Metronidazole and Giardia: In Vitro Viability Assay under Microaerobic Conditions Indicates a Multifactorial Basis for Metronidazole Treatment Failure.

Objectives: Giardiasis, a worldwide intestinal infection caused by the protozoan parasite Giardia intestinalis, is treatable with metronidazole (MTZ). However, MTZ-refractory giardiasis is common. It remains unclear, though, whether MTZ-resistant pathogens cause treatment failure because the natural resistance of Giardia to MTZ has not yet been demonstrated.

Methods: We developed a simple 24-hour viability assay to assess MTZ sensitivity of Giardia parasites in vitro under two atmospheric conditions, microaerobic and anaerobic. The results of the assay were statistically evaluated.

Results: We tested 18 clinical isolates. Based on the minimum lethal concentration (MLC), hierarchical cluster analysis separated the isolates into three categories, sensitive, intermediate resistant, and resistant. The resistant cluster consisted of two isolates with an MLC of 400µg/ml MTZ, which exhibited natural MTZ resistance. Interestingly, this resistance was only manifested under microaerobic conditions. The effect of oxygen on the in vitro drug response was evident in the greater variability of MLC values among the other isolates. Two in vitro-resistant isolates originated from patients with MTZ-refractory giardiasis, suggesting that parasite resistance likely contributes to treatment failure. However, two additional isolates from patients with MTZ-refractory giardiasis showed in vitro susceptibility under both test conditions. This indicates that treatment failure in giardiasis likely stems from multiple factors.

Conclusions: Our study highlights the critical importance of oxygen concentration during the assessment of the Giardia parasite resistance to MTZ. However, it also indicates that MTZ refractory giardiasis may result from other reasons than parasite´s resistance.