Haemoprotozoa surveillance in peri-urban native and introduced wildlife

Vector-borne haemoprotozoa comprise a diverse group of eukaryote single-celled organisms transmitted by haematophagous (blood-feeding) invertebrates. They can cause debilitating diseases that impact wildlife, livestock, companion animals and humans. Recent research has shown that Australian wildlife host a diverse range of haemoprotozoa, however, to date this work has primarily been confined to a few host species or isolated populations in rural habitats. There has been little investigation into the presence of these blood parasites in wildlife inhabiting urban and peri-urban areas. In the study, blood and tissue samples and ticks were collected from wildlife in New South Wales and Western Australia. Extracted DNA samples were screened with pan- specific molecular assays to determine the presence of haemoprotozoa using amplicon metabarcoding and Sanger sequencing approaches. In addition, light microscopy was performed on blood films. Eight haemoprotozoa were identified in the present study, which included Babesia, Hepatozoon, Theileria and Trypanosoma species. Blood samples were collected from 134 animals; 70 black rats (Rattus rattus), 18 common brush-tailed possums (Trichosurus vulpecula), two bush rats (Rattus fuscipes), 22 chuditch (Dasyurus geoffroii), 20 long-nosed bandicoots (Perameles nasuta), one quenda (Isoodon fusciventer) and one swamp rat (Rattus lutreolus). Molecular screening of DNA extracted from blood samples identified 52.2% (95% CIs: 43.8–60.5%) of individuals were positive for at least one haemoprotozoa, with 19.4% (95% CIs: 13.4–26.7%) positive for more than one species. The present study describes the first sequences of Theileria cf. peramelis from black rats and long-nosed bandicoots. Babesia lohae was identified from brush-tailed possums. Two Hepatozoon genotypes were identified from black rats and bush rats. Black rats had the highest diversity of haemoprotozoa, with five species identified. No known human pathogens that have been described in the northern hemisphere were identified in the present study, and future work is required to understand the zoonotic potential of these microbes. This work represents the first large-scale body of research using molecular tools to investigate haemoprotozoa in animals at the urban-wildland interface. Further research is needed to investigate potential consequences of infection in wildlife, particularly effects of pathogen spillover from invasive black rats to native wildlife.

虫媒血生原生动物（Vector-borne haemoprotozoa）是一类多样的真核单细胞生物，通过吸血无脊椎动物（haematophagous invertebrates）传播。它们可引发衰弱性疾病，对野生动物、家畜、伴侣动物及人类健康造成负面影响。近期研究显示，澳大利亚野生动物宿主携带种类丰富的血生原生动物，但截至目前，此类研究主要局限于少数宿主物种或乡村生境中的孤立种群，针对城市及城郊生境野生动物体内此类血液寄生虫的调查仍十分匮乏。
本研究从新南威尔士州与西澳大利亚州的野生动物体内采集血液、组织样本及蜱虫。通过扩增子元条形码（amplicon metabarcoding）与桑格测序（Sanger sequencing）技术，利用泛特异性分子检测方法对提取的DNA样本进行筛查，以确定血生原生动物的存在情况；此外还对血涂片开展了光学显微镜检查。
本研究共鉴定出8种血生原生动物，涵盖巴贝斯虫属（Babesia）、肝簇虫属（Hepatozoon）、泰勒虫属（Theileria）及锥虫属（Trypanosoma）的多个物种。本次共采集134只动物的血液样本，其中包括70只黑家鼠（Rattus rattus）、18只普通刷尾负鼠（Trichosurus vulpecula）、2只澳洲林鼠（Rattus fuscipes）、22只西澳袋鼬（Dasyurus geoffroii）、20只长鼻袋狸（Perameles nasuta）、1只昆达袋狸（Isoodon fusciventer）及1只沼泽鼠（Rattus lutreolus）。
对血液样本提取的DNA进行分子筛查后发现，52.2%（95%置信区间：43.8–60.5%）的个体至少携带一种血生原生动物，其中19.4%（95%置信区间：13.4–26.7%）的个体携带多种血生原生动物物种。本研究首次报道了来自黑家鼠与长鼻袋狸的Peramelis泰勒虫近似种（Theileria cf. peramelis）序列；从刷尾负鼠体内鉴定出洛哈巴贝斯虫（Babesia lohae）；从黑家鼠与澳洲林鼠体内鉴定出2种肝簇虫基因型。黑家鼠体内的血生原生动物多样性最高，共鉴定出5个物种。本研究未检出北半球已报道的已知人类病原体，后续仍需开展研究以明确这些微生物的人畜共患潜力。
本研究是首次利用分子工具对城市-荒野交界带野生动物体内的血生原生动物开展的大规模系统性研究。未来仍需进一步探究野生动物感染此类寄生虫的潜在后果，尤其是入侵性黑家鼠向本土野生动物发生病原体溢出的相关影响。