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Combination therapies have become a standard for the treatment for HIV and hepatitis C virus (HCV) infections. They are advantageous over monotherapies due to better efficacy, reduced toxicity, as well as the ability to prevent the development of resistant viral strains and to treat viral co-infections. Here, we identify new synergistic combinations against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), echovirus 1 (EV1), hepatitis C virus (HCV) and human immunodeficiency virus 1 (HIV-1) in vitro. We observed synergistic activity of nelfinavir with convalescent serum and with purified neutralizing antibody 23G7 against SARS-CoV-2 in human lung epithelial Calu-3 cells. We also demonstrated synergistic activity of nelfinavir with EIDD-2801 or remdesivir in Calu-3 cells. In addition, we showed synergistic activity of vemurafenib with emetine, homoharringtonine, anisomycin, or cycloheximide against EV1 infection in human lung epithelial A549 cells. We also found that combinations of sofosbuvir with brequinar or niclosamide are synergistic against HCV infection in hepatocyte-derived Huh-7.5 cells, and that combinations of monensin with lamivudine or tenofovir are synergistic against HIV-1 infection in human cervical TZM-bl cells. These results indicate that synergy is achieved when a virus-directed antiviral is combined with another virus- or host-directed agent. Finally, we present an online resource that summarizes novel and known antiviral drug combinations and their developmental status. © 2020 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (http://creativecommons.org/licenses/by/4.0/).

Plasmids in conjunction with other mobile elements such as transposons are major players in the genetic adaptation of bacteria in response to changes in environment. Here we show that a large catabolic TOL plasmid, pWW0, from Pseudomonas putida carries genes ( rulAB genes) encoding an error-prone DNA polymerase Pol V homologue which increase the survival of bacteria under conditions of accumulation of DNA damage. A study of population dynamics in stationary phase revealed that the presence of pWW0-derived rulAB genes in the bacterial genome allows the expression of a strong growth advantage in stationary phase (GASP) phenotype of P. putida . When rulAB -carrying cells from an 8-day-old culture were mixed with Pol V-negative cells from a 1-day-old culture, cells derived from the aged culture out-competed cells from the nonaged culture and overtook the whole culture. At the same time, bacteria from an aged culture lacking the rulAB genes were only partially able to out-compete cells from a fresh overnight culture of the parental P. putida strain. Thus, in addition to conferring resistance to DNA damage, the plasmid-encoded Pol V genes significantly increase the evolutionary fitness of bacteria during prolonged nutritional starvation of a P. putida population. The results of our study indicate that RecA is involved in the control of expression of the pWW0-encoded Pol V.

Background The production of recombinant monoclonal antibodies in mammalian cell culture is of high priority in research and medical fields. A critical step in this process is the isolation of the antigen-binding domain sequences of antibodies possessing the desired properties. Many different techniques have been described to achieve this goal, but all have shortcomings; most techniques have problems with robustness, are time-consuming and costly, or have complications in the transfer from isolation to production phase. Here, we report a novel HybriFree technology for the development of monoclonal antibodies from different species that is robust, rapid, inexpensive and flexible and can be used for the subsequent production of antibodies in mammalian cell factories. Results HybriFree technology is illustrated herein via detailed examples of isolating mouse, rabbit and chicken monoclonal antibody sequences from immunized animals. Starting from crude spleen samples, antigen capturing of specific B-cells is performed initially. cDNA of antibody variable domains is amplified from the captured cells and used a source material for simple and rapid restriction/ligation free cloning of expression vector library in order to produce scFv-Fc or intact IgG antibodies. The vectors can be directly used for screening purposes as well as for the subsequent production of the developed monoclonal antibodies in mammalian cell culture. The antibodies isolated by the method have been shown to be functional in different immunoassays, including ELISA, immunofluorescence and Western blot. In addition, we demonstrate that by using a modified method including a negative selection step, we can isolate specific antibodies targeting the desired epitope and eliminate antibodies directed to undesired off-targets. Conclusions HybriFree can be used for the reliable development of monoclonal antibodies and their subsequent production in mammalian cells. This simple protocol requires neither the culturing of B-cells nor single-cell manipulations, and only standard molecular biology laboratory equipment is needed. In principle, the method is applicable to any species for which antibody cDNA sequence information is available. Electronic supplementary material The online version of this article (doi:10.1186/s12896-016-0232-6) contains supplementary material, which is available to authorized users.