Capsid integrity qPCR—an Azo-dye based and culture-independent approach to estimate adenovirus infectivity after disinfection and in the aquatic environment

Recreational, reclaimed and drinking source waters worldwide are under increasing anthropogenic pressure, and often contain waterborne enteric bacterial, protozoan, and viral pathogens originating from non-point source fecal contamination. Recently, the capsid integrity (ci)-qPCR, utilizing the azo-dyes propidium monoazide (PMA) or ethidium monoazide (EMA), has been shown to reduce false-positive signals under laboratory conditions as well as in food safety applications, thus improving the qPCR estimation of virions of public health significance. The compatibility of two widely used human adenovirus (HAdV) qPCR protocols was evaluated with the addition of a PMA/EMA pretreatment using a range of spiked and environmental samples. Stock suspensions of HAdV were inactivated using heat, UV, and chlorine before being quantified by cell culture, qPCR, and ci-qPCR. Apparent inactivation of virions was detected for heat and chlorine treated HAdV while there was no significant difference between ci-qPCR and qPCR protocols after disinfection by UV. In a follow-up comparative analysis under more complex matrix conditions, 51 surface and 24 wastewater samples pre/post UV treatment were assessed for enteric waterborne HAdV to evaluate the ability of ci-qPCR to reduce the number of false-positive results when compared to conventional qPCR and cell culture. Azo-dye pretreatment of non-UV inactivated samples was shown to improve the ability of molecular HAdV quantification by reducing signals from virions with an accessible genome, thereby increasing the relevance of qPCR results for public health purposes, particularly suited to resource-limited low and middle-income settings.Published versio

Oncostatin M Receptor Type II Knockout Mitigates Inflammation and Improves Survival from Sepsis in Mice

Sepsis remains one of the leading causes of death worldwide. Oncostatin M (OSM), an interleukin (IL)-6 family cytokine, can be found at high levels in septic patients. However, little is known about its role in sepsis. This study aimed to determine if the genetic knockout of OSM receptor (OSMR) type II signaling would improve survival in a murine model of sepsis. Aged (>50 weeks) OSMR type II knockout (KO) mice and wild-type (WT) littermates received an intraperitoneal injection of fecal slurry (FS) or vehicle. The KO mice had better survival 48 h after the injection of FS than the WT mice (p = 0.005). Eighteen hours post-FS injection, the KO mice had reduced peritoneal, serum, and tissue cytokine levels (including IL-1β, IL-6, TNFα, KG/GRO, and IL-10) compared to the WT mice (p + F4/80+ Ly6chigh+ macrophages in the peritoneum of KO mice compared to WT mice (34 ± 6 vs. 4 ± 3%, PInt = 0.005). Isolated peritoneal macrophages from aged KO mice had better live E. coli killing capacity than those from WT mice (p 9)/mL; p < 0.001). In summary, deficiency in OSMR type II receptor signaling provided a survival benefit in the progression of sepsis. This coincided with reduced serum levels of pro-inflammatory (IL-1β, TNFα, and KC/GRO) and anti-inflammatory markers (IL-10), increased bacterial killing ability of macrophages, and reduced macrophage infiltration into to site of infection