Isolation and genetic characterization of virulent strains of avian paramyxovirus-1 from multiple avian species in Azad Jammu and Kashmir 2017-2018.

Despite intensive vaccination, endemicity of Avian paramyxoviruses-1 (APMV-1) is a significant problem in developing countries in Africa, Middle East, and Asia. Given the importance of APMV-1 in poultry and multiple non-poultry avian species, it is important to continue surveillance programs, routine monitoring and characterization of field isolates in the region where viruses are endemic. The purpose of this study was to pathotyped and genetically characterized 21 APMV-1s isolated from multiple avian species reared in different regions of Azad Jammu and Kashmir (AJK). Phylogenetic analysis based on complete fusion (F) gene sequences showed that 17 APMV-1 isolates obtained from commercial poultry and backyard birds belonged to sub-genotype VIIi. Though, one pigeon-origin APMV-1 isolate was clustered in sub-genotype VIg and three in recently designated new sub-genotype VIm of genotype VI. The pigeon-origin isolates had the following two motifs 113-RKKR↓F-117 and 113-RQRR↓F-117, while all other isolates had the polybasic amino acid sequence 113-RQKR↓F-117 at the F-cleavage site, which is characteristic of virulent APMV-1 strains. These results are consistent with the five viruses that had intracerebral pathogenicity indices (ICPIs) of between 1.50 and 1.73, corresponding to a velogenic pathotype. The APMV-1s isolated from commercial poultry and backyard birds in this study showed low nucleotide distance (0.3-0.9%) and genetically closely related (> 97%) to viruses repeatedly isolated (2011-2017) from multiple avian species in other states of Pakistan. Strengthened surveillance programs in both commercial poultry and backyard flocks are needed to better assess the commercial-backyard bird interface and form a basis for evidence-based measures to limit and prevent APMV-1 transmission.

Inhibitors of ORAI1 Prevent Cytosolic Calcium-Associated Injury of Human Pancreatic Acinar Cells and Acute Pancreatitis in 3 Mouse Models.

BACKGROUND & AIMS: Sustained activation of the cytosolic calcium concentration induces injury to pancreatic acinar cells and necrosis. The calcium release-activated calcium modulator ORAI1 is the most abundant Ca(2+) entry channel in pancreatic acinar cells; it sustains calcium overload in mice exposed to toxins that induce pancreatitis. We investigated the roles of ORAI1 in pancreatic acinar cell injury and the development of acute pancreatitis in mice. METHODS: Mouse and human acinar cells, as well as HEK 293 cells transfected to express human ORAI1 with human stromal interaction molecule 1, were hyperstimulated or incubated with human bile acid, thapsigargin, or cyclopiazonic acid to induce calcium entry. GSK-7975A or CM_128 were added to some cells, which were analyzed by confocal and video microscopy and patch clamp recordings. Acute pancreatitis was induced in C57BL/6J mice by ductal injection of taurolithocholic acid 3-sulfate or intravenous' administration of cerulein or ethanol and palmitoleic acid. Some mice then were given GSK-7975A or CM_128, which inhibit ORAI1, at different time points to assess local and systemic effects. RESULTS: GSK-7975A and CM_128 each separately inhibited toxin-induced activation of ORAI1 and/or activation of Ca(2+) currents after Ca(2+) release, in a concentration-dependent manner, in mouse and human pancreatic acinar cells (inhibition >90% of the levels observed in control cells). The ORAI1 inhibitors also prevented activation of the necrotic cell death pathway in mouse and human pancreatic acinar cells. GSK-7975A and CM_128 each inhibited all local and systemic features of acute pancreatitis in all 3 models, in dose- and time-dependent manners. The agents were significantly more effective, in a range of parameters, when given at 1 vs 6 hours after induction of pancreatitis. CONCLUSIONS: Cytosolic calcium overload, mediated via ORAI1, contributes to the pathogenesis of acute pancreatitis. ORAI1 inhibitors might be developed for the treatment of patients with pancreatitis.