Transcriptional responses of Norway spruce (Picea abies) inner sapwood against Heterobasidion parviporum.

The white-rot fungus Heterobasidion parviporum Niemelä & Korhonen establishes a necrotrophic interaction with Norway spruce (Picea abies (L.) H.Karst.) causing root and butt rot and growth losses in living trees. The interaction occurs first with the bark and the outer sapwood, as the pathogen enters the tree via wounds or root-to-root contacts. Later, when the fungus reaches the heartwood, it spreads therein creating a decay column, and the interaction mainly occurs in the inner sapwood where the tree creates a reaction zone. While bark and outer sapwood interactions are well studied, little is known about the nature of the transcriptional responses leading to the creation of a reaction zone. In this study, we sampled bark and sapwood both proximal and distal to the reaction zone in artificially inoculated and naturally infected trees. We quantified gene expression levels of candidate genes in secondary metabolite, hormone biosynthesis and signalling pathways using quantitative polymerase chain reaction. An up-regulation of mainly the phenylpropanoid pathway and jasmonic acid biosynthesis was found at the inoculation site, when inoculations were compared with wounding. We found that transcriptional responses in inner sapwood were similar to those reported upon infection through the bark. Our data suggest that the defence mechanism is induced due to direct fungal contact irrespective of the tissue type. Understanding the nature of these interactions is important when considering tree breeding-based resistance strategies to reduce the spread of the pathogen between and within trees.

Host responses in Norway spruce roots induced to the pathogen Ceratocystis polonica are evaded or suppressed by the ectomycorrhizal fungus Laccaria bicolor.

The outcome of a compatible mycorrhizal interaction is different from that in a compatible plant-pathogen interaction; however, it is not clear what mechanisms are used to evade or suppress the host defence. The aim of this work is to reveal differences between the interaction of Norway spruce roots to the pathogen Ceratocystis polonica and the ectomycorrhizal Laccaria bicolor, examine if L. bicolor is able to evade inducing host defence responses typically induced by pathogens, and test if prior inoculation with the ectomycorrhizal fungus affects the outcome of a later challenge with the pathogen. The pathogen was able to invade the roots and caused extensive necrosis, leading to seedling death, with or without prior inoculation with L. bicolor. The ectomycorrhizal L. bicolor colonised primary roots of the Norway spruce seedlings by partly covering, displacing and convoluting the cells of the outer root cortex, leaving the seedlings healthy. We detected increased total peroxidase activity, and staining indicating increased lignification in roots as a response to C. polonica. In L. bicolor inoculated roots there was no increase in total peroxidase activity, but an additional highly acidic peroxidase isoform appeared that was not present in healthy roots, or in roots invaded by the pathogen. Increased protease activity was detected in roots colonised by C. polonica, but little protease activity was detected in L. bicolor inoculated roots. These results suggest that the pathogen efficiently invades the roots despite the induced host defence responses, while L. bicolor suppresses or evades inducing such host responses in this experimental system.

Isolation of the first putative peroxidase cDNA from a conifer and the local and systemic accumulation of related proteins upon pathogen infection.

Peroxidases are associated with the active defence reactions in higher plants in response to foreign organisms. They are involved in the oxidation of phenolic compounds in cell walls, polymerization of lignin and suberin, and in several other oxidation processes but the exact function of individual peroxidases is not known. We have isolated a cDNA encoding the putative defence-related and basic plant peroxidase SPI2 (spruce pathogen-induced 2), with an estimated molecular mass of 34 kDa, from roots of Norway spruce (Picea abies) seedlings. This is the first description of the isolation of a complete cDNA encoding a putative peroxidase from a gymnosperm. The transcript was present in the roots of healthy seedlings, and during infection with the pathogen Pythium dimorphum there was a rapid initial increase followed by a dramatic reduction of the transcript. The 34 kDa mature SPI2 protein was detected in both the developing root and shoot of healthy seedlings and increased amounts of SPI2 and increased accumulation of highly basic peroxidase isoforms was observed in roots after infection. In addition, two SPI2-related proteins with apparent molecular masses of 38 and 39 kDa, were also detected. Both these proteins accumulated in roots only after infection, and the 39 kDa protein was in addition detected in shoots of root-infected seedlings. Thus, both SPI2 and the SPI2-related proteins accumulate as a local response, in roots, and as a systemic response to infection the 39 kDa protein accumulates in the shoot.

Bacteriophage P4 capsid-size determination and its relationship to P2 helper interference.

The sid (size determination) gene product of phage P4 is known to be involved in capsid-size determination. Moreover, the capsid-size determination function interferes with the lytic development of its helper P2, presumably because the helper genome is too large to be packaged into P4-size capsids. In order to study P4-specified helper interference, we cloned the sid gene for expression during phage infection. Even though gpSid restores the capsid-size determination function of a sid defective P4 mutant, we find that gpSid alone is not sufficient to establish full interference of helper P2 phage production. Complete helper interference requires some P4 function in addition to gpSid. Complementation tests show that none of the known P4 genes display this property. We propose that P4 encodes a yet-unidentified function that in concert with gpSid establishes full P2 helper interference at the level of capsid-size determination.