Transgenic Control of Aflatoxin Contamination in Maize Through Host-Induced Gene Silencing Targeting Aspergillus flavus Genes Encoding Polygalacturonase (p2c) and Versicolorin Dehydrogenase (aflM)

Maize (Zea mays L.) is susceptible to Aspergillus flavus infection and subsequent contamination with aflatoxins, the most potent naturally produced carcinogenic secondary metabolites. Here, the A. flavus gene aflM encoding the versicolorin dehydrogenase in aflatoxin biosynthesis and the p2c gene encoding the polygalacturonase that is involved in infection were selected as targets for suppression through host induced gene silencing (HIGS). A HIGS vector targeting these genes was constructed and introduced into immature B104 maize embryos. Thirteen out of fifteen p2c transformation events and six out of seven aflM events were confirmed positive by PCR. Kernels containing the p2c gene from four out of seven events examined from T1 generation and six events out of eleven events from T2 generation examined had less aflatoxin than those without the transgene. Field-inoculated homozygous T3 and T4 generation kernels from four events also revealed significantly lower aflatoxins (p\u3e0.02) than the kernels from the null or B104 controls. Transferring the p2c gene to elite background also resulted in crosses with significantly less aflatoxin production (p\u3e0.02) compared to the controls. For aflM transgenic lines, kernels containing the aflM gene from one (aflM14) out four events examined in T1 generation and from two events (aflM14 and aflM16) out of four examined had less aflatoxin (p≤0.01 and p≤0.08) than those without the transgene than those without the transgene. Homozygous T3 and T4 transgenic kernels of aflM14 and aflM16 showed significantly less aflatoxin production than kernels from the null or B104 controls under both field inoculation and laboratory kernel screening assay conditions. Transferring aflM from these events into four elite inbred lines resulted in F1 crosses with significantly less aflatoxins (p≤0.02) than the controls. P2c13, P2c17, aflM14 and aflM16 were confirmed to contain a single copy insertion of the transgene according to droplet digital PCR analysis. The enhanced aflatoxin resistance in homozygous transgenic lines is associated with high levels of gene specific small RNAs detected in the transgenic leaf and kernel tissues, indicating that it is possible to manage aflatoxin contamination in maize through HIGS targeting p2c or aflM

Keys to success for the advertising media management business on the sky train routes in Thailand

Enhancing the potential of the advertising media management business entrepreneurs enables them to be successful in their businesses on the sky train routes. At present, there are a total of 5 sky train lines and 113 stations in service, covering a distance of more than 157 kilometers. With this reason, advertising media used for the promotion of products and services are displayed both inside the train stations and in the trains in order to create awareness among consumers to make a purchasing decision. At present, the competition of this type of business is very high. Advertising media management business has not been able to adapt and differentiate itself from its competitors nor create its competitive advantage, resulting in its inability to respond to the needs of the customers who travel on the sky trains in Thailand. The objectives of this research were to: 1) study levels of personal norms, purchasing decision, co-branding with customers through social commerce tools, advertising strategies, social support, innovations for trust building, and the success of the advertising media management business on the sky train routes in Thailan

Effect of the inno-life management, participation, and leadership on the organizational climate of community-based tourism in Phatthalung, Thailand

The objective of this study is to check the Effect of the Inno-life management, participation, and leadership on the organizational climate of community-based tourism in Phatthalung, Thailand. The study aims to discover the ways that are affecting community-based tourism to promote the tourism sector in Thailand. It was identified that the tourism sector in Phatthalung, Thailand is facing a crisis from the perceptive of community-based tourism. This study is based on the stratified sampling method in which the data was collected from the diverse respondents on the Likert scale five-point questionnaire to get the results. The study concludes that there is an important role of innovation life management, participation of the employees in the organization, and the leadership qualities of the managerial staff to improve the community-based tourism in Phatthalung, Thailand. This study is significant because it carries critical theoretical as well as practical implications for the community-based tourism business sector to improve the business performance and lead the organization in a prosperous way

Use of Promising Bacterial Strains for Controlling Anthracnose on Leaf and Fruit of Mango Caused by Colletotrichum gloeosporioides

A total 146 isolates of bacteria were taken from leaf surface, fruit skin, and blossom of mango (var. Nam Dorkmai). They were tested for the inhibition of mycelial growth of Colletotrichum gloeosporioides, a causal agent of anthracnose, on potato dextrose agar (PDA). Seventy-four bacterial isolates inhibited the growth of fungal mycelia by 24.51-49.10%. The 40 highly effective isolates out of 74 isolates were further tested for the potential to reduce the development of anthracnose lesion on detached leaves of mango marcotages at 24 h after inoculation of pathogen. Results indicated that 12 isolates provided high efficacy for inhibiting disease by 51.39-86.11%. Application of these bacteria on mango fruits at 24 h prior to the inoculation of the pathogen revealed that isolates B46 and B12 suppressed disease by 50.36 and 44.13% respectively while Trichoderma harzianum CB-Pin-01 provided 37.30% of the inhibition. For controlling post-harvest disease, an isolate B12 or B12 integrated with hot water treatment (55 oC) provided 91.33 and 88.00% of disease severity reduction respectively when applied at 24 h before inoculation of pathogen. Isolates B12 and B44 were identified as Bacillus subtilis while B46 and K112 were B. licheniformis and B. cereus respectively. The mechanism of these isolates for controlling C. gloeosporioides was the reduction of spore germination and the inhibition of germ-tube elongation

Targeting the Aspergillus flavus p2c gene through host-induced gene silencing reduces A. flavus infection and aflatoxin contamination in transgenic maize

Aspergillus flavus is an opportunistic fungal pathogen that infects maize and produces aflatoxins. Using biocontrol or developing resistant cultivars to reduce aflatoxin contamination has only achieved limited success. Here, the A. flavus polygalacturonase gene (p2c) was targeted for suppression through host-induced gene silencing (HIGS) to reduce aflatoxin contamination in maize. An RNAi vector carrying a portion of the p2c gene was constructed and transformed into maize B104. Thirteen out of fifteen independent transformation events were confirmed to contain p2c. The T2 generation kernels containing the p2c transgene had less aflatoxin than those without the transgene in six out of eleven events we examined. Homozygous T3 transgenic kernels from four events produced significantly less aflatoxins (P ≤ 0.02) than the kernels from the null or B104 controls under field inoculation conditions. The F1 kernels from the crosses between six elite inbred lines with P2c5 and P2c13 also supported significantly less aflatoxins (P ≤ 0.02) than those from the crosses with null plants. The reduction in aflatoxin ranged from 93.7% to 30.3%. Transgenic leaf (T0 and T3) and kernel tissues (T4) were also found to have significantly higher levels of p2c gene-specific small RNAs. Further, homozygous transgenic maize kernels had significantly less fungal growth (27~40 fold) than the null control kernels 10 days after fungal inoculation in the field. The calculated suppression of p2c gene expression based on RNAseq data was 57.6% and 83.0% in P2c5 and P2c13 events, respectively. These results indicate clearly that the reduced aflatoxin production in the transgenic kernels is due to RNAi-based suppression of p2c expression, which results in reduced fungal growth and toxin production