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  • Some studies using RNAi have recently

    2021-11-30

    Some studies using RNAi have recently been invalidated by CRISPR/Cas9 [25] due to significant off-target effects of RNAi [26]. In this study, we have used both RNAi and CRISPR/Cas9 techniques and have observed consistent and similar phenotypes thus; knockdown or knockout of PATZ1 resulted in reduction in infection and late RT products of HIV-1 (Fig. 2A–C, H and Fig. 3D, E, G). Moreover, two PATZ1 isoforms rescued reporter virus infection in PATZ1-KO cells, whilst one isoform was sufficient in PATZ1-KD Epigallocatechin mg probably due to traces of both isoforms in knockdown cells, a distinguishing feature between RNAi mediated knockdown and CRISPR/Ca9 frameshifting mutagenesis. Notably, knockdown or knockout of PATZ1 vitiated HIV-1 infection but not MLV infection (Fig. 2, Fig. 3F). Even though MLV and HIV-1 are retroviruses and have many steps in common [1], there are plenty of differences as well. MLV reverse transcriptase (RT), for example, is a simple monomer of 78 kDa whilst that of HIV-1 is a heterodimer containing 66 and 51 kDa subunits with each RT showing a distinctive preference for homopolymeric templates and tRNAs [27]. These variations in structure or function of reverse transcriptase and by extension reverse transcription could mean different interactions of HIV-1 or MLV with cellular proteins and explain different outcomes.
    Conflicts of interest
    Acknowledgements We thank Dr. ISY Chen for pNL4-3luc (env-/nef-), pHCMV-VSV-G and pCMV-DR8.2 packaging plasmids and Dr. Feng Zhang for pSpCas9(BB)-2A-Puro (PX459) V2.0 plasmid. We also recognize the support from Drs. Eiji Ido, William Ampofo, and Jacob Barnor.
    Background Syphilis, a sexually transmitted infection (STI) caused by the bacterium Treponema pallidum, is a significant global health problem with approximately 6 million new cases per year (Newman et al., 2015). However, while syphilis is curable, both male and female rates of primary and secondary syphilis have increased in every region, in every age group and every race/ethnicity group in the U.S. (Centers for Disease Control and Prevention (CDC), 2017), and countries across the world are seeing large increases in syphilis in key populations such as men who have sex with men (MSM) (Abara et al., 2016, Chen et al., 2017, Kojima and Klausner, 2018). HIV co-infection with STIs is common (Kalichman et al., 2011, Rieg et al., 2008). Approximately one-half of all MSM diagnosed with syphilis are also co-infected with HIV (Centers for Disease Control and Prevention (CDC), 2017). Routine screening and subsequent timely treatment are mainstays of HIV and STI control programs. Dual rapid tests for HIV and syphilis antibody detection are available and recommended by the World Health Organization (World Health Organization (WHO), 2017). Those tests can be conducted at the point-of-care and can be done using a drop of blood from a fingerprick. Rapid tests may make screening more feasible and accessible (Gliddon et al., 2017, Tucker et al., 2013). Dual rapid tests for HIV and syphilis allow for the simultaneous screening for the two infections using one specimen and one test device. We aimed to estimate the test accuracy of the Standard Q HIV/Syphilis Combo Test (SD Biosensor, South Korea), a new dual rapid test using stored sera in a laboratory setting in Lima, Peru. The Standard Q HIV/Syphilis Combo uses a lateral flow format similar to other rapid tests on the market, however it tests for and distinguishes between antibodies to HIV-1, HIV-2 and syphilis. The test is not yet commercially available but is under evaluation for CE marking, World Health Organization Pre-qualification and Registration in Brazil, Peru and Paraguay.
    Methods
    Discussion HIV and syphilis continue to cause morbidity and mortality around the globe (Kojima and Klausner, 2018, UNAIDS, 2016). Strategies to improve the uptake and reach of routine screening are urgently needed. Rapid dual testing is one strategy that may have impact. A recent meta-analysis of 18 studies of the diagnostic accuracy of various dual HIV and syphilis tests found that the laboratory performance of dual tests tended to be high, the sensitivity of HIV antibody detection ranged from 98% to 100%, and specificity from 92% to 100% (Gliddon et al., 2017). The meta-analysis also found that for syphilis antibody detection, reported sensitivities ranged from 93% to 100% and specificity values ranged from 93% to 100%. Our findings are within that same range. Because that meta-analysis found that there was a reduction in diagnostic accuracy in field settings compared to laboratory settings for syphilis (Gliddon et al., 2017), the Standard Q HIV/Syphilis Combo test should be evaluated in field settings using fingerprick whole blood specimens.