Testing variability in pathogenicity ofPhytophthora cactorum, P. citrophthora andP. syringae to apple, pear, peach, cherry and plum rootstocks

The relative virulence ofPhytophthora cactorum andP. syringae originating from almond trees, and ofP. citrophthora originating from citrus, to apple, pear, peach, cherry and plum rootstocks, was studiedin vivo andin vitro. Results of the different experiments were in good agreement. All testedPhytophthora isolates showed little virulence to pear rootstocks-causing only minor crown rot symptoms - and no virulence at all to apple rootstocks. In contrast, they were highly virulent to stone fruit rootstocks, causing crown rot disease. The non-pathogenicity of these isolates to pome rootstocks could be interpreted as strict host specificity.     

Effects of inoculum density,plant age and temperature on disease severity caused by pythiaceous fungi on several plants

Alfalfa, maize, sorghum and sugarbeet plants were inoculated with zoospores ofPhytophthora andPythium species in order to assess the effects of inoculum density, plant age and temperature on disease severity. Seedlings were grown axenically in test tubes and inoculated with zoospore suspensions. Disease severity was assessed by measuring the root growth and discoloration of treated and control seedlings. The incremental root length of all plants decreased and root discoloration increased as inoculum concentration of the pathogen increased. Changes were more intensive among low levels of zoospore concentrations and no significant differences in disease severity were found for inoculum densities higher than 10⁴ zoospores ml^-1. Disease severity was negatively related to plant age. Disease development on sugarbeet seedlings infected withPythium andPhytophthora species was affected by temperature, but the pattern of response was determined by the pathogen’s temperature preferences. The incremental root length decreased as temperature increased up to 25°C. The effect ofPythium dissimile andPhytophthora cactorum on root length was significantly lower at 35°C than at 25°C, whereasPythium aphanidermatum andPhytophthora nicotianae caused significant damage to roots even at 35°C. http://www.phytoparasitica.org posting Dec. 3, 2001.     

Occurrence and distribution of <Emphasis Type="Italic">Phytophthora</Emphasis> species in European chestnut stands,and their association with Ink Disease and crown decline

The Phytophthora complex associated with Castanea sativa Mill. was investigated in five European countries in 35 regions and with respect to various domestication levels. Annual precipitation and length of drought season were the main parameters that regulated the presence of Phytophthora species in the chestnut stands. Seven species of Phytophthora were detected; three of these, P. megasperma, P. cryptogea and P. syringae had not been previously reported on sweet chestnut. P. cinnamomi. P. cambivora and P. citricola were most frequently isolated. P. cinnamomi and P. cambivora were the species significantly associated with declining trees with symptoms of Ink Disease. P. cinnamomirequired distinct ecological conditions compared to the other species. P. cinnamomi was never detected in sites characterized by minimum temperatures below 1.4 °C, maximum temperature above 28 °C, or soil pH below 5.4. The results obtained provide useful information for modeling the probability of Ink Disease, crown decline and associated Phytophthora species in chestnut groves in global climatic change scenarios.     

Hot water treatment of Japanese pear trees is effective against white root rot caused by <Emphasis Type="Italic">Rosellinia necatrix</Emphasis> Prillieux

Hot water was dripped into the rhizosphere of Japanese pear trees (Pyrus serotina Rehd. grafted on P. betulifolia Bunge.) infested with the white root rot fungus Rosellinia necatrix Prillieux, to destroy the fungus. Isolates of R. necatrix from diseased roots of Japanese pear were vulnerable to water at temperatures above 35°C, and the fungus was eradicated from the colonized substrate when water at 35°C was provided for 3 days. The time required to eradicate R. necatrix decreased exponentially with increasing temperature. Japanese pear trees tolerated a temperature of 45°C without reduction in vigor. Field experiments demonstrated the practical use of hot water drip irrigation (HWD). HWD at 50°C completely destroyed white root rot mycelia on diseased roots, and many rootlets grew after the treatment. HWD at this temperature caused no injury to the trees. HWD of diseased orchard trees was assessed in Takamori and Iida in southern Nagano, Japan. The fungus recurred in two of four trees 28 months after treatment in Takamori and in two of ten trees 16 months after treatment in Iida. The new mycelia emerged on thick roots deep within the soil. Although there is a possibility of recurrence, HWD treatment is a practical control measure for white root rot.     

<Emphasis Type="Italic">Pythium</Emphasis> and <Emphasis Type="Italic">Phytophthora</Emphasis> species associated with root and stem rots of kalanchoe

Pythium and Phytophthora species were isolated from kalanchoe plants with root and stem rots. Phytophthora isolates were identified as Phytophthora nicotianae on the basis of morphological characteristics and restriction fragment length polymorphism (RFLP) analysis of the rDNA-internal transcribed spacer regions. Similarly, the Pythium isolates were identified as Pythium myriotylum and Pythium helicoides. In pathogenicity tests, isolates of the three species caused root and stem rots. Disease severity caused by the Pythium spp. and Ph. nicotianae was the greatest at 35°–40°C and 30°–40°C, respectively. Ph. nicotianae induced stem rot at two different relative humidities (60% and >95%) at 30°C. P. myriotylum and P. helicoides caused root and stem rots at high humidity (>95%), but only root rot at low humidity (60%).     

Stone Fruit Tree Apoplexy Caused by Phytophthora Collar Rot

Stone fruit tree apoplexy in Greece is often caused by Phytophthora spp. Two types of Phytophthora apoplexy of stone fruit tree can be distinguished, one occurring during the hot summer period and usually caused by P. cactorum (Leb. et Cohn) Schröt. or P. citrophthora (R. et E. Smith) Leonian, and a second occurring in winter or early spring, and caused by P. syringae (Kleb.) Kleb. or sometimes P. megasperma Drechsler. Inoculation tests showed P. syringae to be highly pathogenic to apricot and almond, whereas certain clonal rootstock material showed resistance. Measures of control against Phytophthora attacks are given.     

Apple proliferation resistance of <Emphasis Type="Italic">Malus sieboldii</Emphasis>-based rootstocks in comparison to rootstocks derived from other <Emphasis Type="Italic">Malus</Emphasis> species

In three trials carried out over a period of 24 years, open-pollinated seedlings of Malus sieboldii and M. sargentii and 22 apomictic rootstock selections with either M. sieboldii, M. sargentii or M. hupehensis in their parentage were examined for apple proliferation (AP) resistance in comparison to clonal M. x domestica-based rootstocks M 9, M 11, M 13, stocks of the B (Budagovski) and the Polish P series and M. robusta seedlings. Following experimental inoculation or natural infection the Golden Delicious-grafted trees on most of the M. sieboldii-derived progenies showed a high level of AP resistance expressed by low cumulative disease indices, a high percentage of non or little affected trees, low incidence of the small fruit symptom and non or little effect on vigour. Trees on M 9 and M 11, B 118 and M. robusta seedlings were moderately susceptible while trees on progenies with M. sargentii and M. hupehensis parentage, rootstocks of the P series, B 9, B 490 and M 13 proved highly susceptible. The screening also showed that rootstocks with M. sieboldii and M. sargentii parentage are often highly susceptible to latent apple viruses. Trees on most of the M. sieboldii-based progenies were more vigorous than trees on standard stock M 9, whereas the vigour of some progenies from selections with M. sargentii parentage was in the range of M 9 or even lower. Productivity was often correlated with the vigour.     

Effects of six herbicides on the growth of Phytophthora cactorum and a bacterial antagonist

Six herbicides, incorporated in corn meal agar, were tested for their effects on the growth of Phytophthora cactorum, the growth of a bacterial antagonist B8 (Erwinia sp.), and the antagonistic effect of B8 on P. cactorum Oryzalin and glyphosate significantly reduced the growth of P. cactorum. Neither the growth of B8, nor its antagonistic effect on P. cactorum, was affected by any of the herbicides tested. This suggests that herbicides may not be a limiting factor on the development of bacterial antagonists on a biological control method for the control of crown rot of apple trees.     

Phytophthora crown rot of almonds in Australia 1

Phytophthora crown rot, caused primarily by Phytophthora cambivora, has caused considerable losses in almond orchards in the Willunga and northern plains areas approximately 50 km north and south of Adelaide (AU), respectively. Other Phytophthora species including P. citrophthora, P. cryptogea, P. megasperma and P. syringae have also been associated with diseased trees. Tree losses have been associated with P. cactorum isolated from trunk cankers on scaffold limbs on trees in the Riverland almond-growing area approximately 350 km north-east of Adelaide. The Al mating type of P. cambivora was the most pathogenic to almond seedlings. Cvs Mission and Chellaston, which are commonly used as rootstocks, were highly susceptible to crown rot whereas peach cv. Nemaguard was resistant. An excised twig assay has been developed to screen micropropagated shoots for resistance to P. cambivora. Metalaxyl applied at 5 g active substance per tree in spring, winter and autumn in a shallow trench close to the trunk prevented the development of trunk cankers. Foliar sprays of phosphonate (H₃PO₃) at 2 g active substance per litre in autumn and spring also inhibited canker development.     

Biocontrol of Phytophthora Root Rot of Angelica Trees by Enterobacter cloacae and Serratia ficaria Strains

Bacterial strains isolated from the rhizosphere of angelica trees were evaluated for their antagonistic activity against Phytophthora cactorum, a causal agent of Phytophthora root rot. Of these, three bacterial strains, designated as T-1-8, T-1-14 and T-1-23, strongly inhibited mycelial growth of P. cactorum ARE-862 in a dual-culture plate assay. Biocontrol activity of these strains was then examined by dipping root of young seedlings of angelica trees into a bacterial suspension. The incidence of Phytophthora root rot was markedly suppressed for at least 79 days in pot tests when treated seedlings were planted in naturally infested soil. The suppression was maintained through June of the next year. In addition, these strains significantly reduced the development of Phytophthora root rot up to 47 days in naturally infested field and up to 63 days (the last day of testing) in an artificially (moderately) infested field. Based on their main bacteriological properties, strain T-1-14 was identified as Enterobacter cloacae and T-1-8 and T-1-23 were identified as Serratia ficaria. Received 5 July 1999/ Accepted in revised form 25 October 1999     

Natural hybrids of resident and introduced <Emphasis Type="Italic">Phytophthora</Emphasis> species proliferating on multiple new hosts

Several atypical Phytophthora strains, isolated from a range of horticultural hosts, were tentatively identified as P. cactorum. Numerous abortive oospores were observed in these strains and isozyme analysis showed all were heterozygous for the dimeric malic enzyme (MDHP). More detailed comparisons indicated that their MDHP alleles matched those of both P. cactorum and P. hedraiandra. Cloning and sequencing of the nuclear ribosomal internal transcribed spacer (ITS) regions of the atypical P. cactorum strains demonstrated the presence of sequences characteristic for both P. cactorum and P. hedraiandra. It was concluded that the atypical strains represented hybrids between the resident P. cactorum and the apparently recently introduced P. hedraiandra. Most strains had the mitochondrially inherited cytochrome oxidase I (Cox I) gene typical of one putative parent P. hedraiandra, while one single strain had that of the other putative parent, consistent with the hybrid hypothesis. Our data also suggest that the hybrids are evolving. The hybrids have proliferated on multiple new hosts in the Netherlands.     

The effect of three irrigation practices on phytophthora crown and root rot of apple trees under field conditions

The effect of microjet, drip, and two durations of sprinkler irrigation systems on phytophthora crown and root rot of apple trees was examined under field conditions. This eight year study indicates that crown and root rot caused byPhytophthora cactorum was most severe where young MM. 106 rootstock trees were watered by microjet irrigation for 2.3 h each day. There was no difference in infection byP. cactorum when trees were irrigated either by drip or sprinkler irrigation systems. The MM.106 apple rootstock trees watered by drip irrigation for 2.6 h each day were least affected by phytophthora crown and root rot.     

Effect of Application Timing of Ergosterol Biosynthesis-inhibiting Fungicides on the Suppression of Disease and Latent Infection of Apple White Rot Caused by Botryosphaeria dothidea

To examine the duration of post-infectional activity of ergosterol biosynthesis-inhibiting fungicides (EBIs) against Botryosphaeria dothidea, the cause of apple white rot, a series of experiments was conducted for 3 years at different times during the apple-growing season using apples infected through artificial inoculation or natural infection. Disease incidence at harvest was gradually reduced as the time of chemical application was delayed until early or mid-August, regardless of the time of infection. Latent infections, detected after storing the fruit at 25°C for 4 weeks were also reduced by late season applications, but the reduction was less than that observed on the fruit at harvest. From these results, apple white rot can be effectively controlled by a single application of EBIs around mid-August, after which time, infections no longer occur in Korea. Received 22 August 2001/ Accepted in revised form 7 February 2002     

Detection and Identification of <Emphasis Type="Italic">Phytophthora</Emphasis> Species in Southern Italy by RFLP and Sequence Analysis of PCR-amplified Nuclear Ribosomal DNA

In four neighbouring regions of southern Italy, Basilicata, Campania, Apulia and Calabria, pepper and zucchini plants showing Phytophthora blight symptoms, tomato plants with either late blight or buckeye rot symptoms, plants of strawberry showing crown rot symptoms and declining clementine trees with root and fruit rot were examined for Phytophthora infections by means of polymerase chain reaction (PCR) assays, using primers directed to nuclear ribosomal DNA (rDNA) repeat sequences. All diseased plants and trees examined tested positive. The detected fungal-like organisms were differentiated and characterized on the basis of primer specificity as well as through extensive restriction fragment length polymorphism (RFLP) and sequence analysis of PCR-amplified rDNA. Phytophthora capsici was identified in diseased pepper and zucchini plants, P. infestans was identified in tomato with late blight symptoms whereas buckeye rot-affected tomatoes and diseased strawberry plants proved to be infected by P. nicotianae and P. cactorum, respectively. Declining clementine trees were infected with P. citrophthora and P. nicotianae in about the same proportion. Also, thirty-one pure culture-maintained isolates of Phytophthora which had previously been identified in southern Italy by traditional methods but were never examined molecularly, were examined by RFLP and sequence analysis of PCR-amplified nuclear rDNA. Among these, an isolate from gerbera which had previously been identified by traditional methods only at genus level, was assigned to P. tentaculata. For the remaining pure culture-maintained isolates examined, the molecular identification data obtained corresponded with those delineated by traditional methods. Most of the diseases examined were already known to occur in southern Italy but the pathogens were molecularly detected and fully characterized at nuclear rDNA repeat level only from other geographic areas, very often outside Italy. A new disease to southern Italy was the Phytophthora blight of zucchini. This is also the first report on the presence and molecular identification of P. tentaculata from Italy.     

Survival of strawberry-pathogenic fungi Fusarium oxysporum f. sp. fragariae, Phytophthora cactorum and Verticillium dahliae under anaerobic conditions

The growth and survival of three strawberry pathogens, Fusarium oxysporum f. sp. fragariae (FOF), Phytophthora cactorum, and Verticillium dahliae, were examined in anaerobic (anoxic) conditions at several temperatures (10–40 °C). The growth and survival of these fungi were suppressed by anaerobic conditions in comparison with those in cultured aerobically. Under anaerobic conditions at 22.5 °C, tested isolates of FOF and P. cactorum grew slightly, but V. dahliae did not grow. The three fungi survive for markedly shorter time in the anaerobic conditions compared with the aerobic conditions at all tested temperatures except 40 °C for FOF and P. cactorum. Moreover, survival periods shortened as the cultivation temperature increased. These results demonstrate that anaerobic conditions contribute to eradicating these pathogens during flooding or reductive soil disinfestation.     

Present status of Phytophthora species in the Mediterranean area,especially in relation to citrus1

Of the 44 currently accepted species of Phytophthora, 18 have been reported in the Mediterranean area. The status of each is briefly reviewed. On citrus, P. citrophthora and P. nicotianae var. parasitica are mainly responsible for foot rot and gummosis, and in addition P. hibernalis, P. syringae, P. cifricola and P. cactorum for brown rot of fruits. The incidence of these species is closely linked to their temperature requirements. The use of sour orange as a resistant rootstock has long provided satisfactory control of foot rot and gummosis, while chemical treatments are effective against brown rot. However, there are indications that, with changingcultural practices, the resistance of sour orange is less well expressed, and the species is incidentally also susceptible to citrus tristeza closterovirus. Alternative rootstocks are therefore being sought.     

Molecular Phylogenetic Analysis of Ribosomal DNA Internal Transcribed Spacer Regions and Comparison of Fertility in <Emphasis Type="Italic">Phomopsis </Emphasis>Isolates from Fruit Trees

Sequences of the ribosomal DNA (rDNA) internal transcribed spacer (ITS) regions were examined to infer a molecular phylogeny of small-spored Phomopsis isolates, designated W-type (mainly white colony, weakly virulent, bearing both alpha and beta conidia at 25°C on PDA) and G-type (mainly gray colony, highly virulent, bearing only alpha conidia at 25°C on PDA), and P. amygdali from fruit trees. Phomopsis G-type and P. amygdali were a monophyletic group distinct from the W-type. The W-type isolates were divided into two monophyletic groups. Diaporthe citri, D. tanakae, P. asparagi, P. viticola, P. vitimegaspora and D. nomurai, which are morphologically distinguishable from W- and G-types, differed from the W- and G-types in molecular phylogenetic analyses. PCR-RFLP analysis of rDNA ITS regions was useful to distinguish each of the Phomopsis species and groups using three restriction enzymes. In mating tests, W-type isolates from fruit trees were heterothallic and inter-fertile even between isolates belonging to the different monophyletic groups. Isolates of the G-type and P. amygdali collected in Japan were cross-fertile. Some isolates from Lunaria annua, Ulmus glabra and Juglans regia belonged to one of the two monophyletic groups of the W-type and were cross-fertile with W-type isolates from Rosaceous fruit trees. Received 27 September 1999/ Accepted in revised form 27 January 2000     

Effect of nitrogen form and application method on incidence and severity ofPhytophthora crown and root rot of apple trees

The effect of ammonium nitrate broadcast as a soil or through irrigation, urea applied as a foliar spray, and monoammonium phosphate applied as a planting hole treatment on the incidence ofPhytophthora crown and root rot of apple trees was determined under orchard conditions in the Okanagan Valley of British Columbia, Canada. Results from the eight year study showed that ammonium nitrate applied as a single dose in spring at 240 g tree^–1 year^–1, as a split dose at 120 g tree^–1 each in spring and early autumn, and in irrigation water (fertigation) at 7.5 g tree^–1 wk^–1 for 10 wk year^–1 significantly increasedPhytophthora crown and root rot of Macspur on MM106 rootstock. There was no significant difference inP. cactorum infection between the unfertilized control and treatments with urea applied as a foliar spray at 1.0 kg 100 l^–1 of water in spring and early autumn, and monoammonium phosphate applied as a planting hole treatment at 1 g l^–1 of soil at planting time.     

Control of Phytophthora species in plant stock for habitat restoration through best management practices

Emergent plant pathogens represent one of the most significant threats to biodiversity, and exotic Phytophthora species have recently emerged as a serious problem in restored habitats in California and in nurseries producing the plant stock. It is hypothesized that ‘best management practices’ prescribed through a Phytophthora Prevention Programme (PPP) could be useful in minimizing phytophthora disease incidence. To understand the magnitude of the problem and the efficacy of the PPP, plants in restoration nurseries were evaluated for (i) the Phytophthora species assemblage present in the absence of the PPP, and (ii) the effectiveness of the PPP to reduce them. Sampling included 203 plants grown in the absence of the PPP, and 294 grown implementing the PPP. Only samples collected in the absence of the PPP were Phytophthora-positive, and cumulatively yielded 55 isolates from 13 different taxa, including 1 putative interspecific hybrid genotype. There were 21 novel Phytophthora–plant species combinations. The most common Phytophthora species was P. cactorum. Four plant species had the highest disease incidence, namely: Diplacus aurantiacus (50 ± 11.2%), Heteromeles arbutifolia (33 ± 9.6%), Ceanothus thyrsiflorus (30 ± 8.4%), and Frangula californica (30 ± 8.4%). Disease incidence in nurseries after the implementation of the PPP dropped to zero (P < 0.001), and was unaffected to any significant degree by nursery differences, or plant species tested. This study identifies a large number of novel ‘plant species × Phytophthora species’ combinations, and provides for the first time strong evidence that the PPP significantly reduced Phytophthora in plant stock for habitat restoration.     

Alternative inoculum sources for fire blight: the potential role of fruit mummies and non‐host plants

Fire blight is the most damaging bacterial disease in apple production worldwide. Cankers and symptomless infected shoots are known as sites for the overwintering of Erwinia amylovora, subsequently providing primary inoculum for infection in the spring. In the present work, further potential sources of inoculum were investigated. Real‐time PCR assays covering a 3‐year‐period classified 19·9% of samples taken from fruit mummies as positive. Bacterial abundance in fruit mummies during autumn, winter and spring was up to 10⁹ cells per gram of tissue and correlated well with later infection rates of blossoms. Blossoms of non‐host plants growing close to infected trees were also shown to be colonized by E. amylovora and to enable epiphytic survival and propagation of bacteria. The results indicate a potential role of fruit mummies and buds in overwintering and as a source of primary inoculum for dissemination of the pathogen early in the growing season. Non‐host blossoms may also serve as an inoculum source in the build‐up of the pathogen population. Both aspects may contribute significantly to the epidemiology of E. amylovora. The significance of infected rootstocks as an inoculum source is also discussed. Fruit mummies might be used to determine pathogen pressure in an orchard before the beginning of the blooming period.