Phosphate is one of the most important nutrients for plant growth and development, and only 0.1% of the phosphate in soils is available to plants. Currently, the use of excess phosphate fertilizer has caused surface and ground water pollution and water eutrophication, resulting in algal blooms in lakes and oceans. Therefore, it is imperative to explore alternative ways to solve these problems for sustainable agricultural production and improvement of soil fertility, while protecting the environment. Microorganisms from the rhizosphere and within plants are able to solubilize insoluble soil phosphate, making it available to plants. Five high phosphate solubilizing bacteria from our bacterial endophyte library were chosen for this study and identified as Pantoea vagans IALR611, Pseudomonas psychrotolerans IALR632, Bacillus subtilis IALR1033, Bacillus safensis IALR1035 and Pantoea agglomerans IALR1325. All five bacteria significantly promoted tall fescue growth in vitro. Greenhouse experiments showed that IALR1325 significantly promoted pepper and tomato growth, and IALR632 was the best in promoting tomato growth. In addition, all these bacteria had extracellular acid phosphatase and phytase activities. One of the mechanisms for phosphate solubilization by bacteria is pH reduction caused by gluconic acid production. Our results indicate that P. agglomerans IALR1325 is a promising bacterium for future applications.

Endophytic bacteria could be used as a tool to promote plant growth, enhance abiotic stress tolerance, and inhibit plant pathogen growth. More than 100 bacteria, previously tested to have biocontrol activity against fungal pathogens from our lab, were chosen for pathogen inhibition in vitro with antibiosis assays against strawberry specific pathogens. Three potential endophytic bacteria were tested in the greenhouse for disease reduction and growth performance. Finally, field trials were conducted for fruit yield production. Our results showed that three endophytic bacteria significantly inhibited the growth of the strawberry pathogen Colletotrichum gloeosporioides from antibiosis assays and were identified as Bacillus velezensis strains IALR308, IALR585, and IALR619. Inoculation of these bacteria significantly reduced strawberry disease in greenhouse conditions with pathogen infection. Field trials showed that IALR619 has potential to influence marketable fruit yield when strawberry plants were inoculated twice over the growing season. All three bacteria had the ability to produce auxin and to solubilize phosphate. The antibiotics surfactin and iturin were also detected in IALR585 and IALR619. In conclusion, Bacillus velezensis IALR619 has potential inhibition of strawberry pathogen growth in the greenhouse and possible ability to increase marketable fruit yield in the field.

An important component of conventional sense, antisense, and double-strand RNA-based gene silencing constructs is the transcriptional terminator. Here, we show that this regulatory element becomes obsolete when gene fragments are positioned between two oppositely oriented and functionally active promoters. The resulting convergent transcription triggers gene silencing that is at least as effective as unidirectional promoter-to-terminator transcription. In addition to short, variably sized, and nonpolyadenylated RNAs, terminator-free cassette produced rare, longer transcripts that reach into the flanking promoter. These read-through products did not influence the efficacy and expression levels of the neighboring hygromycin phosphotransferase gene. Replacement of gene fragments by promoter-derived sequences further increased the extent of gene silencing. This finding indicates that genomic DNA may be a more efficient target for gene silencing than gene transcripts.

Controlled environment agriculture hydroponic systems grow plants year-round without restriction from outside environmental conditions. In order to further improve crop yield, plant growth-promoting bacteria were tested on hydroponically grown lettuce (Lactuca sativa) plants. From our bacterial endophyte library, we found one bacterium, Pseudomonas psychrotolerans IALR632, that is promising in promoting lettuce growth in multiple hydroponic systems. When Green Oakleaf lettuce seeds were inoculated with IALR632 during germination, IALR632 significantly increased lateral root development by 164%. When germinated seedlings were inoculated with IALR632 and then transplanted to different hydroponic systems, shoot and root fresh weights of Green Oakleaf increased by 55.3% and 17.2% in a nutrient film technique (NFT) system in the greenhouse, 13.5% and 13.8% in an indoor vertical NFT system, and 15.3% and 13.6% in a deep water cultivation (DWC) system, respectively. IALR632 also significantly increased shoot fresh weights of Rex by 33.9%, Red Oakleaf by 21.0%, Red Sweet Crisp by 15.2%, and Nancy by 29.9%, as well as Red Rosie by 8.6% (no significant difference). Inoculation of IALR632-GFP and subsequent analysis by confocal microscopy demonstrated the endophytic nature and translocation from roots to shoots. The results indicate that P. psychrotolerans IALR632 has a potential application in hydroponically grown lettuce plants.

The use of fungicides to manage disease has led to multiple environmental externalities, including resistance development, pollution, and non-target mortality. Growers have limited options as legacy chemistry is withdrawn from the market. Moreover, fungicides are generally labeled for traditional soil-based production, and not for liquid culture systems. Biocontrol agents for disease management are a more sustainable and environmentally friendly alternative to conventional agroprotectants. Pythium ultimum is a soil borne oomycete plant pathogen with a broad taxonomic host range exceeding 300 plants. Cucumber seedlings exposed to P. ultimum 1 day after a protective inoculation with bacterial endophyte accession IALR1619 (Pseudomonas sp.) recorded 59% survival; with the control assessed at 18%. When the pathogen was added 5 days post endophyte inoculation, 74% of the seedlings treated survived, compared to 36% of the control, indicating a longer-term effect of IALR1619. Under hydroponic conditions, IALR1619 treated leaf type lettuce cv. ‘Cristabel’ and Romaine cv. ‘Red Rosie’ showed 29% and 42% higher shoot fresh weight compared to their controls, respectively. Similar results with less growth decline were observed for a repeat experiment with IALR1619. Additionally, an experiment on hydroponic lettuce in pots with perlite was carried out with a mixture of P. ultimum and P. dissotocum after IALR1619 inoculation. The endophyte treated ‘Cristabel’ showed fresh weight gain, but the second cultivar ‘Pensacola’ yielded no increase. In summary, the endophyte IALR1619 provided short term as well as medium-term protection against Pythium blight in cucumber seedlings and may be used as an alternative to conventional fungicides in a greenhouse setting. This study also demonstrated the potential of ALR1619 as a biocontrol agent against Pythium blight in hydroponic lettuce.