Experimental Frog Virus 3 infection using Brazilian strain: amphibians susceptibility / Infecção experimental utilizando estirpe brasileira de Frog Virus 3: suceptibilidade de anfíbios

An alarming number of global warnings concerning amphibian mortality outbreaks have been released in recent years. Emerging diseases stand out as the main potential causes. Ranavirus is a worldwide-spread highly infectious disease capable of affecting even other ectothermic animals such as fish and reptiles. One major issue regarding this pathology is the lack of clinical signs before it leads up to death. Aiming at having a better understanding of anurans susceptibility, this study analyzed bullfrog (Lithobates catesbeianus) survival rate, when challenged with three doses of a Brazilian strain of Frog Virus 3 (FV3). The qPCR analysis indicated a low infectivity rate in these animals both as larvae and as adults. To elucidate the results, the following hypothesis was performed: 1) The amount of inoculum used on the frogs was insufficient to trigger an infection; 2) For the FV3 to produce clinical signs in this species, there is the need for a cofactor; 3) The animals did undergo FV3 infection but recovered in the course of the experiment, and 4) The inoculum utilized might have been low-virulence. Finally, the presence of actual clinical signs of ranavirus is discussed, with the more likely hypothesis.(AU)

Um número alarmante de notificações globais sobre surtos de mortalidade de anfíbios tem sido realizado nos últimos anos. As doenças emergentes destacam-se como as principais causas potenciais. O ranavírus é uma doença altamente infecciosa disseminada em todo o mundo, capaz de afetar até outros animais ectotérmicos como peixes e répteis. Uma questão importante em relação a essa patologia é a falta de sinais clínicos antes de levar à morte. Com o objetivo de compreender melhor a suscetibilidade dos anuros, o presente trabalho analisou a taxa de sobrevivência de rãs-touro (Lithobates catesbeianus), desafiadas com três doses de uma estirpe brasileira do Frog virus 3 (FV3). A análise de qPCR indicou baixa taxa de infectividade nesses animais, tanto como larvas quanto como adultos. Procurando esclarecer os resultados, foram formuladas as seguintes hipóteses: 1) A quantidade de inóculo aplicada nas rãs foi insuficiente para desencadear uma infecção; 2) Para que o FV3 dê sinais clínicos nesta espécie, é necessário um cofator; 3) Os animais sofreram infecção por FV3, mas se recuperaram no decorrer do experimento, e 4) O inóculo utilizado pode ter sido de baixa virulência. Finalmente, foi discutida a presença de sinais clínicos reais de ranavírus e levantada a hipótese mais provável(AU)

Characterization of an iridovirus detected in rock bream (Oplegnathus fasciatus; Temminck and Schlegel) / 病毒学报

During the summer of 2009, mass mortality was observed in cage-cultured Rock Bream (Oplegnathus fasciatus; Temminck and Schlegel) in the Liaoning Province. Histopathogic studies of the affected fish showed enlarged basophilic cells in the kidney and spleen. These necrotic cells were stained purple using haematoxylin and eosin (HE). GF cell cultures showed advanced cytopathic effects after infection with virus supernatants from diseased fish homogenate. Transmission electron microscopy revealed hexagonal outlines virions in the cytoplasm of the spleen, kidney, liver, intestine cells. The viral particles consisted of a central nucleocapsid (100-110 nm) and envelope, and were 150-180 nm in diameter. These results suggested that the virus belonged to the Iridoviridae. Using polymerase chain reaction (PCR), approximately 570-bp fragments were amplified from the viral DNA in spleen, kidney, gill, intestine, heart and brain of diseased fish with the primers derived from red sea bream Iridovirus (RSIV). In addition, a specific fragment of 1 400 bp of the major capsid protein (MCP) gene of the Iridovirus was amplified by PCR. A phylogenetic tree was constructed to compare the corresponding genetic sequences in Megalocytivirus. The tree demonstrated that RSIV-LN09 virus existed in the same branch as the RSIV-U1 et al. Our present results indicated that RSIV was the causative agent.

Isolation and characterization of an iridovirus from sick giant salamander (Andrias davidianus) / 病毒学报

A virus was isolated from cultured sick giant salmander (Andrias davidianus ) in a farm, Shanxi Province, China. Skin ulceration and necrosis of the distal limbs are main clinical symptoms. Virus propagated and caused CPE at 10 degrees C to 30 degrees C in BF-2, CO, CHSE, FHM cells. The optimum condition of replication was in BF-2 cells at 25 degrees C. The virus was proved to be senstive to chloroform, heat, pH3 and pH10 treatment. Viral replication was inhibited by 5-Fluoro-2-deoxyuridine (FUDR). These results indicated that the virus possessed an envelope and DNA as the genome. Electron-microscopic observation of thin-section showed numerous hexagonal viral particles measuring 130 nm to 150 nm in diameter orderly arranged in a lattice form in cytoplasm of BF-2 cells. The particles showed typical iridovirus morphology. A 413 bp fragment was amplified from the viral main capsid protein gene by PCR. The fragments was sequenced and analysed. The results showed the isolate shared more than 96% nucleotide identity with some Ranaviruses. We suggested that this virus was named as Andrias davidianus iridovirus (ADIV) tentatively.

Natural invertebrate hosts of iridoviruses (Iridoviridae) / Hospederos naturales de los iridovirus de invertebrados

Invertebrate iridescent viruses (IIVs) are icosahedral DNA viruses that infect invertebrates, mainly insects and terrestrial isopods, in damp and aquatic habitats. Exhaustive searches of databases resulted in the identification of 79 articles reporting 108 invertebrate species naturally infected by confirmed or putative iridoviruses. Of these, 103 (95%) were arthropods and the remainder were molluscs, an annelid worm and a nematode. Nine species were from marine habitats. Of the 99 non-marine species, 49 were from terrestrial habitats and 50 were aquatic, especially the aquatic stages of Diptera (44 species). The abundance of records from species of Aedes,Ochlerotatus and Psorophora contrasts markedly with a paucity of records from species of Anopheles,Culex and Culiseta. Records from terrestrial isopods are numerous (19 species), although the diversity of IIVs that infect them is mostly unstudied. IIV infections have been reported from every continent, except Antarctica, but there are few records from Africa, southern Asia and Latin America. Most reports describe patent IIV infections as rare whereas inapparent (covert) infection may be common in certain species. The relationship between particle size and iridescent colour of the host is found to be consistent with optical theory in the great majority of cases. Only 24 reported IIVs from insect hosts have partial characterization data and only two have been subjected to complete genome sequencing. I show that the rate of publication on IIVs has slowed from 1990 to the present, and I draw a number of conclusions and suggestions from the host list and make recommendations for future research efforts.

Los virus iridiscentes de invertebrados (VIIs) son virus icosaedrales de ADN que infectan a invertebrados, principalmente insectos e isópodos terrestres en hábitats húmedos y acuáticos. Búsquedas extensivas de bases de datos resultaron en la identificación de 79 artículos científicos, los cuales reportaron 108 especies de invertebrados infectados naturalmente por iridovirus. De estos, 103 (95%) fueron artrópodos y los otros fueron moluscos, un anélido y un nematodo. Nueve especies fueron de hábitats marinos. De las 99 especies no marinas, 49 fueron terrestres y 50 fueron acuáticas, especialmente los estadios acuáticos de dípteros (44 especies). La abundancia de infecciones en especies de Aedes,Ochlerotatus y Psorophora se contrasta marcadamente con la escasez de casos en especies de Anopheles,Culex y Culiseta. Reportes de infecciones de los isópodos terrestres son numerosos (19 especies), aunque la diversidad de los VII que los infectan es desconocida. Se han reportado infecciones por VIIs de todos los continentes, excepto Antártica, pero se notan pocos ejemplos de África, Asia y Latinoamérica. La mayoría de los artículos señala que las infecciones patentes son poco comunes, mientras que las infecciones enmascaradas (subletales) pueden ser comunes en algunas especies. La relación entre el tamaño de la partícula y el color iridiscente concuerda con la teoría óptica en casi todos los casos. Veinticuatro de los VIIs de insectos han sido caracterizados parcialmente y solo dos de éstos han sido secuenciados completamente. Demuestro que el ritmo de publicación sobre los VIIs ha disminuido en los últimos 15 años, señalo varias conclusiones y sugerencias de la lista de especies de huéspedes y presento algunas recomendaciones para la investigación futura con este grupo de patógenos.

Production of polyclonal antibody against recombinant ORF 112 L of rock bream (Oplegnathus fasciatus) iridovirus (RBIV) and in vitro neutralization.

Rock bream iridovirus (RBIV) is a causative agent of epizootics among cultured rock bream (Oplegnathus fasciatus) in Korea. In this study, the immunogenic property of ankyrin repeats gene (ORF 112L) from RBIV was evaluated to develop vaccines against RBIV. ORF 112L of RBIV was cloned into expression vector of pGEX-4T-1. The recombinant protein was successfully expressed using E. coli BL21 (DE3). The soluble recombinant RBIV protein was applied to affinity column for the purification of the protein. Mice were immunized by the injection of purified recombinant protein to produce polyclonal antibodies. EUSA was carried out to identify the immune reaction abilities of polyclonal antibody to recombinant protein. The antigenic property of this protein was evaluated by using in vitro neutralization with BF-2 cells. In neutralization test, BF-2 cells infected with the mixture of RBIV and antisera containing anti-GST-ORF 112L polyclonal antibody were healthy showing few cytopathic effect (CPE) similar with the negative control (without RBIV). These studies suggest that the protein from the ankyrin repeats gene, ORF 112L of RBIV may play an important role in the mechanism of infection. Also, it can be possible to develop protein or gene vaccines using ORF 112L against RBIV.

Linfoquistosis en peces tetra fantasía (parambassis baculis) de la ciudad de México / Lymphocystis in parambassis baculis in Mexico City

En el presente estudio se informa sobre la presencia de linfoquistosis en peces tetra fantasía (Parambassis baculis) de la ciudad de México. En la revisión macroscópica se observaron tumoraciones de 0.1-0.4 cm de diámetro en las aletas dorsales y opérculos. En el resto de los órganos y tejidos no se observaron lesiones. En histopatología y ultraestructura de las lesiones, se encontraron fibroblastos hipertrofiados con gran cantidad de partículas virales de linfoquistosis, de 200-300 nm de diámetro. Se deberán realizar otros estudios para determinar la prevalencia de linfoquistosis en peces comerciales y ornamentales en México, así como el impacto económico y sanitario en la acuicultura, ya que no se encuentran informes de esta enfermedad en México.