It is possible that the loss of H. pylori cultivability when associated with heterotrophic biofilms had been due to a #4SC-202 concentration randurls[1|1|,|CHEM1|]# negative effect caused by the presence of
other microorganisms [31]. Nevertheless, it is also possible that there were other microorganisms present in the biofilm that could have a beneficial effect on L. pneumophila or H. pylori, as shown by other studies where these pathogens were co-cultured with other microorganisms in liquid media [32, 33]. However, for multi-species biofilms it is technically very challenging to determine which sessile microorganisms could have a positive or negative effect on these pathogens, particularly regarding the intimate associations that occur within biofilms. A particular type of interaction that can facilitate the formation of biofilm is the aggregation of cells, which can occur between cells of the same species (auto-aggregation) or between different species (co-aggregation), and has been well described for isolates of dental plaque species in complex media and aquatic species in potable water [34–36]. The aim
of this work was to study the influence of different autochthonous microorganisms isolated from drinking water biofilms on the incorporation and survival of L. pneumophila and H. pylori in biofilms. For that, the first part of the work tested all the species used for auto and co-aggregation. Subsequently, dual-species biofilms of L. pneumophila and H. pylori were formed selleck compound with the different drinking water bacteria and the results compared with mono-species biofilms formed by L. pneumophila Acyl CoA dehydrogenase and H. pylori. Results Auto and co-aggregation of L. pneumophila and other drinking water bacteria Initially, the selected biofilm strains were tested for auto- and co-aggregation in test tubes as described by Rickard et
al. [35], either alone or with L. pneumophila. No co-aggregation was observed for the strains studied, either alone or in pairs with L. pneumophila (results not shown). L. pneumophila in biofilms For the experiments on biofilm formation on uPVC coupons, an inoculum of L. pneumophila was prepared containing approximately 3.7 × 107 of total cells ml-1 (quantified using SYTO 9 staining). In comparison to total cells, 49% were cultivable on BCYE agar and 50% were detected by PNA-FISH. The inocula of the strains isolated from drinking water biofilms had on average 75% of cultivable cells compared to SYTO 9 stained cells, except in the case of Mycobacterium chelonae where the percentage was considerably lower (2.5%). Figure 1a shows the variation with time of total cells, PNA-cells and cultivable L. pneumophila present in a mono-species biofilm. The attachment of L. pneumophila cells to the surface occurred in the first 24 hours of the experiment. Moreover, the numbers of total cells (stained by SYTO 9) and PNA stained cells did not change significantly between days 1 and 32 (P > 0.05).