Exploring the life cycle of Besnoitia besnoiti - Experimental infection of putative definitive and intermediate host species

The biology of Besnoitia besnoiti, the cause of bovine besnoitiosis, is poorly understood. Its definitive host is unknown, and information on potential intermediate hosts is scarce. In order to investigate potential definitive and intermediate hosts for European isolates of B. besnoiti, domestic dogs, cats, rabbits, guinea pigs (Cavia porcellus), gerbils (Meriones unguiculatus), common voles (Microtus arvalis) and NMRI-mice were inoculated with B. besnoiti isolated from naturally infected German cattle. Dogs and cats were fed 5 x 106 B. besnoiti tachyzoites (isolate Bb-GER1), or tissue cysts containing at least 2 x 107 B. besnoiti bradyzoites obtained from the skin of a naturally infected Limousin cow from the same herd where strain Bb-GER1 was isolated. Rodents and rabbits were subcutaneously inoculated with either 5 x 105 Bb-GER1 tachyzoites or 5 x 105 bradyzoites. Groups of 2-4 non-inoculated animals of each species were monitored as negative controls. Feces from all dogs and cats were daily examined by a sedimentation-flotation technique for at least 11 weeks after inoculation but no B. besnoiti oocysts were identified. Cats fed tachyzoites and dogs did not seroconvert, but specific antibodies to B. besnoiti tachyzoites were detected by IFAT (titer = 100) in 2 out of 3 cats fed tissue cysts since 5-7 weeks post infection. By immunoblot, these two cats exhibited a reaction pattern against tachyzoite antigens similar to that observed in naturally infected cattle. Antibodies against B. besnoiti tachyzoites were detected in all inoculated rodent species and rabbits by both, IFAT and immunoblot since 3 weeks post-inoculation. Rabbits and rodents, subcutaneously inoculated with same doses of inactivated bradyzoites remained serologically negative (IFAT titer < 50). Clinical signs observed in the inoculated rabbits included fever, serous conjunctivitis and transient swelling of the testes. No clinical abnormalities were noticed in the other tested animal species. Voles developed pneumonia as observed by histological examination. B. besnoiti-DNA was detected by PCR in blood from rabbits, gerbils and voles at 9 days post-infection, and in skin, heart, lung, striated muscle and kidney tissues from voles at 19-21 weeks post-infection. Domestic dogs and cats could not be shown to be definitive hosts of B. besnoiti, but cats seroconverted after feeding on B. besnoiti tissue cysts indicating that B. besnoiti stages had invaded the cats' tissues. The molecular and serological results from this study indicate that European B. besnoiti isolates may infect cats, rabbits, guinea pigs, gerbils, mice and voles; however a persistence of the parasite could be demonstrated only in voles.