Isolation, identification, and biocontrol of antagonistic bacterium against Botrytis cinerea after tomato harvest

ABSTRACT Tomato is one of the most important vegetables in the world. Decay after harvest is a major issue in the development of tomato industry. Currently, the most effective method for controlling decay after harvest is storage of tomato at low temperature combined with usage of chemical bactericide; however, long-term usage of chemical bactericide not only causes pathogen resistance but also is harmful for human health and environment. Biocontrol method for the management of disease after tomato harvest has great practical significance. In this study, antagonistic bacterium B-6-1 strain was isolated from the surface of tomato and identified as Enterobacter cowanii based on morphological characteristics and physiological and biochemical features combined with sequence analysis of 16SrDNA and ropB gene and construction of dendrogram. Effects of different concentrations of antagonistic bacterium E. cowanii suspension on antifungal activity after tomato harvest were analyzed by mycelium growth rate method. Results revealed that antifungal activity was also enhanced with increasing concentrations of antagonistic bacterium; inhibitory rates of 1 × 105 colony-forming units (cfu)/mL antagonistic bacterial solution on Fusarium verticillioides, Alternaria tenuissima, and Botrytis cinerea were 46.31%, 67.48%, and 75.67%, respectively. By using in vivo inoculation method, it was further confirmed that antagonistic bacterium could effectively inhibit the occurrence of B. cinerae after tomato harvest, biocontrol effect of 1 × 109 cfu/mL zymotic fluid reached up to 95.24%, and antagonistic bacterium E. cowanii has biocontrol potential against B. cinerea after harvest of fruits and vegetables.

Study of the interactions between Penicillium oxalicum Currie & Thom and Alternaria alternata (Fr. ) Keissler

The aim of this research was the analysis of the possible antagonistic effect of Penicillium oxalicum over the pathogen rice fungus A. alternata under different conditions of temperature, water activity and culture media. The macroscopic study of the dual growth revealed that according to the Index of Dominance P. oxalicum was more competitive that A. alternata at 25ºC whereas at 15ºC was this species. Microscopic analysis showed that P. oxalicum was a mycoparasite of A. alternata at all conditions tested. The antagonist penetrated into A. alternata and disintegrated its conidiophores and conidia. The results suggests that P. oxalicum may be a possible biological control agent of the rice pathogens in a future.

Biological activities of the fermentation extract of the endophytic fungus alternaria alternata isolated from coffea arabica L / Atividades biológicas do extrato da fermentação do fungo endofítico alternaria alternata isoladas de coffea arabica L

A total of 22 endophytic fungi isolated from coffee (Coffea arabica L.) were cultivated in vitro and their crude extracts tested. The screening was carried out using the agar diffusion method against Staphylococcus aureus, Escherichia coli and Candida albicans. The most effective isolate was Alternaria alternata, and subsequently, its extract was assayed. The total phenolic content was 3.44 μg GAE/mg of the crude extract. For the antibacterial and antifungal activity assays, minimum inhibitory concentrations (MIC) and minimum bactericidal and fungicidal concentrations (MBC and MFC) were determined. The ranges of MIC values were 50-100 μg/mL for S. aureus and 400-800 μg/mL for E. coli. The extract did not show activity in the tested concentrations for C. albicans. The fungal crude extract was assayed for antioxidant activities. Its ability to scavenge DPPH radicals and antioxidant activity by β-carotene/linoleic acid system oxidation was not significant. In addition, antitumor activity was studied using the MTT assay. At a dilution of 400 μg/mL, the extract displayed a cytotoxic activity of approximately 50 percent towards HeLa cells in vitro. The results indicate that endophytic fungi could be a promising source of bioactive compounds and warrant further study.

Total de 22 fungos endofíticos isolados de café (Coffea arabica L.) foi cultivado in vitro e seus extratos testados. A triagem foi conduzida pelo método de difusão em agar contra bactérias Gram-positiva, Gram-negativa e uma levedura. O isolado mais efetivo foi Alternaria alternata e, subsequentemente, seu extrato foi analisado. O conteúdo de fenólicos totais do extrato bruto foi de 3,44 μg EAG/mg de extrato. Para os testes de atividade antimicrobiana, a concentração inibitória mínima (CIM) e concentração bactericida e fungicida mínima (CBM e CFM) contra Staphylococcus aureus, Escherichia coli e Candida albicans foram determinadas. Resultados da CIM variaram entre 50-100 μg/mL para S. aureus e 400-800 μg/mL para E. coli. O extrato bruto não apresentou atividade nas concentrações testadas para C. albicans. Foram analisadas as atividades antioxidantes do extrato bruto. Sua habilidade para seqüestrar radicais DPPH e a atividade antioxidante pela oxidação do sistema β-caroteno/ácido linoléico não foram significativas. Além disso, a atividade antitumoral foi estudada pelo teste do MTT. À diluição de 400 μg/mL, o extrato apresentou atividade de aproximadamente 50 por cento sobre as células HeLa in vitro. Os resultados indicam que fungos endófitos poderiam ser uma fonte promissora de compostos bioativos necessitando de estudos futuros.

Hongos anemófilos: viabilidad y alergenicidad / Anemophilous fungi: viability and allergenicity

Se estudiaron tres cepas de hongos anemófilos de distintos géneros, que fueron: Alternaria, Rhizopus y Monilia. Encontramos que el medio de cultivo utilizado en esta investigación (Sabouraud líquido) perdía nutrientes a partir de los siete días del crecimiento de las cepas, pero sin afectarse la viabilidad de las mismas. Esto se comprobó por resiembras a los 15 y 60 días. Se elaboraron extractos alergénicos con micelios de esos hongos con siete, 15 y 60 días de crecimiento y se probaron dichos extractos en pacientes supuestamente sensibilizados, por medio de pruebas cutáneas por intradermorreacción, no encontrando diferencias significativas entre los habones obtenidos con estas pruebas. Se recomiendan investigaciones por periodos más prolongados (90,120,180 ó más días) y ver si ello incide en la viabilidad y alergenicidad de los hongos anemófilos.