Identificação morfológica de organismos semelhantes à anaplasmataceae em monócitos de surubim híbrido (Pseudoplatystoma reticulatum x P. corruscans) – relato de caso

The aim of this study was to report the occurrence of Anaplasmataceae-like organisms in monocytes from the hybrid surubim catfish. During the hematological evaluation of fish infected by Pseudomonas sp. in a fish farm located in the State of Mato Grosso do Sul we observed into the monocytes the presence of numerous pleomorphic inclusions of various sizes, rough in appearance which presented basophilic staining. These inclusions were similar to elementary bodies, initial bodies and morule of the bacteria from Anaplasmataceae family, often diagnosed in domestic mammals. This is the first report of its occurrence possibly belonging to the family of Anaplasmataceae in cultured fish in Brazil. Additional studies are necessary for molecular characterization of this bacteria, pathogenic potential, life cycle and impact on the intensive production of fish.

Molecular diagnosis of Anaplasmataceae organisms in dogs with clinical and microscopical signs of ehrlichiosis

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

Prevalence and molecular characterization of Anaplasmataceae agents in free-ranging Brazilian marsh deer (Blastocerus dichotomus)

Anaplasmataceae organisms comprise a group of obligate intracellular gram-negative, tick-borne bacteria that can infect both animals and humans. In the present work we investigate the presence of Ehrlichia, Anaplasrna, and Neorickettsia species in blood samples from Brazilian marsh deer (Blastocerus dichotomus), using both molecular and serologic techniques. Blood was collected from 143 deer captured along floodplains of the Parana River, near the Porto Primavera hydroelectric power plant. Before and after flooding, marsh deer were captured for a wide range research program under the financial support of São Paulo State Energy Company (CESP), between 1998 and 2001. Samples were divided into four groups according to time and location of capture and named MS01 (n = 99), MS02 (n = 18) (Mato Grosso do Sul, before and after flooding, respectively), PX (n = 9; Peixe River, after flooding), and AGUA (n = 17; Aguapei River, after flooding). The seroprevalences for Ehrlichia chaffeensis and Anaplasma phagocytophilum were 76.76% and 20.2% in MS01, 88.88% and 5.55% in MS02, 88.88% and 22.22% in PX, and 94.12% and 5.88% in AGUA, respectively. Sixty-one animals (42.65% of the total population) were PCR-positive for E. chaffeensis PCR (100.0% identity based on 16S rRNA, dsb, and groESL genes). Seventy deer (48.95% of the total population) were PCR-positive for Anaplasma spp. (99.0% of identity with A. platys, and in the same clade as A. phagocytophilum, A. bovis, and A. platys based on 16S rRNA phylogenetic analysis). Our results demonstrate that Brazilian marsh deer are exposed to E. chaffeensis and Anaplasma spp. and may act as reservoirs for these rickettsial agents, playing a role in disease transmission to humans and other animals. (c) 2012 Elsevier Ltd. All rights reserved.

Identificação e Prevalência de Agentes Rickettsiais (Rickettsiales: Anaplasmataceae) em Cervo-do-pantanal (Blastocerus dichotomus) utilizando métodos sorológico e molecular

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Detecção sorológica e caracterização moleculares de agentes anaplasmataceae, micoplasmas hemotróficos, piroplasmas e Hepatozoon sp. em carnívoros selvagens mantidos em cativeiro no Brasil

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

Caracterização molecular de espécies da família Anaplasmataceae em leucócitos e plaquetas de cães de Jaboticabal-SP e de Campo Grande-MS

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

Identificação morfológica de organismos semelhantes à anaplasmataceae em monócitos de surubim híbrido (Pseudoplatystoma reticulatum x P. corruscans) – relato de caso

The aim of this study was to report the occurrence of Anaplasmataceae-like organisms in monocytes from the hybrid surubim catfish. During the hematological evaluation of fish infected by Pseudomonas sp. in a fish farm located in the State of Mato Grosso do Sul we observed into the monocytes the presence of numerous pleomorphic inclusions of various sizes, rough in appearance which presented basophilic staining. These inclusions were similar to elementary bodies, initial bodies and morule of the bacteria from Anaplasmataceae family, often diagnosed in domestic mammals. This is the first report of its occurrence possibly belonging to the family of Anaplasmataceae in cultured fish in Brazil. Additional studies are necessary for molecular characterization of this bacteria, pathogenic potential, life cycle and impact on the intensive production of fish.

Differential expression of genes in salivary glands of male Rhipicephalus (Boophilus)microplus in response to infection with Anaplasma marginale

Background: Bovine anaplasmosis, caused by the rickettsial tick-borne pathogen Anaplasma marginale (Rickettsiales: Anaplasmataceae), is vectored by Rhipicephalus (Boophilus) microplus in many tropical and subtropical regions of the world. A. marginale undergoes a complex developmental cycle in ticks which results in infection of salivary glands from where the pathogen is transmitted to cattle. In previous studies, we reported modification of gene expression in Dermacentor variabilis and cultured Ixodes scapularis tick cells in response to infection with A. marginale. In these studies, we extended these findings by use of a functional genomics approach to identify genes differentially expressed in R. microplus male salivary glands in response to A. marginale infection. Additionally, a R. microplus-derived cell line, BME26, was used for the first time to also study tick cell gene expression in response to A. marginale infection. Results: Suppression subtractive hybridization libraries were constructed from infected and uninfected ticks and used to identify genes differentially expressed in male R. microplus salivary glands infected with A. marginale. A total of 279 ESTs were identified as candidate differentially expressed genes. Of these, five genes encoding for putative histamine-binding protein (22Hbp), von Willebrand factor (94Will), flagelliform silk protein (100Silk), Kunitz-like protease inhibitor precursor (108Kunz) and proline-rich protein BstNI subfamily 3 precursor (7BstNI3) were confirmed by real-time RT-PCR to be down-regulated in tick salivary glands infected with A. marginale. The impact of selected tick genes on A. marginale infections in tick salivary glands and BME26 cells was characterized by RNA interference. Silencing of the gene encoding for putative flagelliform silk protein (100Silk) resulted in reduced A. marginale infection in both tick salivary glands and cultured BME26 cells, while silencing of the gene encoding for subolesin (4D8) significantly reduced infection only in cultured BME26 cells. The knockdown of the gene encoding for putative metallothionein (93 Meth), significantly up-regulated in infected cultured BME26 cells, resulted in higher A. marginale infection levels in tick cells. Conclusions: Characterization of differential gene expression in salivary glands of R. microplus in response to A. marginale infection expands our understanding of the molecular mechanisms at the tick-pathogen interface. Functional studies suggested that differentially expressed genes encoding for subolesin, putative von Willebrand factor and flagelliform silk protein could play a role in A. marginale infection and multiplication in ticks. These tick genes found to be functionally relevant for tick-pathogen interactions will likely be candidates for development of vaccines designed for control of both ticks and tick-borne pathogens.

First isolation and molecular characterization of Ehrlichia canis in Costa Rica, Central America

The present study investigated Ehrlichia species in blood samples from dogs suspected of clinical ehrlichiosis, using molecular and isolation techniques in cell culture. From a total of 310 canine blood samples analyzed by 16S rRNA nested PCR, 148 (47.7%) were positive for Ehrlichia canis. DNA from Ehrlichia chaffeensis or Ehrlichia ewingii was not detected in any sample using species-specific primers in separated reactions. Leukocytes from five PCR-positive dogs were inoculated into DH82 cells; successful isolation of E. canis was obtained in four samples. Partial sequence of the dsb gene of eight canine blood samples (including the five samples for in vitro isolation) was obtained by PCR and their analyses through BLAST showed 100% of identity with the corresponding sequence of E. canis in GenBank. This study represents the first molecular diagnosis, isolation, and molecular characterization of E. canis in dogs from Costa Rica. (C) 2010 Elsevier Ltd. All rights reserved.

Ehrlichia canis in dogs attended in a veterinary hospital from Botucatu, Sao Paulo State, Brazil

This study investigated the etiology of canine ehrlichiosis and possible clinical and epidemiological data associated with the infection in 70 dogs suspect of ehrlichiosis attended at the Veterinary Hospital of the Sao Paulo State University in Botucatu city during 2001 and 2002. Dogs were evaluated by clinical-epidemiological and hematological data and molecular analysis by partial amplification and DNA sequencing of the ehrlichial dsb gene. E. canes DNA was amplified and sequenced in 28 (40.0%) dogs. Dogs younger than 12 months old showed significantly higher infection rates (65.0%; P < 0.05). Diarrhea, apathy, and anorexia were the major clinical signs observed in 55.2% (P = 0.05), 47.0% (P > 0.05), and 42.4% (P > 0.05) of the PCR-positive dogs, respectively. Twenty-five anemic (<5.5 x 10(6) RBC.mu L(-1)), and 8 leukopenic (<5.5 x 10(3) WBC.mu L(-1)) dogs were PCR-positive (P > 0.05). All 28 PCR-positive dogs showed thrombocytopenia (<175 x 10(3) platelets.mu L(-1)) and revealed statistical significance (P < 0.05). E. canis was the only Ehrlichia species found in dogs in the studied region, with higher infection rates in younger dogs, and statistically associated with thrombocytopenia.

Migratory and Carnivorous Birds in Brazil: Reservoirs for Anaplasma and Ehrlichia Species?

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Molecular and serological detection of Ehrlichia canis and Babesia vogeli in dogs in Colombia

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

Hemoparasite and hematological parameters in Nile tilapia

The accelerated growth of finfish aquaculture has resulted in a series of health problems, including blood disorders by hemoparasites; there are scarce studies about these agents and their impact in actual intensive farming. The aim of this study was to identify and quantify the hemoparasites present in monocytes and erythrocytes in the blood of tilapia and their correlation with the hematological profile. Blood samples were collected from caudal vessels of 15 cage-reared Nile tilapia (Oreochromis niloticus), with 70 ± 10 g weight on average, from Itambaracá Municipality, Parana State, Brazil. The total red blood cells, mean corpuscular volume, hematocrit, total and differential white blood cells counts, and the number of thrombocytes were determined in blood smears stained with May-Grünwald-Giemsa-Wright and Quick Panoptic. The results showed the presence of pleomorphic cytoplasmic inclusions with corrugated appearance and basophilic staining, mainly in monocytes, suggesting Anaplasmataceae parasitemia and inclusions with the same morphological characteristics in erythrocytes of one Nile tilapia. The hematological analysis showed no significant difference (P < 0.05) between infected and not infected fish, and therefore, there was no correlation between parasitemia and hematological profile. These observations allow us to infer that the intracytoplasmic inclusions in monocytes and erythrocytes are compatible with the family Anaplasmataceae. There was no correlation between the blood profile and low level of parasitemia. © 2012 Springer-Verlag London.

Arthropod-borne pathogens circulating in free-roaming domestic cats in a zoo environment in Brazil

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Diagnóstico sorológico e molecular de agentes transmitidos por artrópodes em aves carnívoras

Pós-graduação em Medicina Veterinária - FCAV