Novel Chlamydiales strains isolated from a water treatment plant

Abstract

Chlamydiae are obligate intracellular bacteria infecting free-living amoebae, vertebrates and some invertebrates. Novel members are regularly discovered, and there is accumulating evidence supporting a very important diversity of chlamydiae in the environment. In this study, we investigated the presence of chlamydiae in a drinking water treatment plant. Samples were used to inoculate Acanthamoeba monolayers (Acanthamoeba co-culture), and to recover autochthonous amoebae onto non-nutritive agar. Chlamydiae were searched for by a pan-chlamydia 16S rRNA gene PCR from both Acanthamoeba co-cultures and autochthonous amoebae, and phylotypes determined by 16S rRNA gene sequencing. Autochthonous amoebae also were identified by 18S rRNA gene amplification and sequencing. From a total of 79 samples, we recovered eight chlamydial strains by Acanthamoeba co-culture, but only one of 28 amoebae harboured a chlamydia. Sequencing results and phylogenetic analysis showed our strains belonging to four distinct chlamydial lineages. Four strains, including the strain recovered within its natural host, belonged to the Parachlamydiaceae; two closely related strains belonged to the Criblamydiaceae; two distinct strains clustered with Rhabdochlamydia spp.; one strain clustered only with uncultured environmental clones. Our results confirmed the usefulness of amoeba co-culture to recover novel chlamydial strains from complex samples and demonstrated the huge diversity of chlamydiae in the environment, by identifying several new species including one representing the first strain of a new family.

Enhanced survival but not amplification of Francisella spp. in the presence of free-living amoebae

“…The isolation and identification of amoebae-resisting bacteria (ARB) using the well-established co-culture method involves inoculation of samples onto a monolayer of axenic amoebae and continuous monitoring of cultures for amoebal lysis, resulting from infection by ARBs (reviewed in [46,47]). From soil, drinking water, ground and surface water, and clinical samples, the amoebal co-culture method, with amoebal lysis as an assay endpoint, has been used by numerous groups to identify human pathogens such as those belonging to the class Chlamydiae and the genera Aeromonas, Bacillus, Enterobacter, Legionella, Mycobacterium, Pseudomonas, Rhodococcus, and Streptococcus as well as novel bacteria belonging to the genera Afipia and Bosea [48][49][50][51][52][53][54][55][56]. Thus, the absence of amoebal lysis observed in this study between each FLA and Francisella spp.…”

Development of a New Chlamydiales-Specific Real-Time PCR and Its Application to Respiratory Clinical Samples

“…A percentage of similarity for the best BLAST of Ͼ90% was observed for all 48 sequences obtained, allowing identification at least at the family level. Of the 48 sequences obtained, 26 belonged to the Parachlamydiaceae family, 7 belonged to the Chlamydiaceae family, 5 belonged to the Simkaniaceae family, 5 belonged to the Criblamydiaceae family, 3 belonged to the Rhabdochlamydiaceae family, 1 seemed to belong to the novel E6 lineage (7,11), and 1 other sequence corresponded to an unclassified Chlamydiales (see Table S2 in the supplemental material). Of the 7 sequences corresponding to a Chlamydiaceae species, 6 demonstrated 100% similarity with Chlamydia pneumoniae: five samples were taken from children with pneumonia (Table 5), whereas one was taken from an apparently healthy child (see Table S2 in the supplemental material).…”

Prevalence study of Simkania negevensis in cooling towers in Spain

“…negevensis has also been considered as the etiological agent for several respiratory tract infections in humans, including bronchiolitis in infants (Kahane et al 1998) and community-acquired pneumonia and acute exacerbation of chronic obstructive pulmonary disease in both adults and children (Lieberman et al 1997(Lieberman et al , 2002Petrich et al 2002;Greenberg et al 2003;Fasoli et al 2008). It is important to note that the determination of the prevalence of Parachlamydia and Chlamydia-like organisms has recently been the subject of several studies, in part as a consequence of its association with lung infection (Casson et al 2008;Corsaro et al 2009;Greub 2009). …”