Sequence analysis and expression of a virus-like particle protein,VLP2, from the parasitic wasp Venturia canescens

Endoparasitoid wasps produce maternal protein secretions, which are transported into the body of insect hosts at oviposition to regulate host physiology for successful development of their offspring. Venturia canescens calyx fluid contains so-called virus-like particles (VLPs) that are essential for immune evasion of the developing parasitoid inside the host. VLPs consist of four major proteins. In this paper, we describe the isolation and molecular cloning of a gene (vlp2) that is a constituent of VLPs and discuss its possible role in VLP structure and function.

Isolation and characterization of a neprilysin-like protein from Venturia canescens virus-like particles

Maternal protein secretions from endoparasitoid wasps are evolutionary adaptations to regulate host physiology as part of an extended wasp phenotype. Virus-like particles (VLPs) produced in the calyx region of Venturia canescens wasps are involved in immune evasion of the developing parasitoid inside the host. In contrast to polydnaviruses (PDVs), VcVLPs are devoid of any nucleic acids. To understand the role of these particles in the regulation of host physiology and phylogenetic relationship between VLPs and PDVs, it is essential to identify particle proteins. In this paper, we describe the isolation and molecular cloning of a neprilysin-like gene (VcNEP) coding for a 94 kDa VcVLP protein and discuss its possible role in host regulation.

Virus or not? Phylogenetics of polydnaviruses and their wasp carriers

Our current, still limited, understanding of the comparative biology and evolution of polydnaviruses (PDVs) is reviewed, especially in the context of the possible origins of these parasitoid viruses and of their coevolution with carrier wasps. A hypothetical scenario of evolution of PDVs from ascovirus (or ascovirus-like) ancestors is presented, with examples of apparent extant transitional forms. PDVs appear, in the case of bracoviruses, to show phylogenetic relationships that mirror those of their wasp carriers: with ichno-viruses, the picture is less clear. Ongoing sequencing studies of entire PDV genomes from diverse wasp species are likely to greatly contribute to our understanding of PDV evolution. (C) 2003 Elsevier Science Ltd. All rights reserved.

Evaluation of the effects of Candidatus Liberibacter asiaticus on inoculated citrus plants using laser-induced breakdown spectroscopy (LIBS) and chemometrics tools

This study investigated the organic and inorganic constituents of healthy leaves and Candidatus Liberibacter asiaticus (CLas)-inoculated leaves of citrus plants. The bacteria CLas are one of the causal agents of citrus greening (or Huanglongbing) and its effect on citrus leaves was investigated using laser-induced breakdown spectroscopy (LIBS) combined with chemometrics. The information obtained from the LIBS spectra profiles with chemometrics analysis was promising for the construction of predictive models to identify healthy and infected plants. The major, macro- and microconstituents were relevant for differentiation of the sample conditions. The models were then applied to different inoculation times (from 1 to 8 months). The models were effective in the classification of 82-97% of the diseased samples with a 95% significance level. The novelty of this method was in the fingerprinting of healthy and diseased plants based on their organic and inorganic contents. (C) 2010 Elsevier B.V. All rights reserved.

Varicella zoster virus in bell`s palsy: a prospective study

Although Bell`s palsy is the major cause of acute peripheral facial palsy, its pathogenesis remains unknown. Reactivation of the varicella zoster virus has been implicated as one of the main causes of Bell`s palsy, however, studies which investigate the varicella zoster virus reactivation in Bell`s palsy patients are mostly Japanese and, therefore, personal and geographic characteristics are quite different from our population. Aims: To determine varicella zoster virus frequency in saliva samples from patients with Bell`s palsy, using PCR. Material and Method: One hundred seventy one patients with acute peripheral facial palsy were prospectively enrolled in this study. One hundred twenty were clinically diagnosed with Bell`s palsy, within one week of onset of the disease and no previous anti-viral therapy. We had 20 healthy adults as controls. Three saliva samples were collected from patients and controls at initial examination and at one and two weeks later. The detection of the varicella zoster virus DNA was performed using PCR. Results: Varicella zoster virus was detected in two patients (1.7%). The virus was not identified in saliva samples from the controls. Conclusions: Varicella zoster virus was detected in 1.7% of saliva samples from patients with Bell`s palsy, using PCR.

Improved method for counting virus and virus like particles

An improved method for counting virus and virus like particles by electron microscopy (EM) was developed. The procedure involves the determination of the absolute concentration of pure or semi-pure particles once deposited evenly on EM grids using either centrifugation or antibody capture techniques. The counting of particles was done with a Microfiche unit which enlarged approximately 50 x the image of particles on a developed negative film which had been taken at a relatively low magnification (2500 x) by EM. Initially, latex particles of a known concentration were counted using this approach, to prove the accuracy of the technique. The latex particles were deposited evenly on an EM grid using centrifugation (Modified Beckmen EM-90 Airfuge technique). Subsequently, recombinant Bluetongue virus (BTV) core-like particles (CLPs) captured by a Monoclonal antibody using a hovel sample loading method were counted by the Microfiche unit method and by a direct EM method. Comparison of the simplified counting method developed with a conventional method, showed good agreement. The method is simple, accurate, rapid, and reproducible when used with either pure particles or with particles from crude cell culture extracts.

Macrophomina phaseolina causing leaf spot of mungbean

Macrophomina phaseolina, causing leaf spot of mungbean is reported in Australia. Koch's postulates were fulfilled. The inoculum source was considered to be microsclerotia of the fungus in soil splashed onto the leaves. The disease is not expected to be a problem in Australia in most years.

CD4+T cells from HIV-1-infected patients recognize wild-type and mutant human immunodeficiency virus-1 protease epitopes

P>Human immunodeficiency virus (HIV)-1 protease is a known target of CD8+ T cell responses, but it is the only HIV-1 protein in which no fully characterized HIV-1 protease CD4 epitopes have been identified to date. We investigated the recognition of HIV-1 protease by CD4+ T cells from 75 HIV-1-infected, protease inhibitor (PI)-treated patients, using the 5,6-carboxyfluorescein diacetate succinimidyl ester-based proliferation assay. In order to identify putative promiscuous CD4+ T cell epitopes, we used the TEPITOPE algorithm to scan the sequence of the HXB2 HIV-1 protease. Protease regions 4-23, 45-64 and 73-95 were identified; 32 sequence variants of the mentioned regions, encoding frequent PI-induced mutations and polymorphisms, were also tested. On average, each peptide bound to five of 15 tested common human leucocyte antigen D-related (HLA-DR) molecules. More than 80% of the patients displayed CD4+ as well as CD8+ T cell recognition of at least one of the protease peptides. All 35 peptides were recognized. The response was not associated with particular HLA-DR or -DQ alleles. Our results thus indicate that protease is a frequent target of CD4+ along with CD8+ proliferative T cell responses by the majority of HIV-1-infected patients under PI therapy. The frequent finding of matching CD4+ and CD8+ T cell responses to the same peptides may indicate that CD4+ T cells provide cognate T cell help for the maintenance of long-living protease-specific functional CD8+ T cells.

Human immunodeficiency virus test-seeking blood donors in a large blood bank in Sao Paulo, Brazil

BACKGROUND: Persons with human immunodeficiency virus (HIV) risk behaviors are excluded from donation to reduce the risk of transfusion-transmitted infection. Persons donating to be tested for HIV may therefore deny risk behaviors. STUDY DESIGN AND METHODS: A random sample of donors completed a survey on motivations, knowledge, and attitudes on the screening process. Donors were considered test seekers if they agreed with two statements ""I think that blood donation is a good, fast, and anonymous way to get my blood tested"" and ""I donate to get my test results."" This study was conducted from June to November 2006 at the largest blood bank in Sao Paulo, Brazil. RESULTS: Of 3061 participants, 208 (7%) were test seekers. They tended to be male and had a lower educational level. They were more likely to have incorrect knowledge about blood safety (e.g., not knowing that a unit can test antibody negative and still transmit infection, 60% vs. 42%, p = 0.02), express dissatisfaction with screening questions (e.g., feeling that important questions were not asked, 14% vs. 5%, p < 0.01), and concur that donors do not answer questions truthfully (e.g., donors have more sexual partners than they admit, 29% vs. 18%, p < 0.01). Test seekers were more likely to believe that it is acceptable to donate blood to get tested for HIV (41% vs. 10%, p < 0.01). CONCLUSIONS: Test-seeking motivation, coupled with low knowledge of window period risk, is counter to improving blood safety and to donor prevention needs. Donor education needs to be improved along with availability of appropriate HIV counseling and testing.

Mechanisms of biocontrol by Pantoea dispersa of sugar cane leaf scald disease caused by Xanthomonas albilineans

Albicidins, a family of phytotoxins and antibiotics produced by Xanthomonas albilineans, are important in sugar cane leaf scald disease development. The albicidin detoxifying bacterium Pantoea dispersa (syn. Erwinia herbicola) SB1403 provides very effective biocontrol against leaf scald disease in highly susceptible sugar cane cultivars. The P. dispersa gene (albD) for enzymatic detoxification of albicidin has recently been cloned and sequenced. To test the role of albicidin detoxification in biocontrol of leaf scald disease, albD was inactivated in P. dispersa by site-directed mutagenesis. The mutants, which were unable to detoxify albicidin, were less resistant to the toxin and less effective in biocontrol of leaf scald disease than their parent strain. This indicates that albicidin detoxification contributes to the biocontrol capacity of P. dispersa against X. albilineans. Rapid growth and ability to acidify media are other characteristics likely to contribute to the competitiveness of P. dispersa against X. albilineans at wound sites used to invade sugar cane.

A systemic parvo-like virus in the freshwater crayfish Cherax destructor

Systemic Cowdry Type A inclusions (CAs) were observed in a moribund Cherax destructor collected at an aquaculture farm in South Australia. Inclusions were most common in the gills and were associated with multifocal necrosis of the main gill axis and lamellae. The hepatopancreas was necrotic; however, only one CA was observed in the interstitial tissues. CAs were associated with necrosis in the abdominal and gut musculature. CAs were also observed in the spongy connective tissues and the epicardium. Empty capsids (17.5 +/- 0.5 nm) and microfilaments were most commonly observed within these inclusions by transmission electron microscopy. Complete icosahedral viral particles (20.8 +/- 1.2 nm) were difficult to distinguish within the viroplasm, but were visualised better in aggregates between the viroplasm and the inner nuclear membrane. The nucleolus was closely associated with the developing viroplasm, and was hypertrophied and segregated into its fibrillar and granular components. The virus was named Cherax destructor systemic parvo-like virus (CdSPV) on the basis of its histopathology, cytopathology and morphology. CdSPV is the first systemic virus described in a freshwater crayfish.

Local dispersion of the Eucalyptus leaf beetle Chrysophtharta bimaculata (Coleoptera: Chrysomelidae), and implications for forest protection

1. Chrysophtharta bimaculata is a native chrysomelid species that can cause chronic defoliation of plantation and regrowth Eucalyptus forests in Tasmania, Australia. Knowledge of the dispersion pattern of C. bimaculata was needed in order to assess the efficiency of an integrated pest management (IPM) programme currently used for its control. 2. Using data from yellow flight traps, local populations of C. bimaculata adults were monitored over a season at spatial scales relevant to commercial forestry: within a 50-ha operational management unit (a forestry 'coupe') and between coupes. In addition, oviposition was monitored over a season at a subset of the between-coupe sites. 3. Dispersion indices (Taylor's Power Law and Iwao's Mean Crowding regression method) demonstrated that C. bimaculata adults were spatially aggregated within and between coupes, although the number of egg-batches laid at the between-coupe scale was uniform. Spatial autocorrelation analysis showed that trap-catches at the within-coupe level were similar (positively autocorrelated) to a radius distance of approximately 110 m, and then dissimilar (negatively autocorrelated) at approximately 250 m. At the between-coupe scale, no repeatable spatial autocorrelation patterns were observed. 4. For any individual site, rapid changes in beetle density were observed to be associated with loosely aggregated flights of beetles into and out of that site. Peak adult catches (> the weekly mean plus standard deviation trap-catch) for a site occurred for a period of 2.0 +/- 0.22 weeks at a time (n = 37), with normally only one or two peaks per site per season. Peak oviposition events for a site occurred on average 1.4 +/- 0.11 times per season and lasted 1.5 +/- 0.12 weeks. 5. Analysis of an extensive data set (n = 417) demonstrated that adult abundance at a site was positively correlated with egg density, but negatively correlated with tree damage (caused by conspecifics) and the presence of conspecific larvae. There was no relationship between adult abundance and a visual estimate of the amount of young foliage on trees. 6. Adults of C. bimaculata are show n to occur in relatively small, mobile aggregations. This means that pest surveys must be both regular (less than 2 weeks apart) and intensive (with sampling points no more than 150 m apart) if beetle populations are to be monitored with confidence. Further refinement of the current IPM strategy must recognize the problems posed by this temporal and spatial patchiness, particularly with regard to the use of biological insecticides, such as Bacillus thuringiensis, for which only a very short operational window exists.

Effect of specific leaf nitrogen on radiation use efficiency and growth of sunflower

Specific leaf nitrogen (SLN, g/m(2)) is known to affect radiation use efficiency (RUE, g/MJ) in different crops, However, this association and importance have not been well established over a range of different nitrogen regimes for held-grown sunflower (Helianthus annuus L.). An experiment was conducted to investigate different combinations and rates of applied nitrogen on SLN, RUE, and growth of sunflower, A fully irrigated crop was sown on an alluvial-prairie soil (Fluventic Haplustoll) and treated with five combinations of applied nitrogen, Greater nitrogen increased biomass, grain number, and yield, but did not affect harvest index energy-corrected for oil (0.4) or canopy extinction coefficient (0.88), Decreases in biomass accumulation under low nitrogen treatments were associated,vith reductions in leaf area index (LAI) and light interception, When SLN and RUE were examined together, both were less in the anthesis to physiological maturity period, but relatively stable between bud visible and anthesis, However, the effects of canopy SLN on RUE were confounded by high SLN in the top of the canopy and the crop maintaining SLN by reducing LAI, Measurements of leaf CO2 assimilation and theoretical analyses of RUE supported that RUE was related to SLN, The major effect of nitrogen on early growth of sunflower was mediated by leaf area and the distribution of SLN in the canopy rather than direct effects of canopy SLN on RUE alone. Greater responses of RUE to SLN are more evident later in growth, and may be related to the demand of nitrogen by the grain.

Molecular characterization of the Hepatitis B virus genotypes in Colombia: A Bayesian inference on the genotype F

Hepatitis B is a worldwide health problem affecting about 2 billion people and more than 350 million are chronic carriers of the virus. Nine HBV genotypes (A to I) have been described. The geographical distribution of HBV genotypes is not completely understood due to the limited number of samples from some parts of the world. One such example is Colombia, in which few studies have described the HBV genotypes. In this study, we characterized HBV genotypes in 143 HBsAg-positive volunteer blood donors from Colombia. A fragment of 1306 bp partially comprising HBsAg and the DNA polymerase coding regions (S/POL) was amplified and sequenced. Bayesian phylogenetic analyses were conducted using the Markov Chain Monte Carlo (MCMC) approach to obtain the maximum clade credibility (MCC) tree using BEAST v.1.5.3. Of all samples, 68 were positive and 52 were successfully sequenced. Genotype F was the most prevalent in this population (77%) - subgenotypes F3 (75%) and Fib (2%). Genotype G (7.7%) and subgenotype A2 (15.3%) were also found. Genotype G sequence analysis suggests distinct introductions of this genotype in the country. Furthermore, we estimated the time of the most recent common ancestor (TMRCA) for each HBV/F subgenotype and also for Colombian F3 sequences using two different datasets: (i) 77 sequences comprising 1306 bp of S/POL region and (ii) 283 sequences comprising 681 bp of S/POL region. We also used two other previously estimated evolutionary rates: (i) 2.60 x 10(-4) s/s/y and (ii) 1.5 x 10(-5) s/s/y. Here we report the HBV genotypes circulating in Colombia and estimated the TMRCA for the four different subgenotypes of genotype F. (C) 2010 Elsevier B.V. All rights reserved.