Leandro Quintana Nizoli

Cloning and expression of a truncated form of envelope glycoprotein D of Bovine herpesvirus type 5 in methylotrophic yeast Pichia pastoris.

Meningoencephalitis caused by Bovine herpesvirus type 5 (BoHV-5) is responsible for heavy economic losses in the cattle industry. As in other Alphaherpesviruses, the envelope glycoprotein IV (gD), which mediates penetration into host cells, is one of the major candidate antigens for a recombinant vaccine, since it induces a strong and persistent immune response. The DNA coding for a truncated form of BoHV-5 gD (tgD) has been cloned into the Pichia pastoris expression vector pPICZalphaB to allow protein secretion into the medium. After induction with methanol, a approximately 55kDa protein was obtained. Enzyme deglycosylation with Endo H showed a smaller size band in SDS-PGAE, with approximately 50kDa, suggesting that tgD has N-linked oligosaccharides and that it is not hyperglycosylated. The approximately 55kDa protein was recognized by several polyclonal antibodies, including polyclonal antibody anti-tgD and polyclonal antibodies of different animal species immunized with BoHV-5 and BoHV-1. This is the first report of BoHV-5 gD expression in yeast. It was shown that the recombinant truncated form of BoHV-5 gD has antigenic and immunogenic properties similar to the native BoHV-5 gD. Expression of tgD as a secreted protein allows simple and inexpensive purification methods that can be used for further studies to evaluate its immunogenicity in cattle.

Bovine genetic resistance effects on biological traits of Rhipicephalus (Boophilus) microplus.

This study aimed to verify the influence of bovine genetic resistance on biological traits of the Rhipicephalus (Boophilus) microplus tick. Genetic resistance or susceptibility was determined according to breeding values for tick counts, predicted using a dataset of 9007 Hereford and Braford (Hereford×Zebu) bovines naturally infested and raised under extensive production systems in southern Brazil. From a total of 974 Braford heifers born in 2008, 20 were classified as genetically tick-resistant and 20 classified as genetically tick-susceptible, and used to obtain the ticks samples used in this study. The 40 heifers were exposed to four subsequent artificial infestations with approximately 20,000 larvae at 14-day intervals. From the 19th to 23rd day of each infestation tick counts were performed on the left body side of the heifers. Engorged ticks were manually collected on the day of highest observed burden after each infestation. Tick counts on susceptible heifers were 5.5, 10.5, 11.1 and 6.9 times larger than on resistant heifers, respectively, after the first, second, third and fourth artificial infestations. In the third infestation, ticks from resistant heifers showed lower egg production index (P<0.0001) than ticks from susceptible heifers. In the fourth infestation, ticks from susceptible group showed higher egg mass weight (P<0.05) and nutrient index (P<0.0001) than ticks from resistant heifers. Tick initial weights showed a positive association with egg production index in susceptible heifers (P<0.05) and a negative association in the resistant group (P<0.05), suggesting a host defense mechanism that reduces the conversion efficiency of ingested blood to eggs in engorged ticks from resistant cattle. This shows that bovine genetic tick resistance, in addition to affecting the number of ticks carried by the animals, also affected the egg mass weight, egg production and nutrient indexes of ticks. The results of the present study imply that the selection of resistant animals could be used as a strategic tool for tick control in production systems, reducing infestation levels on cattle and environment.

Presence of Gasterophilus (Leach, 1817) (Díptera: Oestridae) in horses in Rio Grande do Sul State, Brazil

ABSTRACT With the purpose of verify the presence of larvae from the genus Gasterophilus in the Rio Grandedo Sul State, a study was undertaken using 395 horses taken to slaughter in the city of Pelotas. Thematerial analysis revealed that 126 animals (31,90%) presented infection, with 100 animals (25,32%)infected by Gasterophilus nasalis , and 47 animals (11,90%) infected by G . intestinalis . These resultsalso represent the first report of the definite establishment of G. intestinalis as a horse parasite inBrazil. The probable implications of G. intestinalis occurrence in Brazil are discussed. Key words: Gasterophilus; G. nasalis; G. intestinalis. * Laboratorio de Doencas Parasitarias da Faculdade de Veterinaria, UFPel, Pelotas, Brasil.** Setor de Parasitologia, Departamento de Microbiologia, Instituto de Ciencias Basicas da Saude, UFRGS, PortoAlegre, Brasil. INTRODUCTIONThe genus Gasterophilus (Diptera, Oestridae)includes eight species of flies whose larvae causegastrointestinal myiasis in equids

Comparison of three larval bioassays to evaluate susceptibility of Rhipicephalus (Boophilus) microplus to amitraz

Field samples of Rhipicephalus (Boophilus) microplus from the state of Rio Grande do Sul, Brazil, were assessed using the following methods: larval packet test (LPT), larval immersion test (LIT) and syringe immersion test (SIT). The following parameters were determined for each population and for the Mozo susceptible reference strain: lethal concentration for 50% (LC50) with its 95% confidence interval (95% CI), regression line slope and resistance ratio (RR). Using the LPT, only one population was susceptible to amitraz, presenting a RR of 1.9. Using the same technique, the other populations presented RRs of between 92.9 and 3445.8 and were considered resistant. The LC50 of the Mozo strain calculated using the LPT, LIT and SIT was 2.9, 27.3, and 52.7 µg/mL, respectively. In general, a good fit to the probit statistical model was only achieved using the LPT. The results obtained in this study impair recommendations for using the LIT and SIT to diagnose amitraz resistance in R. (B.) microplus populations. Additional studies are required to improve the sensitivity of these tests in relation to the LPT.

Frequency of seropositive equines for Theileria equi in the Southern Rio Grande do Sul State, Brazil

ABSTRACT A study on equine theileriosis was carried out in the southern region of the Brazilian state of RioGrande do Sul (RS). Blood samples were collected from 113 mares from an equine breeding farmlocated in the city of Bage, latitude 31o30’ S and a longitude of 54U10’ W. The serological testing wascarried out with the use of indirect fluorescence test (IFAT) and compared with Nested PolimeraseChain Reaction (nPCR). Among the sera collected from 118 horses, 25 were found positive to Theileria equi by the IFAT, while by nPCR 17 positive for T. equi was observed, corresponding to afrequency of 22.1% and 15.0%, respectively. The racial analysis showed 15.05% (14) thoroughbredand 55% (11) Crioulo breed horses to be positive by IFAT. Key words: Theileria equi, IFAT, prevalence. INTRODUCTIONEquine piroplasmosis, caused by Theileriaequi and Babesia caballi , is considered to be themost important tick-borne disease of horse intropical and subtropical areas 1 . Clinicalmanifestation of the disease is variable includingfever, anemia, icterus, lethargy, and in somecases death