title: Identification of Pathogenicity-Related Genes in Biofilm-Defective Acidovorax citrulli by Transposon Tn5 Mutagenesis.
abstract: Biofilm formation is important for virulence of a large number of plant pathogenic bacteria. Indeed, some virulence genes have been found to be involved in the formation of biofilm in bacterial fruit blotch pathogen Acidovorax citrulli. However, some virulent strains of A. citrulli were unable to format biofilm, indicating the complexity between biofilm formation and virulence. In this study, virulence-related genes were identified in the biofilm-defective strain A1 of A. citrulli by using Tn5 insertion, pathogenicity test, and high-efficiency thermal asymmetric interlaced PCR (hiTAIL-PCR). Results from this study indicated that 22 out of the obtained 301 mutants significantly decreased the virulence of strain A1 compared to the wild-type. Furthermore, sequence analysis indicated that the obtained 22 mutants were due to the insertion of Tn5 into eight genes, including Aave 4244 (cation diffusion facilitator family transporter), Aave 4286 (hypothetical protein), Aave 4189 (alpha/beta hydrolase fold), Aave 1911 (IMP dehydrogenase/GMP reductase domain), Aave 4383 (bacterial export proteins, family 1), Aave 4256 (Hsp70 protein), Aave 0003 (histidine kinase, DNA gyrase B, and HSP90-like ATPase), and Aave 2428 (pyridoxal-phosphate dependent enzyme). Furthermore, the growth of mutant Aave 2428 was unaffected and even increased by the change in incubation temperature, NaCl concentration and the pH of the LB broth, indicating that this gene may be directly involved in the bacterial virulence. Overall, the determination of the eight pathogenicity-related genes in strain A1 will be helpful to elucidate the pathogenesis of biofilm-defective A. citrulli.
authors.affiliation: Department of Plant Quarantine, Shanghai Extension and Service Center of Agriculture Technology, Shanghai 201103, China. toyanzi@126.com.
authors.forename: Jinyan
authors.initials: J
authors.lastname: Luo
authors.affiliation: State Key Laboratory of Rice Biology, Institute of Biotechnology, Zhejiang University, Hangzhou 310058, China. wenwen20101010@163.com.
authors.forename: Wen
authors.initials: W
authors.lastname: Qiu
authors.affiliation: Department of Plant Quarantine, Shanghai Extension and Service Center of Agriculture Technology, Shanghai 201103, China. Chenlei200524@163.com.
authors.forename: Lei
authors.initials: L
authors.lastname: Chen
authors.affiliation: State Key Laboratory of Rice Biology, Institute of Biotechnology, Zhejiang University, Hangzhou 310058, China. ishtiaq@kust.edu.pk.|Department of Zoology, Kohat University of Science and Technology, Kohat 26000, Pakistan. ishtiaq@kust.edu.pk.
authors.forename: Syed Ishtiaq
authors.initials: SI
authors.lastname: Anjum
authors.affiliation: State Key Laboratory of Rice Biology, Institute of Biotechnology, Zhejiang University, Hangzhou 310058, China. mhyu@zju.edu.cn.
authors.forename: Menghao
authors.initials: M
authors.lastname: Yu
authors.affiliation: State Key Laboratory of Rice Biology, Institute of Biotechnology, Zhejiang University, Hangzhou 310058, China. changlin_shan@163.com.|Department of Plant Quarantine, Zhoushan Entry-Exit Inspections and Quarantine Bureau, Hangzhou 310012, China. changlin_shan@163.com.
authors.forename: Changlin
authors.initials: C
authors.lastname: Shan
authors.affiliation: Department of Biotechnology, University of Sargodha, Sargodha 40100, Pakistan. mehmoona.ilyas@gmail.com.
authors.forename: Mehmoona
authors.initials: M
authors.lastname: Ilyas
authors.affiliation: State Key Laboratory of Rice Biology, Institute of Biotechnology, Zhejiang University, Hangzhou 310058, China. libin0571@zju.edu.cn.
authors.forename: Bin
authors.initials: B
authors.lastname: Li
authors.affiliation: State Key Laboratory Breeding Base for Zhejiang Sustainable Plant Pest and Disease Control, Key Laboratory of Detection for Pesticide Residues, Ministry of Agriculture, Zhejiang Academy of Agricultural Sciences, Hangzhou 310021, China. ylwang88@aliyun.com.
authors.forename: Yanli
authors.initials: Y
authors.lastname: Wang
authors.affiliation: State Key Laboratory Breeding Base for Zhejiang Sustainable Plant Pest and Disease Control, Key Laboratory of Detection for Pesticide Residues, Ministry of Agriculture, Zhejiang Academy of Agricultural Sciences, Hangzhou 310021, China. sungc01@sina.com.
authors.forename: Guochang
authors.initials: G
authors.lastname: Sun
mesh_terms: D000595:Amino Acid Sequence; D018441:Biofilms; D042621:Comamonadaceae / Q000235:genetics* / Q000472:pathogenicity*; D019295:Computational Biology / Q000379:methods; D004251:DNA Transposable Elements; D005798:Genes, Bacterial; D008969:Molecular Sequence Data; D016296:Mutagenesis; D016254:Mutagenesis, Insertional; D009154:Mutation; D016415:Sequence Alignment; D013312:Stress, Physiological; D014774:Virulence / Q000235:genetics*
chemical_list: D004251:DNA Transposable Elements
keywords: Acidovorax citrulli; Tn5; bacterial fruit blotch; biofilm; pathogenesis