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The relative quantification of HSV\u20102 was conducted with anal and genital samples from men who have sex with men (MSM), living with HIV. The presence of a single amplification product was validated with a dissociation curves profile and the determination of the melting temperature. The limit of detection for \u03b2\u2010globin was determined as 3.3\u2009\u00d7\u200910<jats:sup>\u22125<\/jats:sup>\u2009ng\/\u03bcL, and for HSV\u20102 at 6.0\u2009\u00d7\u200910<jats:sup>\u22126<\/jats:sup>\u2009ng\/\u03bcL. The efficiency for \u03b2\u2010globin was 100.2% and for HSV\u20102 was 106.8%. From 336 MSM, 2.1% and 3.9% individuals presented anal or genital HSV\u20102 shedding, respectively. The HSV\u20102 viral load was 9.2 RU, individuals with fewer CD4<jats:sup>+<\/jats:sup> presented higher HSV\u20102 viral load. 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