Shewanella oneidensis MR‐1 mutants selected for their inability to produce soluble organic‐Fe(III) complexes are unable to respire Fe(III) as anaerobic electron acceptor

Summary Recent voltammetric analyses indicate that Shewanella putrefaciens strain 200 produces soluble organic‐Fe(III) complexes during anaerobic respiration of sparingly soluble Fe(III) oxides. Results of the present study expand the range of Shewanella species capable of producing soluble organic‐Fe(III) complexes to include Shewanella oneidensis MR‐1. Soluble organic‐Fe(III) was produced by S. oneidensis cultures incubated anaerobically with Fe(III) oxides, or with Fe(III) oxides and the alternate electron acceptor fumarate, but not in the presence of O₂, nitrate or trimethylamine‐N‐oxide. Chemical mutagenesis procedures were combined with a novel MicroElectrode Screening Array (MESA) to identify four (designated Sol) mutants with impaired ability to produce soluble organic‐Fe(III) during anaerobic respiration of Fe(III) oxides. Two of the Sol mutants were deficient in anaerobic growth on both soluble Fe(III)‐citrate and Fe(III) oxide, yet retained the ability to grow on a suite of seven alternate electron acceptors. The rates of soluble organic‐Fe(III) production were proportional to the rates of iron reduction by the S. oneidensis wild‐type and Sol mutant strains, and all four Sol mutants retained wild‐type siderophore production capability. Results of this study indicate that the production of soluble organic‐Fe(III) may be an important intermediate step in the anaerobic respiration of both soluble and sparingly soluble forms of Fe(III) by S. oneidensis.     

Disruption of Putrescine Biosynthesis in Shewanella oneidensis Enhances Biofilm Cohesiveness and Performance in Cr(VI) Immobilization

Although biofilm-based bioprocesses have been increasingly used in various applications, the long-term robust and efficient biofilm performance remains one of the main bottlenecks. In this study, we demonstrated that biofilm cohesiveness and performance of Shewanella oneidensis can be enhanced through disrupting putrescine biosynthesis. Through random transposon mutagenesis library screening, one hyperadherent mutant strain, CP2-1-S1, exhibiting an enhanced capability in biofilm formation, was obtained. Comparative analysis of the performance of biofilms formed by S. oneidensis MR-1 wild type (WT) and CP2-1-S1 in removing dichromate (Cr₂O₇²⁻), i.e., Cr(VI), from the aqueous phase showed that, compared with the WT biofilms, CP2-1-S1 biofilms displayed a substantially lower rate of cell detachment upon exposure to Cr(VI), suggesting a higher cohesiveness of the mutant biofilms. In addition, the amount of Cr(III) immobilized by CP2-1-S1 biofilms was much larger, indicating an enhanced performance in Cr(VI) bioremediation. We further showed that speF, a putrescine biosynthesis gene, was disrupted in CP2-1-S1 and that the biofilm phenotypes could be restored by both genetic and chemical complementations. Our results also demonstrated an important role of putrescine in mediating matrix disassembly in S. oneidensis biofilms.     

Elevated intracellular cyclic-di-GMP level in Shewanella oneidensis increases expression of c-type cytochromes

Electrochemically active biofilms are capable of exchanging electrons with solid electron acceptors and have many energy and environmental applications such as bioelectricity generation and environmental remediation. The performance of electrochemically active biofilms is usually dependent on c-type cytochromes, while biofilm development is controlled by a signal cascade mediated by the intracellular secondary messenger bis-(3ʹ-5ʹ) cyclic dimeric guanosine monophosphate (c-di-GMP). However, it is unclear whether there are any links between the c-di-GMP regulatory system and the expression of c-type cytochromes. In this study, we constructed a S. oneidensis MR-1 strain with a higher cytoplasmic c-di-GMP level by constitutively expressing a c-di-GMP synthase and it exhibited expected c-di-GMP-influenced traits, such as lowered motility and increased biofilm formation. Compared to MR-1 wild-type strain, the high c-di-GMP strain had a higher Fe(III) reduction rate (21.58 vs 11.88 pM of Fe(III)/h cell) and greater expression of genes that code for the proteins involved in the Mtr pathway, including CymA, MtrA, MtrB, MtrC and OmcA. Furthermore, single-cell Raman microspectroscopy (SCRM) revealed a great increase of c-type cytochromes in the high c-di-GMP strain as compared to MR-1 wild-type strain. Our results reveal for the first time that the c-di-GMP regulation system indirectly or directly positively regulates the expression of cytochromes involved in the extracellular electron transport (EET) in S. oneidensis, which would help to understand the regulatory mechanism of c-di-GMP on electricity production in bacteria.     

信号分子AI-2合成关键基因luxS对副溶血性弧菌生物学特性的影响

[背景]副溶血性弧菌是全球范围重要的食源性病原菌,能引起急性肠胃炎。群体感应系统LuxS/AI-2影响细菌的生物学特性,为研究副溶血性弧菌的传播机制和控制技术提供了新的途径。[目的]探讨群体感应信号分子AI-2合成关键基因luxS对海产品中分离的副溶血性弧菌Vp2009027生物学特性的影响。[方法]利用自杀质粒同源重组技术敲除信号分子AI-2合成关键基因luxS,构建副溶血性弧菌Vp2009027的luxS基因缺失株,通过比较野生株与luxS基因缺失株的生长曲线、AI-2活性、运动能力、生物膜形成能力和耐药性,分析LuxS/AI-2系统对副溶血性弧菌生物学特性的影响。[结果]构建了副溶血性弧菌Vp2009027的luxS基因缺失株,野生株和luxS基因缺失株的生长无明显差异,luxS基因的缺失导致AI-2合成受阻、运动能力和生物膜形成能力增强、四环素耐药性降低。[结论]luxS基因对副溶血性弧菌的生物学特性具有重要的调控作用,为进一步研究副溶血性弧菌的传播机制和研发控制技术提供基础。     

LuxS-Based Signaling Affects Streptococcus mutans Biofilm Formation

Streptococcus mutans is implicated as a major etiological agent in human dental caries, and one of the important virulence properties of this organism is its ability to form biofilms (dental plaque) on tooth surfaces. We examined the role of autoinducer-2 (AI-2) on S. mutans biofilm formation by constructing a GS-5 luxS-null mutant. Biofilm formation by the luxS mutant in 0.5% sucrose defined medium was found to be markedly attenuated compared to the wild type. Scanning electron microscopy also revealed that biofilms of the luxS mutant formed larger clumps in sucrose medium compared to the parental strain. Therefore, the expression of glucosyltransferase genes was examined and the gtfB and gtfC genes, but not the gtfD gene, in the luxS mutant were upregulated in the mid-log growth phase. Furthermore, we developed a novel two-compartment system to monitor AI-2 production by oral streptococci and periodontopathic bacteria. The biofilm defect of the luxS mutant was complemented by strains of S. gordonii, S. sobrinus, and S. anginosus; however, it was not complemented by S. oralis, S. salivarius, or S. sanguinis. Biofilm formation by the luxS mutant was also complemented by Porphyromonas gingivalis 381 and Actinobacillus actinomycetemcomitans Y4 but not by a P. gingivalis luxS mutant. These results suggest that the regulation of the glucosyltransferase genes required for sucrose-dependent biofilm formation is regulated by AI-2. Furthermore, these results provide further confirmation of previous proposals that quorum sensing via AI-2 may play a significant role in oral biofilm formation.     

Impact of luxS and Suppressor Mutations on the Gastrointestinal Transit of Lactobacillus rhamnosus GG

It is generally believed that probiotic bacteria need to survive gastrointestinal transit to exert a health-promoting effect. In this study, a genuine luxS mutant and a luxS mutant containing unknown suppressor mutations of the probiotic strain Lactobacillus rhamnosus GG were compared to the wild type for survival and persistence in the murine gastrointestinal tract. The LuxS enzyme, catalyzing the production of the autoinducer-2 signaling molecule, also forms an integral part of the activated methyl cycle and the metabolism of methionine and cysteine. The genuine luxS mutant CMPG5412 showed drastically reduced persistence in mice, which was related to less survival in simulated gastric juice, indicating that LuxS metabolism is crucial for the gastric stress resistance of L. rhamnosus GG. The suppressor mutations in the other luxS mutant, CMPG5413, appear to compensate for the metabolic defects of the luxS mutation and to restore the resistance to gastric juice but cause a defect in adherence, biofilm formation, and exopolysaccharide production. The shorter residence time of this suppressor mutant in the murine gastrointestinal tract indicates a role for biofilm formation and exopolysaccharides in the persistence capacity of L. rhamnosus GG.     

The small regulatory RNA molecule MicA is involved in <Emphasis Type="Italic">Salmonella enterica</Emphasis> serovar Typhimurium biofilm formation

Background  

LuxS is the synthase enzyme of the quorum sensing signal AI-2. In Salmonella Typhimurium, it was previously shown that a luxS deletion mutant is impaired in biofilm formation. However, this phenotype could not be complemented by extracellular addition of quorum sensing signal molecules.     

Involvement of luxS in Biofilm Formation by Capnocytophaga ochracea

Capnocytophaga ochracea is present in the dental plaque biofilm of patients with periodontitis. Biofilm cells change their phenotype through quorum sensing in response to fluctuations in cell-population density. Quorum sensing is mediated by auto-inducers (AIs). AI-2 is involved in intercellular signaling, and production of its distant precursor is catalyzed by LuxS, an enzyme involved in the activated methyl cycle. Our aim was to clarify the role of LuxS in biofilm formation by C. ochracea. Two luxS-deficient mutants, TmAI2 and LKT7, were constructed from C. ochracea ATCC 27872 by homologous recombination. The mutants produced significantly less AI-2 than the wild type. The growth rates of these mutants were similar to that of the wild-type in both undiluted Tryptic soy broth and 0.5 × Tryptic soy broth. However, according to crystal violet staining, they produced significantly less biofilm than the wild type. Confocal laser scanning microscopy and scanning electron microscopy showed that the biofilm of the TmAI2 strain had a rougher structure than that of the wild type. Complementation of TmAI-2 with extrinsic AI-2 from the culture supernatant of wild-type strain did not restore biofilm formation by the TmAI2 strain, but complementation of LKT7 strain with luxS partially restored biofilm formation. These results indicate that LuxS is involved in biofilm formation by C. ochracea, and that the attenuation of biofilm formation by the mutants is likely caused by a defect in the activated methyl cycle rather than by a loss of AI-2.     

Shewanella oneidensis MR-1 Uses Overlapping Pathways for Iron Reduction at a Distance and by Direct Contact under Conditions Relevant for Biofilms

We developed a new method to measure iron reduction at a distance based on depositing Fe(III) (hydr)oxide within nanoporous glass beads. In this “Fe-bead” system, Shewanella oneidensis reduces at least 86.5% of the iron in the absence of direct contact. Biofilm formation accompanies Fe-bead reduction and is observable both macro- and microscopically. Fe-bead reduction is catalyzed by live cells adapted to anaerobic conditions, and maximal reduction rates require sustained protein synthesis. The amount of reactive ferric iron in the Fe-bead system is available in excess such that the rate of Fe-bead reduction is directly proportional to cell density; i.e., it is diffusion limited. Addition of either lysates prepared from anaerobic cells or exogenous electron shuttles stimulates Fe-bead reduction by S. oneidensis, but iron chelators or additional Fe(II) do not. Neither dissolved Fe(III) nor electron shuttling activity was detected in culture supernatants, implying that the mediator is retained within the biofilm matrix. Strains with mutations in omcB or mtrB show about 50% of the wild-type levels of reduction, while a cymA mutant shows less than 20% of the wild-type levels of reduction and a menF mutant shows insignificant reduction. The Fe-bead reduction defect of the menF mutant can be restored by addition of menaquinone, but menaquinone itself cannot stimulate Fe-bead reduction. Because the menF gene encodes the first committed step of menaquinone biosynthesis, no intermediates of the menaquinone biosynthetic pathway are used as diffusible mediators by this organism to promote iron reduction at a distance. CymA and menaquinone are required for both direct and indirect mineral reduction, whereas MtrB and OmcB contribute to but are not absolutely required for iron reduction at a distance.     

Ecological Behavior of Lactobacillus reuteri 100-23 Is Affected by Mutation of the luxS Gene

The luxS gene of Lactobacillus reuteri 100-23C was amplified by PCR, cloned, and then sequenced. To define a physiological and ecological role for the luxS gene in L. reuteri 100-23C, a luxS mutant was constructed by insertional mutagenesis. The luxS mutant did not produce autoinducers AI-2 or AI-3. Complementation of the luxS mutation by a plasmid construct containing luxS restored AI-2 and AI-3 synthesis. In vitro experiments revealed that neither the growth rate, nor the cell yield, nor cell survival in the stationary phase were compromised in the luxS mutant relative to the wild type and complemented mutant. The ATP content of exponentially growing cells of the luxS mutant was, however, 65% of that of wild-type cells. Biofilms formed by the luxS mutant on plastic surfaces in a bioreactor were thicker than those formed by the wild type. Biofilm thickness was not restored to wild-type values by the addition of purified AI-2 to the culture medium. In vivo experiments, conducted with ex-Lactobacillus-free mice, showed that biofilms formed by the mutant strain on the epithelial surface of the forestomach were approximately twice as thick as those formed by the wild type. The ecological performance of the luxS mutant, when in competition with L. reuteri strain 100-93 in the mouse cecum, was reduced compared to that of a xylA mutant of 100-23C. These results demonstrate that LuxS influences important ecological attributes of L. reuteri 100-23C, the consequences of which are niche specific.     

Characterization of the <Emphasis Type="Italic">codY</Emphasis> gene and its influence on biofilm formation in <Emphasis Type="Italic">Bacillus </Emphasis><Emphasis Type="Italic">cereus</Emphasis>

The foodborne pathogen Bacillus cereus can form biofilms on various food contact surfaces, leading to contamination of food products. To study the mechanisms of biofilm formation by B. cereus, a Tn5401 library was generated from strain UW101C. Eight thousand mutants were screened in EPS, a low nutrient medium. One mutant (M124), with a disruption in codY, developed fourfold less biofilm than the wild-type, and its defective biofilm phenotype was rescued by complementation. Addition of 0.1% casamino acids to EPS prolonged the duration of biofilms in the wild-type but not codY mutant. When decoyinine, a GTP synthesis inhibitor, was added to EPS, biofilm formation was decreased in the wild-type but not the mutant. The codY mutant produced three times higher protease activity than the wild-type. Zymogram and SDS-PAGE data showed that production of the protease (∼130 kDa) was repressed by CodY. Addition of proteinase K to EPS decreased biofilm formation by the wild-type. Using a dpp-lacZ fusion reporter system, it was shown that that the B. cereus CodY can sense amino acids and GTP levels. These data suggest that by responding to amino acids and intracellular GTP levels CodY represses production of an unknown protease and is involved in biofilm formation.     

LuxS-mediated signalling in Streptococcus anginosus and its role in biofilm formation

The autoinducer-2 signal (AI-2) produced by several Gram-positive and Gram-negative bacteria mediates interspecies communication. In this study we were able to identify an orthologue of luxS, required for the synthesis of AI-2 signals, in Streptococcus anginosus. Comparative analyses revealed conserved sequences in the predicted S. anginosus LuxS. Expression of luxS was highest during early exponential growth phase. Compared to other oral streptococci, conditioned media from growth of members of the anginosus group were the most efficient in inducing bioluminescence in Vibrio harveyi, indicative of AI-2 signalling. Disruption of luxS in S. anginosus resulted in a mutant deficient in biofilm formation, whereas no effect on planktonic growth rate was observed under various growth conditions. S. anginosus is part of the human flora found in biofilms of the oral cavity, as well as of the upper respiratory, gastrointestinal and urogenital tracts. Such habitats harbour large varieties of bacterial species, among which cell–cell communication may␣play an important role. S. anginosus has also been associated with purulent infections and cancer in the upper digestive tract. Knowledge about the molecular mechanisms involved in S.␣anginosus communication is important for understanding its commensalism and its pathogenic transition.     

Siderophores Are Not Involved in Fe(III) Solubilization during Anaerobic Fe(III) Respiration by Shewanella oneidensis MR-1

Shewanella oneidensis MR-1 respires a wide range of anaerobic electron acceptors, including sparingly soluble Fe(III) oxides. In the present study, S. oneidensis was found to produce Fe(III)-solubilizing organic ligands during anaerobic Fe(III) oxide respiration, a respiratory strategy postulated to destabilize Fe(III) and produce more readily reducible soluble organic Fe(III). In-frame gene deletion mutagenesis, siderophore detection assays, and voltammetric techniques were combined to determine (i) if the Fe(III)-solubilizing organic ligands produced by S. oneidensis during anaerobic Fe(III) oxide respiration were synthesized via siderophore biosynthesis systems and (ii) if the Fe(III)-siderophore reductase was required for respiration of soluble organic Fe(III) as an anaerobic electron acceptor. Genes predicted to encode the siderophore (hydroxamate) biosynthesis system (SO3030 to SO3032), the Fe(III)-hydroxamate receptor (SO3033), and the Fe(III)-hydroxamate reductase (SO3034) were identified in the S. oneidensis genome, and corresponding in-frame gene deletion mutants were constructed. ΔSO3031 was unable to synthesize siderophores or produce soluble organic Fe(III) during aerobic respiration yet retained the ability to solubilize and respire Fe(III) at wild-type rates during anaerobic Fe(III) oxide respiration. ΔSO3034 retained the ability to synthesize siderophores during aerobic respiration and to solubilize and respire Fe(III) at wild-type rates during anaerobic Fe(III) oxide respiration. These findings indicate that the Fe(III)-solubilizing organic ligands produced by S. oneidensis during anaerobic Fe(III) oxide respiration are not synthesized via the hydroxamate biosynthesis system and that the Fe(III)-hydroxamate reductase is not essential for respiration of Fe(III)-citrate or Fe(III)-nitrilotriacetic acid (NTA) as an anaerobic electron acceptor.Bacterial electron transfer to sparingly soluble electron acceptors is a critical component of a wide variety of environmental and energy-generating processes, including biogeochemical cycling of metals, degradation of natural and contaminant organic matter, weathering of clays and minerals, biomineralization of Fe-bearing minerals, reductive precipitation of toxic metals and radionuclides, and electricity generation in microbial fuel cells (17, 33, 34). Anaerobic and facultatively anaerobic bacteria capable of respiring sparingly soluble (<10⁻²⁵ M at pH 7) Fe(III) oxides are ubiquitous in nature and may be found in marine, freshwater, and terrestrial environments, including metal- and radionuclide-contaminated subsurface aquifers (25, 34). Fe(III)-respiring prokaryotes are also deeply rooted and scattered throughout the domains Bacteria and Archaea (possibly indicating an ancient metabolic process) and include hyperthermophiles, psychrophiles, acidophiles, and extreme barophiles (34). Despite their potential environmental, energy-generating, and evolutionary significance, the molecular details of microbial Fe(III) respiration remain unclear.Fe(III)-respiring, neutrophilic bacteria are presented with a unique physiological challenge: they are required to respire anaerobically on electron acceptors found largely as sparingly soluble Fe(III) oxides presumably unable to contact periplasm- or inner membrane (IM)-localized electron transport systems. To overcome this problem, Fe(III)-respiring bacteria are postulated to employ novel respiratory strategies not found in other bacteria (e.g., aerobes, denitrifiers, sulfate-reducing bacteria, and methanogens) that respire soluble electron acceptors (17, 38). The novel respiratory strategies include (i) a direct-contact pathway in which terminal Fe(III) reductases are secreted to the cell outer membrane (OM), where they contact and deliver electrons directly to external Fe(III) oxides (18, 23, 40, 42, 48, 57, 64, 67), (ii) a two-step electron shuttling pathway in which bacterially reduced endogenous or exogenous electron shuttles deliver electrons to external Fe(III) oxides in a second (abiotic) electron transfer reaction (11, 26, 39, 45), and (iii) a two-step Fe(III) chelation (solubilization) pathway in which Fe(III) oxides are first nonreductively dissolved by endogenously synthesized organic ligands prior to reduction of the resulting soluble organic Fe(III) [Fe(III) bound to an organic molecule] complexes (36, 59).Candidate organic ligands for production of soluble organic Fe(III) during anaerobic Fe(III) oxide respiration include siderophores, the Fe(III)-chelating compounds synthesized and secreted by a wide variety of bacteria and fungi for solubilization and subsequent assimilation of otherwise inaccessible Fe(III) substrates (12, 44, 49, 63). Hydroxamate-type siderophores are produced via N⁶ hydroxylation and N⁶ acylation of l-ornithine and, in some cases, cyclization to macrocyclic ring structures (13). The macrocyclic siderophores bisucaberin and putrebactin, for example, are two structural analogs of the cyclic bis(hydroxamate) siderophore alcaligin, synthesized by Aliivibrio salmonicida and Shewanella putrefaciens strain 200, respectively (27, 32, 65). After transport across the cell envelope via a TonB-dependent pathway, Fe(III) is subsequently released from the Fe(III)-siderophore complex by ligand exchange reactions promoted by siderophore ligand hydrolysis and/or protonation or by Fe(III)-siderophore reduction and release of Fe(II) to acceptor ligands (9, 66).The main objectives of the present study were to determine (i) if the Fe(III)-solubilizing organic ligands produced by S. oneidensis during anaerobic Fe(III) oxide respiration are synthesized by Fe(III)-siderophore biosynthesis systems and (ii) if Fe(III)-siderophore reductases are required for respiration of soluble organic Fe(III) as an anaerobic electron acceptor. The experimental strategy for this study included (i) identification of genes encoding the siderophore biosynthesis and Fe(III)-siderophore reductase systems in the S. oneidensis genome, (ii) generation of in-frame deletions in the corresponding siderophore biosynthesis and Fe(III)-siderophore reductase genes, (iii) tests of the resulting siderophore biosynthesis mutants for production of siderophores and soluble organic Fe(III) during aerobic and anaerobic Fe(III) oxide respiration, and (iv) tests of the resulting Fe(III)-siderophore reductase mutants for respiration of soluble organic Fe(III) as an anaerobic electron acceptor.     

LapG mediates biofilm dispersal in Vibrio fischeri by controlling maintenance of the VCBS-containing adhesin LapV

Efficient symbiotic colonization of the squid Euprymna scolopes by the bacterium Vibrio fischeri depends on bacterial biofilm formation on the surface of the squid’s light organ. Subsequently, the bacteria disperse from the biofilm via an unknown mechanism and enter through pores to reach the interior colonization sites. Here, we identify a homolog of Pseudomonas fluorescens LapG as a dispersal factor that promotes cleavage of a biofilm-promoting adhesin, LapV. Overproduction of LapG inhibited biofilm formation and, unlike the wild-type parent, a ΔlapG mutant formed biofilms in vitro. Although V. fischeri encodes two putative large adhesins, LapI (near lapG on chromosome II) and LapV (on chromosome I), only the latter contributed to biofilm formation. Consistent with the Pseudomonas Lap system model, our data support a role for the predicted c-di-GMP-binding protein LapD in inhibiting LapG-dependent dispersal. Furthermore, we identified a phosphodiesterase, PdeV, whose loss promotes biofilm formation similar to that of the ΔlapG mutant and dependent on both LapD and LapV. Finally, we found a minor defect for a ΔlapD mutant in initiating squid colonization, indicating a role for the Lap system in a relevant environmental niche. Together, these data reveal new factors and provide important insights into biofilm dispersal by V. fischeri.     

Going Wireless: Fe(III) Oxide Reduction without Pili by Geobacter sulfurreducens Strain JS-1

Previous studies have suggested that the conductive pili of Geobacter sulfurreducens are essential for extracellular electron transfer to Fe(III) oxides and for optimal long-range electron transport through current-producing biofilms. The KN400 strain of G. sulfurreducens reduces poorly crystalline Fe(III) oxide more rapidly than the more extensively studied DL-1 strain. Deletion of the gene encoding PilA, the structural pilin protein, in strain KN400 inhibited Fe(III) oxide reduction. However, low rates of Fe(III) reduction were detected after extended incubation (>30 days) in the presence of Fe(III) oxide. After seven consecutive transfers, the PilA-deficient strain adapted to reduce Fe(III) oxide as fast as the wild type. Microarray, whole-genome resequencing, proteomic, and gene deletion studies indicated that this adaptation was associated with the production of larger amounts of the c-type cytochrome PgcA, which was released into the culture medium. It is proposed that the extracellular cytochrome acts as an electron shuttle, promoting electron transfer from the outer cell surface to Fe(III) oxides. The adapted PilA-deficient strain competed well with the wild-type strain when both were grown together on Fe(III) oxide. However, when 50% of the culture medium was replaced with fresh medium every 3 days, the wild-type strain outcompeted the adapted strain. A possible explanation for this is that the necessity to produce additional PgcA, to replace the PgcA being continually removed, put the adapted strain at a competitive disadvantage, similar to the apparent selection against electron shuttle-producing Fe(III) reducers in many anaerobic soils and sediments. Despite increased extracellular cytochrome production, the adapted PilA-deficient strain produced low levels of current, consistent with the concept that long-range electron transport through G. sulfurreducens biofilms is more effective via pili.     

Changes in physico-chemical characteristics and viable bacterial communities during fermentation of alfalfa silages inoculated with Lactobacillus plantarum

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.

     

luxS基因缺失对禽致病性大肠杆菌O1、O2和O78三种血清型分离株的生物学特性影响

【目的】LuxS/AI-2型密度群体感应系统产生的自诱导信号分子AI-2(AI-2的产生需要luxS基因编码的Lux S蛋白参与)参与对细菌众多生理功能的调控。探讨luxS对不同血清型禽致病性大肠杆菌(Avian Pathogenicity Escherichia coli,APEC)生物学特性的影响。【方法】本研究以APEC优势血清型APECO_1(O_1血清型)、DE17(O_2血清型)、E940(O_(78)血清型)及其相应luxS缺失株为研究对象,对野生株和缺失株的生长特性、生物被膜形成、rdar(red,dry and rough)形态、运动性和耐药性等特性进行分析。【结果】luxS基因的缺失不影响APEC生长特性,但导致APEC不能产生AI-2;此外,luxS基因的缺失显著降低APECO_1和E940的生物被膜形成(P0.05),而DE17的生物被膜形成无显著变化。对各菌株的rdar形态和运动性检测结果表明,luxS基因的缺失改变了APECO_1的rdar形态,对DE17和E940并无影响;显著降低了APECO_1和DE17运动能力,对E940并无影响。荧光定量PCR检测结果表明,luxS基因的缺失显著降低APECO_1、DE17和E940与细菌运动性相关的鞭毛基因fli G和fli I的转录水平(P0.05)。此外,对各菌株的耐药性检测结果表明,luxS基因缺失导致APECO_1对头孢吡肟和丁胺卡那由耐药变为高敏,同时对氯霉素与E940相同由高敏变为耐药,但对DE17的耐药性无显著改变。【结论】luxS对APEC的生物学特性具有重要的调控作用,且这种调控具有菌株特异性。