Phytophthora captiosa sp. nov. and P. fallax sp. nov. causing crown dieback of Eucalyptus in New Zealand

A locally severe crown disease of exotic plantation Eucalyptus trees has been recorded periodically in New Zealand since 1986. Symptoms include leaf spots, petiole infection and twig and small branch lesions. Outbreaks of disease are episodic and individual trees may show marked variation in crown symptoms ranging from unaffected to total defoliation. Two previously unknown species of Phytophthora are associated with the disease. These are described and formally designated here as P. captiosa, from Eucalyptus botryoides and E. saligna; and P. fallax, from E. delegatensis, E. fastigata, E. nitens and E. regnans. Both P. captiosa and P. fallax have non-papillate, non-caducous sporangia and both are self-fertile. Phylogenetic analysis on the basis of ITS rDNA sequence data indicates they are closely related to each other but evolutionarily distant from the majority of described Phytophthora taxa. They share a common ancestor with another assemblage of Phytophthora lineages that includes P. insolita, P. macrochlamydospora and P. richardiae. Sporulation of P. captiosa and P. fallax has not been observed in the field. The mode of infection and spread of these non-caducous Phytophthora species in the eucalypt tree canopy remains unknown. This issue, and the possible geographic origins of these two Phytophthora species are discussed.     

Endophytic bacterial flora in root and stem tissues of black pepper (Piper nigrum L.) genotype: isolation, identification and evaluation against Phytophthora capsici

Aim:  To isolate and identify black pepper ( Piper nigrum L) associated endophytic bacteria antagonistic to Phytophthora capsici causing foot rot disease.
Methods and Results:  Endophytic bacteria (74) were isolated, characterized and evaluated against P. capsici . Six genera belong to Pseudomonas spp (20 strains), Serratia (1 strain), Bacillus spp. (22 strains), Arthrobacter spp. (15 strains), Micrococcus spp. (7 strains), Curtobacterium sp. (1 strain) and eight unidentified strains were isolated from internal tissues of root and stem. Three isolates, IISRBP 35, IISRBP 25 and IISRBP 17 were found effective for Phytophthora suppression in multilevel screening assays which recorded over 70% disease suppression in green house trials. A species closest match (99% similarity) of IISRBP 35 was established as Pseudomonas aeruginosa ( Pseudomonas EF568931), IISRBP 25 as P. putida ( Pseudomonas EF568932), and IISRBP 17 as Bacillus megaterium ( B. megaterium EU071712) based on 16S rDNA sequencing.
Conclusion:  Black pepper associated P. aeruginosa , P. putida and B. megaterium were identified as effective antagonistic endophytes for biological control of Phytophthora foot rot in black pepper.
Significance and Impact of the Study:  This work provides the first evidence for endophytic bacterial diversity in black pepper stem and roots, with biocontrol potential against P. capsici infection.     

Influence of the medium-solidifying agent, the nutrient, and the genotype on the production of gametangia by Phytophthora ramorum in vitro

The effect of different parameters, including the type of nutrients, the quality of the gelling agent, and the genotype of the strain, were evaluated in the production of gametangia by Phytophthora ramorum in vitro. By comparing different agar sources on a carrot-based medium, a delay or a failure in the production of oospores was observed in pairings carried out on media supplemented with technical agar. In contrast, oospores were produced on other agar types, the production on media supplemented with agarose being slightly higher. The formation of gametangia was also influenced by the genotype of the strains involved in the pairing. A European A1 strain producing very few chlamydospores was found to be a better mating partner than other A1 strains. Using a carrot-agarose medium and selected genotypes, all European isolates were characterized in terms of mating type. A macroscopic experiment highlighted a particular spatial distribution of P. ramorum oospores in vitro. A method using polycarbonate membrane was evaluated to assess the selfing ability of P. ramorum.     

Evaluation of several commercial biocontrol products on European and North American populations of Phytophthora ramorum

Five commercially available biological control products were tested in vitro with seven isolates of Phytophthora ramorum from North American (NA1, NA2), and European (EU1) populations. The in vitro tests included dual culture methods and detached leaf assays on wounded Rhododendron and Camellia leaves. Variability in response to biocontrol agents among isolates of P. ramorum from North American and European populations was examined. In dual culture tests, both Bacillus subtilis products (Companion® and Serenade®) resulted in better inhibition of the NA1 group than NA2 and EU1. Actinovate® (Streptomyces lydicus) was the least effective of the three bacterial biocontrol agents and there was no difference in percent inhibition among P. ramorum lineages. Two products containing Trichoderma spp. were tested: Plant Helper® (T. atroviride) caused 100% inhibition of all lineages of P. ramorum, while SoilGard? (T. virens) was only about 30% effective. There was great variability among P. ramorum isolates in their response to biocontrol agents. All treatments reduced P. ramorum lesion size on both Rhododendron and Camellia. Combined treatments of Actinovate® with one other BCA did not perform as well as either treatment used individually. Best results were obtained with Serenade® on Rhododendron and Camellia foliage, especially against the NA1 group. Lack of a linear relationship between percent inhibition of P. ramorum by BCAs in vitro and foliar treatments on detached Rhododendron and Camellia leaves indicates that in vitro testing is a poor predictor of BCA performance on plant material.     

Development of New Oomycete Taxon‐specific Mitochondrial Cytochrome b Region Primers for Use in Phylogenetic and Phylogeographic Studies

Here, we describe the development of an oomycete‐specific primer pair for amplification of the cytochrome b region in plant pathogenic species that span the order Peronosporales (Phytophthora spp., downy mildews). Because of the high number of variable sites at both inter‐ and intra‐specific levels this marker provides a powerful tool for population genetics and phylogenetic studies in this taxa. We also demonstrate its potential compared with other oomycete‐specific mitochondrial markers currently available.     

大豆疫霉菌一个DNA指纹分析重复序列探针的鉴定

【目的】大豆疫霉菌指纹分析的建立和黑龙江与新疆大豆疫霉菌群体的群体遗传分析。【方法】利用生物信息学方法寻找大豆疫霉菌(Phytophthora sojae)的中度重复序列,并对黑龙江和新疆大豆疫霉菌进行DNA指纹分析。【结果】分析得到一个中度重复序列,定名为PS1227。Southern blot分析表明PS1227在大豆疫霉菌基因组中约有34条可辨的介于1.5-23kb之间的杂交条带,其中21个杂交条带在49个供试菌系中表现多态性。单游动孢子分析表明PS1227指纹特征在病菌无性生殖阶段表现稳定。利用PS1227标记,本实验发现采自黑龙江HP4002、SY6和GJ0105菌系分别与新疆的DW303、71228和71222菌系具有完全相同的指纹特征。【结论】获得一个可用于大豆疫霉菌流行学和群体生物学研究的指纹分析序列PS1227,在分子水平证实了新疆大豆疫霉菌可能由黑龙江传入。     

The role of salicylic acid and jasmonic acid in pathogen defence

Phytohormones are not only instrumental in regulating developmental processes in plants but also play important roles for the plant's responses to biotic and abiotic stresses. In particular, abscisic acid, ethylene, jasmonic acid, and salicylic acid have been shown to possess crucial functions in mediating or orchestrating stress responses in plants. Here, we review the role of salicylic acid and jasmonic acid in pathogen defence responses with special emphasis on their function in the solanaceous plant potato.     

Microbial community responses associated with the development of oomycete plant pathogens on tomato roots in soilless growing systems

AIMS: To determine the spread of different oomycete pathogens in hydroponic, soilless tomato growing systems and their impact on established microbial communities, as baseline studies prior to future introduction of microbial inoculants for disease suppression. METHODS AND RESULTS: The oomycete pathogens, Pythium group F, Pythium aphanidermatum and Phytophthora cryptogea, were introduced into small-scale recirculating tomato growing systems containing rockwool 6 weeks after set-up when roots were well-established. Two weeks later, half of the systems were switched over to run-to-waste. Pythium aphanidermatum spreads the fastest, Pythium group F the slowest and Ph. cryptogea was intermediate in its spread. The switch to run-to-waste had no effect on pathogen recovery. Microbial communities, monitored by dilution plating, were well-established at the first sampling, 6 weeks after set-up and although differences in community levels were found between experiments, changes during any one experiment were small, generally less than 1 log10 CFU g(-1) for bacteria. Pathogen introduction increased microbial community levels in roots but the switch to run-to-waste had no effect. Analysis of bacterial communities through amplification of a fragment of the 16S rRNA gene and DGGE profiling showed that different communities were established within each pathogen experiment and that different communities were established on roots, rockwool and in nutrient solutions. However, no significant changes in microbial profiles were found over time in any experiment. CONCLUSIONS: In these systems, the microbial communities were well-established 6 weeks after set-up and were resistant to biological and physical perturbation. SIGNIFICANCE AND IMPACT OF THE STUDY: The implication for microbial inoculation of such systems for disease suppression is that the micro-organisms would either have to be introduced very early during the set-up of the system or be able to replace an established but variable community.     

Heritability of Phenols in the Resistance of Theobroma cacao against Phytophthora megakarya, the Causal Agent of Black Pod Disease

The black pod disease caused by Phytophthora megakarya is responsible for 80% of the cocoa production loss in Cameroon. To assess the resistance of cocoa plants against this pathogen, necrotic lesions, phenolic content and qualitative alteration of phenolics were conducted in ICS84 and ICS95 clones (two Trinitario introduced from Trinidad) and their hybrids (families F30 and F25) derived from reciprocal cross breeding between these two parental clones after inoculation. The existence of strong hybrid vigour has been shown. Ninety percentage of the hybrid's genotypes manifested a positive heterosis effect for the development of lesion size. This suggests the existence of hybrid vigour with a genetic additive effect. F3086, F2509, F2552 and F2586 hybrids were characterized by localized lesions. Those hybrids genotypes can be considered as elite clones. In relation to analysis of total phenolics and lesion size, no maternal effect was detected in the transmission of these characters. A significant and negative correlation (r = −0.683) (P < 0.01) has been observed between necrosis evolution and phenolics accumulation. The values of the heritability of lesion size and the total phenolic content in offsprings did not permit to show the maternal effect. Qualitative analyses of phenolics showed high flavonones content in cocoa leaves. Qualitative analyses of phenolics in ICS84, ICS95 clones and their hybrids showed a modification of the phenolics profiles, notably concerning apigenin and luteolin derivatives due to the inoculation. These compounds, along with others that were not identified, could have a role in the reaction and mechanism of defence of cocoa against P. megakarya.     

Successful first and second backcrosses of S. nigrum (+) S. tuberosum somatic hybrids with both Solanum parents

Somatic hybrids of Solanum nigrum (+) 2× potato were successfully crossed with S. nigrum and with potato. First and second backcross progeny with S. nigrum could easily be obtained. One of the BC1 genotypes was already self-fertile. Backcrosses with potato had a much lower success rate. Only pollinations with tetraploid potato resulted in seed-containing berries. Two BC1 genotypes were obtained after 4362 pollinations from which 505 ovules were cultured. The first BC1 genotype grew vigorously in vitro and in the greenhouse and flowered abundantly. The second BC1 showed many abnormalities and dropped its flowers before anthesis. The first BC1 was again crossed with tetraploid potato and in this generation also the success rate was low. Over 5000 pollinations resulted in 1750 berries from which over 3000 ovules were obtained. Twelve plants germinated from these ovules and they were not as vigorous in vitro and in vivo as the BC1 parent. Some of the BC2 genotypes were used for further backcrosses but no BC3 plants were obtained. BC1 and BC2 genotypes that resulted from the backcross program with potato were tested for resistance to Phytophthora infestans. The BC1 genotype was as resistant as the S. nigrum fusion parent, but among the eight BC2 genotypes scored six were resistant, whereas two genotypes showing lesions were susceptible. Received: 19 October 1998 / Revision accepted: 20 April 1999