Huarrisson Azevedo Santos

Detection of Anaplasma phagocytophilum in Brazilian dogs by real-time polymerase chain reaction.

Anaplasma phagocytophilum was detected in dogs from Brazil in the municipalities of Seropédica and Itaguaí, Rio de Janeiro state, by real-time polymerase chain reaction (PCR) using SYBR Green to detect the amplification. Of 253 samples, 18 (7.11%) were positive, with a threshold cycle (Ct) ranging from 31 to 35 cycles. The PCR product from a positive sample was cloned and sequenced. The sequence obtained demonstrated 100% identity with other A. phagocytophilum sequences published in the GenBank database. The analytical sensitivity of RT-PCR using SYBR Green system was able to detect 3 plasmid copies when defined numbers of plasmid copies containing 122 base pairs from the msp2 gene were used. The assay was considered specific when DNA from bacteria (Anaplasma platys, Anaplasma marginale, Ehrlichia canis, Neorickettsia risticii, Rickettsia rickettsii) closely related to A. phagocytophilum was placed in the reaction. These results demonstrate that the canine granulocytic anaplasmosis agent is present in regions in which dogs could be a source of infection for tick vectors. The current study reports the detection of A. phagocytophilum, a zoonotic agent responsible for Human granulocytic anaplasmosis, in Brazilian dogs.

Occurrence of anti-Neospora caninum and anti-Toxoplasma gondii IgG antibodies in goats and sheep in western Maranhão, Brazil

Neosporosis and toxoplasmosis are parasitic diseases which can cause reproductive problems in goats and sheep. The current study aimed to determine the occurrence of anti-Neospora caninum and anti-Toxoplasma gondii IgG antibodies in goats and sheep from the districts of Amarante do Maranhão and Buritirana, Imperatriz microregion, western area of Maranhão State, northeastern Brazil, and to assess factors associated to infection by these etiologic agents. Blood samples from 110 animals (46 goats and 64 sheep) from five herds were collected, and indirect immunofluorescence assay was used for serological testing. Of 46 goat samples, 17.39% (n = 8) showed anti-N. caninum antibodies and 4.35% (n = 2) anti-T. gondii, while of 64 sheep samples 4.69% (n = 3) and 18.75% (n = 12) showed anti-N. caninum and anti-T. gondii antibodies, respectively. No significant difference regarding the presence of domestic cats and/or dogs on the property and veterinary care was seen for both etiologic agents studied. However, food supplementation and animal reproductive failure were significantly (p < 0.05) for N. caninum among sheep and goats, respectively. The current study showed that goats and sheep in western Maranhão are exposed to N. caninum and T. gondii. It is the first evidence of these agents in small ruminants in this region.

Factors associated to Theileria equi in equids of two microregions from Rio de Janeiro, Brazil

Serum samples from 714 equids of Itaguaí and Serrana microregions, Rio de Janeiro, southeastern Brazil, were examined by indirect fluorescent antibody test (titer 1:80) for Theileria equi. The prevalence in the microregions and factors associated with seropositivity were evaluated and the prevalence ratio (PR) calculated. The overall prevalence of T. equi infection was 81.09% (n = 579), with higher prevalence (p < 0.05) in the Itaguaí (85.43%) when compared to Serrana microregion (76.92%). The geographic area, altitude, farming condition and area of origin of equids were associated (p < 0.05) with seropositivity for T. equi. Equids reared in the Itaguaí microregion (PR = 1.11, p = 0.003) and at altitudes below 500 m (PR = 1.10; p = 0,014) were more likely to be seropositive for T. equi. Furthermore, when equids were born in the farm (PR = 1.10, p = 0.008) and reared with poor farming conditions (PR = 1.13, p = 0.018) they were more likely to be exposed to T. equi. The main ticks found on equids were Amblyomma cajennense and Dermacentor (Anocentor) nitens. The microregions studied are endemic areas for equine theileriosis and there exists enzootic stability for T. equi. Only factors related to the collection area of serum samples influenced the seropositivity of equids for T. equi in that region.

Avian Spirochetosis in Chickens Following Experimental Transmission of Borrelia anserina by Argas (Persicargas) miniatus

Abstract This study reports the experimental transmission of Borrelia anserina to domestic chickens by infected Argas (Persicargas) miniatus. Clinical alterations as well as prepatent and patent periods were evaluated. Twenty-seven 67-day-old birds were divided into three groups in a randomized experimental design. The first group was exposed to ticks infected with B. anserina, the second group was exposed to noninfected ticks, and the third group was not exposed to ticks. Blood smears from each bird of groups 1 and 2 were prepared daily and examined for 25 days postexposure (PE). Examination of the blood smears from birds in group 1 revealed large numbers of spirochetes from days 5 to 12 PE. In this group the prepatent and patent periods were 5–7 and 4–7 days, respectively. Birds from group 1 presented ruffled feathers, pale combs, drowsiness, inappetence, loss of weight, and greenish diarrhea after day 6 PE. The current study confirms the viability of experimental transmission of B. anserina to domestic chickens by A. (P.) miniatus.

Seroepidemiological aspects of Leishmania spp. in dogs in the Itaguai micro-region, Rio de Janeiro, Brazil.

This study evaluated factors associated with the frequency of Leishmania spp. antibodies in dogs residing in the Itaguai micro-region, State of Rio de Janeiro, Brazil. Blood samples were collected from 524 dogs. The serum samples were submitted to indirect fluorescent antibody test (IFAT) and enzyme-linked immunosorbent assay (ELISA) for Leishmania spp. The frequency of seropositive dogs was 28.24% (n = 148) in the micro-region, and among the three municipalities within that region, the highest frequency (p < 0.05) was observed in Seropedica (59.46%), followed by Itaguai (29.05%) and Mangaratiba (11.49%). Regarding factors associated with the host, mongrel dogs and those over the age of two presented higher frequency of antibodies to Leishmania spp. (p < 0.05). Concerning factors related to the environment and habits of the animal, dogs residing in rural areas (FR = 1.67, p = 0.0002), living outside the residence (FR = 1.42, p = 0.0197), with access to forest, streams and pastures (FR = 2.81, p = 0.0007), remaining loose (FR = 1.66, p = 0.0073), and those that had no shelter (FR = 2.16, p < 0.0001) were more likely to be seropositive. Canine leishmaniasis is a disease with high occurrence in the Itaguai micro-region, and aspects such as the definition of breed, age, habits and care by owners showed significant association in this micro-region.

The influence of the fasting period on the number of nymphal instars and on the sex ratio of Argas (Persicargas) miniatus (Acari: Argasidae)

The current study investigated the biology of nymphs of the first and second instars of Argas (Persicargas) miniatus. Nymphs were deprived of food for 15, 30 or 60 days and held at 27 ± 1 °C and 80 ± 10% relative humidity (controlled conditions) or at room conditions of temperature and relative humidity. Nymphs of first instar deprived of food for 15 or 30 days molted to second and third instars in both controlled and room conditions. Nymphs of the first instar deprived of food for 60 days had 28 and 37% mortality in controlled and room conditions, respectively; and survivors did not attach to the host. Nymphs of the second instar, deprived of food for 60 days, molted either to the third instar or to males after feeding on Gallus gallus, and the nymphs of the third instar developed to adults (42.42% males and 36.36% females when nymphs were held in controlled temperature and humidity conditions, and 40.54% males and 48.65% females when nymphs were held in room conditions). The remainder of the nymphs molted to the fourth instar and then molted to females. In conclusion, the nymphal starvation period of 60 days determined the number of nymph instars in the life cycle of A. miniatus under the experimental conditions studied.

Fatores de risco associados à frequência de anticorpos contra Babesia bovis e B. bigemina em bovinos da região sul de Moçambique

The study aimed to evaluate the risk factors associated with the frequency of IgG antibodies against Babesia bovis and B. bigemina in cattle in southern Mozambique. Eight hundred and nine serum samples were collected from cattle in three provinces namely Maputo, Gaza and Inhambane, and tested by indirect enzyme-linked immunosorbent assay (i-ELISA) to assess the humoral immune response towards B. bovis and B. bigemina. The chi-square test at 5% significance was used to determine whether there was an association between gender, age and geographic origin of seropositive animals. The overall prevalence was 78.8% (548/695) for B. bovis and 76.0% (528/695) for B. bigemina. The origin of the animals showed a significant association (p 0.05). Maputo province had the highest rate of positive animals, with 93.7% (118/126) for B. bovis and 97.6% (123/126) for B. bigemina. In Gaza province 77.3% (321/415) of the animals were positive for B. bovis and 67.5% (280/415) for B. bigemina, while in the province of Inhambane the levels of seropositivity were 70.8% (109/154) and 81.2% (125/154) for B. bovis and B. bigemina respectively. In the present study, the frequency of cattle positive for B. bovis and B. bigemina was shown to increase among older age groups, suggesting that infection and re-infection persisted even after the primary infection. Thus, this region is considered to be in a state of enzootic stability with regards to B. bovis and B. bigemina.

Características biológicas de Boophilus microplus (acari: ixodidae) a partir de infestação experimental em cão

3 ABSTRACT:- FRANQUE, M. P.; SANTOS, H. A.; SILVA, G. V. O.; TAJIRI, J. T.; MASSARD, C. L. (Biological characteristics of Boophilus microplus (Acari: Ixodidae) on dog under experimental infestation.) Caracteris- ticas biologicas de Boophilus microplus (Acari: Ixodidae) a partir de infestacao experimental em cao. Revista Brasileira de Parasitologia Veterinaria, v. 16, n. 4, p. 238-242, 2007. Departamento de Parasitologia Animal, Instituto de Veterinaria, Universidade Federal Rural do Rio de Janeiro, Km 7 da BR 465, Seropedica, RJ 23890.000, Brazil. E-mail: carlosmassard@ufrrj.br Boophilus microplus, a common parasite of cattle, has eventually reported in dogs. To describe biological features of this parasitism, one dog was experimentally infested with 10,000 larvae of B. microplus which were previously held in acclimatized camera at 27 ±1o C and relative humidity up to 80%. The mean of parasitic phase was 24.4 ± 1.50 days, with 0.42% of recovery rate. Of 21 natural detached B. microplus females, six engorged enough (75.1 ± 30.23mg) to achieve posture. The mean period of pre-posture was 4.33 ± 1.37 days and the means period of posture was 9.17 ± 2.32, producing a mean of 18.78 ± 15.34 posture weight. The mean of eggs production index observed was 22.38%. The results showed that B. microplus females fed on dogs to complete their life cycle. The females collected were able to ovoposite viable eggs suggesting that dogs can be a possible alternative hosts to B. microplus, especially when there is no other preferential host species available.

A new quantitative PCR method for the detection of Anaplasma platys in dogs based on the citrate synthase gene

Anaplasma platys is an obligate intracellular bacterium that primarily affects dogs, but it can also infect humans. Our study aimed to standardize a quantitative real-time (q)PCR method using the citrate synthase gene (gltA) as a specific target for A. platys detection in naturally infected dogs. Primers (gltA84F and gltA84R) and probe (PLATYSp) were designed to amplify an 84-bp fragment based on the gltA gene sequences of A. platys available in GenBank. A total of 186 dog blood samples originating from the Brazilian state of Rio de Janeiro were tested by qPCR. Additionally, the same samples were tested by cytology and a nested (n)PCR that targeted the 16S ribosomal DNA to determine the performance of our qPCR method compared to these existing techniques. Among the samples tested with qPCR, 17.2% were considered positive, significantly more than detected by nPCR (14.0%). Under optical microscopy, inclusions were observed in platelets of 25.3% of the samples, and among these samples, only 33.9% were identified as positive for A. platys using qPCR. The qPCR technique proved to be more specific than cytology and to have superior sensitivity to nPCR for detecting A. platys in dogs. The development of this new qPCR method contributes to the advancement of research involving A. platys. Furthermore, it can be used to quantify the presence of this bacterium to evaluate the treatment of infected animals, or even as a more sensitive and specific tool for situations indicating possible clinical disease but with negative cytology.

Infestação experimental de equinos por Rhipicephalus (Boophilus) microplus

Este estudo descreve caracteristicas biologicas da fase parasitaria de Rhipicephalus (Boophilus) microplus em equinos experimentalmente infestados. Quatro equinos foram infestados experimentalmente inicialmente com 20.000 e, posteriormente, tres vezes com 40.000 larvas. Em todas as infestacoes, os animais desenvolveram intensa reacao cutânea no local de fixacao das larvas, caracterizadas pelo intenso prurido local e pela elevacao do pelo, seguida por areas de alopecia com lesoes ulceradas circunscritas. Foi observada mortalidade aproximada de 90% e 60% para larvas e ninfas, respectivamente. Na infestacao inicial com 20.000 larvas, nenhuma femea ingurgitada ou semi-ingurgitada foi recuperada. Apos tres infestacoes realizadas com 40.000 larvas de R. microplus, dois equinos foram considerados resistentes a infestacao por nao terem sido recuperadas nenhuma femea ingurgitada ou semi-ingurgitada. Um equino foi considerado moderadamente resistente, do qual um numero pequeno de femeas semi-ingurgitadas foi recuperado e um susceptivel, no qual R. microplus completou seu ciclo parasitario com mortalidade de 30% dos adultos e duracao media da fase parasitaria de 33,3; 36,9 e 38,5 dias, respectivamente nas tres infestacoes realizadas com 40.000 larvas. A taxa de recuperacao das femeas variou entre 0,9% e 1,8%. Este estudo mostra que e possivel estudar a biologia de R. microplus em equinos, a partir de infestacoes experimentais de R. microplus. Contudo, o numero de larvas a ser usado para infestacao e a existencia de equinos susceptiveis e resistentes devem ser considerados.