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Studies of the survival, recovery and migration of cyathostomin infective larvae in a Bermuda grass (Cynodon dactylon) pasture were carried out in the Baixada Fluminense county, Rio de Janeiro state. Fresh feces (± 1 kg) from naturally infected horses were deposited monthly on Bermuda grass. Samples of feces and surrounding grass were collected every seven days, from March 2005 to March 2007, and larva were counted. In the feces, cyathostomin L 3 survived for up to 15 weeks, with higher recovery rates during the rainy period (46 228/kg dried herbage -dh), and on the grass for up to 12 weeks. The recovery of L 3 was greater during the dry period in the grass base (1 868/kg dh) compared to the apex (809/kg dh). The migration of L 3 from feces to grass varied during the period. Climatic factors, such as temperature and rain, influenced the development and migratory behavior of cyathostomin L 3 . With regard to the grass base, significant differences were observed at the different collection times. The results demonstrate that under local conditions animals are at permanent risk since the infective larvae are always present on pasture.

Sources of contamination such as animal feces runoff, organic fertilizer application, and the release of partially treated or untreated sewage can lead to the contamination of aquatic environments by Cryptosporidium spp. The quality of mussels as food is closely related to the sanitary conditions of the marine environment where these bivalves are found. Marine mollusks are filter feeders that are able to retain Cryptosporidium oocysts in their tissue, thus functioning as bioindicators. A total of 72 pooled mussel samples of the species Perna perna were collected at two sites (A and B) in the municipality of Mangaratiba, Rio de Janeiro State, Brazil. Sampling involved removal of 30 mussels, from each collection site every month for one year. The 30 mussels from each sampling were then allocated into three groups of 10. Two Cryptosporidium spp. genes (18S and GP60) were targeted for DNA amplification from the samples obtained. After purification, all of the products obtained were sequenced and phylogenetic analyses were performed. Of the 72 samples analyzed using the nested-PCR for the 18S gene target, 29.2% were positive for the presence of Cryptosporidium spp. Of these samples, 52.4% were collected at site A (ie 11/21) and 47.6% at site B (ie 10/21). The 18S genes of all the samples considered positive for Cryptosporidium spp. were sequenced, and the following three species were identified: Cryptosporidium parvum, C. meleagridis, and C. andersoni. Three distinct C. parvum subtypes (IIaA19G2R2; IIaA20G2R2; IIaA20G3R2) were identified using the GP60 gene. More studies to evaluate the zoonotic potential of this species should be performed as both sampling locations contain human and/or animal fecal contaminants.

Differentiating between the Cryptosporidium species and their subtypes using only microscopy is impossible. Therefore, molecular tools are indispensable for accurate species and subtype diagnosis. However, if these tools are to be used correctly and accurately, the techniques used must be standardised. In the present study, two molecular techniques for diagnosing Cryptosporidium infection in cows were compared to determine the optimal methods. For each technique, we tested two DNA extraction methods, several annealing temperatures for nested PCR reactions targeting the 18S, SSU rRNA (small subunit ribosomal RNA), and the GP60 (60 kDa glycoprotein) genes, and two types of DNA staining reagents, ethidium bromide and GelRed TM . We determined that one of the tested protocols yields a higher purity of extracted DNA. Additionally, optimised temperatures for the nested PCR of the 18S and GP60 genes were established. Finally, we determined that the GelRed TM dye was more sensitive than ethidium bromide, and its low toxicity facilitates handling and disposal and reduces environmental contamination.

CYATHOSTOMINAE (STRONGYLIDAE-CYATHOSTOMINAE) HORSE PARASITES: EXPERIMENTAL ECOLOGY OF FREE LIVING STAGES ON PASTURE

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The aim of this study was to establish the occurrence of Giardia intestinalis in goat fecal samples from two dairy farms (A and B) located in Rio de Janeiro state, Brazil, assessing possible risk factors for infection. Fecal samples from all young animals (up to one year of age) (n=58) and from 10% of adult animals (n=18) were collected. Samples were collected directly from the rectum and were submitted to centrifuge-flotation in sugar saturate solution for microscopic search of Giardia cysts. From 76 analyzed samples, 17 (22.4%) were positive for Giardia cysts. All positive samples came from young animals of farm A. Also, dog fecal samples (n=9) were collected to evaluate their role in parasite transmission. Three dog samples (33.3%) were positive for Giardia cysts. Presence of helminth eggs was only observed in animals from farm A. Eimeria sp. oocysts were observed in fecal samples from both farms. The comparison of hygienic-sanitary and management conditions of both farms indicated that the high rate of infection in young animals from farm A was due to an association of risk factors: intensive raising of young animals in installations made of wooden slats floor, fecal accumulation underneath it, and the presence of domestic animals inside goat installations. The high Giardia positivity and the observation of risk factors for transmission call attention to the importance of effective prophylactic measures and patterns of management.