New Insights,resistance

The emergence of antibiotic resistance poses a significant global threat, necessitating a deeper understanding of how bacteria evade therapeutic interventions. Among the last-resort antibiotics, colistin plays a crucial role in treating infections caused by multidrug-resistant (MDR) Gram-negative pathogens. However, tolerance to the antimicrobial peptide colistin is a complex phenomenon that can hinder effective treatment. This article delves into the mechanisms and implications of colistin tolerance, drawing upon current scientific understanding to provide a comprehensive overview.

Colistin, a cationic antimicrobial peptide (AMP), exerts its bactericidal effect by disrupting the bacterial cell membrane. Its mechanism of action involves electrostatic interaction with the negatively charged lipopolysaccharide (LPS) on the outer membrane of Gram-negative bacteria, leading to membrane permeabilization and cell death. Despite its efficacy, bacteria can develop various strategies to survive colistin exposure, leading to tolerance and eventual resistance.

One key aspect of colistin tolerance is observed in biofilm-forming bacteria. In mature biofilms, colistin tolerance often reflects penetration limits into the dense matrix and altered physiology rather than changes at the classical target site. Studies have shown that a spatially distinct subpopulation of metabolically active cells within Pseudomonas aeruginosa biofilms can develop tolerance to the antimicrobial peptide colistin. This phenomenon is not solely due to classical target site changes but is influenced by the biofilm's intricate structural organization. The colistin tolerance is influenced by biofilm structural organization, and tolerance induction is basS dependent.

Furthermore, antimicrobial peptides can generate tolerance by lag. This mechanism, observed in bacteria like *Escherichia coli* when treated with different AMPs, particularly those with internal targets, involves a delay in the onset of cell death. This suggests that bacterial populations can adapt to antimicrobial peptide exposure through transient states that reduce immediate susceptibility.

The development of colistin resistance is a significant concern. The evolution of colistin resistance increases bacterial resistance to host antimicrobial peptides and virulence. This implies that widespread use of colistin and other antimicrobials can inadvertently drive the evolution of resistance not only to therapeutic agents but also to the innate immune system's own antimicrobial peptides, such as LL-37. Indeed, data suggest that colistin resistance correlates with increased resistance to the host cationic antimicrobial peptide LL-37.

Mechanisms contributing to colistin tolerance and resistance are diverse. In *P. aeruginosa* biofilms, tolerance to the antimicrobial peptide colistin is linked to metabolically active cells, and can depend on regulatory systems like the pmr and mexAB operons. The mcr-1 gene, a plasmid-mediated colistin resistance determinant, confers resistance to colistin-induced lysis and bacterial cell death, although it may provide minimal protection against other aspects of colistin's action. The discovery of new variants of the mcr-1 gene, such as a new variant with co-resistance to $\beta$-lactam antibiotics, highlights the dynamic nature of resistance evolution and the potential for co-selection of resistance mechanisms.

The clinical implications of colistin tolerance are substantial, leading to difficulties in treating infections caused by biofilm-forming bacteria. Because colistin is considered an antibiotic of last resort, its diminishing efficacy due to tolerance and resistance poses a serious challenge. While colistin is broadly active, its use is associated with side effects like nephrotoxicity and ototoxicity.

Strategies to overcome colistin resistance and tolerance are actively being researched. Combining colistin with other antimicrobial peptides is a promising strategy for bypassing MCR-mediated colistin resistance and enhancing efficacy. For instance, the combination of CDP-B11 plus colistin has shown effectiveness against multiple clinically relevant species *in vitro*. Researchers are exploring novel antimicrobial peptides and strategies to combat MDR, XDR, and colistin-resistant bacteria.

In conclusion, tolerance to the antimicrobial peptide colistin is a multifaceted issue influenced by bacterial physiology, biofilm structure, and evolutionary adaptation. Understanding these mechanisms is critical for developing effective therapeutic strategies against infections caused by resistant pathogens. The ongoing evolution of colistin resistance and the emergence of new resistance genes underscore the urgent need for responsible antibiotic stewardship and continued research into novel antimicrobials and methods to circumvent existing resistance mechanisms.