Are coinfections a threat to drug control programmes for livestock trypanosomes?

非洲锥虫给撒哈拉以南非洲的畜牧生产造成了巨大的经济损失，加剧了受灾地区的贫困。布鲁氏锥虫、刚果锥虫和间日锥虫这三种致病物种共同传播，感染通过化学预防和药物治疗进行管理。然而，物种之间的共感染是常见的，一些文献报道和我们自己的数据表明，锥虫物种和菌株之间的共感染相对于单一物种或菌株的单感染可以改善疾病病理。在暴露于次优剂量的共感染锥虫之间存在不同药物敏感性的环境中，或在某些流行寄生虫种群中存在耐药性的环境中，这会产生风险。具体而言，通过将共感染恢复为仅包含更具抗性的菌株或物种的单感染，药物干预可能导致病理增强。因此，药物应用可能产生增加疾病的反常结果。在本研究中，我们将探讨小鼠模型和疾病相关家畜宿主的共感染、药物敏感性和病理之间的相互作用。具体来说，我们将：1。利用已知的分子耐药/敏感机制（家畜锥虫最常用的治疗方法）对布氏锥虫和刚果锥虫分别进行耐药和敏感工程。具体地说，在布氏体中，核苷转运体TbAT1基因的抗性决定因素将被删除，从而产生耐药的寄生虫，刚果白锥虫将被改造成异源表达布氏体AT1基因，从而产生一个布氏体的超敏感系。这将使我们的研究能够精确地去除实验中与布氏体共感染的刚果螺旋体，避免了不同野生型实验室菌株和野外菌株可能表现出的可变萘敏感性的复杂性。该工程细胞系将用于评估共感染，或共感染后再由咪纳苯诱导的单感染对小鼠中锥虫种群的比例和分布及其病理的影响。这将通过IVIS成像对每个物种和布鲁氏体宿主的寄生虫数量进行定量来实现。小鼠病理将按照既定标准进行评分。利用罗斯林研究所独特和专用的大型动物围护设施，这些工程品系将用于测试共同感染，或共同感染后再由苯甲肼引起的单感染，如何影响与疾病相关的牛宿主的寄生虫流行率和病理学。这些研究将确定当合并感染通过治疗干预重定向为单一感染时对寄生虫流行率和宿主病理的影响。在混合感染情况下存在差异耐药性的情况下，这可能对牲畜健康造成意想不到的危害。我们的实验可以优先考虑这一以前被忽视的威胁的流行病学研究，并促进优化剂量或治疗患病动物和维持治疗效果的策略。这也将加速有针对性地采用可获得的替代锥虫剂。撒哈拉以南非洲的农民和政策制定者没有预料到治疗干预在合并感染环境中可能产生的不利影响，可能被忽视或视为轶事而不予理会。我们将确保通过我们计划的以牲畜锥虫为重点的外联活动和合作，将我们的发现传播给科学界、决策者和农民。这些包括计划中的会议，例如Morrison将于2023年在坦桑尼亚组织的会议，以及与比尔和梅林达·盖茨基金会的合作工作，与肯尼亚的国际牲畜研究所的联系，以及正在非洲进行的实地工作。

African trypanosomes cause substantial economic cost to livestock production in sub-Saharan Africa exacerbating poverty in afflicted regions. Three pathogenic species co-circulate, Trypanosoma brucei, Trypanosoma congolense and Trypanosoma vivax, with infections being managed by chemoprophylaxis and drug therapy. However, coinfections between the species are common and several literature reports and our own data indicate that coinfection between trypanosome species and strains can ameliorate disease pathology with respect to monoinfections with a single species or strain. This generates a risk in settings where there is differential drug sensitivity among coinfecting trypanosomes exposed to suboptimal dosing, or where drug resistance is present in some circulating parasite populations. Specifically, by reverting coinfections to monoinfections comprising only the more resistant strain or species, drug intervention may lead to enhanced pathology. Consequently, drug application may generate a perverse outcome of increased disease.In this proposal we will explore the interaction between coinfection, drug sensitivity and pathology in both a mouse model and disease-relevant livestock host. Specifically, we will:1. Engineer drug resistant and sensitive Trypanosoma brucei and Trypanosoma congolense, respectively, using a known molecular resistance/sensitivity mechanism for diminazene, the most commonly used therapy for livestock trypanosomes. Specifically, the diminazene resistance determinant TbAT1, a nucleoside transporter, will be deleted in T. brucei to generate resistant parasites and T. congolense will be engineered to heterologously express the T. brucei AT1 gene, generating a diminazene super-sensitive line. This will allow the precision removal of T. congolense in experimental coinfections with T. brucei for our studies, avoiding the complexity of variable diminazene sensitivities different wild type laboratory and field strains may exhibit.2. The engineered lines will be used to evaluate the impact of coinfection, or coinfection followed by diminazene induced monoinfections, on the proportion and distribution of the trypanosome populations and their pathology in mice. This will be achieved by quantitating parasite numbers for each species and reservoirs of T. brucei by IVIS imaging. Murine pathology will be scored by established criteria.3. The engineered lines will be used to test how coinfection, or coinfection followed by diminazene-induced monoinfections, affects parasite prevalence and pathology in the disease-relevant bovine host using unique and dedicated large animal containment facilities at Roslin Institute.The studies will determine the consequences for parasite prevalence and host pathology when a coinfection is redirected to a monoinfection through therapeutic intervention. This could be unexpectedly harmful for livestock health where differential resistance exists in mixed infection scenarios. Our experiments could prioritise epidemiological studies of this previously overlooked threat, and promote strategies to optimise dosing, or to treat diseased animals and sustain therapeutic efficacy. This would also accelerate the targeted adoption of alternative trypanocides as they become available.The potential for adverse impact of therapeutic intervention in coinfection settings is unanticipated among farmers and policymakers in sub-Saharan Africa and may have been overlooked or dismissed as anecdote. We will ensure our findings are disseminated to the scientific community, policymakers and famers through our planned outreach activities and collaborations focused on livestock trypanosomes. These include planned meetings, for example a meeting Morrison is organising in Tanzania in 2023, and collaborative work with the Bill and Melinda Gates Foundation, links with the International livestock research Institute in Kenya and ongoing field work in Africa.