Rotavirus Infection Induces Transient Pancreatic Involution and Hyperglycemia in Weanling Mice

Rotavirus is a ubiquitous double-stranded RNA virus responsible for most cases of infantile gastroenteritis. It infects pancreatic islets in vitro and is implicated as a trigger of autoimmune destruction of islet beta cells leading to type 1 diabetes, but pancreatic pathology secondary to rotavirus infection in vivo has not been documented. To address this issue, we inoculated 3 week-old C57Bl/6 mice at weaning with rhesus rotavirus, which is closely related to human rotaviruses and known to infect mouse islets in vitro. Virus was quantified in tissues by culture-isolation and enzyme-linked immunosorbent assay. A requirement for viral double stranded RNA was investigated in toll-like receptor 3 (TLR3)-deficient mice. Cell proliferation and apoptosis, and insulin expression, were analyzed by immunohistochemistry. Following rotavirus inoculation by gavage, two phases of mild, transient hyperglycemia were observed beginning after 2 and 8 days. In the first phase, widespread apoptosis of pancreatic cells was associated with a decrease in pancreas mass and insulin production, without detectable virus in the pancreas. These effects were mimicked by injection of the double-stranded RNA mimic, polyinosinic-polycytidylic acid, and were TLR3-dependent. By the second phase, the pancreas had regenerated but islets were smaller than normal and viral antigen was then detected in the pancreas for several days. These findings directly demonstrate pathogenic effects of rotavirus infection on the pancreas in vivo, mediated initially by the interaction of rotavirus double-stranded RNA with TLR3.     

Vibrio madracius sp. nov. Isolated from Madracis decactis (Scleractinia) in St Peter & St Paul Archipelago,Mid-Atlantic Ridge,Brazil

Three novel isolates (A-354^T, A-328, and A-384) were retrieved from apparently healthy scleractinian Madracis decactis in the remote St Peter & St Paul Archipelago, Mid-Atlantic Ridge, Brazil. The novel isolates formed a distinct lineage based on the phylogenetic reconstruction using the 16S rRNA and pyrH gene sequences. They fell into the Mediterranei clade and their closest phylogenetic neighbour was V. mediterranei species, sharing upto 98.1 % 16S rRNA gene sequence similarity. Genomic analysis including in silico DDH, MLSA, AAI and genomic signature distinguished A-354^T from V. mediterranei LMG 19703 (=AK1) with values of 33.3, 94.2, 92 %, and 11.3, respectively. Phenotypically, the novel isolates can be differentiated from V. mediterranei based on the four following features. They do not grow at 8 % NaCl; use d-gluconic acid but not l-galactonic acid lactone as carbon source; and do not have the fatty acid C_18:0. Differentiation from both the other Mediterranei clade species (V. maritimus and V. variabilis) is supported by fifteen features. The novel species show lysine decarboxylase and tryptophan deaminase, but not gelatinase and arginine dihydrolase activity; produce acetoin; use α-d-lactose, N-acetyl-d-galactosamine, myo-Inositol, d-gluconic acid, and β-hydroxy-d,l-butyric acid; and present the fatty acids C_14:0 iso, C_15:0 anteiso, C_16:0 iso, C_17:0 anteiso, and C_17:1x8c . Whole-cell protein profiles, based on MALDI-TOF, showed that the isolates are not clonal and also distinguished them from the closes phylogenetic neighbors. The name Vibrio madracius sp. nov. is proposed to encompass these novel isolates. The G+C content of the type strain A-354^T (=LMG 28124^T=CBAS 482^T) is 44.5 mol%.     

Apoptosis of CD4<Superscript>+</Superscript>CD25<Superscript>high</Superscript> T cells in response to Sirolimus requires activation of T cell receptor and is modulated by IL-2

Targeted molecular therapies inhibit proliferation and survival of cancer cells but may also affect immune cells. We have evaluated the effects of Sirolimus and Sorafenib on proliferation and survival of lymphoid cell subsets. Both drugs were cytotoxic to CD4⁺CD25^high T cells, and were growth inhibitory for CD4⁺ and CD8⁺ T cells. Cytotoxicity depended on CD3/CD28 stimulation and was detectable within 12 h, with 80–90% of CD4⁺CD25^high cells killed by 72 h. Cell death was due to apoptosis, based on Annexin V and 7AAD staining. Addition of IL-2 prevented the apoptotic response to Sirolimus, potentially accounting for reports that Sirolimus can enhance proliferation of CD4⁺CD25^high cells. These results predict that Sirolimus or Sorafenib would reduce CD4⁺CD25^high cells if administered prior to antigenic stimulation in an immunotherapy protocol. However, administration of IL-2 protects CD4⁺CD25^high T cells from cytotoxic effects of Sirolimus, a response that may be considered in design of therapeutic protocols.     

RNase L plays a role in the antiviral response to West Nile virus

Alleles at the Flv locus determine disease outcome after a flavivirus infection in mice. Although comparable numbers of congenic resistant and susceptible mouse embryo fibroblasts (MEFs) are infected by the flavivirus West Nile virus (WNV), resistant MEFs produce approximately 100- to 150-fold lower titers than susceptible ones and flavivirus titers in the brains of resistant and susceptible animals can differ by >10,000-fold. The Flv locus was previously identified as the 2'-5' oligoadenylate synthetase 1b (Oas1b) gene. Oas gene expression is up-regulated by interferon (IFN), and after activation by double-stranded RNA, some mouse synthetases produce 2-5A, which activates latent RNase L to degrade viral and cellular RNAs. To determine whether the lower levels of intracellular flavivirus genomic RNA from resistant mice detected in cells at all times after infection were mediated by RNase L, RNase L activity levels in congenic resistant and susceptible cells were compared. Similar moderate levels of RNase L activation by transfected 2-5A were observed in both types of uninfected cells. After WNV infection, the mRNAs of IFN-beta and three Oas genes were up-regulated to similar levels in both types of cells. However, significant levels of RNase L activity were not detected until 72 h after WNV infection and the patterns of viral RNA cleavage products generated were similar in both types of cells. When RNase L activity was down-regulated in resistant cells via stable expression of a dominant negative RNase L mutant, approximately 5- to 10-times-higher yields of WNV were produced. Similarly, about approximately 5- to 10-times-higher virus yields were produced by susceptible C57BL/6 RNase L-/- cells compared to RNase L+/+ cells that were either left untreated or pretreated with IFN and/or poly(I) . poly(C). The data indicate that WNV genomic RNA is susceptible to RNase L cleavage and that RNase L plays a role in the cellular antiviral response to flaviviruses. The results suggest that RNase L activation is not a major component of the Oas1b-mediated flavivirus resistance phenotype.     

Microbial communities in wetlands of the Athabasca oil sands: genetic and metabolic characterization

Naphthenic acids are a complex family of naturally occurring cyclic and acyclic carboxylic acids that are present in the acidic fraction of petroleum. Naphthenic acids are acutely toxic to aquatic organisms. Previous studies showed that wetland sediments exposed to oil sands process water containing naphthenic acids had higher rates of naphthenic acid degradation in vitro compared with unexposed wetlands. In this study we compare the microbial community structures in sediments from wetlands exposed to different amounts of oil sands process water using BIOLOG, phospholipid fatty acid analysis and denaturing gradient gel electrophoresis of total bacterial DNA. Community profiles were compared using cluster analysis. BIOLOG profiles were primarily influenced by seasonal trends rather than naphthenic acids content. In contrast, phospholipid fatty acid analysis comparisons clustered communities that had higher levels of residual oil, although this association was not strong. In contrast, cluster diagrams produced from the denaturing gradient gel electrophoresis data clearly separated bacterial communities according to naphthenic acids concentrations, indicating that naphthenic acids content was a major influence on the composition of the bacterial community. In addition, denaturing gradient gel electrophoresis profiles indicated that naphthenic acids-exposed bacterial communities were homogeneous on a scale of meters, whereas unexposed (off-site) wetlands were less homogeneous.     

Natal philopatry does not lead to population genetic differentiation in Buller's albatross (Thalassarche bulleri bulleri)

Genetic variability in the only two existing populations of Buller's albatross (Thalassarche bulleri bulleri) was assessed using six polymorphic microsatellite loci. Large biological samples were obtained from both the Snares (n = 99) and the Solander Islands (n = 27). Several measures of genetic differentiation including F(ST) and R(ST) and a principal coordinates analysis (PCO) suggest a complete absence of genetic structure between three breeding colonies on the Snares Islands, and between them and one breeding colony on the Solander Islands. Mark/recapture studies of recently banded albatross chicks on the Snares found high natal philopatry in T. b. bulleri, but the microsatellite DNA data suggest that sufficient gene flow still occurs between all four breeding colonies to maintain a genetically homogeneous population overall.     

Bilateral Renal Agenesis/Hypoplasia/Dysplasia (BRAHD): Postmortem Analysis of 45 Cases with Breakpoint Mapping of Two De Novo Translocations

Background

Bilateral renal agenesis/hypoplasia/dysplasia (BRAHD) is a relatively common, lethal malformation in humans. Established clinical risk factors include maternal insulin dependent diabetes mellitus and male sex of the fetus. In the majority of cases, no specific etiology can be established, although teratogenic, syndromal and single gene causes can be assigned to some cases.

Methodology/Principal Findings

45 unrelated fetuses, stillbirths or infants with lethal BRAHD were ascertained through a single regional paediatric pathology service (male∶female 34∶11 or 3.1∶1). The previously reported phenotypic overlaps with VACTERL, caudal dysgenesis, hemifacial microsomia and Müllerian defects were confirmed. A new finding is that 16/45 (35.6%; m∶f 13∶3 or 4.3∶1) BRAHD cases had one or more extrarenal malformations indicative of a disoder of laterality determination including; incomplete lobulation of right lung (seven cases), malrotation of the gut (seven cases) and persistence of the left superior vena cava (five cases). One such case with multiple laterality defects and sirelomelia was found to have a de novo apparently balanced reciprocal translocation 46,XY,t(2;6)(p22.3;q12). Translocation breakpoint mapping was performed by interphase fluorescent in-situ hybridization (FISH) using nuclei extracted from archival tissue sections in both this case and an isolated bilateral renal agenesis case associated with a de novo 46,XY,t(1;2)(q41;p25.3). Both t(2;6) breakpoints mapped to gene-free regions with no strong evidence of cis-regulatory potential. Ten genes localized within 500 kb of the t(1;2) breakpoints. Wholemount in-situ expression analyses of the mouse orthologs of these genes in embryonic mouse kidneys showed strong expression of Esrrg, encoding a nuclear steroid hormone receptor. Immunohistochemical analysis showed that Esrrg was restricted to proximal ductal tissue within the embryonic kidney.

Conclusions/Significance

The previously unreported association of BRAHD with laterality defects suggests that renal agenesis may share a common etiology with heterotaxy in some cases. Translocation breakpoint mapping identified ESRRG as a plausible candidate gene for BRAHD.