ARCHIVES OF RAZI INSTITUTE
Year:2007 | Volume:62 | Issue:4|Papers Count:8

Avian Influenza (AI) is a viral respiratory disease of domestic and wild birds. In the diagnostic laboratory, it is essential to have methods for rapid detection of avian respiratory viruses. Cloacal swabs collected from chickens experimentally infected with H9 subtype AI virus, used in a reverse transcription-polymerase chain reaction (RT-PCR) assay for detection of AI. In infected animals, AI viruses are detected most trequently between days 3 and 7 post infection (p.i.). The RT-PCR assay was able to detect, at least, 103.5 EID50 of AI viruses in the allantoic fluid. The RT-PCR assay did not show any cross-reactivity with some other avian respiratory viruses. In comparison with virus isolation (VI) assay, the relative sensitivity, specificity, correlation rate and positive predictive value of the RT-PCR were 80%, 84%, 82% and 83%, respectively. The Kindex of agreement between the two tests were substantial (K= 0.64). The results proved that the RTPCR assay could be a reliable and rapid alternative to VI assay for detection of AI viruses A H9 subtype H9 in fecal specimens.

Leptospirosis is an acute infectious, systemic and septisemic disease which had recent outbreaks in some parts of Iran especially in north provinces. Rapid detection is a critical step for treatment and control ofthis disease. In this research a PCR based method was evaluated for detection of Iranian local endemic serovars. All reported endemic serovars of Leptospira including Leptospira grippotyphosa, Leptospira canicola, Leptospira sejreo harcijo, Leptospira pomona and Leptospira icterohaemorrhagiae which at present are used for Leptospira micro agglutination test (LMAT) in Iran, were subjected to this PCR. Standard representative serovars from ATCC studied in parallel to local serovars. DNA extracted by phenolchloro form- isoamyl alcohol precipitation. After optimization, the sensitivity of PCR was about 1 fg equal to DNA of 1 Leptospira. All studied serovars including non pathogenic L. biflexa produced the reported 285 bp fragment. Restriction analysis with MboI confirmed the PCR product accuracy. In case of L. biflexa the pattern was completely different from the pathogenic ones. None of near matching bacteria had product in this method. This system was able to detect the existence of Leptospira DNA in all of studied LMAT positive serums.

Interferon regulatory factor 7 (IRF7) gene was cloned from a subtractive cDNA library constructed with mRNAs obtained from rainbow trout (Oncorhynchus mykiss) macrophage cell line (RTS-II). Using expressed sequence tag clones of submitted IRF7 amino acid sequences, specific primers were designed. Results showed that IRF7 cDNA contains an ORF of 1251 nucleotides that translates into a 416 residues putative peptide. The 5¢ untranslated region containing 102 nucleotides and the 3¢ UTR of the transcript consists of 645 nucleotides. The start codon of this ORF is located at nt 103 to 105 and lies in favorable sequence context for the initiation of translation. Alignment between the rainbow trout IRF7 and others IRF7s indicated that the predicted trout IRF7 association domain was located in residues 204-371 aa. The rainbow trout IRF7 nucleotide sequence is most similar to fish IRF7 with 41% identity for crucian carp, Carssius auratus, 45% for zebrafish, Danio rerio, and to mammalian IRF7 with 23% for human, Homo sapiens and 7% for mouse, Mus museulus identity overall.

In the February 2006 in two wetlands in northern Iran, the mortality among wild swans was observed, Paralysis was the most prominent feature of the disease. Histologically, diffused necrosis of acinar cells in pancreas, degeneration and necrosis of some neurons in central nervous system (CNS), sever necrotic and hemorrhagic enteritis, foci of haemorrahge and myocardial cell necrosis in the heart, mild to moderate multifocal hepatocytic necrosis, limited focal necrosis in the kidney and testis and morphological evidence of apoptosis in the spleen were observed. Immunohistochemically, influenza virus antigen were detected more often in the acinar cells of pancreas, some neuron and glial cells in CNS, renal tubule of kidney and hepatocytes of the liver. Moreover, antigen was seen in the meisner plexus of the intestine and testis, but seldom in the lung. The RT-PCR tests showed the presence of H5, N1 and NP genes of influenza virus in trachea, lung, liver, kidney, spleen, and cerebellum. Deduced amino acid sequence at cleavage site was PQGERRRKKR G which is typical for highly pathogen avian influenza viruses. The phylogenetic analysis of HA (heamagglutinin) protein showed a very close similarity of Z/101(H5N1) virus with HA proteins of H5 influenza viruses isolates from cat, wild duck, chicken, ostrich and turkey in various geographical regions. Our data confirmed that these new emerging viruses are systemically able to infect wild swans. The high HA protein sequence similarity among H5N1 viruses from various geographical areas of the word shows that the wild aquatic birds are the major cause of worldwide spreading ofH5NI viruses.

With advent and development of DNA recombinant technology and advantages of p. pasloris expression system, fusion (F) protein of PPRV expression, because of effective immunodominant role could be an appropriate candidate for production of recombinant vaccine against PPR disease. In this study, F gene of PPRV Nigeria 75/1 strain (1637 bp) was amplified using RT-PCR and purified. It was then cloned into pPICZaA a secretory expression vector of P. pasloris for first time. The insertion was proved by both production of a 218 bp segment in Nested PCR and isolation of gene from construct by restriction enzyme (XbaI). Finally, it was sequenced. In conclusion, after the expression of fusion (F) gene in p. pastoris expression system, it can be used in production of recombinant vaccine against PPR disease.

Honey bee sting is used in some societies as a treatment of inflammation in joint diseases such as arthritis. To investigate the effect of honey bee venom in reducing inflammation, we selected 30 male Wister rats which were divided in to 6 groups. Except group I, all remaining groups received 0.5 ml of complete Freund's adjuvant to induce arthritis. After 9 days, all animals that received adjuvant were suffering from acute inflammation in their joints especially in knee joint (tibia-tarsal region). Group 2 was not received any treatment. Group 3 was received only saline (0.05 ml) by subcutaneous injection at the site of inflammation. Group 4 received cream without any honey bee venom. Group 5 was received cream containing 200 mg honey bee venom/gram of cream. Group 6 was received 0.05 ml solution containing freshly prepared 7 mg honey bee venom through subcutaneous injection at the site of inflammation. The parameters determined were, arthritis index score (redness, edema, stiffness in movement) and joint diameter, all the parameters were noted before and during experiment. Results obtained in this experiment shows that all animals that received complete freund's adjuvant, suffered from acute inflammation, redness and difficulty in movement. Treatment by honey bee venom at mentioned dose could not reduce either the inflammation or difficulty in movement. This study brings a great doubt for using honey bee venom as an anti inflammatory drug.

A total of 60 neck skin swab samples were taken from 12 different broiler flocks after the chilling stage of processing at a commercial broiler slughtering facility in Mashhad. The presence of Salmonella was assessed by conventional culture method and confirmed by using poly 0 and poly H antiserum in serotyping. PCR amplification of invA gene as a specific method for detection of Salmonella was evaluated.In this study Salmonella was isolated from 11.66% of samples by conventional culture method, then in serological test 28.6% of them detected as serogroup Band 71.4% as serogroup C. In this investigation all positive results by conventional culture method were confirmed by PCR amplification. Because of rapidity and high specificity and sesitivity ofPCR method, by standardization of this method, it could be considered as an alternative to conventional culture method for confirmation of Salmonella presence in raw poultry meat.

This study was carried out at the Razi Institute during 2005-2006 with inspecting 12168 lung tissue of slaughtered sheep and goats at the Ziaran Abattoir. Pneumonia were diagnosed in 282 cases and the affected lung tissue was collected and transferred to the Bacteriology and Pathology Departments for isolating of Pasteurella spp. and interpretation of histopathologic lesions. Pasteurella multocida was isolated from 120 cases. Histopathology sections indicated purulent bronchopneumonia 0.51%, purulent interstitial bronchopneumonia 0.17%, purulent bronchitis / bronchiolitis 0.12%, purulent pleuritis / pleuropneumonia 0.07%, purulent fibrinous bronchopneumonia 0.04%, purulent pneumonia 0.09% and progressive pneumonia 0.01 different in sheep (P<0.001).