The aim of this study was to evaluate the efficacy of a progesterone-based treatment on anoestrus in buffaloes. Primiparous acyclic buffaloes (n = 276), were divided into three classes according to their days in milk (DIM): from 50 to 90 (Class I; n = 86), from 91 to 150 (Class II; n = 102) and from 150 to 200 (Class III; n = 88). Animals were synchronized using P4 vaginal implants, followed by timed artificial insemination (TAI). They were then allowed to enter into a larger group of buffaloes for natural mating 15 days after AI was performed, and pregnancy status was monitored from then on at 15-day intervals. Finally, the temperature–humidity index (THI) was calculated. Statistical analysis was performed by ANOVA by means and both multiple and linear regression. The total pregnancy rate (PR) was 87.7%, with no differences among DIM classes (88.0, 92.4, and 80.0% in Classes I, II, and III, respectively). However, the PR at TAI tended to be higher (p = 0.07) in buffaloes in Class II. The follicle (FL) area in Class II buffaloes was larger (p < 0.01) than that of the other classes. No influence of the THI on the total PR was recorded. The pregnancy outcome at TAI was affected by the FL area (odds ratio = 2.237; p < 0.05) and body condition score (BCS) (odds ratio = 1.256; p < 0.05). In conclusion, treatment with vaginal P4 optimizes pregnancy rates in anoestrus buffaloes, particularly when the animals are in mid-lactation and show an optimal BCS. Furthermore, the THI does not seem to affect the efficiency of the progesterone treatment.