Objective To investigate the staffing status of full-time personnel for healthcare-associated infection (HAI) management department in secondary or above medical institutions in Yunnan Province, analyze the factors affecting the stability of the HAI workforce, and provide a scientific basis for strengthening the department of HAI professionals and improving the standard of the full-time personnel team. Methods In April 2023, a questionnaire survey was conducted to investigate the staffing status of full-time HAI management department personnel across 394 secondary or above medical institutions in Yunnan Province. Results A total of 394 secondary or above medical institutions were included in this survey. Among the institutions participating in the survey, 393 (99.75%) had established a separate hospital-acquired infection management, and 303 (76.90%) met the requirement of having at least one full-time infection control personnel for every 200 beds. Among the 689 full-time HAI management department practitioners surveyed, the majority were female (94.34%), those aged 41-50 were the largest age group (34.69%), the predominant educational background was a bachelor’s degree (72.42%), the primary specialty was nursing (80.84%), and the most common professional title was intermediate level (36.14%). There were statistically significant differences between the full-time HAI practitioners in secondary medical institutions and those in tertiary medical institutions in educational background (Z=–6.220, P<0.001), specialty (χ2=41.612, P<0.001), professional title (Z=–2.432, P=0.015), and years of engagement (χ2=47.298, P<0.001). There was no statistically significant difference in gender or age (P>0.05). The primary reason for full-time practitioners engaging in HAI work was “following leadership arrangements” (79.97%). Tertiary medical institutions had a significantly higher proportion of full-time HAI management personnel who took up their positions due to “graduation recruitment” or “major matching” than secondary medical institutions (P<0.05), while the proportion of those who entered the field due to “leadership arrangement” was significantly lower than that in secondary medical institutions (P<0.05). In the survey on factors contributing to the instability of the full-time personnel team, the top three factors identified were excessive work pressure, difficulties in professional title promotion, and an overwhelming amount of required learning content. Conclusions There is an insufficient allocation of full-time HAI control personnel in secondary or above medical institutions in Yunnan Province, and the professional composition of this workforce requires greater diversification. The primary factors contributing to the instability of the dedicated personnel team are high work pressure and difficulties in professional title promotion. It is recommended to expedite the improvement of career advancement pathways for HAI control personnel to provide support and guarantees for building a robust full-time infection control workforce.