A small-molecule compound belonging to a class of 2,4-disubstituted 1,3,4-thiadiazine-5-ones inhibits intracellular growth and persistence of Chlamydia trachomatis

Abstract

A small-molecule compound belonging to a class of 2,4-disubstituted 1,3,4-thiadiazine-5-ones inhibits intracellular growth and persistence of Chlamydia trachomatis Chlamydia trachomatis is one of the most common sexually transmitted pathogens in the world and often causes chronic inflammatory diseases that are insensitive to antibiotics. The type 3 secretion system (T3SS) of pathogenic bacteria is a promising target for therapeutic intervention aimed at bacterial virulence and can be an attractive alternative for the treatment of chronic infections. Recently, we have shown that a small-molecule compound belonging to a class of 2,4-disubstituted 1,3,4-thiadiazine-5-ones produced through the chemical modification of the thiohydrazides of oxamic acids, designated CL-55, inhibited the intracellular growth of C. trachomatis in a T3SS-dependent manner. To assess the feasibility of CL-55 as a therapeutic agent, our aim was to determine which point(s) in the developmental cycle CL-55 affects. We found that CL-55 had no effect on the adhesion of elementary bodies (EBs) to host cells but significantly suppressed EB internalization. We further found that CL-55 inhibited the intracellular division of reticulate bodies (RBs). An ultrastructural analysis revealed loss of contact between the RBs and the inclusion membrane in the presence of CL-55. Finally, we found that our T3SS inhibitor prevented the persistence of Chlamydia in cell culture and its reversion to the infectious state. Our findings indicate that our T3SS inhibitor may be effective in the treatment of both productive and persistent infections.

Chlamydial Type III Secretion System Needle Protein Induces Protective Immunity againstChlamydia muridarumIntravaginal Infection

“…C. muridarum (strain Nigg) ATCC VR-123 was grown in cycloheximide-treated McCoy cells (a hybrid cell line consisting of human synovial and mouse fibroblasts) as described previously [41, 42]. Chlamydial elementary bodies (EBs) were harvested, purified, quantified, and stored at −70°C in SPG buffer (sucrose/phosphate/glutamic acid: 0.2 M sucrose, 20 mM sodium phosphate, and 5 mM glutamic acid).…”

“…The canal and exocervix were vigorously scraped, and swabbed material was transferred to DMEM and frozen immediately at −70°C. Bacterial loads were determined as previously described [41]. Thawed, undiluted samples (0.5 mL) were plated onto McCoy cells on coverslips (12 mm in diameter) in 24-well plates, and the infection was enhanced by centrifugation at 800 ×g for 1 h. After centrifugation, the inocula were removed, 1 mL of complete DMEM (DMEM with 10% fetal calf serum, 1% nonessential amino acids, 2 mM L-glutamine, 1% vitamins, and 0.5% glucose) was added, and the plates were incubated in a humidified incubator with 5% CO 2 at 37°C for 48 h. Coverslips were incubated with FITC-conjugated monoclonal antibodies against chlamydial lipopolysaccharide (Nearmedic Plus, Moscow, Russia).…”

Preventative treatment with Fluorothiazinon suppressed Acinetobacter baumannii-associated septicemia in mice

“…We continued to study the mechanism of FT action on Acinetobacter infection. For FT, a specific effect on the type three secretion system has been previously shown in Pseudomonas, Salmonella, and Chlamydia [4,[6][7][8]33]. It is likely that FT has another target in A. baumannii because they have not yet been shown to have the T3SS.…”

Anti-Idiotypic Nanobodies Mimicking an Epitope of the Needle Protein of the Chlamydial Type III Secretion System for Targeted Immune Stimulation

“…C. muridarum (Nigg strain) ATCC VR-123 was grown in cycloheximide-treated McCoy cells (a hybrid cell line consisting of human synovial cells and mouse fibroblasts) as described previously [43,53]. The chlamydial elementary bodies (EBs) were collected, purified, quantified and stored at −70 • C in SPG buffer (sucrose/phosphate/glutamic acid: 0.2 mM sucrose, 20 mM sodium phosphate and 5 mM glutamic acid).…”