Development of methodologies for virus detection in soybean and wheat seeds.

Seeds that contain large amounts of oil, starch, fibers and phenols are the most difficult tissues for RNA extraction. Currently, there are some reports of virus detection in seeds using commercial kits for RNA extraction. However, individual seeds were used, which may not be always suitable for analyses that deal with large amounts of seeds. Sangha [1] described a simple, quick and efficient protocol for RNA extraction and downstream applications in a group of seeds of jatropha (Jatropha curcas), mustard (Brassica sp.) and rice (Oryza sativa). We tested this protocol for soybean (Glycine max), maize (Zea mays), wheat (Triticum aestivum) and triticale (×Triticosecale) seeds and further reverse transcription PCR (RT-PCR)/quantitative real-time PCR (qPCR) in order to have a faster and more practical method for virus detection from seeds than the traditional scheme of seed planting and subsequent Elisa/RT-PCR from leaves. The essential points in the method are:•Some modifications in the protocol [1] were done in order to increase performance: Wheat and triticale seeds are incubated with water prior to maceration. An amount of 1.2 g of dry soybean seeds is used to maceration.•RT-PCR is used for detection of Wheat streak mosaic virus from wheat seeds and RT-qPCR for detection of Soybean mosaic virus from soybean seeds.•The method may be tested for other viruses, however, pre-validation will be needed.

HIV seroprevalence and high-risk sexual behavior among female sex workers in Central Brazil.

Female sex workers (FSWs) are considered a high-risk group for human immunodeficiency virus (HIV) infection due to their social vulnerability and factors associated with their work. We estimated the prevalence of HIV, and identified viral subtypes and risk factors among FSWs. A cross-sectional study using respondent-driven sampling (RDS) method was conducted among 402 FSWs in Campo Grande city, Brazil, from 2009 to 2011. Participants were interviewed using a standardized questionnaire about sociodemograpic characteristics and risk behavior. Blood samples were collected for serological testing of HIV. Of the 402 FSWs, median age and age of initiating sex work were 25 years (Interquartile range [IQR]: 9) and 20 years (IQR: 6), respectively. The majority reported use of alcohol (88.5%), had 5-9 years (median: 9; IQR: 3) of schooling (54.5%), 68.6% had tattoos/body piercings, and 45.1% had more than seven clients per week (median: 7; IQR: 10). Only 32.9% of FSW reported using a condom with nonpaying partners in the last sexual contact. Prevalence of HIV infection was 1.0% (95% CI: 0.1-2.6%). Genotyping for HIV-1 performed on three samples detected subtypes B, C, and F1. Sex work in the Midwestern region of Brazil is characterized by reduced education, large numbers of clients per week, and inconsistent condom use, mainly with nonpaying partners. Although prevalence of HIV infection is currently low, elevated levels of high-risk sexual behavior confirm a need to implement prevention measures. Specific interventions targeting FSWs must emphasize the risk associated with both clients and nonpaying partners while providing knowledge about HIV prevention.

Further evidence reveals that okra mottle virus arose from a double recombination event.

As a result of surveys of okra begomoviruses (genus Begomovirus, family Geminiviridae) conducted over the last five years in Central Brazil, we report the complete genome sequence of an isolate of okra mottle virus (OMoV). The DNA-A and DNA-B components were 2660 and 2653 nucleotides (nt) long, respectively, and they were most closely related to the DNA-A (~99 % nt identity) and DNA-B (~98 % nt identity) components of an OMoV isolate from a soybean plant. A phylogenetic tree was generated based on these sequences, and it was shown that both of the OMoV DNA components were grouped in a branch with Brazilian begomoviruses known to infect weeds. By recombination analysis, strong evidence was observed that the OMoV genome may have been the product of a double inter-species recombination event.

Further characterization of tomato-infecting begomoviruses in Brazil.

Tomato cultivation in Brazil is threatened by a number of tomato-infecting viruses belonging to the genus Begomovirus of the family Geminiviridae. Here, we report the full DNA-A sequences of three Brazilian begomoviruses: a potentially new tomato-infecting viruses, tomato interveinal chlorosis virus (ToICV), and two previously proposed begomoviruses for which only partial DNA-A sequences are available in the databases: tomato mottle leaf curl virus (TMoLCV) and tomato golden vein virus (TGVV). The complete sequences of the DNA-B components of TMoLCV and TGVV and the DNA-A components of a number of tomato severe rugose virus variants are also presented. Collectively, all of the analyzed sequences were phylogenetically clustered within the two major groups of Brazilian tomato-infecting begomoviruses.

Molecular and biological characterization of a new Brazilian begomovirus, euphorbia yellow mosaic virus (EuYMV), infecting Euphorbia heterophylla plants.

To date, no begomovirus has been fully characterized from Euphorbia heterophylla, a widely distributed weed, in Brazil. Here, we show the occurrence of a new begomovirus on E. heterophylla plants showing bright yellow mosaic. The bipartite viral genome was cloned from 10 samples, and all clones are almost identical to each other (95.6-98.8% nucleotide sequence identity). The DNA-A sequences shared a maximum nucleotide sequence identity of 87.3% with euphorbia mosaic Peru virus (EuMPV) and thus were classified as belonging to a novel begomovirus species, tentatively named Euphorbia yellow mosaic virus (EuYMV). The EuYMV DNA-B sequences share a maximum nucleotide sequence identity of 56.2% with a euphorbia mosaic virus (EuMV) isolate from Mexico. Phylogenetic analysis demonstrated that this new virus belongs to a different lineage than EuMV isolates from Central America.

Three distinct begomoviruses associated with soybean in central Brazil.

We report the complete nucleotide sequences of geminiviruses of the genus Begomovirus infecting soybean (Glycine max) in central Brazil. Samples obtained from soybean plants collected at Santo Antonio de Goiás, Goiás State, showing typical symptoms of viral infection, were analyzed. Infection was confirmed by PCR-based amplification of a DNA-A fragment with universal begomovirus primers. Total DNA from infected plants was then subjected to rolling-circle amplification (RCA), and 2.6-kb molecules were cloned into plasmid vectors. Sequencing of the three DNA-A and two DNA-B clones thus obtained confirmed infection by three distinct begomoviruses: bean golden mosaic virus, Sida micrantha mosaic virus and okra mottle virus, the last of which was reported recently to be a novel virus infecting okra plants in Brazil. Begomovirus infection of soybean plants has been reported sporadically in Brazil and has generally not been considered to be of economic relevance.