Inhibition de l'expression des gènes de virulence chez Rhodococcus fascians et Pseudomonas aeruginosa par des flavonoïdes isolés chez les genres Dalbergia et Combretum

Plants are continuously confronted with a multitude attack either abiotic but also biotic in nature. Interestingly, despite the abundance of bacteria that plant has to face, only few are able to induce death or disease in the host plant. It is therefore likely that, in addition to secondary metabolites with antimicrobial properties, plants also synthesize secondary metabolites which are able to inhibit the expression of virulence genes in bacteria without affecting either growth or viability, which allows plants to host willingly or not bacterial populations. This work focuses on the identification of such metabolites in Malagasy plants (genera Dalbergia and Combretum) and the demonstration of their inhibitory effect on the expression of virulence genes in two different pathosystems: Rhodococcus fascians (a phytopathogen) and Pseudomonas aeruginosa (an opportunistic pathogen). Thus, two metabolites were isolated using a combination of chromatographic techniques coupled with tests that evaluate the expression of certain genes involved in the virulence mechanisms of these bacteria. The first is a new prenylated isoflavanone, named perbergin, isolated from the bark extract of D. pervillei. It was shown that the perbergin target attR gene expression, encoding a LysR-type transcriptional regulator that plays a key role in regulating the expression of virulence genes of R. fascians and the transition from an epiphytic to a pathogenic lifestyle. Therefore, we have also shown that the expression of all virulence genes known to date in R. fascians is also affected while the expression of genes involved in epiphytic fitness of the bacteria is not altered. In addition, the application of perbergin at the time of infection of plants susceptible to R. fascians shows that this molecule reduces in vivo the virulence of R. fascians, highlighting the potential of perbergin as an anti-infective agent. The second is a flavonoid known as catechin, isolated from the bark extract of C. albiflorum. Catechin significantly inhibits the expression of genes that regulate the mechanism of quorum sensing in P. aeruginosa such as lasI, LasR, rhlI and rhlR but also lasB and rhlA which expression depends on quorum sensing. Therefore, the production of virulence factors such as pyocyanin and elastase is significantly affected. Because of the limited number of our arsenal of antibiotics and their increasing ineffectiveness, the identification of these compounds create a path to an alternative in the fight against pathogenic bacteria and multidrug resistance of pathogenic bacteria to antibiotics. Our results also demonstrate the richness of Malagasy plants as (re)sources of new therapeutic molecules and the importance of widening the range of bacterial targets to be investigated to develop new strategies to fight within the endless war that we are waging against bacteria pathogens.Les plantes sont continuellement confrontées à une multitude d’attaques qu’elles soient de nature abiotique ou surtout biotique. Il est intéressant de noter que malgré la multitude de bactéries auxquelles les plantes doivent faire face, seules quelques unes sont capables d’induire la mort ou une maladie chez la plante hôte. Il est dès lors fort probable que, outre les métabolites secondaires ayant des propriétés antimicrobiennes, les plantes synthétisent également des métabolites secondaires capables d’inhiber l’expression des gènes de virulence chez les bactéries sans toutefois affecter ni leur croissance ni leur viabilité, ce qui permet aux plantes de contenir les populations bactériennes qu’elles hébergent de gré ou de force. Ce travail porte sur l’identification de ce type de métabolites dans des plantes malgaches (genres Dalbergia et Combretum) et la démonstration de leurs effets inhibiteurs sur l’expression de gènes de virulence chez deux pathosystèmes différents: Rhodococcus fascians (un phytopathogène) et Pseudomonas aeruginosa (un pathogène opportuniste). Ainsi, deux métabolites ont été isolés en utilisant une combinaison de techniques chromatographiques couplées avec des tests qui évaluent l’expression de certains gènes impliqués dans les mécanismes de virulence de ces bactéries. Le premier est un nouvel isoflavanone prénylé, nommé perbergine, isolé à partir de l’extrait d’écorces de D. pervillei. Il a été montré que la perbergine cible l’expression du gène attR, codant un régulateur transcriptionnel de type LysR qui joue un rôle clé dans la régulation de l’expression des gènes de virulence de R. fascians et qui assure la transition entre un mode de vie épiphyte et le mode pathogène. En conséquence, nous avons également montré que l’expression de l’ensemble des gènes de virulence connu à ce jour chez R. fascians est également affectée alors que l’expression de gènes impliqués dans l’aptitude épiphyte de la bactérie n’est pas altérée. Par ailleurs, l’application de perbergine au moment de l’infection de plantes sensibles à R. fascians montre que cette molécule atténue la virulence de R. fascians in vivo, mettant en exergue le potentiel de la perbergine comme agent anti-infectieux. Le deuxième est un flavonoïde, connu sous le nom de catéchine, isolé de l’extrait d’écorces de C. albiflorum. La catéchine inhibe significativement l’expression des gènes régulateurs du mécanisme du quorum sensing chez P. aeruginosa tels que lasI, lasR, rhlI et rhlR et également lasB et rhlA dont l’expression dépend du quorum sensing. En conséquence, la production des facteurs de virulence tels que la pyocyanine et l’élastase est significativement affectée. Compte tenu de l’appauvrissement de notre arsenal d’antibiotiques et de leur inefficacité croissante, l’identification de ces composés ouvre une voie alternative de lutte contre les bactéries pathogènes et la multirésistance des bactéries pathogènes aux antibiotiques. Nos résultats démontrent également la richesse des plantes malgaches comme (res)sources de nouvelles molécules thérapeutiques et l’importance d’élargir le champ des cibles bactériennes à investiguer pour développer de nouvelles stratégies de lutte dans la guerre sans fin que nous menons contre les bactéries pathogènes.Doctorat en Sciencesinfo:eu-repo/semantics/nonPublishe

La coopération universitaire, ou comment réunir le Sud et le Nord pour atteindre le quorum afin de mener des recherches de pointe

info:eu-repo/semantics/publishe

Dalbergia pervillei, a legume refractory to Rhodococcus fascians infection, contains new prenylated isoflavones repressing the expression of bacteria loci required for symptom development

info:eu-repo/semantics/publishe

Dalbergia pervillei, a legume refractory to Rhodococcus fascians infection, contains new prenylated isoflavones repressing the expression of bacteria loci required for symptom development

info:eu-repo/semantics/nonPublishe

Identification of Catechin as One of the Flavonoids from Combretum albiflorum Bark Extract That Reduces the Production of Quorum-Sensing-Controlled Virulence Factors in Pseudomonas aeruginosa PAO1 ▿

Quorum-sensing (QS) regulates the production of key virulence factors in Pseudomonas aeruginosa and other important pathogenic bacteria. In this report, extracts of leaves and bark of Combretum albiflorum (Tul.) Jongkind (Combretaceae) were found to quench the production of QS-dependent factors in P. aeruginosa PAO1. Chromatographic fractionation of the crude active extract generated several active fractions containing flavonoids, as shown by their typical spectral features. Purification and structural characterization of one of the active compounds led to the identification of the flavan-3-ol catechin [(2R,3S)-2-(3,4-dihydroxyphenyl)-3,4-dihydro-1(2H)-benzopyran-3,5,7-triol]. The identity of catechin as one of the active molecules was confirmed by comparing the high-pressure liquid chromatography profiles and the mass spectrometry spectra obtained for a catechin standard and for the active C. albiflorum fraction. Moreover, standard catechin had a significant negative effect on pyocyanin and elastase productions and biofilm formation, as well as on the expression of the QS-regulated genes lasB and rhlA and of the key QS regulatory genes lasI, lasR, rhlI, and rhlR. The use of RhlR- and LasR-based biosensors indicated that catechin might interfere with the perception of the QS signal N-butanoyl-l-homoserine lactone by RhlR, thereby leading to a reduction of the production of QS factors. Hence, catechin, along with other flavonoids produced by higher plants, might constitute a first line of defense against pathogenic attacks by affecting QS mechanisms and thereby virulence factor production

The flavanone naringenin reduces the production of quorum sensing-controlled virulence factors in Pseudomonas aeruginosa PAO1

Preliminary screening of the Malagasy plant Combretum albiflorum for compounds attenuating the production of quorum sensing (QS)-controlled virulence factors in bacteria led to the identification of active fractions containing flavonoids. In the present study, several flavonoids belonging to the flavone, flavanone, flavonol and chalcone structural groups were screened for their capacity to reduce the production of QS-controlled factors in the opportunistic pathogen Pseudomonas aeruginosa (strain PAO1). Flavanones (i.e. naringenin, eriodictyol and taxifolin) significantly reduced the production of pyocyanin and elastase in P. aeruginosa without affecting bacterial growth. Consistently, naringenin and taxifolin reduced the expression of several QS-controlled genes (i.e. lasI, lasR, rhlI, rhlR, lasA, lasB, phzA1 and rhlA) in P. aeruginosa PAO1. Naringenin also dramatically reduced the production of the acylhomoserine lactones N-(3-oxododecanoyl)-l-homoserine lactone (3-oxo-C12-HSL) and N-butanoyl-l-homoserine lactone (C4-HSL), which is driven by the lasI and rhlI gene products, respectively. In addition, using mutant strains deficient for autoinduction (ΔlasI and ΔrhlI) and LasR- and RhlR-based biosensors, it was shown that QS inhibition by naringenin not only is the consequence of a reduced production of autoinduction compounds but also results from a defect in the proper functioning of the RlhR-C4-HSL complex. Widely distributed in the plant kingdom, flavonoids are known for their numerous and determinant roles in plant physiology, plant development and in the success of plant-rhizobia interactions, but, as shown here, some of them also have a role as inhibitors of the virulence of pathogenic bacteria by interfering with QS mechanisms.SCOPUS: ar.jinfo:eu-repo/semantics/publishe

Endemic malagasy Dalbergia species inhibit quorum sensing in Pseudomonas aeruginosa PAO1.

Various species of the plant genus Dalbergia are traditionally used as medicine for sundry ailments and some of them have been shown recently to quench the virulence of Gram-positive and Gram-negative bacteria. Cell-to-cell communication mechanisms, quorum sensing (QS) in particular, are key regulators of virulence in many pathogenic bacteria. Screening n-hexane extracts of leaves, roots and bark of endemic Malagasy Dalbergia species for their capacity to antagonize QS mechanisms in Pseudomonas aeruginosa PAO1 showed that many reduced the expression of the QS-regulated genes lasB and rhlA. However, only the extract of Dalbergia trichocarpa bark (DTB) showed a significant reduction of QS gene expression without any effect on the aceA gene encoding a QS-independent isocitrate lyase. Further characterization of DTB impact on QS revealed that the QS systems las and rhl are inhibited and that swarming, twitching, biofilm formation and the production of pyocyanin, elastase and proteases are also hampered in the presence of the DTB extract. Importantly, compared with the known QS inhibitor naringenin, the DTB extract showed a stronger negative effect on twitching, biofilm formation and tobramycin resistance. Preliminary structural characterization of these potent biofilm disrupters suggests that they belong to the phytosterols. The strong inhibition of motility and biofilm formation suggests that the DTB extract contains agents disrupting biofilm architecture, which is an important observation in the context of the design of new drugs targeting biofilm-encapsulated pathogens.Journal ArticleResearch Support, Non-U.S. Gov'tSCOPUS: ar.jinfo:eu-repo/semantics/publishe