Procjena orfvirusa (orfv) in vitro i in vivo elektronskom mikroskopijom

Transmission electron microscopy (TEM) was employed for describing skin and scab lesions in goats affected by orf virus and to demonstrate the parapoxvirus from clinical suspensions by negative staining and ORFV confirmation by immunogold electron microscopy. All samples were confirmed as parapoxvirus by semi-nested PCR amplification of partial gene encoding for the B2L envelope protein. Skin lesions were characterized by ballooning degeneration and loss of desmosomes of the spinosum cells, cytolysis and vesicle formation. Nuclear changes included chromatin margination and increase in electron density. Cytoplasmic changes were typical of cell swelling, vacuolation and the presence of uniform, moderately electron dense viroplasm, situated in the perinuclear region. Various intracellular forms including immature virions (IV), mature virions (IMV) and wrapped virions (WV) were observed in the cytoplasm. All these forms of ORFV were observed morphologically akin to vaccinia virus (VACV). Negative staining of clinical samples and viral suspensions showed typical parapoxvirus morphology with a characteristic criss-cross tubular surface pattern. Both ‘capsule’ form (‘C’ form) and ‘mulberry’ form (‘M’ form) of the virus were demonstrated. ORFV speciation was confirmed by immunogold tagging using a monoclonal antibody in ultra-thin skin section.U radu je za opisivanje lezija na koži i krastama kod koza inficiranih orfvirusom primijenjena transmisijska elektronska mikroskopija (TEM). Dokaz parapoksvirusa iz suspenzije i potvrda ORFV-a obavljeni su tehnikom negativnog bojenja te imunogold elektronskom mikroskopijom. Svi su uzorci potvrđeni kao parapoksvirusi pomoću poluugniježđene lančane reakcije polimerazom kojom je umnožen dio gena B2L odgovornog za protein ovojnice. Lezije na koži karakterizirane su balonskom degeneracijom i gubitkom dezmosoma spinoznih stanica, citolizom te stvaranjem vezikula. Promjene u jezgri uključivale su rub kromatina i povećanje elektronske gustoće. Citoplazmatske promjene bile su tipične za bubrenje stanica, vakuolaciju i prisutnost uniformnog, umjereno elektronički gustog viroplazma smještenog u međumembranskom području. U citoplazmi su zabilježeni različiti unutarstanični oblici, uključujući nezrele virione (IV), zrele virione (IMV) i omotane virione (WV). Svi su oblici ORFV-a pokazali morfološku srodnost s virusom vakcinije (VACV). Negativno bojenje uzoraka i suspenzija pokazali su tipičnu građu parapoksvirusa s karakterističnim križno-cjevastim uzorkom na površini. Također, prikazana su oba oblika virusa, kapsula (oblik C) i dud (oblik M). Specijacija ORFV-a potvrđena je imunogold obilježavanjem pri čemu su korištena monoklonska protutijela u ultratankom rezu kože

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Not AvailableSeroprevalene of Orf in Goats of Assam State of North-East India and Immune Response to a Cell Culture Adapted Orf Virus Vaccine in Goat.Not Availabl

Mosquito abundance and pig seropositivity as a correlate of Japanese encephalitis in human population in Assam, India

Background & objectives: Assam is the most vulnerable state for Japanese encephalitis (JE) in India. The situation warrants characterization of epidemiological patterns of JE in vectors, pigs and human population. This investigation was aimed to determine the relative abundance of mosquito species and seroprevalence of JE in pigs in order to draw an epidemiological association with reported human JE cases in Assam. Methods: Pig sera and mosquitoes from selected farms in Sivasagar and Kamrup districts of Assam were collected fortnightly for one year during June 2015–May 2016. Pig sera were tested for JE antibodies by haemagglutination and virus neurtralization tests. Mosquito species were identified microscopically following the taxonomic keys. The results were analyzed with data on confirmed human JE cases in the selected districts. Results: Culex gelidus (26.07%) and Cx. tritaeniorhynchus (24.07%) were the most abundant species in collected mosquitoes (n = 997). A total of 22.99% of pigs (n = 335) were JEV seropositive and 45.65% of human acute encephalitis syndrome cases (n = 230) were positive for JE virus (JEV) infection. Relative mosquito abundance, pig positivity and human cases were highest during monsoon (June–September) and least during winter (December–February). Rise in mosquito population was observed during pre-monsoon season (March–May) and concurrently higher number of human cases and pig seropositivity were recorded. A good correlation was observed between mosquito number and JEV positivity in pigs/human, and between pigs and human cases (p < 0.05). Human population in Sivasagar was at higher risk for JE infection (OR: 6.46, p < 0.0001) than in Kamrup rural district. Interpretation & conclusion: This study indicates that a seasonal correlation exists between mosquito abundance and JEV seroconversion in pigs with concurrent human JEV outbreaks under field conditions in Sivasagar and Kamrup rural districts of Assam and that monitoring mosquito abundance/density and pig JEV seropositivity may help in predicting JEV outbreak in human population in the region

Experimental infection of pigs with group A rotavirus and enterotoxigenic Escherichia coli in India: gross, histopathological and immunopathological study

The authors describe a detailed study conducted in Assam, India, of gross, histopathological and immunopathological alterations in pigs experimentally infected with rotavirus and enterotoxigenic Escherichia coli (ETEC) expressing K88 pili. A total of 30 Caesarean derived piglets were infected experimentally with rotavirus alone or in combination with ETEC to study the gross and histopathological alterations and the distribution pattern of different B- and T‑cell subsets in the gut. Villus atrophy, especially in the jejunum and ileum, was the consistent lesion in piglets infected with rotavirus, while in piglets simultaneously infected with rotavirus and ETEC, severe necrosis of the intestinal villi was observed. Ultrastructural studies revealed similar pathological alterations in the ileum of the infected piglets. A morphometric study of the intestinal villi and crypts showed a reduction in the ratio between the average villus height and crypt depth (VH:CD ratio) in the group infected with rotavirus (5.95 ± 0.33) and those infected with rotavirus and ETEC (7.90 ± 0.16). A higher (p<0.01) reduction in the VH:CD ratio was observed in the jejunum (8.83 ± 0.79) and ileum (8.46 ± 0.78) compared that in the duodenum (10.03 ± 0.50) of the infected pigs. Piglets infected with rotavirus and sacrificed on day 6 post infection revealed the presence of lymphocytes containing cytoplasmic IgA+ (cIgA+) cells in the villus lamina propria and intra-epithelial CD8+ T‑cells in the villus epithelia. Rotavirus infection of young piglets in association with ETEC was more severe than rotavirus infection alone. Such infection resulted in marked clinico-pathological and immunological alterations in the infected piglets