Characterization of a monothiol glutaredoxin encoded by Chlorella virus PBCV-1

Annotation of the 330-kb Chlorella virus PBCV-1 genome identified a 237 nucleotide gene (a438l) that codes for a protein with ~35% amino acid identity to glutaredoxins (Grx) found in other organisms. The PBCV-1 protein resembles classical Grxs in both size (9 kDa) and location of the active site (N-terminus). However, the PBCV-1 Grx is unusual because it contains a monothiol active site (CPYS) rather than the typical dithiol active site (CPYC). To examine this unique active site, four sitespecific mutants (CPYC, CPYA, SPYC, and SPYS) were constructed to determine if the N-terminal cysteine is necessary for enzyme activity. Wild type and both mutants containing N-terminal cysteines catalyzed the reduction of disulfides in a coupled system with GSH, NADPH, and glutathione reductase. However, both mutants with an altered N-terminal cysteine were inactive. The grx gene is common in the Chlorella viruses. Molecular phylogenetic analyses of the PBCV-1 enzyme support its relatedness to those from other Chlorella viruses and yet demonstrate the divergence of the Grx molecule

Tellurite Resistance in Shiga Toxin-Producing \u3ci\u3eEscherichia coli \u3c/i\u3e

Potassium tellurite ( K2TeO3) is an effective selective agent for O157:H7 Shiga toxin-producing Escherichia coli (STEC), whereas tellurite resistance in non-O157 STEC is variable with information on O45 minimal. High-level K2TeO3 resistance in STEC is attributable to the ter gene cluster with terD an indicator of the cluster’s presence. Polymerase chain reactions for terD and K2TeO3 minimum inhibitory concentration (MIC) determinations in broth cultures were conducted on 70 STEC and 40 non-STEC control organisms. Sixty-six STEC strains (94.3%) were terD+ compared to 28 control organisms (70.0%; P \u3c 0.001). The prevalence of terD in O103 STEC strains was 70%, whereas in all other serogroups it was ≥ 90%. The K2TeO3 geometric mean MIC ranking for STEC serogroups from highest to lowest was O111 \u3e O26 \u3e O145 \u3e O157 \u3e O103 \u3e O12 1 = O45. The K2TeO3 geometric mean MIC was significantly higher in terD+ than in terD− STEC, but not in terD+ versus terD− control strains. Resistance to K2TeO3 (MIC ≥ 25 mg/L) was exhibited by 65/66 terD+ and 0/4 terD− STEC strains, compared to 12/28 terD+ and 8/12 terD− control strains. These results confirm previous studies showing the significantly higher prevalence of the ter gene cluster in STEC strains, and the relationship between these genes and K2TeO3 resistance in STEC and especially intimin (eae)-positive STEC, in contrast to non-STEC organisms. O45 and O121 STEC, although frequently terD positive, on average had significantly lower levels of K2TeO3 resistance than O26, O111, and O145 STEC

Human Ubc9 Contributes to Production of Fully Infectious Human Immunodeficiency Virus Type 1 Virions

Ubc9 was identified as a cellular protein that interacts with the Gag protein of Mason-Pfizer monkey virus. We show here that Ubc9 also interacts with the human immunodeficiency virus type 1 (HIV-1) Gag protein and that their interaction is important for virus replication. Gag was found to colocalize with Ubc9 predominantly at perinuclear puncta. While cells in which Ubc9 expression was suppressed with RNA interference produced normal numbers of virions, these particles were 8- to 10-fold less infectious than those produced in the presence of Ubc9. The nature of this defect was assayed for dependence on Ubc9 during viral assembly, trafficking, and Env incorporation. The Gag-mediated assembly of virus particles and protease-mediated processing of Gag and Gag-Pol were unchanged in the absence of Ubc9. However, the stability of the cell-associated Env glycoprotein was decreased and Env incorporation into released virions was altered. Interestingly, overexpression of the Ubc9 trans-dominant-negative mutant C93A, which is a defective E2-SUMO-1 conjugase, suggests that this activity may not be required for interaction with Gag, virion assembly, or infectivity. This finding demonstrates that Ubc9 plays an important role in the production of infectious HIV-1 virions

Prevalence and Level of Enterohemorrhagic \u3ci\u3eEscherichia coli\u3c/i\u3e in Culled Dairy Cows at Harvest

The primary objective of this study was to determine the prevalence and level of enterohemorrhagic Escherichia coli (EHEC) O26, O45, O103, O111, O121, and O145 (collectively EHEC-6) plus EHEC O157 in fecal, hide, and preintervention carcass surface samples from culled dairy cows. Matched samples (n=300) were collected from 100 cows at harvest and tested by a culture-based method and two molecular methods: NeoSEEK STEC (NS) and Atlas STEC EG2 Combo. Both the culture and NS methods can be used to discriminate among the seven EHEC types (EHEC-7), from which the cumulative prevalence was inferred, whereas the Atlas method can discriminate only between EHEC O157 and non-O157 EHEC, without discrimination of the serogroup. The EHEC-7 prevalence in feces, hides, and carcass surfaces was 6.5, 15.6, and 1.0%, respectively, with the culture method and 25.9, 64.9, and 7.0%, respectively, with the NS method. With the Atlas method, the prevalence of non-O157 EHEC was 29.1, 38.3, and 28.0% and that of EHEC O157 was 29.1, 57.0, and 3.0% for feces, hides, and carcasses, respectively. Only two samples (a hide sample and a fecal sample) originating from different cows contained quantifiable EHEC. In both samples, the isolates were identified as EHEC O157, with 4.7 CFU/1,000 cm2 in the hide sample and 3.9 log CFU/g in the fecal sample. Moderate agreement was found between culture and NS results for detection of EHEC O26 (k=0.58,

Haemorrhagic Colitis Associated with Enterohaemorrhagic \u3ci\u3eEscherichia coli\u3c/i\u3e O165:H25 Infection in a Yearling Feedlot Heifer

Introduction: Enterohaemorrhagic Escherichia coli (EHEC) cause haemorrhagic colitis and haemolytic uraemic syndrome in humans. Although EHEC infection typically results in haemorrhagic colitis in all ages of human patients, in cattle it is usually limited to 1- to 5-week-old nursing calves. Case Presentation: A 1-year-old feedlot beef heifer was moribund with neurological signs and bloody diarrhoea. At necropsy, the colonic mucosa contained multiple grossly visible haemorrhagic erosions, each measuring \u3c1 mm in diameter. Histologically, foci corresponding to the gross erosions had E. coli O165 antigen-positive bacterial rods adherent to the apical surfaces of degenerate and necrotic colonic mucosal epithelial cells in association with attaching and effacing lesions, and also within cytoplasmic vacuoles in some of these cells. An E. coli O165:H25 strain was isolated from the colonic mucosal tissue, and by microarray analysis was found to contain virulence genes corresponding to type III secretion system (T3SS) structure and regulation (cesD, cesT, escD, escF, escN/escV, escR, escT, ler, sepL, sepQ), T3SS effectors (espA, espB, espC, espD, espD, espF, espH, espJ, nleB, nleC, nleD, nleH, tir), serine proteases (eatA, espC, espP), Shiga toxin (stx2), EHEC-haemolysin (ehxA), and adhesins [intimin-ε (eae-ε), type 1 fimbria (fimA, fimB, fimH), type IV pili (pilA, pilB, pilC, pilM, pilP, pilQ) and non-fimbrial adhesin (efa1/lifA)]. Conclusion: To the best of our knowledge, this is the first report of disease in cattle associated with EHEC O165:H25 infection, the oldest bovine EHEC disease case with isolation of the pathogen and the first bovine case to demonstrate grossly evident, haemorrhagic, colonic mucosal erosions associated with EHEC infection

Haemorrhagic Colitis Associated with Enterohaemorrhagic \u3ci\u3eEscherichia coli\u3c/i\u3e O165:H25 Infection in a Yearling Feedlot Heifer

Introduction: Enterohaemorrhagic Escherichia coli (EHEC) cause haemorrhagic colitis and haemolytic uraemic syndrome in humans. Although EHEC infection typically results in haemorrhagic colitis in all ages of human patients, in cattle it is usually limited to 1- to 5-week-old nursing calves. Case Presentation: A 1-year-old feedlot beef heifer was moribund with neurological signs and bloody diarrhoea. At necropsy, the colonic mucosa contained multiple grossly visible haemorrhagic erosions, each measuring \u3c1 mm in diameter. Histologically, foci corresponding to the gross erosions had E. coli O165 antigen-positive bacterial rods adherent to the apical surfaces of degenerate and necrotic colonic mucosal epithelial cells in association with attaching and effacing lesions, and also within cytoplasmic vacuoles in some of these cells. An E. coli O165:H25 strain was isolated from the colonic mucosal tissue, and by microarray analysis was found to contain virulence genes corresponding to type III secretion system (T3SS) structure and regulation (cesD, cesT, escD, escF, escN/escV, escR, escT, ler, sepL, sepQ), T3SS effectors (espA, espB, espC, espD, espD, espF, espH, espJ, nleB, nleC, nleD, nleH, tir), serine proteases (eatA, espC, espP), Shiga toxin (stx2), EHEC-haemolysin (ehxA), and adhesins [intimin-ε (eae-ε), type 1 fimbria (fimA, fimB, fimH), type IV pili (pilA, pilB, pilC, pilM, pilP, pilQ) and non-fimbrial adhesin (efa1/lifA)]. Conclusion: To the best of our knowledge, this is the first report of disease in cattle associated with EHEC O165:H25 infection, the oldest bovine EHEC disease case with isolation of the pathogen and the first bovine case to demonstrate grossly evident, haemorrhagic, colonic mucosal erosions associated with EHEC infection

Increasing dominance of large lianas in Amazonian forests

Ecological orthodoxy suggests that old-growth forests should be close to dynamic equilibrium, but this view has been challenged by recent findings that neotropical forests are accumulating carbon and biomass, possibly in response to the increasing atmospheric concentrations of carbon dioxide. However, it is unclear whether the recent increase in tree biomass has been accompanied by a shift in community composition. Such changes could reduce or enhance the carbon storage potential of old-growth forests in the long term. Here we show that non-fragmented Amazon forests are experiencing a concerted increase in the density, basal area and mean size of woody climbing plants (lianas). Over the last two decades of the twentieth century the dominance of large lianas relative to trees has increased by 1.7–4.6% a year. Lianas enhance tree mortality and suppress tree growth, so their rapid increase implies that the tropical terrestrial carbon sink may shut down sooner than current models suggest. Predictions of future tropical carbon fluxes will need to account for the changing composition and dynamics of supposedly undisturbed forests

Nitrogen fertilizer driven nitrous and nitric oxide production is decoupled from microbial genetic potential in low carbon, semi-arid soil

IntroductionNitrous oxide (N2O) emission from soil is a major concern due to its contribution to global climate change and its function as a loss mechanism of plant-available nitrogen (N) from the soil. This is especially true in intensive agricultural soils with high rates of N fertilizer application such as those on the semi-arid Southern High Plains, USA.MethodsThis study examined emissions of N2O, pore-space concentrations of N2O and nitric oxide (NO), soil chemical properties, water content, and the genetic potential for N cycling five years after conservation system and N management implementation.ResultsFor these semi-arid soils with low N, carbon, and water contents, large soil N2O emissions (up to 8 mL N2O-N m-2 day-1) are directly related to the application of N fertilizer which overwhelms the N2O reducing capacity of the soil. When this fertilizer N is depleted, N2O flux is either low, non-existent, or net-negative and has been observed as early as mid-season for preplant applied N fertilizer (-0.1 mL N2O-N m-2 day-1). Soil pore-space gas concentrations (N2O and NO) remained relatively constant across the growing season (average N2O: 0.78 µL N2O L-1 soil air; NO: 3.3 µL NO L-1 soil air, indicating a base-level of N-cycle activity, but was not directly related to surface emissions of N2O which decreased across the growing season. In addition, genetic potential for N cycle activities increased across the growing season simultaneously with stagnant/reduced N cycle activity. This reflects the difficulty in relating genetic potential to in-situ activity in field research.ConclusionIt is likely that in a nutrient and carbon-poor soil, such as the semi-arid agricultural soil in this study, the microbial processes associated with N cycling are mostly limited by inorganic-N and less directly related to genetic potential at the time of sampling

The Prevalence and Influence of the Combination of Humor and Violence in Super Bowl Commercials

The growing concern over violence in the media has led to vast amounts of research examining the effects of violent media on viewers. An important subset of this research looks at how humor affects this relationship. While research has considered this subset in television programming, almost no research has explored this in the context of advertising. This paper builds on the little research that exists by examining the effects of combining humor and violence, as well as the theoretical approaches that underlie these effects. A content analysis is conducted to identify the prevalence of violence, humor, and the combination of these elements in a longitudinal sample of Super Bowl commercials (2005, 2007, and 2009). Further, we investigate the relationship between the joint occurrence of humor and violence in ads and ad popularity. We conclude that violent acts are rampant in these commercials and that many acts are camouflaged by the simultaneous presence of humor, especially in the most popular ads