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This study analyzes the bacteriological quality of breast milk samples destined to direct milk donation to preterm infants under 34 Gestational Weeks (GW) hospitalized in a neonatology and a neonatal intensive care unit of a French university hospital. All samples of breast milk destined to direct milk donation between April 2007 and December 2016 were included. A sample was defined as compliant if its total flora was less than 106 Colony Forming Units per milliliter (CFU/mL) and in the absence of Staphylococcus aureus and other pathogens. A total of 777 samples were taken from 629 mothers. The overall non-compliance rate for the initial sample was 21.3%; 63 samples (10.0%) had a total flora ≥ 106 CFU/mL, 63 (10.0%) were contaminated by a pathogenic bacteria and 8 (1.3%) were non-compliant because of both. An increase of the non-compliance rate was observed between 2008 and 2016 (10.2%-26.1%). The increase of the total flora non-compliance rate began in 2011, in link with the doubling of the number of samples taken, to reach a peak in 2013 then decreased in link with development of portable pump. No statistically significant difference of the presence of S. aureus in breast milk was observed. For the other pathogenic bacteria, the rate increased significantly in 2014. The increase of the non-compliance rate could be explained by a decrease of best practices in milk collection. Education of mothers should be strengthened. [ABSTRACT FROM AUTHOR]