Comparative analysis of ionic strength tolerance between freshwater and marine Caulobacterales adhesins

Abstract

17Bacterial adhesion is affected by environmental factors, such as ionic strength, pH, 18 temperature, and shear forces, and therefore marine bacteria must have developed holdfasts 19 with different composition and structures than their freshwater counterparts to adapt to their 20 natural environment. The dimorphic a-proteobacterium Hirschia baltica is a marine budding 21 bacterium in the Caulobacterales clade. H. baltica uses a polar adhesin, the holdfast, located at 22 the cell pole opposite the reproductive stalk for surface attachment and cell-cell adhesion. The 23 holdfast adhesin has been best characterized in Caulobacter crescentus, a freshwater member 24 of the Caulobacterales, and little is known about holdfast composition and properties in marine 25 Caulobacterales. Here we use H. baltica as a model to characterize holdfast properties in 26 marine Caulobacterales. We show that freshwater and marine Caulobacterales use similar 27 genes in holdfast biogenesis and that these genes are highly conserved among the two genera. 28We also determine that H. baltica produces larger holdfast than C. crescentus and that those 29 holdfasts have a different chemical composition, as they contain N-acetylglucosamine and 30 galactose monosaccharide residues and proteins, but lack DNA. Finally, we show that H. baltica 31 holdfasts tolerate higher ionic strength than those of C. crescentus. We conclude that marine 32 Caulobacterales holdfasts have physicochemical properties that maximize binding in high ionic 33 strength environments. 34 35 IMPORTANCE 36 Most bacteria spend a large amount of their lifespan attached to surfaces, forming 37 complex multicellular communities called biofilms. Bacteria can colonize virtually any surface, 38 therefore they have adapted to bind efficiently in very different environments. In this study, we 39 compare the adhesive holdfasts produced by the freshwater bacterium C. crescentus and a 40 relative, the marine bacterium H. baltica. We show that H. baltica holdfasts have a different 41 3 morphology and chemical composition, and tolerate high ionic strength. Our results show that H. 42 baltica holdfast is an excellent model to study the effect of ionic strength on adhesion and 43 providing insights on the physicochemical properties required for adhesion in the marine 44 environment. 45 46 48 communities, known as biofilms (1). To irreversibly adhere to surfaces and form these complex 49 mutil-cellular communities, bacteria produce strong adhesins, mainly composed of proteins or 50 polysaccharides (2, 3). Bacterial adhesion is affected by different environmental conditions such 51 as pH, temperature, shear forces, and ionic strength (2,(4)(5)(6). In marine environments, bacteria 52 face 500 times higher ionic strength than in freshwater (7), therefore, marine bacteria have 53 evolved ways to overcome the effect of ionic strength and bind permanently to surfaces in high 54 salt environments such as seas and oceans. 55Caulobacterales are Alphaproteobacteria found in various habitats, from olig...

A polysaccharide deacetylase enhances bacterial adhesion in high ionic strength environments

“…26,27 In C. crescentus, deacetylation of holdfast polysaccharides is important for both the cohesiveness and adhesiveness of holdfast. 27 In our previous work comparing H. baltica and C. crescentus holdfasts, 25 we showed that both species use similar genes to synthesize, export, and anchor holdfast to the cell envelope. We identified H. baltica genes that modify holdfast in C. crescentus, namely the putative acetyltransferase hfsK (hbal_0069) and the polysaccharide deacetylase hfsH (hbal_1965; Fig.…”

“…As holdfast is required for biofilm formation in C. crescentus and H. baltica, 20,25,31 we probed for the presence of holdfasts using fluorescent Alexa Fluor 488 (AF488) conjugated wheat germ agglutinin (WGA) lectin that specifically binds to the GlcNAc component of the holdfast polysaccharide. 20 In exponentially growing planktonic cultures, C. crescentus WT cells produced holdfasts that bound AF488-WGA and formed cell-cell aggregates mediated by holdfasts, called rosettes (Fig.…”

“…The above results indicate that HfsH plays an important role in H. baltica holdfast properties, including their anchoring to the cell envelope. We hypothesized that (1) H. baltica hfsH produces a small amount of holdfast polysaccharide that is insufficient to anchor the cell to the surface, similarly to the under-expression of a glycosyltransferase HfsL, 25 or (2) holdfasts produced by H. baltica hfsH are deficient in adhesion and/or cohesion and are thus dispersed into the medium.…”

“…In order to understand the role of HfsH in H. baltica, we investigated whether varying the level of its expression in H. baltica affects holdfast cohesive and adhesive properties. We used a copper inducible promoter to tightly control the level of hfsH expression 25 (Fig. 4A).…”

Substratum‐associated microbiota

“…Hentchel et al (2019) grew axenic cultures of C. crescentus in complex and chemically defined media to examine the fitness of transposon mutants in freshwater environments and described a new model system to study gene function in ecosystems. In a comparison of the physicochemical properties of adherence between freshwater and marine Caulobacterales , Chepkwony, Berne, and Brun (2019) found the marine Hirschia baltica maximized binding via their holdfast in environments with high ionic strength. Caulobacter crescentus shifts cell morphology in response to changing growth conditions (Heinrich, Leslie, Morlock, Bertilsson, & Jonas, 2019).…”