Quorum sensing (QS) is a bacterial cell-to-cell communication system controlling QS-mediated genes which is synchronized with the population density. The regulation of specific gene activity is dependent on the signaling molecules produced, namely N-acyl homoserine lactones (AHLs). We report here the identification and characterization of AHLs produced by bacterial strain ND07 isolated from a Malaysian fresh water sample. Molecular identification showed that strain ND07 is clustered closely to Pseudomonas cremoricolorata. Spent culture supernatant extract of P. cremoricolorata strain ND07 activated the AHL biosensor Chromobacterium violaceum CV026. Using high resolution triple quadrupole liquid chromatography-mass spectrometry, it was confirmed that P. cremoricolorata strain ND07 produced N-octanoyl-L-homoserine lactone (C8-HSL) and N-decanoyl-L-homoserine lactone (C10-HSL). To the best of our knowledge, this is the first documentation on the production of C10-HSL in P. cremoricolorata strain ND07.
In many species of bacteria, the quorum sensing mechanism is used as a unique communication system which allows them to regulate gene expression and behavior in accordance with their population density. N-Acylhomoserine lactones (AHLs) are known as diffusible autoinducer molecules involved in this communication network. This finding aimed to characterize the production of AHL of a bacterial strain ND04 isolated from a Malaysian waterfall. Strain ND04 was identified as Kluyvera sp. as confirmed by molecular analysis of its 16S ribosomal RNA gene sequence. Kluyvera sp. is closely related to the Enterobacteriaceae family. Chromobacterium violaceum CV026 was used as a biosensor to detect the production of AHL by strain ND04. High resolution triple quadrupole liquid chromatography-mass spectrometry analysis of strain ND04 showed our isolate produced two AHLs which are N-(3-oxohexanoyl)homoserine lactone (3-oxo-C6 HSL) and N-3-oxo-octanoyl-L-homoserine lactone (3-oxo-C8 HSL).
Proteobacteria produce N-acylhomoserine lactones as signaling molecules, which will bind to their cognate receptor and activate quorum sensing-mediated phenotypes in a population-dependent manner. Although quorum sensing signaling molecules can be degraded by bacteria or fungi, there is no reported work on the degradation of such molecules by basidiomycetous yeast. By using a minimal growth medium containing N-3-oxohexanoylhomoserine lactone as the sole source of carbon, a wetland water sample from Malaysia was enriched for microbial strains that can degrade N-acylhomoserine lactones, and consequently, a basidiomycetous yeast strain WW1C was isolated. Morphological phenotype and molecular analyses confirmed that WW1C was a strain of Trichosporon loubieri. We showed that WW1C degraded AHLs with N-acyl side chains ranging from 4 to 10 carbons in length, with or without oxo group substitutions at the C3 position. Re-lactonisation bioassays revealed that WW1C degraded AHLs via a lactonase activity. To the best of our knowledge, this is the first report of degradation of N-acyl-homoserine lactones and utilization of N-3-oxohexanoylhomoserine as carbon and nitrogen source for growth by basidiomycetous yeast from tropical wetland water; and the degradation of bacterial quorum sensing molecules by an eukaryotic yeast.
The antibiotic pyrrolnitrin (PRN) is a tryptophan-derived secondary metabolite that plays an important role in the biocontrol of plant diseases due to its broad-spectrum of antimicrobial activities. The PRN biosynthetic gene cluster remains to be characterised in Serratia plymuthica, though it is highly conserved in PRN-producing bacteria. To better understand PRN biosynthesis and its regulation in Serratia, the prnABCD operon from S. plymuthica G3 was cloned, sequenced and expressed in Escherichia coli DH5α. Furthermore, an engineered strain prnind which is a conditional mutant of G3 prnABCD under the control of the Ptac promoter was constructed. This mutant was able to overproduce PRN with isopropylthiogalactoside (IPTG) induction by overexpressing prnABCD, whilst behaving as a conditional mutant of G3 prnABCD in the absence of IPTG. These results confirmed that prnABCD is responsible for PRN biosynthesis in strain G3. Further experiments involving lux-/dsRed-based promoter fusions, combined with site-directed mutagenesis of the putative σS extended -10 region in the prnA promoter, and liquid chromatography-mass spectrometry (LC-MS) analysis extended our previous knowledge about G3, revealing that quorum sensing (QS) regulates PRN biosynthesis through cross talk with RpoS, which may directly activated prnABCD transcription. These findings suggest that PRN in S. plymuthica G3 is produced in a tightly controlled manner, and has diverse functions, such as modulation of cell motility, in addition to antimicrobial activities. Meanwhile, the construction of inducible mutants could be a powerful tool to improve PRN production, beyond its potential use for the investigation of the biological function of PRN.
Aeromonas is a pathogenic organism that is often found to infect humans. Here we report the draft genome of a clinical isolate in Malaysia, Aeromonas sp. strain 159, which shows N-acylhomoserine lactone production. In the draft genome of strain 159, luxI and luxR homologue genes were found to be located at contig 47, and these genes are believed to be important for the quorum-sensing system present in this pathogen.
Bacterial communication or quorum sensing (QS) is achieved via sensing of QS signaling molecules consisting of oligopeptides in Gram-positive bacteria and N-acyl homoserine lactones (AHL) in most Gram-negative bacteria. In this study, Enterobacteriaceae isolates from Batavia lettuce were screened for AHL production. Enterobacter asburiae, identified by matrix-assisted laser desorption ionization time of flight mass spectrometry (MALDI-TOF-MS) was found to produce short chain AHLs. High resolution triple quadrupole liquid chromatography mass spectrometry (LC/MS) analysis of the E. asburiae spent supernatant confirmed the production of N-butanoyl homoserine lactone (C4-HSL) and N-hexanoyl homoserine lactone (C6-HSL). To the best of our knowledge, this is the first report of AHL production by E. asburiae.
We report the degradation of quorum sensing N-acylhomoserine lactone molecules by a bacterium isolated from a Malaysian marine water sample. MALDI-TOF and phylogenetic analysis indicated this isolate BM1 clustered closely to Labrenzia sp. The quorum quenching activity of this isolate was confirmed by using a series of bioassays and rapid resolution liquid chromatography analysis. Labrenzia sp. degraded a wide range of N-acylhomoserine lactones namely N-(3-hexanoyl)-L-homoserine lactone (C6-HSL), N-(3-oxohexanoyl)-L-homoserine lactone (3-oxo-C6-HSL) and N-(3-hydroxyhexanoyl)-L-homoserine lactone (3-hydroxy-C6-HSL). Re-lactonisation bioassays confirmed Labrenzia sp. BM1 degraded these signalling molecules efficiently via lactonase activity. To the best of our knowledge, this is the first documentation of a Labrenzia sp. capable of degrading N-acylhomoserine lactones and confirmation of its lactonase-based mechanism of action.
One obvious requirement for concerted action by a bacterial population is for an individual to be aware of and respond to the other individuals of the same species in order to form a response in unison. The term "quorum sensing" (QS) was coined to describe bacterial communication that is able to stimulate expression of a series of genes when the concentration of the signaling molecules has reached a threshold level. Here we report the isolation from aquatic environment of a bacterium that was later identified as Enterobacter sp.. Chromobacterium violaceum CV026 and Escherichia coli [pSB401] were used for preliminary screening of N-acyl homoserine lactone (AHL) production. The Enterobacter sp. isolated was shown to produce two types of AHLs as confirmed by analysis using high resolution tandem mass spectrometry. To the best of our knowledge, this is the first documentation of an Enterobacter sp. that produced both 3-oxo-C6-HSL and 3-oxo-C8-HSL as QS signaling molecules.
N-acylhomoserine lactones (AHL) plays roles as signal molecules in quorum sensing (QS) in most Gram-negative bacteria. QS regulates various physiological activities in relation with population density and concentration of signal molecules. With the aim of isolating marine water-borne bacteria that possess QS properties, we report here the preliminary screening of marine bacteria for AHL production using Chromobacterium violaceum CV026 as the AHL biosensor. Strain T33 was isolated based on preliminary AHL screening and further identified by using 16S rDNA sequence analysis as a member of the genus Vibrio closely related to Vibrio brasiliensis. The isolated Vibrio sp. strain T33 was confirmed to produce N-hexanoyl-L-homoserine lactone (C6-HSL) and N-(3-oxodecanoyl)-L-homoserine lactone (3-oxo-C10 HSL) through high resolution tandem mass spectrometry analysis. We demonstrated that this isolate formed biofilms which could be inhibited by catechin. To the best of our knowledge, this is the first report that documents the production of these AHLs by Vibrio brasiliensis strain T33.
A multidrug-resistant clinical bacteria strain GB11 was isolated from a wound swab on the leg of a patient. Identity of stain GB11 as Pseudomonas aeruginosa was validated by using matrix-assisted laser desorption/ionization-time-of-flight mass spectrometry (MALDI-TOF MS). Detection of the production of signaling molecules, N-acylhomoserine lactones (AHLs), was conducted using three different bacterial biosensors. A total of four different AHLs were found to be produced by strain GB11, namely N-butyryl homoserine lactone (C4-HSL), N-hexanoylhomoserine lactone (C6-HSL), N-octanoyl homoserine lactone (C8-HSL) and N-3-oxo-dodecanoylhomoserine lactone (3-oxo-C12-HSL) using high resolution liquid chromatography tandem mass spectrometry (LC-MS/MS). Of these detected AHLs, 3-oxo-C12-HSL was found to be the most abundant AHL produced by P. aeruginosa GB11.
Proteobacteria use quorum sensing to regulate target gene expression in response to population density. Quorum sensing (QS) is achieved via so-called signalling molecules and the best-studied QS signalling system uses N-acyl homoserine lactones (AHLs). This study aimed to identify and characterize the production of AHLs by a bacterium ND03 isolated from a Malaysian tropical rainforest waterfall. Molecular identification showed that ND03 is a Pantoea sp. closely related to Pantoea rodasii. We used Chromobacterium violaceum CV026, an AHL biosensor for preliminary AHL production screening and then used high resolution triple quadrupole liquid chromatography-mass spectrometry, to confirm that P. rodasii strain ND03 produced N-(3-oxo-hexanoyl)-L-homoserine lactone (3-oxo-C6-HSL). To the best of our knowledge, this is the first report for such a discovery in P. rodasii strain ND03.
Strain RB38 was recovered from a former dumping area in Malaysia. MALDI-TOF mass spectrometry and genomic analysis identified strain RB-38 as Pandoraea pnomenusa. Various biosensors confirmed its quorum sensing properties. High resolution triple quadrupole liquid chromatography-mass spectrometry analysis was subsequently used to characterize the N-acyl homoserine lactone production profile of P. pnomenusa strain RB38, which validated that this isolate produced N-octanoyl homoserine lactone as a quorum sensing molecule. This is the first report of the production of N-octanoyl homoserine lactone by P. pnomenusa strain RB38.
Bacterial cell-to-cell communication (quorum sensing) refers to the regulation of bacterial gene expression in response to changes in microbial population density. Quorum sensing bacteria produce, release and respond to chemical signal molecules called autoinducers. Bacteria use two types of autoinducers, namely autoinducer-1 (AI-1) and autoinducer-2 (AI-2) where the former are N-acylhomoserine lactones and the latter is a product of the luxS gene. Most of the reported literatures show that the majority of oral bacteria use AI-2 for quorum sensing but rarely the AI-1 system. Here we report the isolation of Pseudomonas putida strain T2-2 from the oral cavity. Using high resolution mass spectrometry, it is shown that this isolate produced N-octanoylhomoserine lactone (C8-HSL) and N-dodecanoylhomoserine lactone (C12-HSL) molecules. This is the first report of the finding of quorum sensing of P. putida strain T2-2 isolated from the human tongue surface and their quorum sensing molecules were identified.
In the bacteria kingdom, quorum sensing (QS) is a cell-to-cell communication that relies on the production of and response to specific signaling molecules. In proteobacteria, N-acylhomoserine lactones (AHLs) are the well-studied signaling molecules. The present study aimed to characterize the production of AHL of a bacterial strain A9 isolated from a Malaysian tropical soil. Strain A9 was identified as Burkholderia sp. using matrix-assisted laser desorption ionization-time-of-flight mass spectrometry and 16S rDNA nucleotide sequence analysis. AHL production by A9 was detected with two biosensors, namely Chromobacterium violaceum CV026 and Escherichia coli [pSB401]. Thin layer chromatography results showed N-hexanoylhomoserine lactone (C6-HSL) and N-octanoylhomoserine lactone (C8-HSL) production. Unequivocal identification of C6-HSL and C8-HSL was achieved by high resolution triple quadrupole liquid chromatography-mass spectrometry analysis. We have demonstrated that Burkholderia sp. strain A9 produces AHLs that are known to be produced by other Burkholderia spp. with CepI/CepR homologs.
Proteobacteria are known to communicate via signaling molecules and this process is known as quorum sensing. The most commonly studied quorum sensing molecules are N-acylhomoserine lactones (AHLs) that consists of a homoserine lactone moiety and an N-acyl side chain with various chain lengths and degrees of saturation at the C-3 position. We have isolated a bacterium, RB-44, from a site which was formally a landfill dumping ground. Using matrix-assisted laser desorption ionization time-of-flight (MALDI-TOF) mass spectrometry analysis, this isolate was identified as a Pandoraea sp.which was then screened for AHL production using biosensors which indicated its quorum sensing properties. To identify the AHL profile of Pandoraea sp. RB-44, we used high resolution tandem mass spectrometry confirming that this isolate produced N-octanoylhomoserine lactone (C8-HSL). To the best of our knowledge, this is the first report that showed quorum sensing activity exhibited by Pandoraea sp. Our data add Pandoraea sp. to the growing number of bacteria that possess QS systems.
A chemically defined medium called KGm medium was used to isolate from a sample of sea water a bacterial strain, MW3A, capable of using N-3-oxohexanoyl-L: -homoserine lactone as the sole carbon source. MW3A was clustered closely to Pseudomonas aeruginosa by 16S ribosomal DNA sequence analysis. It degraded both N-acylhomoserine lactones (AHLs) with a 3-oxo group substitution and, less preferably, AHLs with unsubstituted groups at C3 position in the acyl side chain, as determined by Rapid Resolution Liquid Chromatography. Its quiP and pvdQ homologue gene sequences showed high similarities to those of known acylases. Spent supernatant of MW3A harvested at 8-h post inoculation was shown to contain long-chain AHLs when assayed with the biosensor Escherichia coli [pSB1075], and specifically N-dodecanoyl-L: -homoserine lactone and N-3-oxotetradecanoyl-L: -homoserine lactone by high resolution mass spectrometry. Hence, we report here a novel marine P. aeruginosa strain MW3A possessing both quorum-quenching and quorum-sensing properties.
Dyella japonica strain A8 is a Malaysian tropical soil bacterial strain which shows N-acylhomoserine lactone-degrading activity. Here, we present its draft genome sequence. A putative quorum-quenching gene was identified based on the genome sequence analysis of strain A8. To the best of our knowledge, this is the first genome announcement of a member from the genus of Dyella, and this is also the first work that reports the quorum-quenching activity of Dyella japonica.
N-Acyl homoserine lactone (AHL) serves as signaling molecule for quorum sensing (QS) in Gram-negative bacteria to regulate various physiological activities including pathogenicity. With the aim of isolating freshwater-borne bacteria that can cause outbreak of disease in plants and portrayed QS properties, environmental water sampling was conducted. Here we report the preliminary screening of AHL production using Chromobacterium violaceum CV026 and Escherichia coli [pSB401] as AHL biosensors. The 16S rDNA gene sequence of isolate M009 showed the highest sequence similarity to Pantoea stewartii S9-116, which is a plant pathogen. The isolated Pantoea sp. was confirmed to produce N-3-oxohexanoyl-L-HSL (3-oxo-C6-HSL) through analysis of high resolution mass tandem mass spectrometry.
Staphylococcus aureus is a Gram-positive pathogen that causes potentially life-threatening nosocomial- and community-acquired infections, such as osteomyelitis and endocarditis. Staphylococcus aureus has the ability to form multicellular, surface-adherent communities called biofilms, which enables it to survive in various sources of stress, including antibiotics, nutrient limitations, heat shock, and immune responses. Biofilm-forming capacity is now recognized as an important virulence determinant in the development of staphylococcal device-related infections. In light of the projected increase in the numbers of elderly patients who will require semi-permanent indwelling medical devices such as artificial knees and hips, we can anticipate an expanded need for new agents and treatment options to manage biofilm-associated infections in an expanding at-risk population. With better understanding of staphylococcal biofilm formation and growth, novel strategies that target biofilm-associated infections caused by S. aureus have recently been described and seem promising as future anti-biofilm therapies.