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1.
ABSTRACT To investigate host specialization in Macrophomina phaseolina, the fungus was isolated from soybean, corn, sorghum, and cotton root tissue and soil from fields cropped continuously to these species for 15 years in St. Joseph, LA. Chlorate phenotype of each isolate was determined after growing on a minimal medium containing 120 mM potassium chlorate. Consistent differences in chlorate sensitivity were detected among isolates from different hosts and from soil versus root. To further explore genetic differentiation among fungal isolates from each host, these isolates were examined by restriction fragment length polymorphism and random amplified polymorphic DNA (RAPD) analysis. No variations were observed among isolates in restriction patterns of DNA fragments amplified by polymerase chain reaction covering the internal transcribed spacer region, 5.8S rRNA and part of 25S rRNA, suggesting that M. phaseolina constitutes a single species. Ten random primers were used to amplify the total DNA of 45 isolates, and banding patterns resulting from RAPD analysis were compared with the neighbor-joining method. Isolates from a given host were genetically similar to each other but distinctly different from those from other hosts. Chlorate-sensitive isolates were distinct from chlorate-resistant isolates within a given host. In greenhouse tests, soybean, sorghum, corn, and cotton were grown separately in soil infested with individual isolates of M. phaseolina that were chosen based on their host of origin and chlorate phenotype. Root colonization and plant weight were measured after harvesting. More colonization of corn roots occurred when corn was grown in soil containing corn isolates compared with isolates from other hosts. However, there was no host specialization in isolates from soybean, sorghum, or cotton. More root colonization in soybean occurred with chlorate-sensitive than with chlorate-resistant isolates.  相似文献   

2.
Phenotypic and genetic diversity of 59 Macrophomina phaseolina isolates collected from various host species growing in or near cluster bean ( Cyamopsis tetragonoloba ) fields in four states of north and north-west India were characterized using RAPD and PCR–RFLPs of the ITS region. These isolates, and 11 from various hosts from culture collections, were classified into three mycelial phenotypes: dense, feathery and restricted, based on variable growth patterns on nutrient agar containing 120 m m chlorate. Pathogenicity of isolates was evaluated by measuring the length of stem lesions 21 days post-inoculation on the susceptible cluster bean genotype FS 277. Isolates showed considerable variation in aggressiveness, with the isolates from cluster bean with dense chlorate phenotype producing relatively higher lesion lengths on cluster bean plants. The results of the RAPD assay clearly distinguished the isolates on the basis of chlorate phenotype and host origin. Isolates from a single host were generally similar to each other, but differed distinctly from those from other hosts. Chlorate-sensitive isolates were distinct from chlorate-resistant isolates within a given host. A high degree of polymorphism in restriction patterns of the ITS region, including part of 25S rDNA, has been reported for the first time in the charcoal rot fungus.  相似文献   

3.
采用RAPD-PCR分子标记技术分析了51株不同地理来源、寄主来源的绿僵菌Metarhizium anisopliae菌株的遗传多态性。从94条RAPD引物中筛选出18条引物,对所有试验菌株进行RAPD-PCR扩增,共获得96条扩增片段,其中81条片段表现多态性,占84.1%。聚类分析表明,供试的51株菌株间的相似性系数范围为0.52~0.98,表明菌株间存在丰富的遗传多态性。供试菌株在相似性系数0.7的水平可分为4个组群。按菌株DNA多态性与地理及寄主来源的聚类分析表明,大多数菌株的DNA多态性与地理或寄主有一定的相关性,即长期的地理环境和寄主适应性可能形成了种群的分化。  相似文献   

4.
Isolates of the take-all fungus, Gaeumannomyces graminis var. avenae , which affects oats, wheat and other grasses, and of G.g. var. tritici , which preferentially affects wheat, rye and barley, contain a high proportion of repeated sequences. Total DNA from 57 fungal isolates collected from many locations and different cereal hosts, and scored for virulence on wheat, rye and oats, revealed many restriction fragment length polymorphisms. These RFLP s were observed either by staining the DNA directly, by hybridization to radioactively labelled total fungal DNA , or by hybridization with labelled wheat ribosomal DNA . With only a few exceptions, the isolates with the same preferred cereal hosts showed more similar patterns of restriction fragments than isolates that had different pathogenicity properties on cereal hosts, irrespective of the geographical origins of the isolates. This was even the case for R isolates of G.g. var. tritici that were virulent on wheat and rye compared with N isolates that were virulent only on wheat. Isolates were identified by hybridizing DNA from infected root samples with 32P-labelled total fungal DNA . The restriction fragment polymorphisms involving families of repeated sequence can therefore be used as a predictive assay for host preference of an isolate, and have probably arisen by host selection of fungal lineages. The variation between isolates in different pathogenicity groups suggests that there is little gene flow between isolates that can infect different hosts, even though they can coexist in the same field.  相似文献   

5.
ABSTRACT Ninety-five isolates of Colletotrichum including 81 isolates of C. acutatum (62 from strawberry) and 14 isolates of C. gloeosporioides (13 from strawberry) were characterized by various molecular methods and pathogenicity tests. Results based on random amplified polymorphic DNA (RAPD) polymorphism and internal transcribed spacer (ITS) 2 sequence data provided clear genetic evidence of two subgroups in C. acutatum. The first subgroup, characterized as CA-clonal, included only isolates from strawberry and exhibited identical RAPD patterns and nearly identical ITS2 sequence analysis. A larger genetic group, CA-variable, included isolates from various hosts and exhibited variable RAPD patterns and divergent ITS2 sequence analysis. Within the C. acutatum population isolated from strawberry, the CA-clonal group is prevalent in Europe (54 isolates of 62). A subset of European C. acutatum isolates isolated from strawberry and representing the CA-clonal and CA-variable groups was assigned to two pathogenicity groups. No correlation could be drawn between genetic and pathogenicity groups. On the basis of molecular data, it is proposed that the CA-clonal subgroup contains closely related, highly virulent C. acutatum isolates that may have developed host specialization to strawberry. C. gloeosporioides isolates from Europe, which were rarely observed were either slightly or nonpathogenic on strawberry. The absence of correlation between genetic polymorphism and geographical origin in Colletotrichum spp. suggests a worldwide dissemination of isolates, probably through international plant exchanges.  相似文献   

6.
Fifty-five isolates of Rosellinia necatrix, the cause of common avocado white root rot disease, were collected from south-east Spain and characterised according to their virulence behaviour and their molecular patterns to assess broader levels of genetic diversity. Virulence properties were revealed by in vitro inoculation on avocado plants. Differences in reaction types showed variability among these isolates. No sequence differences were observed when the internal transcribed spacer 1 (ITS1) and ITS2 regions and DNA fragments of the β-tubulin, adenosine triphosphatase and translation elongation factor 1 genes were explored in representive isolates from five virulence groups. Random amplified polymorphic DNA (RAPD) amplifications were also performed for each isolate using 19 random primers. Four of these primers revealed polymorphism among isolates and repetitive and discriminative bands were used to build an unweighted pair group with arithmetic mean tree. However, RAPD clustering showed low stability, and no correlation between RAPD and virulence groups was observed, possibly indicating high levels of sexual recombination.  相似文献   

7.
Le Cam B  Parisi L  Arene L 《Phytopathology》2002,92(3):314-320
ABSTRACT Genetic relationships, mating crosses, and host specificity of Venturia inaequalis isolates from Malus spp. and of Spilocaea pyracanthae isolates from Pyracantha spp. were evaluated. Sequence analysis of the complete internal transcribed spacer (ITS) region (ITS1-5.8S to ITS2) revealed a total similarity between these two putative species. ITS restriction fragment length polymorphism carried out with five restriction enzymes on a collection of 28 isolates confirmed a lack of diversity in this region between and within these two populations. Additional isolates from three related species (V. pirina, V. nashicola, and S. eriobotryae) were divided into two distinct monophyletic groups in a phylogenetic tree using ITS sequence comparison. These groups were related to their anamorph (i.e., Spilocaea or Fusicladium). When inoculated on their host of origin, fields isolates caused typical symptoms of scab disease, and a host specificity was demonstrated by cross pathogenicity of isolates from Malus x domestica and Pyracantha spp. Mating on dried leaves in vitro between one isolate of each putative species led to production of numerous perithecia. Ninety-six sporulating monoascosporic progenies were isolated from this cross. Based on these genetic and pathogenic data, we proposed that pathogens responsible for scab on Malus spp. and Pyracantha spp. are considered as two formae speciales belonging to V. inaequalis.  相似文献   

8.
ABSTRACT The blackleg disease of oilseed rape is caused by an ascomycete species complex termed Leptosphaeria maculans (anamorph Phoma lingam). L. maculans isolates collected worldwide were gathered in the International Blackleg of Crucifers Network (IBCN) collection. Representative IBCN isolates, along with one P. nigrificans isolate, were further analyzed using polymerase chain reaction (PCR) amplification of the internal transcribed spacer (ITS) region. ITS size polymorphism discriminated three groups: (i) P. nigrificans, (ii) Tox(+) and 'Lepidium' isolates, and (iii) NA1, NA2, NA3, 'Thlaspi', and 'Erysimum' isolates. Digestion of the ITS region with 19 selected endonucleases showed restriction site polymorphism between the different subgroups: digestion with RsaI could discriminate Tox(+) from 'Lepidium' isolates, whereas digestion with four enzymes, i.e., HaeIII, EcoRII, RsaI, and AluI, was needed to discriminate between NA1, NA2, NA3, 'Thlaspi', and 'Erysimum' isolates. No restriction site polymorphism was observed between isolates within the 'Thlaspi', Tox(+), NA1, and NA2 subgroups. Direct amplification of the ITS region could be achieved using intact conidia, collected either in axenic cultures or on leaf lesions, with only a 4-min 95 degrees C denaturation step prior to PCR reaction. A routine identification protocol requiring no DNA extraction and a sequential use of a few restriction enzymes following PCR has been used successfully for large-scale identification of French field isolates.  相似文献   

9.
10.
Qu X  Christ BJ 《Phytopathology》2006,96(10):1157-1163
ABSTRACT Spongospora subterranea f. sp. subterranea causes powdery scab in potatoes and is distributed worldwide. Genetic studies of this pathogen have been hampered due, in part, to its obligate parasitism and the lack of molecular markers for this pathogen. In this investigation, a single cystosorus inoculation technique was developed to produce large amounts of S. subterranea f. sp. subterranea plasmodia or zoosporangia in eastern black nightshade (Solanum ptycanthum) roots from which DNA was extracted. Cryopreservation of zoosporangia was used for long-term storage of the isolates. S. subterranea f. sp. subterranea-specific restriction fragment length polymorphism (RFLP) markers were developed from randomly amplified polymorphic DNA (RAPD) fragments. Cystosori of S. subterranea f. sp. subterranea were used for RAPD assays and putative pathogen-specific RAPD fragments were cloned and sequenced. The fragments were screened for specificity by Southern hybridization and subsequent DNA sequence BLAST search. Four polymorphic S. subterranea f. sp. subterranea-specific probes containing repetitive elements, and one containing single copy DNA were identified. These RFLP probes were then used to analyze 24 single cystosorus isolates derived from eight geographic locations in the United States and Canada. Genetic variation was recorded among, but not within, geographic locations. Cluster analysis separated the isolates into two major groups: group I included isolates originating from western North America, with the exception of those from Colorado, and group II included isolates originating from eastern North America and from Colorado. The techniques developed in this study, i.e., production of single cystosorus isolates of S. subterranea f. sp. subterranea and development of RFLP markers for this pathogen, provide methods to further study the genetic structure of S. subterranea f. sp. subterranea.  相似文献   

11.
DNA isolated from the formae speciales of Erysiphe graminis that grow on barley, wheat, rye and oats was studied using restriction endonucleases and DNA/DNA hybridization procedures. DNA fragments were purified by molecular cloning and a few containing repeated sequences were used to demonstrate the many variations in restriction fragments both within and between the four formae speciales. In an analysis of six single-colony isolates of the barley mildew pathogen collected from different UK sites in different years, more than a quarter of the fragments scored varied among isolates. One isolate, with an uncommon pathogenicity character, differed from the remainder in the distribution of DNA bands. Isolates of rye mildew were also distinct from one another but isolates of oat mildew from a population of similar size appeared to belong to a single clone.
It is concluded that the chromosomes of E. graminis contain many families of dispersed repeated sequences and that there may be extensive polymorphism for restriction endonuclease cleavage sites associated with these repeats. Such unselected polymorphisms could be useful in helping to understand and discriminate among the factors affecting population structure in the pathogen as it responds to different agricultural practices.  相似文献   

12.
Host Range Specificity in Verticillium dahliae   总被引:1,自引:0,他引:1  
Bhat RG  Subbarao KV 《Phytopathology》1999,89(12):1218-1225
ABSTRACT Verticillium dahliae isolates from artichoke, bell pepper, cabbage, cauliflower, chili pepper, cotton, eggplant, lettuce, mint, potato, strawberry, tomato, and watermelon and V. albo-atrum from alfalfa were evaluated for their pathogenicity on all 14 hosts. One-month-old seedlings were inoculated with a spore suspension of about 10(7) conidia per ml using a root-dip technique and incubated in the greenhouse. Disease incidence and severity, plant height, and root and shoot dry weights were recorded 6 weeks after inoculation. Bell pepper, cabbage, cauliflower, cotton, eggplant, and mint isolates exhibited host specificity and differential pathogenicity on other hosts, whereas isolates from artichoke, lettuce, potato, strawberry, tomato, and watermelon did not. Bell pepper was resistant to all Verticillium isolates except isolates from bell pepper and eggplant. Thus, host specificity exists in some isolates of V. dahliae. The same isolates were characterized for vegetative compatibility groups (VCGs) through complementation of nitrate nonutilizing (nit) mutants. Cabbage and cauliflower isolates did not produce nit mutants. The isolate from cotton belonged to VCG 1; isolates from bell pepper, eggplant, potato, and tomato, to VCG 4; and the remaining isolates, to VCG 2. These isolates were also analyzed using the random amplified polymorphic DNA (RAPD) method. Forty random primers were screened, and eighteen of them amplified DNA from Verticillium. Based on RAPD banding patterns, cabbage and cauliflower isolates formed a unique group, distinct from other V. dahliae and V. albo-atrum groups. Minor genetic variations were observed among V. dahliae isolates from other hosts, regardless of whether they were host specific or not. There was no correlation among pathogenicity, VCGs, and RAPD banding patterns. Even though the isolates belonged to different VCGs, they shared similar RAPD profiles. These results suggest that management of Verticillium wilt in some crops through crop rotation is a distinct possibility.  相似文献   

13.
ABSTRACT Alternaria isolates were obtained from various pistachio tissues collected in five orchards in California. For all isolates, morphological characteristics of the colony and sporulation apparatus were determined and compared with those of representative isolates of A. alternata, A. tenuissima, A. arborescens, and A. infectoria. A selection of the pistachio isolates and the representative Alternaria isolates were evaluated for pathogenicity to pistachio. Molecular characteristics of these isolates were determined using random amplified polymorphism DNA (RAPD) analysis, polymerase chain reaction restriction fragment length polymorphism (PCR-RFLP) analysis of nuclear intergenic spacer rDNA, and sequence analysis of nuclear internal transcribed spacer (ITS) rDNA. Based on morphological characteristics, the pistachio isolates were grouped as identical or very similar to either A. alternata, A. tenuissima, A. arborescens, or A. infectoria. Isolates from the alternata, tenuissima, and arborescens species-groups were pathogenic to pistachio and no significant differences in pathogenicity were observed. Isolates from the infectoria species-group were only weakly pathogenic to pistachio. Based on cluster analysis of RAPD and PCR-RFLP data, three distinct clusters were evident; the infectoria cluster, the arborescens cluster, and a combined alternata/tenuissima cluster. Based on analysis of ITS sequence data, the infectoria species-group was phylogenetically distinct from the other species-groups. Isolates of the alternata, tenuissima, and arborescens species-groups comprised a monophyletic clade in which the three species-groups could not be further resolved.  相似文献   

14.
Since its initial detection in Australia in 1979, wheat yellow (stripe) rust ( Puccinia striiformis f.sp. tritici ) has evolved in Australia and New Zealand into more than 20 pathotypes with assorted virulence characteristics. This evolution is believed to have occurred in a stepwise fashion from an original single pathotype, with no subsequent new introductions. A combination of random amplified polymorphic DNA (RAPDs) and amplified fragment length polymorphisms (AFLPs) was used to examine the level of molecular variation in Australian and New Zealand isolates, and to compare this with variation amongst other isolates of P. striiformis . Using 60 RAPD primers on seven Australian isolates representing seven different pathotypes collected between 1979 and 1991, more than 300 potentially polymorphic loci were analysed and no polymorphisms were detected. Using the same primers on two UK isolates, 3% of loci showed a polymorphism. A similar level of polymorphism was found between UK isolates using AFLP primers, and between 5 and 15% of fragments were polymorphic between an isolate from the UK, an isolate from Denmark, and one from Colombia. However, no AFLP polymorphisms were found amongst 14 Australian and New Zealand isolates tested, at over 100 potentially polymorphic loci. The lack of molecular variation in the Australian and New Zealand collection is consistent with the stepwise mutation theory of pathotype evolution from a single introduction.  相似文献   

15.
Pythium group F is a ubiquitous, though minor, pathogen in several soilless and soil cultures; investigations were carried out to analyze different regions of the DNA and better understand the nature of this group. Fourty-two isolates were obtained from a variety of plants (cucumber, lettuce, tomato) grown in soil or soilless cultures collected in various countries (Canada, Denmark, France, Norway, Sweden and United Kingdom). All Pythium group F isolates displayed amplified ITS1-5,8S-ITS2 ribosomal DNA region (rDNA) of similar length, whereas polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) revealed that, among the seven enzymes used, polymorphism was only identified with Hin6I. After cloning of ITS1-5,8S-ITS2 rDNA region from Pythium group F isolates that displayed restriction polymorphism patterns with Hin6I, comparisons of sequence and restriction mapping data showed a slight variation consisting in a single base change. Inter Simple Sequence Repeat (ISSR)-PCR method was also used to obtain data related to the entire genome and not only to a single DNA region. It identified repeated motifs in the genome of Pythium group F isolates. Two primers (CAC)5 and (CCA)5 detected polymorphism, and isolates were classified among 11 molecular clusters. The genetic diversity of this group was not correlated with the geographical locations or the host plants from which the isolates originated. Polymorphism of Pythium group F isolates pointed out by ISSR is discussed  相似文献   

16.
A new Phytophthora disease of common alder (Alnus glutinosa) similar to that previously reported in several countries in Europe has been observed in Hungary. Based on these earlier studies, the alder Phytophthora was considered likely to be a hybrid between P cambivora and a P fragariae-like species: across Europe a range of new alder Phytophthora is spreading that comprise a range of heteroploid hybrids including a 'standard' hybrid type and several other hybrid types termed 'variants'. Phenotypic and molecular features of the pathogen in Hungary were characterised and compared with isolates from elsewhere. The morphologies of five isolates from one region (Hévíz) resembled the common, 'standard' type, whereas the three isolates from another region (Hanság) exhibited traits similar to those of one of the 'variant' types, ie the Swedish 'variant'. Molecular markers of these two groups of Hungarian isolates also represented a good fit to those of the standard type and the Swedish variant, respectively. Isozyme patterns and profiles of restriction fragments of the entire internal transcribed spacer (ITS) region or mitochondrial DNAs and of RAPD-PCR products did not differ within a group, but distinct polymorphisms were exhibited between the two groups of isolates. Southern analysis of random amplified polymorphic DNA (RAPD) revealed the homologous nature of co-migrating bands of P cambivora and the isolates of alder Phytophthora. Furthermore, restriction fragment profiles of the ITS region of ribosomal DNAs and the mtDNAs were consistent with reported biparental origin of alder Phytophthora. The hybrid status of these continuously evolving pathogens raises many issues and challenges concerning efficient control measures.  相似文献   

17.
ABSTRACT Previous studies indicated that incompletely dominant loci determine insensitivity by oomycetes to phenylamide fungicides such as metalaxyl. To compare the bases of insensitivity in different strains of the late blight pathogen, Phytophthora infestans, crosses were performed between sensitive isolates and isolates from Mexico, the Netherlands, and the United Kingdom that displayed varying levels of insensitivity. Segregation analyses indicated that metalaxyl insensitivity was determined primarily by one locus in each isolate, and that two of the isolates were heterozygous and the other homozygous for the insensitive allele. Metalaxyl insensitivity was also affected by the segregation of additional loci of minor effect. DNA markers linked to insensitivity were obtained by bulked segregant analysis using random amplified polymorphic DNA (RAPD) markers and the Dutch and Mexican crosses. By studying the linkage relationships between these markers and the insensitivity in each cross by RAPD or restriction fragment length polymorphism analysis, it appeared that the same chromosomal locus conferred insensitivity in the Mexican and Dutch isolates. However, a gene at a different chromosomal position was responsible for insensitivity in the British isolate.  相似文献   

18.
ABSTRACT Eighty-six isolates of Botryosphaeria dothidea, the causal agent of Botryosphaeria panicle and shoot blight of pistachio, were collected from pistachio and other plant hosts in California. The isolates were characterized by microsatellite-primed polymerase chain reaction (MP-PCR), sequences of the nuclear ribosomal DNA internal transcribed spacer region (ITS1, 5.8S gene, and ITS2), morphological and cultural characters, osmotic and fungicide sensitivity, and pathogenicity on pistachio. Three groups of these isolates were identified based upon analysis of MP-PCR data and ITS sequences. Group I contained 43 pycnidiospore-derived isolates collected from pistachio and other hosts. Group II consisted of 20 ascosporic isolates obtained from a single sequoia plant. Group III consisted of 20 ascosporic isolates from three shoots on a single blackberry plant, two pycnidiospore-derived isolates from incense cedar, and one from pistachio. Group I predominated over the other two groups in California pistachio orchards. B. dothidea isolates of group III grew faster on acidified potato dextrose agar (APDA) than the isolates of groups I and II. Isolates of group III produced pycnidia on both APDA and autoclaved pistachio shoots, but the isolates of the other two groups produced pycnidia on only autoclaved pistachio shoots. Additionally, significant differences in osmotic and fungicide sensitivities were observed among these three groups. Results from lathhouse inoculations demonstrated that the representative isolates for each of the three groups were all capable of infecting pistachio and producing characteristic disease symptoms of Botryosphaeria blight. The virulence of group II isolates on pistachio was, however, significantly lower than that of group I isolates.  相似文献   

19.
The degree of genetic diversity of 127 Colletotrichum gloeosporioides isolates from Stylosanthes guianensis genotypes in South America was measured at the molecular level by random amplified polymorphic DNA (RAPD) with nine arbitrary primers of 10 bases, and by restriction fragment length polymorphism (RFLP) with a non-LTR (long terminal repeats) retrotransposon DNA sequence. The RAPD products revealed scorable polymorphism among the isolates, and a total of 80 band positions were scored. Sixty-three of the 127 isolates were clustered into 13 distinct lineages usually correlating with geographic origin. Where isolates from various regions were clustered together, most had identical host genotype origin. The pathogen population sampled from Carimagua, Colombia, a long-time Stylosanthes breeding and selection site, with a savanna ecosystem, was highly diverse. A set of 12 S. guianensis genotype differentials was used to characterize pathogenic variability of 104 isolates and their virulence patterns were grouped into 57 pathotypes. However, when they were tested on four Australian differentials, they grouped into 11 pathotypes. As shown in previous studies, no strict correlations existed between genetic diversity measured by RAPD or RFLP, and pathotype defined by pathogenicity pattern on the differentials. Southern blot analysis of the 127 isolates revealed 23 hybridizing fragments, resulting in 41 fingerprint patterns among the 127 isolates. Relationships between RFLP and RAPD variables were examined using Spearman's Rank Correlation Coefficient, which showed that the two measures of genotypic variation are in agreement.  相似文献   

20.
ABSTRACT A large epidemiological study of the genetic variation of barley yellow dwarf virus (BYDV) serotype PAV involving different host plant species was conducted. French BYDV PAV isolates were collected from barley and ryegrass, and their capsid protein gene sequences characterized using restriction fragment length polymorphism, single-strand conformation polymorphism, and sequence analyses. The data show that BYDV PAV isolates from five different continents are separated into two distinct groups named cpA and cpB, which are distributed irrespective of geographical location. Amino acid identity of the capsid proteins ranged from 93 to 99.5% in group cpA and from 95 to 99.5% in group cpB, while this value was only from 82 to 88% between the groups. Moreover, isolates from each group were found preferentially (up to 98%) in one of the two plant species examined. These results show that host plant species play a role in isolate selection and maintenance and that they contribute to the genetic diversity of BYDV PAV.  相似文献   

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