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Dairy cow mastitis is an important disease in the dairy industry. Different microbial species have been identified as causative agents in mastitis, and are traditionally diagnosed by bacterial culture. The objective of this study was to use metagenomic pyrosequencing of bacterial 16S rRNA genes to investigate bacterial DNA diversity in milk samples of mastitic and healthy dairy cows and compare the results with those obtained by classical bacterial culture. One hundred and thirty-six milk samples were collected from cows showing signs of mastitis and used for microbiological culture. Additionally, 20 milk samples were collected from healthy quarters. Bacterial DNA was isolated from the same milk samples and the 16S rRNA genes were individually amplified and pyrosequenced. Discriminant analysis showed that the groups of samples that were most clearly different from the rest and thus easily discriminated were the normal milk samples from healthy cows and those characterised by culture as  Trueperella pyogenes and Streptococcus  spp. The mastitis pathogens identified by culture were generally among the most frequent organisms detected by pyrosequencing, and in some cases ( Escherichia coli ,  Klebsiella  spp. and  Streptococcus uberis  mastitis) the single most prevalent microorganism.  Trueperella pyogenes  sequences were the second most prevalent sequences in mastitis cases diagnosed as  Trueperella pyogenes  by culture,  Streptococcus dysgalactiae  sequences were the second most prevalent sequences in mastitis cases diagnosed as  Streptococcus dysgalactiae  by culture, and  Staphyloccocus aureus  sequences were the third most prevalent in mastitis cases diagnosed as  Staphylococcus aureus  by culture. In samples that were aerobic culture negative, pyrosequencing identified DNA of bacteria that are known to cause mastitis, DNA of bacteria that are known pathogens but have so far not been associated with mastitis, and DNA of bacteria that are currently not known to be pathogens. A possible role of anaerobic pathogens in bovine mastitis is also suggested.

Microbial Diversity of Bovine Mastitic Milk as Described by Pyrosequencing of Metagenomic 16s rDNA