Phylogenetic evidence of the intercontinental circulation of a Canine distemper virus lineage in the Americas.

Canine distemper virus (CDV) is the cause of a multisystem disease in domestic dogs and wild animals, infecting more than 20 carnivore and non-carnivore families and even infecting human cell lines in in vitro conditions. Phylogenetic classification based on the hemagglutinin gene shows 17 lineages with a phylogeographic distribution pattern. In Medellín (Colombia), the lineage South America-3 is considered endemic. Phylogenetic studies conducted in Ecuador using fragment coding for the fusion protein signal peptide (Fsp) characterized a new strain belonging to a different lineage. For understanding the distribution of the South America-3 lineage in the north of the South American continent, we characterized CDV from three Colombian cities (Medellín, Bucaramanga, and Bogotá). Using phylogenetic analysis of the hemagglutinin gene and the Fsp region, we confirmed the circulation of CDV South America-3 in different areas of Colombia. We also described, for the first time to our knowledge, the circulation of a new lineage in Medellín that presents a group monophyletic with strains previously characterized in dogs in Ecuador and in wildlife and domestic dogs in the United States, for which we propose the name "South America/North America-4" due its intercontinental distribution. In conclusion, our results indicated that there are at least four different CDV lineages circulating in domestic dogs in South America: the Europe/South America-1 lineage circulating in Brazil, Uruguay, and Argentina; the South America-2 lineage restricted to Argentina; the South America-3 lineage, which has only been reported in Colombia; and lastly an intercontinental lineage present in Colombia, Ecuador, and the United States, referred to here as the "South America/North America-4" lineage.

Experimental characterization of the complete life cycle of Haemoproteus columbae, with a description of a natural host-parasite system used to study this infection.

Characterization of complete life cycles of haemoparasites requires the maintenance of suitable susceptible vertebrate hosts and vectors for long periods in captivity, in order to follow the complete parasitic cycle in definitive and intermediate hosts. Currently, there are few host-parasite models established in avian haemosporidian research, and those have been developed mainly for species of Passeriformes and their parasites. This study aimed to develop an experimental methodology to access the complete life cycle of Haemoproteus columbae (cytb lineage HAECOL1), which parasitizes the Rock Pigeon (Columba livia) and louse fly (Pseudolynchia canariensis). A colony of louse flies, which are the natural vectors of this parasite, was established. Thirty newly emerged insects were exposed to H. columbae infection and used to infect naïve Rock Pigeons. The peak of parasitaemia (acute stage) was seen between 27 and 32 days p.i. when up to 70.8% of red blood cells were infected. The crisis occurred approximately 1 week after the peak, and the long-lasting chronic parasitaemia stage followed. Exo-erythrocytic meronts were seen mainly in the lungs where extensive tissue damage was reported, but also in the kidneys and spleen. In the vector, the sporogonic cycle of H. columbae was completed between 13 and 16 days p.i., at an average temperature ranging between 12 and 15 °C. This host-parasite model is tractable for maintenance in captivity. It is recommended for use in studies aiming for detailed characterization of host-parasite relationships in areas such as physiology, pathology, immunobiology, genetics, as well as for evaluative treatments and to follow the infection in any stage of parasite development both in the vertebrate or invertebrate host.

Lack of Association Between Epstein-Barr Virus and Mammary Tumours in Dogs.

INTRODUCTION: Epstein-Barr virus (EBV) is a γ-herpesvirus associated with various neoplasms in humans and is a probable aetiological agent in breast cancer; however, a causal relationship has not yet been established. Because of the epidemiological and clinicopathological similarities between breast cancer and canine mammary tumours, dogs have been proposed as a valid model for breast cancer. MATERIAL AND METHODS: A total of 47 canine mammary gland tumour tissues were processed by routine histopathological technique with haematoxylin-eosin staining and classified according to the type of neoplasm. DNA was extracted from paraffin-embedded tissues and the EBNA-1 gene and the BamHI-W region specific for EBV were evaluated by nested PCR. RESULTS: The histopathological evaluation revealed 2 benign neoplasms, and many carcinomas: 2 in situ, 9 simple, 3 solid, 10 complex, and 21 mixed. One sample was positive for the EBNA-1 gene, while all were negative for the BamHI-W region. CONCLUSION: No association was found between EBV and mammary tumours in dogs. However, here we report for the first time the presence of an EBV gene sequence in a canine mammary tumour. It is likely that detection of EBV might be affected by the quality and quantity of DNA extracted from paraffin-embedded tissues. Additional studies are necessary to establish any association of EBV with mammary gland cancer in humans and in dogs, which could eventually lead to better public health prevention and control.

A novel loop-mediated isothermal amplification-based test for detecting Neospora caninum DNA.

BACKGROUND: Neospora caninum is a cyst-forming, coccidian parasite which is known to cause neurological disorders in dogs and abortion and neonatal mortality in cows and other livestock. This study reports the development of a loop-mediated isothermal amplification (LAMP) assay based on the Neospora caninum Nc-5 gene and compares its efficacy for detecting DNA to that of a semi-nested PCR test. RESULTS: Six primers were designed based on the Nc-5 repeat region of N. caninum. Specific LAMP primers led to successful amplification of N. caninum DNA at 63 °C in 30 min. The LAMP assay was highly specific (i.e. it did not reveal cross-reactivity with other parasite species) and had a low N. caninum plasmid DNA limit of detection (1 fg), which is ten times higher than that for the semi-nested PCR. LAMP applicability was evaluated using a set of naturally-infected samples (59 from canine faeces and five from bovine abortions). Thirty-nine percent (25/64) of the naturally-infected samples were positive for N. caninum DNA by LAMP and 36% (23/64) by semi-nested PCR. However, the LAMP assay is much faster to perform than semi-nested PCR and provides results in 30 min. CONCLUSION: The optimized reaction conditions described in this study resulted in a sensitive, specific and rapid technique for detecting N. caninum DNA. Considering the advantages of LAMP for detecting N. caninum DNA, further assays aimed at testing its usefulness on a wider range of field samples are recommended.

Sistema inmuney vacunación de peces / Immune System And Vaccination In Fish

Los peces poseen un sistema inmune con muchas de las células y sustancias humorales presentes en vertebrados superiores, pero cuentan también con componentes y funciones especiales, como los centros melanomacrófagos y la capacidad fagocítica de los enterocitos por citar solo algunas de ellas, que difieren profundamente con sus similares en otras especies y que aún hoy son pobremente comprendidas. Sumado a esto, el ambiente acuático y más, el ambiente acuático productivo, es de por si complejo de manera que las posibilidades de las interacciones biológicas son enormes y los procesos difícilmente predecibles. La vacunación es tal vez una de las herramientas más importantes para el control de enfermedades bacterianas en peces, no solo por su potencial preventivo y correctivo sino también por sus bondades con el ambiente y con la salud pública que contrastan notoriamente con los tratamientos antibióticos. El éxito de las vacunas en especies piscícolas depende en buena medida del conocimiento adecuado del sistema inmunológico de los peces, de las interacciones hospedero-ambiente-patógeno, así como de las particularidades de los sistemas productivos. Con todo, la vacunación en ambientes acuáticos se enfrenta a un sinnúmero de dificultades que deben ser abordadas antes que con un enfoque unilateral farmacológico, con una aproximación integral en la que se incluye el uso de herramientas epidemiológicas, clínicas, patológicas, microbiológicas, etc.

The fish immune system posses many of the cells and molecules found in higher vertebrates, but several other components and functions such as the melano-macrophage centers and the phagocityc activity of the epithelial enteric cells which are not found in higher vertebrates are described in fish, though their function is poorly understood. In adition, the aquatic environment, and even most, the productive systems are very complex, so that, the possibilities of the biological interactions are enormous and the biological processes prediction, difficult. Vaccination is perhaps one the most important tools for fish bacterial diseases control, no just for its preventive and corrective potential but also for its beneficial environmental and public health effects which contrast with those of the antibiotic therapy. A great deal of the success of a vaccine in piscine species depends on fish immune system understanding, on the host-environment-pathogen interactions, but also on the productive system particularities. Despite all these considerations, the vaccination in aquatic systems must be approached bearing in mind an integral perspective, where epidemiological, clinical, pathological and microbiological, etc. tools should be taken into account and not only the pharmacological therapy view.

Mycobacterium tuberculosis ESAT-6 antigen immunogenicity in Owl Monkeys

La única vacuna disponible contra la tuberculosis es la cepa Mycobacterium bovis BCG, que ofrece una eficacia protectiva variable (0/100-80/100), siendo urgente un nuevo agente profiláctico. Se han evaluado diversos candidatos a vacuna contra este patógeno, en los modelos animales de experimentación convencionales (murino, cobayo, conejo), obteniéndose información básica sobre el efecto de la vacuna en la carga bacterial frente a un reto infeccioso, así como también la reducción o prevención de la patología en los pulmones u otros órganos blanco; además de los aspectos relacionados con la respuesta inmune hacia el Mycobacterium tuberculosis. Los primates no humanos tienen ventajas sobre los modelos convencionales en la evaluación de vacunas, de hecho se ha verificado el comportamiento de agentes terapéuticos en humanos después de haber sido medida la capacidad protectiva de éstos en monos con tuberculosis inducida. Los primates mas estudiados en la infección por micobacterias son el cynomulgus, y el rhesus, observándose que estos animales mantienen la infección en un estado subclínico, muy similar a la tuberculosis humana donde el 90/100 de la población infectada mantiene la infección en un estado latente. Dado que el modelo animal debe semejar el comportamiento de las proteínas estudiadas en el ser humano, el mono Aotus puede representar ventajas en la investigación de tuberculosis por ser un primate con aproximadamente un 90/100 de similitud al humano en cuanto a las moléculas del sistema inmune estudiadas hasta hoy. La proteína ESAT-6 de (early secretory antigenic target 6 kD) de Mycobacterium tuberculosis es un componente minoritario del filtrado de cultivo de corto tiempo (CFP), ha sido genética y químicamente caracterizada e induce una potente respuestainmunogénica del tipo TH1. Este antígeno es secretado durante la fase inicial de crecimiento siendo fuertementereconocido por animales y humanos infectados por Mycobacterium tuberculosis...

Cutaneous tuberculosis diagnosis in an inhospitable Amazonian region by means of telemedicine and molecular biology.

We report on a 13-year-old boy who displayed a chronic granulomatous inflammatory reaction of 5 years duration. The lesion was resistant to different antibiotic schemes; his routine laboratory tests and chest radiographs were normal. Teledermatologic consultation and histopathologic study of skin biopsy suggested scrofulodermal tuberculosis. Polymerase chain reaction amplification of DNA extracted from lymph node biopsy was taken as starting material for dot-blot hybridization using Mtp-40 and IS 6110 as probes for detecting either Mycobacterium tuberculosis or any mycobacteria belonging to the M tuberculosis complex, respectively. Positive results in both hybridizations were further confirmed by culturing in BACTEC MGIT 960 system. The lesion greatly diminished following isoniazid, rifampin, and ethambutol treatment. Telemedicine allowed a cutaneous tuberculosis diagnosis to be made of a patient living in a remote town located in the Amazon jungle by using molecular biology techniques.

Identifying and structurally characterizing CD1b in Aotus nancymaae owl monkeys.

This study reports the molecular characterization and tissue expression of the non-human Aotus nancymaae primate CD1b isoform in the search for an experimental animal model to be used in evaluating the role of non-peptide antigen-presentation molecules in the immune response to infectious agents. CD1b expression on the surface of A. nancymaae peripheral blood monocyte-derived dendritic cells, shown by flow cytometry, was made possible by using human CD1b isoform antibodies. Studying the expression of CD1b-encoded transcripts revealed this molecule's broad distribution in several tissues. The A. nancymaae CD1b transcript-encoded amino-acid sequence showed 95.5% identity with the human sequence. Such high sequence homology was reflected in the identical structural conservation of how pockets A', C' and F' and tunnel T' conforming the antigen's binding site are organized, the similar arrangement of those amino-acids interacting with the T-cell receptor (TCR) during antigen presentation, and the conservation of YQNI-motif sequence in the cytoplasmatic tail (responsible for the molecule's intracellular trafficking in humans). Comparing the structure of human CD1a and CD1b and mouse CD1d proteins with CD1b structure in A. nancymaae obtained by minimization revealed that changes in the latter molecule's alpha1 and alpha2 domains imposed a narrowing of the antigen-binding groove in A. nancymaae CD1b. The high structural similarity between A. nancymaae CD1b and that from humans presented in this study leads to A. nancymaae being proposed as a suitable experimental animal model for analyzing CD1b in vivo, mainly in bacterial and parasite infections such as tuberculosis and malaria, respectively.