Citations

ABSTRACT We assessed the performance of a duplex real-time PCR assay for bla KPC and bla NDM performed directly (D-PCR) on perianal and perirectal swabs and stool. Spiked specimens and 126 clinical surveillance swabs (comprising a sensitivity panel of 46 perirectal double swabs previously determined to be culture positive for bla KPC -PCR-positive Enterobacteriaceae and a specificity panel of 80 perianal swabs from patients at risk of carbapenemase-producing Enterobacteriaceae [CPE] colonization) were studied. For the surveillance swabs, D-PCR was compared to PCR after broth enrichment (BE-PCR) and two culture-based methods: the HardyCHROM ESBL agar (HC-A) and the CDC screening (CDC-A) methods. PCR was performed on morphologically distinct colonies that were isolated by culture. All of the initial PCR testing was done without extraction using a simple lysis procedure. The analytical sensitivities of D-PCR for bla KPC were 9 CFU/μl (for swabs) and 90 CFU/μl (for stool), and for bla NDM , it was 1.9 CFU/μl (for both swabs and stool). In the clinical sensitivity panel, D-PCR and BE-PCR were initially positive for bla KPC in 41/46 (89.1%) and 43/46 (93.5%) swabs, respectively. The swabs that were initially negative by D-PCR ( n = 5) and BE-PCR ( n = 3) were visibly stool soiled; all swabs were bla KPC positive upon repeat testing after lysate extraction. The CDC-A and HC-A yielded bla KPC -positive Enterobacteriaceae from 36/46 (78.3%) and 35/46 (76.1%) swabs, respectively (sensitivities of D-PCR/BE-PCR postextraction of soiled specimens versus HC-A, P = 0.0009, and versus CDC-A, P = 0.0016). All swabs in the specificity panel were negative for CPE by all four methods. D-PCR allows for the timely detection of bla KPC and bla NDM carriage with excellent sensitivity when specimens visibly soiled with stool undergo preparatory extraction.

The age‐associated decline in growth hormone (GH) secretion may be a consequence of the reduction in the number of GH‐releasing hormone (GHRH) positive neurones. However, it remains unclear whether an alteration in the number or distribution of somatostatin (SST) neurones contributes to this change. In the present study, we characterised the role of SST in modulating the change in pulsatile GH secretion in male C57Bl/6J mice throughout puberty and into early adulthood. We assessed pulsatile GH secretion in mice at 4, 8 and 16 weeks of age. These ages correspond to early pubertal, early adulthood and adulthood, respectively. We show an elevation in peak, total and pulsatile GH secretion coinciding with periods of rapid linear growth. Using in situ hybridisation and morphometric methods, we mapped the distribution of Sst mRNA expression within the mouse brain relative to this change in pulsatile GH secretion. The results obtained show that altered pulsatile GH secretion in male mice from 4–16 weeks of age does not coincide with a significant change in the number of Sst mRNA expressing neurones or an abundance of Sst mRNA expression throughout the arcuate nucleus (ARC) and periventricular nucleus (PeV). Rather, we observed a progressive decline in Sst mRNA expressing neurones within subnuclei of the paraventricular nucleus at this time. We conclude that structural changes in Sst expression within the PeV and ARC may not reflect the observed decline in pulsatile GH secretion in mice from puberty into early adulthood.

ABSTRACT We compared carbapenemase detection among 271 Gram-negative bacilli (of which 131 were carbapenemase producers) using a novel chromogenic rapid test—the Carba NP test (CNP)—and the modified Hodge test (MHT). Sensitivities were comparable (CNP, 100%, versus MHT, 98%; P = 0.08), but CNP was more specific (100% versus 80%; P < 0.0001) and faster.

Consensus abstract Background The management of primary rectal cancer beyond total mesorectal excision planes (PRC-bTME) and recurrent rectal cancer (RRC) is challenging. There is global variation in standards and no guidelines exist. To achieve cure most patients require extended, multivisceral, exenterative surgery, beyond conventional total mesorectal excision planes. The aim of the Beyond TME Group was to achieve consensus on the definitions and principles of management, and to identify areas of research priority. Methods Delphi methodology was used to achieve consensus. The Group consisted of invited experts from surgery, radiology, oncology and pathology. The process included two international dedicated discussion conferences, formal feedback, three rounds of editing and two rounds of anonymized web-based voting. Consensus was achieved with more than 80 per cent agreement; less than 80 per cent agreement indicated low consensus. During conferences held in September 2011 and March 2012, open discussion took place on areas in which there is a low level of consensus. Results The final consensus document included 51 voted statements, making recommendations on ten key areas of PRC-bTME and RRC. Consensus agreement was achieved on the recommendations of 49 statements, with 34 achieving consensus in over 95 per cent. The lowest level of consensus obtained was 76 per cent. There was clear identification of the need for referral to a specialist multidisciplinary team for diagnosis, assessment and further management. Conclusion The consensus process has provided guidance for the management of patients with PRC-bTME or RRC, taking into account global variations in surgical techniques and technology. It has further identified areas of research priority.

Consensus abstract Background The management of primary rectal cancer beyond total mesorectal excision planes (PRC-bTME) and recurrent rectal cancer (RRC) is challenging. There is global variation in standards and no guidelines exist. To achieve cure most patients require extended, multivisceral, exenterative surgery, beyond conventional total mesorectal excision planes. The aim of the Beyond TME Group was to achieve consensus on the definitions and principles of management, and to identify areas of research priority. Methods Delphi methodology was used to achieve consensus. The Group consisted of invited experts from surgery, radiology, oncology and pathology. The process included two international dedicated discussion conferences, formal feedback, three rounds of editing and two rounds of anonymized web-based voting. Consensus was achieved with more than 80 per cent agreement; less than 80 per cent agreement indicated low consensus. During conferences held in September 2011 and March 2012, open discussion took place on areas in which there is a low level of consensus. Results The final consensus document included 51 voted statements, making recommendations on ten key areas of PRC-bTME and RRC. Consensus agreement was achieved on the recommendations of 49 statements, with 34 achieving consensus in over 95 per cent. The lowest level of consensus obtained was 76 per cent. There was clear identification of the need for referral to a specialist multidisciplinary team for diagnosis, assessment and further management. Conclusion The consensus process has provided guidance for the management of patients with PRC-bTME or RRC, taking into account global variations in surgical techniques and technology. It has further identified areas of research priority.

Background Autosomal recessive limb girdle muscular dystrophy (LGMD2) is a heterogeneous group of myopathies characterised by progressive muscle weakness involving proximal muscles of the shoulder and pelvic girdles including at least 17 different genetic entities. Additional loci have yet to be identified as there are families which are unlinked to any of the known loci. Here we have investigated a consanguineous family with LGMD2 with two affected individuals in order to identify the causative gene defect. Methods and results We performed genome wide homozygosity mapping and mapped the LGMD2 phenotype to chromosome 2q35–q36.3. DNA sequence analysis of the highly relevant candidate gene DES revealed a homozygous splice site mutation c.1289-2A>G in the two affected family members. Immunofluorescent staining and western blot analysis showed that the expression and the cytoskeletal network formation of mutant desmin were well preserved in skeletal muscle fibres. Unlike autosomal dominant desminopathies, ultrastructural alterations such as disruption of myofibrillar organisation, formation of myofibrillar degradation products and dislocation/aggregation of membranous organelles were not present. This novel splice site mutation results in addition of 16 amino acids within the tail domain of desmin, which has been suggested to interact with lamin B protein. We also detected a specific disruption of desmin-lamin B interaction in the skeletal muscle of the patient by confocal laser scanning microscopy. Conclusions Our study reveals that autosomal recessive mutations in DES cause LGMD2 phenotype without features of myofibrillar myopathy.

Abstract Purpose: We set out to identify Stearoyl-CoA desaturase 1 (SCD1) as a novel molecular target in clear cell renal cell carcinoma (ccRCC) and examine its role in tumor cell growth and viability in vitro and in vivo independently as well as in combination with current U.S. Food and Drug Administration (FDA)-approved regimens. Experimental Design: Patient normal and ccRCC tissue samples and cell lines were examined for SCD1 expression. Genetic knockdown models and targeted inhibition of SCD1 through use of a small molecule inhibitor, A939572, were analyzed for growth, apoptosis, and alterations in gene expression using gene array analysis. Therapeutic models of synergy were evaluated utilizing pharmacologic inhibition of SCD1 with the tyrosine kinase inhibitors (TKI) sunitinib and pazopanib, and the mTOR inhibitor temsirolimus. Results: Our studies identify increased SCD1 expression in all stages of ccRCC. Both genetic knockdown and pharmacologic inhibition of SCD1 decreased tumor cell proliferation and induced apoptosis in vitro and in vivo. Upon gene array, quantitative real-time PCR, and protein analysis of A939572-treated or SCD1 lentiviral knockdown samples, induction of endoplasmic reticulum stress response signaling was observed, providing mechanistic insight for SCD1 activity in ccRCC. Furthermore, combinatorial application of A939572 with temsirolimus synergistically inhibited tumor growth in vitro and in vivo. Conclusions: Increased SCD1 expression supports ccRCC viability and therefore we propose it as a novel molecular target for therapy either independently or in combination with an mTOR inhibitor for patients whose disease cannot be remedied with surgical intervention, such as in cases of advanced or metastatic disease. Clin Cancer Res; 19(9); 2368–80. ©2013 AACR.

Abstract Barrett's esophagus is the precursor lesion of esophageal adenocarcinoma, whose progression follows sequential stages. However, the low progression rate and the inadequacy and subjective interpretation of histologic grading in predicting Barrett's esophagus progression call for more objective biomarkers that can improve risk prediction. We conducted a genome-wide profiling of 754 human microRNAs (miRNA) in 35 normal epithelium, 34 Barrett's esophagus, and 36 esophageal adenocarcinoma tissues using TaqMan real-time PCR-based profiling. Unsupervised hierarchical clustering using 294 modestly to highly expressed miRNAs showed clear clustering of two groups: normal epithelium versus Barrett's esophagus/esophageal adenocarcinoma tissues. Moreover, there was an excellent clustering of Barrett's metaplasia (without dysplasia) tissues from normal epithelium tissues. However, Barrett's esophagus tissues of different stages and esophageal adenocarcinoma tissues were interspersed. There were differentially expressed miRNAs at different stages. The majority of miRNA aberrations involved upregulation of expression in Barrett's esophagus and esophageal adenocarcinoma tissues, with the most dramatic alterations occurring at the Barrett's metaplasia stage. Known oncomiRs, such as miR-21, miR-25, and miR-223, and tumor suppressor miRNAs, including miR-205, miR-203, let-7c, and miR-133a, showed progressively altered expression from Barrett's esophagus to esophageal adenocarcinoma. We also identified a number of novel miRNAs that showed progressively altered expression, including miR-301b, miR-618, and miR-23b. The significant miRNA alterations that were exclusive to esophageal adenocarcinoma but not Barrett's esophagus included miR-375 downregulation and upregulation of five members of the miR-17-92 and its homologue clusters, which may become promising biomarkers for esophageal adenocarcinoma development. Cancer Prev Res; 6(3); 196–205. ©2013 AACR.

Background Two recent studies identified a mutation (p.Asp620Asn) in the vacuolar protein sorting 35 gene as a cause for an autosomal dominant form of Parkinson disease . Although additional missense variants were described, their pathogenic role yet remains inconclusive. Methods and results We performed the largest multi-center study to ascertain the frequency and pathogenicity of the reported vacuolar protein sorting 35 gene variants in more than 15,000 individuals worldwide. p.Asp620Asn was detected in 5 familial and 2 sporadic PD cases and not in healthy controls, p.Leu774Met in 6 cases and 1 control, p.Gly51Ser in 3 cases and 2 controls. Overall analyses did not reveal any significant increased risk for p.Leu774Met and p.Gly51Ser in our cohort. Conclusions Our study apart from identifying the p.Asp620Asn variant in familial cases also identified it in idiopathic Parkinson disease cases, and thus provides genetic evidence for a role of p.Asp620Asn in Parkinson disease in different populations worldwide.