Biomedical subjects

John P Dekker

Publications and source records attributed to John P Dekker.

7 recordsLinked to original sources

PubMed2025

Genomic determinants of antibiotic resistance for Helicobacter pylori treatment: a retrospective phenotypic and genotypic observational study.

BACKGROUND: Rising antimicrobial resistance of Helicobacter pylori is a public health challenge. Genomic-based susceptibility testing allows for the identification of resistance-associated mutations, complementing conventional diagnostics and advancing towards pathogen-based personalised therapies. Our study aimed to identify genes and mutations involved in antimicrobial resistance in H pylori and evaluate the extent to which these markers can be used as predictors of phenotypic resistance against clarithromycin and levofloxacin. METHODS: In this retrospective phenotypic and genotypic observational study, we included 1011 H pylori whole-genome sequences and strains of known geographical origin from the H pylori Genome Project (HpGP) collection. We performed phenotypic clarithromycin and levofloxacin susceptibility testing on a subset of 419 HpGP strains using Etest at a centralised laboratory. A genomic analysis was conducted to identify 23S rRNA and gyrA variants and build a curated catalogue of mutations associated with resistance to clarithromycin (ie, 23S rRNA 2142A→G, 2142A→C, and 2143A→G) and levofloxacin (ie, gyrA A88V or A88P, N87K or N87I, and D91G, D91N, or D91Y). Genotype-phenotype concordance was assessed to estimate sensitivity and specificity, and the curated catalogue of resistance-associated mutations was applied to the complete HpGP set. Region-specific prevalence of resistance-associated mutations was calculated for a combined dataset including the HpGP genomes and 768 whole-genome sequences retrieved from the US National Center for Biotechnology Information Sequence Read Archive repository. Associations between resistance genotypes, H pylori subpopulations, and minimum inhibitory concentrations (MICs) were tested. FINDINGS: Clarithromycin-resistant and levofloxacin-resistant HpGP strains were estimated with a sensitivity and specificity of 100%, with all confidence intervals ranging from 96% to 100%. The combined analysis (n=1779) found the highest prevalence of clarithromycin resistance in the western Pacific region (173 [51·2%] of 338 in southeast Asia and 75 [29·8%] of 252 in eastern Asia), north African region (seven [38·9%] of 18), and western Asian region (12 [31·6%] of 38), whereas the highest prevalence of levofloxacin resistance was found in south Asia (14 [51·85%] of 27), Central America (48 [38·7%] of 124), eastern Europe (four [36·4%] of 11), and southern Africa (three [33·3%] of nine). Similarly, 23S rRNA and gyrA genotypes are variable across H pylori subpopulations. MIC values changed depending on the specific mutation in 23S rRNA (mean clarithromycin MIC 24·61 mg/L [95% CI 12·27-36·96] for 2143A→G and 142·25 mg/L [95% CI 77·88-206·61] for 2142A→G) and gyrA (mean levofloxacin MIC 9·66 mg/L [95% CI 6·75-12·56] for mutations on codon 91, and 27·97 mg/L [95% CI 25·82-30·11] for mutations on codon 87). INTERPRETATION: Mutations in specific genes are reliable indicators to clarithromycin and levofloxacin resistance in H pylori, making them useful markers for the development of diagnostic assays and molecular monitoring. Our results suggest that using clarithromycin and levofloxacin empirically, without previous susceptibility testing, is unsuitable in all geographical regions covered by this study. FUNDING: Intramural Research Program of the US National Cancer Institute, the European Research Council, and the Spanish Ministry of Science and Innovation.

Helicobacter pylori↗

PubMed2025

Global diversity of integrating conjugative elements (ICEs) in Helicobacter pylori and their influence on genome architecture.

Integrating conjugative elements (ICEs) are mobile genetic elements conferring a wide range of beneficial functions upon their bacterial hosts. Generally, they can be activated from their integrated states to undergo horizontal gene transfer via conjugation. In the case of the human gastric pathogen Helicobacter pylori, a paradigm for extensive genetic diversity, highly efficient natural transformation and recombination processes may superimpose canonical transfer of its two ICEs termed ICEHptfs3 and ICEHptfs4, and thus shape their composition substantially. Here, as a part of the Helicobacter pylori Genome Project (HpGP) initiative, we have analyzed high-quality genome sequences from 1011 clinical strains with respect to their ICE content and variability. We show that both elements are highly prevalent in all H. pylori populations, but have a strong tendency for gene erosion. ICE sequence variations reflect the population structure and show a clear signature of increased horizontal transfer. A detailed map of ICE integration sites revealed local preferences, but also how recombination processes result in hybrid elements or genome rearrangements. Population-specific differences in ICE cargo genes might reflect distinct requirements in the biological functions provided by these mobile elements.

Journal Article↗

PubMed2006

Cooperative gating between single HCN pacemaker channels.

HCN pacemaker channels (I(f), I(q), or I(h)) play a fundamental role in the physiology of many excitable cell types, including cardiac myocytes and central neurons. While cloned HCN channels have been studied extensively in macroscopic patch clamp experiments, their extremely small conductance has precluded single channel analysis to date. Nevertheless, there remain fundamental questions about HCN gating that can be resolved only at the single channel level. Here we present the first detailed single channel study of cloned mammalian HCN2. Excised patch clamp recordings revealed discrete hyperpolarization-activated, cAMP-sensitive channel openings with amplitudes of 150-230 fA in the activation voltage range. The average conductance of these openings was approximately 1.5 pS at -120 mV in symmetrical 160 mM K(+). Some traces with multiple channels showed unusual gating behavior, characterized by a variable long delay after a voltage step followed by runs of openings. Noise analysis on macroscopic currents revealed fluctuations whose magnitudes were systematically larger than predicted from the actual single channel current size, consistent with cooperativity between single HCN channels.

Cell Line↗

PubMed2006

Microneedle-based intradermal delivery of the anthrax recombinant protective antigen vaccine.

The recombinant protective antigen (rPA) of Bacillus anthracis is a promising anthrax vaccine. We compared serum immunoglobulin G levels and toxin-neutralizing antibody titers in rabbits following delivery of various doses of vaccine by microneedle-based intradermal (i.d.) delivery or intramuscular (i.m.) injection using conventional needles. Intradermal delivery required less antigen to induce levels of antibody similar to those produced via i.m. injection during the first 2 weeks following primary and booster inoculation. This dose-sparing effect was less evident at the later stages of the immune response. Rabbits immunized i.d. with 10 mug of rPA displayed 100% protection from aerosol spore challenge, while i.m. injection of the same dose provided slightly lower protection (71%). Groups immunized with lower antigen doses were partially protected (13 to 29%) regardless of the mode of administration. Overall, our results suggest rPA formulated with aluminum adjuvant and administered to the skin by a microneedle-based device is as efficacious as i.m. vaccination.

Administration, Cutaneous↗

PubMed2004

Protective immunization against inhalational anthrax: a comparison of minimally invasive delivery platforms.

A new anthrax vaccine under clinical investigation is based on recombinant Bacillus anthracis protective antigen (rPA). Here, we investigated microneedle-based cutaneous and nasal mucosal delivery of rPA in mice and rabbits. In mice, intradermal (id) delivery achieved up to 90% seroconversion after a single dose, compared with 20% after intramuscular (im) injection. Intranasal (inl) delivery of a liquid formulation required 3 doses to achieve responses that were comparable with those achieved via the id or im routes. In rabbits, id delivery provided complete protection against aerosol challenge with anthrax spores; in addition, novel powder formulations administered inl provided complete protection, whereas a liquid formulation provided only partial protection. These results demonstrate, for the first time, that cutaneous or nasal mucosal administration of rPA provides complete protection against inhalational anthrax in rabbits. The novel vaccine/device combinations described here have the potential to improve the efficacy of rPA and other biodefense vaccines.

Administration, Cutaneous↗

PubMed2004

Intracellular gate opening in Shaker K+ channels defined by high-affinity metal bridges.

Voltage-gated potassium channels such as Shaker help to control electrical signalling in neurons by regulating the passage of K+ across cell membranes. Ion flow is controlled by a voltage-dependent gate at the intracellular side of the pore, formed by the crossing of four alpha-helices--the inner-pore helices. The prevailing model of gating is based on a comparison of the crystal structures of two bacterial channels--KcsA in a closed state and MthK in an open state--and proposes a hinge motion at a conserved glycine that splays the inner-pore helices wide open. We show here that two types of intersubunit metal bridge, involving cysteines placed near the bundle crossing, can occur simultaneously in the open state. These bridges provide constraints on the open Shaker channel structure, and on the degree of movement upon opening. We conclude that, unlike predictions from the structure of MthK, the inner-pore helices of Shaker probably maintain the KcsA-like bundle-crossing motif in the open state, with a bend in this region at the conserved proline motif (Pro-X-Pro) not found in the bacterial channels. A narrower opening of the bundle crossing in Shaker K+ channels may help to explain why Shaker has an approximately tenfold lower conductance than its bacterial relatives.

Bacterial Proteins↗

PubMed2004

A perturbation-based method for calculating explicit likelihood of evolutionary co-variance in multiple sequence alignments.

MOTIVATION: The constituent amino acids of a protein work together to define its structure and to facilitate its function. Their interdependence should be apparent in the evolutionary record of each protein family: positions in the sequence of a protein family that are intimately associated in space or in function should co-vary in evolution. A recent approach by Ranganathan and colleagues proposes to look at subsets of a protein family, selected for their sequence at one position, to see how this affects variation at other positions. RESULTS: We present a quantitative algorithm for assessing covariation with this approach, based on explicit likelihood calculations. By applying our algorithm to 138 Pfam families with at least one member of known structure, we demonstrate that our method has improved power in finding physically close residues in crystal structures, compared to that of Ranganathan and colleagues. SUPPLEMENTARY INFORMATION: www.afodor.net/bioinfosup.html

Algorithms↗