Morphological and molecular characterisation of Streptomyces spp. which suppress pathogenic fungi

Streptomyces species are aerobes and chemoorganotrophic bacteria. These microorganisms produce a wide range of industrially significant compounds, specifically antibiotics and anti fungal substances. The objective of this study was to characterise soil-borne Streptomyces isolates using morphological and molecular traits in order to identify them to species level, and leverage from their potential to suppress the growth of Aspergillus flavus , Fusarium oxysporum and Penicillium italicum . Twenty-seven soil-borne putative Streptomyces, which elicited comprehensive\ua0antimicrobial activity against Aspergillus flavus, Fusarium oxysporum and Penicillium italicum, in a previous study, were evaluated. On the basis of morphology, the bacteria resembled the genus Streptomyces. Initially, colonies phenotypically appeared to have a relatively smooth surface but as growth progressed the bacteria developed a weft of aerial mycelium granular, powdery or velvety in appearance. Bacteria produced a wide variety of pigments which in turn were responsible for the colour of the vegetative and aerial mycelia, colour ranged from white to cream or buff shades and yellow to orange or brown. Microscopic analyses and morphological characteristics generated sub-groups of the isolates and clustered them according to their similarities. One bacterial strain was randomly selected from each cluster and investigated using molecular characteristics. Partial 16S rDNAs from the selected representative isolates from each subgroup, were sequenced and phylogenetic analysis performed. The 16S\ua0rDNA\ua0sequences of the isolates indicated that they were related to\ua0 Streptomyces species: S. bungoensis , S. thermocarboxydus , S. corchorusii and S. lasaliensis, that are known secondary metabolite producers possessing antimicrobial activity against plant pathogens.Les esp\ue8ces de Streptomyces sont des bact\ue9ries a\ue9robies et chimio-organotrophes. Ces micro-organismes produisent une large gamme de compos\ue9s d\u2019importance industrielle, en particulier des antibiotiques et des substances antifongiques. L\u2019objectif de cette \ue9tude \ue9tait de caract\ue9riser les isolats de Streptomyces transmis par le sol \ue0 l\u2019aide de traits morphologiques et mol\ue9culaires afin de les identifier au niveau de l\u2019esp\ue8ce, et de tirer parti de leur potentiel \ue0 supprimer la croissance d\u2019 Aspergillus flavus , Fusarium oxysporum et Penicillium italicum . Vingt-sept Streptomyces putatifs transmis par le sol, qui ont suscit\ue9 une activit\ue9 antimicrobienne compl\ue8te contre Aspergillus flavus, Fusarium oxysporum et Penicillium italicum, dans une \ue9tude pr\ue9c\ue9dente, ont \ue9t\ue9 \ue9valu\ue9s. Sur la base de la morphologie, les bact\ue9ries ressemblaient au genre Streptomyces. Au d\ue9part, les colonies semblaient ph\ue9notypiquement avoir une surface relativement lisse, mais au fur et \ue0 mesure que la croissance progressait, les bact\ue9ries d\ue9veloppaient une trame de myc\ue9lium a\ue9rien d\u2019aspect granuleux, poudreux ou velout\ue9. Les bact\ue9ries produisaient une grande vari\ue9t\ue9 de pigments qui \ue0 leur tour \ue9taient responsables de la couleur des myc\ue9liums v\ue9g\ue9tatifs et a\ue9riens, la couleur variait du blanc au cr\ue8me ou au chamois et du jaune \ue0 l\u2019orange ou au brun. Des analyses microscopiques et des caract\ue9ristiques morphologiques ont g\ue9n\ue9r\ue9 des sous-groupes d\u2019isolats et les ont regroup\ue9s en fonction de leurs similitudes. Une souche bact\ue9rienne a \ue9t\ue9 s\ue9lectionn\ue9e au hasard dans chaque groupe et \ue9tudi\ue9e en utilisant des caract\ue9ristiques mol\ue9culaires. Des ADNr 16S partiels provenant des isolats repr\ue9sentatifs s\ue9lectionn\ue9s de chaque sous-groupe ont \ue9t\ue9 s\ue9quenc\ue9s et une analyse phylog\ue9n\ue9tique a \ue9t\ue9 effectu\ue9e. Les s\ue9quences d\u2019ADNr 16S des isolats ont indiqu\ue9 qu\u2019ils \ue9taient apparent\ue9s aux esp\ue8ces de Streptomyces: S. bungoensis , S. thermocarboxydus , S. corchorusii et S. lasaliensis, qui sont des producteurs de m\ue9tabolites secondaires connus poss\ue9dant une activit\ue9 antimicrobienne contre les phytopathog\ue8nes

In vitro suppression of pathogenic fungi by Streptomyces spp.

The use of living organisms or natural enemies of pathogens to control their populations is called biological disease control. It involves harnessing and introduction of exotic species of microorganism in a natural form, with the intention of controlling pathogens that may exist naturally in the same ecosystem. Prospects for biological control of Aspergillus flavus , Fusarium oxysporum and Penicillium italicum were investigated using Streptomyces spp. isolated from Chinhoyi University of Technology Farm soils in Mashonaland West, Zimbabwe. Twenty seven Streptomyces spp were obtained from the soil, and screened for antimicrobial activity and antagonism in in vitro pathogen inhibition assays, replicated thrice. Although majority of the isolates tested elicited no effect on test pathogens, 22% of the Streptomyces isolates were able to effectively suppress A. flavus, F. oxysporum and P. italicum by at least 55%. There was a significant interaction between Streptomyces isolates and pathogen (A. flavus, F. oxysporum and P. italicum) (P<0.05) on fungal radial growth at days 7, 10 and 14 after pathogen-Streptomyces incubation. Antimicrobial potential against individual and multiple test pathogens was observed, with CUT-Streptomyces 4, CUT-Streptomyces 10, CUT-Streptomyces 11, CUT-Streptomyces 20 and CUT-Streptomyces 23 showing the greatest antimicrobial activity. CUT-Streptomyces isolates have the potential to suppress A. flavus, F. oxysporum and P. italicum in vitro.L\u2019utilisation d\u2019organismes vivants ou d\u2019ennemis naturels d\u2019agents pathog\ue8nes pour contr\uf4ler leurs populations est appel\ue9e contr\uf4le biologique des maladies. Il s\u2019agit d\u2019exploiter et d\u2019introduire des esp\ue8ces exotiques de microorganismes sous une forme naturelle, dans le but de contr\uf4ler les agents pathog\ue8nes pouvant exister naturellement dans le m\ueame \ue9cosyst\ue8me. Les perspectives de lutte biologique contre Aspergillus flavus , Fusarium oxysporum et Penicillium italicum ont \ue9t\ue9 \ue9tudi\ue9es \ue0 l\u2019aide de Streptomyces spp. isol\ue9es des sols agricoles de l\u2019Universit\ue9 de Technologie de Chinhoyi \ue0 Mashonaland West, Zimbabwe. Vingt-sept Streptomyces spp ont \ue9t\ue9 obtenus du sol et cribl\ue9s pour l\u2019activit\ue9 antimicrobienne et l\u2019antagonisme dans des tests d\u2019inhibition des agents pathog\ue8nes in vitro, r\ue9pliqu\ue9s trois fois. Bien que la majorit\ue9 des isolats test\ue9s ne provoquent aucun effet sur les agents pathog\ue8nes test\ue9s, 22% des isolats de Streptomyces sont capables de supprimer efficacement au moins 55% des A. flavus, F. oxysporum et P. italicum. Il y avait une interaction significative entre les isolats de Streptomyces et l\u2019agent pathog\ue8ne (A. flavus, F. oxysporum et P. italicum) (P< 0.05) lors de la croissance radiale des champignons aux 7\ue8me, 10\ue8me et 14\ue8me jours apr\ue8s l\u2019incubation de l\u2019agent pathog\ue8ne-Streptomyces. Un potentiel antimicrobien contre des agents pathog\ue8nes individuels et multiples a \ue9t\ue9 observ\ue9. CUT-Streptomyces 4, CUT-Streptomyces 10, CUT-Streptomyces 11, CUT-Streptomyces 20 et CUT-Streptomyces 23 ont montr\ue9 l\u2019activit\ue9 antimicrobienne la plus \ue9lev\ue9e. Les isolats de CUT-Streptomyces ont le potentiel de supprimer A. flavus, F. oxysporum et P. italicum in vitro

Detecção e análise da variabilidade de seqüências do Banana streak virus (BSV) em bananeiras no Brasil Detection and analysis of Banana streak virus (BSV) sequences variability of banana from Brazil

A técnica de PCR utilizando-se "primers" degenerados para o gênero Badnavirus foi utilizada para a detecção e análise da variabilidade de seqüências do Banana streak virus (BSV) provenientes de bananeiras. A partir desta metodologia seqüências do vírus puderam ser detectadas em cultivares diplóides (AA), triplóides (AAA; AAB) e tetraplóides (AAAB). Foram encontrados quatro padrões de seqüência do BSV (estirpes BSVBR-1, BSVBR-2, BSVBR-3 e BSVBR-4), diferenciadas através da análise do perfil eletroforético das amostras amplificadas. A estirpe BSVBR-1 prevalece nos estados do Acre, Amazonas, Bahia, Ceará, Goiás, Minas Gerais, Piauí, Rio de Janeiro, Rondônia, Santa Catarina, e São Paulo, enquanto que, a estirpe BSVBR-2 foi encontrada em amostras oriundas do Amazonas e do Ceará. As estirpes BSVBR-3 e BSVBR-4 foram encontradas apenas no Ceará. Este trabalho revela a presença de diferentes estirpes do BSV no Brasil, bem como a existência de cultivares de bananeiras sadias e livres de seqüências virais do BSV integradas ao seu genoma.<br>PCR assay using degenerate primers, designed to Badnavirus genus, was used to detect and analyse the variability of BSV strains sequences from banana. The virus was detected in diploid (AA), triploids (AAA; AAB) and tetraploids (AAAB) banana cultivars. Four BSV sequences patterns (BSVBR-1, BSVBR-2, BSVBR-3 and BSVBR-4 strains) were found, and distinguished by eletrophoresis. The strain BSVBR-1 was found in the states of Acre, Amazonas, Bahia, Ceará, Goiás, Minas Gerais, Piauí, Rio de Janeiro, Rondônia, Santa Catarina and São Paulo, while BSVBR-2 strain was detected in the states of Amazonas and Ceará. BSVBR-3 and BSVBR-4 strains were found only in the state of Ceará. This work demonstrated the presence of different BSV strains in Brazil and the existence of health banana cultivars as well as cultivars free of BSV integrated sequences

Identification of Rubus yellow net virus as a distinct badnavirus and its detection by PCR in Rubus species and in aphids

Rubus yellow net virus (RYNV) infects Rubus species and cultivars worldwide and is an essential component of raspberry veinbanding mosaic (RVBMD), a virus disease complex that causes serious decline in plant vigour and productivity. The virus is transmitted, probably in a semi-persistent manner, by the large raspberry aphid, Amphorophora idaei in Europe, and A. agathonica in North America. The particles of RYNV are bacilliform in shape and measure 80-150 × 25-30 nm, similar to those of badnaviruses. A 1.7 kb fragment of the viral DNA was amplified by PCR and then directly sequenced. Analysis of this sequence suggests that RYNV is possibly a distinct species in the genus Badnavirus and is most closely related to Gooseberry vein banding associated virus (GVBAV) and Spiraea yellow leaf spot virus, two other badnaviruses described recently. Using the sequence derived from the PCR-amplified viral DNA fragment, RYNV-specific primers were designed and used in PCR to assay for RYNV in a range of Rubus germplasm infected with RYNV, with other unrelated viruses and virus-like diseases found in Rubus, and in healthy plants. RYNV was detected in all glasshouse cultures of RYNV-infected plants, whether alone or in complex infections with other viruses, but not from healthy Rubus plants, nor from plants infected with other viruses. It was also detected in field-grown raspberry plants with and without symptoms of RVBMD and in raspberry plants infected with RYNV by viruliferous A. idaei. RYNV was also detected by PCR in A. idaei following access feeds on RYNV-infected plants of 1 h or more. PCR failed to amplify DNA from gooseberry infected with GVBAV confirming the specificity of the RYNV analysis. PCR detection of RYNV in dormant raspberry buds allows assays to be made outside the natural growing season, providing a useful application for plant introduction and quarantine programmes