Diguanylate Cyclase Null Mutant Reveals That C-Di-GMP Pathway Regulates the Motility and Adherence of the Extremophile Bacterium Acidithiobacillus caldus

Abstract

An understanding of biofilm formation is relevant to the design of biological strategies to improve the efficiency of the bioleaching process and to prevent environmental damages caused by acid mine/rock drainage. For this reason, our laboratory is focused on the characterization of the molecular mechanisms involved in biofilm formation in different biomining bacteria. In many bacteria, the intracellular levels of c-di-GMP molecules regulate the transition from the motile planktonic state to sessile community-based behaviors, such as biofilm development, through different kinds of effectors. Thus, we recently started a study of the c-di-GMP pathway in several biomining bacteria including Acidithiobacillus caldus. C-di-GMP molecules are synthesized by diguanylate cyclases (DGCs) and degraded by phosphodiesterases (PDEs). We previously reported the existence of intermediates involved in c-di-GMP pathway from different Acidithiobacillus species. Here, we report our work related to At. caldus ATCC 51756. We identified several putative-ORFs encoding DGC and PDE and effector proteins. By using total RNA extracted from At. caldus cells and RT-PCR, we demonstrated that these genes are expressed. We also demonstrated the presence of c-di-GMP by mass spectrometry and showed that genes for several of the DGC enzymes were functional by heterologous genetic complementation in Salmonella enterica serovar Typhimurium mutants. Moreover, we developed a DGC defective mutant strain (Δc1319) that strongly indicated that the c-di-GMP pathway regulates the swarming motility and adherence to sulfur surfaces by At. caldus. Together, our results revealed that At. caldus possesses a functional c-di-GMP pathway which could be significant for ores colonization during the bioleaching process.

Insights into the Quorum Sensing Regulon of the Acidophilic Acidithiobacillus ferrooxidans Revealed by Transcriptomic in the Presence of an Acyl Homoserine Lactone Superagonist Analog

“…These mainly include: (i) induction of the genes encoding the F0-ATPase subunit leading to the PMF allowing AHL efflux and influx, (ii) repression of several genes involved in carbohydrate metabolism to orientate carbon flow to maltodextrin and EPS building block precursor synthesis for adhesion and biofilm formation, respectively; (iii) induction of phosphate and ammonium transporters to anticipate inorganic ion gradient within and around the biofilm structure. Whereas QS and c-di-GMP pathway have been linked in different bacterial species ( Waters et al, 2008 ; Zhang, 2010 ; Kozlova et al, 2011 ; Suppiger et al, 2016 ), it is noteworthy that no change in the transcriptional profiling of the seven genes related to the c-di-GMP pathway in A. ferrooxidans ( Ruiz et al, 2012 ; Castro et al, 2015 ) has been observed in the presence of tetrazole 9c. This result indicates that QS does not modulate c-di-GMP signaling in this Gram-negative species.…”

“…Even if several reports related to biofilm formation regulation by acidophilic bacteria belonging to Acidithiobacillus genus have been released recently ( Farah et al, 2005 ; Bellenberg et al, 2012 , 2014 ; Ruiz et al, 2012 ; Diaz et al, 2013 ; Montgomery et al, 2013 ; Vera et al, 2013 ; Castro et al, 2015 ), the molecular cascade involved in exopolysaccharide production and biofilm formation by Acidithiobacillus species is still undeciphered. While c-di-GMP pathway has been identified in all Acidithiobacillus spp.…”

“…While c-di-GMP pathway has been identified in all Acidithiobacillus spp. ( Ruiz et al, 2012 ; Diaz et al, 2013 ; Castro et al, 2015 ), the species that oxidize only RISC do not possess the genes related to canonical QS systems ( Valdés et al, 2008 ). Indeed, a functional QS system has been reported only in the iron/RISC-oxidizing species A. ferrooxidans ( Farah et al, 2005 ; Rivas et al, 2005 ; Valenzuela et al, 2007 ).…”

The Confluence of Heavy Metal Biooxidation and Heavy Metal Resistance: Implications for Bioleaching by Extreme Thermoacidophiles

“…ferrooxidans and A. ferrooxidans possess chemotaxis systems for sensing Fe(II), which might function to identify specific sites on pyrite surfaces for attachment [308,309]. Quorum sensing functions in bioleaching bacterium allow for swarming behavior on metal sulfides and play a key role in biofilm formation, enhancing dissolution of the mineral substrate [310][311][312][313][314][315]. Early biofilm formation involves capsular polysaccharide production (CPS), up-regulation of genes related to pili and EPS production, motility and quorum sensing, synthesis of cell wall structures, specific proteases, stress response chaperons, and mixed acid fermentation [316][317][318][319].…”

Acidithiobacillus ferrianus sp. nov.: an ancestral extremely acidophilic and facultatively anaerobic chemolithoautotroph

“…For scanning electron microscopy, cells were grown in basal salts/trace elements medium (Ñancucheo et al 2016) containing sulfur as sole electron donor at 30 °C. Sulfur coupons were prepared as described by Castro et al (2015). Samples were critical point dried, coated with gold and observed with a LEO 1420VP scanning electron microscope.…”