Development of a Highly Specific, Selective, and Sensitive Fluorescent Probe for Detection of Mycobacteria in Human Tissues

Abstract

Tuberculosis (TB), including extrapulmonary TB, is responsible for more than one million deaths in a year worldwide. Existing methods of mycobacteria detection have poor sensitivity, selectivity, and specificity, especially in human tissues. Herein, the synthesis of a cholic acid‐derived fluorescent probe (P4) that can specifically stain the mycobacterium species is presented. It is shown that P4 probe specifically binds with mycobacterial lipids, trehalose monomycolate, and phosphatidylinositol mannoside 6. P4 probe can detect mycobacteria in polymicrobial planktonic cultures and biofilms with high specificity, selectivity, and sensitivity. Moreover, it can detect a single mycobacterium in the presence of 10 000 other bacilli. Unlike the probes that depend on active mycobacterial enzymes, the membrane‐specific P4 probe can detect mycobacteria even in formalin‐fixed paraffin‐embedded mice and human tissue sections. Therefore, the ability of the P4 probe to detect mycobacteria in different biological milieu makes it a potential candidate for diagnostic and prognostic applications in clinical settings.

Hydrophobicity of Cholic Acid-Derived Amphiphiles Dictates the Antimicrobial Specificity

“…30,31 Recently, we have shown that the conjugation of DMAP head groups with three hydroxyl groups of CA makes them highly selective for mycobacterial membranes. 32 We further demonstrated the ability of the amphiphile to detect mycobacteria with high selectivity, specificity, and sensitivity. Here, we propose to examine the effect of lipophilicity on the antimicrobial properties of CAderived amphiphiles containing DMAP head groups.…”

“…In-depth mechanistic studies of AMPs against mycobacterial membranes are unknown, but we have shown that molecules like amphiphile 1 perform specific interactions with mycobacterial lipids like trehalose mycolates, making them active. 32 Addition of the hydrophobic alkyl chains disrupts their interactions with mycobacterial lipids, thereby making them less effective.…”

“…In a previous study, we evaluated the impact of hydrophobicity on the antibacterial activities of cholic acid (CA)–glycine conjugates against Gram-positive and Gram-negative bacteria and showed that an increase in the chain length enhances the activity of CA-derived amphiphiles. , Recently, we have shown that the conjugation of DMAP head groups with three hydroxyl groups of CA makes them highly selective for mycobacterial membranes . We further demonstrated the ability of the amphiphile to detect mycobacteria with high selectivity, specificity, and sensitivity.…”

“…Therefore, DMAP charged group in the case of amphiphile 7 might provide the key electrostatic interactions and the alkyl chain allows hydrophobic interactions with bacterial lipids, making it ideal for activity against Gram-negative/Gram-positive bacteria. In-depth mechanistic studies of AMPs against mycobacterial membranes are unknown, but we have shown that molecules like amphiphile 1 perform specific interactions with mycobacterial lipids like trehalose mycolates, making them active . Addition of the hydrophobic alkyl chains disrupts their interactions with mycobacterial lipids, thereby making them less effective.…”

N-methyl Benzimidazole Tethered Cholic Acid Amphiphiles Can Eradicate S. aureus-Mediated Biofilms and Wound Infections

“…The presence of both a steroidal backbone and hydroxyl groups provide a facial amphiphilic nature to CA and also the potential to behave as mimics of AMPs [ 13 ]. Derivatives of CA have been widely exploited as antimicrobial [ 14 ], anticancer [ 15 ], and diagnostic agents [ 16 ]. Savage and coworkers in their pioneering work developed bile acid-based antimicrobial agents (called Ceragenins) that demonstrated broad-spectrum antibacterial properties against different Gram-positive and Gram-negative pathogens [ 17 , 18 , 19 , 20 ].…”