Biology subjects

Paulson, A. R.

Publications and source records attributed to Paulson, A. R..

5 recordsLinked to original sources

bioRxiv2026

Blocking primer improves detection of tick-borne pathogens in Ixodes scapularis (black-legged ticks) from a Lyme disease hotspot region in eastern Ontario, Canada.

Tick-borne pathogen (TBP) surveillance strategies that rely exclusively on targeted methods like PCR (PCR) or immunoblots do not benefit from strain-level sequence variation. Bacterial 16S rRNA metabarcoding offers more agnostic detection but is constrained in I. scapularis by the dominance of a maternally inherited endosymbiont, Rickettsia buchneri. Here we report the design and evaluation of three R. buchneri-specific blocking primers to suppress endosymbiont amplification during full-length 16S rRNA library preparation. Of these, primer 18F-Rb-C3 reduced R. buchneri relative abundance approximately 32-fold. We applied 18F-Rb-C3 with V4-16S metabarcode sequencing on 67 ticks collected from farm animals in Eastern Ontario and compared Borrelia species detection against qPCR. The V4-16S rRNA metabarcoding identified Borrelia species in 21 samples, whereas qPCR detected Borrelia in 24 samples and 11 samples were detected by both methods. Additionally, metabarcoding detected Anaplasma phagocytophilum in 12 samples, including seven samples coinfected with Borrelia, in the same assay. Variation relevant to strain surveillance was also detected by sequencing, though V4-16S was not sufficient to resolve closely related Borrelia genospecies or A. phagocytophilum variants. These findings demonstrate that blocking primer 18F-Rb-C3 enhances sensitivity of amplicon sequencing to the level of qPCR while also detecting other pathogens and sequence variants in a single assay.

genomics↗

bioRxiv2026

Functional genomic analysis reveals mechanisms of epigenetic interference in SARS-CoV-1 and SARS-CoV-2

Small viral RNAs (svRNA) are key factors in host adaptation and virulence and are previously identified in SARS-CoV-2 and SARS-CoV-1. Little is known about svRNA conservation among high-risk coronaviruses (subgenus Sarbecovirus) or how svRNAs might contribute to COVID-19. Here, using functional genomics we identify six novel svRNA-triplex-forming oligonucleotides (svRNA-TFOs) in SARS-CoV-2, and through comparative analyses with SARS-CoV-1, delineate evolutionary pathways for svRNA-TFOs in Sarbecovirus. In addition to significant enrichment of the svRNA-TFO enhancer-associated gene targets among differentially expressed gene sets collected during SARS-CoV-2 infection, we find statistical support for non-random associations with >25 % of recombination breakpoints and hotspots in the Wuhan-Hu-1 genome. Small RNA-sequencing reveals highly abundant svRNA-TFO-N derived from the nucleocapsid of SARS-CoV-2. Using synonymous site conservation analysis, we find svRNA-TFO-N, and other svRNA-TFOs are conserved among both SARS-CoV-2- and SARS-CoV-1-related lineages. Furthermore, BLASTn reveals svRNA-TFO-N, svRNA-TFO-S.1 and svRNA-TFO-E share sequence homology against various mammalian genomes supporting a potential mechanism of host adaptation. Finally, we identify potential links between the putative svRNA-TFO-S.1 and svRNA-TFO-N RNA progenitor structures with variant of concern-associated S:D614G and N:D377Y mutations, respectively. Collectively these findings support a role for epigenetic interference with potential to reveal critical insights into the evolution of SARS-CoV-2 and SARS-CoV-1.

genetics↗

bioRxiv2023

Lysis cassette-mediated exoprotein release in Yersinia entomophaga is controlled by a PhoB-like regulator

Secretion of exoproteins is a key component of bacterial virulence and is tightly regulated in response to environmental stimuli and host-dependent signals. The entomopathogenic bacterium Yersinia entomophaga MH96 produces a wide range of exoproteins including its main virulence factor, the 2.46 MDa insecticidal Yen-Tc toxin complex. Previously, a high-throughput transposon-based screening assay identified the region of exoprotein release (YeRER) as essential to exoprotein release in MH96. The current study defines the role of the YeRER-associated ambiguous holin/endolysin-based lysis cluster (ALC) and the novel RoeA regulator in the regulation and release of exoproteins in MH96. A mutation in the ALC region abolished exoprotein release and caused cell elongation, a phenotype able to be restored through trans-complementation with an intact ALC region. Endogenous ALC did not impact cell growth of the wild type, while artificial expression of an optimised ALC caused cell lysis. Using HolA-sfGFP and Rz1-sfGFP reporter, Rz1 expression was observed in all cells while HolA expression was limited to a small proportion of cells, which increased over time. Transcriptomic assessments found expression of the genes encoding the prominent exoproteins, including the Yen-Tc, was reduced in the roeA mutant and identified a 220 ncRNA of the YeRER intergenic that, when trans complemented in the wildtype, abolished exoprotein release. A model for Y. entomophaga mediated exoprotein regulation and release is proposed. ImportanceWhile theoretical models exist, there is not yet any empirical data that links ALC phage-like lysis cassettes with the release of large macro-molecular toxin complexes such as Yen-Tc in Gram-negative bacteria. In this study, we demonstrate that the novel Y. entomophaga RoeA activates the production of exoproteins (including Yen-Tc) and the ALC at the transcriptional level. The translation of the ALC holin is confined to a subpopulation of cells that then lyse over time, indicative of a complex hierarchical regulatory network. The presence of orthologous RoeA orthologue and a HolA like holin 5 of an eCIS Afp element in Pseudomonas chlororaphis combined with the presented data suggests a shared mechanism is required for the release of some large macromolecular protein assemblies such as the Yen-Tc, and further supports classification of phage-like lysis clusters as type 10 secretion systems.

microbiology↗

bioRxiv2022

Multi-omics analysis identifies symbionts and pathogens of blacklegged ticks (Ixodes scapularis) from a Lyme disease hotspot in southeastern Ontario, Canada

Ticks in the family Ixodidae are important vectors of zoonoses including Lyme disease (LD), which is caused by spirochete bacteria from the Borreliella (Borrelia) burgdorferi sensu lato (Bbsl) complex. The blacklegged tick (Ixodes scapularis) continues to expand across Canada, creating hotspots of elevated LD risk at the leading edge of its expansion range. Current efforts to understand the risk of pathogen transmission associated with I. scapularis in Canada focus primarily on targeted screens, while variation in the tick microbiome remains poorly understood. Using multi-omics consisting of 16S metabarcoding and ribosome-depleted, whole-shotgun RNA transcriptome sequencing, we examined the microbial communities associated with adult I. scapularis (N = 32), sampled from four tissue types (whole tick, salivary glands, midgut, and viscera) and three geographical locations within a LD hotspot near Kingston, Ontario, Canada. The communities consisted of both endosymbiotic and known or potentially pathogenic microbes, including RNA viruses, bacteria, and a Babesia sp. intracellular parasite. We show that {beta}-diversity is significantly higher between individual tick salivary gland and midgut bacterial communities, compared to whole ticks; while linear discriminant analysis (LDA) effect size (LEfSe) determined that the three potentially pathogenic bacteria detected by V4 16S rDNA sequencing also differed among dissected tissues only, including a Borrelia from the Bbsl complex, Borrelia miyamotoi, and Anaplasma phagocytophilum. Importantly, we find co-infection of I. scapularis by multiple microbes, in contrast to diagnostic protocols for LD, which typically focus on infection from a single pathogen of interest (B. burgdorferi sensu stricto). IMPORTANCEA vector of human health concern, blacklegged ticks, Ixodes scapularis, transmit pathogens that cause tick-borne diseases (TBDs), including Lyme disease (LD). Several hotspots of elevated LD risk have emerged across Canada as I. scapularis expands its range. Focusing on a hotspot in southeastern Ontario, we used high-throughput sequencing on whole ticks and dissected salivary glands and midguts. Compared to whole ticks, analysis of salivary glands and midguts revealed greater {beta}-diversity among microbiomes that are less dominated by Rickettsia endosymbiont bacteria and enriched for pathogenic bacteria including a Bbsl- associated Borrelia, Borrelia miyamotoi, and Anaplasma phagocytophilum. We also find evidence of co-infection of I. scapularis in this region by multiple microbes. Overall, our study highlights the challenges and opportunities associated with the surveillance of the microbiome of I. scapularis for pathogen detection using metabarcoding and metatranscriptome approaches.

microbiology↗

bioRxiv2020

In vivo transcriptome analysis provides insights into host-dependent expression of virulence factors by Yersinia entomophaga MH96, during infection of Galleria mellonella

The function of microbes can be inferred from knowledge of genes specifically expressed in natural environments. Here we report the in vivo transcriptome of the entomopathogenic bacterium Yersinia entomophaga MH96, captured during three stages of intrahemocoelic infection in Galleria mellonella. A total of 1,285 genes were significantly upregulated by MH96 during infection; 829 genes responded to in vivo conditions during at least one stage of infection, 289 responded during two stages of infection and 167 transcripts responded throughout all three stages of infection. Genes upregulated during the earliest infection stage included components of the insecticidal toxin complex Yen-TC (chi1, chi2 and yenC1), genes for rearrangement hotspot element containing protein yenC3, cytolethal distending toxin cdtAB and vegetative insecticidal toxin vip2. Genes more highly expressed throughout the infection cycle included the putative heat-stable enterotoxin yenT and three adhesins (usher-chaperone fimbria, filamentous hemagglutinin and an AidA-like secreted adhesin). Clustering and functional enrichment of gene expression data also revealed expression of genes encoding type III and VI secretion system-associated effectors. Together these data provide insight into the pathobiology of MH96 and serve as an important resource supporting efforts to identify novel insecticidal agents.

microbiology↗