Biology subjects

Salama, N. R.

Publications and source records attributed to Salama, N. R..

7 recordsLinked to original sources

bioRxiv2026

Helicobacter pylori allelic variation in cell surface genes influences human exoproteome binding and stomach tissue adherence

Helicobacter pylori, the primary etiological agent of gastric cancer, requires chronic infection to promote severe disease. Throughout colonization, the bacteria accumulate genetic variation, which can reshape the interaction between host and pathogen. In this study, we probed adherence as one important characteristic of this relationship. We leveraged a panel of H. pylori strains, representing both inter- and intra-host diversity, and BASEHIT, a comprehensive barcoded yeast display library of the human exoproteome, to evaluate the human exoprotein binding characteristics of H. pylori. We identified a set of lineage-correlated binding phenotypes and a set of polymorphic cell surface-associated loci that we predict to govern heterogeneous binding. We also identified a general increase in gastric tissue adherence during mouse passage and structural modifications in the O-antigen component lipopolysaccharide. Subsequent sequence analyses identified C-terminal repeat length reduction in either futB or glycosyltransferase family 25 galactose transferases as sufficient to alter lipopolysaccharide. Shorter repeat variants were favored during both acute and chronic colonization and conferred a colonization advantage in coinfection. Our data indicate that shorter enzymes, and the resulting shorter O-antigen repeats, are favored during infection. We hypothesize that repeat length polymorphisms modulate enzyme efficiency and disrupt the balance of competing reactions required for O-antigen synthesis. ImportanceIn this study, we used a high throughput assay to evaluate the adherence of genetically diverse Helicobacter pylori isolates to human proteins they may encounter in the stomach. We showed that more closely related H. pylori strains bind human proteins similarly. We linked changes in bacterial binding to genetic differences affecting cell surface components. In particular, H. pylori strains passaged through mice exhibited increased adherence to stomach tissue, which we associated with structural variation in the enzymes responsible for synthesizing the O-antigen portion of lipopolysaccharide. We also showed that shorter enzyme variants produce distinct O-antigen structures that enhance colonization of the mouse stomach.

microbiology↗

bioRxiv2022

Single-cell profiling uncovers a Muc4-expressing metaplastic gastric cell type sustained by Helicobacter pylori-specific inflammation

AbstractMechanisms for Helicobacter pylori (Hp)-driven stomach cancer are not fully understood. In a transgenic mouse model of gastric preneoplasia, concomitant Hp infection and induction of constitutively active KRAS (Hp+KRAS+) alters metaplasia phenotypes and elicits greater inflammation than either perturbation alone. Gastric single-cell RNA-seq showed that Hp+KRAS+ mice had a large population of metaplastic pit cells that expressed the intestinal mucin Muc4 and the growth factor amphiregulin. Metaplastic pit cells were associated with macrophage and T cell inflammation and prevented by gastric immunosuppression. Lineage tracing showed that Muc4 was not dependent on cell-intrinsic KRAS activity, and lineage-derived cells had a limited propensity for growth as organoids, demonstrating that metaplastic pit cells are largely not self- renewing. Finally, MUC4 expression was significantly associated with proliferation in human gastric cancer samples. These studies identify an Hp-associated metaplastic pit cell lineage, also found in human gastric cancer tissues, whose expansion is driven by Hp-dependent inflammation. Statement of SignificanceUsing a mouse model, we have delineated metaplastic pit cells as a pre-cancerous cell type whose expansion requires H. pylori-driven inflammation. In humans, metaplastic pit cells show enhanced proliferation as well as enrichment in precancer and early cancer tissues, highlighting an early step in the gastric metaplasia to cancer cascade.

microbiology↗

bioRxiv2022

Helicobacter pylori chronic infection selects for effective colonizers of metaplastic glands

Chronic gastric infection with Helicobacter pylori can lead to progressive tissue changes that culminate in cancer, but how H. pylori adapts to the changing tissue environment during disease development is not fully understood. In a transgenic mouse gastric metaplasia model, we found that strains from unrelated individuals differed in their ability to infect the stomach, to colonize metaplastic glands, and to induce proliferation and alter the expression of metaplasia-associated proteins. H. pylori isolates from different stages of disease from a single individual had differential ability to colonize healthy and metaplastic gastric glands. Exposure to the metaplastic environment selected for high gastric colonization by one of these strains. Complete genome sequencing revealed a unique alteration in the frequency of a variant allele of the putative adhesin sabB, arising from a recombination event with the related sialic acid binding adhesin (SabA) gene. Mutation of sabB strongly reduced adherence to both normal and metaplastic gastric tissue in multiple strain backgrounds and highly attenuated stomach colonization. Thus, the changing gastric environment during disease development promotes bacterial adhesin gene variation associated with enhanced gastric colonization. ImportanceChronic infection with Helicobacter pylori is the primary risk factor for developing stomach cancer. As disease progresses H. pylori must adapt to a changing host tissue environment that includes induction of new cell fates in the cells that line the stomach. We tested representative H. pylori isolates collected from the same patient during early and later stages of disease in a mouse model where we can rapidly induce disease-associated tissue changes. Only the later-stage H. pylori strains could robustly colonize the diseased stomach environment. We also found that the ability to colonize the diseased stomach was associated with genetic variation in a putative cell surface adhesin gene called sabB. Additional experiments revealed that SabB promotes binding to stomach tissue and is critical for stomach colonization by the late-stage strains. Thus, H. pylori diversifies its genome during disease progression and these genomic changes highlight critical factors for bacterial persistence.

microbiology↗

bioRxiv2020

Helicobacter pylori diversification during chronic infection within a single host generates sub-populations with distinct phenotypes

Helicobacter pylori chronically infects the stomach of approximately half of the worlds population. Manifestation of clinical diseases associated with H. pylori infection, including cancer, is driven by strain properties and host responses; and as chronic infection persists, both are subject to change. Previous studies have documented frequent and extensive within-host bacterial genetic variation. To define how within-host diversity contributes to phenotypes related to H. pylori pathogenesis, this project leverages a collection of 39 clinical isolates acquired prospectively from a single subject at two time points and from multiple gastric sites. During the six years separating collection of these isolates, this individual, initially harboring a duodenal ulcer, progressed to gastric atrophy and concomitant loss of acid secretion. Whole genome sequence analysis identified 2,232 unique single nucleotide polymorphisms (SNPs) across isolates and a nucleotide substitution rate of 1.3x10-4 substitutions/site/year. Gene ontology analysis identified cell envelope genes among the genes with excess accumulation of nonsynonymous SNPs (nSNPs). A dendrogram based on genetic similarity, clusters isolates from each time point separately. Within time points, there is segregation of subgroups with phenotypic differences in bacterial morphology, ability to induce inflammatory cytokines, and mouse colonization. Higher inflammatory cytokine induction in recent isolates maps to shared polymorphisms in the Cag PAI protein, CagY, while rod morphology in a subgroup of recent isolates mapped to eight mutations in three distinct helical cell shape determining (csd) genes. The presence of subgroups with unique genetic and phenotypic properties suggest complex selective forces and multiple sub-niches within the stomach during chronic infection. Author SummaryHelicobacter pylori, one of the most common bacterial pathogens colonizing humans, is the main agent responsible for stomach ulcers and cancer. Certain strain types are associated with increased risk of disease, however many factors contributing to disease outcome remain unknown. Prior work has documented genetic diversity among bacterial populations within single individuals, but the impact of this diversity for continued bacterial infection or disease progression remains understudied. In our analysis we examined both genetic and functional features of many stomach isolates from a single individual infected over six years. During these six years the subject shifted from having excess acid production and a duodenal ulcer to lower acid production from gastric atrophy. The 39 isolates form sub-populations based on gene sequence changes that accumulated in the different isolates. In addition to having distinguishing genetic features, these sub-populations also have differences in several bacterial properties, including cell shape, ability to activate immune responses, and colonization in a mouse model of infection. This apparent functional specialization suggests that the bacterial sub-populations may have adapted to distinct sub-niches within the stomach during chronic infection.

microbiology↗

bioRxiv2019

Distinct cytoskeletal proteins define zones of enhanced cell wall synthesis in Helicobacter pylori

Helical cell shape is necessary for efficient stomach colonization by Helicobacter pylori, but the molecular mechanisms for generating helical shape remain unclear. We show that the helical centerline pitch and radius of wild-type H. pylori cells dictate surface curvatures of considerably higher positive and negative Gaussian curvatures than those present in straight- or curved-rod bacteria. Quantitative 3D microscopy analysis of short pulses with either N-acetylmuramic acid or D-alanine metabolic probes showed that cell wall growth is enhanced at both sidewall curvature extremes. Immunofluorescence revealed MreB is most abundant at negative Gaussian curvature, while the bactofilin CcmA is most abundant at positive Gaussian curvature. Strains expressing CcmA variants with altered polymerization properties lose helical shape and associated positive Gaussian curvatures. We thus propose a model where CcmA and MreB promote PG synthesis at positive and negative Gaussian curvatures, respectively, and that this patterning is one mechanism necessary for maintaining helical shape.

microbiology↗

bioRxiv2018

Non-helical Helicobacter pylori show altered gland colonization and elicit less gastric pathology during chronic infection

Half of all humans harbor Helicobacter pylori in their stomachs. Helical cell shape is thought to facilitate H. pyloris ability to bore into the protective mucus layer in a corkscrew-like motion, thus enhancing colonization of the stomach. H. pylori cell shape mutants show impaired colonization of the mouse stomach, highlighting the importance of cell shape in infection. To gain a deeper understanding of how helical cell morphology promotes host colonization by H. pylori, we used 3D-confocal microscopy to visualize the clinical isolate PMSS1 and an isogenic straight rod mutant ({Delta}csd6) within thick longitudinal mouse stomach sections and performed volumetric image analysis to quantify the number of bacteria residing within corpus and antral glands in addition to measuring total colony forming units (CFU). We found that straight rods show attenuation during acute colonization of the stomach (one day or one week post-infection) as measured by total CFU. Our quantitative imaging revealed that wild-type bacteria extensively colonized antral glands at one week post-infection, while csd6 mutants showed variable colonization of the antrum at this timepoint. During chronic infection (one or three months post-infection), total CFU were highly variable, but similar for wild-type and straight rods. Both wild-type and straight rods persisted and expanded in corpus glands during chronic infection. However, the straight rods showed reduced inflammation and disease progression. Thus, helical cell shape contributes to tissue interactions that promote inflammation during chronic infection, in addition to facilitating niche acquisition during acute infection.

microbiology↗

bioRxiv2018

Increased H. pylori stool shedding and EPIYA-D cagA alleles are associated with gastric cancer in an East Asian hospital

BackgroundHelicobacter pylori infection induces chronic inflammation and tissue damage in the stomach, increasing risk for gastric cancer. Paradoxically, these tissue alterations may promote loss of H. pylori infection during cancer progression. H. pyloris role in cancer progression beyond initiation is unclear. Geographic variation in gastric cancer risk has been attributed to variation in carriage and type of the H. pylori oncogene cagA.\n\nMethodsTo investigate possible differences in H. pylori load in the stomach and shedding in stool, H. pylori load and cagA genotype were assessed using droplet digital PCR assays on gastric mucosa and stool samples from 49 urea breath test-positive individuals, including 25 gastric cancer and 24 non-cancer subjects at Henan Cancer Hospital, Henan, China.\n\nResultsQuantitation of H. pylori DNA indicated similar gastric loads among cancer and non-cancer cases, but the gastric cancer group had a median H. pylori load in the stool that was six times higher than that of the non-cancer subjects. While the cagA gene was uniformly present among study subjects, only 70% had the East Asian cagA allele, which was significantly associated with gastric cancer (Fishers Exact Test, p = 0.03).\n\nConclusionH. pylori persists in a subset of gastric cancer cases and thus may contribute to cancer progression. In this East Asian population with a high prevalence of the cagA gene, the East Asian allele could still provide a marker for gastric cancer risk.\n\nImpactThis study contributes to our understanding of H. pylori dynamics in the context of pathological changes.

microbiology↗