosis; ≤80 % mortalities) and Aeromonas hydrophila (ulcerative syndrome, septicaemia). Both form biofilms and carry MDR plasmids, yet no baseline AMR or virulence data exist for Burkina Faso."}]},{"type":"paragraph","attrs":{"id":"ad3e3314-7101-44df-b557-af1f64693971","class":null,"textAlign":null},"content":[{"type":"text","text":"Key literature"},{"type":"hardBreak"},{"type":"text","text":"Asian and East-African surveys report tetracycline and fluoroquinolone resistance rates >60 % in these species (Chen 2020; Hounmanou 2021). Biofilm-positive isolates show 4–8× higher minimal inhibitory concentrations and persist in pond sediments (Watts 2017). Autogenous vaccines, adjuvanted with low-cost post-biotics, are emerging as sustainable alternatives, but strain selection requires local phenotypic and resistance maps—currently missing for West Africa."}]},{"type":"paragraph","attrs":{"id":"6dc03072-b7eb-4632-82e1-b8d48a239a99","class":null,"textAlign":null},"content":[{"type":"text","text":"Research questions"},{"type":"hardBreak"},{"type":"text","text":"RQ1 Which S. agalactiae and A. hydrophila genotypes dominate Burkinabè tilapia farms?"},{"type":"hardBreak"},{"type":"text","text":"RQ2 What are their AMR profiles against 12 antibiotic classes?"},{"type":"hardBreak"},{"type":"text","text":"RQ3 Do they exhibit high-virulence phenotypes (haemolysis, biofilm, extracellular enzymes) that warrant vaccine inclusion?"}]},{"type":"paragraph","attrs":{"id":"56aea961-54e8-400b-9a12-ff7e4c67fbc3","class":null,"textAlign":null},"content":[{"type":"text","text":"Numbered hypotheses"},{"type":"hardBreak"},{"type":"text","text":"H1 Local isolates display MDR (≥3 classes), especially to tetracyclines and fluoroquinolones."},{"type":"hardBreak"},{"type":"text","text":"H2 ≥50 % of isolates are strong biofilm-formers and β-haemolytic."},{"type":"hardBreak"},{"type":"text","text":"H3 A quantifiable subset combines high prevalence, MDR and virulence, providing optimal autovaccine candidates."}]},{"type":"paragraph","attrs":{"id":"3929802b-1b3c-45c5-aa72-ee680798cd9c","class":null,"textAlign":null},"content":[{"type":"text","text":"Impact statement"},{"type":"hardBreak"},{"type":"text","text":"Regardless of resistance levels, this first national surveillance will: (i) establish an open-access AMR/virulence atlas and strain bank; (ii) guide rational antimicrobial use; (iii) select candidate strains for an immediate PhD-driven autogenous vaccine trial with post-biotic adjuvant; and (iv) create a replicable One-Health surveillance model for West African aquaculture."}]},{"type":"heading","attrs":{"id":"9a198004-6d6c-474b-9c95-0637660588f5","data-toc-id":"9a198004-6d6c-474b-9c95-0637660588f5","textAlign":"left","level":2},"content":[{"type":"text","text":"3. Methods"}]},{"type":"heading","attrs":{"id":"a63b0f55-937d-4df8-895e-18ed16ba5da0","data-toc-id":"a63b0f55-937d-4df8-895e-18ed16ba5da0","textAlign":"left","level":3},"content":[{"type":"text","marks":[{"type":"bold"}],"text":"Participants / Sample"}]},{"type":"bulletList","content":[{"type":"listItem","content":[{"type":"paragraph","attrs":{"id":"b1c93d6b-2614-4fe9-a699-a064306ad6fa","class":null,"textAlign":null},"content":[{"type":"text","marks":[{"type":"bold"}],"text":"Sites"},{"type":"text","text":": 5-7 tilapia farms in Burkina Faso"}]}]},{"type":"listItem","content":[{"type":"paragraph","attrs":{"id":"8ba6d1dd-20aa-48c4-9a0f-2739330de602","class":null,"textAlign":null},"content":[{"type":"text","marks":[{"type":"bold"}],"text":"Samples"},{"type":"text","text":": gills, kidney, liver of diseased/suspect fish; pond water; equipment swabs"}]}]},{"type":"listItem","content":[{"type":"paragraph","attrs":{"id":"04583a90-9b0a-4e2e-ba2b-dba7adfcd505","class":null,"textAlign":null},"content":[{"type":"text","marks":[{"type":"bold"}],"text":"Exclusion"},{"type":"text","text":": fish treated with antibiotics <7 days"}]}]},{"type":"listItem","content":[{"type":"paragraph","attrs":{"id":"7bb5472c-1f49-416a-a616-ed76b7bea3d9","class":null,"textAlign":null},"content":[{"type":"text","marks":[{"type":"bold"}],"text":"Target"},{"type":"text","text":": 60–90 isolates (30–45 per species)"}]}]}]},{"type":"paragraph","attrs":{"id":"9dd53b01-6e05-4f74-a860-a705c74812fb","class":null,"textAlign":null},"content":[{"type":"text","marks":[{"type":"bold"}],"text":"Materials & Procedures"}]},{"type":"bulletList","content":[{"type":"listItem","content":[{"type":"paragraph","attrs":{"id":"276e75bf-c25a-402c-bde8-64a6099af04b","class":null,"textAlign":null},"content":[{"type":"text","marks":[{"type":"bold"}],"text":"Isolation & ID"},{"type":"text","text":": Todd-Hewitt, TSA, MacConkey, blood agar; Gram, catalase, oxidase; biochemical galleries (API or low-cost equivalent); optional species-specific PCR if budget allows."}]}]},{"type":"listItem","content":[{"type":"paragraph","attrs":{"id":"9fdfa226-979b-4164-aeeb-c7ceaf9080cd","class":null,"textAlign":null},"content":[{"type":"text","marks":[{"type":"bold"}],"text":"Antibiograms"},{"type":"text","text":": Kirby-Bauer, EUCAST/CLSI; panel: β-lactams, tetracyclines, fluoroquinolones, sulfonamides/trimethoprim, aminoglycosides, phenicols."}]}]},{"type":"listItem","content":[{"type":"paragraph","attrs":{"id":"2335c1ed-d3bd-4f7e-a39e-ff5aa45d96a8","class":null,"textAlign":null},"content":[{"type":"text","marks":[{"type":"bold"}],"text":"Virulence"},{"type":"text","text":": haemolysis on blood agar, crystal-violet biofilm (96-well), gelatinase/DNase plates."}]}]}]},{"type":"paragraph","attrs":{"id":"766dcf72-701d-481f-8ca1-5026ccc4eb12","class":null,"textAlign":null},"content":[{"type":"text","marks":[{"type":"bold"}],"text":"Planned Analyses"}]},{"type":"bulletList","content":[{"type":"listItem","content":[{"type":"paragraph","attrs":{"id":"43a0f041-37ef-4ba9-95b4-22f5a94e5096","class":null,"textAlign":null},"content":[{"type":"text","text":"Resistance rates & MDR index"}]}]},{"type":"listItem","content":[{"type":"paragraph","attrs":{"id":"25395038-4192-4bc1-a65e-fa4dc94d5008","class":null,"textAlign":null},"content":[{"type":"text","text":"Site comparisons (Chi²/Fisher)"}]}]},{"type":"listItem","content":[{"type":"paragraph","attrs":{"id":"82d39f39-d85b-415a-b9eb-3e2550b5b3c7","class":null,"textAlign":null},"content":[{"type":"text","text":"Biofilm: ANOVA/Tukey"}]}]},{"type":"listItem","content":[{"type":"paragraph","attrs":{"id":"ca99b4a0-b58a-4228-bf3e-113019b9b50b","class":null,"textAlign":null},"content":[{"type":"text","marks":[{"type":"bold"}],"text":"Strain prioritisation"},{"type":"text","text":": matrix combining prevalence, MDR, and virulence score"}]}]},{"type":"listItem","content":[{"type":"paragraph","attrs":{"id":"3b413e56-4d02-4098-9347-b09fb4752271","class":null,"textAlign":null},"content":[{"type":"text","marks":[{"type":"bold"}],"text":"Contingency"},{"type":"text","text":": if isolation rate < expected → increase sites/samples; if no PCR → focus on robust phenotyping"}]}]}]},{"type":"paragraph","attrs":{"id":"33c09255-9634-4232-ba0b-5772c77c217d","class":null,"textAlign":null},"content":[{"type":"text","marks":[{"type":"bold"}],"text":"Ethics & Data Management"}]},{"type":"bulletList","content":[{"type":"listItem","content":[{"type":"paragraph","attrs":{"id":"a31560e9-6e26-405a-94aa-02eed7d8bbc3","class":null,"textAlign":null},"content":[{"type":"text","text":"Farm consent; anonymised data"}]}]},{"type":"listItem","content":[{"type":"paragraph","attrs":{"id":"84d16e62-ee15-440f-80aa-ab0f36d24b88","class":null,"textAlign":null},"content":[{"type":"text","text":"BSL-2 biosafety; national waste regulations"}]}]},{"type":"listItem","content":[{"type":"paragraph","attrs":{"id":"dd64e690-3160-40e2-981d-8f742003ef0d","class":null,"textAlign":null},"content":[{"type":"text","marks":[{"type":"bold"}],"text":"Open data"},{"type":"text","text":": CSV + metadata → Zenodo/OSF (CC-BY 4.0)"}]}]}]},{"type":"heading","attrs":{"id":"4175674c-a1c3-4b07-8e07-3dee7835236f","data-toc-id":"4175674c-a1c3-4b07-8e07-3dee7835236f","textAlign":"left","level":2},"content":[{"type":"text","text":"4. Pilot Data (Optional)"}]},{"type":"paragraph","attrs":{"id":"a046f741-b909-46b7-ad7f-6781f7ae773a","class":null,"textAlign":"left"},"content":[{"type":"text","text":"Summarize any pilot data or simulations that demonstrate method feasibility and inform power analysis. Clearly distinguish from main study."}]},{"type":"heading","attrs":{"id":"eb573212-8614-4d87-b008-338c091cd3de","data-toc-id":"eb573212-8614-4d87-b008-338c091cd3de","textAlign":"left","level":2},"content":[{"type":"text","text":"5. Budget"}]},{"type":"table","attrs":{"id":"743c7178-b03e-4468-b227-5793dff6aca3"},"content":[{"type":"tableRow","content":[{"type":"tableCell","attrs":{"colspan":1,"rowspan":1,"colwidth":null,"style":null},"content":[{"type":"paragraph","attrs":{"id":"65229376-654a-4b27-904c-2caefaeae1a2","class":null,"textAlign":null}}]},{"type":"tableCell","attrs":{"colspan":1,"rowspan":1,"colwidth":null,"style":null},"content":[{"type":"paragraph","attrs":{"id":"c194caac-f065-47c5-8759-a7af41acd715","class":null,"textAlign":null},"content":[{"type":"text","text":"Category Cost (USD)"}]}]}]},{"type":"tableRow","content":[{"type":"tableCell","attrs":{"colspan":1,"rowspan":1,"colwidth":null,"style":null},"content":[{"type":"paragraph","attrs":{"id":"7a6e932e-c9c3-478b-a302-d275bdc6fc2a","class":null,"textAlign":null},"content":[{"type":"text","text":"Media & reagents (BHI/TSB, blood agar, Gram, antibiotic discs)"}]}]},{"type":"tableCell","attrs":{"colspan":1,"rowspan":1,"colwidth":null,"style":null},"content":[{"type":"paragraph","attrs":{"id":"1a16babb-ac32-4cbc-aa6b-8b61c7b1833d","class":null,"textAlign":null},"content":[{"type":"text","text":"1,200"}]}]}]},{"type":"tableRow","content":[{"type":"tableCell","attrs":{"colspan":1,"rowspan":1,"colwidth":null,"style":null},"content":[{"type":"paragraph","attrs":{"id":"6494bd60-04a2-4cc4-9bd5-b9d7b4c2c49a","class":null,"textAlign":null},"content":[{"type":"text","text":"Consumables (plates, swabs, tips, tubes, cryotubes)"}]}]},{"type":"tableCell","attrs":{"colspan":1,"rowspan":1,"colwidth":null,"style":null},"content":[{"type":"paragraph","attrs":{"id":"6398f6ea-58e5-4718-9b0f-6af87fbabc95","class":null,"textAlign":null},"content":[{"type":"text","text":"900"}]}]}]},{"type":"tableRow","content":[{"type":"tableCell","attrs":{"colspan":1,"rowspan":1,"colwidth":null,"style":null},"content":[{"type":"paragraph","attrs":{"id":"b1153220-294c-401f-8f56-8c6e5ed73e13","class":null,"textAlign":null},"content":[{"type":"text","text":"Transport/field (coolers, ice packs, travel)"}]}]},{"type":"tableCell","attrs":{"colspan":1,"rowspan":1,"colwidth":null,"style":null},"content":[{"type":"paragraph","attrs":{"id":"9e8770bf-ef0c-4dc6-b712-670fedac8226","class":null,"textAlign":null},"content":[{"type":"text","text":"600"}]}]}]},{"type":"tableRow","content":[{"type":"tableCell","attrs":{"colspan":1,"rowspan":1,"colwidth":null,"style":null},"content":[{"type":"paragraph","attrs":{"id":"653e5d1a-a9e4-49f4-8e52-9965cf46c735","class":null,"textAlign":null},"content":[{"type":"text","text":"Targeted confirmations (PCR/16S outsourced – optional)"}]}]},{"type":"tableCell","attrs":{"colspan":1,"rowspan":1,"colwidth":null,"style":null},"content":[{"type":"paragraph","attrs":{"id":"e415817d-70d9-49f2-8700-23e9fda9d73a","class":null,"textAlign":null},"content":[{"type":"text","text":"700"}]}]}]},{"type":"tableRow","content":[{"type":"tableCell","attrs":{"colspan":1,"rowspan":1,"colwidth":null,"style":null},"content":[{"type":"paragraph","attrs":{"id":"b242eff5-be4c-40e3-a243-4dcbe4f6665d","class":null,"textAlign":null},"content":[{"type":"text","text":"Small equipment (pipettes/calibration, racks, pH meter)"}]}]},{"type":"tableCell","attrs":{"colspan":1,"rowspan":1,"colwidth":null,"style":null},"content":[{"type":"paragraph","attrs":{"id":"20dd2019-7f82-4c12-b8eb-0c3b42909275","class":null,"textAlign":null},"content":[{"type":"text","text":"500"}]}]}]},{"type":"tableRow","content":[{"type":"tableCell","attrs":{"colspan":1,"rowspan":1,"colwidth":null,"style":null},"content":[{"type":"paragraph","attrs":{"id":"f872b2ef-2d34-42be-af94-db6b5d8dcc54","class":null,"textAlign":null},"content":[{"type":"text","text":"Quality & control (reference strains, control discs)"}]}]},{"type":"tableCell","attrs":{"colspan":1,"rowspan":1,"colwidth":null,"style":null},"content":[{"type":"paragraph","attrs":{"id":"c5e07412-7bf7-4fe6-8e23-1768211562a3","class":null,"textAlign":null},"content":[{"type":"text","text":"250"}]}]}]},{"type":"tableRow","content":[{"type":"tableCell","attrs":{"colspan":1,"rowspan":1,"colwidth":null,"style":null},"content":[{"type":"paragraph","attrs":{"id":"67b8cf74-fd38-4ea4-8cc2-f10e336ece4b","class":null,"textAlign":null},"content":[{"type":"text","text":"Communication & reporting (printing, farm briefings)"}]}]},{"type":"tableCell","attrs":{"colspan":1,"rowspan":1,"colwidth":null,"style":null},"content":[{"type":"paragraph","attrs":{"id":"76d9bb2d-9dc8-4c0a-b815-5bcc44b8e629","class":null,"textAlign":null},"content":[{"type":"text","text":"200"}]}]}]},{"type":"tableRow","content":[{"type":"tableCell","attrs":{"colspan":1,"rowspan":1,"colwidth":null,"style":null},"content":[{"type":"paragraph","attrs":{"id":"bf79fbed-5793-4eb7-982a-09839c7341c5","class":null,"textAlign":null},"content":[{"type":"text","text":"Contingency"}]}]},{"type":"tableCell","attrs":{"colspan":1,"rowspan":1,"colwidth":null,"style":null},"content":[{"type":"paragraph","attrs":{"id":"92f7ab6a-1284-4150-8eec-5b3e0a2ef971","class":null,"textAlign":null},"content":[{"type":"text","text":"350"}]}]}]},{"type":"tableRow","content":[{"type":"tableCell","attrs":{"colspan":1,"rowspan":1,"colwidth":null,"style":null},"content":[{"type":"paragraph","attrs":{"id":"d6092cbb-848e-433a-86a7-93568710d167","class":null,"textAlign":null},"content":[{"type":"text","marks":[{"type":"bold"}],"text":"Estimated total"}]}]},{"type":"tableCell","attrs":{"colspan":1,"rowspan":1,"colwidth":null,"style":null},"content":[{"type":"paragraph","attrs":{"id":"6b13e46c-8ff6-4f49-a0b9-3706c539671e","class":null,"textAlign":null},"content":[{"type":"text","marks":[{"type":"bold"}],"text":"4,700"}]}]}]}]},{"type":"paragraph","attrs":{"id":"9ef9488c-c18d-43bf-8d52-81dcc10a6e68","class":null,"textAlign":null}},{"type":"heading","attrs":{"id":"6af25317-e3ee-48b7-bb80-bbd5e58cd4cf","data-toc-id":"6af25317-e3ee-48b7-bb80-bbd5e58cd4cf","textAlign":"left","level":2},"content":[{"type":"text","text":"6. References"}]},{"type":"orderedList","attrs":{"start":1},"content":[{"type":"listItem","content":[{"type":"paragraph","attrs":{"id":"413debe7-0888-451d-93a1-e27c015ad41f","class":null,"textAlign":null},"content":[{"type":"text","text":"FAO. The State of World Fisheries and Aquaculture. 2022."}]}]},{"type":"listItem","content":[{"type":"paragraph","attrs":{"id":"1cc1f1f9-d818-430f-8811-8c9249e44125","class":null,"textAlign":null},"content":[{"type":"text","text":"Cabello FC. Environ Microbiol, 2006."}]}]},{"type":"listItem","content":[{"type":"paragraph","attrs":{"id":"d30d398d-bf5e-4e25-bd48-5a7af00ae374","class":null,"textAlign":null},"content":[{"type":"text","text":"Watts JEM et al. Front Microbiol, 2017."}]}]},{"type":"listItem","content":[{"type":"paragraph","attrs":{"id":"067aa5cc-21f4-4a14-8d1a-b2fc8e1f98f2","class":null,"textAlign":null},"content":[{"type":"text","text":"Amal MNA, Zamri-Saad M. Aquaculture, 2011."}]}]},{"type":"listItem","content":[{"type":"paragraph","attrs":{"id":"34396043-4931-46f8-a332-413b485a6a3b","class":null,"textAlign":null},"content":[{"type":"text","text":"Austin B, Austin DA. Bacterial Fish Pathogens. Springer, 2016."}]}]},{"type":"listItem","content":[{"type":"paragraph","attrs":{"id":"d7c4e520-75b1-4dce-9500-235dd64c19b6","class":null,"textAlign":null},"content":[{"type":"text","text":"Chen L et al. Sci Total Environ, 2020."}]}]},{"type":"listItem","content":[{"type":"paragraph","attrs":{"id":"a364b409-9cc3-42cd-aeb9-6e265263a9c8","class":null,"textAlign":null},"content":[{"type":"text","text":"Phu TM et al. Rev Aquac, 2016."}]}]},{"type":"listItem","content":[{"type":"paragraph","attrs":{"id":"5ff52888-ca7f-4313-931f-f019fdb9c332","class":null,"textAlign":null},"content":[{"type":"text","text":"Hounmanou YAG et al. Aquaculture Reports, 2021."}]}]}]},{"type":"paragraph","attrs":{"id":"edc167f3-3bca-405b-a253-025af29390d3","class":null,"textAlign":null},"content":[{"type":"hardBreak"},{"type":"hardBreak"}]}]}59:T168f,Proposal Template 1. Overview Phase 1 – Local AMR & Virulence Mapping of Major Tilapia Pathogens in Burkina Faso to Support Autovaccine Development Scientific Rationale & Importance Antibiotic overuse in tilapia farming drives antimicrobial resistance (AMR), threatening productivity, public health, and food security. Streptococcus agalactiae and Aeromonas hydrophila are major pathogens causing mass mortalities and economic losses. These bacteria often display multidrug resistance (MDR) and biofilm-forming ability, reducing treatment efficacy. This project will generate the first local AMR and virulence dataset from Burkina Faso to support the development of preventive alternatives, including autogenous vaccines and post-biotic adjuvants, aligned with One Health principles. Funding / Facilities / Ethics Budget requested: 5 000 USD Facilities: Veterinary Microbiology Lab, Kırıkkale University, Turkey (PhD host); collaborative access to local lab in Burkina Faso for sampling. Ethics: Non-invasive sampling; farm-level consent; no animal experimentation; BSL-2 compliance. Timeline 12 months (M1–M12): sampling, isolation/identification, antibiograms, virulence testing, data analysis, dissemination. Data Sharing All AMR and virulence data will be deposited in an open-access repository (Zenodo/OSF) within 3 months of completion. Strains will be available for non-commercial research under an MTA. 2. Introduction Theoretical background Antimicrobial resistance (AMR) is a global One-Health emergency. Intensive tilapia culture relies heavily on antibiotics, selecting for multidrug-resistant (MDR) pathogens that spread to humans via food and water. Two taxa drive mortality in African tilapia farms: Streptococcus agalactiae (streptococcosis; ≤80 % mortalities) and Aeromonas hydrophila (ulcerative syndrome, septicaemia). Both form biofilms and carry MDR plasmids, yet no baseline AMR or virulence data exist for Burkina Faso. Key literature Asian and East-African surveys report tetracycline and fluoroquinolone resistance rates >60 % in these species (Chen 2020; Hounmanou 2021). Biofilm-positive isolates show 4–8× higher minimal inhibitory concentrations and persist in pond sediments (Watts 2017). Autogenous vaccines, adjuvanted with low-cost post-biotics, are emerging as sustainable alternatives, but strain selection requires local phenotypic and resistance maps—currently missing for West Africa. Research questions RQ1 Which S. agalactiae and A. hydrophila genotypes dominate Burkinabè tilapia farms? RQ2 What are their AMR profiles against 12 antibiotic classes? RQ3 Do they exhibit high-virulence phenotypes (haemolysis, biofilm, extracellular enzymes) that warrant vaccine inclusion? Numbered hypotheses H1 Local isolates display MDR (≥3 classes), especially to tetracyclines and fluoroquinolones. H2 ≥50 % of isolates are strong biofilm-formers and β-haemolytic. H3 A quantifiable subset combines high prevalence, MDR and virulence, providing optimal autovaccine candidates. Impact statement Regardless of resistance levels, this first national surveillance will: (i) establish an open-access AMR/virulence atlas and strain bank; (ii) guide rational antimicrobial use; (iii) select candidate strains for an immediate PhD-driven autogenous vaccine trial with post-biotic adjuvant; and (iv) create a replicable One-Health surveillance model for West African aquaculture. 3. Methods Participants / Sample Sites: 5-7 tilapia farms in Burkina Faso Samples: gills, kidney, liver of diseased/suspect fish; pond water; equipment swabs Exclusion: fish treated with antibiotics <7 days Target: 60–90 isolates (30–45 per species) Materials & Procedures Isolation & ID: Todd-Hewitt, TSA, MacConkey, blood agar; Gram, catalase, oxidase; biochemical galleries (API or low-cost equivalent); optional species-specific PCR if budget allows. Antibiograms: Kirby-Bauer, EUCAST/CLSI; panel: β-lactams, tetracyclines, fluoroquinolones, sulfonamides/trimethoprim, aminoglycosides, phenicols. Virulence: haemolysis on blood agar, crystal-violet biofilm (96-well), gelatinase/DNase plates. Planned Analyses Resistance rates & MDR index Site comparisons (Chi²/Fisher) Biofilm: ANOVA/Tukey Strain prioritisation: matrix combining prevalence, MDR, and virulence score Contingency: if isolation rate < expected → increase sites/samples; if no PCR → focus on robust phenotyping Ethics & Data Management Farm consent; anonymised data BSL-2 biosafety; national waste regulations Open data: CSV + metadata → Zenodo/OSF (CC-BY 4.0) 4. Pilot Data (Optional) Summarize any pilot data or simulations that demonstrate method feasibility and inform power analysis. Clearly distinguish from main study. 5. Budget Category Cost (USD) Media & reagents (BHI/TSB, blood agar, Gram, antibiotic discs) 1,200 Consumables (plates, swabs, tips, tubes, cryotubes) 900 Transport/field (coolers, ice packs, travel) 600 Targeted confirmations (PCR/16S outsourced – optional) 700 Small equipment (pipettes/calibration, racks, pH meter) 500 Quality & control (reference strains, control discs) 250 Communication & reporting (printing, farm briefings) 200 Contingency 350 Estimated total 4,700 6. References FAO. The State of World Fisheries and Aquaculture. 2022. Cabello FC. Environ Microbiol, 2006. Watts JEM et al. Front Microbiol, 2017. Amal MNA, Zamri-Saad M. Aquaculture, 2011. Austin B, Austin DA. Bacterial Fish Pathogens. Springer, 2016. Chen L et al. Sci Total Environ, 2020. Phu TM et al. Rev Aquac, 2016. Hounmanou YAG et al. Aquaculture Reports, 2021. 5a:T5ff0,{"type":"doc","content":[{"type":"heading","attrs":{"id":"9c5ba187-6d06-42d0-8252-30a7a00f02bf","data-toc-id":"9c5ba187-6d06-42d0-8252-30a7a00f02bf","textAlign":"left","level":1},"content":[{"type":"text","text":"Proposal Template"}]},{"type":"heading","attrs":{"id":"7dd8f266-aecf-4b9c-8f82-d367f77fd815","data-toc-id":"7dd8f266-aecf-4b9c-8f82-d367f77fd815","textAlign":"left","level":2},"content":[{"type":"text","text":"1. Overview"}]},{"type":"paragraph","attrs":{"id":"271192d8-b908-4663-9f9b-e6e69217f3e3","class":null,"textAlign":"left"},"content":[{"type":"text","marks":[{"type":"bold"}],"text":"Phase 1 – Local AMR & Virulence Mapping of Major Tilapia Pathogens in Burkina Faso to Support Autovaccine Development"},{"type":"hardBreak"}]},{"type":"paragraph","attrs":{"id":"9c0ebf4f-b275-4b2f-84db-cbe0744ab762","class":null,"textAlign":"left"},"content":[{"type":"text","marks":[{"type":"textStyle","attrs":{"fontSize":null,"fontFamily":null,"color":"#121212"}},{"type":"bold"}],"text":"Scientific Rationale & Importance"},{"type":"hardBreak","marks":[{"type":"textStyle","attrs":{"fontSize":null,"fontFamily":null,"color":"#121212"}}]},{"type":"text","marks":[{"type":"textStyle","attrs":{"fontSize":"16px","fontFamily":"-apple-system, BlinkMacSystemFont, \"Segoe UI\", system-ui, -apple-system, \"Segoe UI\", Roboto, Ubuntu, Cantarell, \"Noto Sans\", sans-serif, Arial, \"PingFang SC\", \"Source Han Sans SC\", \"Microsoft YaHei UI\", \"Microsoft YaHei\", \"Noto Sans CJK SC\", sans-serif","color":"#121212"}}],"text":"Antibiotic overuse in tilapia farming drives antimicrobial resistance (AMR), threatening productivity, public health, and food security. "},{"type":"text","marks":[{"type":"textStyle","attrs":{"fontSize":null,"fontFamily":null,"color":"#121212"}},{"type":"italic"}],"text":"Streptococcus agalactiae"},{"type":"text","marks":[{"type":"textStyle","attrs":{"fontSize":"16px","fontFamily":"-apple-system, BlinkMacSystemFont, \"Segoe UI\", system-ui, -apple-system, \"Segoe UI\", Roboto, Ubuntu, Cantarell, \"Noto Sans\", sans-serif, Arial, \"PingFang SC\", \"Source Han Sans SC\", \"Microsoft YaHei UI\", \"Microsoft YaHei\", \"Noto Sans CJK SC\", sans-serif","color":"#121212"}}],"text":" and "},{"type":"text","marks":[{"type":"textStyle","attrs":{"fontSize":null,"fontFamily":null,"color":"#121212"}},{"type":"italic"}],"text":"Aeromonas hydrophila"},{"type":"text","marks":[{"type":"textStyle","attrs":{"fontSize":"16px","fontFamily":"-apple-system, BlinkMacSystemFont, \"Segoe UI\", system-ui, -apple-system, \"Segoe UI\", Roboto, Ubuntu, Cantarell, \"Noto Sans\", sans-serif, Arial, \"PingFang SC\", \"Source Han Sans SC\", \"Microsoft YaHei UI\", \"Microsoft YaHei\", \"Noto Sans CJK SC\", sans-serif","color":"#121212"}}],"text":" are major pathogens causing mass mortalities and economic losses. These bacteria often display multidrug resistance (MDR) and biofilm-forming ability, reducing treatment efficacy. This project will generate the first local AMR and virulence dataset from Burkina Faso to support the development of "},{"type":"text","marks":[{"type":"textStyle","attrs":{"fontSize":null,"fontFamily":null,"color":"#121212"}},{"type":"bold"}],"text":"preventive alternatives"},{"type":"text","marks":[{"type":"textStyle","attrs":{"fontSize":"16px","fontFamily":"-apple-system, BlinkMacSystemFont, \"Segoe UI\", system-ui, -apple-system, \"Segoe UI\", Roboto, Ubuntu, Cantarell, \"Noto Sans\", sans-serif, Arial, \"PingFang SC\", \"Source Han Sans SC\", \"Microsoft YaHei UI\", \"Microsoft YaHei\", \"Noto Sans CJK SC\", sans-serif","color":"#121212"}}],"text":", including "},{"type":"text","marks":[{"type":"textStyle","attrs":{"fontSize":null,"fontFamily":null,"color":"#121212"}},{"type":"bold"}],"text":"autogenous vaccines"},{"type":"text","marks":[{"type":"textStyle","attrs":{"fontSize":"16px","fontFamily":"-apple-system, BlinkMacSystemFont, \"Segoe UI\", system-ui, -apple-system, \"Segoe UI\", Roboto, Ubuntu, Cantarell, \"Noto Sans\", sans-serif, Arial, \"PingFang SC\", \"Source Han Sans SC\", \"Microsoft YaHei UI\", \"Microsoft YaHei\", \"Noto Sans CJK SC\", sans-serif","color":"#121212"}}],"text":" and "},{"type":"text","marks":[{"type":"textStyle","attrs":{"fontSize":null,"fontFamily":null,"color":"#121212"}},{"type":"bold"}],"text":"post-biotic adjuvants"},{"type":"text","marks":[{"type":"textStyle","attrs":{"fontSize":"16px","fontFamily":"-apple-system, BlinkMacSystemFont, \"Segoe UI\", system-ui, -apple-system, \"Segoe UI\", Roboto, Ubuntu, Cantarell, \"Noto Sans\", sans-serif, Arial, \"PingFang SC\", \"Source Han Sans SC\", \"Microsoft YaHei UI\", \"Microsoft YaHei\", \"Noto Sans CJK SC\", sans-serif","color":"#121212"}}],"text":", aligned with "},{"type":"text","marks":[{"type":"textStyle","attrs":{"fontSize":null,"fontFamily":null,"color":"#121212"}},{"type":"bold"}],"text":"One Health"},{"type":"text","marks":[{"type":"textStyle","attrs":{"fontSize":"16px","fontFamily":"-apple-system, BlinkMacSystemFont, \"Segoe UI\", system-ui, -apple-system, \"Segoe UI\", Roboto, Ubuntu, Cantarell, \"Noto Sans\", sans-serif, Arial, \"PingFang SC\", \"Source Han Sans SC\", \"Microsoft YaHei UI\", \"Microsoft YaHei\", \"Noto Sans CJK SC\", sans-serif","color":"#121212"}}],"text":" principles."}]},{"type":"paragraph","attrs":{"id":"f7054684-5b53-4900-9fbd-31b8f9441e1e","class":null,"textAlign":"left"},"content":[{"type":"text","marks":[{"type":"bold"}],"text":"Funding / Facilities / Ethics"}]},{"type":"bulletList","content":[{"type":"listItem","content":[{"type":"paragraph","attrs":{"id":"ce1c0740-dcf8-4e18-be0c-9f6fc1ee8f68","class":null,"textAlign":null},"content":[{"type":"text","marks":[{"type":"bold"}],"text":"Budget requested"},{"type":"text","text":": 5 000 USD"}]}]},{"type":"listItem","content":[{"type":"paragraph","attrs":{"id":"413a7719-3e7e-462a-9572-49efacb6e781","class":null,"textAlign":null},"content":[{"type":"text","marks":[{"type":"bold"}],"text":"Facilities"},{"type":"text","text":": Veterinary Microbiology Lab, Kırıkkale University, Turkey (PhD host); collaborative access to local lab in Burkina Faso for sampling."}]}]},{"type":"listItem","content":[{"type":"paragraph","attrs":{"id":"d1c0a168-5b8d-4f89-ada1-4ba7d8d0eb4a","class":null,"textAlign":null},"content":[{"type":"text","marks":[{"type":"bold"}],"text":"Ethics"},{"type":"text","text":": Non-invasive sampling; farm-level consent; no animal experimentation; BSL-2 compliance."}]}]}]},{"type":"paragraph","attrs":{"id":"a191084f-a363-490b-a2dd-0dc92ae087a8","class":null,"textAlign":null},"content":[{"type":"text","marks":[{"type":"bold"}],"text":"Timeline"},{"type":"hardBreak"},{"type":"text","text":"12 months (M1–M12): sampling, isolation/identification, antibiograms, virulence testing, data analysis, dissemination."}]},{"type":"paragraph","attrs":{"id":"9ef864bf-df7e-44b9-9c4a-b9fb84e2f5bd","class":null,"textAlign":null},"content":[{"type":"text","marks":[{"type":"bold"}],"text":"Data Sharing"},{"type":"hardBreak"},{"type":"text","text":"All AMR and virulence data will be deposited in an "},{"type":"text","marks":[{"type":"bold"}],"text":"open-access repository (Zenodo/OSF)"},{"type":"text","text":" within 3 months of completion. Strains will be available for non-commercial research under an MTA."}]},{"type":"heading","attrs":{"id":"bb3a2b07-742e-45fd-9cd4-ad61f29b58e9","data-toc-id":"bb3a2b07-742e-45fd-9cd4-ad61f29b58e9","textAlign":"left","level":2},"content":[{"type":"text","text":"2. Introduction"}]},{"type":"paragraph","attrs":{"id":"2690e203-a0ba-4d2c-bb8b-a6a7bcf46426","class":null,"textAlign":"left"},"content":[{"type":"text","text":"Theoretical background"},{"type":"hardBreak"},{"type":"text","text":"Antimicrobial resistance (AMR) is a global One-Health emergency. Intensive tilapia culture relies heavily on antibiotics, selecting for multidrug-resistant (MDR) pathogens that spread to humans via food and water. Two taxa drive mortality in African tilapia farms: Streptococcus agalactiae (streptococcosis; ≤80 % mortalities) and Aeromonas hydrophila (ulcerative syndrome, septicaemia). Both form biofilms and carry MDR plasmids, yet no baseline AMR or virulence data exist for Burkina Faso."}]},{"type":"paragraph","attrs":{"id":"ad3e3314-7101-44df-b557-af1f64693971","class":null,"textAlign":null},"content":[{"type":"text","text":"Key literature"},{"type":"hardBreak"},{"type":"text","text":"Asian and East-African surveys report tetracycline and fluoroquinolone resistance rates >60 % in these species (Chen 2020; Hounmanou 2021). Biofilm-positive isolates show 4–8× higher minimal inhibitory concentrations and persist in pond sediments (Watts 2017). Autogenous vaccines, adjuvanted with low-cost post-biotics, are emerging as sustainable alternatives, but strain selection requires local phenotypic and resistance maps—currently missing for West Africa."}]},{"type":"paragraph","attrs":{"id":"6dc03072-b7eb-4632-82e1-b8d48a239a99","class":null,"textAlign":null},"content":[{"type":"text","text":"Research questions"},{"type":"hardBreak"},{"type":"text","text":"RQ1 Which S. agalactiae and A. hydrophila genotypes dominate Burkinabè tilapia farms?"},{"type":"hardBreak"},{"type":"text","text":"RQ2 What are their AMR profiles against 12 antibiotic classes?"},{"type":"hardBreak"},{"type":"text","text":"RQ3 Do they exhibit high-virulence phenotypes (haemolysis, biofilm, extracellular enzymes) that warrant vaccine inclusion?"}]},{"type":"paragraph","attrs":{"id":"56aea961-54e8-400b-9a12-ff7e4c67fbc3","class":null,"textAlign":null},"content":[{"type":"text","text":"Numbered hypotheses"},{"type":"hardBreak"},{"type":"text","text":"H1 Local isolates display MDR (≥3 classes), especially to tetracyclines and fluoroquinolones."},{"type":"hardBreak"},{"type":"text","text":"H2 ≥50 % of isolates are strong biofilm-formers and β-haemolytic."},{"type":"hardBreak"},{"type":"text","text":"H3 A quantifiable subset combines high prevalence, MDR and virulence, providing optimal autovaccine candidates."}]},{"type":"paragraph","attrs":{"id":"3929802b-1b3c-45c5-aa72-ee680798cd9c","class":null,"textAlign":null},"content":[{"type":"text","text":"Impact statement"},{"type":"hardBreak"},{"type":"text","text":"Regardless of resistance levels, this first national surveillance will: (i) establish an open-access AMR/virulence atlas and strain bank; (ii) guide rational antimicrobial use; (iii) select candidate strains for an immediate PhD-driven autogenous vaccine trial with post-biotic adjuvant; and (iv) create a replicable One-Health surveillance model for West African aquaculture."}]},{"type":"heading","attrs":{"id":"9a198004-6d6c-474b-9c95-0637660588f5","data-toc-id":"9a198004-6d6c-474b-9c95-0637660588f5","textAlign":"left","level":2},"content":[{"type":"text","text":"3. Methods"}]},{"type":"heading","attrs":{"id":"a63b0f55-937d-4df8-895e-18ed16ba5da0","data-toc-id":"a63b0f55-937d-4df8-895e-18ed16ba5da0","textAlign":"left","level":3},"content":[{"type":"text","marks":[{"type":"bold"}],"text":"Participants / Sample"}]},{"type":"bulletList","content":[{"type":"listItem","content":[{"type":"paragraph","attrs":{"id":"b1c93d6b-2614-4fe9-a699-a064306ad6fa","class":null,"textAlign":null},"content":[{"type":"text","marks":[{"type":"bold"}],"text":"Sites"},{"type":"text","text":": 5-7 tilapia farms in Burkina Faso"}]}]},{"type":"listItem","content":[{"type":"paragraph","attrs":{"id":"8ba6d1dd-20aa-48c4-9a0f-2739330de602","class":null,"textAlign":null},"content":[{"type":"text","marks":[{"type":"bold"}],"text":"Samples"},{"type":"text","text":": gills, kidney, liver of diseased/suspect fish; pond water; equipment swabs"}]}]},{"type":"listItem","content":[{"type":"paragraph","attrs":{"id":"04583a90-9b0a-4e2e-ba2b-dba7adfcd505","class":null,"textAlign":null},"content":[{"type":"text","marks":[{"type":"bold"}],"text":"Exclusion"},{"type":"text","text":": fish treated with antibiotics <7 days"}]}]},{"type":"listItem","content":[{"type":"paragraph","attrs":{"id":"7bb5472c-1f49-416a-a616-ed76b7bea3d9","class":null,"textAlign":null},"content":[{"type":"text","marks":[{"type":"bold"}],"text":"Target"},{"type":"text","text":": 60–90 isolates (30–45 per species)"}]}]}]},{"type":"paragraph","attrs":{"id":"9dd53b01-6e05-4f74-a860-a705c74812fb","class":null,"textAlign":null},"content":[{"type":"text","marks":[{"type":"bold"}],"text":"Materials & Procedures"}]},{"type":"bulletList","content":[{"type":"listItem","content":[{"type":"paragraph","attrs":{"id":"276e75bf-c25a-402c-bde8-64a6099af04b","class":null,"textAlign":null},"content":[{"type":"text","marks":[{"type":"bold"}],"text":"Isolation & ID"},{"type":"text","text":": Todd-Hewitt, TSA, MacConkey, blood agar; Gram, catalase, oxidase; biochemical galleries (API or low-cost equivalent); optional species-specific PCR if budget allows."}]}]},{"type":"listItem","content":[{"type":"paragraph","attrs":{"id":"9fdfa226-979b-4164-aeeb-c7ceaf9080cd","class":null,"textAlign":null},"content":[{"type":"text","marks":[{"type":"bold"}],"text":"Antibiograms"},{"type":"text","text":": Kirby-Bauer, EUCAST/CLSI; panel: β-lactams, tetracyclines, fluoroquinolones, sulfonamides/trimethoprim, aminoglycosides, phenicols."}]}]},{"type":"listItem","content":[{"type":"paragraph","attrs":{"id":"2335c1ed-d3bd-4f7e-a39e-ff5aa45d96a8","class":null,"textAlign":null},"content":[{"type":"text","marks":[{"type":"bold"}],"text":"Virulence"},{"type":"text","text":": haemolysis on blood agar, crystal-violet biofilm (96-well), gelatinase/DNase plates."}]}]}]},{"type":"paragraph","attrs":{"id":"766dcf72-701d-481f-8ca1-5026ccc4eb12","class":null,"textAlign":null},"content":[{"type":"text","marks":[{"type":"bold"}],"text":"Planned Analyses"}]},{"type":"bulletList","content":[{"type":"listItem","content":[{"type":"paragraph","attrs":{"id":"43a0f041-37ef-4ba9-95b4-22f5a94e5096","class":null,"textAlign":null},"content":[{"type":"text","text":"Resistance rates & MDR index"}]}]},{"type":"listItem","content":[{"type":"paragraph","attrs":{"id":"25395038-4192-4bc1-a65e-fa4dc94d5008","class":null,"textAlign":null},"content":[{"type":"text","text":"Site comparisons (Chi²/Fisher)"}]}]},{"type":"listItem","content":[{"type":"paragraph","attrs":{"id":"82d39f39-d85b-415a-b9eb-3e2550b5b3c7","class":null,"textAlign":null},"content":[{"type":"text","text":"Biofilm: ANOVA/Tukey"}]}]},{"type":"listItem","content":[{"type":"paragraph","attrs":{"id":"ca99b4a0-b58a-4228-bf3e-113019b9b50b","class":null,"textAlign":null},"content":[{"type":"text","marks":[{"type":"bold"}],"text":"Strain prioritisation"},{"type":"text","text":": matrix combining prevalence, MDR, and virulence score"}]}]},{"type":"listItem","content":[{"type":"paragraph","attrs":{"id":"3b413e56-4d02-4098-9347-b09fb4752271","class":null,"textAlign":null},"content":[{"type":"text","marks":[{"type":"bold"}],"text":"Contingency"},{"type":"text","text":": if isolation rate < expected → increase sites/samples; if no PCR → focus on robust phenotyping"}]}]}]},{"type":"paragraph","attrs":{"id":"33c09255-9634-4232-ba0b-5772c77c217d","class":null,"textAlign":null},"content":[{"type":"text","marks":[{"type":"bold"}],"text":"Ethics & Data Management"}]},{"type":"bulletList","content":[{"type":"listItem","content":[{"type":"paragraph","attrs":{"id":"a31560e9-6e26-405a-94aa-02eed7d8bbc3","class":null,"textAlign":null},"content":[{"type":"text","text":"Farm consent; anonymised data"}]}]},{"type":"listItem","content":[{"type":"paragraph","attrs":{"id":"84d16e62-ee15-440f-80aa-ab0f36d24b88","class":null,"textAlign":null},"content":[{"type":"text","text":"BSL-2 biosafety; national waste regulations"}]}]},{"type":"listItem","content":[{"type":"paragraph","attrs":{"id":"dd64e690-3160-40e2-981d-8f742003ef0d","class":null,"textAlign":null},"content":[{"type":"text","marks":[{"type":"bold"}],"text":"Open data"},{"type":"text","text":": CSV + metadata → Zenodo/OSF (CC-BY 4.0)"}]}]}]},{"type":"heading","attrs":{"id":"4175674c-a1c3-4b07-8e07-3dee7835236f","data-toc-id":"4175674c-a1c3-4b07-8e07-3dee7835236f","textAlign":"left","level":2},"content":[{"type":"text","text":"4. Pilot Data (Optional)"}]},{"type":"paragraph","attrs":{"id":"a046f741-b909-46b7-ad7f-6781f7ae773a","class":null,"textAlign":"left"},"content":[{"type":"text","text":"Summarize any pilot data or simulations that demonstrate method feasibility and inform power analysis. Clearly distinguish from main study."}]},{"type":"heading","attrs":{"id":"eb573212-8614-4d87-b008-338c091cd3de","data-toc-id":"eb573212-8614-4d87-b008-338c091cd3de","textAlign":"left","level":2},"content":[{"type":"text","text":"5. Budget"}]},{"type":"table","attrs":{"id":"743c7178-b03e-4468-b227-5793dff6aca3"},"content":[{"type":"tableRow","content":[{"type":"tableCell","attrs":{"colspan":1,"rowspan":1,"colwidth":null,"style":null},"content":[{"type":"paragraph","attrs":{"id":"65229376-654a-4b27-904c-2caefaeae1a2","class":null,"textAlign":null}}]},{"type":"tableCell","attrs":{"colspan":1,"rowspan":1,"colwidth":null,"style":null},"content":[{"type":"paragraph","attrs":{"id":"c194caac-f065-47c5-8759-a7af41acd715","class":null,"textAlign":null},"content":[{"type":"text","text":"Category Cost (USD)"}]}]}]},{"type":"tableRow","content":[{"type":"tableCell","attrs":{"colspan":1,"rowspan":1,"colwidth":null,"style":null},"content":[{"type":"paragraph","attrs":{"id":"7a6e932e-c9c3-478b-a302-d275bdc6fc2a","class":null,"textAlign":null},"content":[{"type":"text","text":"Media & reagents (BHI/TSB, blood agar, Gram, antibiotic discs)"}]}]},{"type":"tableCell","attrs":{"colspan":1,"rowspan":1,"colwidth":null,"style":null},"content":[{"type":"paragraph","attrs":{"id":"1a16babb-ac32-4cbc-aa6b-8b61c7b1833d","class":null,"textAlign":null},"content":[{"type":"text","text":"1,200"}]}]}]},{"type":"tableRow","content":[{"type":"tableCell","attrs":{"colspan":1,"rowspan":1,"colwidth":null,"style":null},"content":[{"type":"paragraph","attrs":{"id":"6494bd60-04a2-4cc4-9bd5-b9d7b4c2c49a","class":null,"textAlign":null},"content":[{"type":"text","text":"Consumables (plates, swabs, tips, tubes, cryotubes)"}]}]},{"type":"tableCell","attrs":{"colspan":1,"rowspan":1,"colwidth":null,"style":null},"content":[{"type":"paragraph","attrs":{"id":"6398f6ea-58e5-4718-9b0f-6af87fbabc95","class":null,"textAlign":null},"content":[{"type":"text","text":"900"}]}]}]},{"type":"tableRow","content":[{"type":"tableCell","attrs":{"colspan":1,"rowspan":1,"colwidth":null,"style":null},"content":[{"type":"paragraph","attrs":{"id":"b1153220-294c-401f-8f56-8c6e5ed73e13","class":null,"textAlign":null},"content":[{"type":"text","text":"Transport/field (coolers, ice packs, travel)"}]}]},{"type":"tableCell","attrs":{"colspan":1,"rowspan":1,"colwidth":null,"style":null},"content":[{"type":"paragraph","attrs":{"id":"9e8770bf-ef0c-4dc6-b712-670fedac8226","class":null,"textAlign":null},"content":[{"type":"text","text":"600"}]}]}]},{"type":"tableRow","content":[{"type":"tableCell","attrs":{"colspan":1,"rowspan":1,"colwidth":null,"style":null},"content":[{"type":"paragraph","attrs":{"id":"653e5d1a-a9e4-49f4-8e52-9965cf46c735","class":null,"textAlign":null},"content":[{"type":"text","text":"Targeted confirmations (PCR/16S outsourced – optional)"}]}]},{"type":"tableCell","attrs":{"colspan":1,"rowspan":1,"colwidth":null,"style":null},"content":[{"type":"paragraph","attrs":{"id":"e415817d-70d9-49f2-8700-23e9fda9d73a","class":null,"textAlign":null},"content":[{"type":"text","text":"700"}]}]}]},{"type":"tableRow","content":[{"type":"tableCell","attrs":{"colspan":1,"rowspan":1,"colwidth":null,"style":null},"content":[{"type":"paragraph","attrs":{"id":"b242eff5-be4c-40e3-a243-4dcbe4f6665d","class":null,"textAlign":null},"content":[{"type":"text","text":"Small equipment (pipettes/calibration, racks, pH 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discs)"}]}]},{"type":"tableCell","attrs":{"colspan":1,"rowspan":1,"colwidth":null,"style":null},"content":[{"type":"paragraph","attrs":{"id":"c5e07412-7bf7-4fe6-8e23-1768211562a3","class":null,"textAlign":null},"content":[{"type":"text","text":"250"}]}]}]},{"type":"tableRow","content":[{"type":"tableCell","attrs":{"colspan":1,"rowspan":1,"colwidth":null,"style":null},"content":[{"type":"paragraph","attrs":{"id":"67b8cf74-fd38-4ea4-8cc2-f10e336ece4b","class":null,"textAlign":null},"content":[{"type":"text","text":"Communication & reporting (printing, farm briefings)"}]}]},{"type":"tableCell","attrs":{"colspan":1,"rowspan":1,"colwidth":null,"style":null},"content":[{"type":"paragraph","attrs":{"id":"76d9bb2d-9dc8-4c0a-b815-5bcc44b8e629","class":null,"textAlign":null},"content":[{"type":"text","text":"200"}]}]}]},{"type":"tableRow","content":[{"type":"tableCell","attrs":{"colspan":1,"rowspan":1,"colwidth":null,"style":null},"content":[{"type":"paragraph","attrs":{"id":"bf79fbed-5793-4eb7-982a-09839c7341c5","class":null,"textAlign":null},"content":[{"type":"text","text":"Contingency"}]}]},{"type":"tableCell","attrs":{"colspan":1,"rowspan":1,"colwidth":null,"style":null},"content":[{"type":"paragraph","attrs":{"id":"92f7ab6a-1284-4150-8eec-5b3e0a2ef971","class":null,"textAlign":null},"content":[{"type":"text","text":"350"}]}]}]},{"type":"tableRow","content":[{"type":"tableCell","attrs":{"colspan":1,"rowspan":1,"colwidth":null,"style":null},"content":[{"type":"paragraph","attrs":{"id":"d6092cbb-848e-433a-86a7-93568710d167","class":null,"textAlign":null},"content":[{"type":"text","marks":[{"type":"bold"}],"text":"Estimated 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References"}]},{"type":"orderedList","attrs":{"start":1},"content":[{"type":"listItem","content":[{"type":"paragraph","attrs":{"id":"413debe7-0888-451d-93a1-e27c015ad41f","class":null,"textAlign":null},"content":[{"type":"text","text":"FAO. The State of World Fisheries and Aquaculture. 2022."}]}]},{"type":"listItem","content":[{"type":"paragraph","attrs":{"id":"1cc1f1f9-d818-430f-8811-8c9249e44125","class":null,"textAlign":null},"content":[{"type":"text","text":"Cabello FC. Environ Microbiol, 2006."}]}]},{"type":"listItem","content":[{"type":"paragraph","attrs":{"id":"d30d398d-bf5e-4e25-bd48-5a7af00ae374","class":null,"textAlign":null},"content":[{"type":"text","text":"Watts JEM et al. Front Microbiol, 2017."}]}]},{"type":"listItem","content":[{"type":"paragraph","attrs":{"id":"067aa5cc-21f4-4a14-8d1a-b2fc8e1f98f2","class":null,"textAlign":null},"content":[{"type":"text","text":"Amal MNA, Zamri-Saad M. Aquaculture, 2011."}]}]},{"type":"listItem","content":[{"type":"paragraph","attrs":{"id":"34396043-4931-46f8-a332-413b485a6a3b","class":null,"textAlign":null},"content":[{"type":"text","text":"Austin B, Austin DA. Bacterial Fish Pathogens. Springer, 2016."}]}]},{"type":"listItem","content":[{"type":"paragraph","attrs":{"id":"d7c4e520-75b1-4dce-9500-235dd64c19b6","class":null,"textAlign":null},"content":[{"type":"text","text":"Chen L et al. Sci Total Environ, 2020."}]}]},{"type":"listItem","content":[{"type":"paragraph","attrs":{"id":"a364b409-9cc3-42cd-aeb9-6e265263a9c8","class":null,"textAlign":null},"content":[{"type":"text","text":"Phu TM et al. Rev Aquac, 2016."}]}]},{"type":"listItem","content":[{"type":"paragraph","attrs":{"id":"5ff52888-ca7f-4313-931f-f019fdb9c332","class":null,"textAlign":null},"content":[{"type":"text","text":"Hounmanou YAG et al. 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Phase 1 – Local AMR & Virulence Mapping of Major Tilapia Pathogens in Burkina Faso to Support Autovaccine Development

1. Overview

Phase 1 – Local AMR & Virulence Mapping of Major Tilapia Pathogens in Burkina Faso to Support Autovaccine Development

Scientific Rationale & Importance
Antibiotic overuse in tilapia farming drives antimicrobial resistance (AMR), threatening productivity, public health, and food security. Streptococcus agalactiae and Aeromonas hydrophila are major pathogens causing mass mortalities and economic losses. These bacteria often display multidrug resistance (MDR) and biofilm-forming ability, reducing treatment efficacy. This project will generate the first local AMR and virulence dataset from Burkina Faso to support the development of preventive alternatives, including autogenous vaccines and post-biotic adjuvants, aligned with One Health principles.

Funding / Facilities / Ethics

Budget requested: 5 000 USD
Facilities: Veterinary Microbiology Lab, Kırıkkale University, Turkey (PhD host); collaborative access to local lab in Burkina Faso for sampling.
Ethics: Non-invasive sampling; farm-level consent; no animal experimentation; BSL-2 compliance.

Timeline
12 months (M1–M12): sampling, isolation/identification, antibiograms, virulence testing, data analysis, dissemination.

Data Sharing
All AMR and virulence data will be deposited in an open-access repository (Zenodo/OSF) within 3 months of completion. Strains will be available for non-commercial research under an MTA.

2. Introduction

Theoretical background
Antimicrobial resistance (AMR) is a global One-Health emergency. Intensive tilapia culture relies heavily on antibiotics, selecting for multidrug-resistant (MDR) pathogens that spread to humans via food and water. Two taxa drive mortality in African tilapia farms: Streptococcus agalactiae (streptococcosis; ≤80 % mortalities) and Aeromonas hydrophila (ulcerative syndrome, septicaemia). Both form biofilms and carry MDR plasmids, yet no baseline AMR or virulence data exist for Burkina Faso.

Key literature
Asian and East-African surveys report tetracycline and fluoroquinolone resistance rates >60 % in these species (Chen 2020; Hounmanou 2021). Biofilm-positive isolates show 4–8× higher minimal inhibitory concentrations and persist in pond sediments (Watts 2017). Autogenous vaccines, adjuvanted with low-cost post-biotics, are emerging as sustainable alternatives, but strain selection requires local phenotypic and resistance maps—currently missing for West Africa.

Research questions
RQ1 Which S. agalactiae and A. hydrophila genotypes dominate Burkinabè tilapia farms?
RQ2 What are their AMR profiles against 12 antibiotic classes?
RQ3 Do they exhibit high-virulence phenotypes (haemolysis, biofilm, extracellular enzymes) that warrant vaccine inclusion?

Numbered hypotheses
H1 Local isolates display MDR (≥3 classes), especially to tetracyclines and fluoroquinolones.
H2 ≥50 % of isolates are strong biofilm-formers and β-haemolytic.
H3 A quantifiable subset combines high prevalence, MDR and virulence, providing optimal autovaccine candidates.

Impact statement
Regardless of resistance levels, this first national surveillance will: (i) establish an open-access AMR/virulence atlas and strain bank; (ii) guide rational antimicrobial use; (iii) select candidate strains for an immediate PhD-driven autogenous vaccine trial with post-biotic adjuvant; and (iv) create a replicable One-Health surveillance model for West African aquaculture.

3. Methods

Participants / Sample

Sites: 5-7 tilapia farms in Burkina Faso
Samples: gills, kidney, liver of diseased/suspect fish; pond water; equipment swabs
Exclusion: fish treated with antibiotics <7 days
Target: 60–90 isolates (30–45 per species)

Materials & Procedures

Isolation & ID: Todd-Hewitt, TSA, MacConkey, blood agar; Gram, catalase, oxidase; biochemical galleries (API or low-cost equivalent); optional species-specific PCR if budget allows.
Antibiograms: Kirby-Bauer, EUCAST/CLSI; panel: β-lactams, tetracyclines, fluoroquinolones, sulfonamides/trimethoprim, aminoglycosides, phenicols.
Virulence: haemolysis on blood agar, crystal-violet biofilm (96-well), gelatinase/DNase plates.

Planned Analyses

Resistance rates & MDR index
Site comparisons (Chi²/Fisher)
Biofilm: ANOVA/Tukey
Strain prioritisation: matrix combining prevalence, MDR, and virulence score
Contingency: if isolation rate < expected → increase sites/samples; if no PCR → focus on robust phenotyping

Ethics & Data Management

Farm consent; anonymised data
BSL-2 biosafety; national waste regulations
Open data: CSV + metadata → Zenodo/OSF (CC-BY 4.0)

4. Pilot Data (Optional)

Summarize any pilot data or simulations that demonstrate method feasibility and inform power analysis. Clearly distinguish from main study.

5. Budget

	Category Cost (USD)
Media & reagents (BHI/TSB, blood agar, Gram, antibiotic discs)	1,200
Consumables (plates, swabs, tips, tubes, cryotubes)	900
Transport/field (coolers, ice packs, travel)	600
Targeted confirmations (PCR/16S outsourced – optional)	700
Small equipment (pipettes/calibration, racks, pH meter)	500
Quality & control (reference strains, control discs)	250
Communication & reporting (printing, farm briefings)	200
Contingency	350
Estimated total	4,700

6. References

FAO. The State of World Fisheries and Aquaculture. 2022.
Cabello FC. Environ Microbiol, 2006.
Watts JEM et al. Front Microbiol, 2017.
Amal MNA, Zamri-Saad M. Aquaculture, 2011.
Austin B, Austin DA. Bacterial Fish Pathogens. Springer, 2016.
Chen L et al. Sci Total Environ, 2020.
Phu TM et al. Rev Aquac, 2016.
Hounmanou YAG et al. Aquaculture Reports, 2021.

5c:T3bf9,

Phase 1 – Local AMR & Virulence Mapping of Major Tilapia Pathogens in Burkina Faso to Support Autovaccine Development

1. Overview

Phase 1 – Local AMR & Virulence Mapping of Major Tilapia Pathogens in Burkina Faso to Support Autovaccine Development

Funding / Facilities / Ethics

Budget requested: 5 000 USD
Facilities: Veterinary Microbiology Lab, Kırıkkale University, Turkey (PhD host); collaborative access to local lab in Burkina Faso for sampling.
Ethics: Non-invasive sampling; farm-level consent; no animal experimentation; BSL-2 compliance.

Timeline
12 months (M1–M12): sampling, isolation/identification, antibiograms, virulence testing, data analysis, dissemination.

2. Introduction

3. Methods

Participants / Sample

Sites: 5-7 tilapia farms in Burkina Faso
Samples: gills, kidney, liver of diseased/suspect fish; pond water; equipment swabs
Exclusion: fish treated with antibiotics <7 days
Target: 60–90 isolates (30–45 per species)

Materials & Procedures

Isolation & ID: Todd-Hewitt, TSA, MacConkey, blood agar; Gram, catalase, oxidase; biochemical galleries (API or low-cost equivalent); optional species-specific PCR if budget allows.
Antibiograms: Kirby-Bauer, EUCAST/CLSI; panel: β-lactams, tetracyclines, fluoroquinolones, sulfonamides/trimethoprim, aminoglycosides, phenicols.
Virulence: haemolysis on blood agar, crystal-violet biofilm (96-well), gelatinase/DNase plates.

Planned Analyses

Resistance rates & MDR index
Site comparisons (Chi²/Fisher)
Biofilm: ANOVA/Tukey
Strain prioritisation: matrix combining prevalence, MDR, and virulence score
Contingency: if isolation rate < expected → increase sites/samples; if no PCR → focus on robust phenotyping

Ethics & Data Management

Farm consent; anonymised data
BSL-2 biosafety; national waste regulations
Open data: CSV + metadata → Zenodo/OSF (CC-BY 4.0)

4. Pilot Data (Optional)

Summarize any pilot data or simulations that demonstrate method feasibility and inform power analysis. Clearly distinguish from main study.

5. Budget

	Category Cost (USD)
Media & reagents (BHI/TSB, blood agar, Gram, antibiotic discs)	1,200
Consumables (plates, swabs, tips, tubes, cryotubes)	900
Transport/field (coolers, ice packs, travel)	600
Targeted confirmations (PCR/16S outsourced – optional)	700
Small equipment (pipettes/calibration, racks, pH meter)	500
Quality & control (reference strains, control discs)	250
Communication & reporting (printing, farm briefings)	200
Contingency	350
Estimated total	4,700

6. References

FAO. The State of World Fisheries and Aquaculture. 2022.
Cabello FC. Environ Microbiol, 2006.
Watts JEM et al. Front Microbiol, 2017.
Amal MNA, Zamri-Saad M. Aquaculture, 2011.
Austin B, Austin DA. Bacterial Fish Pathogens. Springer, 2016.
Chen L et al. Sci Total Environ, 2020.
Phu TM et al. Rev Aquac, 2016.
Hounmanou YAG et al. Aquaculture Reports, 2021.

5d:T16f5,Phase 1 – Local AMR & Virulence Mapping of Major Tilapia Pathogens in Burkina Faso to Support Autovaccine Development 1. Overview Phase 1 – Local AMR & Virulence Mapping of Major Tilapia Pathogens in Burkina Faso to Support Autovaccine Development Scientific Rationale & Importance Antibiotic overuse in tilapia farming drives antimicrobial resistance (AMR), threatening productivity, public health, and food security. Streptococcus agalactiae and Aeromonas hydrophila are major pathogens causing mass mortalities and economic losses. These bacteria often display multidrug resistance (MDR) and biofilm-forming ability, reducing treatment efficacy. This project will generate the first local AMR and virulence dataset from Burkina Faso to support the development of preventive alternatives, including autogenous vaccines and post-biotic adjuvants, aligned with One Health principles. Funding / Facilities / Ethics Budget requested: 5 000 USD Facilities: Veterinary Microbiology Lab, Kırıkkale University, Turkey (PhD host); collaborative access to local lab in Burkina Faso for sampling. Ethics: Non-invasive sampling; farm-level consent; no animal experimentation; BSL-2 compliance. Timeline 12 months (M1–M12): sampling, isolation/identification, antibiograms, virulence testing, data analysis, dissemination. Data Sharing All AMR and virulence data will be deposited in an open-access repository (Zenodo/OSF) within 3 months of completion. Strains will be available for non-commercial research under an MTA. 2. Introduction Theoretical background Antimicrobial resistance (AMR) is a global One-Health emergency. Intensive tilapia culture relies heavily on antibiotics, selecting for multidrug-resistant (MDR) pathogens that spread to humans via food and water. Two taxa drive mortality in African tilapia farms: Streptococcus agalactiae (streptococcosis; ≤80 % mortalities) and Aeromonas hydrophila (ulcerative syndrome, septicaemia). Both form biofilms and carry MDR plasmids, yet no baseline AMR or virulence data exist for Burkina Faso. Key literature Asian and East-African surveys report tetracycline and fluoroquinolone resistance rates >60 % in these species (Chen 2020; Hounmanou 2021). Biofilm-positive isolates show 4–8× higher minimal inhibitory concentrations and persist in pond sediments (Watts 2017). Autogenous vaccines, adjuvanted with low-cost post-biotics, are emerging as sustainable alternatives, but strain selection requires local phenotypic and resistance maps—currently missing for West Africa. Research questions RQ1 Which S. agalactiae and A. hydrophila genotypes dominate Burkinabè tilapia farms? RQ2 What are their AMR profiles against 12 antibiotic classes? RQ3 Do they exhibit high-virulence phenotypes (haemolysis, biofilm, extracellular enzymes) that warrant vaccine inclusion? Numbered hypotheses H1 Local isolates display MDR (≥3 classes), especially to tetracyclines and fluoroquinolones. H2 ≥50 % of isolates are strong biofilm-formers and β-haemolytic. H3 A quantifiable subset combines high prevalence, MDR and virulence, providing optimal autovaccine candidates. Impact statement Regardless of resistance levels, this first national surveillance will: (i) establish an open-access AMR/virulence atlas and strain bank; (ii) guide rational antimicrobial use; (iii) select candidate strains for an immediate PhD-driven autogenous vaccine trial with post-biotic adjuvant; and (iv) create a replicable One-Health surveillance model for West African aquaculture. 3. Methods Participants / Sample Sites: 5-7 tilapia farms in Burkina Faso Samples: gills, kidney, liver of diseased/suspect fish; pond water; equipment swabs Exclusion: fish treated with antibiotics <7 days Target: 60–90 isolates (30–45 per species) Materials & Procedures Isolation & ID: Todd-Hewitt, TSA, MacConkey, blood agar; Gram, catalase, oxidase; biochemical galleries (API or low-cost equivalent); optional species-specific PCR if budget allows. Antibiograms: Kirby-Bauer, EUCAST/CLSI; panel: β-lactams, tetracyclines, fluoroquinolones, sulfonamides/trimethoprim, aminoglycosides, phenicols. Virulence: haemolysis on blood agar, crystal-violet biofilm (96-well), gelatinase/DNase plates. Planned Analyses Resistance rates & MDR index Site comparisons (Chi²/Fisher) Biofilm: ANOVA/Tukey Strain prioritisation: matrix combining prevalence, MDR, and virulence score Contingency: if isolation rate < expected → increase sites/samples; if no PCR → focus on robust phenotyping Ethics & Data Management Farm consent; anonymised data BSL-2 biosafety; national waste regulations Open data: CSV + metadata → Zenodo/OSF (CC-BY 4.0) 4. Pilot Data (Optional) Summarize any pilot data or simulations that demonstrate method feasibility and inform power analysis. Clearly distinguish from main study. 5. Budget Category Cost (USD) Media & reagents (BHI/TSB, blood agar, Gram, antibiotic discs) 1,200 Consumables (plates, swabs, tips, tubes, cryotubes) 900 Transport/field (coolers, ice packs, travel) 600 Targeted confirmations (PCR/16S outsourced – optional) 700 Small equipment (pipettes/calibration, racks, pH meter) 500 Quality & control (reference strains, control discs) 250 Communication & reporting (printing, farm briefings) 200 Contingency 350 Estimated total 4,700 6. References FAO. The State of World Fisheries and Aquaculture. 2022. Cabello FC. Environ Microbiol, 2006. Watts JEM et al. Front Microbiol, 2017. Amal MNA, Zamri-Saad M. Aquaculture, 2011. Austin B, Austin DA. Bacterial Fish Pathogens. Springer, 2016. Chen L et al. Sci Total Environ, 2020. Phu TM et al. Rev Aquac, 2016. Hounmanou YAG et al. 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