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Bacteria of the genus  Legionella  cause water-based infections resulting in severe pneumonia. Here we analyze and compare the bacterial microbiome of sputum samples from pneumonia patients in relation to the presence and abundance of the genus  Legionella . The prevalence of  Legionella  species was determined by culture, PCR, and Next Generation Sequencing (NGS). Nine sputum samples out of the 133 analyzed were PCR-positive using  Legionella  genus-specific primers. Only one sample was positive by culture. Illumina MiSeq 16S rRNA gene sequencing analyses of  Legionella- positive and  Legionella- negative sputum samples, confirmed that indeed,  Legionella  was present in the PCR-positive sputum samples. This approach allowed the identification of the sputum microbiome at the genus level, and for  Legionella  genus at the species and sub-species level. 42% of the sputum samples were dominated by  Streptococcus. Legionella  was never the dominating genus and was always accompanied by other respiratory pathogens. Interestingly, sputum samples that were  Legionella  positive were inhabited by aquatic bacteria that have been observed in an association with amoeba, indicating that amoeba might have transferred  Legionella  from the drinking water together with its microbiome. This is the first study that demonstrates the sputum major bacterial commensals and pathogens profiles with regard to  Legionella  presence.

Comparison of sputum microbiome of legionellosis-associated patients and other pneumonia patients: indications for polybacterial infections