Biology subjects

Penttinen, R.

Publications and source records attributed to Penttinen, R..

8 recordsLinked to original sources

bioRxiv2026

Otenabant is a Selective Antagonist of Human PIEZO1

Background and purposePIEZO1 mechanosensitive cation channels translate mechanical cues into intracellular Ca2+ and Na+ elevations, enabling cells to respond to physical alterations in their environment. PIEZO1 contributes to red blood cells (RBC) volume homeostasis and gain-of-function PIEZO1 mutations cause hereditary xerocytosis (HX), a rare mostly compensated hemolytic anemia, and aberrant channel activation exacerbates sickling and vascular dysfunction in sickle cell disease. Despite strong genetic and physiological evidence supporting PIEZO1 as a therapeutic target, potent and selective inhibitors are limited, and existing compounds show modest specificity or poorly explored mechanisms. Improved pharmacological tools are needed. Experimental approachWe conducted a high-throughput screen of FDA-approved drugs to identify PIEZO1 inhibitors. Compounds were tested at concentrations of 10 {micro}M in a monocytic cell line, using intracellular Ca2+ elevations evoked by the PIEZO1 agonist Yoda1 as read-out. The inhibitory activity of the best hit was validated and compared to existing PIEZO1 inhibitors using electrophysiological analysis, orthogonal PIEZO1-dependent assays across cell lines and human RBCs. As functional proof, we investigated the impact of three PIEZO1 inhibitors on RBC deformability by ektacytometry, after Yoda1 pre-stimulation. Key resultsThis screen identified Otenabant, a selective Cannabinoid Receptor Type 1 (CB1) antagonist, as a potent PIEZO1 inhibitor. Otenabant dose-dependently inhibited Ca2+ elevations mediated by endogenous or exogenously expressed human PIEZO1, but was ineffective against mouse Piezo1, revealing species-specific channel differences. Otenabant inhibited mechanosensitive currents elicited by shear stress in fibroblasts and by repeated poking in PIEZO1-expressing HEK-293 cells, altering the currents activation and inactivation kinetics, and prevented Yoda1-induced hyperpolarization in RBCs. Otenabant was able to reverse the negative impact of Yoda1 on RBC deformability. Conclusions and implicationsThese findings demonstrate the utility of Yoda-based screening for discovering PIEZO1 antagonists and identify Otenabant as a promising chemical scaffold for developing selective PIEZO1 inhibitors with therapeutic potential.

pharmacology and toxicology↗

bioRxiv2026

Mucin modulates phage infection dynamics and biofilm formation in enteropathogenic Yersinia enterocolitica

Mucosal barriers serve as a multifunctional interface and nutrient-rich habitat for diverse microbes, including bacteria and bacteriophages. Some phages can bind to mucin glycoproteins via carbohydrate-interacting modules and provide an additional layer of mucosal immunity by shielding the underlying epithelium from invading bacteria. However, the role of mucins in shaping phage-bacterium interactions remains poorly understood. We investigated the dynamics between highly pathogenic Yersinia enterocolitica serotype O:8 and its mucus-adherent phage fMtkYen801 under in vitro mucosal environment. We assessed how mucin supplementation, varying phage doses, nutrient and temperature conditions influence phage-bacterium dynamics and biofilm formation. We found that pre-exposure to mucins led to enhanced phage replication in the bacterial host, with a 2-log increase in phage titers, and high abundance of surviving bacteria. Interestingly, mucin glycoproteins also provided Y. enterocolitica a nutrient source and a chemical cue to modulate its growth and biofilm biogenesis. Genomic analysis of phage-resistant bacterial variants revealed mutations in virulence, quorum sensing, and antibiotic resistance genes in both mucin enrichment and control groups, suggesting potential fitness tradeoffs during resistance evolution. Collectively, these findings highlight the importance of mucosal surfaces as an important ecological driver of phage-host interactions in Y. enterocolitica, a significant enteric pathogen, and emphasize the need for investigating these dynamics under complex, physiologically relevant systems to inform better phage therapy strategies against mucosal bacterial infections.

microbiology↗

bioRxiv2025

ssDNA phage FLiP resides in dsDNA form in resistant Flavobacterium host

Most of the environmental flavobacteria decompose organic matter, playing a crucial role in ecosystem balance. Phages infecting these bacteria regulate the host populations and thereby the ecosystem functions, however, bacterial resistance against phage may cause changes in bacterial phenotypic characteristics. ssDNA phage Finnlakevirus FLiP infects three known environmental Flavobacterium sp. isolates: B330, B167, and B114. Building on our previous FLiP-host interaction studies, we aimed to broaden our understanding of the host perspective by exploring phage resistance mechanisms against FLiP and similar phages. We compared growth dynamics of ancestral and resistant variants to detect the effect of resistance on host fitness. Genomic comparison between the variants was used to identify resistance-related mutations. Additionally, we analysed genomic differences among the three host strains, screening for anti-phage systems. Sequence analyses, PCR, and nuclease treatments of resistant host genomes and plasmids were used to assess the possibility of lysogeny and superinfection immunity. No single mutation or anti-phage system consistently explained FLiP resistance, as these varied across hosts. Resistant B114 contained FLiP genome in circular extrachromosomal dsDNA form, suggesting possible lysogeny. Surprisingly, we found that low quantities of FLiP sequences exist in bacterial populations not exposed to FLiP in the laboratory. This suggests that residing as extrachromosomal dsDNA elements in a small percentage of host cells may be a natural strategy for FLiP type phages to endure unfavourable conditions.

microbiology↗

bioRxiv2025

New Finnlakevirus isolate FLiP-2 provides insight into the ecology of ssDNA phages in Flavobacterium hosts

Life cycle details or ecological impact are well characterized only for a few ssDNA phages. The Finnlakeviridae family includes one species, Finnlakevirus FLiP. Here, using the same Flavobacterium host and sampling location, we isolated a new strain designated FLiP-2, with 96,7 % genetic identity to the original isolate FLiP. To understand the ecology of this Flavobacterium-infecting phage species, we explored the host interactions of the two FLiP strains and a dsDNA Flavobacterium phage MaF61 under various conditions representing those encountered in their natural habitats in boreal lakes including different temperatures, anoxic conditions, and in the presence of different nutrients and antibiotics. While FLiP and FLiP-2 had similar virion stability outside the host, they exhibited significant differences in plaque morphology and infectivity. FLiP-2 could not replicate in the presence of ampicillin, whereas FLiP thrived even at high concentrations. Both strains of Finnlakevirus FLiP propagated better under or after stress exposure compared to MaF61. Additionally, Finnlakevirus FLiP plaques appeared far from the original infection site, particularly in response to stress, suggesting latent presence within a motile or filamentous bacterium. In conclusion, Finnlakevirus FLiP showed remarkable flexibility in host-interactions being well adapted to fluctuating conditions in boreal freshwaters.

microbiology↗

bioRxiv2025

ESBL plasmid compatibility with the surrounding microbial community influences ESBL gene survival under CRISPR-antimicrobial targeting

Bacteria often acquire resistance against antibiotics through the transfer of conjugative resistance plasmids. Hence, it is vital to develop strategies to mitigate the dispersal of antimicrobial resistance (AMR). CRISPR-based antimicrobial tools offer a sequence-specific solution to diminish and restrict the dissemination of antimicrobial resistance genes among bacteria. CRICON (CRISPR via conjugation) is an antimicrobial CRISPR tool that has been shown to efficiently reduce multi-resistance when targeting ESBL (Extended Spectrum Beta-Lactamase) harboring plasmids. However, conjugatively delivered genetic elements may be subjected to bacterial defense, lead to resistance development, and revert the efficiency of the CRISPR tools. Here, we studied the evolutionary consequences of four ESBL-harboring Escherichia coli strains targeted by CRICON in a 10-day multispecies microcosm experiment. We show that CRICON reduces the ESBL prevalence within the bacterial community, while the final ESBL persistence depends on the initial community composition. We observed an unexpected survival strategy of an ESBL-plasmid by escaping into a more competitive host species. Further, we show the development of partial resistance against the CRISPR-antimicrobials during the experiment. Our results underline the importance of the ecological and evolutionary factors in multispecies bacterial communities, as they may disrupt the effective use of CRISPR-based antimicrobial strategies via undesired outcomes of targeted therapies against plasmid-bearing multi-resistant bacteria.

microbiology↗

bioRxiv2025

Longitudinal dynamics of gut plasmidome and antibiotic resistance during antibiotic therapy: a case report

The human gut microbiome is composed of diverse microbes, and its association with human health is well-recognized. Antibiotic therapies for treating infectious diseases often mediate adverse influences on gut microbial ecosystems. Further, ESBL (extended-spectrum beta-lactamase)-producing bacteria potentially residing in the gut benefit from antibiotic-induced environmental changes via positive selection for resistance traits. As antimicrobial resistance (AMR) genes are often harbored by mobile plasmids, the importance of these extrachromosomal mobile genetic elements during antibiotic exposure is evident. However, there is still a knowledge gap in how microbiomes respond to antibiotic treatment especially in terms of plasmid carriage and how gut plasmid populations evolve following and revive after antibiotic therapy. To address these questions, in this case report we investigated the changes in plasmid population in the gut microbiome of a single ESBL-carrying patient during antibiotic therapy for uncomplicated acute appendicitis. Employing longitudinal sampling, we collected E. coli strains and performed metagenomic analysis before, during, and after the treatment. Our findings indicate that the antibiotic treatment is associated with a transient alteration in the microbial composition, AMR profile, and plasmid population. An extensive but temporary domination of ESBL-E. coli within the gut microbiome was observed parallel to the ongoing antibiotic treatment. However, this was not sustained in the follow-up period, indicating a slight restoration of both the microbial composition and the plasmid population. The research underscores the temporary impact of antibiotic therapy on the dynamics of the gut plasmidome which essentially mediate the spread of AMR within the gut microbiome.

microbiology↗

bioRxiv2022

Single-cell resolution genetic association analysis of heterogeneous bacterial communities by utilizing droplet digital PCR

Microbial communities often respond to environmental challenges, such as the presence of antibiotics, as a whole. Dissecting these community-level effects into separate acting entities requires the identification of organisms that carry functional genes for the observed feature. However, unculturable microbes are abundant in various environments, hence making the identification challenging. Moreover, while at present the development and application of single-cell tools for eukaryotic cells are enhancing, the comparable methodologies applicable for prokaryotic cells are still scarce and have not gained broad and solid status as tools for investigating microbial populations. Here, we present a cultivation-free technique that can be utilized to link functional genes with the carrying bacterial species at single-cell resolution. The developed protocol is relatively simple to use, utilizes commercially available droplet microfluidics devices, does not require toxic reagents, and eliminates invalid signals emerging from extracellular DNA. We validate the methodology by studying the conjugative transfer of antibiotic resistance plasmids in an environment challenged by antibiotics. Furthermore, the method can be customized for any given genetic trait to accurately identify its hosting subpopulation from a heterogeneous and potentially uncultivable bacterial community. ImportanceBacterial systems usually contain numerous different species that may harbor highly similar or identical genes that confer same phenotypic qualities for the community. To decipher the functions of these systems, we report the development of a novel methodology that enables investigating microbial communities at single-cell level. This user-friendly method utilizes droplet digital PCR (ddPCR) to find and identify carriers of specific genes potentially from various microbial sample types. By pinpointing gene carriers, such as those responsible for antibiotic resistance, this method can provide insights to the behavior of microbial communities and gene transfer therein. By strategically combining the use of common methods (ddPCR and amplicon sequencing), the workflow is highly accessible. Thus, it allows also the researchers without a background in single-cell techniques or access to special equipment to adopt the method for producing single-cell data to serve their own research, enabling new research avenues in microbial genetics and ecology.

microbiology↗

bioRxiv2021

Multi-plasmid clash in a bacterial community: plasmid viability depends on the ecological setting of hosts

Plasmids are genetic elements that disperse horizontally between different strains and species of bacteria and a major factor in the dissemination of virulence factors and antibiotic resistance. Understanding the ecology of plasmids has a notable anthropocentric value and therefore the interactions between bacterial hosts and individual plasmids have been studied in detail. However, bacterial systems often carry multiple genetically distinct plasmids, but dynamics of these multiplasmid "clashes" has remained unstudied. Here, we set to investigate the survival of 11 mobilizable or conjugative plasmids in five different ecological settings. The key incentive was to determine whether plasmid dynamics are reproducible and whether there are trade-offs in plasmid fitness that stem from the ecological situation of their initial hosts. Growth rates and maximum population densities increased in all communities and treatments over the 42-day evolution experiment although plasmid contents at the end varied notably. We show that large multiresistance conferring plasmids are unfit when the community also contains smaller plasmids with fewer resistance genes. This suggests that restraining the use to few antibiotics can make bacterial communities sensitive to others. The hosts also appear to react to the presence of multiple genetically different plasmids by enhancing fimbriae production instead of alleviating costs of individual plasmids. In general, the survivors of the here-studied multi-plasmid clash are significantly affected by the presence or absence of antibiotic selection and plasmid-free hosts of varying fitness. Therefore, these trade-offs in different settings can explain for example why some resistance plasmids have an advantage during a rapid proliferation of antibiotic sensitive pathogen whereas others dominate in alternative situations.

microbiology↗