Comprehensive analysis of hairpin small RNAs involved in resistance and pathogenesis during wheat-Puccinia triticina interactions.

Wheat leaf rust, caused by the fungus Puccinia triticina (Pt), severely affects the grain quality and quantity of bread wheat (Triticum aestivum L.). Hairpin small(s)RNAs, like micro(mi)RNAs and their variants [including isomiRNAs (isomiRs) and microRNA-like RNAs (milRNAs)], along with their corresponding target genes, bestow leaf rust disease resistance, development and progression from both interacting species. However, the regulatory networks remain inadequately understood. Thirteen differentially expressed novel miRNAs, including two isomiRs and three milRNAs were discerned from induced reads of wheat sRNA libraries, and a further 5,393 and 1,275 candidate target genes were predicted in wheat and Pt, respectively. Functional annotation divulged that wheat-originated miRNAs/isomiRs were involved in resistance, while Pt-derived milRNAs imparted pathogenesis. The identified milRNAs- Tae-Pt-milR5, Tae-Pt-milR12, and Tae-Pt-milR14b and their cleavage sites on Pt target gene MEP5 were confirmed through degradome library screening, suggesting cross-kingdom translocation of Pt virulent genes in wheat host. Co-expression analysis of miRNAs/isomiRs-target genes provided insights into combating leaf rust disease, while co-expression analysis of milRNAs-target gene pairs reflected the extent of pathogenicity exerted by Pt with varied expression levels at the analyzed time points. The analysis pinpointed leaf rust-responsive candidate hairpin sRNAs- Tae-miR8, Tae-Pt-miR12, Tae-Pt-miR14a, and Tae-Pt-miR14b in wheat and Tae-Pt-milR12 in Pt. This study provides new insights into the hairpin sRNAs involved in the resistance and pathogenesis of wheat and Pt, respectively. Furthermore, crucial hairpin sRNAs and their promising targets for future biotechnological interventions to augment stress resilience have been identified.

5-mC methylation study of sORFs in 3'UTR of transcription factor JUNGBRUNNEN 1-like during leaf rust pathogenesis in wheat.

BACKGROUND: JUB1, a NAC domain containing hydrogen peroxide-induced transcription factor, plays a critical role in plant immunity. Little is known about how JUB1 responds to leaf rust disease in wheat. Recent discoveries in genomics have also unveiled a multitude of sORFs often assumed to be non-functional, to argue for the necessity of including them as potential regulatory players of translation. However, whether methylation on sORFs spanning the 3'UTR of regulatory genes like JUB1 modulate gene expression, remains unclear. METHODS AND RESULTS: In this study, we identified the methylation states of two sORFs in 3'UTR of a homologous gene of JUB1 in wheat, TaJUB1-L, at cytosine residues in CpG, CHH and CHG sites at different time points of disease progression in two near-isogenic lines of wheat (HD2329), with and without Lr24 gene during leaf rust pathogenesis. Here, we report a significant demethylation of the CpG dinucleotides occurring in the sORFs of the 3'UTR in the resistant isolines after 24 h post-infection. Also, the up-regulated gene expression observed through RT-qPCR was directly proportional to the demethylation of the CpG sites in the sORFs. CONCLUSIONS: Our findings indicate that TaJUB1-L might be a positive regulator in providing tolerance during leaf rust pathogenesis and cytosine methylation at 3'UTR might act as a switch for its expression control. These results enrich the potential benefit of conventional methylation assay techniques for unraveling the unexplored enigma in epigenetics during plant-pathogen interaction in a cost-effective and confidentially conclusive manner.

Contraceptive knowledge, attitude and practice among rural women.

OBJECTIVE: To assess the knowledge, attitude regarding family planning and the practice of contraceptives among rural women. STUDY DESIGN: A cross-sectional observational study. PLACE AND DURATION OF STUDY: The gynaecological outpatient clinic of Fatima Hospital, Baqai Medical University, Karachi, from July to December 2005. METHODOLOGY: One-hundred women between the ages 15-45, living with their husbands and coming from rural area (villages) were interviewed. Women who were pregnant, had a child younger than 2 years, or had any medical disorder were excluded. Their knowledge, attitude and practice on contraceptives were evaluated with the help of a predesigned questionnaire. The other variables used were the age of women, parity and educational status. Descriptive analysis was conducted to obtain percentages. RESULTS: Out of 100 interviewed women with mean age of 29.7 years, 81(81%) had some knowledge about family planning methods. The media provided information of contraceptives in 52 out of 81 (64%) women. Regarding the usage of contraceptive methods, only 53 (53%) of the respondents were using some sort of contraception. Barrier method (condoms) was in practice by 18 (33.9%) and 12 (22.6%) of women had already undergone tubal ligation. The women using injectables and intrauterine contraceptive devices were 10 (18.8%) and 7 (13.2%) respectively. Six were using oral contraceptive pills (11.3%). Positive attitude towards contraception was shown by 76 (76%) of them, while 41(41%) stated their husbands' positive attitude towards contraception. CONCLUSION: In the present study, there was a low contraceptive use among women of rural origin despite good knowledge. Motivation of couples through media and health personnel can help to achieve positive attitude of husbands for effective use of contraceptives.