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Fungal pathogens are one of the most important agents affecting crop production and food safety, and agrochemical application is one of the main approaches to reduce phytopathogenic fungi contamination in agricultural products. However, excessive and inadequate use can cause environmental damage, human and animal hazard, and increased phytopathogen resistance to fungicides. Biological control using lactic acid bacteria (LAB) and Bacillus spp. is an environmentally friendly strategy for phytopathogenic fungi management. Several molecules produced by these bacteria indeed affect fungal growth and viability in different plant crops. In this article, the activity spectra are reviewed along with the antifungal effect and antifungal compounds produced by LAB (e.g. organic acids, peptides, cyclic dipeptides, fatty acids, and volatile compounds) and Bacillus spp. (e.g. peptides, enzymes, and volatile compounds).

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In recent decades, various bacterial species have been characterized as biocontrol agents for plant crop diseases; however, only a few genera have been predominantly reported in the literature. Therefore, the identification of new antagonists against phytopathogens is essential for boosting sustainable food production systems. In this study, we evaluated the role of strain SER3 from the recently discovered Rouxiella badensis as a biocontrol agent. SER3 was isolated from the phyllosphere of decaying strawberry fruit (Fragaria × ananassa) and showed different grades of antagonism against 20 fungal pathogens of berries, based on confrontation assays, due to the action of its diffusible and volatile compounds. These fungal pathogens were isolated from decayed strawberry, blackberry, and blueberry fruit and were characterized through internal transcribed spacer (ITS) sequencing and homology searches, exhibiting similarity with well-known postharvest pathogens such as Botrytis, Fusarium, Geotrichum, Mucor, Penicillium, Alternaria, and Botryosphaeria. Koch's postulates were confirmed for most pathogens by reinfecting berry fruit. SER3 showed good capacity to inhibit the growth of Botrytis cinerea and Fusarium brachygibbosum in strawberry fruit, affecting mycelial development. To gain better understanding of the genetic and metabolic capacities of the SER3 strain, its draft genome was determined and was found to comprise a single chromosome of 5.08 Mb, 52.8% G + C content, and 4,545 protein-coding genes. Phylogenetic analysis indicated that the SER3 strain is affiliated with the R. badensis species, with an average nucleotide identity >96% and a genome-to-genome distance >70%. A comparison of the genomic properties of R. badensis SER3 and other close bacterial relatives showed several genes with potential functions in biocontrol activities, such as those encoding siderophores, non-ribosomal peptide synthetases, and polyketide synthases. This is the first study to demonstrate a novel role of the recently discovered R. badensis species (and any other species of the genus Rouxiella) as a biocontrol agent against postharvest fungal pathogens.

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The fungal species Trichoderma is frequently found in soil antagonizing plant-pathogenic fungi as well as parasitizing plant-pathogenic nematodes. Metarhizium species are insect-pathogenic fungi that are used throughout the world to control agricultural insect pests. Here, we determine whether the antagonism (A) of Trichoderma atroviride to Metarhizium robertsii during growth and spore formation can impact the stress biology of M. robertsii conidia. Cultures of M. robertsii were either produced without exposure to T. atroviride (control) or in the presence of T. atroviride. M. robertsii was grown in dual culture with T. atroviride on potato dextrose agar (PDA) using the following treatments: 1) Trichoderma inoculated at the same time with Metarhizium (A0); 2) Trichoderma inoculated two days after the inoculation of Metarhizium (A2); 3) Trichoderma inoculated four days after Metarhizium (A4); 4) Trichoderma inoculated 6 d after Metarhizium (A6); 5) M. robertsii grown alone on PDA medium (control); and 6) M. robertsii grown alone on minimal medium (Czapek-Dox medium without sucrose) (MM). Germination of M. robertsii conidia from all six treatments was then assessed under osmotic, oxidative, UV-B, and thermal stress. M. robertsii conidia produced on MM were the most tolerant to all stress conditions. For all stress conditions, conidia from treatments A0 and A2 were not viable. For osmotic stress, conidia produced in treatment A4 were the most tolerant, followed by conidia from treatment A6, which were both more tolerant than the control. For oxidative stress, conidia produced in both A4 and A6 treatments were similarly tolerant and more tolerant than conidia produced in the control. For thermal stress, conidia produced in treatments A4, A6, and control (PDA) were similarly heat-tolerant. For UV-B stress, conidia produced in treatments A4 and A6 were equally tolerant and more tolerant than conidia produced in the control. The germination speed of conidia produced in all treatments, A0, A2, A4, and A6 was also tested. Conidia produced on MM germinated faster than the other treatments. Conidia produced in the A4 treatment were the second fastest, followed by conidia produced in treatment A6. Both A4 and A6 conidia germinated faster than conidia produced in the control treatment. Conidia produced in the treatments A0 and A2 did not germinate in 24 h. In summary, moderate levels of biotic stress from a fungal competitor or low-nutrient conditions can enhance the stress tolerance of M. robertsii conidia.

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Grapevine Trunk Diseases (GTDs) are a major challenge to the grape industry worldwide. GTDs are responsible for considerable loss of quality, production, and vineyard longevity. Seventy-five percent of Chilean vineyards are estimated to be affected by GTDs. GTDs are complex diseases caused by several fungi species, including members of the Botryosphaeriaceae family and Phaeomoniella chlamydospora, considered some of the most important causal agents for these diseases in Chile. In this study, we isolated 169 endophytic and 209 rhizospheric fungi from grapevines grown under organic and conventional farming in Chile. Multiple isolates of Chaetomium sp., Cladosporium sp., Clonostachys rosea, Epicoccum nigrum, Purpureocillium lilacinum, and Trichoderma sp. were evaluated for their potential of biocontrol activity against Diplodia seriata, Neofusicoccum parvum, and Pa. chlamydospora. Tests of antagonism were carried out using two dual-culture-plate methods with multiple media types, including agar containing grapevine wood extract to simulate in planta nutrient conditions. Significant pathogen growth inhibition was observed by all isolates tested. Clonostachys rosea showed 98.2% inhibition of all pathogens in the presence of grapevine wood extract. We observed 100% pathogen growth inhibition when autoclaved lignified grapevine shoots were pre-inoculated with either C. rosea strains or Trichoderma sp. Overall, these results show that C. rosea strains isolated from grapevines are promising biocontrol agents against GTDs.

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Legumes establish symbiotic relationships with different microorganisms, which could function as plant growth promotion microorganisms (PGPM). The finding of new PGPM strains is important to increase plant production avoiding or diminishing the use of industrial fertilizers. Thus, in this work we evaluated the plant growth promotion traits of ten strains isolated from Mimosa pudica root nodules. According to the 16S rDNA sequence, the microorganisms were identified as Enterobacter sp. and Serratia sp. To the best of our knowledge this is the first report describing and endophytic interaction between Mimosa pudica and Enterobacter sp. These strains have some plant growth promoting traits such as phosphate solubilization, auxin production and cellulase and chitinase activity. Strains identified as Serratia sp. inhibited the growth of the phytopathogenic fungi Fusarium sp., and Alternaria solani and the oomycete Phytophthora capsici. According to their biochemical characteristics, three strains were selected to test their plant growth promoting activity in a medium with an insoluble phosphate source. These bacteria show low specificity for their hosts as endophytes, since they were able to colonize two very different legumes: Phaseolus vulgaris and M. pudica. Seedlings of P. vulgaris were inoculated and grown for fifteen days. Enterobacter sp. NOD1 and NOD10, promoted growth as reflected by an increase in shoot height as well as an increase in the size and emergence of the first two trifolia. We could localize NOD5 as an endophyte in roots in P. vulgaris by transforming the strain with a Green Fluorescent Protein carrying plasmid. Experiments of co-inoculation with different Rhizobium etli strains allowed us to discard that NOD5 can fix nitrogen in the nodules formed by a R. etli Fix- strain. The isolates described in this work show biotechnological potential for plant growth promoting activity and production of indoleacetic acid and siderophores.

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Interactions among microorganisms may be the cause of morphological modifications, particularly in fungal cells. The aim of this work was to examine the changes that occur in cells of the fungus Fonsecaea pedrosoi after in vitro co-culturing with Bacillus subtilis and to explore the results of this interaction in vivo in an experimental murine infection. B. subtilis strain was inoculated into a 15-day pure culture of F. pedrosoi. In vitro, after 48 hours of co-culturing, the fungal cells were roundish. The secretion of fungal dark pigments and production of terminal chlamydoconidia were observed in hyphae after one week. In the in vivo study, two animal groups of 30 BALB/c mice each were employed. One group was inoculated intraperitoneally with hyphal fragments from the co-culture of bacteria and fungi; the other group was infected only with F. pedrosoi hyphae. After seven days of infection, both animal groups developed neutrophilic abscesses. Phagocytosis of bacilli by macrophages occurred at three days. At later periods, generally after 25 days, only roundish cells similar to sclerotic bodies remained in the tissues while hyphae were eliminated by 15 to 20 days. These fungal forms originated mainly from terminal chlamydoconidia. The co-culturing between bacteria and fungi may constitute a mechanism to rapidly obtain resistant fungal forms for host defenses, especially for chromoblastomycosis (CBM) experimental infections.(AU)

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RESUMO

Interactions among microorganisms may be the cause of morphological modifications, particularly in fungal cells. The aim of this work was to examine the changes that occur in cells of the fungus Fonsecaea pedrosoi after in vitro co-culturing with Bacillus subtilis and to explore the results of this interaction in vivo in an experimental murine infection. B. subtilis strain was inoculated into a 15-day pure culture of F. pedrosoi. In vitro, after 48 hours of co-culturing, the fungal cells were roundish. The secretion of fungal dark pigments and production of terminal chlamydoconidia were observed in hyphae after one week. In the in vivo study, two animal groups of 30 BALB/c mice each were employed. One group was inoculated intraperitoneally with hyphal fragments from the co-culture of bacteria and fungi; the other group was infected only with F. pedrosoi hyphae. After seven days of infection, both animal groups developed neutrophilic abscesses. Phagocytosis of bacilli by macrophages occurred at three days. At later periods, generally after 25 days, only roundish cells similar to sclerotic bodies remained in the tissues while hyphae were eliminated by 15 to 20 days. These fungal forms originated mainly from terminal chlamydoconidia. The co-culturing between bacteria and fungi may constitute a mechanism to rapidly obtain resistant fungal forms for host defenses, especially for chromoblastomycosis (CBM) experimental infections.

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