Biology subjects

Ibranosyan, M.

Publications and source records attributed to Ibranosyan, M..

4 recordsLinked to original sources

bioRxiv2026

Genetic diversity of Legionella species in culture-negative clinical and environmental specimens by sequencing the 23S-5S ribosomal intergenic spacer region

The diagnosis of Legionnaires' disease (LD) caused by Legionella non-pneumophila species is likely to increase with broader use of PCR targeting Legionella spp. In this context, accurate species identification in PCR-positive but culture-negative samples is essential to improve understanding of disease epidemiology and to support source attribution. Here, we presented a validated and user-friendly bioinformatic pipeline compatible with next-generation sequencing (NGS) for analyzing the hypervariable 23S-5S region, paired with a curated database encompassing all described Legionella species as of January 2026. Parameters were optimized for sensitivity and specificity using both strains and culture-positive clinical and environmental samples. We then applied the pipeline retrospectively to 92 culture-negative PCR-positive samples collected from 2023 to 2025. Legionella species were successfully assigned in 60% (55/92) of tested samples and revealed a high diversity. Co-infections were detected in clinical samples, including combinations of L. pneumophila with L. longbeachae or L. bozemanii, while environmental samples contained up to six different species. These results demonstrate that 23S-5S amplicon NGS enables species-level identification in the absence of cultured isolates, improving surveillance of non-pneumophila Legionella cases. The proposed pipeline, implemented in QIIME2 and accompanied by a publicly available database, provides a practical framework for routine molecular monitoring and outbreak investigation.

microbiology↗

bioRxiv2026

Importance of taking Single Amino Acid Variant and accessory proteome variability into account in Data Independent Acquisition Proteomics: illustrated with Legionella pneumophila analysis

Understanding allelic variability is crucial for elucidating intrinsic bacterial mechanisms and distinguishing phenotypic profiles. However, such variability poses a major challenge for the reliable identification of proteins in data-independent acquisition (DIA) proteomics. To address this, we developed an analytical workflow that integrates protein sequence variability to enhance proteome coverage. Fifteen Legionella pneumophila isolates were analyzed using DIA-NN, with spectral libraries generated either from a reference proteome or incorporating allelic variability. Our workflow includes protein clustering and subsequent protein inference from these clusters, allowing the accurate assignment of shared and variant-specific peptides. Integration of variability enabled the identification of a comparable number of proteins as the reference proteome while capturing between 28 and 77 % of variant-specific sequences in each isolate, all while maintaining a low false positive rate. These findings demonstrate that accounting for allelic variability substantially improves proteomic coverage and identification confidence, providing a more comprehensive view of the proteome. This approach facilitates a deeper understanding of biological mechanisms and enables precise bacterial proteotyping of Legionella pneumophila isolates.

bioinformatics↗

bioRxiv2026

Functional alterations of immune gene expression in ICU and non-ICU patients with Legionnaires' disease, a prospective observational study

Legionnaires disease (LD), a pneumonia caused by Legionella pneumophila intracellular bacterium, leads to intensive care unit (ICU) admission in 20-40% of cases. While these ICU-LD patients display severe lung injury or septic shock, their functional immune response remains poorly understood. The present study aimed, through a large immune gene expression assessment, to improve the understanding of immune cell functionality after whole blood LPS ex vivo stimulation in ICU-LD patients compared with non-ICU. Both ICU and non-ICU-LD displayed altered gene expression indicating that both patients immune cells are less able to respond to the LPS ex vivo stimulus than a healthy population. ICU-LD patients had 1.6-fold greater number of less-expressed genes (35/93 vs 22/93, p=0.039), and lower Log2(FC) of these genes (median [IQR]: -1.9 [-2.6;-1.5] vs -1.2 [-1.7;-0.9], p=0.0011) than non-ICU-LD. Seven genes were significantly less expressed by ICU-LD patients (IRF7, MX1, NFKBI2, NFKBIA, RELB, SRC, TIM3; p-value range: 0.029-0.0080). Top five gene ontology biological processes, subcellular localisations, and reactome pathways (STRING database) uniquely enriched in ICU-LD-patients and related less-expressed genes were cellular response to LPS (CCL2, NFKBIA, IRAK2, TIM3, SRC, NFKB1), regulation of IFN-{beta} production (IRF7, RIG1, OAS2, RELB), I-{kappa}B/NF-{kappa}B complex (NFKBIA, NFKB1, NFKB2), IFN regulatory factor complex (RIG1, IRF7), and TRAF6-mediated NF-{kappa}B activation pathway (NFKBIA, NFKB1, NFKB2, RIG1). Immune gene expression alterations in LD after LPS stimulation were found herein, with more pronounced alterations in ICU-LD patients. A reduced expression of key genes and pathways involved in controlling Legionella proliferation in ICU-LD patients may contribute to increased disease severity.

immunology↗

bioRxiv2021

Zebrafish larvae as a powerful model to dissect protective innate immunity in response to Legionella pneumophila infection

The zebrafish has become a powerful model organism to study host-pathogen interactions. Here, we developed a zebrafish model of Legionella pneumophila infection to dissect innate immune responses. We show that L. pneumophila cause zebrafish larvae death in a dose dependent manner, and that macrophages are the first line of defence, with neutrophils cooperating to clear the infection. When either macrophages or neutrophils are depleted, these "immunocompromised" larvae become lethally sensitive to L. pneumophila similar to what is known for humans that develop pneumonia. Also as observed in human infections, the adaptor signalling molecule Myd88 is not required to control disease in the larvae. Furthermore, proinflammatory cytokine genes il1{beta} and tnf were upregulated during infection, recapitulating key immune responses seen in human infection. Strikingly, we uncovered a previously undescribed infection phenotype in zebrafish larvae, whereby bloodborne, wild type L. pneumophila invade and grow in the larval yolk region, a phenotype not observed with a type IV secretion system deficient mutant that cannot translocate effectors into its host cell. Thus, zebrafish larva represents an innovative L. pneumophila infection model that that on one hand mimics important aspects of the human immune response to L. pneumophila infection and that on the other hand will allow to elucidate the mechanisms by which type IV secretion effectors allow L. pneumophila to cross membranes and to obtain nutrients from nutrient rich environments.

microbiology↗