Prevalence of Toxoplasma gondii and Neospora caninum infections in sheep from Federal District, central region of Brazil

Serum samples from 1028 sheep were collected from 32 herds within Federal District, in the central region of Brazil. The samples were examined by indirect fluorescent antibody test (IFAT) using sera diluted 1:64 and 1:50 as cut-off values for the detection of antibodies against Toxoplasma gondii and Neospora caninum, respectively. The observed prevalence for T. gondii infection was 38.22% (26.81%< CI 0.95 < 49.62%), and the titers ranged from 64 to 65536. The observed prevalence for N. caninum infection was 8.81% (7.08%< CI 0.95 < 10.53%). The titers ranged from 50 to 51200. The reactant sera to both pathogens corresponded to 4.67% of the samples. The risk factors were not determined because of the absence of negative herds for T. gondii and the high proportion of positive herds for N. caninum (87.50%). The prevalence for T. gondii infection was significantly higher among males than in females. The present work is the first report on seroprevalence of T. gondii and N. caninum in sheep from Federal District and shows that infection by both parasites is widespread in the ovine population from this region.

Occurrence of ticks (Acari: Ixodidae) in the municipality of Goiatins, Tocantins

Due to a suspected human case of Brazilian Lyme-like disease in the city of Goiatins, Tocantins State, an epidemiological survey was carried out in eight counties in this region during September 2007 and February 2008, where 1,890 ticks were collected from domestic animals and from the environment. A total of eight tick species were identified: Rhipicephalus sanguineus, Rhipicephalus (Boophilus) microplus, Dermacentor nitens, Amblyomma cajennense, Amblyomma oblongoguttatum, Amblyomma ovale, Amblyomma parvum and Amblyomma tigrinum. The last four species were described for the first time in this region. Although human parasitism by ticks is frequently described in Goiatins, no ticks collected from humans were analyzed. The Study of ixodids in this region contributes with the survey of Brazilian ticks, as well as the elucidation of the possible transmission of the agent that caused the Brazilian Lyme-like disease case in Goiatins.

Shedding of Neospora caninum oocysts by dogs fed different tissues from naturally infected cattle

Neospora caninum is one of the most important causes of abortion in dairy cattle worldwide. The distribution of N. caninum in tissues of adult cattle is unknown and the parasite has not been demonstrated histologically in tissues of cows. In the present study the distribution of N. caninum in different tissues of adult cattle was evaluated by bioassays in dogs. Seventeen dogs (2-3 month-old) were fed different tissues of 4 naturally exposed adult cattle (indirect fluorescent antibody test N. caninum titer >= 400): 5 were fed with masseter: 5 with heart, 3 with liver, 4 with brain, and 3 pups were used as non-infected control. Two dogs fed masseter, 2 fed heart, 1 fed liver, and 3 fed brain shed oocysts, and all dogs presented no seroconvertion to N. caninum during the observation period of 4 weeks. The oocysts were confirmed as N. caninum based on the detection of N. caninum-specific DNA by PCR and sequencing. The results indicate that dogs can be infected by N. caninum with different tissues of infected cattle. (C) 2011 Elsevier B.V. All rights reserved.

Use of the real time RT-PCR for immune related gene expression quantitation in experimentally infected Neospora caninum bovine calves

Neospora caninum is one of the main causes of abortion and natimortality in cattle. Host immune defense is capable to inhibit tachyzoite activity during acute infection, but there is no action against bradyzoites in tissue cysts. Activation and modulation of this response is controlled by cell mediators. The real-time RT-PCR technique was employed to detect some of those mediators during N. caninum infection. Holstein and Nelore calves intramuscularly infected with tachyzoites and uninfected controls were slaughtered at the sixth day post-infection and popliteal lymph node, liver and brain cortex samples were analyzed. Real-time RT-PCR detected gene expression in all tissues. No significant variation of GAPDH gene expression was detected among groups, its amplification efficiency was similar to the other genes tested and it was used as the endogenous control for the analysis. Comparisons between infected and uninfected groups allowed the relative gene expression quantification. IFN-gamma and TNF-alpha genes showed increased expression in some samples. iNOS and TGF-beta 1 genes had some non-significant variations and IL-4 and IL-10 stayed pratically inaltered.

Molecular diversity of Brazilian strains of porcine circovirus type 2 (PCV-2)

Porcine circovirus 2 (PCV-2) is associated with a broad range of syndromes. In this study, eight pig tissue samples from two Brazilian states were analyzed using six PCR primer pairs amplifying a 1705-bp fragment of the PCV-2 genome. The NJ distance-based method was used for the phylogenetic analysis with the eight field strains herein, 15 GenBank sequences and using PCV-1 as an out-group. This yielded two major clusters (A and B) for this viral species, with the Brazilian strains segregating with European and Asian sequences. Nucleotide identity was 99.7 to 100% among the sequences. This information can be used in further studies of pathogenesis related to PCV-2 in Brazil. (C) 2007 Elsevier Ltd. All rights reserved.

Differential diagnosis of oocysts of Hammondia-like organisms of dogs and cats by PCR-RFLP analysis of 70-kilodalton heat shock protein (HSP70) gene

Organisms of the genera Toxoplasma, Hammondia and Neospora, the Hammondia-like organisms, are closely related coccidian with similarly sized oocysts. Therefore, a diagnosis based on microscopy of oocysts in feces is not a method of choice for species identification of these important parasites. In this paper, we present a polymerase chain reaction coupled with restriction fragment length polymorphism (PCR-RFLP) method to differentially diagnose oocysts of Toxoplasma gondii from oocyst of Hammondia hammondi. Another PCR-RFLP was designed to differentiate oocysts of Hammondia heydorni from oocysts of Neospora spp. Both PCR-RFLP are based on nucleotide sequences of the Hsp70 coding gene. In conclusion, we presented two alternative molecular diagnostic assays that can be successfully applied for the differentiation of oocysts of Hammondia-like organisms shed by felids and canids.

OCCURRENCE AND MOLECULAR DIAGNOSIS OF CRYPTOSPORIDIUM SERPENTIS IN CAPTIVE SNAKES IN SAO PAULO, BRAZIL

The present study aimed to determine whether Cryptosporidium oocysts were present in stools from captive snakes at Fundacao Parque Zoo logic (Zoological Park Foundation) in Sno Paulo, Brazil. Two collections were performed; the first in July 2008 and the second in February 2009. Fecal samples were collected from 74 enclosures that housed 101 individuals of 23 snake species. The stool specimens collected from 16 out of the 74 enclosures (21.6%) contained Cryptosporidium spp. oocysts; all of them were confirmed as Cryptosporidium serpent is, using molecular techniques. Only in three (18.7%) out of the 16 enclosures with positive samples were there animals with clinical signs compatible with infection by C. serpentis, such as regurgitation and significant progressive weight loss. From the results, it was concluded that diagnostic examinations need to be performed periodically, even on clinically healthy animals, as a preventive measure.

Genotyping of Toxoplasma gondii isolates from free range chickens in the Pantanal area of Brazil

The aim of this paper was to genetically characterize Toxoplasma gondii isolates from free range chickens in regions of Brazilian territory in the state of Mato Grosso do Sul (MS) where T. gondii strains have never been studied. In total, T. gondii isolates from 22 free range chickens were included in this study. Fifty chickens from Eldorado, thirty from Rio Verde and ten from Aquidauana were sampled between January and April 2007. In relation to the genetic diversity of T. gondii isolates from chickens in MS, the magnitude of the diversity in the isolates sampled in this study was comparable to the overall diversity in a composite data set. These 22 isolates in MS revealed 11 genotypes, whereas the 321 isolates ever genotyped in Brazil have revealed 95 genotypes. The values of Simpson`s Diversity Index for the whole population of T. gondii isolates in Brazil, the whole population of T. gondii isolates from chickens in Brazil and the population surveyed in this study were 0.97, 0.95 and 0.90, respectively. Seven of the 11 genotypes revealed from chicken isolates from MS are newly described genotypes and six of them each have a single isolate. In conclusion, the results obtained from isolates in MS corroborate previous studies on T. gondii isolates in Brazil, thus confirming their diversity and atypicality. Nonetheless, the applicability of PCR-RFLP markers for epidemiological inferences remains controversial. (c) 2011 Elsevier B.V. All rights reserved.

Cross-mating experiments with geographically different populations of Amblyomma cajennense (Acari: Ixodidae)

The present study evaluated the reproductive compatibility of the crosses between adult ticks of the following three geographically different populations of Amblyomma cajennense: State of So Paulo (SP), southeastern Brazil; State of Rondnia (RO), northern Brazil; and Colombia (CO). In addition, crosses between A. cajennense ticks from Argentina (AR) and SP ticks were also performed. The Argentinean population (AR) was compatible with SP because their crosses resulted in high % egg hatching (mean values ranging from 71.5 to 93.5%), similarly to all homologous (intrapopulational) crosses. In contrast, the tick populations SP, RO, and CO were shown to be incompatible with each other, since their heterologous (interpopulational) crosses always resulted in very low % egg hatching (range: 0-5%). The F(1) larval offspring derived from some of these females that yielded 5% egg hatching were reared until the F(1) adult stage. In all cases, only adult females molted from engorged nymphs. These F(1) females were likely to be a product of thelytokous parthenogenesis of the SP, RO, and CO females that were used in the heterologous crosses. Reproductive incompatibility is not expected to occur between different populations of a single species. Thus, our results suggest that the taxon A. cajennense might be represented by a complex of different species, whereas SP and AR ticks might represent a single species. Further populational genetic studies, coupled with extensive morphological analyses, are needed to clarify and determine a possible complex of valid species that might have been classified under the taxon A. cajennense.

Effects of the utilization of homeopathic elements in commercial diluent on swine sperm viability

It has been speculated that the homeopathic treatment of sperm cells in order to improve semen quality could be promising. However, few data is available and its use in spermatozoa requires investigation. It is well established that mitochondrial membrane potential is an important viability parameter of spermatozoa and it is intimately related to reproductive efficiency. In this manner, new technologies in order to improve the activity of sperm cells and, finally, the fecundity of swine herds are of extremely importance. Due to the lack of knowledge of homeopathic treatment effect on spermatozoa, the aim of the present study was to verify the effect of three different homeopathic treatments on viability of boar sperm cells. Three homeopathic treatments composed by Pulsatila CH6, Pulsatila and Avena CH6, Avena CH6 and one control treatment (sucrose) were added to diluted boar semen, which were cooled for 24 or 48 h. Interestingly, no positive effect of homeopathic treatments was observed over semen viability. However, it was demonstrated that the 24 h of cooling storage provided more viable sperm cells when compared to the 48-h period. This effect of storage period on sperm viability was assessed by intact plasmatic membrane, intact acrosome and mitochondrial membrane potential evaluation.

Tick toxicosis in a dog bitten by Ornithodoros brasiliensis

Ticks are hematophagous parasites of people and animals and are a public health hazard in several countries. They are vectors of infectious diseases; in addition, the bite of some ticks, mainly from the Ornithodoros genus, may lead to local lesions and systemic illness, referred to as tick toxicosis. In this report, we describe a dog bitten by Ornithodoros brasiliensis, popularly known as the mouro tick. The main clinical findings were disseminated skin rash, pruritus, mucosal hyperemia, lethargy, and fever. Laboratory abnormalities 48 hours after the bites occurred included mild nonregenerative anemia, eosinophilia, basophilia, increased serum creatine kinase activity, increased serum C-reactive protein concentration, and prolonged coagulation times. Tick-borne pathogens were not detected by PCR analysis or serologic testing, supporting the diagnosis of a noninfectious syndrome due to tick bite, compatible with tick toxicosis.

Ticks Infesting Wildlife Species in Northeastern Brazil With New Host and Locality Records

From September 2008 to March 2010, 397 ticks (315 larvae, 33 nymphs, 23 females, and 26 males) were collected from captive and free-living wildlife species in northeastern Brazil. Six tick species were identified, including Amblyomma auricularium (Conil) on Tamandua tetradactyla (L.),Amblyomma dubitalum Neumann on Hydrochaeris hydrochaeris (L.), Nectomys rattus (Pelzen) and T. tetradactyla, Amblyomma parvim A ragao on T. tatradactyla, Amblyomma rotundatum Koch on Boa constrictor L., Chelonoidis carbonaria (Spix), Kinosternon scorpioides (L.) and Rhinella jimi (Stevaux), Amblyomma cerium Koch on Bradypus variegatus Schinz, and Rhipicephalus sanguineus (Latreille) on Lycalopex vetulus (Lund). Nectomys rattus and T. tetradactyla are new hosts for A. dubitatum This study extends the known distribution of A. dubitatum in South America and provides evidence that its geographical range has been underestimated because of the lack of research. Four (A. dubitatum, A. parvum, A. rotundatum, and R. sanguineus) of six tick species identified in this study have previously been found on humans in South America, some of them being potentially involved in the transmission of pathogens of zoonotic concern.

Genotyping of potentially zoonotic Giardia duodenalis from exotic and wild animals kept in captivity in Brazil

We have studied the variability of glutamate dehydrogenase (gdh) and small subunit ribosomal (SSU) rRNA coding genes of Giardia species in fecal samples isolated from wild and exotic animals in Brazil, and compared with homologous sequences of isolates from human and domestic animals characterized in previous studies. Cysts of Giardia duodenalis were obtained from feces of naturally infected monkeys (Alouatta fusca) (n = 20), chinchillas (Chinchilla lanigera) (n = 3), ostriches (Struthio camelus) (n = 2) and jaguar (Panthera onca) (n = 1). Assemblage AI was assigned to the unique isolate of jaguar. All the samples from monkeys, chinchillas, and ostriches were assigned to Assemblage B. There was little evolutionary divergence between the referred isolates and isolates described elsewhere. The Assemblage B isolates identified in this study were closely related to Assemblage BIV isolated from humans. The molecular identification of Assemblages A and B of G. duodenalis isolates from exotic and wild animals demonstrates that such hosts may be a potential reservoir for zoonotic transmission of G. duodenalis. (C) 2011 Elsevier B.V. All rights reserved.

Nymphs of the genus Amblyomma (Acari: Ixodidae) of Brazil: descriptions, redescriptions, and identification key

Together with the larval stage, the nymphal stage of ticks of the genus Amblyomma are the most aggressive ticks for humans entering areas inhabited by wildlife and some domestic animals in Brazil. However, due to the absence of morphological descriptions of the nymphal stage of most Brazilian Amblyomma species, plus the lack of an identification key, little or nothing is known about the life history of Amblyomma spp. nymphs in the country. In the present study, morphological description of the nymphal stage, illustrating important external characters through scanning electron microscopy, is provided for nymphs of 15 Amblyomma species that occur in Brazil, for which the nymphal stage had never been described: A. aureolatum, A. auricularium, A. calcaratum, A. coelebs, A. fuscum, A. humerale, A. incisum, A. latepunctatum, A. naponense, A. nodosum, A. ovate, A. pacae, A. pseudoconcolor, A. scalpturatum, A. varium. In addition, the nymphal stage of 12 Amblyomma species, which had been previously described, are redescribed: A. brasiliense, A. cajennense, A. dissimile, A. dubitatum, A. longirostre, A. oblongoguttatum, A. parked, A. parvum, A. romitii, A. rotundatum, A. tigrinum, A. triste. The descriptions and redescriptions totalized 27 species. Only 2 species (A. geayi, A. goeldii) out of the 29 Amblyomma species established in Brazil are not included in the present study. A dichotomous identification key is included to support taxonomic identification of the nymphal stage of 27 Amblyomma species established in Brazil. (C) 2010 Elsevier GmbH. All rights reserved.

Acquisition and transmission of Hepatozoon canis (Apicomplexa: Hepatozoidae) by the tick Amblyomma ovale (Acari: Ixodidae)

The present study aimed to evaluate under controlled conditions the acquisition of Hepatozoon canis by Amblyomma ovale after feeding on infected dogs, and the subsequent induction of infection in uninfected dogs that ingested the experimentally infected ticks. Two H. canis naturally infected dogs were infested with A. ovate adult ticks derived from an uninfected laboratory tick colony. After feeding, two A. ovale females presented H. canis oocysts in the hemolymph at the first and fourth days after removal of ticks from dogs. The oocysts had an average size of 244.34 mu m x 255.46 mu m. Three uninfected dogs were fed with ticks previously fed on the infected dogs. Only one dog became infected 32 days after oral inoculation, presenting circulating gametocytes, parasitemia less than 1%, and positive PCR confirmed to be H. canis by DNA sequencing. The results obtained indicated A ovale ticks as potential vector of H. canis in rural areas of Brazil. (C) 2009 Elsevier B.V. All rights reserved.