Enhanced detection of low-expressed miRNAs in Leishmania-infected macrophages through RNA fractionation and RT-qPCR optimization
Maestrini, S.; Lecoeur, H.; Gharsallah, C.; Diotallevi, A.; Späth, G. F.; Galluzzi, L.; Prina, E.
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MicroRNAs (miRNAs) play critical roles in regulating host responses to Leishmania infections, yet accurate detection of low-abundance miRNAs remains challenging. This study evaluated the impact of RNA fractionation and RT-qPCR protocol optimization on miRNA quantification in Leishmania amazonensis-infected murine macrophages. Using a panel of nine infection-associated miRNAs, we compared small RNA and total RNA fractions for their ability to detect weakly expressed targets. Small RNA consistently provided greater sensitivity and specificity, particularly when combined with a modified RT-qPCR protocol. These findings underscore the importance of RNA preparation methods for studying miRNA dynamics in infectious disease contexts and support improved approaches for detecting biologically relevant, low-expressed miRNAs.
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