Biology subjects

Upadhyaya, N.

Publications and source records attributed to Upadhyaya, N..

5 recordsLinked to original sources

bioRxiv2023

High prevalence of Bovine Tuberculosis reported in Cattle and Buffalo of Eastern Nepal

Livestock farming, specifically of cattle and buffalo, is crucial to Nepals economy, with more than 66% of the population involved in agriculture and animal husbandry. Animal tuberculosis (TB) caused by Mycobacterium bovis is a chronic disease that affects these animals and results in economic losses due to reduced milk and meat productivity, fertility and mortality. M. bovis also infects humans, non-human primates, goats and other mammals, and can afflict both cattle and buffalo. Our study is a part of routine surveillance of prevalent diseases, including M. bovis, in cattle and buffalo. We collected blood samples (n=400, 100 samples from each district) from selected eastern districts of Nepal. We used a Rapid Bovine TB Test Kit to test these samples for presence of M. bovis. Of the 400 samples collected, 74 animals (18.75%) tested positive for M. bovis, with the majority of positive samples coming from cattle (n=71, 17.75%) and only three from buffalo (<1%). Among the screened breeds of cattle and buffalo, Holstein Friesian cattle (HF) (n=43, 58%), Jersey-cross cattle (JX) (n=20, 27%), Local buffalo (n=8, 10.8%) and Murra breed buffalo (n=3, 4.1%) were found to carry M. bovis. The majority (50%) of infected animals were between 3-6 years old. Morang (n= 24 positive in cattle; n=1 positive in buffalo) and Jhapa (n= 22 positive in cattle; n=2 positive in buffalo) had the highest prevalence of M. bovis, while all the positive cases in Sunsari (n=19) and Udaypur (n=6) were in cattle. The fact that over 18% of the samples tested positive for M. bovis is of great concern. It is critical to thoroughly test animal products from these livestock prior to human consumption. To prevent and mitigate M. bovis-related infections in Nepal, a more comprehensive screening strategy coupled with more effective animal husbandry practices needs to be adapted.

microbiology↗

bioRxiv2021

Identification and correction of phase switches with Hi-C data in the Nanopore and HiFi chromosome-scale assemblies of the dikaryotic leaf rust fungus Puccinia triticina

BackgroundMost animals and plants have more than one set of chromosomes and package these haplotypes into a single nucleus within each cell. In contrast, many fungal species carry multiple haploid nuclei per cell. Rust fungi are such species with two nuclei (karyons) that contain a full set of haploid chromosomes each. The physical separation of haplotypes in dikaryons means that, unlike in diploids, Hi-C chromatin contacts between haplotypes are false positive signals. ResultsWe generate the first chromosome-scale, fully-phased assembly for the dikaryotic leaf rust fungus Puccinia triticina and compare Nanopore MinION and PacBio HiFi sequence-based assemblies. We show that false positive Hi-C contacts between haplotypes are predominantly caused by phase switches rather than by collapsed regions or Hi-C read mis-mappings. We introduce a method for phasing of dikaryotic genomes into the two haplotypes using Hi-C contact graphs, including a phase switch correction step. In the HiFi assembly, relatively few phase switches occur, and these are predominantly located at haplotig boundaries and can be readily corrected. In contrast, phase switches are widespread throughout the Nanopore assembly. We show that haploid genome read coverage of 30-40 times using HiFi sequencing is required for phasing of the leaf rust genome (~0.7% heterozygosity) and that HiFi sequencing resolves genomic regions with low heterozygosity that are otherwise collapsed in the Nanopore assembly. ConclusionsThis first Hi-C based phasing pipeline for dikaryons and comparison of long-read sequencing technologies will inform future genome assembly and haplotype phasing projects in other non-haploid organisms.

genomics↗

bioRxiv2019

Emergence of the Ug99 lineage of the wheat stem rust pathogen through somatic hybridisation

Parasexuality contributes to diversity and adaptive evolution of haploid (monokaryotic) fungi. However non-sexual genetic exchange mechanisms are not defined in dikaryotic fungi (containing two distinct haploid nuclei). Newly emerged strains of the wheat stem rust pathogen, Puccinia graminis f. sp. tritici (Pgt), such as Ug99, are a major threat to global food security. Here we show that Ug99 arose by somatic hybridisation and nuclear exchange between dikaryons. Fully haplotype-resolved genome assembly and DNA proximity analysis revealed that Ug99 shares one haploid nucleus genotype with a much older African lineage of Pgt, with no recombination or reassortment. Generation of genetic variation by nuclear exchange may favour the evolution of dikaryotism by providing an advantage over diploidy.

genetics↗

bioRxiv2018

The stem rust fungus Puccinia graminis f. sp. tritici induces waves of small RNAs with opposing profiles during wheat infection

BackgroundSilencing of transposable elements (TEs) is essential for maintaining genome stability. Plants use small RNAs (sRNAs) to direct DNA methylation to TEs (RNA-directed DNA methylation; RdDM). Similar mechanisms of epigenetic silencing in the fungal kingdom have remained elusive. ResultsWe use sRNA sequencing and methylation data to gain insight into epigenetics in the dikaryotic fungus Puccinia graminis f. sp. tritici (Pgt), which causes the devastating stem rust disease on wheat. We use Hi-C data to define the Pgt centromeres and show that they are repeat-rich regions ([~]250 kb) that are highly diverse in sequence between haplotypes and, like in plants, are enriched for young TEs. DNA cytosine methylation is particularly active at centromeres but also associated with genome-wide control of young TE insertions. Strikingly, over 90% of Pgt sRNAs and several RNAi genes are differentially expressed during infection. Pgt induces waves of functionally diversified sRNAs during infection. The early wave sRNAs are predominantly 21 nts with a 5 uracil derived from genes. In contrast, the late wave sRNAs are mainly 22 nt sRNAs with a 5 adenine and are strongly induced from centromeric regions. TEs that overlap with late wave sRNAs are more likely to be methylated, both inside and outside the centromeres, and methylated TEs exhibit a silencing effect on nearby genes. ConclusionsWe conclude that rust fungi use an epigenetic silencing pathway that resembles RdDM in plants. The Pgt RNAi machinery and sRNAs are under tight temporal control throughout infection and might ensure genome stability during sporulation.

microbiology↗

bioRxiv2017

De novo assembly and phasing of dikaryotic genomes from two isolates of Puccinia coronata f. sp. avenae, the causal agent of oat crown rust

Oat crown rust, caused by the fungus Puccinia coronata f. sp. avenae (Pca), is a devastating disease that impacts worldwide oat production. For much of its life cycle, Pca is dikaryotic, with two separate haploid nuclei that may vary in virulence genotype, highlighting the importance of understanding haplotype diversity in this species. We generated highly contiguous de novo genome assemblies of two Pca isolates, 12SD80 and 12NC29, from long-read sequences. In total, we assembled 603 primary contigs for a total assembly length of 99.16 Mbp for 12SD80 and 777 primary contigs with a total length of 105.25 Mbp for 12NC29, and approximately 52% of each genome was assembled into alternate haplotypes. This revealed structural variation between haplotypes in each isolate equivalent to more than 2% of the genome size, in addition to about 260,000 and 380,000 heterozygous single-nucleotide polymorphisms in 12SD80 and 12NC29, respectively. Transcript-based annotation identified 26,796 and 28,801 coding sequences for isolates 12SD80 and 12NC29, respectively, including about 7,000 allele pairs in haplotype-phased regions. Furthermore, expression profiling revealed clusters of co-expressed secreted effector candidates, and the majority of orthologous effectors between isolates showed conservation of expression patterns. However, a small subset of orthologs showed divergence in expression, which may contribute to differences in virulence between 12SD80 and 12NC29. This study provides the first haplotype-phased reference genome for a dikaryotic rust fungus as a foundation for future studies into virulence mechanisms in Pca.\n\nImportanceDisease management strategies for oat crown rust are challenged by the rapid evolution of Puccinia coronata f. sp. avenae (Pca), which renders resistance genes in oat varieties ineffective. Despite the economic importance of understanding Pca, resources to study the molecular mechanisms underpinning pathogenicity and emergence of new virulence traits are lacking. Such limitations are partly due to the obligate biotrophic lifestyle of Pca as well as the dikaryotic nature of the genome, features that are also shared with other important rust pathogens. This study reports the first release of a haplotype-phased genome assembly for a dikaryotic fungal species and demonstrates the amenability of using emerging technologies to investigate genetic diversity in populations of Pca.

genomics↗