RESUMO
After a time away from the classrooms and laboratories due to the global pandemic, the return to teaching activities during the semester represented a challenge to both teachers and students. Our particular situation in a Microbial Physiology course was the necessity of imparting in shorter time, laboratory practices that usually take longer. This article describes a 2-week-long laboratory exercise that covers several concepts in an interrelated way: conjugation as a gene transfer mechanism, regulation of microbial physiology, production of secondary metabolites, degradation of macromolecules, and biofilm formation. Utilizing a Quorum Quenching (QQ) strategy, the Quorum Sensing (QS) system of Pseudomonas aeruginosa is first attenuated. Then, phenotypes regulated by QS are evidenced. QS is a regulatory mechanism of microbial physiology that relies on signal molecules. QS is related in P. aeruginosa to several virulence factors, some of which are exploited in the laboratory practices presented in this work. QQ is a phenomenon by which QS is interrupted or attenuated. We utilized a QQ approach based on the enzymatic degradation of the P. aeruginosa QS signals to evidence QS-regulated traits that are relevant to our Microbial Physiology course. Results obtained with the same test performed by a random group of students before and after the activities show the positive effectiveness of the approach presented in this work.
Assuntos
Laboratórios , Pseudomonas aeruginosa , Percepção de Quorum , Pseudomonas aeruginosa/fisiologia , Pseudomonas aeruginosa/metabolismo , Humanos , Estudantes , Biofilmes/crescimento & desenvolvimentoRESUMO
Vinasse is a by-product with a key role in circular economy. In this work, we analyze sugarcane vinasse as culture medium for obtaining single and mixed inoculants. Trichoderma harzianum MT2 was cultured in single and sequential co-culture with Pseudomonas capeferrum WCS358 or Rhizobium sp. N21.2. Fungal biomass in single culture was more than three folds higher in vinasse than in a standard medium, and was higher in co-culture with Rhizobium sp. N21.2 than with P. capeferrum WCS358. Bacterial growths in vinasse, in particular P. capeferrum WCS358, were improved in co-culture with T. harzianum MT2. Residual vinasses, obtained after microbial growth, presented almost neutral pH and lower conductivities and toxicity than raw vinasse. Fertigation with residual vinasses modifies characteristics of soil evidenced in the total N, cation exchange capacity, urease and acid phosphatase, and microbial metabolic diversity, in comparison to raw vinasse. In general, soil fertigation with residual vinasse from co-culture with P. capeferrum WCS358 is more similar to irrigation with water. Treatment evaluation indicates that vinasse is suitable for the production of mixed inoculants containing T. harzianum. The co-culture with P. capeferrum WCS358 improves the characteristics of the residual vinasse allowing a fertigation with less detrimental effect in soil in comparison to Rhizobium sp. N21.2. Obtaining valuable biomass of single or mixed inoculants in vinasse with lower ecological impact is relevant for the circular and green economy.
Assuntos
Rhizobium , Saccharum , Solo , Conservação de Recursos EnergéticosRESUMO
Beyond their biological roles, metals have a strong impact on the environment. It has been reported that metals are also inhibitory of Quorum Sensing (QS) mechanisms, ones of the best characterized signaling systems in bacteria and fungi. We analyzed the effect of CuSO4, CdCl2, and K2Cr2O7, on QS systems sharing or differing in the bacterial host or the QS signal. The results in this study show that CuSO4 can not only be inhibitory, but also stimulatory of QS activity: at 0.2 mM increased six fold the activity in Chromobacterium subtsugae CV026. This behavior is related to the concentration of the metal and the particular QS system: E. coli MT102 (pJBA132) was no affected, but CuSO4 decreased the QS activity of Pseudomonas putida F117 (pKR-C12) to half its control values. K2Cr2O7 increased four and three folds the QS activities of E. coli MT102 (pJBA132) and P. putida F117 (pAS-C8), respectively, but without effect when combined with CuSO4 or CdCl2. CdCl2 only showed a positive effect in CV026 when combined with CuSO4. Results suggest that factors related with the culture conditions impact on the influence of the metals, and reinforce the importance of the environment in the modulation of QS activity.
Assuntos
Técnicas Biossensoriais , Percepção de Quorum , Cloreto de Cádmio/farmacologia , Dicromato de Potássio/farmacologia , Sulfato de Cobre/farmacologia , Escherichia coli , Bactérias , Chromobacterium , Antibacterianos/farmacologia , Pseudomonas aeruginosaRESUMO
Agrobacterium tumefaciens is considered a prominent phytopathogen, though most isolates are nonpathogenic. Agrobacteria can inhabit plant tissues interacting with other microorganisms. Yeasts are likewise part of these communities. We analyzed the quorum sensing (QS) systems of A. tumefaciens strain 6N2, and its relevance for the interaction with the yeast Meyerozyma guilliermondii, both sugarcane endophytes. We show that strain 6N2 is nonpathogenic, produces OHC8-HSL, OHC10-HSL, OC12-HSL and OHC12-HSL as QS signals, and possesses a complex QS architecture, with one truncated, two complete systems, and three additional QS-signal receptors. A proteomic approach showed differences in QS-regulated proteins between pure (64 proteins) and dual (33 proteins) cultures. Seven proteins were consistently regulated by quorum sensing in pure and dual cultures. M. guilliermondii proteins influenced by QS activity were also evaluated. Several up- and down- regulated proteins differed depending on the bacterial QS. These results show the QS regulation in the bacteria-yeast interactions.
Assuntos
Percepção de Quorum , Saccharomycetales , Agrobacterium tumefaciens/genética , Agrobacterium tumefaciens/metabolismo , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Proteômica , Saccharomycetales/genética , Saccharomycetales/metabolismoRESUMO
Quorum sensing modulates bacterial collective behaviors including biofilm formation, motility and virulence in the important human pathogen Acinetobacter baumannii. Disruption of quorum sensing has emerged as a promising strategy with important therapeutic potential. In this work, we show that light modulates the production of acyl-homoserine lactones (AHLs), which were produced in higher levels in the dark than under blue light at environmental temperatures, a response that depends on the AHL synthase, AbaI, and on the photoreceptor BlsA. BlsA interacts with the transcriptional regulator AbaR in the dark at environmental temperatures, inducing abaI expression. Under blue light, BlsA does not interact with AbaR, but induces expression of the lactonase aidA and quorum quenching, consistently with lack of motility at this condition. At temperatures found in warm-blooded hosts, the production of AHLs, quorum quenching as well as abaI and aidA expression were also modulated by light, though in this case higher levels of AHLs were detected under blue light than in the dark, in a BlsA-independent manner. Finally, AbaI reduces A. baumannii's ability to kill C. albicans only in the dark both at environmental as well as at temperatures found in warm-blooded hosts. The overall data indicate that light directly modulates quorum network in A. baumannii.