In Planta Detection of Beauveria bassiana (Ascomycota: Hypocreales) Strains as Endophytes in Bean (Phaseolus vulgaris L.).

Beauveria bassiana (B. bassiana) is a significant entomopathogenic fungus (EPF) in agriculture as a sprayable biocontrol agent. It has the potential to be established as an endophyte (ENP) in various crops, resulting in beneficial effects for the host plants, including resistance to pest insects and increased growth and yield. However, it is not known whether a B. bassiana strain has such a favorable impact on the plant, since it is a common soil microorganism. Therefore, techniques that allow strain monitoring will be advantageous. To date, methods for detecting or monitoring a specific EPF strain after external application are scarce. In the present study, an in planta nested PCR technique was standardized to differentiate between three B. bassiana strains (GHA, PTG4, and BB37) established as endophytes in bean plants under laboratory conditions by detecting the insertion profile of four group I introns located in the 28S gene of B. bassiana ribosomal DNA. This technique recognized a distinct pattern of bands of different sizes for each strain, with a sensitivity of 1 pg per 10 ng of plant DNA. This molecular approach may be more effective monitoring B. bassiana strains after application to evaluate their significance on crops.

Entomopathogenic fungi tested in planta on pepper and in field on sorghum, to control commercially important species of aphids.

Given the aphids high reproductive capacity, assessing their biocontrol by using entomopathogenic fungi is crucial; to determine their potential, fungi were tested in planta and in field conditions. Significant decrease of Myzus persicae (Sulzer) population was observed in planta after applying Beauveria bassiana (strain 7R), Trichoderma gamsii (strain Z) or Metarhizium brunneum (strain Meta Br1) at 1 × 107 or 1 × 108 conidia/mL on pepper plants. Significant differences of aphids' populations were detected between fungus concentration and control (F = 68.743, df = 6.980, P < 0.001), where M. brunneum at 1 × 108 conidia/mL reduced aphids population close to zero. At 20 °C, dead aphids' mycosis by B. bassiana and T. gamsii was 78% and 84%; at 25 °C was 83% and 88%; and at 30 °C was 75% and 79%, respectively. In field conditions, Mexican PTG4 and commercial GHA B. bassiana strains were tested [(1 × 106 conidia/mL + corn starch) seed treatments] against the Melanaphis sacchari (Zehntner) aphid populations, on naturally infested sorghum plants. Results showed that plant germination and emergence were not affected, whereas yield (grams of sugar/plant) was significantly higher among treated compared with untreated plants. The aphid population decreased in plants from PTG4 treated seeds; indeed, this treatment had a significant positive effect on the flowering index, whereas the stem fresh weight and juice volume was significantly increased among plants from GHA treated seeds. Taken together, tested strains can be used as a tool to control aphids' population on several crops such as pepper and even increase the yield in sorghum.

Isolation and enzyme bioprospection of bacteria associated to Bruguiera cylindrica, a mangrove plant of North Sumatra, Indonesia.

Mangrove-associated bacteria are of industrial interest due to their diverse and versatile enzyme properties. This study investigates the culturable bacteria from a wide range of habitat in a Bruguiera cylindrica mangrove ecosystem in North Sumatra. Screening of extracellular hydrolytic enzymes showed multiple potential traits in amylase, cellulase, chitinase, phosphatase, protease, and urease production by bacterial isolates. Molecular identification based on 16S rDNA region of a potential strain, Vibrio alginolyticus Jme3-20 is then reported as a newly proteolytic agent. The strain also showed a stable growth under salinity (NaCl) stress with considerable phosphate solubilization activities. Protease activity was enhanced by optimizing the 0.5 % (w/v) sucrose and soy peptone in the fermentation medium. SDS-PAGE and zymogram analysis showed the presence of a 35-kDa MW protease. Hence, our study revealed important insights into the bacterial diversity and activity in mangrove ecosystems, evidencing the importance of microbial exploration in this ecosystem.

Bacterial and Fungal Microbiome Profiling in Chilhuacle Negro Chili (Capsicum annuum L.) Associated With Fruit Rot Disease.

Chilhuacle negro chili (Capsicum annuum L.) is an ancient Mexican landrace that is deeply linked to the culinary heritage of the country. Because of the high profitability and uniqueness of this crop, the Universidad Autónoma del Estado de Morelos is exploring its production in controlled environments. In the crop cycles of 2018 to 2019, the production of chilhuacle negro plants was seriously affected by an unidentified pathogen causing fruit rot, which reduced its quality, yield, and market value. Therefore, the main objective of this work was to study and characterize the fruit microbiota, which could help reveal the causal agent of this disease. Using DNA metabarcoding coupled with Illumina and nanopore sequencing technologies, we collected and analyzed both healthy and infected chili fruit, along with greenhouse bioaerosols. We also explored the bacterial and fungal microbiota by using microbiological techniques to isolate some of the culturable bacterial and fungal species. Our results suggest that the seedborne fungus Alternaria alternata is activated during the maturation stage of chilhuacle negro fruit, triggering a microbiome imbalance, which may in turn enable the establishment of other opportunistic pathogenic fungi during fruit decay, such as Mucor sp. To our knowledge, this is the first study of the chilhuacle negro chili microbiome, which can shed some light on our understanding of one of the main diseases that affect this valuable crop.

Endophytic Beauveria bassiana promotes drought tolerance and early flowering in corn.

Beauveria bassiana (Bals.) Vuillemin (B. bassiana) is an entomopathogenic fungus that establishes endophytic symbiosis with plants. In the present study, the effects of B. bassiana strains colonization in growing Zea mays L. (Z. mays), crop production, and drought tolerance were evaluated. Z. mays seeds were inoculated with B. bassiana strains (GHA, PTG4, and PTG6), using 1 × 106 blastospores/mL and methyl cellulose (MC) or cornstarch (CS) as adherents. Colonization was determined by B. bassiana recovery from plant tissues plated on PDA medium. Plant height, fresh and dry weight, and flowering time were analyzed to assess plant performance. Drought tolerance was evaluated by stopping watering for 10 days, watering again, and determining vigor recovery after 24 h. Results showed 100% endophytic roots colonization, regardless of adherent type or strain tested. Colonization was variable in shoots and leaves, but GHA strain achieved the highest inoculation rates, including 88% in stems and 50% in leaves, which did not depend on adherent type used; for PTG4 strain, adherent type had an important effect (MC = 100% stems and leaves; CS = 63% stems and 25% leaves). For PTG6 strain, the best adherent type was CS (71% stems and 75% leaves), whereas MC showed variable inoculation percentage (25% stems and 75% leaves). Interestingly, only MCPTG4 treatment showed consistent positive effects on germination percentage (day 5 = 46 ± 2%; day 14 = 87 ± 7%) compared with controls (CC = 63 ± 4%, CMC = 50 ± 3%, CCS = 47 ± 0%). In addition, the other treatments showed low germination percentages at day 5 (7 ± 7% to 46 ± 2%), which recovered at day 14 (53 ± 0% to 73 ± 8%), except for MCPTG6 treatment with 23 ± 10% germination. About plant performance, not significant effects on plant height and fresh/dry weight in all the treatments were observed. However, B. bassiana-treated plants, using either GHA, PTG4 or PTG6 strains, and MC as adherent, showed tolerance to drought and flowered one to two weeks earlier, providing evidence supporting further applications of these seed treatments in agricultural systems, for abiotic stress sustainable management practices.

Endophytic fungi alter sucking bug responses to cotton reproductive structures.

All plants including cotton host a wide range of microorganisms as endophytes. There is a growing appreciation of the prevalence, ecological significance and management potential of facultative fungal endophytes in protecting plants from pests, pathogens and environmental stressors. Hemipteran sucking bugs have emerged as major pests across the U.S. cotton belt, reducing yields directly by feeding on developing reproductive structures and indirectly by vectoring plant pathogens. We used no-choice and simultaneous choice assays to examine the host selection behavior of western tarnished plant bugs (Lygus hesperus) and southern green stink bugs (Nezara viridula) in response to developing flower buds and fruits from cotton plants colonized by 1 of 2 candidate beneficial fungal endophytes, Phialemonium inflatum or Beauveria bassiana. Both insect species exhibited strong negative responses to flower buds (L. hesperus) and fruits (N. viridula) from plants that had been colonized by candidate endophytic fungi relative to control plants under both no-choice and choice conditions. Behavioral responses of both species indicated that the insects were deterred prior to contact with plant tissues from endophyte-colonized plants, suggesting a putative role for volatile compounds in mediating the negative response. Our results highlight the role of fungal endophytes as plant mutualists that can have positive effects on plant resistance to pests.

The entomopathogenic fungal endophytes Purpureocillium lilacinum (formerly Paecilomyces lilacinus) and Beauveria bassiana negatively affect cotton aphid reproduction under both greenhouse and field conditions.

The effects of two entomopathogenic fungal endophytes, Beauveria bassiana and Purpureocillium lilacinum (formerly Paecilomyces lilacinus), were assessed on the reproduction of cotton aphid, Aphis gossypii Glover (Homoptera:Aphididae), through in planta feeding trials. In replicate greenhouse and field trials, cotton plants (Gossypium hirsutum) were inoculated as seed treatments with two concentrations of B. bassiana or P. lilacinum conidia. Positive colonization of cotton by the endophytes was confirmed through potato dextrose agar (PDA) media plating and PCR analysis. Inoculation and colonization of cotton by either B. bassiana or P. lilacinum negatively affected aphid reproduction over periods of seven and 14 days in a series of greenhouse trials. Field trials were conducted in the summers of 2012 and 2013 in which cotton plants inoculated as seed treatments with B. bassiana and P. lilacinum were exposed to cotton aphids for 14 days. There was a significant overall effect of endophyte treatment on the number of cotton aphids per plant. Plants inoculated with B. bassiana had significantly lower numbers of aphids across both years. The number of aphids on plants inoculated with P. lilacinum exhibited a similar, but non-significant, reduction in numbers relative to control plants. We also tested the pathogenicity of both P. lilacinum and B. bassiana strains used in the experiments against cotton aphids in a survival experiment where 60% and 57% of treated aphids, respectively, died from infection over seven days versus 10% mortality among control insects. Our results demonstrate (i) the successful establishment of P. lilacinum and B. bassiana as endophytes in cotton via seed inoculation, (ii) subsequent negative effects of the presence of both target endophytes on cotton aphid reproduction using whole plant assays, and (iii) that the P. lilacinum strain used is both endophytic and pathogenic to cotton aphids. Our results illustrate the potential of using these endophytes for the biological control of aphids and other herbivores under greenhouse and field conditions.

Potential role of Arabidopsis PHP as an accessory subunit of the PAF1 transcriptional cofactor.

Paf1C is a transcriptional cofactor that has been implicated in various transcription-associated mechanisms spanning initiation, elongation and RNA processing, and is important for multiple aspects of development in Arabidopsis. Our recent studies suggest Arabidopsis Paf1C is crucial for proper regulation of genes within H3K27me3-enriched chromatin, and that a protein named PHP may act as an accessory subunit of Paf1C that promotes this function.