J Microbiol Methods 2006, 65:194–201.PubMedCrossRef 75. Amann RI, Binder BJ, ML323 manufacturer Olson RJ, Chisholm

SW, Devereux R, Stahl DA: Combination of 16S ribosomal-RNA-targeted oligonucleotide probes with flow-cytometry for analyzing mixed microbial-populations. Appl Environ Microbiol 1990, 56:1919–1925.PubMed Authors’ contributions NJF, MH and BMW conceived and designed the study. NJF and BMW collected samples. NJF carried out the experiments, evaluated the results and drafted the manuscript. BMW and MH provided guidance during the whole study and revised the manuscript. All authors read and approved the final manuscript.”
“Background Klebsiella pneumoniae, an opportunistic pathogen responsible for a wide range of nosocomial infections that include pneumonia, bacteremia and urinary tract infections, is estimated to cause approximately 8% of hospital acquired infections [1–5]. This Gram-negative bacterium can also be found in the environment

in association with plants, as well as in soil and in water [2, 6]. One important factor associated with virulence in K. pneumoniae is its capacity to adhere to surfaces and form biofilms. Although the formation of biofilms by ATM inhibitor K. pneumoniae is still not fully understood, several key determinants have been identified such as pili, polysaccharides, quorum sensing and transport and regulatory proteins [7–13]. More recently, it has been shown that c-di-GMP controls type 3 fimbria expression and biofilm formation in K. pneumoniae by binding to and modulating the activity of the transcriptional regulator MrkH [14,

15]. The second messenger c-di-GMP is known to play a key role in several cellular functions as well as in biofilm formation in bacteria where it modulates the transition between planktonic and sessile lifestyles. Low levels of c-di-GMP result in increased motility Dynein while high levels promote adhesion to surfaces, production of exopolysaccharides and biofilm formation [16, 17]. The intracellular levels of c-di-GMP are regulated by the C188-9 concentration antagonistic activity of diguanylate cyclase (DGC) enzymes and phosphodiesterases (PDEs) that catalyze synthesis and hydrolysis of this molecule, respectively [16, 18]. Several genetic and biochemical studies have shown that besides their C-terminal catalytically active A site, most of these proteins harbor N-terminal sensory domains that can respond to different internal and external signals, triggering activation of DGCs or PDEs. When enough c-di-GMP is available, it binds different effector molecules, proteins or RNAs, which influence cell behavior [18]. The active site of DGCs contains a conserved GGDEF domain, characterized by the GG(D/E)EF motif, while PDE activity is associated with C-terminal EAL or HD-GYP domains [16, 17]. These domains can be found separately or together, forming hybrid proteins that have both GGDEF and EAL domains.