An atypical clinical presentation for the first isolation of canid herpesvirus 1 in Argentina / Uma apresentação clínica atípica para o primeiro isolamento do herpesvirus canino 1 na Argentina

Relatam-se o primeiro isolamento de herpesvirus canino 1 (CaHV-1) e a localização atípica das lesões vesiculares associadas a este vírus na Argentina. A amostra foi recuperada de lesões vesiculares, localizadas na parte interna da coxa direita, em uma fêmea de raça Labrador. A cadela tinha quatro anos de idade e era de propriedade privada. O primeiro diagnóstico foi realizado pela reação em cadeia da polimerase e, posteriormente, o vírus foi isolado e sua identificação confirmada por imunofluorescência indireta e pelo teste de neutralização viral.

Evaluation of neutralization patterns of the five unique Argentine equine arteritis virus field strains reported / Evaluación de los patrones de neutralización de las únicas cinco cepas argentinas descritas de arteritis viral equina

Equine viral arteritis (EVA) is a contagious viral disease that frequently causes mild or subclinical infections in adult horses. Only one EAV serotype has been described. However, there are differences in antigenicity, pathogenicity and neutralization characteristics of virus field strains. The interaction of two viral proteins, GP5 and M, is critical for infectivity and amino acid changes in the GP5 sequences have an effect on the neutralizing phenotype, regardless the effects of other viral proteins. The objective of the present study was to evaluate the neutralization phenotypes of the 5 unique Argentine EAV strains reported and to compare them with the neutralization phenotypes of the EAV-UCD reference strain, with special emphasis on the analysis of M and GP5 proteins. The strains had a similar neutralization phenotype pattern when anti-EAV serum, derived from EAV seropositive horses, was used in the analysis. Meanwhile, low titers were observed when equine polyclonal anti-EAV reference sera were used in the assay. Argentine strains have almost the same amino acid substitutions, with the exception of LP01 strain, that mainly involves the first variable region V1, especially in neutralization sites B and C. However, they are fairly different from the EAV-UCD strain. Nevertheless, the nucleotide and amino acid differences observed among the Argentine strains LP02/R, LP02/C, LP02/P and LP-LT-ARG did not show any variations in the neutralization phenotype.

La arteritis viral equina (AVE) ocasiona infecciones, en su mayoría subclínicas, pero puede causar abortos y enfermedad respiratoria. Si bien se ha descrito un solo serotipo de AVE, existen diferencias en cuanto a la antigenicidad, patogenicidad y patrones de neutralización en las cepas de campo. Los ORF5 y ORF6 del virus codifican las proteínas de envoltura GP5 y M; la interacción entre estas proteínas es crítica para la infectividad. Los cambios en las secuencias de aminoácidos en la proteína GP5, especialmente en la región V1, afectan el fenotipo neutralizante, sin tener en cuenta variaciones aminoacídicas de otras proteínas virales. En este estudio evaluamos los fenotipos neutralizantes de las 5 únicas cepas de arteritis viral equina aisladas en Argentina y los comparamos con los de la cepa de referencia EAV-UCD por virus neutralización cruzada y análisis de secuencias aminoacídicas de las proteínas M y GP5. Las cepas argentinas presentaron un patrón de neutralización similar cuando se utilizaron sueros positivos del banco de sueros, mientras que fueron neutralizadas en menor medida por los sueros policlonales de referencia anti-AVE. A excepción de la cepa LP01, las cepas argentinas tienen casi las mismas sustituciones aminoacídicas en la primera región variable V1 de la proteína GP5, específicamente en los sitios neutralizantes B y C, pero difieren en gran medida respecto de la cepa de referencia EAV-UCD. Las diferencias encontradas en los aislamientos LP02/R, LP02/C, LP02/P y LT-LP-ARG no se reflejaron en variaciones en el fenotipo neutralizante.

Aplicación de PCR-RFLP para subtipificar Campylobacter jejuni / [PCR-RFLP for Campylobacter jejuni subtyping]

Ten Campylobacter jejuni isolates, 8 identified as C. jejuni biotype II of Lior and 2 as C. jejuni biotipe I, were recovered from aborted pig fetuses. In order to discriminate among strains, restriction fragment length polymorphism (RFLP) using Ddel of polymerase chain reaction (PCR) products of flaA gen was used. C. jejuni biotype II strains could be diferenciated in 6 by PCR-RFLP, and one subtype was obtained from C. jejunibiotype I. Although there is great variability of molecular techniques applied to the Campylobacter epidemiological studies, PCR-RFLP demonstrated to be a simple and accessible technique to discriminate Campylobacter jejuni isolates.

Campylobacter jejuni y C. coli en suinos abortados: comparación entre la identificación fenotípica y los perfiles proteicos en geles de poliacrilamida / [Campylobacter jejuni and C. coli in aborted swine: comparison between phenotypic identification and polyacrylamide gel protein profiles]

Campylobacter jejuni and Campylobacter coli were isolated from aborted pig fetuses which proceeded from different animals and farms between February 2000 and March 2001. Seven Campylobacter jejuni biotype II, three biotype I and one Campylobacter coli biotype I were identified by phenotypic tests and Lior’s scheme. To corroborate and compare the phenotypic results, 7.5, 10 and 12.5

polyacrilamide gel electrophoresis (SDS-PAGE) were used under reducing conditions. Characteristic bands of hypervariable dense zone within C. jejuni and C. coli species were observed in all the whole cell protein extracts with differences in mobility. It was possible to establish differences between identical phenotypic Campylobacter isolates and different protein profile from fetuses of the same litter. SDS-PAGE is a stable and reproducible method to establish differences between Campylobacter strains and is considered applicable for the differentiation of the wide variability of Campylobacter species for epidemiologic purposes.

Rapid diagnosis of pseudorabies virus infection in swine tissues using the polymerase chain reaction (PCR)

In Argentina pseudorabies is an endemic disease. Routine diagnosis is made by virus isolation. It is a very long procedure to carry out and gives variable results depending on the quality of sample, hence the need for effective techniques, which are rapid and not dependent on the isolation of infectious virus. The polymerase chain reaction (PCR) technique has provided a sensitive, specific and rapid mean to detect DNA sequences. This study describes a PCR method for detection of pseudorabies virus sequences in swine tissues. In order to determine the presence of suid herpesvirus-1 DNA and antigens, 36 tissue samples collected from 19 dead pigs, with signs of pseudorabies infection, were examined by PCR, virus isolation and indirect immunofluorescence, respectively. Fifteen out of 19 pigs were positive at least for one tissue by PCR (15/19) while only three pseudorabies virus strains were isolated (3/19). All the amplified products were identified by digestion with Sa/l and hybridization. The method described herein circumvents tedious viral isolation and DNA purification and would be a valuable tool for rapid diagnosis, since it would take less than 5 h to reach an accurate result even in poorly preserved tissue samples.

ELISA indirecto para el diagnóstico rápido de Influenza Equina / Indirect ELISA for the rapid diagnosis of Equine Influenza

An indirect enzyme linked immunosorbent assay was developed. Infected and non infected allantoic fluids precipitated with polyetilenglycol 6000 were used as antigen and control antigen, respectively. Serum samples were diluted 1/20 and a commercial horse radish peroxidase-labelled rabbit anti-equine IgG was used as second antibody. The reaction was developed using azino-diethylbenzotyazol-sulfonate (ABTS). Cut-off was determined by ratio sample (Rs). The hemagglutination inhibition test was used as a reference test for the 391 samples analyzed. Of these, 301 sera were positive by hemagglutination inhibition test and indirect ELISA, 75 were negative by both techniques, and 15 were positive by indirect ELISA and negative by hemagglutination inhibition test. Using hemagglutination inhibition test as standard, the indirect ELISA showed a relative specificity and sensitivity of 83.3 and 100, respectively. This indirect ELISA is useful as screening test.

Avaliaçäo da técnica de western blot no diagnóstico da leucose enzoótica bovina / Evaluation of western blotting for the diagnosis of enzootic bovine leukemia

Um sistema de western blotting (WB) foi desenvolvido para detecçäo de anticorpos contra o vírus da leucose em soros bovinos. Foram utilizadas amostras de soros de 233 animais naturalmente infectados e soros de vacas experimentalmente infectadas. O teste de imunodifusäo em ágar (AGID) foi usado para comparaçäo dos resultados. Graus diferentes de reatividade foram observados em soros positivos ao AGID, quando testados em WB frente a um ou aos dois antígenos mais importantes (gp51 e p24). Outras proteínas (gp30, p15, p12 e p10) näo foram detectadas por nenhum soro positivo ao AGID, sendo que três bandas correspondentes à proteína p24 foram observadas ocasionalmente. Em soros obtidos por inoculaçäo experimental, anticorpos contra a proteína p24 foram detectados entre a segunda e a quarta semanas após a inoculaçäo e, em alguns animais, detectaram-se anticorpos anti-gp51 mais tardiamente. O estudo de soros de campo com AGID e WB mostrou concordância de 90,9 por cento sendo que apenas 1,7 por cento dos soros negativos pelo AGID foram positivos ao WB e 7,2 por cento dos resultados näo conclusivos por AGID foram definidos por WB (4,2 por cento como positivos e 3 por cento como negativos)

Diversity of genomic electropherotypes of naturally occurring equine herpesvirus 1 isolates in Argentina

The genomes of 10 equine herpesvirus 1 (EHV-1) strains isolated in Argentina from 1979 to 1991, and a Japanese HH1 reference strain were compared by restriction endonuclease analysis. Two restriction enzymes, Bam HI and Bgl II, were used and analysis of the electropherotypes did not show significant differences among isolates obtained from horses with different clinical signs. This suggests that the EHV-1 isolates studied, which circulated in Argentina for more than 10 years, belong to a single genotype