In this study, we enlarged our previous investigation focusing on the biodiversity
of chlamydiae and amoebae in a drinking water treatment plant, by the inclusion of
two additional plants and by searching also for the presence of legionellae and mycobacteria.
Autochthonous amoebae were recovered onto non-nutritive agar, identified by 18S rRNA
gene sequencing, and screened for the presence of bacterial endosymbionts. Bacteria
were also searched for by Acanthamoeba co-culture. From a total of 125 samples, we
recovered 38 amoebae, among which six harboured endosymbionts (three chlamydiae and
three legionellae). In addition, we recovered by amoebal co-culture 11 chlamydiae,
36 legionellae (no L. pneumophila), and 24 mycobacteria (all rapid-growers). Two plants
presented a similar percentage of samples positive for chlamydiae (11%), mycobacteria
(20%) and amoebae (27%), whereas in the third plant the number of recovered bacteria
was almost twice higher. Each plant exhibited a relatively high specific microbiota.
Amoebae were mainly represented by various Naegleria species, Acanthamoeba species
and Hartmannella vermiformis. Parachlamydiaceae were the most abundant chlamydiae
(8 strains in total), and in this study we recovered a new genus-level strain, along
with new chlamydiae previously reported. Similarly, about 66% of the recovered legionellae
and 47% of the isolated mycobacteria could represent new species. Our work highlighted
a high species diversity among legionellae and mycobacteria, dominated by putative
new species, and it confirmed the presence of chlamydiae in these artificial water
systems.