Distribution and expression of elicitin genes in the interspecific hybrid oomycete Phytophthora alni

Phytophthora alni subsp. alni, P. alni subsp. multiformis, and P. alni subsp. uniformis are responsible for alder disease in Europe. Class I and II elicitin gene patterns of P. alni subsp. alni, P. alni subsp. multiformis, P. alni subsp. uniformis, and the phylogenetically close species P. cambivora and P. fragariae were studied through mRNA sequencing and 3' untranslated region (3'UTR)-specific PCRs and sequencing. The occurrence of multiple 3'UTR sequences in association with identical elicitin-encoding sequences in P. alni subsp. alni indicated duplication/recombination events. The mRNA pattern displayed by P. alni subsp. alni demonstrated that elicitin genes from all the parental genomes are actually expressed in this allopolyploid taxon. The complementary elicitin patterns resolved confirmed the possible involvement of P. alni subsp. multiformis and P. alni subsp. uniformis in the genesis of the hybrid species P. alni subsp. alni. The occurrence of multiple and common elicitin gene sequences throughout P. cambivora, P. fragariae, and P. alni sensu lato, not observed in other Phytophthora species, suggests that duplication of these genes occurred before the radiation of these species.     

Involvement of Phytophthora species in the decline of beech Fagus sylvatica in Wallonia (Belgium)

During the last decade, typical symptoms of Phytophthora diseases were observed in beech stands of several European countries. The main symptoms were the presence of bleeding cankers on the stem, a low crown density as well as the yellowing of foliage and the small size of leaves. Several species of Phytophthora, such as Phytophthora citricola, P. cambivora and P. cactorum, were reported as the causal agents. In order to evaluate the implication of the different Phytophthora species in beech decline in the southern part of Belgium (Wallonia), a monitoring was undertaken with the help of managers of public and private forests. Phytophthora strains isolated from beech of different stands as well as from soil were characterized through morphological and molecular analyses (PCR-RFLP of ITS). All the isolated strains were identified as P. cambivora, except for one strain whose identification is ongoing. Molecular analysis was also directly applied to necrosed tissues of bleeding beeches and enabled the detection of additional cases. All positive cases exhibited a profile characteristic of the species P. cambivora, except for one of the sampled trees showing a different Phytophthora profile also corresponding to the unidentified isolated strain. Identification of the Phytophthora species linked to this different RFLP profile is also ongoing. Both complementation types (A1 and A2) of P. cambivora were identified, sometimes in the same sampling site. Ornamented oogonia characteristic of this species were produced by pairing A1 and A2 strains isolated from the same site.     

Specificity of cytoplasmic and cell-wall antigens from four species of Phytophthora.

Cytoplasmic and cell-wall antigens and antisera were prepared from four Phytophthora species, and cell-wall antigens were prepared from two Pythium species. Immunodiffusion of the Pythium and Phytophthora cell-wall antigens showed that the two Pythium species did not cross-react with the Phytophthora cell-wall antisera. Immunodiffusion analysis of both cell-wall and cytoplasmic antigens of Phytophthora revealed some degree of specificity between species but not between A1 and A2 mating types in Phytophthora cinnamomi. Species specificity was improved by using indirect fluorescent antibody techniques and by the use of cross-absorbed sera. Agglutination and quantitative precipitation techniques did not significantly improve specificity. It was possible to distinguish serologically between Phytophthora cinnamomi and Phytophthora cambivora and the Phytophthora cryptogea-Phytophthora drechsleri group. The absence of consistent serological variation between P. cryptogea and P dreschsleri is consistent with the suggestion (Bumbieris, 1974) that P. cryptogea and P. drechsleri should be considered as one species.     

Relationship between occurrence of Phytophthora cambivora on plants in hardy ornamental nursery stocks and detection of the pathogen from water ponds

Phytophthora cambivora was isolated from rooted bases and roots of Castanea sativa, Chamaecyparis lawsoniana and Cotoneaster spp. as well as from water ponds Located in HNS. In the laboratory trials the species colonised leaf blades and stem parts of 3 tested plant species. Differentiated reaction of tested plant species on P. cambivora was observed.     

Detection and identification of Phytophthora alni

In 2004 Brasier et al. described new species--Phytophthora alni, which was especially aeggressive to alder. Now, this Phytophthora disease of alder is widely distributed in Europe as well as in Poland. In this research note we report on identification and detection of P. alni from water and soil samples using PCR method with species-specific primers. Dilution series of P. alni zoospore were used to test the potential sensitivity of the PCR detection methods. Zoospores of P. alni were produced by flooding of 1-week-old Frozen Pea Medium (FPM) cultures in Petri dishes with 30 ml distilled water. The dishes were incubated at 20 degrees C. After 5 days, sporangial production was checked using a binocular microscope and plates were placed at 4 degrees C for 1 h to enhance zoospore release. Zoospores were counted under the microscope using Burker's cabin. A dilution series of zoospores ranging from 5 to 5000 per 200 microl was prepared in autoclaved distilled water and in 1 g samples of autoclaved soil. DNA was extracted from artificially infected water and soil, and purified using the CleanUp Kit (A&A Biotechnology). Zoospores of P. alni in the water were detected by PCR in 5 x 10(3), 5 x 10(2), 5 x 10(1) concentrations. In case of detecting spores in the artificially infected soil it succeeded only for two highest concentrations, i.e. 5 x 10(3), 5 x 10(2) and only when the DNA was additionally purified.     

Phytophthora polonica, a new species isolated from declining Alnus glutinosa stands in Poland

In a survey of Phytophthora associated with alder decline in Poland, several isolates of a homothallic Phytophthora sp., which could not be assigned to other taxa including Phytophthora alni subspecies, were consistently recovered from rhizosphere soil samples. Their morphology and pathogenicity, as well as sequence data for three nuclear regions (internal transcribed spacer rDNA, elongation factor-1alpha and beta-tubulin) and a coding mitochondrial DNA region (nadh1), were examined. The new Phytophthora species is characterized by the moderate to slow growth rate of its colony in carrot agar at 20 degrees C, high optimal (c. 30 degrees C) and maximum (c. 38 degrees C) growth temperatures, formation of catenulate, often lateral, hyphal swellings, large chlamydospores in agar media and in soil extract, persistent, ovoid to ellipsoid nonpapillate sporangia and large oogonia with paragynous and sometimes amphigynous antheridia. Phytophthora polonica was slightly pathogenic to alder twigs and not pathogenic to trunks of several tree species. In a phylogenetic analysis using either Bayesian inference or maximum likelihood methods, P. polonica falls in clade 8 'sensu Kroon et al. (2004)' of Phytophthora.     

Safflower Seedling a Selective Host to Discriminate Phytophthora melonis from Phytophthora drechsleri

Safflower seedlings were used to discriminate two morphologically similar species of Phytophthora namely P. melonis and P. drechsleri . All isolates of P. melonis from different sources could not infect safflower seedlings under high inoculum potential whereas all isolates of P. drechsleri from various hosts attacked safflower within a short period. Of 31 authentic Phytophthora species inoculated to safflower seedlings only seven species including Phytophthora asparagi , Phytophthora cactorum , Phytophthora cryptogea , Phytophthora drechsleri , Phytophthora erythroseptica , Phytophthora palmivora and Phytophthora quercina caused hypocotyls infection. All cucurbit isolates of Phytophthora from different parts of Iran and one from China could not infect safflower seedlings and were identified as P. melonis which had been confirmed previously by molecular analysis.     

Touchdown nested multiplex PCR detection of Phytophthora cinnamomi and P. cambivora from French and English chestnut grove soils

Soil borne Phytophthora cinnamomi and Phytophthora cambivora are considered the most pathogenic species associated with chestnut (Castanea sativa) decline in Europe. Mapping their incidence and distribution from nursery and plantation soils may offer valuable information for limiting spread. As conventional biological baiting and taxonomic confirmation is generally time consuming, labour, logistically and space intensive, we have focused on the development of a specific touchdown nested multiplex Polymerase Chain Reaction (PCR) approach for the simultaneous detection of both species direct from soil. Pre-existing and novel primers, based on Internal Transcribed Spacer (ITS) sequences, have been evaluated for their specificity and use in a multiplex capacity in various combinations. Coupled to this we have modified a mechanical lysis procedure for DNA extraction from up to 10 g of chestnut under storey soils (ranging from 0.5 to 25 μg DNA g(-1)fresh soil). Using serial dilutions and/or polyvinylpolypyrrolidone chromatography purification, both species have been successfully detected, in artificially and naturally infected soils. Levels of assay detection are comparable to other Phytophthora species where PCR based diagnostic systems have been reported. A qualitative evaluation of this approach against conventional baiting is presented.     

Single-strand-conformation polymorphism of ribosomal DNA for rapid species differentiation in genus Phytophthora

Single-strand-conformation polymorphism (SSCP) of ribosomal DNA of 29 species (282 isolates) of Phytophthora was characterized in this study. Phytophthora boehmeriae, Phytophthora botryosa, Phytophthora cactorum, Phytophthora cambivora, Phytophthora capsici, Phytophthora cinnamomi, Phytophthora colocasiae, Phytophthora fragariae, Phytophthora heveae, Phytophthora hibernalis, Phytophthora ilicis, Phytophthora infestans, Phytophthora katsurae, Phytophthora lateralis, Phytophthora meadii, Phytophthora medicaginis, Phytophthora megakarya, Phytophthora nicotianae, Phytophthora palmivora, Phytophthora phaseoli, Phytophthora pseudotsugae, Phytophthora sojae, Phytophthora syringae, and Phytophthora tropicalis each showed a unique SSCP pattern. Phytophthora citricola, Phytophthora citrophthora, Phytophthora cryptogea, Phytophthora drechsleri, and Phytophthora megasperma each had more than one distinct pattern. A single-stranded DNA ladder also was developed, which facilitates comparison of SSCP patterns within and between gels. With a single DNA fingerprint, 277 isolates of Phytophthora recovered from irrigation water and plant tissues in Virginia were all correctly identified into eight species at substantially reduced time, labor, and cost. The SSCP analysis presented in this work will aid in studies on taxonomy, genetics, and ecology of the genus Phytophthora.     

The plant pathogen Phytophthora andina emerged via hybridization of an unknown Phytophthora species and the Irish potato famine pathogen, P. infestans

Emerging plant pathogens have largely been a consequence of the movement of pathogens to new geographic regions. Another documented mechanism for the emergence of plant pathogens is hybridization between individuals of different species or subspecies, which may allow rapid evolution and adaptation to new hosts or environments. Hybrid plant pathogens have traditionally been difficult to detect or confirm, but the increasing ease of cloning and sequencing PCR products now makes the identification of species that consistently have genes or alleles with phylogenetically divergent origins relatively straightforward. We investigated the genetic origin of Phytophthora andina, an increasingly common pathogen of Andean crops Solanum betaceum, S. muricatum, S. quitoense, and several wild Solanum spp. It has been hypothesized that P. andina is a hybrid between the potato late blight pathogen P. infestans and another Phytophthora species. We tested this hypothesis by cloning four nuclear loci to obtain haplotypes and using these loci to infer the phylogenetic relationships of P. andina to P. infestans and other related species. Sequencing of cloned PCR products in every case revealed two distinct haplotypes for each locus in P. andina, such that each isolate had one allele derived from a P. infestans parent and a second divergent allele derived from an unknown species that is closely related but distinct from P. infestans, P. mirabilis, and P. ipomoeae. To the best of our knowledge, the unknown parent has not yet been collected. We also observed sequence polymorphism among P. andina isolates at three of the four loci, many of which segregate between previously described P. andina clonal lineages. These results provide strong support that P. andina emerged via hybridization between P. infestans and another unknown Phytophthora species also belonging to Phytophthora clade 1c.     

New antifungal activity of penicillic acid against Phytophthora species

Penicillic acid was isolated from a culture filtrate of Aspergillus sclerotiorum. It had a high in vitro antifungal activity against Phytophthora spp., which has not been previously reported. MICs of penicillic acid were from 1 to 25 microg ml(-1) against Phytophthora spp. Penicillic acid induced abnormal branch formation, apical branching, and swelling in P. capsici, in P. cactorum mycelia contained irregular branching and small spherical swelling at apices, in P. cambivora there was irregular branching and swelling, and in P. drechsleri there was irregular multiple spherical swelling at or near hyphal apices.     

云南三种同域分布的棘蛙（蛙科Ranidae：无尾目Anura）的核型和银带研究

本文比较分析了云南景东地区三种同域分布棘蛙的核型和Ａｇ－ＮＯＲｓ。花棘蛙２ｎ＝２６（１６Ｍ＋ １０ＳＭ）,ＮＦ＝５２,次缢痕和Ａｇ－ＮＯＲｓ位于１ｐｉｎｔｅｒ,Ｎｏｓ．２－４,８,９等为ＳＭ。棘肛蛙２ｎ＝４０（１６Ｍ＋ ２０ＳＭ＋２ＳＴ＋２Ｔ）,ＮＦ＝７８,Ｎｏｓ．５－９,１１－１３,１５,１７等１０对为ＳＭ,Ｎｏ．３为ＳＴ,Ｎｏ．１８为Ｔ,其余 均为Ｍ,Ａｇ－ＮＯＲｓ位于１１Ｐ。二种的Ａｇ－ＮＯＲｓ都有异形现象。双团棘胸蛙２ｎ＝６４Ｔ,次缢痕和Ａｇ－ ＮＯＲＳ在４ｑｐｅｒ。都未发现异形性染色体。最后,对棘蛙属的核型演化机制和物种形成方式作了讨论。     

中国木层孔菌属三个新记录种

报道了木层孔菌属Phellinus 3个中国新记录种。赤杨木层孔菌Phellinus alni典型特征为菌盖具有较宽的同心环带和清晰的密集环纹菌核;黑木层孔菌P. nigricans具有较大的担孢子;东方木层孔菌P. orienticus具有平伏的子实体,担孢子相对较小。对这3个种进行了详细的描述和显微结构绘图。     

The repertoire of transfer RNA genes is tuned to codon usage bias in the genomes of Phytophthora sojae and Phytophthora ramorum

In all, 238 and 155 transfer (t)RNA genes were predicted from the genomes of Phytophthora sojae and P. ramorum, respectively. After omitting pseudogenes and undetermined types of tRNA genes, there remained 208 P. sojae tRNA genes and 140 P. ramorum tRNA genes. There were 45 types of tRNA genes, with distinct anticodons, in each species. Fourteen common anticodon types of tRNAs are missing altogether from the genome in the two species; however, these appear to be compensated by wobbling of other tRNA anticodons in a manner which is tied to the codon bias in Phytophthora genes. The most abundant tRNA class was arginine in both P. sojae and P. ramorum. A codon usage table was generated for these two organisms from a total of 9,803,525 codons in P. sojae and 7,496,598 codons in P. ramorum. The most abundant codon type detected from the codon usage tables was GAG (encoding glutamic acid), whereas the most numerous tRNA gene had a methionine anticodon (CAT). The correlation between the frequencies of tRNA genes and the codon frequencies in protein-coding genes was very low (0.12 in P. sojae and 0.19 in P. ramorum); however, the correlation between amino acid tRNA gene frequency and the corresponding amino acid codon frequency in P. sojae and P. ramorum was substantially higher (0.53 in P. sojae and 0.77 in P. ramorum). The codon usage frequencies of P. sojae and P ramorum were very strongly correlated (0.99), as were tRNA gene frequencies (0.77). Approximately 60% of orthologous tRNA gene pairs in P sojae and P. ramorum are located in regions that have conserved synteny in the two species.     

A revision of the Paeonia suffruticosa complex (Paeoniaceae)

The taxonomical concept of the Paeonia suffruticosa complex i.e. Sect. Moutan Subsect. Vaginatae, has changed greatly since 1990. Six species and four subspecies have been described as new and two subspecies raised to specific level. Five species and two subspecies are recognized in the present revision, viz. P. suffruticosa subsp. suffruticosa and subsp. yinpingmudan, P. jishanensis, P. qiui, P. ostii, P. rockii subsp. rockii and subsp. taibaishanica. P. yananensis, P. ridleyi, P. spontanea, P. moutan subsp. atava, P. suffruticosa subsp. atava, P. rockii subsp. linyanshanii and P. ostii var. lishizhenii are treated as synonyms. P. papaveracea and P. baokangensis are proposed to be interspecific hybrids. A key to the recognized species and subspecies is provided. Biological features of the species are described and their distributions are mapped. The relationships between species are inferred and the origins of commonly cultivated tree peonies ( P. suffruticosa and P. ostii ) are discussed.     

Mitochondrial protein import motor: differential role of Tim44 in the recruitment of Pam17 and J-complex to the presequence translocase

The presequence translocase of the mitochondrial inner membrane (TIM23 complex) mediates the import of preproteins with amino-terminal presequences. To drive matrix translocation the TIM23 complex recruits the presequence translocase-associated motor (PAM) with the matrix heat shock protein 70 (mtHsp70) as central subunit. Activity and localization of mtHsp70 are regulated by four membrane-associated cochaperones: the adaptor protein Tim44, the stimulatory J-complex Pam18/Pam16, and Pam17. It has been proposed that Tim44 serves as molecular platform to localize mtHsp70 and the J-complex at the TIM23 complex, but it is unknown how Pam17 interacts with the translocase. We generated conditional tim44 yeast mutants and selected a mutant allele, which differentially affects the association of PAM modules with TIM23. In tim44-804 mitochondria, the interaction of the J-complex with the TIM23 complex is impaired, whereas unexpectedly the binding of Pam17 is increased. Pam17 interacts with the channel protein Tim23, revealing a new interaction site between TIM23 and PAM. Thus, the motor PAM is composed of functional modules that bind to different sites of the translocase. We suggest that Tim44 is not simply a scaffold for binding of motor subunits but plays a differential role in the recruitment of PAM modules to the inner membrane translocase.     

Investigations on Net CO2 Assimilation, Transpiration and Root Growth of Fagus sylvatica Infested with Four Different Phytophthora Species

Abstract: In this contribution, we compare the influence of four different Phytophthora species on root development, net CO₂ assimilation and transpiration of beech seedlings and saplings. Some few days after inoculation, photosynthesis and transpiration of seedlings infected with either P. citricola or P. cambivora were strongly reduced. In parallel, about 60 % of their root systems was destroyed compared to control plants. Three weeks after infection, all seedlings were dead, showing severe wilt symptoms on leaves. Remarkably, P. syringae and P. undulata infected seedlings and older beech plants did not differ from controls regarding photosynthesis and transpiration, although the root systems were damaged. However, a significant influence on net CO₂ assimilation and transpiration of P. citricola infected beech saplings was visible after bud break in the following year. Some days before plants started to wilt, photosynthesis and transpiration were reduced to almost zero. Water use efficiency data (WUE) clearly indicated that infected plants suffered from severe drought.     

刘氏棘蛙的分类修订

采集了云南宁蒗和四川盐源县交界泸沽湖的刘氏棘蛙Paa liui地模标本进行形态学研究,并与形态上相似的双团棘胸蛙P. yunnanensis、多疣棘蛙P.verrucospinosa、察隅棘蛙P. chayuensis开展分子生物学研究,结果表明:刘氏棘蛙作为一个独立物种不存在,分布于泸沽湖的刘氏棘蛙为双团棘胸蛙的一个地理居群,应作为双团棘胸蛙的同物异名进行归并.