Global and Phylogenetic Distribution of Quorum Sensing Signals,Acyl Homoserine Lactones,in the Family of Vibrionaceae

Bacterial quorum sensing (QS) and the corresponding signals, acyl homoserine lactones (AHLs), were first described for a luminescent Vibrio species. Since then, detailed knowledge has been gained on the functional level of QS; however, the abundance of AHLs in the family of Vibrionaceae in the environment has remained unclear. Three hundred and one Vibrionaceae strains were collected on a global research cruise and the prevalence and profile of AHL signals in this global collection were determined. AHLs were detected in 32 of the 301 strains using Agrobacterium tumefaciens and Chromobacterium violaceum reporter strains. Ethyl acetate extracts of the cultures were analysed by ultra-high performance liquid chromatography-high resolution mass spectrometry (MS) with automated tandem MS confirmation for AHLs. N-(3-hydroxy-hexanoyl) (OH-C6) and N-(3-hydroxy-decanoyl) (OH-C10) homoserine lactones were the most common AHLs found in 17 and 12 strains, respectively. Several strains produced a diversity of different AHLs, including N-heptanoyl (C7) HL. AHL-producing Vibrionaceae were found in polar, temperate and tropical waters. The AHL profiles correlated with strain phylogeny based on gene sequence homology, however not with geographical location. In conclusion, a wide range of AHL signals are produced by a number of clades in the Vibrionaceae family and these results will allow future investigations of inter- and intra-species interactions within this cosmopolitan family of marine bacteria.     

Quorum Quenching Agents: Resources for Antivirulence Therapy

The continuing emergence of antibiotic-resistant pathogens is a concern to human health and highlights the urgent need for the development of alternative therapeutic strategies. Quorum sensing (QS) regulates virulence in many bacterial pathogens, and thus, is a promising target for antivirulence therapy which may inhibit virulence instead of cell growth and division. This means that there is little selective pressure for the evolution of resistance. Many natural quorum quenching (QQ) agents have been identified. Moreover, it has been shown that many microorganisms are capable of producing small molecular QS inhibitors and/or macromolecular QQ enzymes, which could be regarded as a strategy for bacteria to gain benefits in competitive environments. More than 30 species of marine QQ bacteria have been identified thus far, but only a few of them have been intensively studied. Recent studies indicate that an enormous number of QQ microorganisms are undiscovered in the highly diverse marine environments, and these marine microorganism-derived QQ agents may be valuable resources for antivirulence therapy.     

Quorum Sensing Inhibition by Asparagopsis taxiformis,a Marine Macro Alga: Separation of the Compound that Interrupts Bacterial Communication

The majority of the marine algal species, though completing their life cycle in seawater, are rarely susceptible to fouling, making them an important source of quorum sensing (QS) inhibitory substances. The separation and characterization of QS inhibitors are crucial for any potential application. Thirty marine macroalgae were tested for QS inhibition activity by using Chromobacterium violaceum CV026 as the reporter strain, and among them, Asparagopsis taxiformis showed antibacterial, as well as antiquorum, sensing activities. Cinnamaldehyde (75 mM) and methanol were used as positive and negative controls, respectively. The antiquorum sensing activity of A. taxiformis was further confirmed using the sensor strain, Serratia liquefaciens MG44, having green fluorescent protein (gfp). Methanolic extract of the alga was fractionated by solid phase extraction (SPE), and each fraction was tested for QS inhibition. Two types of activities were observed—zone of clearance (antibacterial activity) and zone of inhibition with or without finger-like projections (QS inhibition). Out of five SPE cartridges, Bond Elut PH showed clear separation of these two fractions. The Ion Cyclotron Resonance Fourier Transformation Mass Spectrometer (ICR-FT/MS) analysis of the fractions further supported the bioassay results. The presence of strong QS inhibitory compound in A. taxiformis indicates its potential use in antifouling preparations.     

AhaP,A Quorum Quenching Acylase from Psychrobacter sp. M9-54-1 That Attenuates Pseudomonas aeruginosa and Vibrio coralliilyticus Virulence

Although Psychrobacter strain M9-54-1 had been previously isolated from the microbiota of holothurians and shown to degrade quorum sensing (QS) signal molecules C6 and C10-homoserine lactone (HSL), little was known about the gene responsible for this activity. In this study, we determined the whole genome sequence of this strain and found that the full 16S rRNA sequence shares 99.78–99.66% identity with Psychrobacter pulmonis CECT 5989^T and P. faecalis ISO-46^T. M9-54-1, evaluated using the agar well diffusion assay method, showed high quorum quenching (QQ) activity against a wide range of synthetic N-acylhomoserine lactone (AHLs) at 4, 15, and 28 °C. High-performance liquid chromatography-mass-spectrometry (HPLC-MS) confirmed that QQ activity was due to an AHL-acylase. The gene encoding for QQ activity in strain M9-54-1 was identified from its genome sequence whose gene product was named AhaP. Purified AhaP degraded substituted and unsubstituted AHLs from C4- to C14-HSL. Furthermore, heterologous expression of ahaP in the opportunistic pathogen Pseudomonas aeruginosa PAO1 reduced the expression of the QS-controlled gene lecA, encoding for a cytotoxic galactophilic lectin and swarming motility protein. Strain M9-54-1 also reduced brine shrimp mortality caused by Vibrio coralliilyticus VibC-Oc-193, showing potential as a biocontrol agent in aquaculture.     

Inhibition of Bacterial Quorum Sensing by Extracts from Aquatic Fungi: First Report from Marine Endophytes

In our search for quorum-sensing (QS) disrupting molecules, 75 fungal isolates were recovered from reef organisms (endophytes), saline lakes and mangrove rhizosphere. Their QS inhibitory activity was evaluated in Chromobacterium violaceum CVO26. Four strains of endophytic fungi stood out for their potent activity at concentrations from 500 to 50 μg mL⁻¹. The molecular characterization, based on the internal transcribed spacer (ITS) region sequences (ITS1, 5.8S and ITS2) between the rRNA of 18S and 28S, identified these strains as belonging to four genera: Sarocladium (LAEE06), Fusarium (LAEE13), Epicoccum (LAEE14), and Khuskia (LAEE21). Interestingly, three came from coral species and two of them came from the same organism, the coral Diploria strigosa. Metabolic profiles obtained by Liquid Chromatography-High Resolution Mass Spectrometry (LC-HRMS) suggest that a combination of fungal secondary metabolites and fatty acids could be the responsible for the observed activities. The LC-HRMS analysis also revealed the presence of potentially new secondary metabolites. This is, to the best of our knowledge, the first report of QS inhibition by marine endophytic fungi.     

Marine-Source Quorum Quenching Enzyme YtnP to Improve Hygiene Quality in Dental Units

Biofilm in dental unit water lines may pose a health risk to patients and dental practitioners. An AdiC-like quorum quenching enzyme, YtnP, was cloned from a deep-sea probiotic Bacillus velezensis, and heterologously expressed in E. coli to examine the application on the improvement of hygiene problems caused by biofilm infection of Pseudomonas aeruginosa in dental units. Pseudomonas bacteria were isolated from dental chair units and used to grow static biofilms in the laboratory. A water filter system was designed to test the antifouling activity of YtnP in Laboratory, to simulate the biofilm contamination on water filter in dental unit water lines. The results demonstrated that the enzyme of YtnP was able to degrade the N-acyl homoserine lactones, significantly inhibited the EPS generation, biofilm formation, and virulence factors production (pyocyanin and rhamnolipid) of P. aeruginosa, and was efficient on the antifouling against P. aeruginosa. The findings in this study indicated the possibility of YtnP as novel disinfectant reagent for hygiene treatment in dental units.     

EGCG抑制波罗的海希瓦氏菌生物被膜和腐败活性的研究

以海产品腐败菌波罗的海希瓦氏菌(Shewanella baltica)为对象,评价了儿茶素单体抑制生物被膜的效果,并分析效果最佳的单体对该菌生物被膜和腐败活性的影响。结果表明,5种儿茶素单体均能抑制S.baltica生物被膜形成,其中表没食子儿茶素没食子酸酯(Epigallocatechin gallate,EGCG)抑制率最高,其最小抑菌质量浓度为40μg·m L-1,亚抑菌质量浓度10~30μg·m L-1的EGCG对细菌生长无显著影响。在30μg·m L-1 EGCG作用下,群体感应(Quorum sensing,QS)信号分子呋喃酮酰硼酸二酯(Autoinducer-2,AI-2)、二酮哌嗪类化合物(Diketopiperazine,DKPs)cyclo-(L-Pro-L-Leu)和cyclo-(L-Pro-L-Phe)活性显著下降(P0.05),其抑制率分别为63.95%、27.43%和21.94%。亚抑菌浓度EGCG能显著抑制S.baltica生物被膜形成、泳动能力和蛋白酶活性(P0.05),呈现浓度依赖性,在30μg·m L-1时抑制率分别为55.18%,66.71%和46.67%。因此,EGCG在亚抑菌浓度下能够干扰S.baltica的QS系统,有效抑制生物被膜形成,减弱致腐能力,研究为EGCG作为新型的QS抑制剂调控食品腐败菌奠定基础。     

Emerging Concepts Promising New Horizons for Marine Biodiscovery and Synthetic Biology

The vast oceans of the world, which comprise a huge variety of unique ecosystems, are emerging as a rich and relatively untapped source of novel bioactive compounds with invaluable biotechnological and pharmaceutical potential. Evidence accumulated over the last decade has revealed that the diversity of marine microorganisms is enormous with many thousands of bacterial species detected that were previously unknown. Associated with this diversity is the production of diverse repertoires of bioactive compounds ranging from peptides and enzymes to more complex secondary metabolites that have significant bioactivity and thus the potential to be exploited for innovative biotechnology. Here we review the discovery and functional potential of marine bioactive peptides such as lantibiotics, nanoantibiotics and peptidomimetics, which have received particular attention in recent years in light of their broad spectrum of bioactivity. The significance of marine peptides in cell-to-cell communication and how this may be exploited in the discovery of novel bioactivity is also explored. Finally, with the recent advances in bioinformatics and synthetic biology, it is becoming clear that the integration of these disciplines with genetic and biochemical characterization of the novel marine peptides, offers the most potential in the development of the next generation of societal solutions.     

Marine-Derived Quorum-Sensing Inhibitory Activities Enhance the Antibacterial Efficacy of Tobramycin against Pseudomonas aeruginosa

Bacterial epiphytes isolated from marine eukaryotes were screened for the production of quorum sensing inhibitory compounds (QSIs). Marine isolate KS8, identified as a Pseudoalteromonas sp., was found to display strong quorum sensing inhibitory (QSI) activity against acyl homoserine lactone (AHL)-based reporter strains Chromobacterium violaceum ATCC 12472 and CV026. KS8 supernatant significantly reduced biofilm biomass during biofilm formation (−63%) and in pre-established, mature P. aeruginosa PAO1 biofilms (−33%). KS8 supernatant also caused a 0.97-log reduction (−89%) and a 2-log reduction (−99%) in PAO1 biofilm viable counts in the biofilm formation assay and the biofilm eradication assay respectively. The crude organic extract of KS8 had a minimum inhibitory concentration (MIC) of 2 mg/mL against PAO1 but no minimum bactericidal concentration (MBC) was observed over the concentration range tested (MBC > 16 mg/mL). Sub-MIC concentrations (1 mg/mL) of KS8 crude organic extract significantly reduced the quorum sensing (QS)-dependent production of both pyoverdin and pyocyanin in P. aeruginosa PAO1 without affecting growth. A combinatorial approach using tobramycin and the crude organic extract at 1 mg/mL against planktonic P. aeruginosa PAO1 was found to increase the efficacy of tobramycin ten-fold, decreasing the MIC from 0.75 to 0.075 µg/mL. These data support the validity of approaches combining conventional antibiotic therapy with non-antibiotic compounds to improve the efficacy of current treatments.     

Haloperoxidase Mediated Quorum Quenching by Nitzschia cf pellucida: Study of the Metabolization of N-Acyl Homoserine Lactones by a Benthic Diatom

Diatoms are known to produce a variety of halogenated compounds, which were recently shown to have a role in allelopathic interactions between competing species. The production of these compounds is linked to haloperoxidase activity. This research, has shown that this system may also be involved in diatom-bacteria interactions via the H₂O₂ dependent inactivation of a type of quorum sensing (QS) molecule, i.e., N-β-ketoacylated homoserine lactones (AHLs), by a natural haloperoxidase system from the benthic diatom Nitzschia cf pellucida. The AHL degradation pathway towards corresponding halogenated derivatives was elucidated via HPLC-MS analysis and the synthesis of a broad series of novel halogenated AHL analogues as reference compounds. Furthermore, their biological activity as quorum sensing modulators was directly compared and evaluated against a series of naturally occurring β-keto-AHLs. It has been demonstrated that the loss of the QS activity results from the final cleavage of the halogenated N-acyl chain of the signal molecules.     

Identification of Four New agr Quorum Sensing-Interfering Cyclodepsipeptides from a Marine Photobacterium

During our search for new natural products from the marine environment, we discovered a wide range of cyclic peptides from a marine Photobacterium, closely related to P. halotolerans. The chemical fingerprint of the bacterium showed primarily non-ribosomal peptide synthetase (NRPS)-like compounds, including the known pyrrothine antibiotic holomycin and a wide range of peptides, from diketopiperazines to cyclodepsipeptides of 500–900 Da. Purification of components from the pellet fraction led to the isolation and structure elucidation of four new cyclodepsipeptides, ngercheumicin F, G, H, and I. The ngercheumicins interfered with expression of virulence genes known to be controlled by the agr quorum sensing system of Staphylococcus aureus, although to a lesser extent than the previously described solonamides from the same strain of Photobacterium.     

Ecological and Industrial Implications of Dynamic Seaweed-Associated Microbiota Interactions

Seaweeds are broadly distributed and represent an important source of secondary metabolites (e.g., halogenated compounds, polyphenols) eliciting various pharmacological activities and playing a relevant ecological role in the anti-epibiosis. Importantly, host (as known as basibiont such as algae)–microbe (as known as epibiont such as bacteria) interaction (as known as halobiont) is a driving force for coevolution in the marine environment. Nevertheless, halobionts may be fundamental (harmless) or detrimental (harmful) to the functioning of the host. In addition to biotic factors, abiotic factors (e.g., pH, salinity, temperature, nutrients) regulate halobionts. Spatiotemporal and functional exploration of such dynamic interactions appear crucial. Indeed, environmental stress in a constantly changing ocean may disturb complex mutualistic relations, through mechanisms involving host chemical defense strategies (e.g., secretion of secondary metabolites and antifouling chemicals by quorum sensing). It is worth mentioning that many of bioactive compounds, such as terpenoids, previously attributed to macroalgae are in fact produced or metabolized by their associated microorganisms (e.g., bacteria, fungi, viruses, parasites). Eventually, recent metagenomics analyses suggest that microbes may have acquired seaweed associated genes because of increased seaweed in diets. This article retrospectively reviews pertinent studies on the spatiotemporal and functional seaweed-associated microbiota interactions which can lead to the production of bioactive compounds with high antifouling, theranostic, and biotechnological potential.     

Inhibition of virulence gene expression in Staphylococcus aureus by novel depsipeptides from a marine photobacterium

During a global research expedition, more than five hundred marine bacterial strains capable of inhibiting the growth of pathogenic bacteria were collected. The purpose of the present study was to determine if these marine bacteria are also a source of compounds that interfere with the agr quorum sensing system that controls virulence gene expression in Staphylococcus aureus. Using a gene reporter fusion bioassay, we recorded agr interference as enhanced expression of spa, encoding Protein A, concomitantly with reduced expression of hla, encoding α-hemolysin, and rnaIII encoding RNAIII, the effector molecule of agr. A marine Photobacterium produced compounds interfering with agr in S. aureus strain 8325-4, and bioassay-guided fractionation of crude extracts led to the isolation of two novel cyclodepsipeptides, designated solonamide A and B. Northern blot analysis confirmed the agr interfering activity of pure solonamides in both S. aureus strain 8325-4 and the highly virulent, community-acquired strain USA300 (CA-MRSA). To our knowledge, this is the first report of inhibitors of the agr system by a marine bacterium.     

洋葱根腐病防治药剂筛选试验

[目的]筛选防治洋葱根腐病的有效药剂。[方法]在室内药剂筛选基础上,进行化学药剂筛选的同时,将其与生防菌复配,开展不同药剂田间药效试验。[结果]哈茨木霉菌和15%多菌灵复配对洋葱根腐病的防治效果明显,可增产27.6%。[结论 ]哈茨木霉菌和15%多菌灵复配进行洋葱根腐病的防治,具有防治过程农药毒性小,持效期长,能有效防治洋葱根腐病,降低环境污染的特点,生产中可作为防治洋葱根腐病的有效药剂推广使用。     

基于多源遥感数据的海南岛2007—2018年森林时空变化研究

海南岛作为中国仅有的两大热区之一,森林资源极其丰富,具有非常重要的生态和经济价值。本世纪以来,海南省的经济一直呈高速增长态势,橡胶林作为当地森林的重要组成部分,其种植面积因全球胶价的快速攀升增加了约50%,经济的快速发展和橡胶等人工林的增长将对森林的结构和分布产生重要影响,在此背景下监测森林现状及动态变化可为海南岛森林可持续经营管理和国家生态文明试验区建设提供重要依据。本文联合Landsat/Sentinel-2光学和PALSAR/PALSAR-2雷达多源遥感影像为数据源,以海南岛为研究对象,基于谷歌地球引擎（Google Earth Engine, GEE）监测海南岛2007—2018年的森林变化特征。研究结果表明：（1）2007—2018年海南岛森林面积增长显著,从2007年的1.94×10⁴ km²增长至2018年的2.15×10⁴ km²,覆盖率从56.43%上升到62.59%,森林面积年平均增长率为0.99%;（2）2007—2018年,在海南岛18个市（县）中有15个市（县）的森林面积呈正增长趋势,其中增长排前5的市（县）（儋州市、澄迈县、琼海市、临高县、定安县）增长面积占正增长市（县）总增长面积的68.78%,仅有东方市、三亚市和琼中县3个市（县）的森林面积有所下降,分别下降了13、76、113 km²;（3）森林增长集中分布在海拔小于200 m和坡度小于8°的区域,分别占增长总面积的92.88%和91.43%;（4）橡胶林的增长是森林面积快速增长的主要原因,橡胶林面积增长最多的5个市（县）其森林面积也增长最多,这5个市（县）的橡胶增长总面积占森林总增长面积的63.40%;（5）森林损失区域集中在海南岛南部、西南部和东北部沿海地区,主要是因为高郁闭度森林（如橡胶林）逐步转换为郁闭度较底的热带果园（如芒果园等）,另外旅游建设开发、城市化和水利建设等也导致了部分森林损失。研究总体显示,海南岛2007—2018年森林在低海拔平缓区域增长显著,森林消长具有一定的空间差异性,中部山区森林持续保持较高的覆盖度且较少大面积森林损失,总体增长模式良好。     

2000～2014年闽东海岸带土地利用动态变化

海岸带地区因其特殊的海陆生态系统环境而成为土地利用/覆被变化(LUCC)研究的热点区域.本文以位于海峡西岸经济区东北翼的闽东海岸带作为研究对象,利用2000、2009和2014年3期Landsat TM/OLI影像作为数据源,对该区2000～2014年的土地利用动态变化特征进行分析.结果表明:研究区土地利用类型以林草地和耕地为主,其中林草地面积迅速扩增,研究期间共增加了1 047.24 km2,相比之下,耕地面积显著减少,近14a间共减少了959.45 km2;在人类活动影响下,水产养殖的动态变化最为剧烈,稳定性最差,而由于政府加强对建筑用地的集约利用,使其成为近14 a研究区最为稳定的土地利用类型,面积基本保持不变;闽东海岸带土地利用的集约化程度远超全国平均水平,土地利用程度综合指数达255.19,但2000～2009年间,研究区土地利用仍处于调整期,2009～2014年,随着经济的快速发展及前期的调整,土地利用效率有所提高,其变化量和变化率分别为4.86和0.019,土地利用步入发展期.本研究对闽东及其他地区的土地集约化利用及生态环境可持续具有指导意义.     

Continuous Application of Biochar Inoculated with Root Nodule Bacteria to Subsoil Enhances Yield of Soybean by the Nodulation Control using Crack Fertilization Technique

Abstract

Crack fertilization is a soybean cultivation technique for nodulation control in which midterm subsoiling is used to supply fertilizing materials to deep soil just before the flowering stage. This study examined the effects of fertilizing materials and the continuous application of nodulation control, on soybean yield enhancement in two field experiments. The survival of nodule bacteria in deep soil was also evaluated by a bioassay of nodule bacteria in a root box. When the nodule bacteria on biochar were continuously applied without any other chemical fertilizers for three successive years, seed weight was significantly heavier being up to 1.34 times that of the control. The application of nodulation control in the previous year but not in the experimental year did not have residual effects on seed weight. The enhancement of seed weight in a farm field converted from a paddy was much lower. This may be partly attributed to the midterm tillage practice, whichdestroys the crack structure after the nodulation control, together with soil water status and cultivar differences. Nodule growth and nitrogen fixation activities significantly increased in the soybean plants grown on the soil collected from the subsoil to which nodule bacteria on biochar had been applied the previous year. This suggests that nodule bacteria in the subsoil survived in the biochar habitat for at least a year after application. These results indicate that nodulation control by the crack fertilization technique leads to yield enhancement when nodule bacteria on biochar are continuously applied.     

Impact of Agronomic Strategies (Seed Tuber Pre-sprouting,Cultivar Choice) to Control Late Blight (<Emphasis Type="Italic">Phytophthora infestans</Emphasis>) on Tuber Growth and Yield in Organic Potato (<Emphasis Type="Italic">Solanum tuberosum</Emphasis> L.) Crops

In order to optimize potato crop management in organic farming systems, knowledge of crop growth processes in relation to limitations and reductions by nitrogen (N) and disease is necessary. This paper examines the effect of different prevention measures (seed tuber pre-sprouting, choice of cultivars: resistance to Phytophthora infestans; earliness of tuber initiation) against disease-related constraints on yields, depending on the N-mediated growing conditions of organic potato crops. Under conditions of a relatively high N supply, accelerating the early development and tuber initiation of potato crops by seed tuber pre-sprouting (yield increase of c. 18–23%) or the selective choice of cultivars with an earlier tuber initiation (yield increase of c. 0–21%) are the most effective strategies in combating late blight. They bring forward the crop development c. 7–10 days, escaping the negative impact of the fungus on tuber growth. Under conditions of a relatively low N supply, these strategies do not affect final tuber yield. The reason for the differences of the effect depending on N supply of the crops is, that the lower the N supply, the shorter the period of time over which tuber growth takes place, independent of whether P. infestans becomes severe or not. Tuber growth of organic crops low in N has mostly ceased by the time late blight becomes potentially important for limiting yield, with the consequence that preventative measures against P. infestans are meaningless. Surprisingly, the level of cultivar haulm resistance to late blight did not affect tuber yields in years with an early and strong late blight development. Probably, the positive effect of a longer resistance to the fungus (c. 1 week) was counterbalanced by a later tuber initiation (also c. 1 week). In years with a moderate late blight incidence, less susceptible cultivars were able to produce higher tuber yields on fields with a lower N availability (c. 17–20%), probably due to higher N use efficiency and a better match of N mineralization and N uptake. Under conditions of a relatively high N supply, the tested cultivars with a higher haulm resistance to late blight were not able to improve tuber yields.