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Pectobacterium atrosepticum and Pectobacterium carotovorum Harbor Distinct, Independently Acquired Integrative and Conjugative Elements Encoding Coronafacic Acid that Enhance Virulence on Potato Stems.

Panda P, Vanga BR, Lu A, Fiers M, Fineran PC, Butler R, Armstrong K, Ronson CW, Pitman AR - Front Microbiol (2016)

Bottom Line: Much is known about the functions of the virulence determinants that ICEs harbor, but little is understood about the cryptic effects of ICEs on their host cell.As expected, deletion of HAI2 resulted in reduced blackleg symptoms in potato stems.Thus, the future spread of these ICEs via lateral gene transfer might contribute to an increase in the prevalence of blackleg-causing strains of P. carotovorum.

View Article: PubMed Central - PubMed

Affiliation: The Bio-Protection Research CentreLincoln, New Zealand; Plant Pathology, The New Zealand Institute for Plant and Food Research LimitedLincoln, New Zealand.

ABSTRACT
Integrative and conjugative elements (ICEs) play a central role in the evolution of bacterial virulence, their transmission between bacteria often leading to the acquisition of virulence factors that alter host range or aggressiveness. Much is known about the functions of the virulence determinants that ICEs harbor, but little is understood about the cryptic effects of ICEs on their host cell. In this study, the importance of horizontally acquired island 2 (HAI2), an ICE in the genome of Pectobacterium atrosepticum SCRI1043, was studied using a strain in which the entire ICE had been removed by CRISPR-Cas-mediated genome editing. HAI2 encodes coronafacic acid, a virulence factor that enhances blackleg disease of potato stems caused by P. atrosepticum SCRI1043. As expected, deletion of HAI2 resulted in reduced blackleg symptoms in potato stems. A subsequent screen for HAI2-related ICEs in other strains of the Pectobacterium genus revealed their ubiquitous nature in P. atrosepticum. Yet, HAI2-related ICEs were only detected in the genomes of a few P. carotovorum strains. These strains were notable as blackleg causing strains belonging to two different subspecies of P. carotovorum. Sequence analysis of the ICEs in different strains of both P. atrosepticum and P. carotovorum confirmed that they were diverse and were present in different locations on the genomes of their bacterial host, suggesting that the cfa cluster was probably acquired independently on a number of occasions via chromosomal insertion of related ICEs. Excision assays also demonstrated that the ICEs in both P. atrosepticum and P. carotovorum are mobilized from the host chromosome. Thus, the future spread of these ICEs via lateral gene transfer might contribute to an increase in the prevalence of blackleg-causing strains of P. carotovorum.

No MeSH data available.


Related in: MedlinePlus

Excision of HAI2-like elements in various strains of P. atrosepticum grown in vitro. Back-transformed mean log10 ratios (normalized to ECA0515) for attP are plotted against those for attB0515 on a log10 spaced axes for P. atrosepticum strains SCRI1043, NZEC22, ICMP 11525, ICMP 4398, and ICMP 1526. Cultures were grown in MM for 24 h, in triplicate. Three qPCR reactions were performed for each DNA sample and the mean ratio was plotted. The LSD between two means on the transformed scale was 0.29 for attP and was 0.44 for attB (at the 5% significance level with 10 degrees of freedom).
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Figure 2: Excision of HAI2-like elements in various strains of P. atrosepticum grown in vitro. Back-transformed mean log10 ratios (normalized to ECA0515) for attP are plotted against those for attB0515 on a log10 spaced axes for P. atrosepticum strains SCRI1043, NZEC22, ICMP 11525, ICMP 4398, and ICMP 1526. Cultures were grown in MM for 24 h, in triplicate. Three qPCR reactions were performed for each DNA sample and the mean ratio was plotted. The LSD between two means on the transformed scale was 0.29 for attP and was 0.44 for attB (at the 5% significance level with 10 degrees of freedom).

Mentions: At the same time, the formation of attP and attB0515 was measured by qPCR to determine the dynamics of ICE excision in P. atrosepticum. In these experiments, both attP and attB0515 were detected in all isolates (Figure 2). The ratios for attP and attB0515 were 2.25 × 10-5 and 8.45 × 10-6 in P. atrosepticum SCRI1043, consistent with the low frequency of HAI2 excision from the phe-tRNA gene immediately downstream of ECA0515 and the formation of the circular extrachromosomal form reported previously (Vanga et al., 2012, 2015). The relative proportion of attP compared to attB0515 calculated using these ratios was 2.67. A ratio above 1 suggested that the ICE may be replicating at low copy number.


Pectobacterium atrosepticum and Pectobacterium carotovorum Harbor Distinct, Independently Acquired Integrative and Conjugative Elements Encoding Coronafacic Acid that Enhance Virulence on Potato Stems.

Panda P, Vanga BR, Lu A, Fiers M, Fineran PC, Butler R, Armstrong K, Ronson CW, Pitman AR - Front Microbiol (2016)

Excision of HAI2-like elements in various strains of P. atrosepticum grown in vitro. Back-transformed mean log10 ratios (normalized to ECA0515) for attP are plotted against those for attB0515 on a log10 spaced axes for P. atrosepticum strains SCRI1043, NZEC22, ICMP 11525, ICMP 4398, and ICMP 1526. Cultures were grown in MM for 24 h, in triplicate. Three qPCR reactions were performed for each DNA sample and the mean ratio was plotted. The LSD between two means on the transformed scale was 0.29 for attP and was 0.44 for attB (at the 5% significance level with 10 degrees of freedom).
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4814525&req=5

Figure 2: Excision of HAI2-like elements in various strains of P. atrosepticum grown in vitro. Back-transformed mean log10 ratios (normalized to ECA0515) for attP are plotted against those for attB0515 on a log10 spaced axes for P. atrosepticum strains SCRI1043, NZEC22, ICMP 11525, ICMP 4398, and ICMP 1526. Cultures were grown in MM for 24 h, in triplicate. Three qPCR reactions were performed for each DNA sample and the mean ratio was plotted. The LSD between two means on the transformed scale was 0.29 for attP and was 0.44 for attB (at the 5% significance level with 10 degrees of freedom).
Mentions: At the same time, the formation of attP and attB0515 was measured by qPCR to determine the dynamics of ICE excision in P. atrosepticum. In these experiments, both attP and attB0515 were detected in all isolates (Figure 2). The ratios for attP and attB0515 were 2.25 × 10-5 and 8.45 × 10-6 in P. atrosepticum SCRI1043, consistent with the low frequency of HAI2 excision from the phe-tRNA gene immediately downstream of ECA0515 and the formation of the circular extrachromosomal form reported previously (Vanga et al., 2012, 2015). The relative proportion of attP compared to attB0515 calculated using these ratios was 2.67. A ratio above 1 suggested that the ICE may be replicating at low copy number.

Bottom Line: Much is known about the functions of the virulence determinants that ICEs harbor, but little is understood about the cryptic effects of ICEs on their host cell.As expected, deletion of HAI2 resulted in reduced blackleg symptoms in potato stems.Thus, the future spread of these ICEs via lateral gene transfer might contribute to an increase in the prevalence of blackleg-causing strains of P. carotovorum.

View Article: PubMed Central - PubMed

Affiliation: The Bio-Protection Research CentreLincoln, New Zealand; Plant Pathology, The New Zealand Institute for Plant and Food Research LimitedLincoln, New Zealand.

ABSTRACT
Integrative and conjugative elements (ICEs) play a central role in the evolution of bacterial virulence, their transmission between bacteria often leading to the acquisition of virulence factors that alter host range or aggressiveness. Much is known about the functions of the virulence determinants that ICEs harbor, but little is understood about the cryptic effects of ICEs on their host cell. In this study, the importance of horizontally acquired island 2 (HAI2), an ICE in the genome of Pectobacterium atrosepticum SCRI1043, was studied using a strain in which the entire ICE had been removed by CRISPR-Cas-mediated genome editing. HAI2 encodes coronafacic acid, a virulence factor that enhances blackleg disease of potato stems caused by P. atrosepticum SCRI1043. As expected, deletion of HAI2 resulted in reduced blackleg symptoms in potato stems. A subsequent screen for HAI2-related ICEs in other strains of the Pectobacterium genus revealed their ubiquitous nature in P. atrosepticum. Yet, HAI2-related ICEs were only detected in the genomes of a few P. carotovorum strains. These strains were notable as blackleg causing strains belonging to two different subspecies of P. carotovorum. Sequence analysis of the ICEs in different strains of both P. atrosepticum and P. carotovorum confirmed that they were diverse and were present in different locations on the genomes of their bacterial host, suggesting that the cfa cluster was probably acquired independently on a number of occasions via chromosomal insertion of related ICEs. Excision assays also demonstrated that the ICEs in both P. atrosepticum and P. carotovorum are mobilized from the host chromosome. Thus, the future spread of these ICEs via lateral gene transfer might contribute to an increase in the prevalence of blackleg-causing strains of P. carotovorum.

No MeSH data available.


Related in: MedlinePlus