PHYLOGENETIC ANALYSIS OF PROTOZOA AND SISTAN COW'S RUMEN BACTERIA FED WITH FORAGE RATIONS BY MOLECULAR AND LABORATORY METHODS

The present study was conducted to investigate the properties of rumen Microbiome in Sistani cattle by performing molecular experiments to clone the 18S rRNA genes of protozoans, culture rumen bacteria and then sequencing their 16S rRNA gene. Rumen liquid samples were collected from 10 Sistani cattle from the Sistan region, which were kept in a similar feeding group and fed on forage rations, through stomach tubes before feeding in the morning. The protozoa genome was extracted by the ASL buffer of the QiaAmp DNA stool kit (Qiagen), and their 18S rRNA gene was amplified and isolated with specific primers by the PCR method. The quantity and quality of DNA extracted with nanodrop 1000 and electrophoresis on 1% agarose gel, respectively, have been in The 18S rRNA protozoan gene was cloned using the T/A cloning technique in the PTZ plasmid and then the recombinant plasmid was transferred to E. coli.vestigated. The bacteriawere cultured with the dependent method and then their 16S rRNA gene was sequenced. Rumen bacteria culture was performed in an anaerobic culture medium using an anaerobic chamber with flexible plastic gloves. After purification, the bacteria were partially studied morphologically. Then the bacterial genome was extracted using a kit, and the 16S rRNA gene was propagated by PCR methods. The sequences have been sent to determine the final structure of the gene for sequencing. In the end, the phylogenetic tree is drawn using the MEGAX software.Investigating of the microbial sequences obtained from the rumen of the cattle showed that the results are somewhat consistent with the results of other researchers in other countries and in other animals, but in some cases, there are significant differences. In this study, the Entodinium genus was the dominantprotozoan group in the Sistani cattle's rumen. In the library, OTUs had a similarity of over 98.5% with the protozoan sequences identified in the database. The results of the culture of bacteria showed Ruminococcus albus, Ruminococcus flavefaciense, Clostridium colinum, Streptococcus equinus and butyrivibrio genus, which had a high similarity of 96.54% with the bacterial sequences identified in the database. Ruminal microbial ecology is very complex. The real scope of this diversity is determined by the use of molecular identification methods for species. The complexity of this diversity is determined by the use of 16S rRNA genes and 18S rRNA genes. Since each microbial species occupies a small area and is found only in some animals

Salinity stress endurance of the plants with the aid of bacterial genes

The application of plant growth-promoting bacteria (PGPB) is vital for sustainable agriculture with continuous world population growth and an increase in soil salinity. Salinity is one of the severe abiotic stresses which lessens the productivity of agricultural lands. Plant growth-promoting bacteria are key players in solving this problem and can mitigate salinity stress. The highest of reported halotolerant Plant growth-promoting bacteria belonged to Firmicutes (approximately 50%), Proteobacteria (40%), and Actinobacteria (10%), respectively. The most dominant genera of halotolerant plant growth-promoting bacteria are Bacillus and Pseudomonas. Currently, the identification of new plant growth-promoting bacteria with special beneficial properties is increasingly needed. Moreover, for the effective use of plant growth-promoting bacteria in agriculture, the unknown molecular aspects of their function and interaction with plants must be defined. Omics and meta-omics studies can unreveal these unknown genes and pathways. However, more accurate omics studies need a detailed understanding of so far known molecular mechanisms of plant stress protection by plant growth-promoting bacteria. In this review, the molecular basis of salinity stress mitigation by plant growth-promoting bacteria is presented, the identified genes in the genomes of 20 halotolerant plant growth-promoting bacteria are assessed, and the prevalence of their involved genes is highlighted. The genes related to the synthesis of indole acetic acid (IAA) (70%), siderophores (60%), osmoprotectants (80%), chaperons (40%), 1-aminocyclopropane-1-carboxylate (ACC) deaminase (50%), and antioxidants (50%), phosphate solubilization (60%), and ion homeostasis (80%) were the most common detected genes in the genomes of evaluated halotolerant plant growth-promoting and salinity stress-alleviating bacteria. The most prevalent genes can be applied as candidates for designing molecular markers for screening of new halotolerant plant growth-promoting bacteria

Genome mining conformance to metabolite profile of Bacillus strains to control potato pathogens

Abstract Biocontrol agents are safe and effective methods for controlling plant disease pathogens, such as Fusarium solani, which causes dry wilt, and Pectobacterium spp., responsible for potato soft rot disease. Discovering agents that can effectively control both fungal and bacterial pathogens in potatoes has always presented a challenge. Biological controls were investigated using 500 bacterial strains isolated from rhizospheric microbial communities, along with two promising biocontrol strains: Pseudomonas (T17-4 and VUPf5). Bacillus velezensis (Q12 and US1) and Pseudomonas chlororaphis VUPf5 exhibited the highest inhibition of fungal growth and pathogenicity in both laboratory (48%, 48%, 38%) and greenhouse (100%, 85%, 90%) settings. Q12 demonstrated better control against bacterial pathogens in vivo (approximately 50%). Whole-genome sequencing of Q12 and US1 revealed a genome size of approximately 4.1 Mb. Q12 had 4413 gene IDs and 4300 coding sequences, while US1 had 4369 gene IDs and 4255 coding sequences. Q12 exhibited a higher number of genes classified under functional subcategories related to stress response, cell wall, capsule, levansucrase synthesis, and polysaccharide metabolism. Both Q12 and US1 contained eleven secondary metabolite gene clusters as identified by the antiSMASH and RAST servers. Notably, Q12 possessed the antibacterial locillomycin and iturin A gene clusters, which were absent in US1. This genetic information suggests that Q12 may have a more pronounced control over bacterial pathogens compared to US1. Metabolic profiling of the superior strains, as determined by LC/MS/MS, validated our genetic findings. The investigated strains produced compounds such as iturin A, bacillomycin D, surfactin, fengycin, phenazine derivatives, etc. These compounds reduced spore production and caused deformation of the hyphae in F. solani. In contrast, B. velezensis UR1, which lacked the production of surfactin, fengycin, and iturin, did not affect these structures and failed to inhibit the growth of any pathogens. Our findings suggest that locillomycin and iturin A may contribute to the enhanced control of bacterial pectolytic rot by Q12