Background: Needlesticks and sharp injuries are occupational hazards for healthcare workers that result from the accidental piercing of the skin. Needlestick injuries expose healthcare workers to blood and body fluids that may be infected and can be transmitted to them. Healthcare workers have been exposed to blood-borne pathogens through contaminated needles and other sharp materials every day. Around 20 blood-borne diseases can be transmitted through casual needlesticks and sharp injuries. Objective: To assess needlestick and sharp injuries and its associated factors among healthcare workers in Southern Ethiopia, 2021. Methods: The hospital-based cross-sectional study design was conducted among 341 healthcare workers in Worabe Comprehensive Specialized Hospital from June 10 to July 6, 2021. A stratified sampling technique was used and data were collected using standardized structural questionnaires by BSc nursing professionals. The collected data were checked for completeness and consistency by the investigator. The completed questionnaire was given an identification number and entered into EpiData version 3.5.1. The data were coded and analyzed using SPSS version 26 using a binary logistic regression model and presented with texts, tables, and graphs. Results: The finding revealed that 30.6% of healthcare workers had experienced needlestick and sharp injuries within their working area. Healthcare workers not trained on safety measures of needlestick and sharp injury (adjusted odds ratio: 7.179 (3.494–14.749)), working in the delivery unit (adjusted odds ratio: 6.528 (3.171–11.834)), being older age (adjusted odds ratio: 3.394 (1.775–7.126)), working in inpatient unit (3.278 (1.804–5.231)), working in an emergency unit (adjusted odds ratio: 5.718 (4.326–6.398)), working in an operation room theater (adjusted odds ratio: 2.359 (1.781–4.430)), working as a medical laboratory technician (adjusted odds ratio: 1.070 (1.432–3.304)), working in pediatrics unit (adjusted odds ratio: 1.063 (1.431–2.843)), working as cleaners (adjusted odds ratio: 0.018 (0.002–0.195)), working <40 h per week (adjusted odds ratio: 0.036 (0.004–0.345)), and seldom needle recapping (adjusted odds ratio: 0.043 (0.015–0.125)) were statistically associated with needlestick and sharp injury. Conclusions: In this study, there is a high magnitude of needlestick or sharp injuries among healthcare workers. Lack of training on work-related safety measures; working in delivery; being older age; working in the inpatient unit, emergency, operation room, and pediatrics units; being laboratory technicians, and cleaners; working hours per week; and seldom needle recapping were significant predictors of needlestick and sharp injury.