Iraildes P. Assunção

Circular DNA genomics (circomics) exemplified for geminiviruses in bean crops and weeds of northeastern Brazil

Circomics was coined to describe the combination of rolling circle amplification (RCA), restriction fragment length polymorphism (RFLP) and pyro-sequencing to investigate the genome structures of small circular DNAs. A batch procedure is described using 61 plant samples from Asia, South America and Central America which revealed 83 contig sequences of geminiviral DNA components and 4 contig sequences of DNA satellites. The usefulness of this approach is validated for the Brazilian begomoviruses, and the sequence fidelity is determined by comparing the results with those of conventional cloning and sequencing of Bolivian begomoviruses reported recently. Therefore, circomics has been proven to be a major step forward to economize costs and labor and to characterize reliably geminiviral genomes in their population structure of the quasispecies.

Species of Lasiodiplodia associated with papaya stem-end rot in Brazil

This study aims to identify and characterize species of Lasiodiplodia associated with stem-end rot of papaya in six different populations in the Northeast of Brazil. Fungal identifications were made using a combination of morphology together with a phylogenetic analysis based on partial translation elongation factor 1-α sequence (EF-1α) and internal transcribed spacers (ITS). Five species of Lasiodiplodia were identified: Lasiodiplodia brasiliense sp. nov., L. hormozganensis, L. marypalme sp. nov., L. pseudotheobromae and L. theobromae. Only L. theobromae had previously been reported in papaya, while all the other species are reported for the first time in association with this host in Brazil and worldwide. Lasiodiplodia theobromae was the most prevalent species. All species of Lasiodiplodia were pathogenic on papaya fruit, with L. hormozganensis being the most virulent.

High genetic variability and recombination in a begomovirus population infecting the ubiquitous weed Cleome affinis in northeastern Brazil

Diseases caused by begomoviruses are a serious constraint to crop production in many tropical and subtropical areas of the world, including Brazil. Begomoviruses are whitefly-transmitted, single-stranded DNA viruses that are often associated with weed plants, which may act as natural reservoirs of viruses that cause epidemics in crop plants. Cleome affinis (family Capparaceae) is an annual weed that is frequently associated with leguminous crops in Brazil. Samples of C. affinis were collected in four states in the northeast of Brazil. Analysis of 14 full-length DNA-A components revealed that only one begomovirus was present, with 91-96% identity to cleome leaf crumple virus (ClLCrV). In a phylogenetic tree, ClLCrV forms a basal group relative to all other Brazilian begomoviruses. Evidence of multiple recombination events was detected among the ClLCrV isolates, which also display a high degree of genetic variability. Despite ClLCrV being the only begomovirus found, its phylogenetic placement, high genetic variability and recombinant nature suggest that C. affinis may act as a source of novel viruses for crop plants. Alternatively, ClLCrV could be a genetically isolated begomovirus. Further studies on the biological properties of ClLCrV should help to clarify the role of C. affinis in the epidemiological scenario of Brazilian begomoviruses.

Further molecular characterization of weed-associated begomoviruses in Brazil with an emphasis on Sida spp

Begomoviruses are whitefly-transmitted, single-stranded DNA viruses that are often associated with weed plants. The aim of this study was to further characterize the diversity of begomoviruses infecting weeds (mostly Sida spp.) in Brazil. Total DNA was extracted from weed samples collected in Vicosa (Minas Gerais state) and in some municipalities of Alagoas state in 2009 and 2010. Viral genomes were amplified by RCA, cloned and sequenced. A total of 26 DNA-A clones were obtained. Sequence analysis indicated the presence of 10 begomoviruses. All viral isolates from Blainvillea rhomboidea belonged to the same species, Blainvillea yellow spot virus (BlYSV ), thereby suggesting that BlYSV may be the only begomovirus present in this weed species. Four isolates represent new species, for which the following names are proposed: Sida yellow blotch virus (SiYBV), Sida yellow net virus (SiYNV), Sida mottle Alagoas virus (SiMoAV) and Sida yellow mosaic Alagoas virus (SiYMAV). Recombination events were detected among the SiYBV isolates and in the SiYNV isolate. These results constitute further evidence of the high species diversity of begomoviruses in Sida spp. However, the role of this weed species as a source of begomoviruses infecting crop plants remains to be determined.

Reaction of faba bean genotypes to Rhizoctonia solani and resistance stability

The production of faba bean (Phaseolus lunatus) is limited due to the occurrence of Rhizoctonia canker, caused by the fungus Rhizoctonia solani. In order to select materials with potential for use in faba beans breeding programs and/or disease integrated management, 72 genotypes were evaluated and also stability of the disease resistance in relation to different pathogen isolates, inoculum densities and soil type. The seeds were sown in soil infested with the pathogen by the addition of colonized substrate (rice grains). The evaluation of genotypes was done after 15 days, using a note scale to discriminate the genotypes into five reaction classes. None of the 72 genotypes showed immunity to R. solani and only four genotypes (F-42, F-49, F-53 and F-58) behaved as highly resistant. The reactions presented by these genotypes varied according to the R. solani isolate, inoculum density and soil used. The genotype F-58 showed more stable resistance to pathogen isolates and inoculum densities. The genotype F-53 showed resistance stability in soils collected in different locations, remaining highly resistant in all situations. The genotype F-58 did not show high resistance to only one tested soil. Therefore, the genotypes F-58 and F-53 are promising sources of resistance to R. solani and should be indicated in areas infested by the pathogen.

Variabilidade genética de isolados de badnavírus infectando inhame (Dioscorea spp.) no nordeste do Brasil

Viroses causadas por virus do genero Badnavirus sao responsaveis por grandes prejuizos a cultura do inhame no Nordeste brasileiro. O conhecimento da variabilidade destes patogenos pode fornecer informacoes importantes sobre seu potencial evolutivo, permitindo a elaboracao de melhores estrategias de manejo da doenca. A analise de 425 amostras foliares de inhame coletadas em tres estados do Nordeste brasileiro, em 2010, revelou uma alta incidencia (93,3%) de badnaviroses. Para avaliar a variabilidade genetica dos badnavirus infectando inhame, um fragmento de 579 nucleotideos correspondente a regiao codificante da transcriptase reversa (RT)/RNaseH dos isolados amostrados foi amplificada por PCR e sequenciada. A analise filogenetica das sequencias de nucleotideos revelou que os isolados dividem-se em dois grupos. Um e altamente relacionado com Dioscorea bacilliform AL virus (DBALV), enquanto o outro forma um clado altamente divergente dentro do genero Badnavirus. Os isolados de DBALV apresentam 70-98% de identidade nucleotidica entre si e foram detectados em todas as areas avaliadas e em D. alata e D. cayennensis, as duas especies de inhame mais cultivadas no Nordeste. Os isolados do outro grupo compartilham 47-58% de identidade com isolados de DBALV e 78-95% entre si e foram encontrados apenas em D. alata na Paraiba.

Genes diferentes podem conferir resistência ao Cowpea severe mosaic virus em caupi

Cowpea (Vigna unguiculata) is an important vegetable crop in Northeast Brazil and has been traditionally cultivated by small farmers. Virus diseases are considered to be the main factor in yield limiting cowpea yield in the region. The severe mosaic disease caused by Cowpea severe mosaic virus (CpSMV), family Comoviridae, genus Comovirus, seems to be one of the most prevalent diseases responsible for high losses. Resistant cultivars may be considered the best alternative for disease control, and several promising sources of resistance such as Macaibo, CNC 0434 cultivars, and the line L 254.008 have been reported. More recent investigations into the genetic basis of these resistant plant genotypes have pointed to similar results, and the inheritance of this resistance has been recognized as monogenic recessive. On the other hand, any research aimed to investigate if the resistance genes are alleles or not. In the present investigation a test was conducted to elucidate this question. Plants of the genotypes Macaibo, CNC 0434 and L 254.008 were bred in reciprocal and direct ways producing six populations. Evaluation of those cowpea genotypes using one isolate of CpSMV indicated that the resistance gene of Macaibo is the same as that for CNC 0434 and distinct from the gene L 254.008 found in that line.

Colletotrichum species causing anthracnose on lima bean in Brazil

The taxonomy of the genus Colletotrichum has undergone various changes. These alterations arise from the use of modern molecular tools. Currently, Colletotrichum species are grouped into complexes. Species of Colletotrichum associated with diseases in lima bean have been investigated very little. In Brazil, anthracnose in lima bean has often been associated with Colletotrichum truncatum, but only morphological characteristics have been used for identification purposes. In this study, samples of lima bean with symptoms of spots on the leaves and pods have been collected in Piauí and Alagoas states. Comparisons of morphological characteristics revealed nine isolates with cylindrical or curved conidia. The ITS region and partial sequences of GAPDH, β-tubulin and Actin regions were amplified by PCR, sequenced and submitted to multilocus phylogenetic analysis. The isolates analyzed grouped with reference specimens from Colletotrichum truncatum, C. cliviae and C. fructicola. All isolates were pathogenic to 25-day-old lima bean seedlings. Therefore, this work confirms the pathogenicity of C. truncatum to lima bean and for the first time records the occurrence of C. cliviae and C. fructicola as pathogens of this host in Brazil.

Influência da intensidade da murcha-de-fusário no rendimento do caupi

A murcha-de-fusario, causada por Fusarium oxysporum f.sp. tracheiphilum, e uma importante doenca do caupi (Vigna unguiculata) no Nordeste brasileiro. Foi conduzido um experimento em microparcelas para avaliar a influencia da intensidade da murcha-de-fusario nas reducoes de rendimento do caupi. O solo acondicionado nas microparcelas foi infestado com a mistura do inoculo de dois isolados do patogeno, obtendo-se diferentes gradiente de densidade, variando de 1,2 x 102 a 5,3 x 104 ufc/g. Cinco dias apos a infestacao, foram plantadas as cultivares BR-17 Gurgueia e IPA-206, classificadas como altamente e moderadamente suscetiveis a F. oxysporum f.sp. tracheiphilum, respectivamente. Na fase de colheita, o rendimento de cada cultivar foi determinado por microparcela, obtendo-se o numero de vagens por planta, o peso total de sementes por planta e o peso de 100 sementes. Apos a colheita, a severidade da murcha-de-fusario foi avaliada em todas as plantas com o auxilio de uma escala descritiva, sendo calculado o indice de intensidade da doenca. Nao foram verificadas diferencas significativas (P=0,05) na intensidade de doenca e na taxa de reducao de rendimento entre as duas cultivares. As reducoes no rendimento de sementes variaram de 9,11 a 80,30% e de 8,30 a 86,51% nas cultivares BR-17 Gurgueia e IPA-206, respectivamente. O modelo de regressao linear simples, sem a transformacao dos dados, possibilitou o ajuste adequado aos dados na analise da relacao entre a intensidade da murcha-de-fusario e as reducoes de rendimento das duas cultivares.

Molecular and morpho-cultural characterization of Colletotrichum spp. associated with anthracnose on Capsicum spp. in northeastern Brazil

Anthracnose, caused by Colletotrichum spp., is one of the major fungal diseases that affect Capsicum spp. and leads to severe crop losses. The aim of this study was to identify species within Colletotrichum that cause anthracnose in bell and hot pepper using phylogenetic and morphological criteria. Pathogenic isolates were obtained from bell and hot peppers exhibiting anthracnose symptoms and grown in the northeastern region of Brazil. These were initially identified based on the sequences of GAPDH, ACT, CAL, β-TUB genes and the ITS-rDNA region. Mycelial growth, coloration of the colony, size and shape of conidia and appressoria were determined for the selected isolates. It was concluded that five species: C. brevisporum, C. scovillei, C. siamense, C. tropicale and C. truncatum compose the species complex associated with anthracnose of Capsicum spp. in northeastern Brazil.