Objective To obtain information on the level of exposure and protection of workers to biological agents working in the health sector, farms, meat industry, laboratories, sewage treatment plants, food industry and veterinary centers, valuing their differences in relation to the size of the company. Method A study in which 590 workers belonging to 59 Spanish companies. An initial questionnaire with 3 questions was sent to companies and answered by those responsible for prevention, and another questionnaire with 34 questions to be answered by exposed workers was also sent. Validation was performed by analysis of overall alpha of the second questionnaire to assess the internal consistency and reliability of Cronbach of the questionnaires. Results The final sample included 518 workers in 51 companies in which there was exposure to biological agents. The questionnaire obtained a Cronbach's alpha greater than 0.7. Significant differences in biological risk management work depending on the size of the company (p-value <0.05) were found. Conclusions Our results provide an adequate internal consistency of the questionnaire. Furthermore occupational exposure to biological agents is a factor to be taken into account since it is not fully developed. This is because neither businesses nor prevention professionals have tools available that allow easy implementation, and, therefore, management of overall biological hazards is inadequate.