First record of Bulimulus tenuissimus (Mollusca) as potential experimental intermediate host of Angiostrongylus cantonensis (Nematoda)

Abstract Snails are essential to complete the life cycle of the metastrongylid nematode Angiostrongylus cantonensis, the causative agent of infections in domestic and wild animals, mainly rodents, and also of neural angiostrongyliasis or eosinophilic meningitis in humans. There are many reports of mollusks that can act as intermediate hosts of this parasite, especially freshwater snails and the African giant Achatina fulica. The terrestrial gastropod Bulimulus tenuissimus is widely distributed in Brazil and other species of the same genus occur in Brazil and other countries, overlapping regions in which there are reports of the occurrence of A. cantonensis and angiostrongyliasis. In spite of this, there are no records in the literature of this species performing the role of intermediate host to A. cantonensis. The present study analyzed the experimental infection with first-stage larvae of A. cantonensis, under laboratory conditions, of B. tenuissimus, by using histology and electron microscopy techniques. Three weeks after exposure to L1 larvae, it was possible to recover L3 larvae in small numbers from the infected snails. Developing larvae were observed in the cephalopedal mass (foot), ovotestis, and mantle tissues, being located inside a granulomatous structure composed of hemocyte infiltration, but there was no calcium or collagen deposition in these structures in significant amounts. In the third week post exposure, it was possible observe a sheath around the developing larvae. The infected snails presented reduction in the fibrous muscular tissue in the foot region, loss of the acinar organization in the digestive gland, with increase of amorphous material inside the acini and loss of epithelial pattern of nuclear organization in the acinar cells. However, the ovotestis seemed unaffected by the infection, since there was a large number of developing oocytes and spermatozoa in different stages of formation. The digestion of infected snails allows us the third-stage recovery rate of 17.25%, at 14 days post exposure to the L1. These L3 recovered from B. tenuissimus were used to infect rats experimentally, and 43 days post infection first-stage (L1) larvae of A. cantonensis were recovered from fresh feces. The results presented constituted the first report of the role of B. tenuissimus as an experimental intermediate host to A. cantonensis and shed some light on a possible problem, since the overlapping distribution of B. tenuissimus and A. cantonensis in Brazil and other countries where different species of Bulimulus occur enables the establishment and maintenance of the life cycle of this parasite in nature, with wild rodents as reservoirs, acting as a source of infection to humans, causing neural angiostrongyliasis.

摘要 蜗牛是完成后圆线虫科（Metastrongylidae）线虫广州管圆线虫（Angiostrongylus cantonensis）生活史的必需宿主，该寄生虫可引发家养与野生动物（主要为啮齿类）感染，同时也是人类神经型广州管圆线虫病或嗜酸性粒细胞性脑膜炎的致病菌。目前已有诸多关于软体动物可作为该寄生虫中间宿主的报道，尤以淡水螺类与非洲大蜗牛（Achatina fulica）最为常见。陆生腹足类细芽蜗牛（Bulimulus tenuissimus）在巴西分布广泛，该属其他物种亦见于巴西及其他国家，其分布区域与广州管圆线虫及广州管圆线虫病的报道区域存在重叠。尽管如此，目前尚无文献记录该物种可充当广州管圆线虫的中间宿主。本研究在实验室条件下，采用组织学与电子显微镜技术，对细芽蜗牛（Bulimulus tenuissimus）进行广州管圆线虫一期幼虫（L1）的实验感染分析。暴露于L1幼虫三周后，可从感染螺体内少量分离出三期幼虫（L3）。发育中的幼虫见于头足部组织、两性腺及外套膜中，定位于由血细胞浸润形成的肉芽肿结构内，但此类结构中并无大量钙或胶原沉积。感染后第三周，可观察到发育幼虫周围形成鞘膜。感染螺的足部纤维肌肉组织出现萎缩，消化腺的腺泡结构消失，腺泡腔内无定形物质增多，腺泡细胞的细胞核上皮样排布模式丧失。但两性腺似乎未受感染影响，可见大量处于不同发育阶段的卵母细胞与精子。在暴露于L1幼虫后14天，通过消化感染螺的方式，可获得17.25%的L3回收率。从细芽蜗牛中分离得到的L3被用于实验感染大鼠，感染后43天可从新鲜粪便中分离出广州管圆线虫的一期幼虫（L1）。本研究结果首次证实细芽蜗牛（Bulimulus tenuissimus）可作为广州管圆线虫的实验性中间宿主，该发现也揭示了一个潜在的公共卫生问题：细芽蜗牛与广州管圆线虫在巴西及其他分布有Bulimulus属物种的国家存在分布重叠，这使得该寄生虫能够以野生啮齿类为储存宿主，在自然环境中建立并维持其生活史，进而成为人类感染源，引发神经型广州管圆线虫病。