For a study, it was determined that during 96 hours of refrigerated storage in a clinical environment, microbial development in unfortified and fortified Holder pasteurized donor human milk (HPDHM) was assessed. To imitate a real-life feeding situation, 36 unfortified HPDHM samples and 77 fortified HPDHM samples were made in a neonatal intensive care milk preparation room and kept at 4°C in the NICU refrigerator. At 24-hour intervals, one-milliliter aliquots were extracted and grown in duplicate on a solid blood agar medium for bacterial growth. Standard microbiological procedures were used to identify viable bacterial colonies.

In a vertical laminar flow hood, 96.5% of milk samples tested negative for bacterial growth. The greatest growth was one colony-forming unit/0.1 ml plated in the remaining 3.5% of the samples. When the laminar hood was not employed, colony numbers were higher. In every case, the colonies were common skin bacteria that grew in an erratic and unsustainable manner. Increased bacterial growth was not significantly linked with fortifier quality or storage period (P>0.05). In NICU settings, HPDHM, both unfortified and fortified, were essentially free of bacterial development for up to 96 hours of refrigerated storage. Microbial contamination was avoided by using proper sample-handling practices.