Rapid Detection of Wheat Blast Pathogen Magnaporthe oryzae Triticum Pathotype Using Genome-Specific Primers and Cas12a-mediated Technology

Wheat blast,caused by the fungus Magnaporthe oryzae Triticum(MoT)pathotype,is a devastating disease persistent in South America and Bangladesh.Since MoT generally fails to cause visual symptoms in wheat until the heading stage when the infection would have advanced,disease control by fungicide application solely based on the detection of visual symptoms is ineffective.To develop an accurate and sensitive method to detect MoT at the seedling and vegetative stages for disease control,we sequenced the genomes of two MoT isolates from Brazil and identified two DNA fragments,MoT-6098 and MoT-6099,that are present in the MoT genome but not in the genome of the rice-infecting Magnaporthe oryzae Oryzae(MoO)pathotype.Using polymerase chain reaction(PCR),we confirmed the specificity of the two markers in 53 MoT and MoO isolates from South America and Bangladesh.To test the efficiency of the two markers,we first established a loop-mediated isothermal amplification(LAMP)method to detect MoT at isothermal conditions,without the use of a PCR machine.Following this,we used the Cas12a protein and guide RNAs(gRNAs)to target the MoT-6098 and MoT-6099 sequences.The activated Cas12a showed indiscriminate single-stranded deoxyribonuclease(ssDNase)activity.We then combined targetdependent Cas12a ssDNase activation with recombinase polymerase amplification(RPA)and nucleic acid lateral flow immunoassay(NALFIA)to develop a method that accurately,sensitively,and cost-effectively detects MoT-specific DNA sequences in infected wheat plants.This novel technique can be easily adapted for the rapid detection of wheat blast and other important plant diseases in the field.

Enhancement of Growth and Grain Yield of Rice in Nutrient Deficient Soils by Rice Probiotic Bacteria

Plant associated bacteria are promising alternatives to chemical fertilizers for plant growth and yield improvement in an eco-friendly manner.In this study,rice associated bacteria were isolated and assessed for mineral phosphate solubilization and indole-3-acetic acid(IAA) production activity in vitro.Six promising strains,which were tentatively identified as phylotaxon Pseudochrobactrum sp.(BRRh-1),Burkholderia sp.(BRRh-2),Burkholderia sp.(BRRh-3),Burkholderia sp.(BRRh-4),Pseudomonas aeruginosa(BRRh-5 and BRRh-6) based on their 16 S rRNA gene phylogeny,exhibited significant phosphate solubilizing activity in National Botanical Research Institute phosphate growth medium,and BRRh-4 displayed the highest phosphate solubilizing activity,followed by BRRh-5.The p H of the culture broth declined,resulting in increase of growth rate of bacteria at p H 7,which might be due to organic acid secretion by the strains.In presence of L-tryptophan,five isolates synthesized IAA and the maximum IAA was produced by BRRh-2,followed by BRRh-1.Application of two most efficient phosphate solubilizing isolates BRRh-4 and BRRh-5 by root dipping(colonization) of seedling and spraying at the flowering stage significantly enhanced the growth and grain yield of rice variety BRRI dhan-29.Interestingly,application of both strains with 50% of recommended nitrogen,phosphorus and potassium fertilizers produced equivalent or higher grain yield of rice compared to the control grown with full recommended fertilizer doses,which suggests that these strains may have the potential to be used as bioinoculants for sustainable rice production.

Variability in total antioxidant capacity, antioxidant leaf pigments and foliage yield of vegetable amaranth

Twenty vegetable amaranth genotypes were evaluated for total antioxidant capacity, antioxidant leaf pigments, vitamins, and selection of suitable genotypes for extraction of juice in a randomized complete block design（RCBD） with three replications. Vegetable amaranth was rich in chlorophyll, β-cyanins, β-xanthins, betalains, carotene, ascorbic acid and total antioxidant. The genotypes VA14, VA16, VA18, VA15, and VA20 could be selected as amaranth vegtable varieties with high yields and abundance antioxidant leaf pigments and vitamins to produce juice. The genotypes VA13 and VA19 had above-average foliage yield and high antioxidant profiles while the genotypes VA2, VA3, VA9, VA11, VA12, and VA17 had a high antioxidant profiles and below-average foliage yield. These genotypes could be used as a donor parent for integration of potential high antioxidant profiles genes into other genotypes. The correlation study revealed a strong positive association among all the antioxidant leaf pigments, total antioxidant capacity and foliage yield. Selection based on total antioxidant capacity, antioxidant leaf pigments could economically viable to improve the yield potential of vegetable amaranth genotypes. Total carotene and ascorbic acid exhibited insignificant genotypic correlation with all the traits except total antioxidant capacity. This indicates that selection for antioxidant vitamins might be possible without compromising yield loss.