Signal Autoinducer-2 Research Articles

Atlantic Salmon Mucins Inhibit LuxS-Dependent A. Salmonicida AI-2 Quorum Sensing in an N-Acetylneuraminic Acid-Dependent Manner.

One of the most important bacterial diseases in salmonid aquaculture is furunculosis, caused by Aeromonas salmonicida. Bacterial communication through secreted autoinducer signals, quorum sensing, takes part in the regulation of gene expression in bacteria, influencing growth and virulence. The skin and mucosal surfaces, covered by a mucus layer, are the first point of contact between fish and bacteria. Mucins are highly glycosylated and are the main components of mucus. Here, we validate the Vibrio harveyi BB170 bioreporter assay for quantifying A. salmonicida quorum sensing and study the effects of Atlantic salmon mucins as well as mono- and disaccharides on the AI-2 levels of A. salmonicida. Atlantic salmon mucins from skin, pyloric ceca, proximal and distal intestine reduced A. salmonicida AI-2 levels. Among the saccharides abundant on mucins, fucose, N-acetylneuraminic acid and GlcNAcβ1-3Gal inhibited AI-2 A. salmonicida secretion. Removal of N-acetylneuraminic acid, which is the most abundant terminal residue on mucin glycans on Atlantic salmon mucins, attenuated the inhibitory effects on AI-2 levels of A. salmonicida. Deletion of A. salmonicida luxS abolished AI-2 production. In conclusion, Atlantic salmon mucins regulate A. salmonicida quorum sensing in a luxS and N-acetylneuraminic acid-dependent manner.

Aromatic Agriculture: Volatile Compound-Based Plant Disease Diagnosis and Crop Protection

Volatiles exist ubiquitously in nature. Volatile compounds produced by plants and microorganisms confer inter-kingdom and intra-kingdom communications. Autoinducer signaling molecules from contact-based chemical communication, such as bacterial quorum sensing, are relayed through short distances. By contrast, biogenic volatiles derived from plant-microbe interactions generate long-distance (>20 cm) alarm signals for sensing harmful microorganisms. In this review, we discuss prior work on volatile compound-mediated diagnosis of plant diseases, and the use of volatile packaging and dispensing approaches for the biological control of fungi, bacteria, and viruses. In this regard, recent developments on technologies to analyze and detect microbial volatile compounds are introduced. Furthermore, we survey the chemical encapsulation, slow-release, and bio-nano techniques for volatile formulation and delivery that are expected to overcome limitations in the application of biogenic volatiles to modern agriculture. Collectively, technological advances in volatile compound detection, packaging, and delivery provide great potential for the implementation of ecologically-sound plant disease management strategies. We hope that this review will help farmers and young scientists understand the nature of microbial volatile compounds, and shift paradigms on disease diagnosis and management to aromatic (volatile-based) agriculture.

Quorum sensing inhibitory effect of hexanal on Autoinducer‐2 (AI‐2) and corresponding impacts on biofilm formation and enzyme activity in Erwinia carotovora and Pseudomonas fluorescens isolated from vegetables

Hexanal is an essential oil component which has quorum sensing (QS) inhibitor effect on vegetable spoilage bacteria such as Erwinia carotovora (E. carotovora) and Pseudomonas fluorescens (P. fluorescens). However, the QS inhibitory (QSI) effect of hexanal on Autoinducer-2 (AI-2), biofilm formation, and enzyme activity is not well understood. Our results showed that the contents of AI-2 produced by E. carotovora were higher than P. fluorescens, and the 4,5-Dihydroxy-2,3-pentanedione (DPD) concentrations were significantly (p < .05) inhibited by hexanal. Adding DPD exogenously could form more biofilm when inhibited by 1/2 of minimum inhibitory concentration (MIC) of hexanal, which showed that QSI effect of hexanal on biofilm was due to its inhibition on AI-2. There was no significant increase in enzyme activity, which suggested that enzyme inhibition by hexanal may not due to AI-2. Understanding the relationship between QSI effect of hexanal on AI-2, biofilm formation, and enzyme activity can improve to suppress decomposition of vegetables after harvest. Practical applications Hexanal is an EO component which has shown good QSI effect on biofilm formation and exoenzyme activities of P. fluorescens and E. carotovora. However, the reason for the QSI effect of hexanal was still unclear. In this study, hexanal as a QS inhibitor on these two vegetable spoilage bacteria was studied from the perspective of AI-2 signal molecule. Understanding the QS inhibition of hexanal on AI-2 signal molecule of spoilage bacteria and its relationship with biofilm formation and enzyme activity was necessary for the application of more EO components as QS inhibitor to suppress corruption of vegetables after harvest.

Agr Quorum Sensing influences the Wood-Ljungdahl pathway in Clostridium autoethanogenum

Acetogenic bacteria are capable of fermenting CO2 and carbon monoxide containing waste-gases into a range of platform chemicals and fuels. Despite major advances in genetic engineering and improving these biocatalysts, several important physiological functions remain elusive. Among these is quorum sensing, a bacterial communication mechanism known to coordinate gene expression in response to cell population density. Two putative agr systems have been identified in the genome of Clostridium autoethanogenum suggesting bacterial communication via autoinducing signal molecules. Signal molecule-encoding agrD1 and agrD2 genes were targeted for in-frame deletion. During heterotrophic growth on fructose as a carbon and energy source, single deletions of either gene did not produce an observable phenotype. However, when both genes were simultaneously inactivated, final product concentrations in the double mutant shifted to a 1.5:1 ratio of ethanol:acetate, compared to a 0.2:1 ratio observed in the wild type control, making ethanol the dominant fermentation product. Moreover, CO2 re-assimilation was also notably reduced in both hetero- and autotrophic growth conditions. These findings were supported through comparative proteomics, which showed lower expression of carbon monoxide dehydrogenase, formate dehydrogenase A and hydrogenases in the ∆agrD1∆agrD2 double mutant, but higher levels of putative alcohol and aldehyde dehydrogenases and bacterial micro-compartment proteins. These findings suggest that Agr quorum sensing, and by inference, cell density play a role in carbon resource management and use of the Wood-Ljungdahl pathway as an electron sink.

A Facile HPLC-UV-Based Method for Determining the Concentration of the Bacterial Universal Signal Autoinducer-2 in Environmental Samples

As a universal quorum sensing (QS) signal, autoinducer-2 (AI-2) is utilized by both Gram-negative and Gram-positive bacteria to coordinate several group behaviors, such as biofilm formation, virulence, and motility, when the bacterial cell density exceeds the thresholds. The determination of the AI-2 level is essential to understand the physiological and biochemical processes involved in bacterial communication. However, the current methods for AI-2 determination are complicated, time-consuming, and require costly equipment, such as a mass spectrometer (MS) or fluorescence detector (FLD). In this study, we present a new and easily applicable method for AI-2 determination. This method, based on the primary derivatization of AI-2 with 2,3-diaminonaphthalene (DAN), uses an affordable high-performance liquid chromatography (HPLC) instrument with a UV detector. Under optimized conditions, our method showed a good linearity (r2 = 0.999) and demonstrated the effective detection of AI-2 levels in various environmental samples, as follows: 0.38 (±0.05) μM for E. coli K12, 0.48 (±0.05) μM for Aeromonas sp. YB-2, 0.32 (±0.06) μM for the Enterobacter sp. YB-3, and 0.28 (±0.16) μM for activated sludge.

Roles of autoinducer-2 mediated quorum sensing in wastewater treatment

Abstract Quorum sensing (QS) is considered to be a promising regulation method for biological wastewater treatment (WWT) due to its regulation in extracellular substances (EPS) production, biofilm formation, granulation, colonization and bacterial activity and stability. Recently, autoinducer-2 (AI-2), a kind of interspecies communication QS signal molecule, is being increasingly reported for its roles in regulating bacterial gene expression and aggregation. Consequently, AI-2 mediated QS system is considered as a promising regulatory approach in WWT processes. This article systematically reviews the effects of AI-2-mediated QS system on bacterial behavior and its high potential for real-world applications in different WWT systems. Given the extensive presence of AI-2, AI-2 mediated QS could cooperate with other signal molecules in WWT processes, which suggests that the interactions among multiple signal molecules might be underestimated in the previous studies. The differences between AI-2 and AHL signaling molecules are also compared. Furthermore, the attempts at AI-2 regulated QS in pollution control of different WWT systems are summarized, while some challenges and defects still require targeted research in the future.

The abaI/abaR Quorum Sensing System Effects on Pathogenicity in Acinetobacter baumannii.

Acinetobacter baumannii is a Gram-negative pathogen that has emerged as one of the most troublesome pathogens for healthcare institutions globally. Bacterial quorum sensing (QS) is a process of cell-to-cell communication that relies on the production, secretion, and detection of autoinducer (AI) signals to share information about cell density and regulate gene expression accordingly. The molecular and genetic bases of A. baumannii virulence remains poorly understood. Therefore, the contribution of the abaI/abaR QS system to growth characteristics, morphology, biofilm formation, resistance, motility, and virulence of A. baumannii was studied in detail. RNA sequencing (RNA-seq) analysis indicated that genes involved in various aspects of energy production and conversion; valine, leucine, and isoleucine degradation; and lipid transport and metabolism are associated with bacterial pathogenicity. Our work provides a new insight into the abaI/abaR QS system effects on pathogenicity in A. baumannii. We propose that targeting the acyl homoserine lactone (AHL) synthase enzyme abaI could provide an effective strategy for attenuating virulence. On the contrary, interdicting the AI synthase receptor abaR elicits unpredictable consequences, which may lead to enhanced bacterial virulence.

Exogenous autoinducer-2 inhibits biofilm development of Desulfovibrio sp. Huiquan2017.

Sulfate-reducing bacteria (SRB) are culprits for microbiologically influenced corrosion, and biofilms are believed to play essential roles in the corrosion induced by SRB. However, little is known about the regulation of SRB biofilms. Quorum sensing signal molecules acyl-homoserine lactones (AHLs) and autoinducer-2 (AI-2) regulate biofilm formation of many bacteria. In this study, the production of AHLs and AI-2 by one SRB strain, Desulfovibrio sp. Huiquan2017, was detected, and the effect of exogenous AI-2 on bacterial biofilm formation was discussed. It was found that the cell-free supernatants of Desulfovibrio sp. Huiquan2017 induced luminescence in a ∆luxS mutant strain Vibrio harveyi BB170, indicating the production of functional AI-2 by the bacterium. In the presence of exogenous AI-2, the growth of Desulfovibrio sp. Huiquan2017 and early biofilm formation were not affected, but the later stage of biofilm development was inhibited significantly. The biofilms became looser, smaller, and thinner, and contained less bacteria and extracellular polymeric substances (EPS). The inhibition effect of AI-2 on the biofilm development of Desulfovibrio sp. Huiquan2017 was mainly achieved through reducing the amount of EPS in biofilms. These findings shed light on the biofilm regulation of SRB.

ACYL-HOMOSERINE LACTONES FOR CROP PRODUCTION AND STRESS TOLERANCE OF AGRICULTURAL PLANTS (review)

Acyl homoserine lactones (AHL) are a class of mediator molecules coordinating cell activity in the gram-negative bacteria population. AHLs synchronize individual genomes due to which bacterial populations function as a multicellular organism. AHLs provide a remote signaling between bacteria colonizing the phytosphere that enables the bacterial population to respond to external influences and establish symbiotic or antagonistic relationships with the host plant (A.R. Stacy et al., 2018; A. Shrestha et al., 2020). Autoreception of quantitative parameters of the bacterial population is called quorum sensing (QS) (R.G. Abisado et al., 2018). QS systems form autoinducer signaling molecules that easily penetrate from cells into the environment and back into the cell (M.B. Miller et al., 2001; B. Bassler, 2002). QS systems play a key role in the regulation of metabolic and physiological processes in a bacterial cell (M. Frederix et al., 2011; M. Whiteley et al., 2017). Bacterial signaling is perceived by eukaryotes, which form a symbiosis with microbial communities (A. Schenk et al., 2015; L.M. Babenko et al., 2016, 2017). Plant growth and development, nutrients assimilation, and stress resistance are largely determined by the pattern of this interaction (H.P. Bais et al., 2006; R. Ortíz-Castro et al., 2009; S. Basu et al., 2017). In the plant, bacterial signaling is controlled by the quorum quenching (QQ) system (N. Calatrava-Morales et al., 2018), whose mechanism of action is to suppress AHL synthesis by plant metabolites, compete with AHL for binding to receptor proteins, and repression of QS-controlled genes (H. Zhu et al., 2008; R. Sarkar et al., 2015). However, to date, the molecular mechanisms by which plants respond to bacterial signaling are not fully understood. Individual metabolites of AHL signaling have been characterized, but their role in the chemical interaction of partners in most cases requires further study. It has been shown that the QS phenomenon and its participants are involved in the regulation of prokaryotic-eukaryotic interactions, including the formation of biofilms, the synthesis of phytohormones, the transfer of plasmids, the production of virulence factors, bioluminescence, sporulation, and the formation of nodules (L.M. Babenko et al., 2017). Differences in the structure of molecules ensure that bacteria recognize their own AHL and separate foreign ones. The transfer of AHL from a bacterium to a host plant is carried out by means of membrane vesicles (M. Toyofuku, 2019). In recent years, there has been an active study of genetics, genomics, biochemistry, and signaling diversity of QS molecules. The regulation of the functions of the rhizosphere, the most dynamic site of interaction between the plant and the associated microflora with the participation of AHL, is of particular importance in the development of new biotechnological approaches aimed at increasing the yield and stress resistance of agricultural crops. One of the effective technologies for increasing resistance to biotic and abiotic stresses is pre-sowing treatment (priming) of seeds (A. Shrestha et al., 2020). Both direct (on plants) and indirect (on rhizosphere microflora) effects of AHL priming was established (O.V. Moshynets et al., 2019). AHL induce an increase of growth, of photosynthetic pigments content, as well as cause changes in the ratio of phytohormones in organs and tissues, affect the formation of defense mechanisms, which increases the productivity of agricultural crops (A. Schikora, S.T. Schenk, 2016; A. Shrestha et al., 2020). AHL meet the requirements of intensive organic farming, they are considered as promising ecological phytostimulants and phytomodulators capable of safely increasing the quantity and quality of agricultural products.

АЦИЛГОМОСЕРИНЛАКТОНЫ КАК РЕГУЛЯТОРЫ УРОЖАЙНОСТИ И СТРЕССОУСТОЙЧИВОСТИ СЕЛЬСКОХОЗЯЙСТВЕННЫХ КУЛЬТУР (обзор)

Acyl homoserine lactones (AHL) are a class of mediator molecules coordinating cell activity in the gram-negative bacteria population. AHLs synchronize individual genomes due to which bacterial populations function as a multicellular organism. AHLs provide a remote signaling between bacteria colonizing the phytosphere that enables the bacterial population to respond to external influences and establish symbiotic or antagonistic relationships with the host plant (A.R. Stacy et al., 2018; A. Shrestha et al., 2020). Autoreception of quantitative parameters of the bacterial population is called quorum sensing (QS) (R.G. Abisado et al., 2018). QS systems form autoinducer signaling molecules that easily penetrate from cells into the environment and back into the cell (M.B. Miller et al., 2001; B. Bassler, 2002). QS systems play a key role in the regulation of metabolic and physiological processes in a bacterial cell (M. Frederix et al., 2011; M. Whiteley et al., 2017). Bacterial signaling is perceived by eukaryotes, which form a symbiosis with microbial communities (A. Schenk et al., 2015; L.M. Babenko et al., 2016, 2017). Plant growth and development, nutrients assimilation, and stress resistance are largely determined by the pattern of this interaction (H.P. Bais et al., 2006; R. Ortíz-Castro et al., 2009; S. Basu et al., 2017). In the plant, bacterial signaling is controlled by the quorum quenching (QQ) system (N. Calatrava-Morales et al., 2018), whose mechanism of action is to suppress AHL synthesis by plant metabolites, compete with AHL for binding to receptor proteins, and repression of QS-controlled genes (H. Zhu et al., 2008; R. Sarkar et al., 2015). However, to date, the molecular mechanisms by which plants respond to bacterial signaling are not fully understood. Individual metabolites of AHL signaling have been characterized, but their role in the chemical interaction of partners in most cases requires further study. It has been shown that the QS phenomenon and its participants are involved in the regulation of prokaryotic-eukaryotic interactions, including the formation of biofilms, the synthesis of phytohormones, the transfer of plasmids, the production of virulence factors, bioluminescence, sporulation, and the formation of nodules (L.M. Babenko et al., 2017). Differences in the structure of molecules ensure that bacteria recognize their own AHL and separate foreign ones. The transfer of AHL from a bacterium to a host plant is carried out by means of membrane vesicles (M. Toyofuku, 2019). In recent years, there has been an active study of genetics, genomics, biochemistry, and signaling diversity of QS molecules. The regulation of the functions of the rhizosphere, the most dynamic site of interaction between the plant and the associated microflora with the participation of AHL, is of particular importance in the development of new biotechnological approaches aimed at increasing the yield and stress resistance of agricultural crops. One of the effective technologies for increasing resistance to biotic and abiotic stresses is pre-sowing treatment (priming) of seeds (A. Shrestha et al., 2020). Both direct (on plants) and indirect (on rhizosphere microflora) effects of AHL priming was established (O.V. Moshynets et al., 2019). AHL induce an increase of growth, of photosynthetic pigments content, as well as cause changes in the ratio of phytohormones in organs and tissues, affect the formation of defense mechanisms, which increases the productivity of agricultural crops (A. Schikora, S.T. Schenk, 2016; A. Shrestha et al., 2020). AHL meet the requirements of intensive organic farming, they are considered as promising ecological phytostimulants and phytomodulators capable of safely increasing the quantity and quality of agricultural products.

The Primary Physiological Roles of Autoinducer 2 in Escherichia coli Are Chemotaxis and Biofilm Formation.

Autoinducer 2 (AI-2) is a ubiquitous metabolite but, instead of acting as a “universal signal,” relatively few phenotypes have been associated with it, and many scientists believe AI-2 is often a metabolic byproduct rather than a signal. Here, the aim is to present evidence that AI-2 influences both biofilm formation and motility (swarming and chemotaxis), using Escherichia coli as the model system, to establish AI-2 as a true signal with an important physiological role in this bacterium. In addition, AI-2 signaling is compared to the other primary signal of E. coli, indole, and it is shown that they have opposite effects on biofilm formation and virulence.

An Efficient Synthesis of Optically Active [4-13C] Labelled Quorum Sensing Signal Autoinducer-2.

A new synthetic route for the quorum sensing signal Autoinducer-2 (AI-2) is described and used for the preparation of [4-13C]-AI-2 starting from [1-13C]-bromoacetic acid. The key step in this process was the enantioselective reduction of an intermediate ketone. This synthesis provides, selectively, both enantiomers of the labelled or unlabelled parent compound, (R) or (S)-4,5-dihydroxypentane-2,3-dione (DPD) and was used for an improved synthesis of [1-13C]-AI-2.

Quorum sensing signal autoinducer-2 promotes root colonization of Bacillus velezensis SQR9 by affecting biofilm formation and motility.

Root colonization of beneficial rhizobacteria is critical for their beneficial effects. Quorum sensing (QS) has been reported to affect the colonization of many plant pathogens. However, how QS signals regulate root colonization of beneficial rhizobacteria is unclear. In this study, the QS signal AI-2 synthetase-encoding gene luxS was completely deleted from the genome of the plant beneficial rhizobacterium Bacillus velezensis SQR9, and bioluminescence experiments showed that AI-2 production was blocked. Deletion of luxS reduced biofilm formation, motility, and root colonization of B. velezensis SQR9, while addition of exogenous AI-2 to the mutant restored this phenomenon. These results indicated that AI-2 positively affects the root colonization of B. velezensis SQR9. This study provided new insights for enhancing the colonization of beneficial rhizobacteria. KEY POINTS: • LuxS participated in the synthesis of the quorum sensing signal AI-2 in B. velezensis. • AI-2 enhanced motility, biofilm formation, and root colonization of B. velezensis. • AI-2 stimulated the production of γ-polyglutamic acid by B. velezensis.

Metagenomic Insight Towards Vanillin-Mediated Membrane Biofouling Prevention: In Silico Docking Validation.

Biofouling leads to water quality deterioration and higher maintenance cost for cleaning of membranes. The present study has demonstrated the application of a biomolecule (vanillin) in scrubbing and destabilizing biofilms of drinking water reverse osmosis (RO) membrane module in lab scale reactor set-up. Reverse osmosis membrane reactor was connected with tap water supply and subjected with optimal concentration of vanillin. The pressure drop was delayed by 17-20days as compared to control reactor. Real-time PCR analysis of metagenome indicated the reduced copy number of functional biofilm-associated genes (bdlA, lasI, pgaC) in treated membrane. SEM and metagenome analysis revealed that the sticky biofilm communities shifted to loosely bound emboli after vanillin treatment. Metagenome sequence analysis revealed the inhibitory activity against major biofouling biota like members of Proteobacteria, Acidobacteria, Acnitobacteria, Bacteroidetes, Candidatus, Nitrospira, and Firmicutes. Biofouled membrane metagenome sequence was also compared with real-life (brackish water, waste water, domestic drinking water) biofouled membrane communities. In silico docking of vanillin to receptor proteins and chemical configuration simulation along with other phenolic derivatives were performed, which suggested that the autoiducer signal capability of vanillin was effective against representative broad spectrum biofilm population. Vanillin exhibited the quorum-quenching mode of action by virtue of docking towards similar amino acid (Thr 131, Ilu 214) responsible of autoinducer signal anchoring in the transcriptional regulatory proteins.

Cannabigerol Prevents Quorum Sensing and Biofilm Formation of Vibrio harveyi.

Cannabigerol (CBG) is a non-psychoactive cannabinoid naturally present in trace amounts in the Cannabis plant. So far, CBG has been shown to exert diverse activities in eukaryotes. However, much less is known about its effects on prokaryotes. In this study, we investigated the potential role of CBG as an anti-biofilm and anti-quorum sensing agent against Vibrio harveyi. Quorum sensing (QS) is a cell-to-cell communication system among bacteria that involves small signaling molecules called autoinducers, enabling bacteria to sense the surrounding environment. The autoinducers cause alterations in gene expression and induce bioluminescence, pigment production, motility and biofilm formation. The effect of CBG was tested on V. harveyi grown under planktonic and biofilm conditions. CBG reduced the QS-regulated bioluminescence and biofilm formation of V. harveyi at concentrations not affecting the planktonic bacterial growth. CBG also reduced the motility of V. harveyi in a dose-dependent manner. We further observed that CBG increased LuxO expression and activity, with a concomitant 80% downregulation of the LuxR gene. Exogenous addition of autoinducers could not overcome the QS-inhibitory effect of CBG, suggesting that CBG interferes with the transmission of the autoinducer signals. In conclusion, our study shows that CBG is a potential anti-biofilm agent via inhibition of the QS cascade.

Fatty Acids as Antibiofilm and Antivirulence Agents.

Fatty acids (FAs) have emerged as a potential alternative to conventional antibiotics. Since many microbes respond differently to a variety of natural and synthetic FAs, substantial efforts have been made to understand the unique features of FAs that function as antimicrobials at high doses and biofilm inhibitors at low doses. Here, we provide an overview of the emerging antibiofilm properties of FAs and their relations to virulence and quorum sensing, such as diffusible signal factors (DSFs), acyl-homoserine lactones, and autoinducer-2 systems. Several FAs mimic DSFs and control motility, fimbriae, hyphae, and biofilm development as well as virulence characteristics of diverse microbes. The findings of FAs as antibiofilm and antivirulence agents will provide a new paradigm to cope with recalcitrant pathogens.

Autoinducer-2 influences tetracycline resistance in Streptococcus suis by regulating the tet(M) gene via transposon Tn916

The concern over increasing resistance to tetracyclines (TCs), such as tetracycline and chlortetracycline, necessitates exploration of new approaches to combating infection in antimicrobial therapy. Given that bacteria use the chemical language of autoinducer 2 (AI-2) signaling molecules in order to communicate and regulate group behaviors, we asked whether the AI-2 signaling influence the tetracyclines antibiotics susceptibility in S. suis. Our present work demonstrated that MIC increased when exogenous AI-2 was added, when compared to the wild type strain. When grown in the presence of sub-MIC of antibiotics, it has been shown that exogenous AI-2 increases growth rate and biofilm formation. These results suggest that the TCs resistance in S. suis could involve a signaling mechanism. Base on the above observations, transcriptomic analyses showed significant differences in the expression of tet(M) of tetracyclines resistance genes, as well as differences in Tn916 transposon related genes transcription, as judged by RT-PCR. Our results provide strong evidence that AI-2 signaling molecules is may involve in TCs antibiotic resistance in S. suis by regulating tet(M) gene via Tn916 transposon. This study may suggest that targeting AI-2 signaling in bacteria could represent an alternative approach in antimicrobial therapy.

Ιn Vitro Screening of γ-Aminobutyric Acid and Autoinducer-2 Signalling in Lactic Acid Bacteria Exhibiting Probiotic Potential Isolated from Natural Black Conservolea Olives.

In the present study, 33 strains of lactic acid bacteria (LAB) previously isolated from natural black Conservolea olives were assessed for their probiotic potential in vitro, as well as for their autoinducer-2 (AI-2) activity under standard growth conditions and the production of γ-aminobutyric acid (GABA). The probiotic tests included the in vitro resistance to low pH and resistance to bile salts, the evaluation of bile salt hydrolase activity, as well as safety tests regarding their possible haemolytic activity and their antimicrobial activity against pathogens. The results indicated that 17 strains were able to survive in low pH and in the presence of bile, with 15 of them also exhibiting partial bile salt hydrolase activity. None of the strains exhibited haemolytic activity or inhibited the growth of any of the examined pathogens. Moreover, the strains displayed generally low AI-2 activity under the growth conditions tested, regardless of the species. Interestingly, in contrast to what has been found in most foods, none of the isolates were found to produce GABA after 48 h of growth. The results from the AI-2 activity and extracellular GABA detection were considered as unexpected for LAB with probiotic attributes.

Inhibition of biofouling in membrane bioreactor by metabolic uncoupler based on controlling microorganisms accumulation and quorum sensing signals secretion.

Biofouling is a limiting bottleneck in the development of membrane bioreactor (MBR) since the birth of this technology. Recently, the biofouling control strategy based on interfering with the bacterial quorum sensing (QS) system is highly desirable for biofouling control in MBR. In this study, three lab-scale parallel MBR systems were operated over 100 days to investigate the inhibitory effect of a metabolic uncoupler (3,3',4',5-tetrachlorosalicylanilide, TCS) on biofouling and the potential mechanism for biofouling control. Compared to the control MBR, the fouling cycle duration of MBR 2 with 100μg/L TCS extended over two times. The attached biomass on membrane in MBR 2 decreased over 50% at the end of each operating period, which indicated that the addition of TCS significantly mitigated microorganisms accumulation on membrane. The content of interspecies QS signal (autoinducer-2) and intraspecific QS signals (N-octanoyl-dl-homoserine lactone, C8-HSL) was reduced by the TCS, suggesting the secretion of QS signals in MBR were affected by uncoupler. Although the addition of TCS induced brief fluctuations of extracellular proteins and polysaccharides, microorganisms seemed to rapidly acclimatize to the presence of TCS and then the secretion of extracellular polymeric substances (EPS) was inhibited by 100μg/L TCS. The continuous operation of MBR was not be affected by the low-concentration uncoupler via the analysis of substrate removal and sludge growth. This study systematically evaluated the effect and inhibitory efficiency of TCS on biofouling, biomass accumulation, QS signals, EPS and treatment performances, demonstrating the feasibility of metabolic uncoupler for biofouling control in MBR.

N-Acyl Homoserine Lactone Analog Modulators of the Pseudomonas aeruginosa Rhll Quorum Sensing Signal Synthase.

Virulence in the Gram-negative pathogen Pseudomonas aeruginosa relies in part on the efficient functioning of two LuxI/R dependent quorum sensing (QS) cascades, namely, the LasI/R and RhlI/R systems that generate and respond to N-(3-oxo)-dodecanoyl-l-homoserine lactone and N-butyryl-l-homoserine lactone, respectively. The two acyl homoserine lactone (AHL) synthases, LasI and RhlI, use 3-oxododecanoyl-ACP and butyryl-ACP, respectively, as the acyl-substrates to generate the corresponding autoinducer signals for the bacterium. Although AHL synthases represent excellent targets for developing QS modulators in P. aeruginosa, and in other related bacteria, the identification of potent and signal synthase specific inhibitors has represented a significant technical challenge. In the current study, we sought to test the utility of AHL analogs as potential modulators of an AHL synthase and selected RhlI in P. aeruginosa as an initial target. We systematically varied the chemical functionalities of the AHL headgroup, acyl chain tail, and head-to-tail linkage to construct a small library of signal analogs and evaluated them for RhlI modulatory activity. Although the native N-butyryl-l-homoserine lactone did not inhibit RhlI, we discovered that several of our long-chain, unsubstituted acyl-d-homoserine lactones and acyl-d-homocysteine thiolactones inhibited while a few of the 3-oxoacyl-chain counterparts activated the enzyme. Additional mechanistic investigations with acyl-substrate analogs and docking experiments with AHL analogs revealed two distinct inhibitor and activator binding pockets in the enzyme. This study provides the first evidence of the yet untapped potential of AHL analogs as signal synthase modulators of QS pathways.