Biology subjects

Withey, J. H.

Publications and source records attributed to Withey, J. H..

6 recordsLinked to original sources

bioRxiv2026

Effect of Temperature on Gene Expression of Escherichia marmotae

Increased temperature is one of the first environmental cues encountered by bacteria upon entering a mammalian host. Here, we investigated the effects of temperature on the transcriptome and proteome of Escherichia marmotae and E. coli. Previous studies demonstrated that temperature affects motility in E. marmotae; therefore, we examined how temperature alters gene expression at 37 {degrees}C versus 28 {degrees}C and whether this response is conserved in E. coli. Strains were grown under static conditions at both temperatures, and gene expression and protein abundance were assessed by RNA transcriptome analysis and global proteomics. Temperature altered the expression of 111 genes (2.7%) in E. marmotae and 99 genes (2.5%) in E. coli (adjusted p < 0.05, [≥]2-fold change), with changes concentrated within specific functional pathways. In E. marmotae, flagellar and chemotaxis genes and operons involved in cellulose-dependent biofilm formation and nitrate respiration were markedly downregulated at 37 {degrees}C. In contrast, genes associated with fimbrial adhesion and immune evasion, including fimA/fimB, ompT, and prophage-associated loci, were upregulated. Proteomic analysis corroborated these trends, showing reduced flagellar and chemotaxis proteins and increased stress-adaptation and host-interaction proteins. E. coli showed a distinct response, with stronger enrichment of metabolic and amino-acid biosynthesis pathways and minimal changes in motility regulation. Together, these findings demonstrate that E. marmotae motility is temperature-dependent and may represent a mechanism for immune evasion within the host.

genomics↗

bioRxiv2025

Exposure to third generation cephalosporin induces L-form transition in Shigella sonnei, potentially acting as a bacterial reservoir for persistent infection

Shigellosis remains a major global health burden, and the increasing prevalence of multidrug-resistant (MDR) Shigella strains is complicating effective antibiotic therapy. Bacteria may survive antibiotics by transitioning into cell wall-deficient L-forms, which are intrinsically resistant to {beta}-lactams and can revert to a virulent state, potentially causing relapsing infections. Here we characterized a clinical MDR isolate Shigella sonnei HK8, a.k.a. PD552A, whose genome contains key resistance (gyrA, PBP3) and virulence (icsA) genes. Exposure to ceftriaxone induced a transition into a viable L-form state that was hyper-adhesive to macrophages in vitro. However, this survival adaptation was linked to a profound loss of pathogenicity. Using murine and guinea pig models, the L-form variant was shown to be profoundly attenuated, failing to cause the keratoconjunctivitis, diarrheal disease, or significant histopathology characteristic of the wild-type strain. These findings reveal a critical virulence-survival trade-off, positioning the L-form as a "stealth" phenotype that enables bacterial persistence at the expense of acute virulence. This offers a potential mechanism for asymptomatic carriage and recurrent infections, highlighting a previously underappreciated mechanism by which antibiotic treatment may resolve acute symptoms while permitting the persistence of a cryptic bacterial reservoir capable of driving recurrent infection. ImportanceOur research provides critical insight into the challenge of antibiotic treatment failure in shigellosis. By integrating experimental validation of reversible L-form transitions with mathematical modelling, we uncover a crucial virulence-survival trade-off. We show that MDR S. sonnei survives ceftriaxone by adopting a "stealth" phenotype, quantified by a high Stealth Index - maintaining bacterial burden while evading host inflammatory detection. These findings imply that standard antibiotics may resolve acute symptoms while inadvertently selecting for a cryptic, persistent reservoir poised for relapse. This work challenges the conventional view of therapeutic success and highlights an urgent need to develop novel diagnostic and therapeutic strategies capable of identifying and eliminating these resilient, "stealth" L-form persisters to achieve true bacterial clearance and prevent chronic infections.

microbiology↗

bioRxiv2025

In vivo efficacy of L-ascorbic acid in restricting cholera pathogenesis

In the last three centuries, our world has experienced seven cholera pandemics. Cholera is a deadly diarrheal disease caused by Vibrio cholerae, an important member of the gamma-proteobacteria. Despite conventional treatments like antibiotics, vaccines, and ORS, the 7th cholera pandemic is still a major threat to developing nations. L-ascorbic acid was recently shown to effectively kill the Vibrio cholerae cells in vitro under various growth conditions mimicking the in vivo host conditions, including growth in the presence of bile salts, growth of acid-adapted V. cholerae, and growth in the presence of various ORS components. In the current study, we extend this work and test the efficacy of L-ascorbic acid in animal models (rabbit ileal loop and the removable intestinal tie adult rabbit diarrhea). We show that L-ascorbic acid can effectively reduce the bacterial load in both the rabbit models as well as fast-track the recovery from the diarrheal symptoms. ImportanceCholera is a severe human diarrheal disease that affects millions each year. Cholera treatment is primarily oral rehydration to balance the voluminous fluid loss. Previous studies have found that L-ascorbic acid (L-AA) is effective at inhibiting growth of Vibrio cholerae, the causative agent of cholera. Thus L-AA has potential as am inexpensive cholera therapeutic. Here, L-AA is tested in rabbit models to determine whether it can inhibit the effects of V. cholerae infection. Results suggest that L-AA treatment inhibits bacterial proliferation and leads to shortened recovery from disease.

microbiology↗

bioRxiv2021

Transition of Vibrio cholerae through a natural host induces resistance to environmental changes

The pandemic-related strains of Vibrio cholerae are known to cause diarrheal disease in animal hosts. These bacteria must overcome rapid changes in their environment, such as the transition from fresh water to the gastrointestinal system of their host. To study the morphological adjustments during environmental transitions, we used zebrafish as a natural host. Using a combination of fluorescent light microscopy, cryogenic electron tomography and serial block face scanning electron microscopy, we studied the structural changes that occur during the infection cycle. We show that the transition from an artificial nutrient-rich environment to a nutrient-poor environment has a dramatic impact on the cell shape, most notably membrane dehiscence. In contrast, excreted bacteria from the host retain a uniform distance between the membranes as well as their vibrioid shape. Inside the intestine, V. cholerae cells predominantly colonized the anterior to mid-gut, forming micro-colonies associated with the microvilli as well as within the lumen. The cells retained their vibrioid shape but changed their cell-length depending on their localization. Our results demonstrate dynamic changes in morphological characteristics of V. cholerae during the transition between the different environments, and we propose that these structural changes are critical for the pathogens ability to colonize host tissues.

microbiology↗

bioRxiv2021

Neutrophil-associated responses to Vibrio cholerae infection in a natural host model

Vibrio cholerae, the cause of human cholera, is an aquatic bacterium found in association with a variety of animals in the environment, including many teleost fish species. V. cholerae infection induces a pro-inflammatory response followed by a non-inflammatory convalescent phase. Neutrophils are integral to this early immune response. However, the relationship between the neutrophil-associated protein calprotectin and V. cholerae has not been investigated, nor have the effects of limiting transition metals on V. cholerae growth. Zebrafish are useful as a natural V. cholerae model as the entire infectious cycle can be recapitulated in the presence of an intact intestinal microbiome and mature immune responses. Here, we demonstrate that zebrafish produce a significant neutrophil, IL-8, and calprotectin response following V. cholerae infection. Bacterial growth was completely inhibited by purified calprotectin protein or the chemical chelator TPEN, but growth was recovered by addition of transition metals zinc and manganese. Expression of downstream calprotectin targets also significantly increased in the zebrafish. These findings are the first to illuminate the role of calprotectin and nutritional immunity in combating V. cholerae infection. Inhibition of V. cholerae growth through metal limitation may provide new approaches in the development of anti-V. cholerae therapeutics. This study also establishes a major role for calprotectin in combating infectious diseases in zebrafish.

microbiology↗

bioRxiv2021

Elucidating the correlation between the number of TTTTGAT heptamer repeats and cholera toxin promoter activity in Vibrio cholerae O1 pandemic strains

A complex regulatory cascade controls expression of the cholera toxin genes (ctxAB) in Vibrio cholerae; which eventually leads to choleragen (CT) production and secretion, resulting in rice watery diarrhoea. The cholera toxin promoter (PctxAB) contains a series of heptad repeats (5-TTTTGAT-3); which have been previously shown to play crucial role in ctxAB transcriptional regulation by recruiting the transcriptional activators ToxT, ToxR, and the nucleoid-associated protein H-NS along the ctx promoter. The numbers of these repeats vary between the two biotypes of V. cholerae O1 strains, and even among strains of the same biotype. In this study, we examined PctxAB activation of V. cholerae O1 pandemic strains to understand the significance of the distal heptad repeats in regulating ctx expression. Interestingly, we found that ctx activation may depend on the number of TTTTGAT heptad repeats within PctxAB, and we posit that the occupation of the distal repeats by H-NS could further prevent transcriptional activation of ctx genes in V. cholerae. We hypothesize that ToxT-dependent transcriptional activation may not require entire displacement of H-NS and propose a revision in the currently accepted model of ToxT dependent PctxAB transcriptional activation. IMPORTANCECT production by pathogenic V. cholerae O1 strains is regulated through the transcriptional silencing of CTX promoter by H-NS and counter repression by ToxT. The highly AT rich PctxAB is composed of the tandem repeats 5{square} TTTTGAT 3{square}; the numbers of which differ among the classical and El Tor biotypes. However, it is still not very clear whether the numbers of these repeats could be correlated with promoter activation of the ctx operon. Here we report the role of the distal repeats in ctxAB expression levels. We demonstrate that PctxAB activation changes with the number of the heptad repats within the core promoter element, thereby suggesting a model for ctx regulation in the toxigenic strains of V. cholerae.

microbiology↗