Biology subjects

Granek, J. A.

Publications and source records attributed to Granek, J. A..

5 recordsLinked to original sources

bioRxiv2026

From external-input sensitivity to resident persistence: community assembly in a sink p-trap model

Microbial habitats that receive repeated external input may not remain shaped by that input forever if local retention allows resident communities to build up over time. Here, we used a controlled bench-scale sink p-trap system to examine how community assembly unfolded during initial establishment in new, bleach-treated p-traps. Two p-traps received repeated handwashing-water input, while one received tap water as baseline. The treated p-traps, but not the control, showed clear successional change toward later resident-like states. Nested-model comparisons further showed that recent external input had its greatest influence early in succession, but the p-traps own prior state remained the stronger predictor throughout. Final-day post-flush trajectories indicated short-term displacement from pre-flush positions, with later time points tending to move back toward late-stage resident centroids. Together, these results show that repeated inoculation does not necessarily keep communities under continued outside influence. Instead, retentive microbial habitats can shift over time from early sensitivity to external input toward persistence shaped more by local history.

ecology↗

bioRxiv2025

A simple library preparation modification significantly reduces barcode crosstalk in ONT multiplexed sequencing

Barcode crosstalk is a potential source of error in indexed multiplex sequencing that can mimic cross-contamination and generate false-positive signals, particularly in experiments with low-biomass samples and imbalanced designs. Here we identify and quantify barcode crosstalk in multiplexed Oxford Nanopore sequencing and introduce post-ligation pooling (PLP). PLP is a drop-in library-preparation modification that prevents barcode crosstalk, rather than simply mitigating its effects. These results highlight the potential for barcode crosstalk to have detrimental effects in multiplexed long-read sequencing and establish PLP as an immediately adoptable mitigation, especially where negative controls and low-abundance signals inform biological interpretation.

genomics↗

bioRxiv2025

A nonhuman primate model mirrors human congenital cytomegalovirus infection and reveals a spectrum of vertical transmission outcomes

Congenital cytomegalovirus (cCMV) is the leading infectious cause of birth defects worldwide, yet immune determinants of protection to inform design of a maternal vaccine remain elusive. Here, we characterized the outcome of primary rhesus CMV (RhCMV) infection during pregnancy in an immune competent nonhuman primate (NHP) model. RhCMV DNA was detected in amniotic fluid and/or fetal tissues in six of 12 (50% placental transmission) CMV-naive rhesus macaque dams inoculated intravenously with RhCMV in early second trimester gestation. Widespread tissue dissemination dominated by one of two inoculated RhCMV strains was present in one fetus (8.3% cCMV disease). Placental RhCMV transmission was associated with elevated fetal and maternal plasma TNF-alpha and reduced maternal brain-derived neurotrophic factor and IL-10 levels. CMV exposure during pregnancy had a broad impact on the placenta and fetus even in the absence of congenital infection as evidenced by RhCMV infection at the maternal-fetal interface in all 12 dams, along with significantly reduced placental efficiency and fetal growth metrics compared to gestation-matched control pregnancies. This NHP model recapitulates key aspects of human cCMV and provides new insight into barriers and biomarkers of successful vertical transmission. One sentence summary: The nonhuman primate model mirrors the epidemiology of human congenital CMV (cCMV) after primary infection and reveals its transmission bottlenecks.

immunology↗

bioRxiv2024

Malassezia responds to environmental pH signals through the conserved Rim/Pal pathway

During mammalian colonization and infection, microorganisms must be able to rapidly sense and adapt to changing environmental conditions including alterations in extracellular pH. The fungus-specific Rim/Pal signaling pathway is one process that supports microbial adaptation to alkaline pH. This cascading series of interacting proteins terminates in the proteolytic activation of the highly conserved Rim101/PacC protein, a transcription factor that mediates microbial responses that favor survival in neutral/alkaline pH growth conditions, including many mammalian tissues. We identified the putative Rim pathway proteins Rim101 and Rra1 in the human skin colonizing fungus Malassezia sympodialis. Gene deletion by transconjugation and homologous recombination revealed that Rim101 and Rra1 are required for M. sympodialis growth at higher pH. Additionally, comparative transcriptional analysis of the mutant strains compared to wild-type suggested mechanisms for fungal adaptation to alkaline conditions. These pH-sensing signaling proteins are required for optimal growth in a murine model of atopic dermatitis, a pathological condition associated with increased skin pH. Together these data elucidate both conserved and phylum-specific features of microbial adaptation to extracellular stresses. ImportanceThe ability to adapt to host pH has been previously associated with microbial virulence in several pathogenic fungal species. Here we demonstrate that a fungal-specific alkaline response pathway is conserved in the human skin commensal fungus Malassezia sympodialis (Ms). This pathway is characterized by the pH-dependent activation of the Rim101/PacC transcription factor that controls cell surface adaptations to changing environmental conditions. By disrupting genes encoding two predicted components of this pathway, we demonstrated that the Rim/Pal pathway is conserved in this fungal species as a facilitator of alkaline pH growth. Moreover, targeted gene mutation and comparative transcriptional analysis supports the role of the Ms Rra1 protein as a cell surface pH sensor conserved within the basidiomycete fungi, a group including plant and human pathogens. Using an animal model of atopic dermatitis, we demonstrate the importance of Ms Rim/Pal signaling in this common inflammatory condition characterized by increased skin pH.

microbiology↗

bioRxiv2021

Comparative analysis of RNA enrichment methods for preparation of Cryptococcus neoformans RNA sequencing libraries

Ribosomal RNA (rRNA) is the major RNA constituent of cells, therefore most RNA sequencing (RNA-Seq) experiments involve removal of rRNA. This process, called RNA enrichment, is done primarily to reduce cost: without rRNA removal, deeper sequencing would need to be performed to balance the sequencing reads wasted on rRNA. The ideal RNA enrichment method would remove all rRNA without affecting other RNA in the sample. We have tested the performance of three RNA enrichment methods on RNA isolated from Cryptococcus neoformans, a fungal pathogen of humans. We show that the RNase H depletion method unambiguously outperforms the commonly used Poly(A) isolation method: the RNase H method more efficiently depletes rRNA while more accurately recapitulating the expression levels of other RNA observed in an unenriched "gold standard". The RNase H depletion method is also superior to the Ribo-Zero depletion method as measured by rRNA depletion efficiency and recapitulation of protein-coding gene expression levels, while the Ribo-Zero depletion method performs moderately better in preserving non-coding RNA (ncRNA). Finally, we have leveraged this dataset to identify novel long non-coding RNA (lncRNA) genes and to accurately map the C. neoformans mitochondrial rRNA genes. ARTICLE SUMMARYWe compare the efficacy of three different RNA enrichment methods for RNA-Seq in Cryptococcus neoformans: RNase H depletion, Ribo-Zero depletion, and Poly(A) isolation. We show that the RNase H depletion method, which is evaluated in C. neoformans samples for the first time here, is highly efficient and specific in removing rRNA. Additionally, using data generated through these analyses, we identify novel long non-coding RNA genes in C. neoformans. We conclude that RNase H depletion is an effective and reliable method for preparation of C. neoformans RNA-Seq libraries.

microbiology↗