Comparative performance evaluation of four different methods for diagnosing Giardia infection in dogs and zoonotic assemblages' identification.

Giardia duodenalis (syn. G. intestinalis or G. lamblia) is a parasitic protozoan that infects the upper intestinal tract of a broad range of hosts, including humans and domestic animals. Thus, it has raised concerns about the public health risk due to companion animals. Recently, with the improvement of living standards and increasing contacts between pets and humans, the zoonotic transmission of Giardia has dramatically increased. From a genetic point of view, G. duodenalis should be viewed as a complex species that includes eight different species-specific genetic assemblages. The laboratory diagnosis is mainly based on the finding of microscopic cysts in stool samples by coprological examination. Other methods include the detection of antigens, immunoassays or PCR protocols, which allow the identification of Giardia assemblages. The study aimed to compare the performance of Direct Fluorescence Antibody test (DFA), zinc sulfate flotation technique (ZnSO4), rapid diagnostic test (RDT), end-point PCR amplification (PCR) for the detection of Giardia and to identify the concerning assemblages in a canine population from Central Italy. Direct fluorescence antibody test is the reference standard for laboratory diagnosis of Giardia in fecal samples from dogs, despite the microscopic examination after flotation remains the most useful method in many veterinary diagnostic centers. The present findings demonstrate the high performance of DFA and ZnSO4 in detecting Giardia, while RDT may be useful as alternative or complementary method to the DFA and ZnSO4. PCR performance was low, but it allowed determining Giardia B zoonotic assemblage in 25% of the PCR-positive specimens (15 out of 60), while the remaining PCR-positive isolates belonged to the dog-specific assemblage C. The 26% prevalence of G. duodenalis detected by DFA in owned dogs and the identification of potentially zoonotic assemblages underline the potential risk for public health and indicate frequent cross-species transmission of the parasite between humans and dogs.

Naturally acquired bovine besnoitiosis: Disease frequency, risk and outcome in an endemically infected beef herd.

The recent spread of bovine besnoitiosis warrants further epidemiological investigations to improve the knowledge on disease development. Thus, a 4-year longitudinal open cohort study was conducted in the first German cattle herd naturally infected with Besnoitia besnoiti. At seven herd-visits between 2008 and 2012, fourteen breeding bulls (>1.5 years) and 131 females (>1 year) were examined clinically and serologically. In females, clinical and serological prevalences, incidence and remission rates were determined. In addition, the association of age, antibody levels and number of visible parasitic cysts with clinical and serological outcome was investigated. The seroprevalence (89.4%-100%) and serological incidence rate (140.5 per 100 animal-years) were considerably higher than the clinical prevalence (23.5%-36.6%) and clinical incidence rate (16.7 per 100 animal-years). Of 33 new clinical and 12 new serological cases, only 6.7% (3/45) attracted attention with clinical signs of acute bovine besnoitiosis. The apparent serological remission rate (1.9 per 100 animal-years) was considerably lower than the clinical remission rate (37.3 per 100 animal-years). A median cyst score of <1 and mean immunofluorescent antibody test (IFAT) titre of ≤1,600 over the entire observation period was significantly associated with a negative clinical outcome at the end. Overall cyst score was not significantly associated with serological outcome and age had no significant influence on clinical and serological outcome. Within 4 years, there was a significant reduction in cyst scores and IFAT titres in the same animals, leading to eight clinically and serologically negative animals in the end. Two initially negative animals achieved clinical and apparent serological remission in about 2.5 years. In bulls, the time between herd entry and seroconversion was 7-30 months and the serological incidence rate was nearly identical to the rate in females (142.0 per 100 animal-years). This shows that a high B. besnoiti prevalence leads to infection of bulls within a short time period.

Epidemiological survey in Leczynsko-Wlodawskie Lake District of eastern Poland reveals new evidence of zoonotic potential of Giardia intestinalis.

Faecal samples from 297 farm animals were collected from 18 households in distinct sites of the Leczynsko-Wlodawskie Lake District of eastern Poland. They included samples from 86 cattle (Bos taurus), 84 pigs (Sus scrofa f. domestica), 81 sheep (Ovis aries), 10 horses (Equus caballus), and 36 dogs (Canis lupus familiaris). The samples were examined for the presence of Giardia intestinalis by the Direct Fluorescence Assay (DFA) and semi-nested PCR. All amplicons were sequenced on both strands. By DFA, cysts of Giardia spp. were detected in 66 of 297 faecal samples (22.2%). Positive specimens for Giardia spp. were derived from 29.8% of examined pigs, 21.0% of sheep, 18.6% of cattle, 10% of horses, and 19.4% of dogs. Based on the detection of the ß-giardin gene by PCR, 39 (13.1%) of the 297 examined samples were recognized as positive. Detection of the presence of Giardia cysts by DFA test was overall significantly higher compared to PCR (p=0.0045). By PCR, Giardia was found in 28.1% of sheep, 11.6% of cattle, 10% of horses, 9.5% of pigs and 5.6% of dogs. Partial ß-giardin gene sequences were obtained for 73.7% of the PCR positive samples. From sequenced samples derived from the studied animals, Giardia were identified as assemblage A (8 samples), B (1 sample) and E (18 samples). As assemblages A and B may be zoonotic, the farm animals living in eastern Poland could be regarded as a potential source of Giardia infection for humans.

Comparison of diagnostic techniques for the detection of Cryptosporidium oocysts in animal samples.

While a large number of laboratory methods for the detection of Cryptosporidium oocysts in faecal samples are now available, their efficacy for identifying asymptomatic cases of cryptosporidiosis is poorly understood. This study was carried out to determine a reliable screening test for epidemiological studies in livestock. In addition, three molecular tests were compared to identify Cryptosporidium species responsible for the infection in cattle, sheep and horses. A variety of diagnostic tests including microscopic (Kinyoun's staining), immunological (Direct Fluorescence Antibody tests or DFAT), enzyme-linked immunosorbent assay (ELISA), and molecular methods (nested PCR) were compared to assess their ability to detect Cryptosporidium in cattle, horse and sheep faecal samples. The results indicate that the sensitivity and specificity of each test is highly dependent on the input samples; while Kinyoun's and DFAT proved to be reliable screening tools for cattle samples, DFAT and PCR analysis (targeted at the 18S rRNA gene fragment) were more sensitive for screening sheep and horse samples. Finally different PCR primer sets targetedat the same region resulted in the preferential amplification of certain Cryptosporidium species when multiple species were present in the sample. Therefore, for identification of Cryptosporidium spp. in the event of asymptomatic cryptosporidiosis, the combination of different 18S rRNA nested PCR primer sets is recommended for further epidemiological applications and also tracking the sources of infection.

Occurrence of Giardia and Cryptosporidium in captive chimpanzees (Pan troglodytes), mandrills (Mandrillus sphinx) and wild Zanzibar red colobus monkeys (Procolobus kirkii).

BACKGROUND: The aim of this study was to investigate the occurrence of Giardia duodenalis and Cryptosporidium spp. in primates and determine their zoonotic or anthropozoonotic potential. METHODS: Direct immunofluorescence was used to identify Giardia and Cryptosporidium from faecal samples. PCR and DNA sequencing was performed on positive results. RESULTS: Giardia cysts were identified from 5.5% (5/90) of captive chimpanzees and 0% (0/11) of captive mandrills in the Republic of Congo; 0% (0/10) of captive chimpanzees in Norway; and 0% of faecal samples (n = 49) from wild Zanzibar red colobus monkeys. Two Giardia positive samples were also positive on PCR, and sequencing revealed identical isolates of Assemblage B. Cryptosporidium oocysts were not detected in any of the samples. CONCLUSIONS: In these primate groups, in which interactions with humans and human environments are quite substantial, Giardia and Cryptosporidium are rare pathogens. In chimpanzees, Giardia may have a zoonotic or anthropozoonotic potential.

Utility of feline coronavirus antibody tests.

Eight different tests for antibodies to feline coronavirus (FCoV) were evaluated for attributes that are important in situations in veterinary practice. We compared four indirect immunofluorescent antibody tests (IFAT), one enzyme-linked immunosorbent assay (ELISA) (FCoV Immunocomb; Biogal) and three rapid immunochromatographic (RIM) tests against a panel of samples designated by consensus as positive or negative. Specificity was 100% for all but the two IFATs based on transmissible gastroenteritis virus (TGEV), at 83.3% and 97.5%. The IFAT and ELISA tests were best for obtaining an antibody titre and for working in the presence of virus. The RIM tests were the best for obtaining a result quickly (10-15 mins); of these, the Speed F-Corona was the most sensitive, at 92.4%, followed by FASTest feline infectious peritonitis (FIP; 84.6%) and Anigen Rapid FCoV antibody test (64.1%). Sensitivity was 100% for the ELISA, one FCoV IFAT and one TGEV IFAT; and 98.2% for a second TGEV IFA and 96.1% for a second FCoV IFAT. All tests worked with effusions, even when only blood products were stipulated in the instruction manual. The ELISA and Anigen RIM tests were best for small quantities of sample. The most appropriate FCoV antibody test to use depends on the reason for testing: in excluding a diagnosis of FIP, sensitivity, specificity, small sample quantity, rapidity and ability to work in the presence of virus all matter. For FCoV screening, speed and sensitivity are important, and for FCoV elimination antibody titre is essential.

Concomitant infection of Neospora caninum and Bovine Herpesvirus type 5 in spontaneous bovine abortions / Infecção simultânea de Neospora caninum e Herpesvirus bovino tipo 5 em casos espontâneos de aborto bovino

Bovine Herpesvirus type 5 (BoHV-5) has not been conclusively demonstrated to cause bovine abortion. Brain lesions produced by Neospora caninum and Bovine Herpesvirus type 1 (BoHV-1) exhibit common features. Therefore, careful microscopic evaluation and additional diagnostic procedures are required to achieve an accurate final etiological diagnosis. The aim of the present work was to investigate the occurrence of infections due to BoHV-1, BoHV-5 and N. caninum in 68 cases of spontaneous bovine abortions which showed microscopic lesions in the fetal central nervous system. This study allowed the identification of 4 (5.9%) fetuses with dual infection by BoHV-5 and N. caninum and 33 (48.5%) cases in which N. caninum was the sole pathogen identified. All cases were negative to BoHV-1. The results of this study provide evidence that dual infection by BoHV-5 and N. caninum occur during pregnancy in cattle; however, the role of BoHV-5 as a primary cause of bovine abortion needs further research. Molecular diagnosis of BoHV-5 and N. caninum confirmed the importance of applying complementary assays to improve the sensitivity of diagnosing bovine abortion.

Não está demonstrado até ao momento, que o Herpesvírus bovino tipo 5 (BoHV-5) seja um agente causal de aborto bovino. Uma vez que as lesões cerebrais tanto de Neospora caninum como de Herpesvírus bovino tipo 1(BoHV-1) têm características similares, é necessária uma avaliação microscópica cuidadosa, bem como exames laboratoriais adicionais, para obter um diagnóstico final preciso. O objetivo do presente trabalho foi investigar a presença de infeções por BoHV-1, BoHV-5 e N. caninum em 68 casos de aborto espontâneo, nos quais se verificaram lesões microscópicas no sistema nervoso central. Foram encontrados 4 (5,9%) fetos com infeção simultânea de BoHV-5 e N. caninum e 33 (48,5%) casos com infeção exclusiva de N. caninum. Todos os casos foram negativos a BoHV-1. Os resultados deste estudo demonstram que a infeção dual por BoHV-5 y N. caninum está presente durante a gestçao dos bovinos. Apesar disso, o papel de BoHV-5 como agente primário causal de aborto, carece de mais investigaçao. O diagnóstico molecular de BoHV-5 e N. caninum confirmou a importância de se aplicar ensaios complementares para melhorar a sensibilidade do diagnóstico de aborto bovino.

Diagnostic utility of a direct immunofluorescence test to detect feline coronavirus antigen in macrophages in effusive feline infectious peritonitis.

The antemortem diagnosis of feline infectious peritonitis (FIP) remains challenging in clinical practice, since current testing methods have suboptimal diagnostic accuracy. Immunohistochemical testing of biopsy specimens and postmortem examination are the standard diagnostic methods, although direct immunofluorescence (DIF) testing to detect feline coronavirus in macrophages in effusion specimens has been reported to have 100% specificity and has been recommended as an antemortem confirmatory test. The aim of this study was to compare the results of DIF testing in antemortem feline effusions with postmortem results using field samples. Effusion specimens were collected antemortem from 17 cats and tested by DIF, followed by postmortem examination. Histopathological examination of specimens collected at postmortem confirmed FIP in 10/17 cases and ruled out FIP out in 7/17 cases. Antemortem DIF testing was positive in all 10 cases confirmed as FIP at postmortem examination. In the seven cats where FIP was ruled out at postmortem examination, DIF was negative in five cases and positive in the remaining two cases. The calculated sensitivity of DIF testing was 100% and the specificity was 71.4%. Duplicate effusion specimens from eight cats that were initially DIF positive were stored refrigerated (4 °C) or at room temperature (22-25 °C) and subjected to serial DIF testing to determine the duration of positive results. DIF-positive specimens stored at both temperatures retained their positive status for at least 2 days.

Scrapie e seu diagnóstico diferencial em ovinos no Mato Grosso do Sul / Scrapie and differential diagnosis in sheep in Mato Grosso do Sul, Brazil

Scrapie é uma doença infecciosa, neurodegenerativa fatal, causada pelo príon scrapie (PrPsc). Apresenta-se tanto na forma clássica em ovinos e caprinos geneticamente susceptíveis quanto na forma atípica em ovinos. A primeira notificação oficial do Brasil à Organização Mundial de Saúde Animal (OIE), um caso da forma clássica diagnosticado no Rio Grande do Sul ocorreu em 1985, mas a doença já havia sido diagnosticada no mesmo Estado em 1978. Este trabalho objetivou descrever dois surtos de Scrapie em ovinos em Mato Grosso do Sul (MS), Brasil e investigar, por meio de imuno-histoquímica (IHQ) a presença de PrPsc no Sistema Nervoso Central (SNC) de ovinos examinados entre 2003 e 2010. Na primeira parte observaram-se dois ovinos com sinais clínicos típicos de scrapie, detalhando-se os sinais neurológicos, dados epidemiológicos, histopatológicos e amostras teciduais em duplicata desses ovinos foram encaminhadas para realização de diagnóstico de Raiva e para diagnóstico IHQ para príon. Na segunda parte realizou-se levantamento de laudos de necropsia e diagnósticos histopatológicos de ovinos, no período de maio de 2003 a março de 2010. Amostras de sistema nervoso central de 51 casos foram selecionados, incluindo os dois já com diagnóstico de Scrapie mencionados acima; os tecido de todos esses ovinos foram submetidos à IHQ para detecção de proteína priônica. Os 49 ovinos avaliados apresentaram resultado negativo na IHQ para príon.

Scrapie is a fatal neurodegenerative infectious disease, caused by the scrapie prion (PrPsc), that can both in the as the classic form in genetically susceptible sheep and goats and in the atypical form in sheep. The first official notification of scrapie from Brazil was made to the World Organization for Animal Health (OIE) in 1985, in the state of Rio Grande do Sul, although the disease was first documented in this Brazilian state in 1978. The objective this paper was to describe two outbreaks of scrapie in sheep from Mato Grosso do Sul (MS), Brazil, and to investigate by immunohistochemistry (IHC) the presence of PrPsc in samples from the CNS of sheep examined during 2003 and 2010. The study was conducted in two stages; the first was the observation of two sheep with typical clinical signs of scrapie that underwent clinical examination with emphasis on neurological parameters, epidemiological data collection, necropsy and collection of samples in duplicate forwarded to the diagnosis of rabies, and for the IHC diagnosis of Transmissible Spongiform Encephalopathies. In the second part of the study, a survey was made the necropsy reviewing gross findings and histopathological diagnoses in sheep from May 2003 to March 2010. Samples of the central nervous system of fifty-one cases, including the two sheep mentioned above were subjected to IHC for detection of prion protein. The other 49 sheep, although displaying neurological-disease which should be included as scrapie differential diagnosis, had their tissues submitted to IHC resulting negative.

Coprological survey in pet reptiles in Italy.

Faecal samples were collected from 324 pet reptiles showing no clinical signs, including 28 saurian species (n=192), three ophidian species (n=74) and three chelonian species (n=58). Samples were examined for the presence of intestinal parasites by direct smear and faecal flotation, while direct immunofluorescence assays were used to reveal the presence of Cryptosporidium oocysts and Giardia cysts. Overall, 57.4 per cent of the reptiles were harbouring intestinal parasites. These included oxyurids (16 per cent), coccidia (12.3 per cent), flagellates (9.3 per cent), strongyles (6.8 per cent), coccidia plus oxyurids (4.9 per cent), coccidia plus flagellates (1.8 per cent), coccidia plus strongyles (1.8 per cent), oxyurids plus strongyles (1.2 per cent), oxyurids plus flagellates (1.2 per cent), Cryptosporidium species (1.2 per cent) and strongyles plus flagellates (0.6 per cent). Intestinal parasites were more prevalent in saurians than in ophidians and chelonians, in insectivores than in carnivores, omnivores and herbivores, and in wild-caught than in captive-born reptiles. A highly significant difference was observed for saurians versus chelonians (odds ratio [OR]=2.20, 95 per cent confidence interval [CI] 1.21 to 3.99), insectivores versus herbivores (OR=2.38, 95 per cent CI 1.26 to 4.49) and in wild-caught versus captive-born pet reptiles (OR=2.36, 95 per cent CI 1.27 to 4.40).

Chlamydophila psittaci and Chlamydophila pecorum infections in goats and sheep in Egypt.

The aim of this study was to investigate the epidemiology of chlamydiosis in free-ranging asymptomatic and diarrhoeic sheep and goats in Egypt. Faecal swabs were examined for the presence of Chlamydiae by culture in Vero cells and chick embryos, and staining with Giménez, direct fluorescein-conjugated monoclonal antibodies, and immunoperoxidase. Specific chlamydial DNA was identified by amplification of the omp2 gene. The asymptomatic goats were 50% positive for the presence of the omp2 gene of the family Chlamydiaceae, and all isolates were Chlamydophila psittaci. The percentage of diseased goats in which Chlamydiaceae were identified was 16.2%, and all were positive for Cp. psittaci. Of the asymptomatic sheep, 6.7% were positive for the omp2 gene of the family Chlamydiaceae, and again all were positive for Cp. psittaci. In contrast, 42.9% of the samples that were collected from the diseased sheep were positive for Chlamydiaceae, of which 25.7% were Cp. psittaci and 4.8% Cp. pecorum.

Epidemiological survey on equine cryptosporidium and giardia infections in Italy and molecular characterization of isolates.

Cryptosporidium and Giardia are two of the most common enteric pathogens of domestic and wild animals and humans. However, little is known on the prevalence, clinical manifestations and economic and zoonotic significance of these infections in horses. This study was undertaken to investigate the prevalence, excretion patterns and risk factors related to the faecal shedding of Cryptosporidium oocysts and Giardia cysts in horses and the zoonotic potential of species/genotypes isolated. The survey was performed on 120 foals and 30 broodmares reared in five Italian farms. Foals were divided in four homogeneous groups of 30 animals each (age classes: 0-2, 2-4, 4-8, >8 weeks). Three sequential faecal samples were collected from each animal and analysed by three techniques: direct fluorescent antibody test (DFA), faecal flotation (FF) and stained faecal smears (SFS). The DFA results showed a prevalence of 8% for Cryptosporidium and of 13.33% for Giardia; the prevalence values obtained by FF and SFS were lower and in poor agreement with DFA results. Giardia and Cryptosporidium infections were more common in foals (23.33% and 26.66% respectively) and higher excretions were observed in the youngest foals. Distribution of Cryptosporidium prevalence was statistically related to farms (P < 0.01), age of animals (P < 0.01), but was unrelated to the presence of diarrhoea. In the case of Giardia, the prevalence was only related to age (P < 0.01). Pattern sheddings were related to intestinal diseases and horse age (P < 0.01). Risk factors for shedding included residence farms and age older than 8 weeks for both parasites. All DFA-positive faecal samples were submitted to DNA extraction and PCR to determine Giardia and Cryptosporidium species/genotypes. Sequence analysis of the COWP gene of Cryptosporidium and of the SSU-rRNA gene of Giardia revealed that they were identical to each other and identified Cryptosporidium parvum and Giardia duodenalis assemblage E. The potential role of infected horses in zoonotic transmission of Cryptosporidium was supported by the findings of this study.

Aspectos clínicos e patológicos em bovinos afetados por raiva com especial referência ao mapeamento do antígeno rábico por imuno-histoquímica / Clinical and pathological aspects in cattle affected by rabies with special reference to the rabies antigen mapping by immunohistochemistry

Este estudo retrospectivo incluiu achados clínicos e patológicos de 15 bovinos afetados por raiva. Em treze dos quinze casos, raiva foi confirmada por imunofluorescência direta. Bovinos entre 4 meses e 8 anos foram afetados. O curso clínico variou de três a sete dias. A forma paralítica foi a mais frequente e incluiu incoordenação, paresia e paralisia dos membros pélvicos, decúbito, movimentos de pedalagem e morte. Os principais achados histopatológicos foram meningoencefalite linfoplasmocitária associada com corpúsculos de Negri em 86,6 por cento dos casos. Todos os casos foram positivos na imuno-histoquímica para raiva, cujas reações foram mais evidentes no tronco encefálico, incluindo bulbo, ponte e mesencéfalo, além de gânglio trigêmeo. A imuno-histoquímica demonstrou o vírus da raiva em axônios, dendritos e pericário de neurônios, como agregados de grânulos ou em formações arredondadas associadas com números variáveis de corpúsculos de inclusão virais nos neurônios. Houve também marcação nos neurônios de Purkinje e de seus processos na camada molecular, nos núcleos do tronco encefálico e camada profunda do córtex telencefálico. A imuno-histoquímica pode ser importante ferramenta diagnóstica no diagnóstico da raiva, especialmente em situações nas quais não é possível manter refrigeração adequada das amostras e em casos com meningoencefalite não-supurativa e ausência de corpúsculos de inclusão.

This retrospective study included clinical and pathological findings from 15 cattle affected by rabies. Thirteen of the 15 cases were confirmed by direct immunofluorescence. Cattle between 4 months and 8 years of age were affected. Clinical course ranged from 3 to 7 days. Paralytical form was the most common clinical picture and included incoordination, paresis, and paralysis of the pelvic members, besides recumbence, paddling, and death. The main histopathological findings were lymphoplasmacytic meningoencephalitis associated with characteristic Negri bodies in 86.6 percent of the cases. All cases showed anti-rabies immunostaining, which were most prominent in the brainstem including medulla oblongata, pons, and midbrain, besides trigeminal ganglion. Positive labeling was present within axons, dendrites, and perikarya of neurons as aggregates of granules or round formations associated with varying numbers of inclusion bodies. Immunostaining was also observed in the Purkinje neurons and their processes in the molecular layer, in the neurons of of the brainstem, and deep layer of the telencephalic cortex. Immunohistochemistry may be an important auxiliary tool for the diagnosis of rabies, especially in circumstances in which refrigeration cannot be adequately maintained, and in cases characterized by nonsuppurative meningoencephalitis with absence of inclusion bodies.

Expression of serotonin and its 5-HT1A receptor in canine cutaneous mast cell tumours.

Mast cells of a number of different animal species have been reported to contain serotonin (5-hydroxytryptamine; 5-HT), a monoamine capable of numerous and complex actions, which may include an impact on tumour growth. Limited previous studies have suggested that normal or neoplastic canine mast cells do not express 5-HT. In the present study, canine cutaneous mast cell tumours (MCTs) of Patnaik histological grades I-III were investigated immunohistochemically for expression of 5-HT and its receptor (R) 5-HT1A. The proportion of positively labelled cells and the intensity of labelling of individual cells were determined. Both 5-HT and the 5-HT1A receptor were expressed by non-neoplastic dermal mast cells and neoplastic mast cells. More neoplastic mast cells expressed 5-HT than the 5-HT1AR. Poorly differentiated tumours expressed fewer of both molecules, but the better differentiated mast cells at the periphery of such lesions had more consistent 5-HT expression. 5-HT and the 5-HT1A receptor may be involved in the differentiation of canine MCTs and in the microvascular complications associated with these neoplasms.

Cellular phenotypes in the abomasal mucosa and abomasal lymph nodes of goats infected with Haemonchus contortus.

The distribution of T-cell subsets (CD2, CD4, CD8, and gammadelta) and B cells (IgM) was examined at 3, 6, 10 and 13 days post-infection (dpi) in the abomasal mucosa and abomasal lymph nodes of goats primarily infected with Haemonchus contortus. In the abomasal mucosa a mild (3 and 6 dpi) or marked (10 and 13 dpi) increase of T cells, particularly CD4+ and gammadelta+ lymphocytes, was observed, whereas the increase in CD8+ cells was less pronounced. B cells and IgG+ plasma cells also showed a marked increase in the abomasal mucosa at 10 and 13 dpi. The abomasal lymph nodes showed an increase in size, particularly at 10 and 13 dpi, and a decrease in the proportion of T cells, particularly CD8+ lymphocytes, due to the increased proportion of B cells. The proportion of CD4+ and gammadelta+ lymphocytes did not change significantly during the infection in the abomasal lymph nodes, but their absolute numbers were augmented as a result of the enlargement of the nodes. The results revealed a strong cellular and humoral immune response during the early post-infection stages. However, as indicated by the worm burdens, this rapid host response was unable to induce larval expulsion.

Detection of several clostridia by a direct fluorescent antibody test in formalin-fixed, paraffin-embedded tissues

Descreve-se a aplicabilidade de uma técnica de imunofluorescência direta, para o diagnóstico de mionecroses causadas por clostrídios, a partir de tecidos fixados em formol e incluídos em parafina. Essa técnica pode auxiliar no diagnóstico do carbúnculo sintomático e da gangrena gasosa, contribuindo para determinar a real prevalência dessas doenças no país

Steroid receptors in canine and human female genital tract tumours with smooth muscle differentiation.

The expression of oestrogen receptor-alpha (ERalpha) and progesterone receptor (PR) was examined in 32 canine genital tract tumours diagnosed as smooth muscle tumours (benign or malignant, pure or mixed). The immunohistochemical expression of calponin was used to assess the smooth muscle differentiation of the tumours. Nineteen human uterine leiomyomas were also examined. Calponin expression was detected in 89.3% of canine and 100% of human genital tract tumours diagnosed as leiomyomas, as well as in the majority of other tumours examined (canine or human, genital or extragenital, benign or malignant) with the exception of canine negative control tumours (cutaneous fibroma and hepatoid gland adenoma). ERalpha was found in 56.3% of canine and 52.6% of human leiomyomas, while PR was found in 84.4% of canine and 94.7% of human tumours. These results indicate that calponin is a good marker for differentiating neoplasia of the canine genital system of uncertain origin, as in human patients. They also show that canine tumours with smooth muscle differentiation of the genital tract of the bitch express steroid hormone receptors, a finding that opens up the possibility of hormone therapy.

Comparison of parasitological, immunological and molecular methods for the diagnosis of leishmaniasis in dogs with different clinical signs.

Aiming to improve the diagnosis of canine leishmaniasis (CanL) in an endemic area of the Northwest region of São Paulo State, Brazil, the efficacy of parasitological, immunological and molecular diagnostic methods were studied. Dogs with and without clinical signs of the disease and positive for Leishmania, by direct parasite identification on lymph node smears and/or specific antibody detection by ELISA, were selected for the study. According to the clinical signs, 89 dogs attending the Veterinary Hospital of UNESP in Araçatuba (SP, Brazil) were divided into three groups: symptomatic (36%), oligosymptomatic (22%) and asymptomatic (22%). Twenty-six dogs from an area non-endemic for CanL were used as negative controls (20%). Fine-needle aspiration biopsies (FNA) of popliteal lymph nodes were collected and Diff-Quick-stained for optical microscopy. Direct immunofluorescence, immunocytochemistry and parasite DNA amplification by PCR were also performed. After euthanasia, fragments of popliteal lymph nodes, spleen, bone marrow and liver were collected and processed for HE and immunohistochemistry. Parasite detection by both HE and immunohistochemistry was specifically more effective in lymph nodes, when compared with the other organs. Immunolabeling provided higher sensitivity for parasite detection in the tissues. In the symptomatic group, assay sensitivity was 75.61% for direct parasite search on Diff-Quick-stained FNAs, 92.68% for direct immunofluorescence, 92.68% for immunocytochemistry and 100% for PCR; the corresponding values in the other clinical groups were: 32, 60, 76 and 96% (oligosymptomatic), and 39.13, 73.91, 100 and 95.65% (asymptomatic). Results of the control animals from the CanL non-endemic area were all negative, indicating that the methods used were 100% specific.

Prevalence and factors associated with fecal shedding of Giardia spp. in domestic cats.

The prevalence of cats shedding Giardia cysts (13.6%) in the present study was found to be higher than previously reported (1% to 11%) and may reflect a higher sensitivity for the diagnostic test used. The presence of Cryptosporidium spp. oocysts, coccidial oocysts, and a clinical history of chronic (>2 weeks) gastrointestinal signs were significantly associated with the presence of Giardia spp. cysts in the feces. There were no associations between the presence of Giardia spp. cysts and type of housing, acute gastrointestinal signs, vomiting, gender, source of cat (i.e., animal shelter versus private breeder), or gastrointestinal parasites other than Cryptosporidium spp. and intestinal coccidial agents.

Sarcolemma-specific autoantibodies in canine inflammatory myopathy.

Inflammatory myopathies (IM) are relatively common in dogs with an increased incidence in the Boxer and Newfoundland breeds. Here, we show that a high proportion of affected Boxers and Newfoundlands have circulating autoantibodies against unknown sarcolemma antigens, that are muscle-specific but not species specific. We further show that the autoantigen can be extracted from muscle membranes with non-ionic detergent, and that such detergent extracts can be used in a sensitive ELISA for detection and quantitation of antibodies. The relatively high incidence of IM with autoantibodies in selected breeds of dogs indicates a genetic predisposition for a particular form of IM. In these breeds, this form of IM could be diagnosed and monitored with a simple serum assay.