Background: The emergence of multi-drug resistant tuberculosis (MDR-TB) is a challenge for global prevention and control of the disease. MDR-TB case management is difficult because it exposes to further economic and social costs that the patients face while seeking help and treatment. Ethiopia is one of the 30 MDR-TB burden countries. There is little research evidence of MDR-TB from the peripheral parts of the country.Objective: The study was designed to identify determinants of MDR-TB among patients attending anti tuberculosis treatment from peripheral districts, Southern Nations, Nationalities and People Region, Ethiopia, 2019. Methods: Hospital based case control study was conducted from March to April, 2019 in Southern Ethiopia. Cases were confirmed MDR-TB patients, while controls were those who declared cured or completed first line ant-tuberculosis treatment. The study participants were recruited by stratified random sampling. The data were entered into Epi data 4.4.3, cleaned and analyzed by SPSS 24. Bivariate and multivariable analyses was used to identify determinants of MDR-TB. Determinants with P-value <0.05 were declared as having significant association with MDR-TB and adjusted odd ratio with 95% CI was used to measure degree of association. Result: A total of 180 study participants were recruited (90 cases and 90 controls) and participated in this study. The median age for cases and controls was 29 and 30.5 years respectively. More than two third (78.89%) of case participants were from rural and 49(54.44%) of the controls were from rural. Uneducated [AOR:5.18, 95%CI (1.69-15.80)], rural resident [AOR:2.60,95%CI(1.14-6.88)], body mass index(BMI)<18.5kg/m2[AOR:3.11,95%CI(1.41-6.88], pulmonary tuberculosis[AOR:3.98,95%CI(1.11-14.22)], contact history with tuberculosis patient [AOR:3.99,95%CI (1.75-9.07)] and history of previous treatment[AOR:9.5,95%CI(4.08-22)] were found independent determinants of MDR-TB.Conclusions and Recommendation: Uneducated, rural residence, body mass index <18.5kg/m2, pulmonary tuberculosis, contact history with tuberculosis patient and history of previous treatment were associated with MDR-TB. A community level education to enhance public awareness about MDR-TB, nutritional counseling and support, strengthening contact tracing and directly observed treatment strategies with treatment adherence interventions were recommended.