Acidovorax citruli: genetic analyses and protocol for its detection in seeds

Bacterial fruit blotch of cucurbits (BFB), caused by the seed borne Gramnegative bacterium Acidovorax citrulli is a serious threat to cucurbit industry worldwide. Since late 1980`s after devastating outbreaks in watermelon fields in southern United States, BFB has spread worldwide and has been reported in other cucurbit crops such as melon, pumpkin, cucumber and squash. To date, there is evidence for the existence of at least two genetically and pathogenically distinct populations of A. citrulli. In Brazil, the first report of BFB was in 1991, in a watermelon field in São Paulo. Although widespread in the country, BFB has been a major problem to melon production. More precisely, BFB has caused significant yield losses to melon production in northeastern Brazil, which concentrates > 90% of the country`s melon production. Despite the management efforts and the recent advances in A. citrulli research, BFB is still a continuous threat to the cucurbit industry, including seed producers, growers and transplant nurseries. To better understand the population structure of A. citrulli strains in Brazil, and to provide a basis for the integrated management of BFB, we used pulsed-field gel electrophoresis (PFGE), multilocus sequence analysis (MLSA) of housekeeping and virulence-associated genes and pathogenicity tests on different cucurbit seedlings to characterize a Brazilian population of A. citrulli strains from different hosts and regions. Additionally, we conducted for the first time a comparative analysis of the A. citrulli group I and II population at genomic level and showed that these two groups differ on their genome sizes due to the presence of eight DNA segments, which are present in group II and absent in group I genomes. We also provide the first evidence to suggest that temperature might be a driver in the ecological adaptation of A. citrulli populations under nutrient-rich or -depleted conditions. Finally, in order to improve the routine detection of A. citrulli on melon seedlots, we designed a new primer set that is able to detect the different Brazilian haplotypes, thus minimizing the risk of false-negatives on PCR-based seed health testing.

Simbioses de rizóbios com leguminosas arbóreas na pré-amazônia maranhense

The tree leguminous gliricídia (Gliricidia sepium), acácia (Acacia mangium), leucena (Leucaena leucocephala) and sombreiro (Clitoria fairchildiana) are indicated for agroforestry systems, reclamation of degraded lands, reforestation and other purposes in the wet tropic. Despite the importance of legumes the preamazon region it is lacking in information about the symbiotic capacity and diversity of indigenous rhizobia of these legumes. The aim of this work was to evaluate the phenotypic and genetic diversity of rhizobia species nodulating gliricidia sombreiro, leucena and acacia in the Maranhão pre-Amazon region and authenticate isolates of these species in siratro (Macroptilium atropurpureum). For this they were installed two experiments. Sampling was carried out on a alley cropping system, was sampled 20 plants of each species by collecting 10 nodules per plant. It was made isolation, cultural characterization, partial 16S rRNA gene sequencing and analysis of the symbiotic ability of bacterial strains with siratro. The authentication experiment was done in two steps for each legume (gliricidia, acácia, sombreiro and leucena), in the greenhouse and in a completely randomized design with three replications with sterile Hoagland nutrient solution as substrate. Gliricídia, Sombreiro, leucena and acacia are colonized by distinct groups of rhizobia. Gliricidia nodulate preferably with Rhizobium, sombreiro and acacia nodulate preferably with Bradyrhizobium and leucena has Mesorhizobium main symbiote. Endophytic strains of ten genera were found colonizing the gliricidia, sombrero, leucena and acacia nodules and a strain of Arthrobacter sp. had a positive nodulation with siratro. This is the first report on isolation of Methylobacterium sp. in gliricidia nodules and endophytic ability of Terriglobus sp. strains. Indigenous strains of pre-Amazon region of Bradyrhizobium, Mesorhizobium and Rhizobium genus nodulate with siratro, but are ineficiente or had low efficiency to promote their growth.