Application of DNA Chip Scanning Technology for Automatic Detection of Chlamydia trachomatis and Chlamydia pneumoniae Inclusions

Abstract

e Chlamydiae are obligate intracellular bacteria that propagate in the inclusion, a specific niche inside the host cell. The standard method for counting chlamydiae is immunofluorescent staining and manual counting of chlamydial inclusions. High-or medium-throughput estimation of the reduction in chlamydial inclusions should be the basis of testing antichlamydial compounds and other drugs that positively or negatively influence chlamydial growth, yet low-throughput manual counting is the common approach. To overcome the time-consuming and subjective manual counting, we developed an automatic inclusion-counting system based on a commercially available DNA chip scanner. Fluorescently labeled inclusions are detected by the scanner, and the image is processed by ChlamyCount, a custom plug-in of the ImageJ software environment. ChlamyCount was able to measure the inclusion counts over a 1-log-unit dynamic range with a high correlation to the theoretical counts. ChlamyCount was capable of accurately determining the MICs of the novel antimicrobial compound PCC00213 and the already known antichlamydial antibiotics moxifloxacin and tetracycline. ChlamyCount was also able to measure the chlamydial growth-altering effect of drugs that influence host-bacterium interaction, such as gamma interferon, DEAE-dextran, and cycloheximide. ChlamyCount is an easily adaptable system for testing antichlamydial antimicrobials and other compounds that influence Chlamydia-host interactions.

Nonactivated titanium-dioxide nanoparticles promote the growth ofChlamydia trachomatisand decrease the antimicrobial activity of silver nanoparticles

“…As described previously (Bogdanov et al . ), the slide was scanned with a DNA‐chip scanner, and the ChlamyCount software was used to enumerate the chlamydial inclusions. ChlamyCount inclusion number counts supported the qPCR results.…”

“…Image processing and automatic counting of chlamydial inclusions were performed by the ChlamyCount software, as described previously (Bogdanov et al . ).…”

“…TiO 2 NP treatment of HeLa cells and monitoring of C. trachomatis growth on a chamber slide system Chamber slides with 16 wells were used to culture HeLa cells for infection with the C. trachomatis as described previously (Bogdanov et al 2014). Briefly, C. trachomatis (MOI 8) was preincubated with TiO 2 NPs for 1 h at 37°C with a concentration range from 100 to 3Á12 lg ml À1 with twofold dilutions.…”

Impact of antiseptics on Chlamydia trachomatis growth

“…The reason could be that the dynamic range of our immunofluorescent ChlamyCount method was about 2 log 10 (Bogdanov et al . ), while the qPCR method's dynamic range was approx. 5 log 10 (Eszik et al .…”

“…Inclusion counts were determined by the ChlamyCount software as it was described before (Bogdanov et al . ).…”