During the COVID-19 pandemic, sex workers (SW) were one of the vulnerable groups affected by lockdown measures. COVID-19 had also disrupted HIV/Sexually transmitted infection (STI) testing and treatment services for sex workers due to numerous restrictions in specialist medical care. This study aims to assess the seroprevalence of HIV, syphilis, HBV, and HCV and associated factors among SW as COVID-19 restrictions were lifted. The SW aged over 18 years residing in Chiangmai, Thailand, were recruited between March and December 2022. An interview-based questionnaire was administered. Blood was collected for HIV, syphilis, HBV, and HCV serological testing. Logistic regression models were used to examine factors associated with these serological markers. Of 264 SW recruited, 52.3% were male. The median age was 31 years. Male sex workers (MSW) had higher seroprevalence of HIV (13% vs. 4.8%), syphilis (23.9% vs. 6.4%) and HCV (6.5% vs. 2.4%). Female sex workers (FSW) had higher seroprevalence of HBsAg (9.5% vs. 4.4%). A high proportion were unaware of their HIV/STI infection. MSW reporting receptive anal sex were more likely to be HIV and Treponema Ab positive. MSW reporting drug injection history were more likely to be HCV Ab positive. FSW reporting younger age at first sex were more likely to be HIV Ab positive. In conclusion, SW remains particularly affected by HIV/STIs. Despite the lockdown, HIV/STIs continued to spread, highlighting the need to provide access to HIV/STIs testing, prevention, and treatment services for this population, particularly young men.