Viable Spore-Forming Obligate Anaerobes Are Rare in Pasteurized Donor Human Milk: A Pilot Study

Background: When access to mother's own milk is limited, pasteurized donor human milk (PDHM) is the best alternative source of nutrition for high-risk preterm infants. Microbial screening of PDHM is essential to ensure its safety, as spore-forming bacteria may survive pasteurization. Standard screening will detect spore-forming bacteria that grow aerobically, such as Bacillus cereus, but may miss obligate anaerobes, such as Clostridium species. Although milk banking guidelines globally recommend microbial screening of milk batches, they do not specifically recommend anaerobic testing. This study aimed to determine the proportion of PDHM batches containing viable anaerobic bacteria after pasteurization. Materials and Methods: In this prospective cohort study, 150 batches of PDHM from unique donors were sampled (August-December 2024) at Australian Red Cross Lifeblood and tested at an accredited food safety laboratory. Prepasteurization samples were tested for aerobic bacteria, and postpasteurization samples were tested for both aerobic and anaerobic bacteria using a validated method (detection limit ≤1 CFU/mL). Results: No bacteria were recovered from any of the 150 postpasteurization samples tested using an anaerobic culture method. Using standard aerobic culture, 4.7% (7/150) of samples failed prepasteurization microbial screening according to local acceptance guidelines, due to a total colony count ≥ 10⁵ CFU/mL (n = 6) and/or the presence of Enterobacteriaceae ≥ 10⁴ CFU/mL (n = 3), and none failed postpasteurization testing. Conclusions: This study confirmed that obligate anaerobic bacteria are rarely cultured from PDHM. Additional process control through routine testing for anaerobes in PDHM is therefore not considered essential, particularly when PDHM is stored frozen (<18°C) after pasteurization. Clinicians should maintain vigilance for potential recipient adverse events and promptly report these to the source milk bank.