Dataset Information

Changes in OMPs of Leptospira under simulated physiological conditions of infection

ABSTRACT: Leptospirosis is a worldwide zoonotic disease caused bypathogenic Leptospiraspread through urine of infected animals. There are many gaps in understanding the pathophysiology of leptospirosis and the host immunity.This lack of knowledge hinders taking necessary steps in the development of a potential vaccine, effective diagnostic tool, and potential biomarkers.The Outer Membrane Proteins (OMPs) of Leptospira are significant response regulators during the phase of invasive dissemination and their consecutive differential expression allows adjustment and survival advantage for the microorganism to the host environment which may increase its infectivity and cause disease. In the present study, we were trying to represent the dynamic change (both qualitatively & quantitatively) of differentially expressed proteins of Leptospirainterrogans and their localization on the OM under simulated pathophysiological conditions. We had challenged mid-log phase leptospires to 37 ºC, 300 mOsm, and 37 °C + 300 mOsm and standard culture conditions as a control in EMJH medium for 24 h. The leptospires were washed with PBS containing 5mM MgCl2 three times followed by extraction of the cellular proteins through the phase partition method with the help of Triton X-114 which resulted in three distinct cellular fractions including aqueous, detergent, and pellet. The proteins from the three distinct fractions concerning their respective treatments were tagged with Tandem Mass Tag (TMT) (Thermo Fisher Scientific) followed by LC-MS/MS analysis. The analysis showed a significant number of differentially expressed proteins in the three cellular fractions with respect to the standard culture condition. It also allowed us to analyze the differentially regulated proteins expressed on the outer membrane during the physiological condition of infection.

INSTRUMENT(S):

ORGANISM(S): Leptospira Interrogans Serovar Icterohaemorrhagiae

SUBMITTER: Madathiparambil G. Madanan

LAB HEAD: Dr Madathiparambil G. Madanan,

PROVIDER: PXD035782 | Pride | 2025-09-08

REPOSITORIES: Pride

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Publications

Exoproteome Profiling Reveals Increased Secretion of Adhesins and Proteases by to Facilitate Host Colonization and Immune Modulation.

Sarma Abhijit A Phukan Homen H Devasahayam Arokia Balaya Rex R Thoduvayil Sikha S Dhandapani Gunasekaran G Christie Swamidurai Arul Diana SAD Prasad Thottethodi Subrahmanya Keshava TSK Madanan Madathiparambil Gopalakrishnan MG

ACS omega 20250727 30

Leptospirosis, a re-emerging zoonotic disease, is challenging human and animal health due to the lack of early and rapid diagnostic tools and effective vaccines. The exoproteome of the pathogen expressed under pathogenic conditions possesses a rational diagnostic significance due to its consistent presence in body fluids. were challenged to conditions simulating infection using physiological temperatures and osmolarity. Using state-of-the-art extraction techniques, efficient enrichment of nonabu ...[more]

PMID: 40787326

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Project description:Leptospirosis, an emerging zoonotic disease with worldwide distribution, is caused by spirochetes belonging to the genus Leptospira. More than 500,000 cases of severe leptospirosis are reported annually, with .10% of these being fatal. Leptospires can survive for weeks in suitably moist conditions before encountering a new host. Reservoir hosts, typically rodents, exhibit little to no signs of disease but shed large numbers of organisms in their urine. Transmission occurs when mucosal surfaces or abraded skin come into contact with infected urine or urine-contaminated water or soil. In humans, leptospires can cause a variety of clinical manifestations, ranging from asymptomatic or mild fever to severe icteric (Weil’s) disease and pulmonary haemorrhage. Currently, little is known about how Leptospira persist within a reservoir host. Prior in vitro studies have suggested that leptospires alter their transcriptomic and proteomic profiles in response to environmental signals encountered during mammalian infection. However, no study has examined gene expression by leptospires within a mammalian host-adapted state. To obtain a more faithful representation of how leptospires respond to host-derived signals, we used RNA-Seq to compare the transcriptome of L. interrogans cultivated within dialysis membrane chambers (DMCs) implanted into the peritoneal cavities of rats with that of organisms grown in vitro. In addition to determining the relative expression levels of ‘‘core’’ housekeeping genes under both growth conditions, we identified 166 genes that are differentially-expressed by L. interrogans in vivo. Our analyses highlight physiological aspects of host adaptation by leptospires relating to heme uptake and utilization. We also identified 11 novel non-coding transcripts that are candidate small regulatory RNAs. The DMC model provides a facile system for studying the transcriptional and antigenic changes associated with mammalian host-adaption, selection of targets for mutagenesis, and the identification of previously unrecognized virulence determinants. Transcriptome analysis of L. interrogans Copenhageni FIOCRUZ L1-130 using RNA from 2 different conditions using RNA-seq. Also, the reproducibility and robustness of data is ensured by three biological replicates from each condition.

Project description:A large body of work has demonstrated that leptospires regulate and modify gene expression in response to environmental cues, as encountered during disease transmission, including changes in temperature, osmolarity, concentration of iron, the presence of serum, and interaction with macrophages. However, since leptospires are not readily amenable to genetic manipulation, the functional and biological significance of these differentially expressed genes often remains unclear. More recently, it is been shown that proteins, in response to changing environmental conditions, can be modified further by specific post-translational modifications. Indeed, both saprophytic and pathogenic leptospires have comprehensive systems to modify proteins. The paucibacillary nature of spirochetal infections, combined with the challenges associated with acquiring pathogens free from contaminating host proteins makes the study of these bacteria in a mammalian host-adapted state inherently difficult. As an alternative approach, we developed a model in which leptospires are cultivated in a dialysis membrane chamber (DMC) implanted within the peritoneal cavity of rats, where they are exposed to the environmental cues encountered during host infection. This strategy has been successfully applied to compare the transcriptome of L. interrogans cultivated within DMC with that of leptospires grown under standard in vitro conditions. In addition to determining the relative expression levels of ‘‘core’’ housekeeping genes under both growth conditions, we identified 166 genes that were differentially-expressed by L. interrogans in response to mammalian host signals. The proteome of DMC (dialysis membrane chamber) cultivated leptospires was compared to that of in vivo derived leptospires and with leptospires cultivated in vitro at 30°C or 37°C by 2-dimensional difference in gel electrophoresis (2-D DIGE). Our analysis indicates that the abundance of leptospire-proteins is modulated the expression of a range of proteins which are differentially expressed in response to mammalian host signals, and not temperature alone . In addition, we confirm that in several proteins there is a change in the presence of modify the expression of the post-translational modifications trimethyllysine and acetyllysine in response to environmental cues encountered during persistent renal colonization in a reservoir host of infection. These results provide novel insights in the proteome level changes, including to differential protein and post-translational modifications, expressed in response to mammalian host signals which can be used to further define the unique equilibrium that exists between pathogenic leptospires and their reservoir host of infection.

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