Worldwide forest surveys reveal forty-three new species in Phytophthora major Clade 2 with fundamental implications for the evolution and biogeography of the genus and global plant biosecurity.

During 25 surveys of global Phytophthora diversity, conducted between 1998 and 2020, 43 new species were detected in natural ecosystems and, occasionally, in nurseries and outplantings in Europe, Southeast and East Asia and the Americas. Based on a multigene phylogeny of nine nuclear and four mitochondrial gene regions they were assigned to five of the six known subclades, 2a-c, e and f, of Phytophthora major Clade 2 and the new subclade 2g. The evolutionary history of the Clade appears to have involved the pre-Gondwanan divergence of three extant subclades, 2c, 2e and 2f, all having disjunct natural distributions on separate continents and comprising species with a soilborne and aquatic lifestyle and, in addition, a few partially aerial species in Clade 2c; and the post-Gondwanan evolution of subclades 2a and 2g in Southeast/East Asia and 2b in South America, respectively, from their common ancestor. Species in Clade 2g are soilborne whereas Clade 2b comprises both soil-inhabiting and aerial species. Clade 2a has evolved further towards an aerial lifestyle comprising only species which are predominantly or partially airborne. Based on high nuclear heterozygosity levels ca. 38 % of the taxa in Clades 2a and 2b could be some form of hybrid, and the hybridity may be favoured by an A1/A2 breeding system and an aerial life style. Circumstantial evidence suggests the now 93 described species and informally designated taxa in Clade 2 result from both allopatric non-adaptive and sympatric adaptive radiations. They represent most morphological and physiological characters, breeding systems, lifestyles and forms of host specialism found across the Phytophthora clades as a whole, demonstrating the strong biological cohesiveness of the genus. The finding of 43 previously unknown species from a single Phytophthora clade highlight a critical lack of information on the scale of the unknown pathogen threats to forests and natural ecosystems, underlining the risk of basing plant biosecurity protocols mainly on lists of named organisms. More surveys in natural ecosystems of yet unsurveyed regions in Africa, Asia, Central and South America are needed to unveil the full diversity of the clade and the factors driving diversity, speciation and adaptation in Phytophthora. Taxonomic novelties: New species: Phytophthora amamensis T. Jung, K. Kageyama, H. Masuya & S. Uematsu, Phytophthora angustata T. Jung, L. Garcia, B. Mendieta-Araica, & Y. Balci, Phytophthora balkanensis I. Milenkovic, Z. Tomic, T. Jung & M. Horta Jung, Phytophthora borneensis T. Jung, A. Durán, M. Tarigan & M. Horta Jung, Phytophthora calidophila T. Jung, Y. Balci, L. Garcia & B. Mendieta-Araica, Phytophthora catenulata T. Jung, T.-T. Chang, N.M. Chi & M. Horta Jung, Phytophthora celeris T. Jung, L. Oliveira, M. Tarigan & I. Milenkovic, Phytophthora curvata T. Jung, A. Hieno, H. Masuya & M. Horta Jung, Phytophthora distorta T. Jung, A. Durán, E. Sanfuentes von Stowasser & M. Horta Jung, Phytophthora excentrica T. Jung, S. Uematsu, K. Kageyama & C.M. Brasier, Phytophthora falcata T. Jung, K. Kageyama, S. Uematsu & M. Horta Jung, Phytophthora fansipanensis T. Jung, N.M. Chi, T. Corcobado & C.M. Brasier, Phytophthora frigidophila T. Jung, Y. Balci, K. Broders & I. Milenkovic, Phytophthora furcata T. Jung, N.M. Chi, I. Milenkovic & M. Horta Jung, Phytophthora inclinata N.M. Chi, T. Jung, M. Horta Jung & I. Milenkovic, Phytophthora indonesiensis T. Jung, M. Tarigan, L. Oliveira & I. Milenkovic, Phytophthora japonensis T. Jung, A. Hieno, H. Masuya & J.F. Webber, Phytophthora limosa T. Corcobado, T. Majek, M. Ferreira & T. Jung, Phytophthora macroglobulosa H.-C. Zeng, H.-H. Ho, F.-C. Zheng & T. Jung, Phytophthora montana T. Jung, Y. Balci, K. Broders & M. Horta Jung, Phytophthora multipapillata T. Jung, M. Tarigan, I. Milenkovic & M. Horta Jung, Phytophthora multiplex T. Jung, Y. Balci, K. Broders & M. Horta Jung, Phytophthora nimia T. Jung, H. Masuya, A. Hieno & C.M. Brasier, Phytophthora oblonga T. Jung, S. Uematsu, K. Kageyama & C.M. Brasier, Phytophthora obovoidea T. Jung, Y. Balci, L. Garcia & B. Mendieta-Araica, Phytophthora obturata T. Jung, N.M. Chi, I. Milenkovic & M. Horta Jung, Phytophthora penetrans T. Jung, Y. Balci, K. Broders & I. Milenkovic, Phytophthora platani T. Jung, A. Pérez-Sierra, S.O. Cacciola & M. Horta Jung, Phytophthora proliferata T. Jung, N.M. Chi, I. Milenkovic & M. Horta Jung, Phytophthora pseudocapensis T. Jung, T.-T. Chang, I. Milenkovic & M. Horta Jung, Phytophthora pseudocitrophthora T. Jung, S.O. Cacciola, J. Bakonyi & M. Horta Jung, Phytophthora pseudofrigida T. Jung, A. Durán, M. Tarigan & M. Horta Jung, Phytophthora pseudoccultans T. Jung, T.-T. Chang, I. Milenkovic & M. Horta Jung, Phytophthora pyriformis T. Jung, Y. Balci, K.D. Boders & M. Horta Jung, Phytophthora sumatera T. Jung, M. Tarigan, M. Junaid & A. Durán, Phytophthora transposita T. Jung, K. Kageyama, C.M. Brasier & H. Masuya, Phytophthora vacuola T. Jung, H. Masuya, K. Kageyama & J.F. Webber, Phytophthora valdiviana T. Jung, E. Sanfuentes von Stowasser, A. Durán & M. Horta Jung, Phytophthora variepedicellata T. Jung, Y. Balci, K. Broders & I. Milenkovic, Phytophthora vietnamensis T. Jung, N.M. Chi, I. Milenkovic & M. Horta Jung, Phytophthora ×australasiatica T. Jung, N.M. Chi, M. Tarigan & M. Horta Jung, Phytophthora ×lusitanica T. Jung, M. Horta Jung, C. Maia & I. Milenkovic, Phytophthora ×taiwanensis T. Jung, T.-T. Chang, H.-S. Fu & M. Horta Jung. Citation: Jung T, Milenkovic I, Balci Y, Janousek J, Kudlácek T, Nagy ZÁ, Baharuddin B, Bakonyi J, Broders KD, Cacciola SO, Chang T-T, Chi NM, Corcobado T, Cravador A, Dordevic B, Durán A, Ferreira M, Fu C-H, Garcia L, Hieno A, Ho H-H, Hong C, Junaid M, Kageyama K, Kuswinanti T, Maia C, Májek T, Masuya H, Magnano di San Lio G, Mendieta-Araica B, Nasri N, Oliveira LSS, Pane A, Pérez-Sierra A, Rosmana A, Sanfuentes von Stowasser E, Scanu B, Singh R, Stanivukovic Z, Tarigan M, Thu PQ, Tomic Z, Tomsovský M, Uematsu S, Webber JF, Zeng H-C, Zheng F-C, Brasier CM, Horta Jung M (2024). Worldwide forest surveys reveal forty-three new species in Phytophthora major Clade 2 with fundamental implications for the evolution and biogeography of the genus and global plant biosecurity. Studies in Mycology 107: 251-388. doi: 10.3114/sim.2024.107.04.

Extensive morphological and behavioural diversity among fourteen new and seven described species in Phytophthora Clade 10 and its evolutionary implications.

During extensive surveys of global Phytophthora diversity 14 new species detected in natural ecosystems in Chile, Indonesia, USA (Louisiana), Sweden, Ukraine and Vietnam were assigned to Phytophthora major Clade 10 based on a multigene phylogeny of nine nuclear and three mitochondrial gene regions. Clade 10 now comprises three subclades. Subclades 10a and 10b contain species with nonpapillate sporangia, a range of breeding systems and a mainly soil- and waterborne lifestyle. These include the previously described P. afrocarpa, P. gallica and P. intercalaris and eight of the new species: P. ludoviciana, P. procera, P. pseudogallica, P. scandinavica, P. subarctica, P. tenuimura, P. tonkinensis and P. ukrainensis. In contrast, all species in Subclade 10c have papillate sporangia and are self-fertile (or homothallic) with an aerial lifestyle including the known P. boehmeriae, P. gondwanensis, P. kernoviae and P. morindae and the new species P. celebensis, P. chilensis, P. javanensis, P. multiglobulosa, P. pseudochilensis and P. pseudokernoviae. All new Phytophthora species differed from each other and from related species by their unique combinations of morphological characters, breeding systems, cardinal temperatures and growth rates. The biogeography and evolutionary history of Clade 10 are discussed. We propose that the three subclades originated via the early divergence of pre-Gondwanan ancestors > 175 Mya into water- and soilborne and aerially dispersed lineages and subsequently underwent multiple allopatric and sympatric radiations during their global spread. Citation: Jung T, Milenkovic I, Corcobado T, et al. 2022. Extensive morphological and behavioural diversity among fourteen new and seven described species in Phytophthora Clade 10 and its evolutionary implications. Persoonia 49: 1-57. https://doi.org/10.3767/persoonia.2022.49.01.

Population structure of the ash dieback pathogen, Hymenoscyphus fraxineus, in relation to its mode of arrival in the UK.

The ash dieback fungus, Hymenoscyphus fraxineus, a destructive, alien pathogen of common ash (Fraxinus excelsior), has spread across Europe over the past 25 years and was first observed in the UK in 2012. To investigate the relationship of the pathogen's population structure to its mode of arrival, isolates were obtained from locations in England and Wales, either where established natural populations of ash had been infected by wind-dispersed ascospores or where the fungus had been introduced on imported planting stock. Population structure was determined by tests for vegetative compatibility (VC), mating type and single-nucleotide polymorphisms (SNPs). VC heterogeneity was high at all locations, with 96% of isolate pairings being incompatible. Frequencies of the MAT1-1-1 and MAT1-2-1 idiomorphs were approximately equal, consistent with H. fraxineus being an obligate outbreeder. Most SNP variation occurred within study location and there was little genetic differentiation between the two types of location in the UK, or between pathogen populations in the UK and continental Europe. There was modest differentiation between UK subpopulations, consistent with genetic variation between source populations in continental Europe. However, there was no evidence of strong founder effects, indicating that numerous individuals of H. fraxineus initiated infection at each location, regardless of the route of pathogen transmission. The ssRNA virus HfMV1 was present at moderate to high frequencies in all UK subpopulations. The results imply that management of an introduced plant pathogen requires action against its spread at the continental level involving coordinated efforts by European countries.

Phytophthora ramorum on Quercus ilex in the United Kingdom.

Phytophthora ramorum causes bleeding cankers of trunks of trees native to the west coast of the United States (i.e., Quercus kelloggii, Q. parvula var. shrevei, and Lithocarpus densiflorus). In the United Kingdom so far, bleeding cankers caused by inner bark infections have been found on Aesculus hippocastanum, Fagus sylvatica, Q. cerris, Q. falcata, and Q. petraea ( http://rapra.csl.gov.uk [2005]). Shoot tip dieback (ramorum dieback) and foliar necrosis (ramorum leaf blight) are other diseases caused by the pathogen on understory and ornamental plants (3). Inoculum is produced on infected shoots and leaves of foliar hosts but not on bole cankers (1). Foliar hosts are thus critical in initiating and maintaining epidemics of tree mortality resulting from lethal bark cankers. Ramorum dieback and blight occurs in Europe on genera Rhododendron, Camellia, Kalmia, Pieris, and Viburnum (http://rapra.csl.gov.uk [2005]), and now we report these diseases on foliage and shoots of holm oaks (Quercus ilex) in Cornwall (UK). First discovered in November 2003, infected young leaves had a water-soaked, dull gray appearance, and petioles were blackened. Lesions started at leaf margins, tips, or petioles, often progressing into the midrib veins. Initial infections also occurred on shoots and extended into the petioles. If shoots were infected, they were blackened at first, but later in the season clusters of dry, dead leaves and twigs characterized branch tips. Infected mature leaves bore dry, reddish-brown, restricted lesions. P. ramorum (A1 sexual compatibility type belonging to the European population) was isolated and confirmed by morphological studies, ITS sequence (GenBank Accession No. AY924253), and amplified fragment length polymorphism analyses. Lesions developed on detached leaves dipped for 10 sec in inoculum (4 × 105 zoospores per ml) and incubated in moist chambers at 20°C for 6 days (2). Two isolates were used (four leaves per isolate). The pathogen was reisolated, and the tests were repeated twice. Koch's postulates were also successfully completed once on foliage attached to saplings. To our knowledge, this is the first report of P. ramorum on holm oak. So far, at least 24 holm oaks are infected at various woodland and garden sites in the United Kingdom; infected rhododendrons have also been found at these sites. P. ramorum has also been recorded on saplings in nurseries. The high sporulation potential, the evergreen nature of leaves, and susceptible shoots indicate that holm oak could be a significant source of inoculum for other hosts. References: (1) J. M. Davidson et al. Phytopathology 95:587, 2005. (2) S. Denman et al. Plant Pathol. 54:512, 2005. (3) E. M. Hansen et al. Plant Dis. 89:63, 2005.

A molecular phylogeny of Phytophthora and related oomycetes.

Phylogenetic relationships among 50 Phytophthora species and between Phytophthora and other oomycetes were examined on the basis of the ITS sequences of genomic rDNA. Phytophthora grouped with Pythium, Peronospora, and Halophytophthora, distant from genera in the Saprolegniales. Albugo was intermediate between these two groups. Unlike Pythium, Phytophthora was essentially monophyletic, all but three species forming a cluster of eight clades. Two clades contained only species with nonpapillate sporangia. The other six clades included either papillate and semipapillate, or semipapillate and nonpapillate types, transcending traditional morphological groupings, which are evidently not natural assemblages. Peronospora was related to P. megakarya and P. palmivora and appears to be derived from a Phytophthora that has both lost the ability to produce zoospores and become an obligate biotroph. Three other Phytophthoras located some distance from the main Phytophthora-Peronospora cluster probably represent one or more additional genera.

Detection of an RNA-dependent RNA polymerase in mitochondria from a mitovirus-infected isolate of the Dutch Elm disease fungus, Ophiostoma novo-ulmi.

RNA-dependent RNA polymerase (RdRp) activity has been detected in mitochondria from an isolate of Ophiostoma novo-ulmi infected with O. novo-ulmi mitovirus 6 (OnuMV6). The reaction products corresponded to the double-stranded and single-stranded forms of OnuMV6 RNA. Western blot analysis using antibodies raised against a conserved RdRp region has detected a protein of ca. 80 kDa in OnuMV6-infected mitochondria, close to the predicted size of the OnuMV6 RdRp. No RdRp activity or protein was detected in mitochondria from an uninfected O. novo-ulmi isolate. This is the first detection of a virus RdRp in fungal mitochondria and the results are consistent with the use of UGA tryptophan codons in its synthesis.

Regeneration of phenotypically normal English elm (Ulmus procera) plantlets following transformation with an Agrobacterium tumefaciens binary vector.

A transformation system was developed for English elm (Ulmus procera Salisbury) using Agrobacterium tumefaciens C58 pMP90 p35SGUS/INTRON, allowing for the transfer of foreign genes and regeneration of phenotypically normal elm plantlets. The PCR analysis indicated that both nptII and uidA genes were stably inserted in the plant genome. beta-Glucuronidase histochemical and fluorimetric assays revealed expression of the uidA gene in the shoots, leaves, stems and roots of regenerated transgenic plants. The DNA-DNA hybridizations confirmed the presence of the uidA gene in regenerant plants. Factors influencing successful transformation and regeneration of elms included: identifying gene-transfer-proficient Agrobacterium strains for use with elms; developing an infection protocol allowing T-DNA transfer while retaining the ability to remove inciting bacteria; and identifying selection conditions to eliminate non-transformed material and choice of regeneration medium to allow shoot production. The potential utility of an effective elm transformation and regeneration system in the control of Dutch elm disease is discussed.

Two natural cerato-ulmin (CU)-deficient mutants of Ophiostoma novo-ulmi: one has an introgressed O. ulmi cu gene, the other has an O. novo-ulmi cu gene with a mutation in an intron splice consensus sequence.

Summary The nucleotide sequences of the cerato-ulmin (cu) genes of two naturally occurring pathogenic CU-deficient mutants, PG470 and MAFf8, of the Dutch elm disease fungus, Ophiostoma novo-ulmi, were determined. The PG470 cu gene sequence was identical to that of CU-secreting isolates of O. novo-ulmi, except for a G to A mutation in the GT splice consensus sequence at the start of intron 1, suggesting that the CU deficiency was due to a splicing defect in the premRNA. In contrast, the MAFf8 cu gene showed a 99.1% sequence identity with cu genes of O. ulmi isolates, but only 92.8% sequence identity with cu genes of CU-secreting isolates of O. novo-ulmi, and in a dendrogram clustered with cu gene sequences of O. ulmi isolates with 100% bootstrap support. Restriction fragment length polymorphisms of the ribosomal RNA region, random amplified polymorphic DNA markers, and many biological properties of MAFf8, including pathogenicity, were typical of O. novo-ulmi. It is therefore likely that the cu gene of MAFf8 has been introgressed from O. ulmi, probably as a result of rare hybrid formation between O. ulmi and O. novo-ulmi, followed by backcrossing of the hybrid with O. novo-ulmi. The presence of an O. ulmi-like cu gene in MAFf8 is consistent with its CU deficiency, since the O. ulmicu gene is known to be poorly expressed and O. ulmi isolates secrete little or no CU in culture.

Origin of a new Phytophthora pathogen through interspecific hybridization.

Plant disease epidemics resulting from introductions of exotic fungal plant pathogens are a well known phenomenon. An associated risk-that accelerated pathogen evolution may be occurring as a consequence of genetic exchange between introduced, or introduced and resident, fungal pathogens-is largely unrecognized. This is, in part, because examples of natural, interspecific hybridization in fungi are very rare. Potential evolutionary developments range from the acquisition of new host specificities to emergence of entirely new pathogen taxa. We present evidence from cytological behavior, additive nucleotide bases in repetitive internal transcribed spacer regions of the rRNA-encoding DNA (rDNA), and amplified fragment length polymorphisms of total DNA that a new, aggressive Phytophthora pathogen of alder trees in Europe comprises a range of heteroploid-interspecific hybrids involving a Phytophthora cambivora-like species and an unknown taxon similar to Phytophthora fragariae. The hybrids' marked developmental instabilities, unusual morphological variability, and evidence for recombination in their internal transcribed spacer profiles indicates that they are of recent origin and that their evolution is continuing. The likelihood of such evolutionary events may be increasing as world trade in plants intensifies. However, routine diagnostic procedures currently in use are insufficiently sensitive to allow their detection.

Multiple mitochondrial viruses in an isolate of the Dutch Elm disease fungus Ophiostoma novo-ulmi.

The nucleotide sequences of three mitochondrial virus double-stranded (ds) RNAs, RNA-4 (2599 nucleotides), RNA-5 (2474 nucleotides), and RNA-6 (2343 nucleotides), in a diseased isolate Log1/3-8d2 (Ld) of the Dutch elm disease fungus Ophiostoma novo-ulmi have been determined. All these RNAs are A-U-rich (71-73% A + U residues). Using the fungal mitochondrial genetic code in which UGA codes for tryptophan, the positive-strand of each of RNAs 4, 5, and 6 contains a single open reading frame (ORF) with the potential to encode a protein of 783, 729, and 695 amino acids, respectively, all of which contain conserved motifs characteristic of RNA-dependent RNA polymerases (RdRps). Sequence comparisons showed that these RNAs are related to each other and to a previously characterized RNA, RNA-3a, from the same O. novo-ulmi isolate, especially within the RdRp-like motifs. However, the overall RNA nucleotide and RdRp amino acid sequence identities were relatively low (43-55% and 20-32%, respectively). The 5'- and 3'-terminal sequences of these RNAs are different, but they can all be folded into potentially stable stem-loop structures. Those of RNA-4 and RNA-6 have inverted complementarity, potentially forming panhandle structures. Their molecular and biological properties indicate that RNAs 3a, 4, 5, and 6 are the genomes of four different viruses, which replicate independently in the same cell. These four viruses are also related to a mitochondrial RNA virus from another fungus, Cryphonectria parasitica, recently designated the type species of the Mitovirus genus of the Narnaviridae family, and to a virus from the fungus Rhizoctonia solani. It is proposed that the four O. novo-ulmi mitochondrial viruses are assigned to the Mitovirus genus and designated O. novo-ulmi mitovirus (OnuMV) 3a-Ld, 4-Ld, 5-Ld, and 6-Ld, respectively. Northern blot analysis indicated that O. novo-ulmi Ld nucleic acid extracts contain more single-stranded (ss, positive-stranded) RNA than dsRNA for all three newly described mitoviruses. O. novo-ulmi RNA-7, previously believed to be a satellite-like RNA, is shown to be a defective RNA, derived from OnuMV4-Ld RNA by multiple internal deletions. OnuMV4-Ld is therefore the helper virus for the replication of both RNA-7 and another defective RNA, RNA-10. Sequence comparisons indicate that RNA-10 could be derived from RNA-7, as previously suggested, or derived directly from RNA-4.

Evolutionary relationships among putative RNA-dependent RNA polymerases encoded by a mitochondrial virus-like RNA in the Dutch elm disease fungus, Ophiostoma novo-ulmi, by other viruses and virus-like RNAs and by the Arabidopsis mitochondrial genome.

The nucleotide sequence (2617 nucleotides) of virus-like double-stranded (ds) RNA 3a in a diseased isolate, Log1/3-8d2 (Ld), of the ascomycete fungus Ophiostoma novo-ulmi has been determined. One strand of the dsRNA contains an open reading frame (ORF) with the potential to encode a protein of 718 amino acids, and the complementary strand contains two smaller ORFs with the potential to encode proteins of 178 and 182 amino acids, respectively. The large ORF contains 12 UGA codons which code for tryptophan in ascomycete mitochondria and has a codon bias typical of mitochondrial genes, consistent with the localization of Ld dsRNAs within the mitochondria. The amino acid sequence contains motifs characteristic of RNA-dependent RNA polymerases (RdRps). This putative RdRp was shown to be related to putative RdRps of mitochondrial dsRNAs of another ascomycete and a basidiomycete fungus and also to a putative RdRp encoded by the mitochondrial genome of Arabidopsis thaliana. In multiple sequence alignments, the fungal mitochondrial dsRNA-encoded RdRp-like proteins formed a cluster, ancestrally related to the RdRps of the yeast 20S and 23S RNA replicons and of the positive-stranded RNA bacteriophages of the Leviviridae family, but distinct from RdRps of other families and genera of fungal RNA viruses and related plant and animal RNA viruses. Northern blot analysis with RNA 3a strand-specific probes indicated that nucleic acid extracts of Ld contain more single-stranded (positive-stranded) RNA than dsRNA, consistent with an evolutionary relationship between RNA 3a and positive-stranded RNA phages.

Novel structures of two virus-like RNA elements from a diseased isolate of the Dutch elm disease fungus, Ophiostoma novo-ulmi.

The nucleotide sequences of 2 of the 10 mitochondrial double-stranded (ds) RNA segments in a diseased isolate, Log 1/3-8d2 (Ld), of Ophiostoma novo-ulmi, RNA-7 (1057 nucleotides) and RNA-10 (317-330 nucleotides), have been determined. Both RNAs are A-U-rich, but in Southern and Northern blots, no hybridization with mitochondrial DNA or RNA could be detected. Only very short open reading frames were found in both RNAs. As most of its sequence is unrelated to any of the other Ld dsRNAs, RNA-7 may be regarded as a satellite RNA. Northern blotting detected a full-length single-stranded (ss) form of RNA-7 in nucleic acid extracts from Ld. The 5'- and 3'-terminal 39 nucleotides of ssRNA-7 are imperfect inverted complementary repeats of each other, which could cause ssRNA-7 to form a panhandle structure. In addition, the 5'-terminal nucleotides 1-28 and 3'-terminal nucleotides 1032-1057 of ssRNA-7 each contained inverted complementary sequences, allowing the possibility for each terminus to form separate stem-loop structures. The combination of these two structural features has not been found previously in any dsRNA or ssRNA virus. RNA-10 was shown to have an unusual structure, consisting of a mosaic of sequences derived from regions of the 5'- and 3'-termini, or just the 5'-terminus, of RNA-7, RNA-10 has a high degree of inverted complementarity, with the potential to be folded into a very stable hairpin structure. A model for the formation of RNA-10 is presented, involving replicase-driven strand switching between (-)-strand and (+)-strand templates during RNA synthesis, followed by utilization of the nascent strand as a primer and template to form a snap-back RNA.

The influence of temperature and light on defoliation levels of elm by dutch elm disease.

ABSTRACT The amount of defoliation of elm (Ulmus procera) caused by three Ophiostoma novoulmi Eurasian race isolates over 14 seasons of field trials was found to be strongly correlated with mean air temperature and mean number of sunshine hours over the 12-week period from inoculation to assessment, and with tree age. The coefficient of determination for the regression of percent defoliation on the environmental and tree factors was 0.76, P < 0.001 (33 df). Levels of defoliation were greatest when mean air temperatures exceeded 17 degrees C with moderate light (5 to 7 h of sunshine), and lowest under conditions of either high light (>7.5 h of sunshine) at all air temperatures or low light (<4.5 h of sunshine) and air temperatures of less than 15.5 degrees C. The model varied in its intercept for the three isolates, reflecting their different levels of aggressiveness. The role of environmental factors in the development of Dutch elm disease symptoms and the implications for elm resistance breeding are discussed.

Nucleotide-sequence analysis indicates that a DNA plasmid in a diseased isolate of Ophiostoma novo-ulmi is derived by recombination between two long repeat sequences in the mitochondrial large subunit ribosomal RNA gene.

The nucleotide sequence of a mitochondrial plasmid (2234 bp) in a diseased isolate of Ophiostoma novo-ulmi, and sequences of the mitochondrial DNA that overlap and flank the plasmid end-points, have been determined. The plasmid was shown to be derived from the O. novo-ulmi mitochondrial large subunit ribosomal RNA gene and contained most of intron 1, the whole of exon 2, and probably the first part of intron 2. Within intron 1 there is an open reading frame with the potential to encode a 323 amino-acid polypeptide which contained dodecapeptide sequences typical of RNA maturases and DNA endonucleases. The endpoints of the plasmid in the mtDNA were located within two 90-bp direct imperfect repeat sequences, one of which comprised the last 7 bp of exon 1 and the first 83 bp of intron 1 whilst the other comprised the last 7 bp of exon 2 and the first 83 bp of intron 2. It is proposed that the Ld plasmid was generated by intramolecular recombination between these two repeats with the crossover point probably within the last 15 bp.

De-novo generation of mitochondrial DNA plasmids following cytoplasmic transmission of a degenerative disease in Ophiostoma novo-ulmi.

A mitochondrial DNA plasmid was detected in an isolate of Ophiostoma novo-ulmi with a degenerative disease. The DNA plasmid was shown to be derived from the mitochondrial DNA and to map to a region corresponding to the large ribosomal RNA coding region. The DNA plasmid was not transmitted into sexual (ascospore) progeny, irrespective of whether the diseased isolate acted as the female or male parent. Transmission of the disease to healthy, plasmid-free, "recipient" isolates by hyphal anastomosis was not accompanied by transfer of mitochondrial DNA or DNA plasmid from the diseased "donor" isolate, but resulted in de-novo generation of different plasmids, derived from the recipient's mitochondrial DNA.

Variation in a natural population of Schizophyllum commune. Variation within the extreme isolates for growth rate.

Two extreme dikaryotic idolates chosen from a large sample of a localised population of Schizophyllum commune exhibited a considerable amount of genetical variation for growth rate at the near ambient temperature of 20 degrees C and at the higher temperature of 30 degrees C. The potential variation within these extreme isolates were greater than the variation observed in the whole sample. Regression analysis of the variation in growth rate of the dikaryotic progeny of the extreme isolates on that of their component monokaryons showed that the nature of gene action was not the same in these two stages of the life cycle. The simple additive-dominance model of gene action was adequate to explain the variation in growth rate in both of the extreme isolates at both of the temperatures. The small deviations from this model could be accounted for by unequal gene frequencies due to small sample size although a low incidence of non-allelic interactions could not be rule out. Directional dominance for growth rate was detected in both isolates at the more normal temperature and it was opposing in direction in the two isolates. In the slow growing isolate the dominance was for faster growth and in the fast growing isolate it was for slower growth. This is expected for a character which displays overall ambi-directional dominance if isolates with more extreme growth rates than those recovered in the population sample are eliminated by stabilising selection. The dominance is temperature dependent being ambi-directional in both isolates at the higher temperature. Environmental heterogeneity, the buffering effects of directional dominance and genotype-environment interactions and opposing selective forces operating on the monokaryotic and dikaryotic stages of the life cycle are possible contributory factors to the considerable free and potential variability displayed in this small, localised population.