Infectious diseases have been the leading cause of death for many centuries. The development of vaccination and antibiotic treatments combined with improved hygiene has decreased the number of deaths, but the morality and morbidity associated with infections remain considerable, requiring constant societal awareness and scientific research. An increasing concern are the latent and chronic infections that are often refractory to treatments. As the frequency of latent infections increases with age, it is a major concern for aging societies. The great diversity of the infectious agents, and the multidisciplinary nature of the infectious biology research demand a convergence of various competencies: microbiology, cell biology, animal infection models, immunology etc, emphasizing the need for collaboration between research centres. Especially important are joint activities for smaller countries, e.g. Estonia, where establishment of full- scale stand- alone programs is not economically feasible. Despite the strong positions in basic molecular biology, virology and microbiology, Estonia often fails to capitalize on the excellence in basic research by transitioning to the development of therapeutics targeting medically relevant processes. The main objective of the MIBEst project is to strenghten the research capacity on latent and chronic infections of Institute of Technology at University of Tartu by creating long-lasting links with internationally-leading research institutions: Molecular Infection Medicine Sweden at Umea University, Sweden, and Basel Biozentrum, University of Basel, Switzerland. As an outcome of MIBEst, Estonian scientists will have new knowledge in infection biology with particular focus on advancement in models for latent infections and high throughput screening for promising candidates for antiinfective compounds. Altogether, it enables development of new anti- infection strategies that will have major impact at the national, European and global scale.
Nakkushaigused on olnud paljude sajandite jooksul peamine surmapõhjus. Vaktsineerimise ja antibiootikumiravi arendamine koos parema hügieeniga on vähendanud surmade arvu, kuid infektsioonidega seotud suremus ja tüsistused on endiselt märkimisväärsed, nõudes pidevat ühiskondlikku teadlikkust ja teaduslikku uurimistööd. Suurenev mure kroonilised infektsioonid, mis on sageli ravile ei allu. Kuna krooniliste infektsioonide esinemissagedus suureneb koos vanusega, on see vananeva ühiskonna jaoks suur probleem. Nakkusetekitajate suur mitmekesisus ja nakkustekitajate bioloogia uuringute multidistsiplinaarne olemus nõuavad erinevate pädevuste ühendamist: mikrobioloogia, rakubioloogia, loomade nakatumise mudelid, immunoloogia jne. Seega on suur vajadus uurimiskeskuste vahelise koostöö järele. Eriti oluline on koostöö väiksemate riikide jaoks, nt. Eesti, kus täieulatuslike iseseisvate programmide loomine ei ole majanduslikult teostatav. Vaatamata tugevale positsioonile molekulaarbioloogias, viroloogias ja mikrobioloogias, ei suuda Eesti teadlased on tulemusi efektiivselt rakendada. MIBEsti projekti peamine eesmärk on tugevdada Tartu Ülikooli Tehnoloogiainstituudi uurimissuutlikkust krooniliste infektsioonide alal, luues pikaajalisi sidemeid rahvusvaheliselt juhtivate teadusasutustega: Rootsi Molekulaarinfektsioon Keskus Umea Ülikoolis, ja Baseli Biokeskus, Baseli ülikoolis, Šveitsis. MIBEst projekti tulemusena on Eesti teadlastel uusi teadmisi nakkusbioloogias, see juures rõhuasetusega kroonilistele infektsioonidele. Kokkuvõttes võimaldab see välja töötada uusi infektsioonivastaseid strateegiaid, millel on suur mõju riiklikul, Euroopa ja ülemaailmsel tasandil.
Azithromycin is a clinically important drug for treating invasive salmonellosis despite poor activity in laboratory assays for MIC. Addition of the main buffer in blood, bicarbonate, has been proposed for more physiologically relevant and more predictive testing conditions. However, we show here that bicarbonate-triggered lowering of azithromycin MIC is entirely due to alkalization of insufficiently buffered media. In addition, bicarbonate is unlikely to be altering efflux pump activity.
RelA-SpoT Homolog (RSH) enzymes control bacterial physiology through synthesis and degradation of the nucleotide alarmone (p)ppGpp. We recently discovered multiple families of small alarmone synthetase (SAS) RSH acting as toxins of toxin-antitoxin (TA) modules, with the FaRel subfamily of toxSAS abrogating bacterial growth by producing an analog of (p)ppGpp, (pp)pApp. Here we probe the mechanism of growth arrest used by four experimentally unexplored subfamilies of toxSAS: FaRel2, PhRel, PhRel2, and CapRel. Surprisingly, all these toxins specifically inhibit protein synthesis. To do so, they transfer a pyrophosphate moiety from ATP to the tRNA 3′ CCA. The modification inhibits both tRNA aminoacylation and the sensing of cellular amino acid starvation by the ribosome-associated RSH RelA. Conversely, we show that some small alarmone hydrolase (SAH) RSH enzymes can reverse the pyrophosphorylation of tRNA to counter the growth inhibition by toxSAS. Collectively, we establish RSHs as RNA-modifying enzymes.
Toxin–antitoxin (TA) gene pairs are ubiquitous in microbial chromosomal genomes and plasmids as well as temperate bacteriophages. They act as regulatory switches, with the toxin limiting the growth of bacteria and archaea by compromising diverse essential cellular targets and the antitoxin counteracting the toxic effect. To uncover previously uncharted TA diversity across microbes and bacteriophages, we analyzed the conservation of genomic neighborhoods using our computational tool FlaGs (for flanking genes), which allows high-throughput detection of TA-like operons. Focusing on the widespread but poorly experimentally characterized antitoxin domain DUF4065, our in silico analyses indicated that DUF4065-containing proteins serve as broadly distributed antitoxin components in putative TA-like operons with dozens of different toxic domains with multiple different folds. Given the versatility of DUF4065, we have named the domain Panacea (and proteins containing the domain, PanA) after the Greek goddess of universal remedy. We have experimentally validated nine PanA-neutralized TA pairs. While the majority of validated PanA-neutralized toxins act as translation inhibitors or membrane disruptors, a putative nucleotide cyclase toxin from a Burkholderia prophage compromises transcription and translation as well as inducing RelA-dependent accumulation of the nucleotide alarmone (p)ppGpp. We find that Panacea-containing antitoxins form a complex with their diverse cognate toxins, characteristic of the direct neutralization mechanisms employed by Type II TA systems. Finally, through directed evolution, we have selected PanA variants that can neutralize noncognate TA toxins, thus experimentally demonstrating the evolutionary plasticity of this hyperpromiscuous antitoxin domain.
(Microbiol Mol Biol Rev, 2020-11-11) Kaldalu, N; Hauryliuk, V; Turnbull, Kathryn Jane; Putrinš, M; Tenson, Tanel
Many bacterial pathogens can permanently colonize their host and establish either chronic or recurrent infections that the immune system and antimicrobial therapies fail to eradicate. Antibiotic persisters (persister cells) are believed to be among the factors that make these infections challenging. Persisters are subpopulations of bacteria which survive treatment with bactericidal antibiotics in otherwise antibiotic-sensitive cultures and were extensively studied in a hope to discover the mechanisms that cause treatment failures in chronically infected patients; however, most of these studies were conducted in the test tube. Research into antibiotic persistence has uncovered large intrapopulation heterogeneity of bacterial growth and regrowth but has not identified essential, dedicated molecular mechanisms of antibiotic persistence. Diverse factors and stresses that inhibit bacterial growth reduce killing of the bulk population and may also increase the persister subpopulation, implying that an array of mechanisms are present. Hopefully, further studies under conditions that simulate the key aspects of persistent infections will lead to identifying target mechanisms for effective therapeutic solutions.