scholarly journals Genetic Causes of Non-pathogenic Pseudomonas syringae pv. actinidiae Isolates in Kiwifruit Orchards

2021 ◽  
Vol 12 ◽  
Author(s):  
Yue Li ◽  
Qiaomei Zhu ◽  
Taihui Zhi ◽  
Rong Fan ◽  
Ting Xie ◽  
...  
Keyword(s):  
Pseudomonas Syringae ◽  
Dna Sequences ◽  
Host Plants ◽  
Gene Encoding ◽  
Low Frequencies ◽  
Canker Disease ◽  
Type Iii ◽  
Transposon Insertion ◽  
Typing Methods ◽  
Transposon Insertions

Bacterial canker disease has become the largest threat to kiwifruit cultivation and production. A monomorphic subpopulation of Pseudomonas syringae pv. actinidiae biovar 3 (Psa3) is responsible for the pandemic worldwide. Diversity in pathogenicity has been found in the pandemic subpopulation and in other Psa3 subpopulations causing epidemics in China. However, the genetic bases have not yet been elucidated. In this study, 117 Psa3 isolates were identified by Psa- and Psa3-specific primers, and evaluated for pathogenicity. Three isolates G4, G40, and S2 are not pathogenic to kiwifruit and do not elicit hypersensitivity responses (HRs) in non-host Nicotiana benthamiana leaves. Two isolates, G25 and G35, exhibited attenuated HR-eliciting activity in non-host N. benthamiana, but they exhibited greatly and slightly reduced pathogenicity in host plants, respectively. The genomes of the five isolates were sequenced and compared with closely related isolates revealed by MLVA and whole-genome typing methods. The candidate genetic loci responsible for the changes in pathogenicity and HR elicitation, were further evaluated by allele replacement experiments. We found that the three non-pathogenic isolates were formed due to the independent, identical insertion events of ISPsy36 transposon in the hrpR gene, encoding a key regulator of type III secretion system (T3SS) and type III effectors (T3Es). In the symptomatic sample from which G4 was isolated, 27% HR negative isolates were detected. In isolate G25, transposon insertion of ISPsy32 at the non-coding sequence upstream of the hrpR gene was detected, similar to a previously reported low-virulent Psa3 strain M227. In isolate G35, we detected disruptions of T3Es hopBB1-1 and hopBB1-2, which induce HR in N. benthamiana leaves revealed by Agrobacterium tumefaciens infiltration. These phenotype-changed isolates were formed at low frequencies during the course of pathogen infection in host plants, supported by the binding assay of ISPsy32 and the non-coding DNA sequences upstream of the hrpR gene, the co-isolation of the virulent isolates belonging to the same MLVA clade, and the low levels of transcription of the transposon genes. Taken together, in terms of short-term field evolution, transposon insertions in the T3SS-related genes resulted in the formation of non-pathogenic and low-virulent Psa3 isolates.

scholarly journals The type III effector repertoire of Pseudomonas syringae pv. syringae B728a and its role in survival and disease on host and non-host plants

2006 ◽  
Vol 62 (1) ◽  
pp. 26-44 ◽  
Author(s):  
Boris A. Vinatzer ◽  
Gail M. Teitzel ◽  
Min-Woo Lee ◽  
Joanna Jelenska ◽  
Sara Hotton ◽  
...  
Keyword(s):  
Pseudomonas Syringae ◽  
Host Plants ◽  
Type Iii Effector ◽  
Type Iii

scholarly journals Identification of a Virulence-Associated Determinant, Dihydrolipoamide Dehydrogenase (lpd), in Mycoplasma gallisepticum through In Vivo Screening of Transposon Mutants

2006 ◽  
Vol 74 (2) ◽  
pp. 931-939 ◽  
Author(s):  
P. Hudson ◽  
T. S. Gorton ◽  
L. Papazisi ◽  
K. Cecchini ◽  
S. Frasca ◽  
...  
Keyword(s):  
Insertion Site ◽  
Mycoplasma Gallisepticum ◽  
Dihydrolipoamide Dehydrogenase ◽  
Gene Encoding ◽  
Transposon Insertion ◽  
Transposon Mutants ◽  
Genomic Regions ◽  
Transposon Insertions ◽  
Biologic Function

ABSTRACT To effectively analyze Mycoplasma gallisepticum for virulence-associated determinants, the ability to create stable genetic mutations is essential. Global M. gallisepticum mutagenesis is currently limited to the use of transposons. Using the gram-positive transposon Tn4001mod, a mutant library of 110 transformants was constructed and all insertion sites were mapped. To identify transposon insertion points, a unique primer directed outward from the end of Tn4001mod was used to sequence flanking genomic regions. By comparing sequences obtained in this manner to the annotated M. gallisepticum genome, the precise locations of transposon insertions were discerned. After determining the transposon insertion site for each mutant, unique reverse primers were synthesized based on the specific sequences, and PCR was performed. The resultant amplicons were used as unique Tn4001mod mutant identifiers. This procedure is referred to as signature sequence mutagenesis (SSM). SSM permits the comprehensive screening of the M. gallisepticum genome for the identification of novel virulence-associated determinants from a mixed mutant population. To this end, chickens were challenged with a pool of 27 unique Tn4001mod mutants. Two weeks postinfection, the birds were sacrificed, and organisms were recovered from respiratory tract tissues and screened for the presence or absence of various mutants. SSM is a negative-selection screening technique whereby those mutants possessing transposon insertions in genes essential for in vivo survival are not recovered from the host. We have identified a virulence-associated gene encoding dihydrolipoamide dehydrogenase (lpd). A transposon insertion in the middle of the coding sequence resulted in diminished biologic function and reduced virulence of the mutant designated Mg 7.

scholarly journals Translocation and Functional Analysis of Pseudomonas savastanoi pv. savastanoi NCPPB 3335 Type III Secretion System Effectors Reveals Two Novel Effector Families of the Pseudomonas syringae Complex

2014 ◽  
Vol 27 (5) ◽  
pp. 424-436 ◽  
Author(s):  
Isabel M. Matas ◽  
M. Pilar Castañeda-Ojeda ◽  
Isabel M. Aragón ◽  
María Antúnez-Lamas ◽  
Jesús Murillo ◽  
...  
Keyword(s):  
Pseudomonas Syringae ◽  
Dna Sequences ◽  
Type Iii Secretion ◽  
Type Iii Secretion System ◽  
Secretion System ◽  
Callose Deposition ◽  
Type Iii ◽  
T3ss Effector ◽  
Olive Knot Disease ◽  
Relevant Role

Pseudomonas savastanoi pv. savastanoi NCPPB 3335 causes olive knot disease and is a model pathogen for exploring bacterial infection of woody hosts. The type III secretion system (T3SS) effector repertoire of this strain includes 31 effector candidates plus two novel candidates identified in this study which have not been reported to translocate into plant cells. In this work, we demonstrate the delivery of seven NCPPB 3335 effectors into Nicotiana tabacum leaves, including three proteins from two novel families of the P. syringae complex effector super-repertoire (HopBK and HopBL), one of which comprises two proteins (HopBL1 and HopBL2) that harbor a SUMO protease domain. When delivered by P. fluorescens heterologously expressing a P. syringae T3SS, all seven effectors were found to suppress the production of defense-associated reactive oxygen species. Moreover, six of these effectors, including the truncated versions of HopAA1 and HopAZ1 encoded by NCPPB 3335, suppressed callose deposition. The expression of HopAZ1 and HopBL1 by functionally effectorless P. syringae pv. tomato DC3000D28E inhibited the hypersensitive response in tobacco and, additionally, expression of HopBL2 by this strain significantly increased its competitiveness in N. benthamiana. DNA sequences encoding HopBL1 and HopBL2 were uniquely detected in a collection of 31 P. savastanoi pv. savastanoi strains and other P. syringae strains isolated from woody hosts, suggesting a relevant role of these two effectors in bacterial interactions with olive and other woody plants.

scholarly journals Role of Ornibactin Biosynthesis in the Virulence ofBurkholderia cepacia: Characterization of pvdA, the Gene Encoding l-OrnithineN5-Oxygenase

1999 ◽  
Vol 67 (9) ◽  
pp. 4443-4455 ◽  
Author(s):  
P. A. Sokol ◽  
P. Darling ◽  
D. E. Woods ◽  
E. Mahenthiralingam ◽  
C. Kooi
Keyword(s):  
Burkholderia Cepacia ◽  
Respiratory Infections ◽  
Cosmid Library ◽  
Gene Encoding ◽  
Transposon Insertion ◽  
Peptide Synthetase ◽  
Transposon Insertions ◽  
Siderophore Activity

ABSTRACT Burkholderia cepacia is a frequent cause of respiratory infections in cystic fibrosis patients. B. cepacia has been shown to produce at least four siderophores which may play a role in the virulence of this organism. To characterize genes involved in the synthesis of siderophores, Tn5-OT182 mutants were isolated in strain K56-2, which produces two siderophores, salicylic acid (SA) and ornibactins. Two mutants were characterized that did not produce zones on Chrome Azurol S agar in a commonly used assay to detect siderophore activity. These mutants were determined to produce sevenfold more SA than K56-2 yet did not produce detectable amounts of ornibactins. These mutants, designated I117 and T10, had a transposon insertion in genes with significant homology to pyoverdine biosynthesis genes of Pseudomonas aeruginosa. I117 contained an insertion in a pvdA homolog, the gene for the enzymel-ornithine N 5-oxygenase, which catalyzes the hydroxylation of l-ornithine. Ornibactin synthesis in this mutant was partially restored when the precursorl-N 5-OH-Orn was added to the culture medium. T10 contained an insertion in a pvdDhomolog, which is a peptide synthetase involved in pyoverdine synthesis. β-Galactosidase activity was iron regulated in both I117 and T10, suggesting that the transposon was inserted downstream of an iron-regulated promoter. Tn5-OT182 contains alacZ gene that is expressed when inserted downstream of an active promoter. Both I117 and T10 were deficient in uptake of iron complexed to either ornibactins or SA, suggesting that transposon insertions in ornibactin biosynthesis genes also affected other components of the iron transport mechanism. The B. cepacia pvdA homolog was approximately 47% identical and 59% similar tol-ornithine N 5-oxygenase fromP. aeruginosa. Three clones were identified from a K56-2 cosmid library that partially restored ornibactin production, SA production, and SA uptake to parental levels but did not affect the rate of 59Fe-ornibactin uptake in I117. A chromosomalpvdA deletion mutant was constructed that had a phenotype similar to that of I117 except that it did not hyperproduce SA. ThepvdA mutants were less virulent than the parent strain in chronic and acute models of respiratory infection. A functionalpvdA gene appears to be required for effective colonization and persistence in B. cepacia lung infections.

scholarly journals A Draft Genome Sequence of Pseudomonas syringae pv. tomato T1 Reveals a Type III Effector Repertoire Significantly Divergent from That of Pseudomonas syringae pv. tomato DC3000

2009 ◽  
Vol 22 (1) ◽  
pp. 52-62 ◽  
Author(s):  
Nalvo F. Almeida ◽  
Shuangchun Yan ◽  
Magdalen Lindeberg ◽  
David J. Studholme ◽  
David J. Schneider ◽  
...  
Keyword(s):  
Host Range ◽  
Host Specificity ◽  
Pseudomonas Syringae ◽  
Genome Sequence ◽  
Draft Genome ◽  
Draft Genome Sequence ◽  
Tomato Dc3000 ◽  
Type Iii ◽  
Host Range Determination ◽  
Range Determination

Diverse gene products including phytotoxins, pathogen-associated molecular patterns, and type III secreted effectors influence interactions between Pseudomonas syringae strains and plants, with additional yet uncharacterized factors likely contributing as well. Of particular interest are those interactions governing pathogen-host specificity. Comparative genomics of closely related pathogens with different host specificity represents an excellent approach for identification of genes contributing to host-range determination. A draft genome sequence of Pseudomonas syringae pv. tomato T1, which is pathogenic on tomato but nonpathogenic on Arabidopsis thaliana, was obtained for this purpose and compared with the genome of the closely related A. thaliana and tomato model pathogen P. syringae pv. tomato DC3000. Although the overall genetic content of each of the two genomes appears to be highly similar, the repertoire of effectors was found to diverge significantly. Several P. syringae pv. tomato T1 effectors absent from strain DC3000 were confirmed to be translocated into plants, with the well-studied effector AvrRpt2 representing a likely candidate for host-range determination. However, the presence of avrRpt2 was not found sufficient to explain A. thaliana resistance to P. syringae pv. tomato T1, suggesting that other effectors and possibly type III secretion system–independent factors also play a role in this interaction.

scholarly journals What the Wild Things Do: Mechanisms of Plant Host Manipulation by Bacterial Type III-Secreted Effector Proteins

2021 ◽  
Vol 9 (5) ◽  
pp. 1029
Author(s):  
Karl J. Schreiber ◽  
Ilea J. Chau-Ly ◽  
Jennifer D. Lewis
Keyword(s):  
Pseudomonas Syringae ◽  
Structural Data ◽  
Enzymatic Activities ◽  
Effector Proteins ◽  
Host Recognition ◽  
Plant Host ◽  
Type Iii ◽  
Xanthomonas Species ◽  
Pathogen Fitness ◽  
Enzymatic Mechanisms

Phytopathogenic bacteria possess an arsenal of effector proteins that enable them to subvert host recognition and manipulate the host to promote pathogen fitness. The type III secretion system (T3SS) delivers type III-secreted effector proteins (T3SEs) from bacterial pathogens such as Pseudomonas syringae, Ralstonia solanacearum, and various Xanthomonas species. These T3SEs interact with and modify a range of intracellular host targets to alter their activity and thereby attenuate host immune signaling. Pathogens have evolved T3SEs with diverse biochemical activities, which can be difficult to predict in the absence of structural data. Interestingly, several T3SEs are activated following injection into the host cell. Here, we review T3SEs with documented enzymatic activities, as well as T3SEs that facilitate virulence-promoting processes either indirectly or through non-enzymatic mechanisms. We discuss the mechanisms by which T3SEs are activated in the cell, as well as how T3SEs modify host targets to promote virulence or trigger immunity. These mechanisms may suggest common enzymatic activities and convergent targets that could be manipulated to protect crop plants from infection.

scholarly journals Characterization of a Unique Chromosomal Copper Resistance Gene Cluster from Xanthomonas campestris pv. vesicatoria

2005 ◽  
Vol 71 (12) ◽  
pp. 8284-8291 ◽  
Author(s):  
Huseyin Basim ◽  
Gerald V. Minsavage ◽  
Robert E. Stall ◽  
Jaw-Fen Wang ◽  
Savita Shanker ◽  
...  
Keyword(s):  
Pseudomonas Syringae ◽  
Resistance Genes ◽  
Xanthomonas Campestris ◽  
Sinorhizobium Meliloti ◽  
The United States ◽  
Copper Resistance ◽  
Pcr Analysis ◽  
Pepper Plant ◽  
Gene Encoding ◽  
Copper Resistance Genes

ABSTRACT We characterized the copper resistance genes in strain XvP26 of Xanthomonas campestris pv. vesicatoria, which was originally isolated from a pepper plant in Taiwan. The copper resistance genes were localized to a 7,652-bp region which, based on pulsed-field gel electrophoresis and Southern hybridization, was determined to be located on the chromosome. These genes hybridized only weakly, as determined by Southern analysis, to other copper resistance genes in Xanthomonas and Pseudomonas strains. We identified five open reading frames (ORFs) whose products exhibited high levels of amino acid sequence identity to the products of previously reported copper genes. Mutations in ORF1, ORF3, and ORF4 removed copper resistance, whereas mutations in ORF5 resulted in an intermediate copper resistance phenotype and insertions in ORF2 had no effect on resistance conferred to a copper-sensitive recipient in transconjugant tests. Based on sequence analysis, ORF1 was determined to have high levels of identity with the CopR (66%) and PcoR (63%) genes in Pseudomonas syringae pv. tomato and Escherichia coli, respectively. ORF2 and ORF5 had high levels of identity with the PcoS gene in E. coli and the gene encoding a putative copper-containing oxidoreductase signal peptide protein in Sinorhizobium meliloti, respectively. ORF3 and ORF4 exhibited 23% identity to the gene encoding a cation efflux system membrane protein, CzcC, and 62% identity to the gene encoding a putative copper-containing oxidoreductase protein, respectively. The latter two ORFs were determined to be induced following exposure to low concentrations of copper, while addition of Co, Cd, or Zn resulted in no significant induction. PCR analysis of 51 pepper and 34 tomato copper-resistant X. campestris pv. vesicatoria strains collected from several regions in Taiwan between 1987 and 2000 and nine copper-resistant strains from the United States and South America showed that successful amplification of DNA was obtained only for strain XvP26. The organization of this set of copper resistance genes appears to be uncommon, and the set appears to occur rarely in X. campestris pv. vesicatoria.

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