Red squirrels in the British Isles are infected with leprosy bacilli

Leprosy, caused by infection with Mycobacterium leprae or the recently discovered Mycobacterium lepromatosis, was once endemic in humans in the British Isles. Red squirrels in Great Britain (Sciurus vulgaris) have increasingly been observed with leprosy-like lesions on the head and limbs. Using genomics, histopathology, and serology, we found M. lepromatosis in squirrels from England, Ireland, and Scotland, and M. leprae in squirrels from Brownsea Island, England. Infection was detected in overtly diseased and seemingly healthy animals. Phylogenetic comparisons of British and Irish M. lepromatosis with two Mexican strains from humans show that they diverged from a common ancestor around 27,000 years ago, whereas the M. leprae strain is closest to one that circulated in Medieval England. Red squirrels are thus a reservoir for leprosy in the British Isles

Implementation considerations for a point-of-care Neisseria gonorrhoeae rapid diagnostic test at primary healthcare level in South Africa : a qualitative study

ADDITIONAL FILE 1 : IDI protocols. Interview protocols used in this study during the pre-implementation, initial use, 3-month (mid-assessment) and 6-month (post-evaluation) implementation phase.ADDITIONAL TABLE 1 : Consolidated criteria for reporting qualitative studies (COREQ): 32-item checklist for interviews and focus groups.ADDITIONAL TABLE 2 : NPT Constructs as defined for the implementation of the NG-LFA.BACKGROUND : South Africa maintains an integrated health system where syndromic management of sexually transmitted infections (STI) is the standard of care. An estimated 2 million cases of Neisseria gonorrhoeae (N. gonorrhoeae) occur in South Africa every year. Point-of-care diagnostic tests (POCT) may address existing STI control limitations such as overtreatment and missed cases. Subsequently, a rapid lateral flow assay with fluorescence-based detection (NG-LFA) with a prototype reader was developed for N. gonorrhoeae detection showing excellent performance and high usability; however, a better understanding is needed for device implementation and integration into clinics. METHODS : A qualitative, time-series assessment using 66 in-depth interviews was conducted among 25 trained healthcare workers involved in the implementation of the NG-LFA. Findings were informed by the Normalization Process Theory (NPT) as per relevant contextual (strategic intentions, adaptive execution, and negotiation capacity) and procedural constructs (coherence, cognitive participation, collective action, reflexive monitoring) to examine device implementation within primary healthcare levels. Interviews were audio-recorded, transcribed, and then analyzed using a thematic approach guided by NPT to interpret results. RESULTS : Overall, healthcare workers agreed that STI POCT could guide better STI clinical decision-making, with consideration for clinic integration such as space constraints, patient flow, and workload. Perceived NG-LFA benefits included enhanced patient receptivity and STI knowledge. Further, healthcare workers reflected on the suitability of the NG-LFA given current limitations with integrated primary care. Recommendations included sufficient STI education, and appropriate departments for first points of entry for STI screening. CONCLUSIONS : The collective action and participation by healthcare workers in the implementation of the NG-LFA revealed adaptive execution within the current facility environment including team compositions, facility-staff receptivity, and STI management experiences. User experiences support future clinic service integration, highlighting the importance of further assessing patient-provider communication for STI care, organizational readiness, and identification of relevant departments for STI screening.Sub-award via FIND (primary recipient Cecilia Ferreyra), from the Global Antimicrobial Resistance Innovation Fund (GAMRIF) (https://www.gov.uk/government/groups/theglobal-amr-innovation-fund).https://bmchealthservres.biomedcentral.comhj2024Medical MicrobiologySDG-03:Good heatlh and well-bein

Association of mutation patterns in gyrA/B genes and ofloxacin resistance levels in Mycobacterium tuberculosis isolates from East China in 2009

<p>Abstract</p> <p>Background</p> <p>This study aimed to analyze the association of mutation patterns in <it>gyrA </it>and <it>gyrB </it>genes and the ofloxacin resistance levels in clinical <it>Mycobacterium tuberculosis </it>isolates sampled in 2009 from East China.</p> <p>Methods</p> <p>The quinolone resistance-determining region of <it>gyrA/B </it>were sequenced in 192 <it>M. tuberculosis </it>clinical isolates and the minimal inhibitory concentrations (MICs) of 95 ofloxacin-resistant <it>M. tuberculosis </it>isolates were determined by using microplate nitrate reductase assays.</p> <p>Results</p> <p>Mutations in <it>gyrA </it>(codons 90, 91 and 94) and in <it>gyrB </it>(G551R, D500N, T539N, R485C/L) were observed in 89.5% (85/95) and 11.6% (11/95) of ofloxacin-resistant strains, respectively. The <it>gyrB </it>mutations G551R and G549D were observed in 4.1% (4/97) of ofloxacin-susceptible strains and no mutation was found in <it>gyrA </it>in ofloxacin-susceptible strains. The MICs of all ofloxacin-resistant strains showed no significant difference among strains with mutations at codons 90, 91 or 94 in <it>gyrA </it>(F = 1.268, <it>p </it>= 0.287). No differences were detected among strains with different amino acid mutations in the quinolone resistance-determining region of <it>gyrA </it>(F = 1.877, <it>p </it>= 0.123). The difference in MICs between ofloxacin-resistant strains with mutations in <it>gyrA </it>only and ofloxacin-resistant strains with mutations in both <it>gyrA </it>and <it>gyrB </it>genes was not statistically significant (F = 0.549, <it>p </it>= 0.461).</p> <p>Conclusions</p> <p>Although <it>gyrA/B </it>mutations can lead to ofloxacin resistance in <it>M. tuberculosis</it>, there were no associations of different mutation patterns in <it>gyrA/B </it>and the level of ofloxacin resistance in <it>M. tuberculosis </it>isolates from East China in 2009.</p

Structural Insights into the Quinolone Resistance Mechanism of Mycobacterium tuberculosis DNA Gyrase

Mycobacterium tuberculosis DNA gyrase, an indispensable nanomachine involved in the regulation of DNA topology, is the only type II topoisomerase present in this organism and is hence the sole target for quinolone action, a crucial drug active against multidrug-resistant tuberculosis. To understand at an atomic level the quinolone resistance mechanism, which emerges in extensively drug resistant tuberculosis, we performed combined functional, biophysical and structural studies of the two individual domains constituting the catalytic DNA gyrase reaction core, namely the Toprim and the breakage-reunion domains. This allowed us to produce a model of the catalytic reaction core in complex with DNA and a quinolone molecule, identifying original mechanistic properties of quinolone binding and clarifying the relationships between amino acid mutations and resistance phenotype of M. tuberculosis DNA gyrase. These results are compatible with our previous studies on quinolone resistance. Interestingly, the structure of the entire breakage-reunion domain revealed a new interaction, in which the Quinolone-Binding Pocket (QBP) is blocked by the N-terminal helix of a symmetry-related molecule. This interaction provides useful starting points for designing peptide based inhibitors that target DNA gyrase to prevent its binding to DNA

Etudes structurales et fonctionnelles du cœur catalytique de l'ADN gyrase de M. tuberculosis

Les topoisomérases de type II sont des enzymes qui régulent la topologie de l ADN dans la cellule pour permettre l exécution des processus informationnels. De manière générale, deux topoisomérases de type II aux actions synergiques coexistent chez les bactéries, l ADN gyrase et la topoisomérase IV. M. tuberculosis ne possède qu une seule topoisomérase de type II au sein de son génome, l ADN gyrase. qui doit donc accomplir seule, les fonctions nécessaires à la survie de la cellule généralement effectuées par deux topoisomérases de type II et possède, par conséquent, un spectre d activité hybride entre une ADN gyrase classique et une topoisomérase IV. Au cours de ce travail, nous avons résolu les structures cristallographiques de trois des quatre domaines de l ADN gyrase de M. tuberculosis, à savoir le domaine TOPRIM, le domaine de coupure-ligation et le domaine CTD, et complété les données structurales obtenues par des études biophysiques et fonctionnelles. Enfin, nous avons étudié le mécanisme d action des fluoroquinolones, antibiotiques utilisés dans le traitement de la tuberculose, et dont la cible unique chez M. tuberculosis est le cœur catalytique de l ADN gyrase. Ce dernier, composé du domaine de coupure-ligation et le domaine TOPRIM, a été modélisé en complexe avec l ADN et la moxifloxacine, qui est la fluoroquinolone la plus efficace contre M. tuberculosis. L analyse de ce modèle a permis de proposer un modèle d interaction entre les fluoroquinolones, l ADN et l ADN gyrase mettant en lumière l importance de la nature des acides aminés bordant la poche d interaction aux quinolones, souvent responsable de résistance à ces antibiotiquesPARIS-BIUSJ-Physique recherche (751052113) / SudocSudocFranceF

High resolution CryoEM structure of the ring-shaped virulence factor EspB from Mycobacterium tuberculosis

The EspB protein of Mycobacterium tuberculosis is a 60 kDa virulence factor, implicated in conjugation and exported by the ESX-1 system of which it may also be a component. Previous attempts to obtain high-resolution maps of EspB by cryo-electron microscopic examination of single particles have been thwarted by severe orientation bias of the particles. This was overcome by using detergent as a surfactant thereby allowing reconstruction of the EspB structure at 3.37 Å resolution. The final structure revealed the N-terminal domain of EspB to be organized as a cylindrical heptamer with dimensions of 90 Å x 90 Å and a central channel of 45 Å diameter whereas the C-terminal domain was unstructured. New atomic insight was obtained into the helical packing required for protomer interactions and the overall electrostatic potential. The external surface is electronegatively charged while the channel is lined with electropositive patches. EspB thus has many features of a pore-like transport protein that might allow the passage of an ESX-1 substrate such as the 35 Å diameter EsxA-EsxB heterodimer or B-form DNA consistent with its proposed role in DNA uptake

The Exchangeable Apolipoprotein Nplp2 Sustains Lipid Flow and Heat Acclimation in Drosophila

Summary: In ectotherms, increased ambient temperature requires the organism to consume substantial amounts of energy to sustain a higher metabolic rate, prevent cellular damage, and respond to heat stress. Here, we identify a heat-inducible apolipoprotein required for thermal acclimation in Drosophila. Neuropeptide-like precursor 2 (Nplp2) is an abundant hemolymphatic protein thought to be a neuropeptide. In contrast, we show that Nplp2 contributes to lipid transport, functioning as an exchangeable apolipoprotein. More precisely, Nplp2-deficient flies accumulate lipids in their gut, have reduced fat stores, and display a dyslipoproteinemia, showing that Nplp2 is required for dietary lipid assimilation. Importantly, Nplp2 is induced upon thermal stress and contributes to survival upon heat stress. We propose that Nplp2 associates with lipoprotein particles under homeostatic and high energy-demand conditions to optimize fat transport and storage. Our study also shows that modulation of the lipid uptake and transport machinery is part of an integrated cytoprotective response. : Rommelaere et al. identify a small exchangeable apolipoprotein in Drosophila that facilitates dietary lipid extraction and fat storage. Nplp2 sustains lipid flow to fuel the heat stress response, promoting insect survival at elevated temperatures. Optimization of lipid transport during stress via dedicated lipoproteins likely applies to other species including vertebrates. Keywords: exchangeable lipoprotein, neuropeptide, lipid transport, fat storage, heat acclimatio

Mycobacterium tuberculosis DNA gyrase possesses two functional GyrA-boxes.

International audienceIn contrast with most bacteria which possess two type II topoisomerases (topoisomerase IV and DNA gyrase), Mycobacterium tuberculosis possesses only one, DNA gyrase, which is functionally a hybrid enzyme. Functional differences between the two type IIA topoisomerases are thought to be specified by a CTD (C-terminal DNA-binding domain), which controls DNA recognition. To explore the molecular mechanism responsible for the hybrid functions of the M. tuberculosis DNA gyrase, we conducted a series of sequence analyses and structural and biochemical experiments with the isolated GyrA CTD and the holoenzyme. Although the CTD displayed a global structure similar to that of bona fide GyrA and ParC paralogues, it harbours a second key motif similar in all respects to that of the conserved GyrA-box sequence motif. Biochemical assays showed that the GyrA-box is responsible for DNA supercoiling, whereas the second GyrA-box-l (GyrA-box-like motif) is responsible for the enhanced decatenation activity, suggesting that the mechanistic originality of M. tuberculosis DNA gyrase depends largely on the particular DNA path around the CTD allowed for by the presence of GyrA-box-l. The results of the present study also provide, through phylogenetic exploration of the entire Corynebacterineae suborder, a new and broader insight into the functional diversity of bacterial type IIA topoisomerases

Implementation considerations for a point-of-care Neisseria gonorrhoeae rapid diagnostic test at primary healthcare level in South Africa: a qualitative study.

BACKGROUND South Africa maintains an integrated health system where syndromic management of sexually transmitted infections (STI) is the standard of care. An estimated 2 million cases of Neisseria gonorrhoeae (N. gonorrhoeae) occur in South Africa every year. Point-of-care diagnostic tests (POCT) may address existing STI control limitations such as overtreatment and missed cases. Subsequently, a rapid lateral flow assay with fluorescence-based detection (NG-LFA) with a prototype reader was developed for N. gonorrhoeae detection showing excellent performance and high usability; however, a better understanding is needed for device implementation and integration into clinics. METHODS A qualitative, time-series assessment using 66 in-depth interviews was conducted among 25 trained healthcare workers involved in the implementation of the NG-LFA. Findings were informed by the Normalization Process Theory (NPT) as per relevant contextual (strategic intentions, adaptive execution, and negotiation capacity) and procedural constructs (coherence, cognitive participation, collective action, reflexive monitoring) to examine device implementation within primary healthcare levels. Interviews were audio-recorded, transcribed, and then analyzed using a thematic approach guided by NPT to interpret results. RESULTS Overall, healthcare workers agreed that STI POCT could guide better STI clinical decision-making, with consideration for clinic integration such as space constraints, patient flow, and workload. Perceived NG-LFA benefits included enhanced patient receptivity and STI knowledge. Further, healthcare workers reflected on the suitability of the NG-LFA given current limitations with integrated primary care. Recommendations included sufficient STI education, and appropriate departments for first points of entry for STI screening. CONCLUSIONS The collective action and participation by healthcare workers in the implementation of the NG-LFA revealed adaptive execution within the current facility environment including team compositions, facility-staff receptivity, and STI management experiences. User experiences support future clinic service integration, highlighting the importance of further assessing patient-provider communication for STI care, organizational readiness, and identification of relevant departments for STI screening