MINI-REVIEW Focus on recently developed assays for detection of resistance/sensitivity to reverse transcriptase inhibitors Francesca Marino-Merlo 1 & Beatrice Macchi 2 & Daniele Armenia 3 & Maria Concetta Bellocchi 3 & Francesca Ceccherini-Silberstein 3 & Antonio Mastino 4,5 & Sandro Grelli 3 Received: 12 July 2018 /Revised: 7 September 2018 /Accepted: 9 September 2018 # Springer-Verlag GmbH Germany, part of Springer Nature 2018 Abstract The biology of HIV is rather complex due to high rate of replication, frequent recombination, and introduction of mutations. This gives rise to a number of distinct variants referred as quasispecies. In addition, the latency within reservoir allows the periodic reactivation of virus replication. The rapid replication of HIV allows immune response escape and establishment of resistance to therapy that can be acquired through drug selection and/or transmitted among individuals. This prompted, over the years, the development of a range of assays aimed to determine drug resistance and sensitivity, to be used both in clinical practice and in antiviral research. Reverse transcriptase (RT) inhibitors have an eminent place among the anti-HIV drugs, being constantly present from the beginning until today in the most commonly used antiviral regimens. This mini-review seeks to provide an up- to-date overview of recent efforts in developing even more reliable and simple methods, of both genotypic and phenotypic types, for specifically detecting drug resistance and sensitivity to RT inhibitors. Keywords HIV . Reversetrancriptase . Nucleosidereversetranscriptaseinhibitors . Non nucleosidereverse transcriptase inhibitors Introduction HIV replication consists in a complex sequence of steps giving rise to a huge amount of viral particles estimated around 10 10 virions per day per infected, untreated individual. The high replicative ability of HIV explains both transmission and spreading of infection and the outcome of mutant strains. Nevertheless, the introduction of antiretroviral therapy (ART) has deeply changed the survival expectancy of HIV- infected, treated individuals. The first evidence for an effective outcome on clinical progression and death of a monotherapy regimen based on the nucleoside reverse transcriptase (RT) inhibitor (NRTI) zidovudine dates back more than 30 years ago. Although, the inadequateness of a NRTI-monotherapy based ART strategy was soon clear, resulting in incomplete virological suppression and resistance mutations onset. Thus, in the mid-1990s, the introduction of a novel therapeutic strat- egy consisting of two NRTIs plus a protease inhibitor (PI) or a non-nucleoside RT inhibitor (NNRTI) dramatically changed the course of the disease by highly affecting morbidity and mortality of treated patients. With the ongoing of time several drawbacks, such as unfavorable pharmacokinetic of PIs, tox- icity and emergence of high-level class resistance of NNRTIs, toxicity of first-generation NRTIs, drug-drug interactions, problems of patient compliance due to high pill burden, and difficulties in achieving a complete virological suppression, as highlighted by sensitive assays to measure viraemia, led to reconsider first-generation, combination-based, ART regi- mens. This occurred by developing second-generation NRTIs, NNRTIs, and PIs, less toxic and longer-acting, and, more recently, by introducing new classes of licensed drugs, such as integrase inhibitors (INIs), entry inhibitors, chemokine receptor inhibitors, ordered according to their approval, aimed to substitute or to flank the other classes of drugs. In particular, Francesca Marino-Merlo and Beatrice Macchi contributed equally to this work. * Antonio Mastino antonio.mastino@unime.it 1 IRCCS Centro Neurolesi Bonino-Pulejo, Messina, Italy 2 Department of Systems Medicine, University of Rome BTor Vergata^, Rome, Italy 3 Department of Experimental Medicine and Surgery, University of Rome BTor Vergata^, Rome, Italy 4 Department of Chemical, Biological, Pharmaceutical, and Environmental Sciences, University of Messina, Via F. Stagno d’Alcontres 31, 98166 Messina, Italy 5 The Institute of Translational Pharmacology, CNR, Rome, Italy Applied Microbiology and Biotechnology https://doi.org/10.1007/s00253-018-9390-x