Atmospheric air is a microbial habitat of pathogenic bioaerosols that may pose serious risks to humans. A commonly laboratory-based approach for the diagnosis of such infections in the bloodstream is the blood culture analysis. Its clinical relevance is attributed to the fact that these infections are characterized by high rates of morbidity and mortality, requiring the need for efficient methods for rapid diagnosis. For this reason, our study aimed to develop a method of manometric monitoring for the rapid detection of viable microorganisms in blood culture vials. A methodology was developed to detect pressure variation in intra-vials through a manometric instrument that was coupled to vials of blood culture containing culture broth that allowed microbial growth. This device allowed the early detection of microbial activity based on the production or use of intra-flask gases as a result of microbial metabolic activity. The analyzed variables were the pressure as a function of time, microbial species, and culture medium. The highest pressure found in the flasks without microorganisms was 40 mmHg between 2 and 6 h, and the lowest pressure was −42 mmHg between 21 and 24 h. The variation of the internal pressure in blood culture flasks according to different groups of microorganisms as a function of time demonstrated that the fermentative gram-negative bacilli and gram-positive cocci exhibited a significant increase in relation to their respective control groups (p < 0.001). The non-fermenting gram-negative bacilli showed expected results in relation to the pressure variation in which the production of negative pressures was noticed during the period of analysis, with a significant difference with respect to their control groups (p < 0.001). The developed methodology for the early detection of microorganisms responsible for bloodstream infection was demonstrated to be effective.