OR02

Granulomas can efficiently contain Mtb infection, but not completely eradicate it, due to Mtb’s ability to persist in this environment. Thus, they represent a battlefield that dictates both the host immune and bacterial responses. Recently, human PBMC-related in vitro models were developed to mimic the granuloma microenvironment.

In this study, a GLS assay was used to evaluate drug efficacy. PBMCs isolated from donors were infected with Mtb and cells were observed up to 13 days post-infection. After 6 days, formation of GLSs were observed in wells containing infected cells. The drugs were added at concentrations of 1, 2 and 4xMIC to the infected PBMCs, at day 7 post-infection. CFU counts on cell lysates were performed at different days both in the base and the supernatant of the wells.

The trend of Mtb growth was similar in the samples coming from different donors. It was observed that the first-line drug isoniazid had a bacterial growth-reducing effect in the granuloma, while rifampicin had a greater effect in the supernatant. Using GFP-expressing Mtb and confocal microscopy, it was seen that most of the bacteria localize inside GLS. Finally, removal of the supernatant and replacement with supernatant from another not-infected well had no significant decrease of Mtb growth at the base and eliminated bacteria in the supernatant.

In conclusion, the GLS assay represents a promising model to study early events in host-pathogen interaction and to evaluate drug efficacy.

This work has received support from the Innovative Medicines Initiatives 2 Joint Undertaking (grant No 853989).