The Western honey bee (Apis mellifera) is a key pollinator in many natural and managed ecosystems, and is under pressure due to multiple factors, including the infestation with the ectoparasitic mite Varroa destructor. This mite causes serious economic damages for beekeeping, the global honey bee colony losses increasing continuously. As no study investigating the Varroa infestation levels in Romanian apiaries was conducted so far, the objective of this study was to evaluate the Varroa infestation levels in apiaries from southeast of the country and to investigate the effects of different apiary management practices on Varroa infestation rates. Fifty apiaries (five hives/apiary) were monitored between June and October 2024, across eight counties in southeast Romania, to assess the V. destructorinfestation rates and beekeeping practices potentially associated with them. Field data were collected through a questionnaire applied to all 44 beekeepers and Varroa infestation rate (VIR %) was calculated as number of mites per 100 honey bees. Generalized linear mixed models (GLMMs) were performed in R to investigate the relationship between VIR and different apiary characteristics. 52.3% of the beekeepers never monitored the Varroa infestation in their apiaries and almost 30% of them have never participated in any training on beekeeping practices, while 60% of the neighboring beehives were located at less than 1 km from the surveyed apiary. During the five months monitoring, the average VIR across the 250 collected samples was 5.03%. Of these samples, 38.4% exceeded the 3% threshold and 14.4% exhibited a VIR higher than 10%. Regarding the applied anti- Varroatreatments, hard acaricides were the most frequently used, while soft acaricides based on oxalic acid were implemented in 7 organic apiaries and during the honey harvest period or in winter in some conventional apiaries. Based on statistical analyses, VIR decreased significantly as the number of adult honey bees increased, with the presence of the anti- Varroa bottom and with increasing distance from the neighbouring apiaries, while increased significantly during the sampling period, from June to October. Our results highlighted the significant gaps that persist in this sector in Romania regarding the evaluation of the VIR in apiaries across the entire country, the monitoring of mite infestation level, the use of adequate anti- Varroa treatments or the participation in training activities. In addition, future studies are needed to determine to what extent the parasite has developed resistance to some acaricides in Romania.