Morphine tolerance in vivo is reduced following blockade of the orphanin FQ/nociceptin (OFQ/N)/opioid receptor-like 1 (ORL1) receptor system, suggesting that OFQ/N contributes to the development of morphine tolerance. We previously reported that a 60-min activation of ORL1 receptors natively expressed in BE(2)-C cells desensitized both and ORL1 receptor-mediated inhibition of cAMP. Investigating the mechanism(s) of OFQ/N-mediated and ORL1 receptor cross-talk, we found that pretreatment with the protein kinase C inhibitor, chelerythrine chloride (1 M), blocked OFQ/N-mediated homologous desensitization of ORL1 and heterologous desensitization of opioid receptors. Furthermore, depletion of PKC by 12-O-tetradecanoylphorbol-13-acetate exposure (48 h, 1 M) also prevented OFQ/N-mediated and ORL1 desensitization. OFQ/N pretreatment resulted in translocation of PKC-␣, G proteincoupled receptor kinase 2 (GRK2) and GRK3 from the cytosol to the membrane, and this translocation was also blocked by chelerythrine. Reduction of GRK2 and GRK3 levels by antisense, but not sense DNA treatment blocks ORL1 and receptor desensitization. This suggests that PKC-␣ is required for GRK2 and GRK3 translocation to the membrane, where GRK can inactivate ORL1 and opioid receptors upon rechallenge with the appropriate agonist. Our results demonstrate for the first time the involvement of conventional PKC isozymes in OFQ/N-induced -ORL1 cross-talk, and represent a possible mechanism for OFQ/N-induced anti-opioid actions.Orphanin FQ/nociceptin (OFQ/N), an exquisitely selective agonist at the opioid receptor-like 1 (ORL1) receptor, modulates both behavioral (nociception, anxiety, learning, reward) and immune (cell proliferation) responses (Harrison and Grandy, 2000;Peluso et al., 2001). Although ORL1 and OFQ/N share greater than 40% homology with other opioid receptors and endogenous opioid peptides, many of their actions are anti-opioid in nature (Harrison and Grandy, 2000). One cellular mechanism for their anti-opioid effect is attributed, at least partially, to actions of OFQ/N on different populations of neurons in the brainstem (Pan et al., 2000). The molecular basis of their anti-opioid effects still remains to be determined, but heterologous cross-talk is a possibility since and ORL1 receptors are colocalized on several cell populations within the descending analgesic pathway (Connor et al

Background. Clostridium difficile infection is often considered to result from recent acquisition of a C difficile isolate in a healthcare setting. However, C difficile spores can persist for long periods of time, suggesting a potentially large community environmental reservoir. The objectives of this study were to assess community environmental contamination of toxigenic C difficile and to assess strain distribution in environmental versus clinical isolates.Methods.From 2013 to 2015, we collected community environmental swabs from homes and public areas in Houston, Texas to assess C difficile contamination. All positive isolates were tested for C difficile toxins A and B, ribotyped, and compared with clinical C difficile isolates obtained from hospitalized patients in Houston healthcare settings.Results.A total of 2538 environmental samples were collected over the study period. These included samples obtained from homes (n = 1079), parks (n = 491), chain stores (n = 225), fast food restaurants (n = 123), other commercial stores (n = 172), and hospitals (n = 448). Overall, 418 environmental isolates grew toxigenic C difficile (16.5%; P < .001) most commonly from parks (24.6%), followed by homes (17.1%), hospitals (16.5%), commercial stores (8.1%), chain stores (7.6%), and fast food restaurants (6.5%). A similar distribution of ribotypes was observed between clinical and environmental isolates with the exception that ribotype 027 was more common in clinical isolates compared with environmental isolates (P < .001).Conclusions.We identified a high prevalence of toxigenic C difficile from community environs that were similar ribotypes to clinical isolates. These findings suggest that interventions beyond isolation of symptomatic patients should be targeted for prevention of C difficile infection.

BACKGROUND Conflicting reports have been published on the association between Clostridium difficile ribotypes and severe disease outcomes in patients with C. difficile infection (CDI); several so-called hypervirulent ribotypes have been described. We performed a multicenter study to assess severe disease presentation and severe outcomes among CDI patients infected with different ribotypes. METHODS Stool samples that tested positive for C. difficile toxin were collected and cultured from patients who presented to any of 7 different hospitals in Houston, Texas (2011–2013). C. difficile was characterized using a fluorescent PCR ribotyping method. Medical records were reviewed to determine clinical characteristics and ribotype association with severe CDI presentation (ie, leukocytosis and/or hypoalbuminemia) and severe CDI outcomes (ie, ICU admission, ileus, toxic megacolon, colectomy, and/or in-hospital death). RESULTS Our study included 715 patients aged 61 ± 18 years (female: 63%; median Charlson comorbidity index: 2.5 ± 2.4; hospital-onset CDI: 45%; severe CDI: 36.7%; severe CDI outcomes: 12.3%). The most common ribotypes were 027, 014-020, FP311, 002, 078-126, and 001. Ribotype 027 was a significant independent predictor of severe disease (adjusted odds ratio [aOR], 2.24; 95% confidence interval [CI], 1.53–3.29; P < .001) and severe CDI outcomes (aOR, 1.71; 95% CI, 1.02–2.85; P = .041) compared with all other ribotypes in aggregate. However, in an analysis using all common ribotypes as individual variables, ribotype 027 was not associated with severe CDI outcomes more often than other ribotypes. CONCLUSION Ribotype 027 showed virulence equal to that of other ribotypes identified in this endemic setting. Clinical severity markers of CDI may be more predictive of severe CDI outcomes than a particular ribotype.

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