شناسایی ژن‌های احتمالی مرتبط با سیستم ضد احساس حد نصاب (توانایی کوئنچری) در باکتری‌های دارای قابلیت مهار زیستی

نوع مقاله: علمی پژوهشی-فارسی

نویسندگان

1 استادیار دانشگاه ایلام

2 استادیار، دانشگاه لرستان

10.22055/ppr.2019.15297

چکیده

یکی از راه­ های جدید کنترل باکتری‌ های بیمارگر، استفاده از باکتری ­های دارای توانایی مهار زیستی مبتنی بر سیستم ضداحساس حد نصاب (توانایی کوئنچری) است. بر این اساس جدایه­ های باکتریایی دارای قدرت کوئنچری از ریزوسفر و فیلوسفر گیاهان مختلف جداسازی گردیدند. در بین جدایه ­های باکتریایی مورد بررسی، چهار جنس و گونه باکتریایی شامل Pseudomonas chlororaphis، Pseudomonas putida،Acinetobacter sp.  و Bacillus sp. به عنوان جدایه ­های باکتریایی برتر با توانایی کوئنچری و قابلیت مهار زیستی بیمارگرهای باکتریایی جنس Pectobacterium معرفی شدند. فراوان­ترین­ و قوی­ترین­ جدایه ­های دارای توانایی کوئنچری، فعالیت آنزیمی مرتبط با این پدیده از خود نشان دادند. سپس به منظور شناسایی ژن ­های آسیلازی و لاکتونازی این باکتری ­ها، آغازگرهایی طراحی و مورد آزمایش قرار گرفت. بر اساس نتایج حاصل از بررسی آغازگرها، در جنس باسیلوس ژن لاکتونازی aiia، در جدایه­­های  P. chlororaphis سه ژن آسیلازی همولوگ pvdQ، quiP و  hacB(یا فقط دو ژن اول)، درP. putida دو ژن همولوگ pvdQ  وquiP و در Acinetobacter یک ژن آسیلازی ردیابی گردید.  

کلیدواژه‌ها


عنوان مقاله [English]

Detection and identification of potentially anti-quorum sensing related genes in biocontrol bacteria

نویسندگان [English]

  • M. R. Alimanesh 1
  • H. Mirzaei Najafgholi 2
1 Assistant Professor, Ilam University
2 Assistant Professor, Lorestan University
چکیده [English]

Background and Objectives
Quorum sensing (QS) is a process of cell to cell communication that allows bacteria to be aware about cell density. In this phenomenon bacteria communicate with each other through signaling molecules such as N-acylhomoserine lactones (AHLs). Also, virulence gene expression in many pathogenic bacteria like Pectobacterium is under control of QS. Some bacteria can degrade AHLs molecules by a process called anti quorum sensing or quorum quenching (QQ); therefore, QQ can be used in biocontrol of plant pathogenic bacteria. The main purpose of this research was to detect QQ genes in strong QQ-based biocontrol bacteria.
Materials and Methods
In this investigation, QQ bacteria were isolated from rhizosphere and phyllosphere of some agricultural and non-agricultural plants using minimum media containing AHL. QQ bacteria were identified using biosensors CV026 and VIR07. Mechanism of QQ action was determined by heating, proteinase treatment and filtering methods. Afterwards, in order to find acylase and lactonase genes in these bacteria, homology searches were performed using BLASTn and BLASTp in NCBI. Then, several primers were designed by several softwares including CLC main workbench 5.5, Primer Premier 6, Primer3, Oligo7 and primer BLAST.
Results
The most abundant and strongest QQ isolates were shown enzymatic activity. Main twenty-seven isolates with high QQ activity were detected. Four genera and species including Pseudomonas chlororaphis, Pseudomonas putida, Acinetobacter sp. and Bacillus sp. were found as the bacteria with highest QQ and biocontrol ability against Pectobacterium. As a results of this study, in the Bacillus Aiia lactonase gene;in P. chlororaphis three acylase genes of pvdQ، quiP and hacB; in P.putida two acylase genes of  pvdQ and quiP  and in Acinetobacter one acylase genes were traced.
Discussion
According to the results of this investigation, Pseudomonas genus was determined as the frequent QQ bacteria. Also, P. chlororaphis species with the strongest QQ activity, were divided in to two groups; the first isolates with two acylase genes (pvdQ and quiP) and the second isolates containing extra homologous acylase hacB. Isolates of the latter group with the most detected genes involved in QQ, were identified as the best QQ-based biocontrol bacteria against Pectobacterium.

کلیدواژه‌ها [English]

  • Cell communication
  • Pseudomonas
  • Pectobacterium
  • acylase and lactonase genes
Ahmed, S. A., Rudden, M., Smyth, T. J., Dooley, J. S., Marchant, R., and Banat, I. M. 2019. Natural quorum sensing inhibitors effectively downregulate gene expression of Pseudomonas aeruginosa virulence factors. Applied Microbiology and Biotechnology, 103: 3521-3535.
Andersson, R. A., Eriksson, A. R., Heikinheimo, R., Mäe, A., Pirhonen, M., Kõiv, V., Hyytiäinen, H., Tuikkala, A., and Palva, E. T. 2000. Quorum sensing in the plant pathogen Erwinia carotovora subsp. carotovora: the role of expREcc. Molecular Plant-Microbe Interactions, 13 (4): 384-393.
Berger-Bächi, B. 2002. Resistance mechanisms of gram-positive bacteria. International Journal of Medical Microbiology, 292 (1): 27-35.
Bokhove, M., Jimenez, P. N., Quax, W. J., and Dijkstra, B. W. 2010. The quorum-quenching N-acyl homoserine lactone acylase PvdQ is an Ntn-hydrolase with an unusual substrate-binding pocket. Proceedings of the National Academy of Sciences, 107:686-691.
Chan, K. G., Yin, W. F., Sam, C. K., and Koh, C. L. 2009. A novel medium for the isolation of N-acylhomoserine lactone-degrading bacteria. Journal of Industrial Microbiology and Biotechnology36: 247-251.
Chen, B., Peng, M., Tong, W., Zhang, Q., and Song, Z. 2019. The Quorum Quenching Bacterium Bacillus licheniformis T-1 Protects Zebrafish against Aeromonas hydrophila Infection. Probiotics and Antimicrobial Proteins, 4: 1-2.
Christiaen, S. E., Brackman, G., Nelis, H. J., and Coenye, T. 2011. Isolation and identification of quorum quenching bacteria from environmental samples. Journal of Microbiological Methods, 87 (2): 213-219.
Chu, Y, Y., Nega, M., Wölfle, M., Plener, L., Grond, S., Jung, K., Götz, F. A. 2013. New class of quorum quenching molecules from Staphylococcus species affects communication and growth of gram-negative bacteria. PLoS Pathogens, 26 (9): e1003654.
D’Angelo‐Picard, C., Faure, D., Penot, I., and Dessaux, Y. 2005. Diversity of N‐acyl homoserine lactone‐producing and‐degrading bacteria in soil and tobacco rhizosphere. Environmental Microbiology. 7 (11): 1796-1808.
Das, R., and Mehta, D. K. 2018. Microbial biofilm and quorum sensing inhibition: endowment of medicinal plants to combat multidrug-resistant bacteria. Current Drug Targets, 19 (16): 1916-1932.
Dong, Y. H., Xu, J. L., Li, X. Z., and Zhang, L. H. 2000. AiiA, an enzyme that inactivates the acylhomoserine lactone quorum-sensing signal and attenuates the virulence of Erwinia carotovora. Proceedings of the National Academy of Sciences, 97: 3526-3531.
Dong, Y. H., Gusti, A. R., Zhang, Q., Xu, J. L., and Zhang, L. H. 2002. Identification of quorum-quenching N-acyl homoserine lactonases from Bacillus species. Applied and Environmental Microbiology, 68: 1754-1759.
Dong, Y. H., Zhang, X. F., Xu, J. L., and Zhang, L. H. 2004. Insecticidal Bacillus thuringiensis silences Erwinia carotovora virulence by a new form of microbial antagonism, signal interference. Applied and Environmental Microbiology, 70: 954-960.
Fetzner, S. 2015. Quorum quenching enzymes. Journal of Biotechnology, 201: 2-14.
Fuqua, C., Parsek, M. R., and Greenberg, E. P. 2001. Regulation of gene expression by cell-to-cell communication: acyl-homoserine lactone quorum sensing. Annual Review of Genetics, 35 (1): 439-468.
Grandclément, C., Tannières, M., Moréra, S., Dessaux, Y., and Faure, D.D. 2015. Quorum quenching: role in nature and applied developments. FEMS Microbiology Reviews, 40:86-116.
Jones, S., Yu, B., Bainton, N. A., Birdsall, M., Bycroft, B. W., Chhabra, S. R., Cox, A. J., Golby, P., Reeves, P. J., and Stephens, S. 1993. The lux autoinducer regulates the production of exoenzyme virulence determinants in Erwinia carotovora and Pseudomonas aeruginosa. The EMBO Journal. 12 (6): 2477-2482.
Kalia, V. C. 2013. Quorum sensing inhibitors: an overview. Biotechnology Advances, 31(2): 224-245.
Kang, B. R., Lee, J. H., Ko, S. J., Lee, Y. H., Cha, J. S., Cho, B. H., and Kim, Y.C. 2004. Degradation of acyl-homoserine lactone molecules by Acinetobacter sp. strain C1010. Canadian Journal of Microbiology, 50: 935-941.
Kaur, J., and Yogalakshmi, K. N. 2018. Screening of quorum quenching activity of the bacteria isolated from dairy industry waste activated sludge. International Journal of Environmental Science and Technology 16: 5421-5428.
Koch, G., Nadal-Jimenez, P., Reis, C. R., Muntendam, R., Bokhove, M., Melillo, E., Dijkstra, B. W., Cool, R. H., and Quax, W. J. 2014. Reducing virulence of the human pathogen Burkholderia by altering the substrate specificity of the quorum-quenching acylase PvdQ. Proceedings of the National Academy of Sciences, 111: 1568-1573.
Lasarre, B., and Federle, M. J. 2013. Exploiting quorum sensing to confuse bacterial pathogens. Microbiology and Molecular Biology Reviews, 77(1): 73-111.
Lin, Y. H., Xu, J. L., Hu, J., Wang, L. H., Ong, S. L., Leadbetter, J. R., and Zhang, L. H. 2003. Acyl‐homoserine lactone acylase from Ralstonia strain XJ12B represents a novel and potent class of quorum‐quenching enzymes. Molecular Microbiology, 47: 849-860.
Lwin, M., and Ranamukhaarachchi, S. 2006. Development of biological control of Ralstonia solanacearum through antagonistic microbial populations. International Journal of Agriculture and Biology, 8: 657-660.
Mahmoudi, E., Tabatabaei, B. E. S. and Venturi, V. 2011. Virulence attenuation of Pectobacterium carotovorum using N-Acyl-homoserine lactone degrading bacteria isolated from potato rhizosphere. The Plant Pathology Journal, 27 (3): 242-248.
McClean, K. H., Winson, M. K., Fish, L., Taylor, A., Chhabra, S. R., Camara, M., Daykin, M., Lamb, J. H., Swift, S., and Bycroft, B. W. 1997. Quorum sensing and Chromobacterium violaceum: exploitation of violacein production and inhibition for the detection of N-acylhomoserine lactones. Microbiology, 143: 3703-3711.
Mei, G. Y., Yan, X. X., Turak, A., Luo, Z. Q., and Zhang, L. Q., 2010. AidH, an alpha/beta-hydrolase fold family member from an Ochrobactrum sp. strain, is a novel N-acylhomoserine lactonase. Applied and Environmental Microbiology, 76 (15): 4933-4942.
Morohoshi, T., Kato, M., Fukamachi, K., Kato, N., and Ikeda, T. 2008. N-acylhomoserine lactone regulates violacein production in Chromobacterium violaceum type strain ATCC 12472. FEMS Microbiology Letters, 279: 124-130.
Morohoshi, T., Someya, N. and Ikeda, T. 2009. Novel N-acylhomoserine lactone-degrading bacteria isolated from the leaf surface of Solanum tuberosum and their quorum-quenching properties. Bioscience, Biotechnology and Biochemistry, 73 (9): 2124-2127.
Oldfield, E., and Feng, X. 2014. Resistance-resistant antibiotics. Trends in Pharmacological Sciences, 35 (12): 664-674.
Otto, M. 2004. Quorum‐sensing control in Staphylococci–a target for antimicrobial drug therapy. FEMS Microbiology Letters,241: 135-141.
Ouyang, L., and Li, L. 2016. Effects of an inducible aiiA gene on disease resistance in Eucalyptus urophylla× Eucalyptus grandis. Transgenic Research, 25: 441-452.
Papenfort, K., and Bassler, B. L. 2016. Quorum sensing signal–response systems in Gram-negative bacteria. Nature Reviews Microbiology, 14: 576.
Pirhonen, M., Flego, D., Heikinheimo, R., and Palva, E. T. 1993. A small diffusible signal molecule is responsible for the global control of virulence and exoenzyme production in the plant pathogen Erwinia carotovora. The EMBO Journal, 12: 2467-2476.
Põllumaa, L., Alamäe, T., and Mäe, A. 2012. Quorum sensing and expression of virulence in Pectobacteria. Sensors, 12: 3327-3349.
Rajesh, P., and Rai, V.R. 2014. Molecular identification of aiiA homologous gene from endophytic Enterobacter species and in silico analysis of putative tertiary structure of AHL-lactonase. Biochemical and Biophysical Research Communications, 443: 290-295.
Reina, J. C., Torres, M., and Llamas, I. 2019. Stenotrophomonas maltophilia AHL-degrading strains isolated from marine invertebrate microbiota attenuate the virulence of Pectobacterium carotovorum and Vibrio coralliilyticus. Marine Biotechnology, 21: 276-290.
Schaad, N, W., Jones, J, B., and Chun, W. 2001. Laboratory guide for the identification of plant pathogenic bacteria. American Phytopathological Society (APS Press), 3: 20-59.
Truchado, P., López-Gálvez, F., Gil, M., Tomás-Barberán, F., and Allende, A. 2009. Quorum sensing inhibitory and antimicrobial activities of honeys and the relationship with individual phenolics. Food Chemistry, 115: 1337-1344.
Uroz, S., Oger, P., Chhabra, S.R., Cámara, M., Williams, P. and Dessaux, Y., 2007. N-acyl homoserine lactones are degraded via an amidolytic activity in Comamonas sp. strain D1. Archives of Microbiology, 187 (3): 249-256.
Vinoj, G., Vaseeharan, B., Thomas, S., Spiers, A.J., and Shanthi, S. 2014. Quorum-quenching activity of the AHL-Lactonase from Bacillus licheniformis DAHB1 inhibits vibrio biofilm formation in vitro and reduces shrimp intestinal colonisation and mortality. Marine Biotechnology, 16: 707-715.
Wahjudi, M., Papaioannou, E., Hendrawati, O., Van Assen, A. H., van Merkerk, R., Cool, R. H. Poelarends, G. j., and Quax, W.J. 2011. PA0305 of Pseudomonas aeruginosa is a quorum quenching acylhomoserine lactone acylase belonging to the Ntn hydrolase superfamily. Microbiology, 157: 2042-2055.
Waters, C. M., and Bassler, B. L. 2005. Quorum sensing: cell-to-cell communication in bacteria. Annual Review of Cell and Developmental Biology, 21: 319-346.
Weisburg, W. G., Barns, S. M., Pelletier, D. A., and Lane, D. J. 1991. 16S ribosomal DNA amplification for phylogenetic study. Journal of Bacteriology, 173 (2): 697-703.