Anaemia is a major public health concern in developing countries, particularly among children, adolescents, and women of reproductive age. The study aimed to assess the anaemia status among adolescent girls, pregnant, and lactating women with their contributing factors in the southern rural regions of Bangladesh. This cross-sectional study was conducted among 400 adolescent girls, 375 pregnant, and 375 lactating women using a multistage cluster-random sampling technique. Anaemia was measured through haemoglobin concentration in blood capillaries collected with a Hemocue 301 machine. Multinomial logistic regression was used to determine the factors associated with anaemia. The average age of pregnant and lactating women was 24 years and 15.2 years for girls. Overall, the prevalence of anaemia was 50% among pregnant women, 46% among lactating women, and 38% among adolescent girls. The risk of anaemia among adolescent girls was higher among non-Muslim (aOR = 2.13, 95%CI:1.05–4.31), belonged to families having >5 members (aOR = 2.24, 95%CI:1.16–4.31) while exposure to media reduced their risk (aOR = 0.33, 95%CI:0.15–0.74). Pregnant women who consumed a diversified diet, washed their hands after toilet, and received ≥4 ANC visits had a lower likelihood of developing anaemia. Lactating women who were employed, consumed a diversified diet, washed their hands before preparing food, and after toilet, had been exposed to media, received ≥4 ANC visits, and consumed ≥90 IFA, had a lower risk of developing anaemia. However, anaemia was more likely to be associated with lactating women who were non-Muslim (aOR = 3.75; 95%CI:1.26–11.22). The high prevalence of anaemia emphasizes the need to reconsider the existing strategy for the prevention and control of micronutrient deficiencies in Bangladesh.