The swimming defect caused by the absence of the transcriptional regulator LdtR in Sinorhizobium meliloti is restored by mutations in the motility genes motA and motS.

The flagellar motor is a powerful macromolecular machine used to propel bacteria through various environments. We determined that flagellar motility of the alpha-proteobacterium Sinorhizobium meliloti is nearly abolished in the absence of the transcriptional regulator LdtR, known to influence peptidoglycan remodeling and stress response. LdtR does not regulate motility gene transcription. Remarkably, the motility defects of the ΔldtR mutant can be restored by secondary mutations in the motility gene motA or a previously uncharacterized gene in the flagellar regulon, which we named motS. MotS is not essential for S. meliloti motility and may serve an accessory role in flagellar motor function. Structural modeling predicts that MotS comprised an N-terminal transmembrane segment, a long-disordered region, and a conserved ß-sandwich domain. The C terminus of MotS is localized in the periplasm. Genetics based substitution of MotA with MotAG12S also restored the ΔldtR motility defect. The MotAG12S variant protein features a local polarity shift at the periphery of the MotAB stator units. We propose that MotS may be required for optimal alignment of stators in wild-type flagellar motors but becomes detrimental in cells with altered peptidoglycan. Similarly, the polarity shift in stator units composed of MotB/MotAG12S might stabilize its interaction with altered peptidoglycan.

Involvement of ArlI, ArlJ, and CirA in archaeal type IV pilin-mediated motility regulation.

Many prokaryotes use swimming motility to move toward favorable conditions and escape adverse surroundings. Regulatory mechanisms governing bacterial flagella-driven motility are well-established; however, little is yet known about the regulation underlying swimming motility propelled by the archaeal cell surface structure, the archaella. Previous research showed that the deletion of the adhesion pilins (PilA1-6), subunits of the type IV pili cell surface structure, renders the model archaeon Haloferax volcanii non-motile. In this study, we used ethyl methanesulfonate mutagenesis and a motility assay to identify motile suppressors of the ∆pilA[1-6] strain. Of the eight suppressors identified, six contain missense mutations in archaella biosynthesis genes, arlI and arlJ. In trans expression of arlI and arlJ mutant constructs in the respective multi-deletion strains ∆pilA[1-6]∆arlI and ∆pilA[1-6]∆arlJ confirmed their role in suppressing the ∆pilA[1-6] motility defect. Additionally, three suppressors harbor co-occurring disruptive missense and nonsense mutations in cirA, a gene encoding a proposed regulatory protein. A deletion of cirA resulted in hypermotility, while cirA expression in trans in wild-type cells led to decreased motility. Moreover, quantitative real-time PCR analysis revealed that in wild-type cells, higher expression levels of arlI, arlJ, and the archaellin gene arlA1 were observed in motile early-log phase rod-shaped cells compared to non-motile mid-log phase disk-shaped cells. Conversely, ∆cirA cells, which form rods during both early- and mid-log phases, exhibited similar expression levels of arl genes in both growth phases. Our findings contribute to a deeper understanding of the mechanisms governing archaeal motility, highlighting the involvement of ArlI, ArlJ, and CirA in pilin-mediated motility regulation.IMPORTANCEArchaea are close relatives of eukaryotes and play crucial ecological roles. Certain behaviors, such as swimming motility, are thought to be important for archaeal environmental adaptation. Archaella, the archaeal motility appendages, are evolutionarily distinct from bacterial flagella, and the regulatory mechanisms driving archaeal motility are largely unknown. Previous research has linked the loss of type IV pili subunits to archaeal motility suppression. This study reveals three Haloferax volcanii proteins involved in pilin-mediated motility regulation, offering a deeper understanding of motility regulation in this understudied domain while also paving the way for uncovering novel mechanisms that govern archaeal motility. Understanding archaeal cellular processes will help elucidate the ecological roles of archaea as well as the evolution of these processes across domains.

The FabA-FabB Pathway Is Not Essential for Unsaturated Fatty Acid Synthesis but Modulates Diffusible Signal Factor Synthesis in Xanthomonas campestris pv. campestris.

Most bacteria use type II fatty acid synthesis (FAS) systems for synthesizing fatty acids, of which the conserved FabA-FabB pathway is considered to be crucial for unsaturated fatty acid (UFA) synthesis in gram-negative bacteria. Xanthomonas campestris pv. campestris, the phytopathogen of black rot disease in crucifers, produces higher quantities of UFAs under low-temperature conditions for increasing membrane fluidity. The fabA and fabB genes were identified in the X. campestris pv. campestris genome by BLAST analysis; however, the growth of the X. campestris pv. campestris fabA and fabB deletion mutants was comparable to that of the wild-type strain in nutrient and minimal media. The X. campestris pv. campestris ΔfabA and ΔfabB strains produced large quantities of UFAs and, altogether, these results indicated that the FabA-FabB pathway is not essential for growth or UFA synthesis in X. campestris pv. campestris. We also observed that the expression of X. campestris pv. campestris fabA and fabB restored the growth of the temperature-sensitive Escherichia coli fabA and fabB mutants CL104 and CY242, respectively, under non-permissive conditions. The in-vitro assays demonstrated that the FabA and FabB proteins of X. campestris pv. campestris catalyzed FAS. Our study also demonstrated that the production of diffusible signal factor family signals that mediate quorum sensing was higher in the X. campestris pv. campestris ΔfabA and ΔfabB strains and greatly reduced in the complementary strains, which exhibited reduced swimming motility and attenuated host-plant pathogenicity. [Formula: see text] Copyright © 2023 The Author(s). This is an open access article distributed under the CC BY 4.0 International license.

FliL and its paralog MotF have distinct roles in the stator activity of the Sinorhizobium meliloti flagellar motor.

The bacterial flagellum is a complex macromolecular machine that drives bacteria through diverse fluid environments. Although many components of the flagellar motor are conserved across species, the roles of FliL are numerous and species-specific. Here, we have characterized an additional player required for flagellar motor function in Sinorhizobium meliloti, MotF, which we have identified as a FliL paralog. We performed a comparative analysis of MotF and FliL, identified interaction partners through bacterial two-hybrid and pull-down assays, and investigated their roles in motility and motor rotation. Both proteins form homooligomers, and interact with each other, and with the stator proteins MotA and MotB. The ∆motF mutant exhibits normal flagellation but its swimming behavior and flagellar motor activity are severely impaired and erratic. In contrast, the ∆fliL mutant is mostly aflagellate and nonmotile. Amino acid substitutions in cytoplasmic regions of MotA or disruption of the proton channel plug of MotB partially restored motor activity to the ∆motF but not the ∆fliL mutant. Altogether, our findings indicate that both, MotF and FliL, are essential for flagellar motor torque generation in S. meliloti. FliL may serve as a scaffold for stator integration into the motor, and MotF is required for proton channel modulation.

Inhibitory effect of resveratrol on swimming motility and adhesion ability against Salmonella enterica serovar Typhimurium infection.

Salmonella enterica serovar Typhimurium (S. typhimurium) is a common Gram-negative foodborne pathogen that threatens public health and hinders the development of livestock industry. Resveratrol, an important component in grape fruits and seeds, has been shown to possess multiple biological activities, but its potential effects on S. typhimurium-mediated virulence have been rarely reported. In this study, we investigated the effect of resveratrol on S. typhimurium flagella -mediated virulence. The results showed that resveratrol significantly reduced the transcription of flagella genes and swimming motility of S. typhimurium, and also inhibited the transcription of T3SS-related virulence genes with varying degrees inhibiting bacterial growth. Simultaneously, resveratrol significantly reduced the adhesion of S. typhimurium to HeLa cells. Unfortunately, resveratrol does not improve the survival rate of S. typhimurium-infected mice, but it reduces the bacterial load in the liver and spleen of infected mice, and it also has a certain degree of anti-inflammatory activity. In summary, these results indicated that resveratrol has the potential to be developed as an alternative drug or antibacterial agent to prevent Salmonella infection.

Assessment of the potential of phage-antibiotic synergy to induce collateral sensitivity in Salmonella Typhimurium.

This study was designed to evaluate the synergistic effect of phage and antibiotic on the induction of collateral sensitivity in Salmonella Typhimurium. The synergistic effects of Salmonella phage PBST32 combined with ciprofloxacin (CIP) against S. Typhimurium KCCM 40253 (STKCCM) were evaluated using a fractional inhibitory concentration (FIC) assay. The CIP susceptibility of STKCCM was increased when combined with PBST32, showing 16-fold decrease at 7 log PFU/mL. The combination of 1/2 × MIC of CIP and PBST32 (CIP[1/2]+PBST32) effectively inhibited the growth of STKCCM up to below the detection limit (1.3 log CFU/mL) after 12 h of incubation at 37 °C. The significant reduction in bacterial swimming motility was observed for PBST32 and CIP[1/4]+PBST32. The CIP[1/4]+PBST32 increased the fitness cost (relative fitness = 0.57) and decreased the cross-resistance to different classes of antibiotics. STKCCM treated with PBST32 alone treatment exhibited the highest coefficient of variation (90%), followed by CIP[1/4]+PBST32 (75%). These results suggest that the combination of PBST32 and CIP can be used to control bacterial pathogens.

Metformin promotes bacterial surface aggregation by inhibiting the swimming motility of flagellated Escherichia coli.

The antiglycemic drug metformin, which is widely prescribed as a first-line drug for the treatment of type 2 diabetes, has become a concern for emerging pollutants in natural ecosystems. However, its effects on bacterial swimming motility remain unclear. In this study, we showed that metformin promotes bacterial surface aggregation by tracking swimming and by measuring the density distribution of Escherichia coli cultured with metformin near a surface in a homogeneous environment. Flagella are essential for the promotion of bacterial surface aggregation by metformin. Swimming motility, which is mediated by flagella, determines bacterial surface aggregation. The promotion of bacterial surface aggregation by metformin is caused by a reduction in swimming motility, which is governed by a decrease in the proton motive force. Our results reveal that metformin has a pronounced effect on flagellated bacterial motility associated with surface sensing and aggregation.

Evolutionary bioenergetics of ciliates.

Understanding why various organisms evolve alternative ways of living requires information on both the fitness advantages of phenotypic modifications and the costs of constructing and operating cellular features. Although the former has been the subject of a myriad of ecological studies, almost no attention has been given to how organisms allocate resources to alternative structures and functions. We address these matters by capitalizing on an array of observations on diverse ciliate species and from the emerging field of evolutionary bioenergetics. A relatively robust and general estimator for the total cost of a cell per cell cycle (in units of ATP equivalents) is provided, and this is then used to understand how the magnitudes of various investments scale with cell size. Among other things, we examine the costs associated with the large macronuclear genomes of ciliates, as well as ribosomes, various internal membranes, osmoregulation, cilia, and swimming activities. Although a number of uncertainties remain, the general approach taken may serve as blueprint for expanding this line of work to additional traits and phylogenetic lineages.

Xanthomonas oryzae pv. oryzae Exoribonuclease R Is Required for Complete Virulence in Rice, Optimal Motility, and Growth Under Stress.

Exoribonuclease R (RNase R) is a 3' hydrolytic exoribonuclease that can degrade structured RNA. Mutation in RNase R affects virulence of certain human pathogenic bacteria. The aim of this study was to determine whether RNase R is necessary for virulence of the phytopathogen that causes bacterial blight in rice, Xanthomonas oryzae pv. oryzae (Xoo). In silico analysis has indicated that RNase R is highly conserved among various xanthomonads. Amino acid sequence alignment of Xoo RNase R with RNase R from various taxa indicated that Xoo RNase R clustered with RNase R of order Xanthomonadales. To study its role in virulence, we generated a gene disruption mutant of Xoo RNase R. The Xoo rnr- mutant is moderately virulence deficient, and the complementing strain (rnr-/pHM1::rnr) rescued the virulence deficiency of the mutant. We investigated swimming and swarming motilities in both nutrient-deficient minimal media and nutrient-optimal media. We observed that RNase R mutation has adversely affected the swimming and swarming motilities of Xoo in optimal media. However, in nutrient-deficient media only swimming motility was noticeably affected. Growth curves in optimal media at suboptimal temperature (15°C cold stress) indicate that the Xoo rnr- mutant grows more slowly than the Xoo wild type and complementing strain (rnr-/pHM1::rnr). Given these findings, we report for the first time that RNase R function is necessary for complete virulence of Xoo in rice. It is also important for motility of Xoo in media and for growth of Xoo at suboptimal temperature.

Characterization of the response of Escherichia coli to l-fucose in bacterial swimming motility.

l-Fucose, as a monosaccharide in nature, plays a crucial role in bacteria colonization. Escherichia coli (E. coli), as a common microorganism in environment, utilize bacterial flagellar motor to drive the rotation of flagella, which is regulated by chemotactic signal transduction signals. Yet the effect of l-fucose to bacterial motility remains unclear. The effect of l-fucose on the swimming motility of bacteria was investigated from the level of single flagellar motor to individual cell and cell population by employing a bead assay, a high-throughput 2D tracking assay and a high-throughput dark-field flicker microscopy. The results showed that the swimming motility of the bacteria cultured with l-fucose was decreased, while the tumble frequency increased. Furthermore, the behavioral alterations of bacteria affected by l-fucose were directly reveled by measuring the cell distribution of bacteria swimming near surfaces and bacterial surface adhesion, suggesting that l-fucose promotes bacterial surface aggregation and surface adhesion. The effect of l-fucose on bacterial swimming motility characterized in this study are consistent with the key role that l-fucose plays in bacterial colonization.

Can only one physiological trait determinate the adverse effect of green fluorescent protein (GFP) incorporation on Vibrio virulence?

Green fluorescent protein (GFP) has been used extensively for in situ animal studies that follow up bacterial infection under epifluorescence microscopy. It is assumed that GFP is acting as a "neutral" protein with no influence on the bacterial physiology. To verify this hypothesis, the virulence of Vibrio splendidus ME9, Vibrio anguillarum NB10, and their respective GFP-tagged strains ME9-GFP and NB10-GFP (transconjugants) was compared in vitro and tested in vivo towards blue mussel (Mytilus edulis) larvae. Results showed that the incorporation of GFP negatively impacted the growth and swimming motility of NB10 in vitro. Correspondingly, the mRNA levels of genes involved in bacterial swimming motility (flaA, flaE, and cheR) were significantly down-regulated in NB10-GFP. As for the strain ME9 on the other hand, GFP incorporation only had a negative effect on swimming motility. However, both the strains NB10-GFP and ME9-GFP showed almost the same virulence as their respective parental strain towards mussel larvae in vivo. Overall, the data presented here demonstrated that incorporation of GFP may cause modifications in cell physiology and highlight the importance of preliminary physiological tests to minimize the negative influence of GFP tagging when it is used to monitor the target localization. The study also supports the idea that the virulence of Vibrio species is determined by complex regulatory networks. Notwithstanding the change of a single physiological trait, especially growth or swimming motility, the GFP-tagged Vibrio strain can thus still be considered usable in studies mainly focusing on the virulence of the strain. KEY POINTS: • The effect of GFP incorporation on physiological trait of Vibrio strains. • The virulence in vibrios could be multifactorial. • The stable virulence of Vibrio strains after GFP incorporation.

Swimming behavior of the multicellular magnetotactic prokaryote 'Candidatus Magnetoglobus multicellularis' near solid boundaries and natural magnetic grains.

The magnetotactic yet uncultured species 'Candidatus Magnetoglobus multicellularis' is a spherical, multicellular ensemble of bacterial cells able to align along magnetic field lines while swimming propelled by flagella. Magnetotaxis is due to intracytoplasmic, membrane-bound magnetic crystals called magnetosomes. The net magnetic moment of magnetosomes interacts with local magnetic fields, imparting the whole microorganism a torque. Previous works investigated 'Ca. M. multicellularis' behavior when free swimming in water; however, they occur in sediments where bumping into solid particles must be routine. In this work, we investigate the swimming trajectories of 'Ca. M. multicellularis' close to solid boundaries using video microscopy. We applied magnetic fields 0.25-8.0 mT parallel to the optical axis of a light microscope, such that microorganisms were driven upwards towards a coverslip. Because their swimming trajectories approach cylindrical helixes, circular profiles would be expected. Nevertheless, at fields 0.25-1.1 mT, most trajectory projections were roughly sinusoidal, and net movements were approximately perpendicular to applied magnetic fields. Closed loops appeared in some trajectory projections at 1.1 mT, which could indicate a transition to the loopy profiles observed at magnetic fields ≥ 2.15 mT. The behavior of 'Ca. M. multicellularis' near natural magnetic grains showed that they were temporarily trapped by the particle's magnetic field but could reverse the direction of movement to flee away. Our results show that interactions of 'Ca. M. multicellularis with solid boundaries and magnetic grains are complex and possibly involve mechano-taxis.

Simple mechanosense and response of cilia motion reveal the intrinsic habits of ciliates.

An important habit of ciliates, namely, their behavioral preference for walls, is revealed through experiments and hydrodynamic simulations. A simple mechanical response of individual ciliary beating (i.e., the beating is stalled by the cilium contacting a wall) can solely determine the sliding motion of the ciliate along the wall and result in a wall-preferring behavior. Considering ciliate ethology, this mechanosensing system is likely an advantage in the single cell's ability to locate nutrition. In other words, ciliates can skillfully use both the sliding motion to feed on a surface and the traveling motion in bulk water to locate new surfaces according to the single "swimming" mission.

Cinnamomum cassia and Syzygium aromaticum Essential Oils Reduce the Colonization of Salmonella Typhimurium in an In Vivo Infection Model Using Caenorhabditis elegans.

The regulation of intestinal colonization in livestock by means of non-bactericidal additives is an important management lever for zoonotic bacteria such as Salmonella spp. Caenorhabditis elegans is proposed here as a model for the evaluation of five essential oils (EOs) as anti-colonization products against Salmonella Typhimurium. An evaluation of the toxicity of EOs for C. elegans showed LD50 values ranging from 74.5 ± 9.6 µg/mL for Cinnamomum cassia (CEO) to 271.6 ± 14.9 µg/mL for Syzygium aromaticum (SyEO). Both EOs significantly inhibited bacterial colonization in the digestive tract of C. elegans with reductions of 0.88 and 0.70 log CFU/nematode at nontoxic concentrations of 50 µg/mL and 150 µg/mL, respectively. With the minimal bactericidal concentrations of CEO and SyEO against S. Typhimurium being 312.5 µg/mL and 625 µg/mL, respectively, an antibacterial effect can be excluded to explain the inhibition of the bacterial load. The anti-colonizing activity of these two EOs could, however, be related to an inhibition of the swimming motility, which was significantly reduced by 23.47% for CEO at 50 µg/mL and 19.56% for SyEO at 150 µg/mL. This study shows the potential of C. elegans as a predictive in vivo model of anti-colonizing activities that is suitable for the evaluation of essential oils.

Relationship of biofilm-forming ability of with swimming motility, twitching motility and virulence gene distribution.

To investigate the relationship of biofilm-forming ability of (PA) with swimming motility, twitching motility and virulence gene distribution. A total of 192 clinical isolates of PA were collected consecutively. Microtiter plate method was used to evaluate the ability to form biofilm. The swimming and twitching motilities were detected by plate method. Polymerase chain reaction (PCR) was used to detect virulence genes. Of the 192 PA clinical isolates, 186 (96.9%) showed biofilm-forming ability. Among them, 36 isolates showed weak biofilm-forming ability, 84 exhibited moderate biofilm-forming ability and 66 showed strong biofilm-forming ability. The diameters of the swimming ring for PA with none biofilm-forming ability, weak biofilm-forming ability, moderate biofilm-forming ability, strong biofilm-forming ability were (9.12±6.76), (18.42±7.51), (19.10±4.77) and respectively. The diameters of the twitching ring for PA in above groups were (8.38±1.50), (17.21±7.42), (18.49±5.62) and respectively. The swimming motility and twitching motility of none biofilm-forming ability group were weaker than biofilm-forming ability groups (all <0.05). Among 192 PA strains, 163 were positive (84.9%), 40 were positive (20.8%), 183 were positive (95.3%), and 189 were positive (98.4%). The positive rate of PA virulence gene , and were different in strains with different biofilm-forming abilities (<0.05). The rate of in the strong biofilm-forming ability group was lower than that in the moderate biofilm-forming ability group (=9.293, <0.01) and the weak biofilm-forming ability group (=9.997, <0.01). The rate of in the strong biofilm-forming ability group was higher than that in the weak biofilm-forming ability group (=10.803, <0.01). Most clinical isolates of PA can form biofilm. Swimming and twitching motilities are related to the formation of biofilm, but not significantly related to strength of biofilm-forming ability. The virulence genes of type Ⅲ secretion system for PA may be related to the biofilm-forming ability.

Inhibitory effects of 1,3-diaminopropane on the biofilm formation of Pseudomonas aeruginosa via interaction with quorum sensing system. / 1,3-丙二胺通过群体密度感应系统对铜绿假单胞菌生物膜形成的抑制作用.

OBJECTIVES: To study the inhibitory effects of 1,3-diaminopropane on the biofilm formation of Pseudomonas aeruginosa and the underlying mechanisms. METHODS: The experiment was divided into an experimental group and a control group. Crystal violet staining was used to examine the inhibitory effects of 1,3-diaminopropane on the biofilm formation of Pseudomonas aeruginosa, and the biofilm formation was compared between the 2 groups.Initial adherence inhibition assay and swimming plate assay were used to determine the inhibitory effects of 1,3-diaminopropane on the initial adherence and swimming motility of Pseudomonas aeruginosa,and the quantification of adhered cells and swimming diameter were compared between the 2 groups. Meanwhile, Western blotting was used to detect the Flagellin production of Pseudomonas aeruginosa; real-time RT-PCR was used to detect the quorum sensing system relative genes and flagellum regulative related genes expression in the 2 groups. Finally, molecular docking assay was used to calculate the interaction between 1,3-diaminopropane and LasI. RESULTS: Compared with the control group, the biofilm formation of Pseudomonas aeruginosa was significantly inhibited in the experimental group in a dose-dependent manner (t=6.07, P<0.01).Compared with the control group, the initial adherence of Pseudomonas aeruginosa could significantly inhibit from (0.890±0.389)×106 to (0.245±0.076)×106 CFU/mL (t=3.257, P<0.05) in the experimental group (2.0 mmol/L).Compared with the control group, the swimming motility of Pseudomonas aeruginosa flagellar mediation could also inhibit in the experimental group (2.0 mmol/L). The swimming motility diameter was from (1.840±0.144) to (0.756±0.222) cm (t=7.099, P<0.01). Compared with the control group, the Flagellin production was inhibited in the experimental group. Finally, the molecular docking assay showed that the potential target of 1,3-diaminopropane was LasI. CONCLUSIONS: 1,3-diaminopropane can significantly inhibit the biofilm formation of Pseudomonas aeruginosa, which potentially targets LasI protein.

The LysR-Type Transcriptional Regulator CrgA Negatively Regulates the Flagellar Master Regulator flhDC in Ralstonia solanacearum GMI1000.

The invasion and colonization of host plants by the destructive pathogen Ralstonia solanacearum rely on its cell motility, which is controlled by multiple factors. Here, we report that the LysR-type transcriptional regulator CrgA (RS_RS16695) represses cell motility in R. solanacearum GMI1000. CrgA possesses common features of a LysR-type transcriptional regulator and contains an N-terminal helix-turn-helix motif as well as a C-terminal LysR substrate-binding domain. Deletion of crgA results in an enhanced swim ring and increased transcription of flhDC In addition, the ΔcrgA mutant possesses more polar flagella than wild-type GMI1000 and exhibits higher expression of the flagellin gene fliC Despite these alterations, the ΔcrgA mutant did not have a detectable growth defect in culture. Yeast one-hybrid and electrophoretic mobility shift assays revealed that CrgA interacts directly with the flhDC promoter. Expressing the ß-glucuronidase (GUS) reporter under the control of the crgA promoter showed that crgA transcription is dependent on cell density. Soil-soaking inoculation with the crgA mutant caused wilt symptoms on tomato (Solanum lycopersicum L. cv. Hong yangli) plants earlier than inoculation with the wild-type GMI1000 but resulted in lower disease severity. We conclude that the R. solanacearum regulator CrgA represses flhDC expression and consequently affects the expression of fliC to modulate cell motility, thereby conditioning disease development in host plants.IMPORTANCERalstonia solanacearum is a widely distributed soilborne plant pathogen that causes bacterial wilt disease on diverse plant species. Motility is a critical virulence attribute of R. solanacearum because it allows this pathogen to efficiently invade and colonize host plants. In R. solanacearum, motility-defective strains are markedly affected in pathogenicity, which is coregulated with multiple virulence factors. In this study, we identified a new LysR-type transcriptional regulator (LTTR), CrgA, that negatively regulates motility. The mutation of the corresponding gene leads to the precocious appearance of wilt symptoms on tomato plants when the pathogen is introduced using soil-soaking inoculation. This study indicates that the regulation of R. solanacearum motility is more complex than previously thought and enhances our understanding of flagellum regulation in R. solanacearum.

The cold shock family gene cspD3 is involved in the pathogenicity of Ralstonia solanacearum CQPS-1 to tobacco.

Cold shock proteins (Csps) are small and highly conserved proteins that have target RNA- and DNA-binding activities. Csps play roles in different cellular processes and show functional redundancy. Ralstonia solanacearum, the agent of bacterial wilt, has 4 or 5 Csps based on genome analysis. However, the functions of all Csps in R. solanacearum remain unclear. According to phylogenetic analysis, the Csps from R. solanacearum are clustered into a group with CspD from E. coli. Here, we studied the role of CspD3, which was closer to CspD of E. coli in the phylogenetic tree. A cspD3 deletion strain was constructed to assess its effect on the phenotype of R. solanacearum, including growth, biofilm formation, motility, and virulence. The results showed that cspD3 of R. solanacearum was not necessary for normal growth, cold-shock adaptation, or biofilm formation. However, deletion of cspD3 in R. solanacearum CQPS-1 led to increased swimming motility, and the mean diameters of swimming haloes produced by the ΔcspD3 mutant were 1.3-fold larger than those produced by wild-type strain and 1.2-fold larger than those produced by the complemented strain. More importantly, the virulence of the cspD3 deletion mutant on susceptible tobacco plants was significantly attenuated compared to the wild-type strain. At 20 days after inoculation, the disease index of the ΔcspD3 mutant was 2.27, which was reduced by 1.6-fold relative to the wild-type strain. To assess the molecular response influenced by cspD3, the expressions of the main motility-associated genes and virulence-associated genes including flgM, fliA, pehS, pehR, hrpG, xpsR, and prhI in R. solanacearum were measured. The results showed that the expressions of hrpG, xpsR, and prhI were significantly decreased in cspD3 deletion mutant. Collectively, our findings showed that Csps are involved in the regulation of motility and virulence in R. solanacearum.

Identification of the genes controlling biofilm formation in the plant commensal Pseudomonas protegens Pf-5.

Determinant genes controlling biofilm formation in a plant commensal bacterium, Pseudomonas protegens Pf-5, were identified by transposon mutagenesis. Comprehensive screening of 7500 transposon-inserted mutants led to the isolation of four mutants exhibiting decreased and five mutants exhibiting increased biofilm formation. Mutations in the genes encoding MFS drug resistance transporter, LapA adhesive protein, RetS sensor histidine kinase/response regulator, and HecA adhesin/hemagglutinin led to decreased biofilm formation, indicating that these genes are necessary for biofilm formation in Pf-5. The mutants exhibiting increased biofilm formation had transposon insertions in the genes coding for an outer membrane protein, a GGDEF domain-containing protein, AraC transcriptional regulator, non-ribosomal peptide synthetase OfaB, and the intergenic region of a DNA-binding protein and the Aer aerotaxis receptor, suggesting that these genes are negative regulators of biofilm formation. Some of these mutants also showed altered swimming and swarming motilities, and a negative correlation between biofilm formation and swarming motility was observed. Thus, sessile-motile lifestyle is regulated by divergent regulatory genes in Pf-5.

Evaluation of growth and motility in non-photosynthetic Azospirillum brasilense exposed to red, blue, and white light.

Azospirillum brasilense is a non-photosynthetic rhizobacterium that promotes the growth of plants. In this work, we evaluated the effects of different light qualities on the growth, viability, and motility in combination to other culture conditions such as temperature or composition of the culture medium. Exponential cultures of A. brasilense Az39 were inoculated by drop-plate method on nutritionally rich (LB) or chemically defined (MMAB) media in the presence or absence of Congo Red indicator (CR) and exposed continuously to white light (WL), blue light (BL), and red light (RL), or maintained in dark conditions (control). The exposure to BL or WL inhibited growth, mostly in LB medium at 36 °C. By contrast, the exposure to RL showed a similar behavior to the control. Swimming motility was inhibited by exposure to WL and BL, while exposure to RL caused only a slight reduction. The effects of WL and BL on plant growth-promoting rhizobacteria should be considered in the future as deleterious factors that could be manipulated to improve the functionality of foliar inoculants, as well as the bacterial effects on the leaf after inoculation.