Phoma stem canker disease on oilseed rape (Brassica napus) in China is caused by Leptosphaeria biglobosa ‘brassicae’

This document is the Accepted Manuscript version of the following article: Ze Liu, Akinwunmi O. Latunde-Dada, Avice M. Hall, Bruce D. L. Fitt, ‘Phoma stem canker disease on oilseed rape (Brassica napus) in China is caused by Leptosphaeria biglobosa ‘brassicae’’, European Journal of Plant Pathology, Vol. 140(4): 841-857, December 2014. The final publication is available at Springer via: http://dx.doi.org/10.1007/s10658-014-0513-7 © Koninklijke Nederlandse Planteziektenkundige Vereniging 2014Phoma stem canker of oilseed rape (Brassica napus) is a globally important disease that is caused by the sibling ascomycete species Leptosphaeria maculans and L. biglobosa. Sixty fungal isolates obtained from oilseed rape stems with phoma stem canker disease symptoms collected from four provinces in China in 1999, 2005 and 2006 were all identified as Leptosphaeria biglobosa, not L. maculans, by PCR diagnostics based on species-specific primers. There were no differences in cultural characteristics (e.g. pigmentation and in vitro growth) between these L. biglobosa isolates from China and those of 37 proven L. biglobosa isolates from Europe or Canada. In studies using amplified fragment length polymorphism (AFLP) markers, Chinese L. biglobosa populations were genetically more similar to European L. biglobosa populations than to the more diverse Canadian L. biglobosa populations. Sequencing of gene fragments of β-tubulin, actin and the internal transcribed spacer (ITS) region of rDNA from L. biglobosa isolates from China, Europe, Australia and Canada showed a closer taxonomic similarity of Chinese L. biglobosa to the European L. biglobosa ‘brassicae’ than to Canadian L. biglobosa ‘canadensis’ or to the Australian L. biglobosa ‘occiaustralensis’ or ‘australensis’ subclades. These results suggest that the Chinese L. biglobosa population in this study is in the same subclade as European L. biglobosa ‘brassicae’ populationsPeer reviewe

DNA-PK inhibitor wortmannin enhances DNA damage induced by bleomycin in V79 Chinese hamster cells

The fungal metabolite wortmannin (WM) is a potent and irreversible inhibitor of the enzyme DNA-dependent protein kinase (DNA-PK), a nuclear serine-threonine kinase, member of the phosphaditylinositol-3 kinase related kinase family. WM has been used in the last few years as a promising radiosensitizer mainly throughout cell survival experiments. However, few studies have addressed the role of DNA-PK inhibition in the repair of DNA lesions generated by antitumor agents, Bleomycin (BLM) is an antitumor agent used in the treatment of various neoplasia with a unique genotoxicity profile that mimics the ionizing radiation effects. In this study, we evaluated the effect of different concentrations of WM on the DNA damage induced by BLM, The cytokinesis-block micronucleus assay (CBMN) in V79 Chinese hamster cells was used as the end-point, WM significantly increased the frequency of micronucleated cells (%MNBN) by about 2.2-fold, the number of micronuclei per binucleated cell (MN/BN) by about 14-fold, and also changed the pattern of the distribution of micronuclei induced by BLM, The frequency of micronucleated cells with 2 MN per cell and with greater than or equal to3 MN per cell increased, whereas the frequency of micronucleated cells with 1 MN per cell decreased. WM was not genotoxic but decreased cell proliferation as assessed by the frequency of binucleated cells. Our results show that WM clearly enhances the efficacy of BLM in terms of DNA damage inflicted and therefore reinforces its use as a chemosensitizer. (C) 2002 Wiley-Liss, Inc

Wortmannin enhances the induction of micronuclei by low and high LET radiation

In mammalian cells, the repair of DNA double-strand breaks (DSBs) is mainly mediated by DNA non-homologous end joining. DNA-dependent protein kinase (DNA-PK), a nuclear serine-threonine kinase and a member of the phosphaditylinositol-3 kinase-related kinase family that is activated by DSBs, is a key component of this pathway. Wortmannin (WM) is known to be an irreversible and potent inhibitor of DNA-PK and has thus been proposed as an effective sensitizer for ionizing radiation and for radiomimetic compounds. The present study, using the cytokinesis block micronucleus assay, reports on the differential effect of WM on the repair of the DNA damage induced by low LET (Co-60 gamma-radiation) and high LET radiation by the boron neutron capture reaction (alpha and Li particles) in V79 Chinese hamster cells. Significant increases in the number of micronuclei per binucleated cell as well as in the frequency of micronucleated binucleated cells were observed in the presence of different concentrations of WM for high LET radiation from the boron neutron capture reaction. The increases observed reached a maximum of similar to2-fold in comparison with the respective controls. WM, however, had a more pronounced effect on Co-60 gamma-radiation-induced micronuclei, increasing the genotoxic damage from this radiation by similar to3- to 4-fold. These results are in general in agreement with the concept that DSBs induced by high LET radiation are not a more suitable substrate for the end joining processes mediated by DNA-PK, yet they do not preclude a role for DNA-PK in high LET-induced damage repair

Evaluation of the genotoxic effects of the boron neutron capture reaction in human melanoma cells using the cytokinesis block micronucleus assay

The present work reports on the genotoxicity of the boron neutron capture (BNC) reaction in human metastatic melanoma cells (A2058) assessed by the cytokinesis block micronucleus assay (CBMN) using p-borono-L-phenylalanine (BPA) as the boron delivery agent. Different concentrations of BPA (0.48, 1.2 and 2.4 mM) and different fluences of thermal neutrons were studied. Substantial genotoxic potential of alpha and lithium particles generated inside or near the malignant cell by the BNC reaction was observed in a dose-response manner as measured by the frequency of micronucleated binucleated melanoma cells and by the number of micronuclei (MN) per binucleated cell. The distribution of the number of MN per micronucleated binucleated cell was also studied. The BNC reaction clearly modifies this distribution, increasing the frequency of micronucleated cells with 2 and, especially, less than or equal to3 MN and conversely decreasing the frequency of micronucleated cells with 1 MN. A decrease in cell proliferation was also observed which correlated with MN formation. A discrete genotoxic and anti-proliferative contribution from both thermal neutron irradiation and BPA was observed and should be considered secondary. Additionally, V79 Chinese hamster cells (chromosomal aberrations assay) and human lymphocytes (CBMN assay) incubated with different concentrations of BPA alone did not show any evidence of genotoxicity. The presented results reinforce the usefulness of the CBMN assay as an alternative method for assessment of the deleterious effects induced by high LET radiation produced by the BNC reaction in human melanoma cells

World-wide importance of phoma stem canker (Leptosphaeria maculans and L-Biglobosa) on oilseed rape (Brassica napus)

Phoma stem canker is an internationally important disease of oilseed rape (Brassica napus, canola, rapeseed), causing serious losses in Europe, Australia and North America. UK losses of Euro56M per season are estimated using national disease survey data and a yield loss formula. Phoma stem canker pathogen populations comprise two main species, Leptosphaeria maculans, associated with damaging stem base cankers, and Leptosphaeria biglobosa, often associated with less damaging upper stem lesions. Both major gene and quantitative trait loci mediated resistance to L. maculans have been identified in B. napus, but little is known about resistance to L. biglobosa. Leptosphaeria maculans, which has spread into areas in North America and eastern Europe where only L. biglobosa was previously identified, now poses a threat to large areas of oilseed rape production in Asia. Epidemics are initiated by air-borne ascospores; major gene resistance to initial infection by L. maculans operates in the leaf lamina of B. napus. It is not clear whether the quantitative trait loci involved in the resistance to the pathogen that can be assessed only at the end of the season operate in the leaf petioles or stems. In countries where serious phoma stem canker epidemics occur, a minimum standard for resistance to L. maculans is included in national systems for registration of cultivars. This review provides a background to a series of papers on improving strategies for managing B. napus resistance to L. maculans, which is a model system for studying genetic interactions between hemi-biotrophic pathogens and their hostsPeer reviewe