Molecular detection and genotyping of Fusarium oxysporum f. sp. psidii isolates from different agro-ecological regions of India

Twenty one isolates of Fusarium oxysporum f. sp. psidii (Fop), causing a vascular wilt in guava (Psidium guajava L.), were collected from different agro-ecological regions of India. The pathogenicity test was performed in guava seedlings, where the Fop isolates were found to be highly pathogenic. All 21 isolates were confirmed as F. oxysporum f. sp. psidii by a newly developed, species-specific primer against the conserved regions of 28S rDNA and the intergenic spacer region. RAPD and PCR-RFLP were used for genotyping the isolates to determine their genetic relationships. Fifteen RAPD primers were tested, of which five primers produced prominent, polymorphic, and reproducible bands. RAPD yielded an average of 6.5 polymorphic bands per primer, with the amplified DNA fragments ranging from 200–2,000 bp in size. A dendrogram constructed from these data indicated a 22–74% level of homology. In RFLP analysis, two major bands (350 and 220 bp) were commonly present in all isolates of F. oxysporum. These findings provide new insight for rapid, specific, and sensitive disease diagnosis. However, genotyping could be useful in strain-level discrimination of isolates from different agro-ecological regions of India.     

Suppression of Fusarium oxysporum with recombinant polygalacturonase inhibiting proteins (BvPGIPs) extracted from sugar beet roots

Soil-borne fungus Fusarium oxysporum f. sp. betae (Fob) is the causative agent of Fusarium yellows in sugar beet. Leaf interveinal yellowing and root vascular discoloration significantly reduce root yield as well as sucrose content and juice purity. Fob, like other fungal pathogens, initiates disease development by secreting polygalacturonase (PG) enzymes to break down plant cell walls during early stages of infection. To protect themselves, plants produce polygalacturonase-inhibiting proteins (PGIPs). In our study of sugar beet root defense responses, several PGIP genes (BvPGIPs) were identified. To determine if BvPGIPs inhibit Fob PGs, genes BvPGIP1, BvPGIP2 and Bv(FC607)PGIP1 were fused with the CaMV 35S promoter and each was expressed individually in sugar beet hairy roots. We demonstrate that all three recombinant BvPGIP proteins inhibited Fob and F. oxysporum f. sp. gladioli (Fog) PGs. A comparable level of BvPGIP activity was observed against Fob PGs, while BvPGIP2 showed higher activity against Fog PGs. Similar results were obtained when recombinant PGIPs were used to bioassay effects on Fob and Fog spore germination and hyphal growth. This is a first report that documents F. oxysporum inhibition by overexpressing BvPGIPs that may lead to improved Fusarium yellows resistance in sugar beet.

     

Characterization of virulent gene locus in the genome of Fusarium spp. causing wilt disease of Psidium guajava L. in India

Wilt of Psidium guajava L., incited by Fusarium oxysporum f. sp. psidii and Fusarium solani is a serious soil borne disease of guava in India. Forty-two isolates, each of F. oxysporum f. sp. psidii (Fop) and F. solani (Fs), collected from different agro climatic zones of India showing pathogenicity were subjected to estimate their virulence factor in terms of analysis using virulent gene-related microsatellite loci. The erratic spread and occurrence of guava wilt in different areas may be due to variable aggressiveness or virulence of different pathogenic isolates in the soil. Out of 10 virulent gene locus related microsatellite markers ofFusarium spp., only six marker viz. Xyl, KHS1, PelA1, PG6/7, CHS1/2 and FMK1/MAPK1 were successfully amplified. This indicates that all the tested Fusarium sp. isolates of guava are having virulence gene in their genome. Microsatellite marker for virulence factor genes of Xyl loci was amplified in both Fop and Fs isolates. Product size of 281 bps was exactly amplified with a single banding pattern in all the isolates of Fop and Fs. It has been observed that other five microsatellite marker for virulence factor genes such as KHS1, PelA1, PG6/7, CHS1/2 and FMK1/MAPK1 were amplified with specific band pattern. PG6/7, CHS1/2 and FMK1/MAPK1 were only amplified in Fop isolates with a product size of 765 bps, 1566 bps; 1010 bps and 1244 bps. PelA1 and KHS1were amplified only in Fs isolates with the product size of 586 bps; 1359 bps, respectively. The results indicate that virulence factor genes are in response to produce wilt disease like symptoms in guava plants and also having pathogenic gene-related locus.     

Genetic Diversity of Fusarium oxysporum Strains from Common Bean Fields in Spain

Fusarium wilt is an endemic disease in El Barco de Avila (Castilla y León, west-central Spain), where high-quality common bean cultivars have been cultured for the last century. We used intergenic spacer (IGS) region polymorphism of ribosomal DNA, electrophoretic karyotype patterns, and vegetative compatibility and pathogenicity analyses to assess the genetic diversity within Fusarium oxysporum isolates recovered from common bean plants growing in fields around El Barco de Avila. Ninety-six vegetative compatibility groups (VCGs) were found among 128 isolates analyzed; most of these VCGs contained only a single isolate. The strains belonging to pathogenic VCGs and the most abundant nonpathogenic VCGs were further examined for polymorphisms in the IGS region and electrophoretic karyotype patterns. Isolates belonging to the same VCG exhibited the same IGS haplotype and very similar electrophoretic karyotype patterns. These findings are consistent with the hypothesis that VCGs represent clonal lineages that rarely, if ever, reproduce sexually. The F. oxysporum f. sp. phaseoli strains recovered had the same IGS haplotype and similar electrophoretic karyotype patterns, different from those found for F. oxysporum f. sp. phaseoli from the Americas, and were assigned to three new VCGs (VCGs 0166, 0167, and 0168). Based on our results, we do not consider the strains belonging to F. oxysporum f. sp. phaseoli to be a monophyletic group within F. oxysporum, as there is no correlation between pathogenicity and VCG, IGS restriction fragment length polymorphism, or electrophoretic karyotype.     

Genetic Diversity of Fusarium oxysporum Populations Isolated from Tomato Plants in Tunisia

Fusarium crown and root rot of tomato (Lycopersicon esculentum) caused by Fusarium oxysporum f. sp. radicis‐lycopersici is a new devastative disease of tomato greenhouse crops in Tunisia. Nothing is known neither about the population of this pathogen in this region, nor about the population of F. oxysporum f. sp. lycopersici the causal agent of Fusarium wilt of tomato. In order to examine the genetic relatedness among the F. oxysporum isolates by intergenic spacer restriction fragment length polymorphism (IGS‐RFLP) analysis and to elucidate the origin of the formae specialesradicis‐lycopersici in Tunisia by looking for genetic similarity of Tunisians isolates with isolates from a foreign source, the genetic diversity among F. oxysporum f. sp. radicis‐lycopersici and F. oxysporum f. sp. lycopersici populations was investigated. A total of 62 isolates of F. oxysporum, obtained from symptomless tomato plants, were characterized using IGS typing and pathogenicity tests on tomato plants. All Fusarium isolates were highly pathogenic on tomato. Fusarium oxysporum f. sp. radicis‐lycopersici isolates were separated into five IGS types. From the 53 F. oxysporum f. sp. radicis‐lycopersici isolates, 34 isolates have the same IGS types (IGS type 25), and the remaining 19 isolates were distributed into four IGS types. However, the only nine isolates of F. oxysporum f. sp. lycopersici have six different IGS types. This difference of diversity between the two formae speciales suggests that F. oxysporum f. sp. radicis‐lycopersici isolates have a foreign origin and may have been accidentally introduced into Tunisia.     

Molecular characterisation of Fusarium oxysporum causing rot diseases in small cardamom (Elettaria cardamomum Maton)

Incidence of root rot and foliar yellowing, rhizome rot, panicle wilt and stem rot diseases of small cardamom (Elettaria cardamomum Maton) are caused by Fusarium oxysporum Schlecht., and were surveyed in the high ranges of Idukki district, Kerala during 2010–2011. The diseases were noticed in different areas to varying degrees. Root rot was found to be most severe, followed by pseudostem rot, rhizome rot and panicle wilt. The Fusarium infections were prevalent throughout the year (January–December) and varied from 1.5 to 10.6%. Even though the pathogen was isolated from different plant parts, during pathogenicity studies, all the isolates could cross-infect other plant parts too. Twenty different isolates of F. oxysporum were obtained from diseased samples, and five morphologically distinct isolates were analysed with Randomly Amplified Polymorphic DNA (RAPD) markers to study the genetic variability, if any, among them. PCR amplification of total genomic DNA with random oligonucleotide primers generated unique banding patterns, depending upon primers and isolates. Nine oligunucleotide primers were selected for the RAPD assays, which resulted in 221 bands for the five isolates of F. oxysporum. The number of bands obtained was entered into an NTSYS, and the results showed moderate genetic variability among F. oxysporum isolates causing root rot, rhizome rot, panicle wilt and pseudostem rot, collected from different locations. The dendrogram of different isolates into groups resulted in one major cluster at 0.61 similarity index comprising of four isolates (CRT 3, CRR 3, CPW 2 and CSR 1) and one isolate (CRT 5) formed in a separate cluster. Among the five isolates of F. oxysporum, CRT 5 was entirely different from the other four isolates. The isolates also differ according to the geographical area, as revealed from the genetic variability observed in different root rot isolates (CRT 3 and CRT 5). It is inferred that despite moderate variability, F. oxysporum, infecting small cardamom in Idukki district of Kerala, consists of a single clonal lineage.     

Use of RAPD and ISSR Markers in Detection of Genetic Variation and Population Structure among Fusarium oxysporum f. sp. ciceris Isolates on Chickpea in Turkey

Genetic variation among the isolates of Fusarium oxysporum f. sp. ciceris, the causal agent of chickpea wilt worldwide, was analysed using pathogenicity tests and molecular markers – random amplified polymorphic DNA (RAPD) and inter‐simple sequence repeat (ISSR) polymorphism. Hundred and eight isolates were obtained from diseased chickpea plants in 13 different provinces of Turkey, out of which 74 isolates were assessed using 30 arbitrary decamer primers and 20 ISSR primers. Unweighted pair‐grouped method by arithmetic average cluster analysis of RAPD, ISSR and RAPD + ISSR datasets provided a substantially similar discrimination among Turkish isolates and divided into three major groups. Group 1, 2 and 3 consisted of 41, 18 and 15 isolates, respectively. These methods revealed a considerable genetic variation among Turkish isolates, but no correlation with regard to the clustering of isolates from different geographic regions. Analysis of molecular variance confirmed that most genetic variability resulted from the differences among isolates within regions. Our results also indicated that the low‐genetic differentiation (F_ST) and high gene flow (Nm) among populations had a significant effect on the emergence and evolutionary development of F. oxysporum f. sp. ciceris. This is the first report on genetic diversity and population structure of F. oxysporum isolates on chickpea in Turkey.     

Molecular Characterization of Fusarium oxysporum f. melongenae by ISSR and RAPD Markers on Eggplant

Fusarium oxysporum f. melongenae is a major soil-borne pathogen of eggplant (Solanum melongena). ISSR and RAPD markers were used to characterize Fusarium oxysporum f. melongenae isolates collected from eggplant fields in southern Turkey. Those isolates were not pathogenic to tomato. Pathogens were identified by their morphology, and their identity was confirmed by PCR amplification using the specific primer PF02-3. The isolates were classified into groups on the basis of ISSR and RAPD fingerprints, which showed a level of genetic specificity and diversity not previously identified in Fusarium oxysporum f. melongenae, suggesting that genetic differences are related to the pathogen in the Mediterranean region. The primers selected to characterize Fusarium oxysporum f. melongenae may be used to determine genetic differences and pathogen virulence. This study is the first to characterize eggplant F. oxysporum species using ISSR and RAPD.     

Analysis of Genetic Diversity of Fusarium oxysporum f. sp. phaseoli Isolates, Pathogenic and Non-pathogenic to Common Bean (Phaseolus vulgaris L.)

Twenty isolates of Fusarium oxysporum from Brazil, pathogenic and non‐pathogenic to common bean, were analysed using random amplified polymorphic DNA (RAPDs) to study the genetic diversity. RAPD analysis using 23 oligonucleotides resulted in the amplification of 229 polymorphic and 7 monomorphic DNA fragments ranging from 234 to 2590 bp. High genetic variability was observed among the isolates, with the distances varying between 8% and 76% among pathogenic, 2% and 63% among the non‐pathogenic and 45% and 76% between pathogenic and non‐pathogenic isolates. The analysis of genetic distance data showed that the pathogenic isolates tended to group in one group and the non‐pathogenic in another. The genetic distance values of 30% among the pathogenic isolates in cluster A are compatible with the genetic distance values observed within the physiological races, but the distance values among the pathogenic isolates in clusters B and G are not compatible with the distance values observed within the race. Although our results are preliminary, it was not possible to exclude the existence of more than one race of this fungus in Brazil.     

Characterization of Colletotrichum lindemuthianum Isolates from the State of Minas Gerais, Brazil

Anthracnose disease of common bean (Phaseolus vulgaris), caused by Colletotrichum lindemuthianum, is responsible for extensive yield losses worldwide. This pathogen is known to vary greatly in its pathogenicity. Control strategies include chemical control and, mainly, the development of resistant cultivars, taking into account the population structure of C. lindemuthianum. The objective of this study was to investigate the pathogenic and genetic diversity and population structure among C. lindemuthianum isolates collected in Minas Gerais state, Brazil. When these isolates were inoculated on 12 differential cultivars, a total of 10 races were identified within a series of 48 isolates collected in Minas Gerais, Brazil. Races 65, 81 and 73 were the most frequent races and occurred in most of the regions. This study also detected race 337, which had not been reported previously in the literature. Random amplified polymorphic DNA (RAPD) analysis performed on the same 48 isolates revealed great genetic diversity, clustering the series into five groups at a maximum similarity value of 89.6%. There was no clear relationship between the loci sampled by RAPD markers and the pathogenic characterization. Analysis of molecular variance showed that 96.06% of the variability was contained within regions and 3.94% among regions, indicating a high exchange of genetic material among the regions of the State. Most of the variability was detected within races (75.24%). The pathogenicity and RAPD assays corroborated the broad genetic diversity of the pathogen and the results have been useful in breeding for resistance to anthracnose.     

Microsatellite marker-based detection of Fusarium wilt pathogens of Psidium guajava L.

Wilt of Psidium guajava L., incited by Fusarium oxysporum f. sp. psidii and Fusarium solani is a serious soil-borne disease of guava in India. Forty-two isolates each of F. oxysporum f. sp. psidii (Fop) and F. solani (Fs) collected from different agro climatic zones of India showing pathogenicity were subjected to estimate the genetic and molecular characterisation in terms of analysis of microsatellite marker studies. Out of eight microsatellite markers, only four microsatellite markers, viz. MB 13, MB 17, RE 102 and AY212027 were amplified with single band pattern showing the character of identical marker for molecular characterisation and genetic identification. Microsatellite marker MB 13 was amplified in F. oxysporum f. sp. psidii and F. solani isolates. Product size of 296 bps and 1018 bps were exactly amplified with a single banding pattern in all the isolates of F. oxysporum f. sp. psidii and F. solani, respectively. Microsatellite markers, viz. MB 17, RE 102 and AY212027 were also exactly amplified with a single banding pattern. MB 17 was amplified in F. oxysporum f. sp. psidii isolates with a product size of 300 bp. RE 102 and AY212027 were amplified in F. solani isolates with the product size of 153 bp and 300 bp, respectively. Therefore, amplified microsatellite marker may be used as identifying DNA marker.     

Analysis of genetic variability of Fusarium oxysporum f. sp. cepae the causal agent of basal rot on onion using RAPD markers

Genetic variability among isolates of Fusarium oxysporum f. sp. cepae was obtained from different onion-growing areas of Tamil Nadu, India. Random amplified polymorphic DNA (RAPD) analysis was carried out using 12 random primers, each of them consisting of 10 base pairs. Four out of the 12 primers were differentiated between some of the tested F. oxysporum f. sp. cepae isolates. Analysis of the genetic coefficient matrix derived from the scores of RAPD profile showed that minimum and maximum per cent similarities among the F. oxysporum f. sp. cepae isolates were in the range of 14–85%. Cluster analysis, using the unweighted pair-group method with arithmetic average, clearly separated the isolates into two clusters (A and B) confirming the genetic diversity among the isolates of F. oxysporum f. sp. cepae from onion.     

Interaction between Meloidogyne incognita and Fusarium oxysporum f. sp. phaseoli on Selected Bean Genotypes

Four bean genotypes (IPA-1, A-107, A-211, and Calima), representing all possible combinations of resistance and susceptibility to Fusarium oxysporum f. sp. phaseoli (Fop) and Meloidogyne incognita, were each inoculated with three population densities of these pathogens. Calima and A-107 were resistant to Fop; A-107 and A-211 were resistant to M. incognita; and IPA-1 was susceptible to both pathogens. In Fop-susceptible lines (IPA-1 and A-211), the presence of M. incognita contributed to an earlier onset and increased severity of Fusarium wilt symptoms and plant stunting. However, the Fop-resistant Calima developed symptoms of Fusarium wilt only in the presence of M. incognita. Genotype A-107 (resistant to both M. incognita and Fop) exhibited Fusarium wilt symptoms and a moderately susceptible reaction to Fop only after the breakdown of its M. incognita resistance by elevated incubation temperatures (27 C). Root galling and reproduction of M. incognita was generally increased as inoculum density of M. incognita was increased on the M. incognita susceptible cultivars. However, these factors were decreased as the inoculum density of Fop was increased. It was concluded that severe infections of bean roots by M. incognita increase the severity of Fusarium wilt on Fop-susceptible genotypes and may modify the resistant reaction to Fop.     

RAPD based genetic diversity among different isolates of <Emphasis Type="Italic">Fusarium oxysporum</Emphasis> f. sp<Emphasis Type="Italic">. lycopersici</Emphasis> and their comparative biocontrol

Fusarium wilt of tomato (Solanum lycopersicum Mill.) caused by Fusarium oxysporum f. sp. lycopersici (Sacc.) W. C. Snyder and H. N. Hans (Fol.), is most serious and versatile pathogen. Chemical control of disease is not satisfactory and biological control is an attractive and potential alternative to the use of chemicals to control fusarium wilt of tomato. No any bioagent is universally effective everywhere therefore, search for potential biocontrol agent is continuous process and mandatory for several and individual ecological niches. In this experiment biocontrol efficacy of five species of Aspergillus and five species of Trichoderma were evaluated in vitro against Fusarium oxysporum f. sp. lycopersici. In both the experiments (dual culture and culture filtrates) T. harzianum was found to be highly effective against the isolates of Fol. followed by A. niger biocontrol potential of A. terreus is least among all the isolates tested. Culture filtrates obtained from A. luchuensis exerted least inhibition of Fol. The most sensitive isolate of Fol. against all the antagonists tested was identified as IIVR-2 (Fol. 9). Inherent diversity among Fol. isolates, from different tomato growing regions in India, was determined using RAPD primers. The genetic similarity coefficients ranged from 0.20 to 0.96, indicating that no any two or more isolates were 100% similar. RAPD profiles revealed up to 20% genetic diversity among ten isolates of Fusarium oxysporum f. sp. lycopersici.     

Molecular diversity of native Trichoderma isolates against Fusarium oxysporum f. sp. lycopersici (Sacc.). A casual agent of Fusarium wilt in tomato (Lycopersicon esculentum Mill.)

Fusarium wilt in tomato caused by Fusarium oxysporum is the one of the problematic diseases. In this study, 12 native Trichoderma isolates were isolated from different land use types in Rayalaseema region of Andhrapradesh, India and were tested for antagonistic activity against F. oxysporum using dual culture method; the maximum inhibition occurred in WT₂ (78.4%) compared to the control. Molecular characterisation using random amplified polymorphic DNA (RAPD) technique reported 91.8% polymorphism among 12 isolates of Trichoderma. Internal transcribed spacer (ITS) region of rDNA amplification with genus-specific ITS1 and ITS4 universal primers produced amplicon size from 569 bp in all the isolates. The study resulted in identification of good competitive Trichoderma isolates against F. oxysporum. A relationship was found between the polymorphism showed by the Trichoderma isolates and their hardness to F. oxysporum during antagonism. Also, exhibition of sufficient genetic polymorphism aids further exploitation in genomic fingerprinting.     

Pathogenicity of Fusarium oxysporum to Easter lily, narcissus and gladiolus

Isolates of Fusarium oxysporum from roots, bulbs and stems of Easter lilies (Lilium longiflorum) differed widely in pathogenicity and also, apparently, in tissue specificity. Virulent isolates caused a typical basal rot and root rot (but not a wilt) in which the mycelium advanced intercellularly through the scales and basal plates. Mildly pathogenic isolates became established in mature or senescent outer scales, at first producing only superficial effects, but further growth of mycelium occurred as the outer scales died and sometimes continued until the dead tissues were permeated and chlamydospores were formed. The underlying scales were then colonized. The modes of pathogenicity and survival in Easter lily were compared with those of the F. oxysporum formae causing bulb rots of gladiolus and narcissus. It is suggested that advance of hyphae by penetration between the cells of the vascular parenchyma, which is common in isolates causing rots in bulbs and corms, represents a stage in the evolution of the truly vascular habit among fusaria.     

Genetic and virulence variation in Fusarium oxysporum f. sp. cepae causing root and basal rot of common onion in Iran

Root and basal rot of common onion (Allium cepae L.) caused by Fusarium oxysporum f. sp. cepae is one of the most important diseases causing tremendous losses in onion‐growing areas worldwide. In this study, random amplified polymorphic DNA (RAPD), intersimple sequence repeats (ISSR) and virulence studies were conducted to analyse 26 F. oxysporum f. sp. cepae isolates obtained from the main onion‐growing regions of Iran, including Fars, Azerbaijan and Isfahan states. Cluster analysis using UPGMA method for both RAPD and ISSR markers revealed no clear grouping of the isolates obtained from different geographical regions, and the isolates were observed to derive probably from the same clonal lineage. Pathogenicity test indicated that all F. oxysporum f. sp. cepae isolates were pathogenic on onion; however, virulence variability was observed among the isolates. The grouping based on virulence variability was not correlated with the results of RAPD and ISSR analyses.     

Genetic Diversity and Pathogenicity of Fusarium oxysporum f.sp. melonis Races from Different Areas of Italy

Fusarium wilt caused by Fusarium oxysporum f.sp. melonis (FOM) is a devastating disease of melon worldwide. Pathogenicity tests performed with F. oxysporum isolates obtained from Italian melon‐growing areas allowed to identify thirty‐four FOM isolates and the presence of all four races. The aims of this work were to examine genetic relatedness among FOM isolates by race determination and to perform phylogenetic analyses of identified FOM races including also other formae speciales of F. oxysporum of cucurbits. Results showed that FOM race 1,2 was the most numerous with a total of eighteen isolates, while six and nine isolates were identified as race 0 and 1, respectively, and just one isolate was assigned to race 2. Phylogenetic analysis was performed by random amplified polymorphic DNA (RAPD) profiling and by translation elongation factor‐1α (TEF‐1α) sequencing. The analysis of RAPD profiles separated FOM races into two distinct clades. Clade 1, which included races 0, 1 and 1,2, was further divided into ‘subclade a’ which grouped almost all race 1,2 isolates, and into ‘subclade b’ which included race 0 and 1 isolates. Clade 2 comprised only race 2 isolates. The phylogenetic analysis based on TEF‐1α separated FOM from the other formae speciales of F. oxysporum. Also with TEF‐1α analysis, FOM races 0, 1 and 1,2 isolates grouped in one single clade clearly separated from FOM race 2 isolates which grouped closer to F. oxysporum f.sp. cucumerinum. RAPD technique was more effective than TEF‐1α in differentiating FOM race 1,2 isolates from those belonging to the closely related races 0 and 1. Both phylogenetic analyses supported the close relationship between the three different FOM races which might imply the derivation from one another and the different origin of FOM race 2.     

Presence of Fusarium oxysporum f. sp. melonis Race 1 in Soils Cultivated with Melon in the State of Colima (Mexico)

During years 2001, 2002 and 2003 the gravity of the Fusarium wilt in 1000 hectares of melon culture was evaluated in Colima (Mexico). In spite of the soil disinfections with methyl bromide, the losses could reach 25% of the final production. The analysis of 4 soil samples from the fields with ill plants, in a selective medium for Fusarium, allowed to detect the presence of F. oxysporum. By means of the presented technique “soil phytopathometry”, 31 isolates of F. oxysporum f. sp. melonis were obtained from the soil samples. The isolates were inoculated on melon plants to evaluate their pathogenicity. The 31 isolates inoculated, produced the symptoms of chlorosis and wilting, in melon cultivars that allowed us to affirm that all isolates were race 1 of F. oxysporum f. sp. melonis. Being this the first news of the presence of F. oxysporum f. sp. melonis in the state of Colima (Mexico).     

Antagonistic activity of bacteria isolated from crops cultivated in a rotation system and a monoculture against <Emphasis Type="Italic">Pythium debaryanum</Emphasis> and <Emphasis Type="Italic">Fusarium oxysporum</Emphasis>

The effect of crop rotation and monocropping on the occurrence of bacteria with antagonistic activity toward Pythium debaryanum and Fusarium oxysporum was shown. Arthrobacter spp., fluorescent Pseudomonas spp. and actinomycetes were isolated from winter rape, sugar beet and winter barley rhizosphere and bulk soil from the plots of a long-term crop rotation experiment (18 years). The occurrence of mycoantagonistic isolates and their antibiosis level exhibited specificity for the site, crop and crop rotation. Mycoantagonistic activity was common among actinomycetes and fluorescent Pseudomonas spp. and less frequent among Arthrobacter spp. Antibiosis of fluorescent Pseudomonas spp. and Arthrobacter spp. was in general stronger against P. debaryanum than F. oxysporum. The highest percentage of antagonistic Pseudomonas spp. against P. debaryanum was in the plots of barley crop, while plots of winter rape showed higher frequency of antagonists against F. oxysporum. The highest antibiosis activity of Arthrobacter spp. against both pathogens occurred in isolates from barley and winter rape monoculture, and there were no F. oxysporum antagonists among these bacteria in sugar beet monoculture. Most of actinomycete isolates strongly inhibited growth of P. debaryanum and F. oxysporum. The percentage of mycoantagonistic actinomycetes and their antibiosis level were the highest in the 6-year crop rotation system.