Design and Expression of Specific Hybrid Lantibiotics Active Against Pathogenic Clostridium spp.

Abstract

Clostridium difficile has been reported as the most common cause of nosocomial diarrhea (antibiotic-associated diarrhea), resulting in significant morbidity and mortality in hospitalized patients. The resistance of the clostridial spores to antibiotics and their side effects on the gut microbiota are two factors related to the emergence of infection and its relapses. Lantibiotics provide an innovative alternative for cell growth inhibition due to their dual mechanism of action (membrane pore-forming and cell wall synthesis inhibition) and low resistance rate. Based on the fact that bacteriocins are usually active against bacteria closely related to the producer strains, a new dual approach combining genome mining and synthetic biology was performed, by designing new lantibiotics with high activity and specificity toward Clostridium. We first attempted the heterologous expression of putative lantibiotics identified following Clostridium genome mining. Subsequently, we designed new hybrid lantibiotics combining the start or end of the putative clostridial peptides and the start or end parts of nisin. The designed peptides were cloned and expressed using the nisin biosynthetic machinery in Lactococcus lactis. From the 20 initial peptides, only 1 fulfilled the requirements established in this work to be considered as a good candidate: high heterologous production level and high specificity/activity against clostridial species. The high specificity and activity observed for the peptide AMV10 makes it an interesting candidate as an alternative to traditional antibiotics in the treatment of C. difficile infections, avoiding side effects and protecting the normal gut microbiota.

Combating Antimicrobial Resistance With New-To-Nature Lanthipeptides Created by Genetic Code Expansion

“…Using lantibiotic-sensitive strains such as L. lactis strain NZ9000, Listeria monocytogenes, Clostridioides difficile, etc. have been reported in the literature (Field et al, 2010;Cebrián et al, 2019).…”

Cell wall homeostasis in lactic acid bacteria: threats and defences

“…Also, deep understanding on the mode of action of bacteriocins targeting the CW, their specific interactions with receptors and the mechanisms behind resistance allow the rational design of potent bacteriocin variants with wider spectra and able to circumvent resistance mechanisms (Li, Montalban-Lopez and Kuipers 2018 ; Cebrián et al . 2019 ; Field et al . 2019 ).…”

Bacteriocins From LAB and Other Alternative Approaches for the Control of Clostridium and Clostridiodes Related Gastrointestinal Colitis

“…The genus Clostridium belongs to the family Clostridiaceae and, with presently an estimated of more than 203 metabolically diverse species, only a few species are recognized as being pathogenic to humans (NHS, 2016). Representatives are Gram-positive, endospore-forming anaerobic bacteria that are ubiquitously found in virtually all anoxic habitats where organic compounds are present, including soils, aquatic sediments, and the intestinal tracts of animals and humans (Udaondo et al, 2017;Cebrian et al, 2019). By far the majority of the Clostridium spp.…”

“…are considered to be commensal, but a few such as C. perfringens, C. botulinum, C. tetani, and Clostridiodes (Cd.) difficile are known to be opportunistic, toxin-producing pathogens in both animals and humans and may be associated with a high mortality rate (Cassir et al, 2016;Cebrian et al, 2019;Czepiel et al, 2019). One particular species of the pathogenic 'clostridia, ' Cd.…”

“…difficile, is receiving increased attention from the clinical community, due to its high resistance to antibiotics and the growing number of especially nosocomial infections with Cd. difficile as one of the most common causes (Cebrian et al, 2019;Czepiel et al, 2019). A recent proposal to restrict the genus Clostridium to the C. butyricum clade and related species resulted in ramifications for "unrelated" Clostridium spp.…”