Biology subjects

Fernando, L.

Publications and source records attributed to Fernando, L..

4 recordsLinked to original sources

bioRxiv2026

m6A positions polyadenylation in Plasmodium falciparum

N6-methyladenosine (m6A) is the most abundant modification in eukaryotic mRNA and can influence gene expression, but the molecular mechanisms by which it does this at an individual gene level are poorly understood. The eukaryote with the highest known percentage of adenosine in its RNA is P. falciparum, causative agent of the deadliest form of malaria. P. falciparum has a high genomic AT content (>80% A+T) and a particular A-bias in mRNA - coding strands have an average adenosine content of 44%. Plasmodium parasites employ extensive transcriptional and post-transcriptional control over gene expression to differentiate within and between sexual and asexual stages in response to changing environments and hosts. We used direct Nanopore RNA sequencing and GLORI-seq to interrogate m6A methylation of the asexual Plasmodium falciparum transcriptome, revealing m6A depletion in protein coding regions relative to the 3UTR, where m6A is deposited in precise patterns approximately 50 nt upstream of the polyadenylation site. We used an inducible protein mislocalisation system to disrupt the methyltransferase, which writes m6A to mRNA, and observed m6A depletion associated with transcriptional readthrough and chimeric transcripts. We show that m6A in transcript 3UTRs is required for faithful positioning of polyadenylation and transcription termination. This work highlights the importance of m6A to mRNA processing in P. falciparum, and potentially a wider role in post-transcriptional regulation.

molecular biology↗

bioRxiv2020

North American deer mice are susceptible to SARS-CoV-2

The zoonotic spillover of the pandemic SARS-coronavirus 2 (SARS-CoV-2) from an animal reservoir, currently presumed to be the Chinese horseshoe bat, into a naive human population has rapidly resulted in a significant global public health emergency. Worldwide circulation of SARS-CoV-2 in humans raises the theoretical risk of reverse zoonosis events with wildlife, reintroductions of SARS-CoV-2 into permissive non-domesticated animals, potentially seeding new host reservoir species and geographic regions in which bat SARS-like coronaviruses have not historically been endemic. Here we report that North American deer mice (Peromyscus maniculatus) and some closely related members of the Cricetidae family of rodents possess key amino acid residues within the angiotensin-converting enzyme 2 (ACE2) receptor known to confer SARS-CoV-2 spike protein binding. Peromyscus rodent species are widely distributed across North America and are the primary host reservoirs of several emerging pathogens that repeatedly spill over into humans including Borrelia burgdorferi, the causative agent of Lyme disease, deer tick virus, and Sin Nombre orthohantavirus, the causative agent of hantavirus pulmonary syndrome (HPS). We demonstrate that adult deer mice are susceptible to SARS-CoV-2 infection following intranasal exposure to a human isolate, resulting in viral replication in the upper and lower respiratory tract with little or no signs of disease. Further, shed infectious virus is detectable in nasal washes, oropharyngeal and rectal swabs, and viral RNA is detectable in feces and occasionally urine. We further show that deer mice are capable of transmitting SARS-CoV-2 to naive deer mice through direct contact. The extent to which these observations may translate to wild deer mouse populations remains unclear, and the risk of reverse zoonosis and/or the potential for the establishment of Peromyscus rodents as a North American reservoir for SARS-CoV-2 is unknown. Nevertheless, efforts to monitor wild, peri-domestic Peromyscus rodent populations are likely warranted as the SARS-CoV-2 pandemic progresses.

microbiology↗

bioRxiv2018

Modeling the relative role of human mobility, land-use and climate factors on dengue outbreak emergence in Sri Lanka

BackgroundMore than 80,000 dengue cases including 215 deaths were reported nationally in less than seven months between 2016-2017, a fourfold increase in the number of reported cases compared to the average number over 2010-2016. The region of Negombo, located in the Western province, experienced the greatest number of dengue cases in the country and is the focus area of our study, where we aim to capture the spatial-temporal dynamics of dengue transmission.\n\nMethodsWe present a statistical modeling framework to evaluate the spatial-temporal dynamics of the 2016-2017 dengue outbreak in the Negombo region of Sri Lanka as a function of human mobility, land-use, and climate patterns. The analysis was conducted at a 1 km x 1 km spatial resolution and a weekly temporal resolution.\n\nResultsOur results indicate human mobility to be a stronger indicator for local outbreak clusters than land-use or climate variables. The minimum daily temperature was identified as the most influential climate variable on dengue cases in the region; while among the set of land-use patterns considered, urban areas were found to be most prone to dengue outbreak, followed by areas with stagnant water and then coastal areas. The results are shown to be robust across spatial resolutions.\n\nConclusionsOur study highlights the potential value of using travel data to target vector control within a region. In addition to illustrating the relative relationship between various potential risk factors for dengue outbreaks, the results of our study can be used to inform where and when new cases of dengue are likely to occur within a region, and thus help more effectively and innovatively, plan for disease surveillance and vector control.

epidemiology↗

bioRxiv2018

Optimizing guide RNA selection and CRISPR/Cas9 methodology for efficient generation of deletions in C. elegans.

The Caenorhabditis elegans Gene Knockout (KO) Consortium is tasked with obtaining null mutations in each of the more than 20,000 open reading frames (ORFs) of this organism. To date, approximately15,000 ORFs have associated putative null alleles. A directed approach using CRISPR/Cas9 methodology is the most promising technique to complete the task. While there has been substantial success in using CRISPR/Cas9 in C. elegans, there has been little emphasis on optimizing the method for generating large insertions/deletions in this organism. To enhance the efficiency of using CRISPR/Cas9 to generate gene knockouts in C. elegans we have developed an online species-specific guide RNA selection tool (http://genome.sfu.ca/crispr). When coupled with previously developed selection vectors, optimization for homology arm length, and the use of purified Cas9 protein, we demonstrate a robust, efficient and effective protocol for generating deletions. Debate and speculation in the larger scientific community about off- target effects due to non-specific Cas9 cutting has prompted us to investigate through whole genome sequencing the occurrence of single nucleotide variants and indels accompanying targeted deletions. We did not detect any off-site variants above the natural spontaneous mutation rate and therefore conclude this modified protocol does not generate off-target events to any significant degree in C. elegans.

genetics↗