Infection Dynamics of the Lungworm Rhabdias Pseudosphaerocephala in Its Natural Host, the Cane Toad (Bufo Marinus), and in Novel Hosts (Native Australian Frogs)
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Journal of Wildlife Diseases, 46(4), 2010, pp. 1152–1164 # Wildlife Disease Association 2010 INFECTION DYNAMICS OF THE LUNGWORM RHABDIAS PSEUDOSPHAEROCEPHALA IN ITS NATURAL HOST, THE CANE TOAD (BUFO MARINUS), AND IN NOVEL HOSTS (NATIVE AUSTRALIAN FROGS) Ligia Pizzatto,1,3 Catherine M. Shilton,2 and Richard Shine1 1 School of Biological Sciences A08, University of Sydney, NSW 2006, Australia 2 Berrimah Veterinary Laboratories, Northern Territory Government, Berrimah, NT 0801, Australia 3 Corresponding author (email: [email protected]) Downloaded from http://meridian.allenpress.com/jwd/article-pdf/46/4/1152/2239199/0090-3558-46_4_1152.pdf by guest on 01 October 2021 ABSTRACT: Host-parasite systems have often evolved over time, such that infection dynamics may become greatly modified from the time of initial contact of the host with the parasite. Biological invasions may be useful to clarify processes in the initial contact of hosts with parasites, and allow us to compare parasite uptake between the ancestral (coevolved) host and novel (noncoevolved) hosts. Cane toads (Bufo marinus) are spreading rapidly through tropical Australia, carrying with them a nematode lungworm (Rhabdias pseudosphaerocephala) congeneric with those found in Australian frogs. We investigated the dynamics of infections of the toad parasite by conducting histologic examinations of cane toads and three native Australian frogs (Litoria dahlii, Litoria nasuta, and Opisthodon ornatus) at 2, 6, and 10 days after experimental exposure to the toad lungworm. More worms were found in toads than in frogs, especially at longer periods postexposure. In toads, the infective larvae entered the skin and muscles within 2 days postexposure, passed into the coelom in 6 days, and reached the lungs at 10 days. In frogs, larvae were found in many organs rather than migrating to consistent target tissues; a few larvae reached the lungs of L. dahlii. Migratory larvae caused increasing inflammation (primarily granulomatous admixed with granulocytes then lymphocytes) through time, especially in frogs. Evolution has resulted in an enhanced ability of the lungworm to locate the target organ (the lungs) of the toad, and an increase in rates of parasite survival within this host. Key words: Anura, host switch, immunologic response, inflammation, migratory pathway, nematode, parasite. INTRODUCTION environment along with their hosts. For example, the microsporidian parasite Fi- Invasive species can exert an impact on brillanosema crangonycis was introduced native taxa via many pathways, including to the UK with its host, the amphipod predation, competition, and habitat mod- Crangonyx pseudogracilis (Galbreath et ification (Mack et al., 2000; Sakai et al., al., 2004). Such parasites may be useful for 2001). One of the most important but controlling their host (alien) populations, poorly understood processes may be the but if the parasites are not host specific, introduction of novel pathogens into they can also pose a threat to the native highly susceptible populations (Prenter et fauna by switching to other hosts (Prenter al., 2004), a process responsible for the et al., 2004; Taraschewski, 2006). For immense impact of European colonization example, a pathogenic protozoan (the on the indigenous human population of rosette agent) was introduced with an many Pacific (Willcox, 1980) and South AsiancyprinidfishtoEuropeandis American (Willcox, 1980; Hopkins, 1983; threatening an endangered native fish Rothschild, 2005) countries. Although (Gozlan et al., 2005). The cattle and sheep accidents of the introduction process often liver fluke is native to the Palearctic region result in invasive species arriving in their (see Taraschewski, 2006) but has been new range without many of their native- introduced to North, Central, and South range parasites (Mitchell and Power, 2003; America, Africa, Australia, and New Zea- Torchin et al., 2003), there are many cases land, where it has switched hosts to native of parasites being introduced to a novel marsupials (Spratt and Presidente, 1981), 1152 PIZZATTO ET AL.—NEMATODE INFECTION DYNAMICS IN NATURAL AND NOVEL HOSTS 1153 rodents (Fuentes et al., 1997), and even the lungs during larval migration? More humans (Esteban et al., 1999). Its patho- generally, how do infection dynamics of genicity in some new hosts can be as high the parasite in a coevolved (natural) host as in the original hosts (Viggers and Spratt, (the cane toad) differ from those in novel 1995). hosts (Australian frogs)? To answer these When the cane toad (Bufo marinus,or questions we looked at a system where Rhinella marina in an alternative nomen- toads have arrived within the last few clatural scheme; Pramuk, 2006) was intro- years, leaving little opportunity for coevo- duced to Australia in 1935, it brought with lution. it the rhabditoid lungworm Rhabdias Downloaded from http://meridian.allenpress.com/jwd/article-pdf/46/4/1152/2239199/0090-3558-46_4_1152.pdf by guest on 01 October 2021 pseudosphaerocephala (Dubey and Shine, MATERIALS AND METHODS 2008). These parasites are found in cane We used three species of abundant non- toads across most of the toads’ Australian arboreal frogs: Litoria dahlii, Litoria nasuta range (Barton, 1998), but have not been (family Hylidae), and Opisthodon ornatus reported in Australian frogs, which are (family Limnodynastidae), as well as the cane infected with other Rhabdias species toad, B. marinus (family Bufonidae). The (Dubey and Shine, 2008). Rhabdias pseu- anurans were collected as eggs in ponds near Fogg Dam (12.6uS, 131.3uE) about 60 km east dosphaerocephala have a direct life cycle, of Darwin, in the wet-dry tropics of the similar to other species in the genus. The Northern Territory, Australia, where the first larvae (L3) infect the host via cutaneous cane toads arrived in early 2005. Tadpoles penetration and migrate through the body were raised outdoors in plastic containers to the lungs, where they develop into (1103110364 cm; each species in a different container) containing 650 l of nonchlorinated hermaphroditic adults. The adult nema- water, fitted with screened lids. Tadpoles were todes lay eggs, which are released into the fed on frozen lettuce, fish flakes (TetraminH lungs and pass through the intestines. In Tropical Flakes, Blacksburg, Virginia, USA) and pellets for algae eaters (HBH Veggie the intestines, the embryonated eggs hatch TM into (L1) larvae (Baker, 1979), which are Wafers , Springville, Utah, USA). After completing metamorphosis, nine individuals usually first seen in the toad metamorph’s of each species were placed in a single feces after 30 days of infection (C. container (as above) that was placed over a Kelehear, unpubl. data [University of plastic tray containing iodine scrub (BetadineH Sydney]; Pizzatto, pers. obs.). After being Surgical Scrub; 7.5% weight/volume povi- defecated, L1 larvae develop into L3 done-iodine) to avoid contamination and kept indoors under ultraviolet light (natural light/ larvae or free-living dioecious adults that dark cycles), at 28.262.68 C. Each container produce L3 larvae (Baker, 1979). In had a substrate of preboiled sand and tree bark anurans, migrating rhabditoid larvae can (for refuge) and a pool of nonchlorinated induce granulomatous inflammation, and water. They were fed ad libitum on termites adult worms in the lung can cause and small cockroaches every third day throughout the experiment. In other work, inflammation and scarring (Williams, we have found that this maintenance environ- 1960; Green, 2001; Ladds, 2009). ment for the eggs, tadpoles, and metamorphs Infection with R. pseudosphaerocephala enables us to eliminate any accidental contact can reduce the fitness (survival, locomotor of our study animals with Rhabdias sp. performance, food intake, and growth (Pizzatto, unpubl. data). On 30 May 2009, cultured infective larvae of rates) of metamorphs of its natural host, R. pseudosphaerocephala (about 3,000) were the cane toad (Kelehear et al., 2009), but added to the container. These larvae originat- their interactions with native frogs remain ed from the eggs of three–four adult lung- unclear. Does the lack of records of R. worms from local cane toads, smeared in a pseudosphaerocephala in Australian frogs small amount of toad feces (for details on culture preparation see Kelehear et al., 2009). (Dubey and Shine, 2008) reflect a lack of Because metamorphs were obtained as eggs infection, or an inability of the lungworms and reared in uncontaminated conditions, any to survive in nontoad hosts, or to locate nematodes infecting the study animals were 1154 JOURNAL OF WILDLIFE DISEASES, VOL. 46, NO. 4, OCTOBER 2010 assumed to be R. pseudosphaerocephala (con- proximal humerus or femur distally), including firmed by DNA sequence analysis in compan- feet, were rarely visualized in section (total of six ion studies using the same methods of distal front leg sections, and we were unable to infection; L. Pizzatto, unpubl. data). The identify front or hind leg section in six frogs). duration of the experiment (10 days) was not Distal limb portions were generally lost in long enough to allow the larvae to develop into processing because they were very small adults in the lungs of metamorphs (minimum segments of tissue that were isolated after being is 30 days: Pizzatto, pers. obs.). Thus, the severed from the rest of the body during original infective dose was the only contact transverse sectioning for histology. Because of between hosts and parasites. Larvae usually this paucity of distal limb sections, and the lack survive for up to 7 days under the conditions in of larvae in any of these sections, data on distal the containers (Pizzatto, pers. obs.); therefore limbs were not included in the analysis. Downloaded from http://meridian.allenpress.com/jwd/article-pdf/46/4/1152/2239199/0090-3558-46_4_1152.pdf by guest on 01 October 2021 hosts were potentially exposed to infective For subadults free in the lumen of a lung, the larvae during the entire period. Because all total number of sections containing lungworms anurans were housed together, all individuals was used to quantify the number of individuals had the same exposure to parasites.