Biology subjects

Saini, J. S.

Publications and source records attributed to Saini, J. S..

5 recordsLinked to original sources

bioRxiv2025

Decoding efficacy and resistance space at a drug binding site

Interactions between drugs and their targets impact efficacy and, when altered by mutation, can result in resistance 1-3. Assessing and understanding the impacts of all possible mutations at a drug binding site remain challenging, however 4-6. Here, we used Multiplex Oligo Targeting (MOT) for mutational profiling, and computational modelling, to decode efficacy and resistance space at the otherwise native binding site for a low nanomolar potency, anti-trypanosomal, proteasome inhibitor 7. We saturation-edited twenty codons in the Trypanosoma brucei proteasome {beta}5 subunit and subjected the resulting MOT libraries to stepwise drug selection. Amplicon sequencing, and codon variant scoring, yielding dose-response profiles for >100 resistance-conferring mutants, among 1,280 possible codon variants. Codon variant scores were predictive of relative resistance observed using a bespoke set of mutants, while fitness profiling revealed otherwise extensive constraints on mutational fitness and resistance space. The resistance profile that emerged allowed us to readily predict routes to spontaneous drug resistance observed within accessible, single nucleotide mutational space. In silico analysis of {beta}5 subunit mutations predicted impacts on ligand affinity via steric effects, hydrogen-bonding and lipophilicity, which when combined with predictions of proteasome function perturbing mutations, were closely aligned with observed impacts on drug resistance. We conclude that MOT-library profiling facilitates assessment of all possible mutations at a drug binding site. Further decoding of drug target structure-activity relationships and drug resistance space will facilitate the design of more effective and durable drugs.

molecular biology↗

bioRxiv2024

Multiple extracellular polymeric substances pathways expressed by Accumulibacter and the flanking community during aerobic granule formation and after influent modification

Aerobic granular sludge is a biological wastewater treatment process in which a microbial community forms a granular biofilm. The role of Candidatus Accumulibacter in the production of a biofilm matrix composed of extracellular polymeric substances was studied in a sequencing batch reactor enriched with polyphosphate-accumulating organisms. The metabolisms of the microbial populations were investigated using de novo metatranscriptomics analysis. Finally, the effect of decreasing the influent phosphate concentration was investigated. A few weeks after the reactor start-up, the microbial community was dominated by Accumulibacter. Up to nine species were active in parallel. However, the most active species differed according to sampling time. Reducing the phosphate concentration led to a dominance of the glycogen-accumulating organism Propionivibrio, with some Accumulibacter species still abundant. De novo metatranscriptomic analysis indicated a high diversity of potential extracellular substances produced mainly by Azonexus, Accumulibacter, Candidatus Contendobacter, and Propionivibrio. Moreover, the results suggest that Azonexus, Contendobacter and Propionivibrio recycle the neuraminic acid produced by Accumulibacter. Changes in the microbial community did not cause the granules to disintegrate, indicating that a Propionivibrio-dominated community can maintain stable granules.

microbiology↗

bioRxiv2023

Benchmarking combined informatics approaches for virus discovery: Caution is needed when combining in silico identification methods

BackgroundThe identification of viruses from environmental metagenomic samples using informatics tools has offered critical insights in microbiome studies. However, it remains difficult for researchers to know for their specific study which tool(s) and settings are best suited to maximize capture of viruses while minimizing false positives. Studies are increasingly combining multiple tool outputs attempting to recover more viruses, but no combined approach has been benchmarked for accuracy. Here, we benchmarked 63 viral identification rulesets against mock metagenomes composed of publicly available viral, bacterial, archaeal, fungal, and protist sequences. These rulesets are based on combinations of four single-tool rules and two multi-tool tuning rules. We applied these rulesets to various aquatic metagenomes and filtering strategies to evaluate the impact of habitat and viral enrichment on individual and combined tool performance. We provide a packaged pipeline for researchers that want to replicate our process. ResultsWe found that combining rules increased viral recall, but at the expense of increased false positives. Six of the 63 combinations tested had equivalent accuracies to the highest one (MCC=0.77, padj [≥] 0.05). All of the six high accuracy rulesets included VirSorter2, five included our "tuning removal" rule, and no high performing rulesets used more than four of our six rules. DeepVirFinder, VIBRANT, and VirSorter were each found once in these high accuracy rulesets, but never in combination with each other. Our validation suggests that the MCC plateau at 0.77 is caused by inaccurate labeling of the data that viral identification tools rely on for training and validation. In the aquatic metagenomes, our "highest MCC" ruleset identified a higher proportion of viral sequences in the virus-enriched samples (44-46%) than the non-enriched, cellular metagenomes (7-19%). ConclusionWhile improved algorithms may lead to more accurate viral identification tools, this should be done in tandem with curating accurately labeled viral gene and sequence databases. For most applications, we recommend the use of the ruleset that uses VirSorter2 and our empirically derived tuning removal rule. By providing a rigorous overview of the behavior of in silico viral identification strategies, our findings guide the use of existing viral identification tools and offer a blueprint for feature engineering of new tools that will lead to higher-confidence viral discovery in microbiome studies.

bioinformatics↗

bioRxiv2021

Bacterial, phytoplankton, and viral dynamics of meromictic Lake Cadagno offer insights into the Proterozoic ocean microbial loop

Lake Cadagno, a permanently stratified high-alpine lake with a persistent microbial bloom in its anoxic chemocline, has long been considered a model for the low-oxygen, high-sulfide Proterozoic ocean where early microbial life gave rise to Earths oxygenated atmosphere. Although the lake has been studied for over 25 years, the absence of concerted study of the bacteria, phytoplankton, and viruses, together with primary and secondary production, has hindered a comprehensive understanding of its microbial food web. Here, the identities, abundances, and productivity of microbes were evaluated in the context of Lake Cadagno biogeochemistry. Photo-synthetic pigments and chloroplast 16S rRNA gene phylogenies suggested high abundances of eukaryotic phytoplankton, primarily Chlorophyta, through the water column. Of these, a close relative of Ankyra judayi, a high-alpine adapted chlorophyte, peaked with oxygen in the mixolimnion, while Closteriopsis-related chlorophytes peaked in the chemocline and monimolimnion. Anoxygenic phototrophic sulfur bacteria, Chromatium, dominated the chemocline along with Lentimicrobium, a newly observed genus of known fermenters. Secondary production peaked in the chemocline, suggesting anoxygenic primary producers depended on heterotrophic nutrient remineralization. Virus-to-microbe ratios spanned an order of magnitude, peaking with high phytoplankton abundances and at a minimum at the peak of Chromatium, dynamic trends that suggest viruses may play a role in the modulation of oxygenic and anoxygenic photo- and chemosynthesis in Lake Cadagno. Through the combined analysis of bacterial, eukaryotic, viral, and biogeochemical dynamics of Lake Cadagno, this study provides a new perspective on the biological and geochemical connections that comprised the food webs of the Proterozoic ocean. IMPORTANCEAs a window to the past, the study offers insights into the role of microbial guilds of Proterozoic ocean chemoclines in the production and recycling of organic matter of sulfur- and ammonia-containing ancient oceans. The new observations described here suggest that eukaryotic algae were persistent in the low oxygen upper-chemocline in association with purple and green sulfur bacteria in the lower half of the chemocline. Further, this study provides the first insights into Lake Cadagno viral ecology. High viral abundances suggested viruses may be essential components of the chemocline where their activity may result in the release and recycling of organic matter. The framework developed in this study through the integration of diverse geochemical and biological data types lays the foundation for future studies to quantitatively resolve the processes performed by discrete populations comprising the microbial loop in this early anoxic ocean analogue.

microbiology↗

bioRxiv2021

Microbial loop of Proterozoic ocean analogue

Meromictic Lake Cadagno, an ancient ocean analogue, is known for its permanent stratification and persistent anoxygenic microbial bloom within the chemocline. Although the anaerobic microbial ecology of the lake has been extensively studied for at least 25 years, a comprehensive picture of the microbial food web linking the bacterial layer to phytoplankton and viruses, with explicit measures of primary and secondary production, is still missing. This study sought to better understand the abundances and productivity of microbes in the context of nutrient biogeochemical cycling across the stratified zones of Lake Cadagno. Photosynthetic pigments and chloroplast 16S rRNA gene phylogenies suggested the presence of eukaryotic phytoplankton through the water column. Evidence supported high abundances of Ankyra judayi, a high-alpine adapted chlorophyte, in the oxic mixolimnion where oxygenic-primary production peaked. Through the no- and low-oxygen chemocline and monimolimnion, chlorophytes related to Closteriopsis acicularis, a known genus of meromictic lakes, and Parachlorella kessleri were observed. Chromatium, anoxygenic phototrophic sulfur bacteria, dominated the chemocline along with Lentimicrobium, a genus of known fermenters whose abundance was newly reported in Lake Cadagno. Secondary production peaked in the chemocline suggesting primary producers depend on hetrotrophs for nutrient remineralization. As previously observed, sulfur reducing bacteria (SRBs), especially Desulfocapsa and Desulfobulbus, were present in the chemocline and anoxic monimolimnion. Virus-to-microbe ratios (VMR) peaked in the zone of phytoplankton, yet were at a minimum at the peak of Chromatium. These dynamic trends suggest viruses may play a role in the modulation of oxygenic and anoxygenic photo- and chemosynthesis in Lake Cadagno and other permanently stratified systems. ImportanceAs a window to the past, the study offers insights into the role of microbial guilds of Proterozoic ocean chemoclines in the production and recycling of organic matter of sulfur- and ammonia-containing ancient oceans. The new observations described here suggest that eukaryotic algae were persistent in the low oxygen upper-chemocline in association with purple and green sulfur bacteria in the lower half of the chemocline. Further, this study provides the first insights into Lake Cadagno viral ecology. High viral abundances suggested viruses may be essential components of the chemocline where their activity may result in the release and recycling of organic matter. The framework developed in this study through the integration of diverse geochemical and biological data types lays the foundation for future studies to quantitatively resolve the processes performed by discrete populations comprising the microbial loop in this early anoxic ocean analogue.

microbiology↗