Pathogenic variability in Ethiopian isolates of Fusarium oxysporum f. sp. ciceris and reaction of chickpea improved varieties to the isolates

Twenty-four isolates of Fusarium oxysporum f. sp. ciceris were isolated from wilted chickpea plants obtained from different districts and ‘wilt sickplots’ of central Ethiopia to assess variability in pathogenecity of the populations. Each isolate was tested on 10 different chickpea lines and eight improved chickpea varieties. Isolates showed highly significant variation in wilt severity on the differential lines and improved varieties. Based on the reaction types induced on differential lines, isolates were grouped into four corresponding races. Of the 24 isolates, F13, F20 and F22 were the most virulent. Isolates of race 3 were found in all of the districts and ‘wilt sickplots’ studied. Improved chickpea varieties also showed differential reactions to the isolates. All varieties were resistant to isolates of race 3, while varieties Arerti and DZ-10-4 were resistant to all isolates tested, showing the lowest mean wilt severity. Varieties DZ-10-11 and Maryie were susceptible to isolates F13, F20 and F22 and showed the highest mean wilt severity. Identification of races can be useful in breeding chickpea varieties resistant to wilt. The differential reactions of the improved varieties against different races might be important in managing chickpea wilt through gene deployment.     

Plant defence reactions against fusarium wilt in chickpea induced by incompatible race 0 of Fusarium oxysporum f.sp. ciceris and nonhost isolates of F. oxysporum

Germinated seeds of 'kabuli' chickpea cv. ICCV 4 were inoculated with a conidial suspension of the incompatible race 0 of Fusarium oxysporum f.sp. ciceris (Foc) or of nonhost F. oxysporum resistance 'inducers', and 3 days later were challenged by root dip with a conidial suspension of highly virulent Foc race 5. Prior inoculation with inducers delayed the onset of symptoms and/or significantly reduced the final amount of fusarium wilt caused by race 5. However, the extent of disease suppression varied with the nature of the inducing agent; the nonhost isolates of F. oxysporum were more effective at disease suppression than the incompatible Foc race 0. Inoculation with the inducers gave rise to synthesis of maackiain and medicarpin phytoalexins in inoculated seedlings; these did not accumulate in plant tissues but were released into the inoculum suspension. Inoculation with inducers also resulted in accumulation of chitinase, β-1,3-glucanase and peroxidase activities in plant roots. These defence-related responses were induced more consistently and intensely by nonhost isolates of F. oxysporum than by incompatible Foc race 0. The phytoalexins and, to a lesser extent, the antifungal hydrolases, were also induced after challenge inoculation with Foc race 5. However, in this case the defence responses were induced in both preinduced and noninduced plants infected by the pathogen. It is concluded that the suppression of fusarium wilt in this study possibly involved an inhibitory effect on the pathogen of preinduced plant defences, rather than an increase in the expression of defence mechanisms of preinduced plants following a subsequent challenge inoculation.     

Genetic variation and population structure of Fusarium oxysporum f.sp. vasinfectum in Australia

Genetic variation among 348 isolates of Fusarium oxysporum f.sp. vasinfectum (Fov) collected from diseased cotton plants in 31 fields in six cotton-growing regions in New South Wales and Queensland in 2002 and 2004 was analysed using amplified fragment length polymorphisms (AFLPs). Twenty-eight haplotypes were identified based on 146 polymorphic bands generated with four Eco RI and Mse I and four Hind III and Mse I primer combinations. The haplotypes separated into two distinct groups (37% similarity), with 21 in group I and seven in group II. The two unique vegetative compatibility groups of Fov known to occur in Australia (VCG 01111 and VCG 01112) were correlated to the two AFLP groups, with both VCG 01111 reference isolates being included in group I and both VCG 01112 reference isolates in group II. Group I was widespread, occurring in all regions sampled and all but one of the fields, while group II was limited to three fields in the Boggabilla region. Group I was further divided into two subgroups. The two haplotypes in subgroup I-B (I-20 and I-21) may represent the emergence of a new form of Fov based on their marked genetic discrimination from the subgroup I-A haplotypes. No spatial population differentiation was discernible at the national level, as only 3·9% of total genetic variation was attributed to differences among regions ( P =  0·4868). When each region was analysed separately, clear differentiation was found in the Boggabilla region, with 86·3% of total genetic variation resulting from differences among fields ( P <  0·0001).     

Pathogenicity and race characterization of Fusarium oxysporum f. sp. phaseoli isolates from Spain and Greece

Virulence (≡ severity of disease) and physiological specialization of nine isolates of Fusarium oxysporum f. sp. phaseoli recovered in El Barco de Avila (Castilla y León, west-central Spain) and of two isolates from Chryssoupolis (Greece) were determined. The susceptibility/resistance response showed by a differential set of common bean cultivars ( Phaseolus vulgaris ) selected at the Centro Internacional de Agricultura Tropical (CIAT) delineated the isolates into two new races: races 6 and 7. The results of pathogenicity tests did not show any significant differences in virulence among the isolates. However, the reactions of several Spanish common bean cultivars indicated the presence of two groups of isolates, highly virulent and weakly virulent, among the Spanish isolates analysed. These results indicate that isolates classified in the same race are not homogeneous with respect to virulence, and suggests that race analysis using the CIAT differential cultivars is insufficient to describe the physiological specialization of F. oxysporum f. sp. phaseoli .     

Variation in the response of melon genotypes to Fusarium oxysporum f.sp. melonis race 1 determined by inoculation tests and molecular markers

Screening of genotypes of melon ( Cucumis melo ) for resistance to wilt caused by Fusarium oxysporum f.sp. melonis is often characterized by wide variability in their responses to inoculation, even under carefully controlled conditions. The variability at the seedling stage of 17 genotypes susceptible to race 1 was examined in growth-chamber experiments. Disease incidence varied from 0 to 100% in a genotype-dependent manner. Using four combinations of light (60 and 90  µ E m⁻² s⁻¹) and temperatures of (27 and 31°C), only light intensity showed a statistically significant effect. Marker-assisted selection for fusarium resistance breeding using cleaved amplified polymorphic sequence (CAPS) and sequence-characterized amplified region (SCAR) markers were compared using a single set of genotypes that included 24 melon accessions and breeding lines whose genotype regarding the Fom-2 gene was well characterized. The practical value of the markers for discriminating a range of genotypes and clarifying the scoring of phenotypes was also tested using a segregating breeding population which showed codominant SCAR markers to be useful in marker-assisted selection.     

A new race of Fusarium oxysporum f. sp. lactucae of lettuce

Fusarium oxysporum f. sp. lactucae, the causal agent of fusarium wilt of lettuce (Lactuca sativa), occurs in most countries in which lettuce is grown and causes serious economic losses. Three races (1, 2 and 3) of the pathogen have previously been identified on the basis of their ability to cause disease on differential lettuce cultivars, as well as by means of molecular tools developed to characterize different races of this pathogen. Only race 1 has been detected in Europe so far. In this study, two isolates of F. oxysporum, obtained from lettuce plants grown in the Netherlands showing symptoms of wilt, have been characterized by combining the study of pathogenicity with differential cultivars of lettuce and molecular assays to determine whether the isolates are different from the known races of F. oxysporum f. sp. lactucae. This study reports the presence of F. oxysporum f. sp. lactucae for the first time in the Netherlands. The causal pathogen has been identified, using the IRAP‐SCAR technique, as a new race of F. oxysporum f. sp. lactucae. Specific primers have been designed to identify this new race.     

Genetic diversity in Fusarium oxysporum f.sp. dianthi and Fusarium redolens f.sp. dianthi

Pathogenic isolates were selected representing all known vegetative compatibility groups (VCGs) and races of Fusarium oxysporum sensu lato from Dianthus spp. On basis of differences in the internal transcribed spacer region of the ribosomal DNA, six VCGs were classified as F. oxysporum f.sp. dianthi and four as F. redolens f.sp. dianthi. All VCGs of F. oxysporum f.sp. dianthi were characterized by unique restriction fragment length polymorphisms (RFLPs), unique overall esterase profiles, and unique virulence spectra, supporting a clonal lineage concept. Two VCGs of F. oxysporum f.sp. dianthi nevertheless comprised more than one race, but races within the same VCG shared the same distinct overall virulence spectrum. VCGs belonging to F. redolens f.sp. dianthi also had unique RFLPs and unique virulence spectra, but had grossly identical esterase profiles. Three new races (9, 10 and 11) are described for F. oxysporum f.sp. dianthi, and four for F. redolens f.sp. dianthi. Two races previously considered lost were recovered; race 7 was identified as a member of VCG 0021 of F. oxysporum f.sp. dianthi while race 3 was identified as a distinct VCG and race of F. redolens f.sp. dianthi. A summary of races and VCGs in F. oxysporum f.sp. dianthi and F. redolens f.sp. dianthi is presented.     

Influence of Inoculum Density of Races 0 and 5 of Fusarium oxysporum f. sp. ciceris on Development of Fusarium Wilt in Chickpea Cultivars

Artificial inoculation experiments were carried out at 25°C to determine the effects of inoculum density of Fusarium oxysporum f.sp. ciceris races 0 (Foc-0) and 5 (Foc-5) and susceptibility of chickpea cultivars P-2245 and PV-61 on development of Fusarium wilt. Foc-5 proved much more virulent than Foc-0. Increasing the inoculum density of F. oxysporum f.sp. ciceris caused an exponential reduction in disease incubation period and a monomolecular increase of disease incidence and the area under the disease intensity progress curve. The extent of these effects was highest in the most conducive P-2245/Foc-5 combination and decreased in the less susceptible PV-61 and for the less virulent Foc-0, in that order. For P-2245/Foc-5, the highest disease intensity was attained with 6 chlamydospores g^–1 of soil, the lowest inoculum density in the study. One thousand chlamydospores g^–1 of soil of the same race were needed to attain a comparable disease intensity in PV-61. Twenty thousand chlamydospores g^–1 of soil of Foc-0 were required for maximum disease intensity in P-2245.The disease intensity curves were adequately described by the Gompertz model. Using this model, a response surface for disease intensity was developed, in which the model parameters are expressed as a function of both time from inoculation and inoculum density. This response surface confirmed that the final amount of disease intensity increases in a monomolecular relationship with increasing inoculum density and showed that the relative rate of disease progress increases exponentially with increasing inoculum density of the pathogen.     

Influence of chickpea genotype and Bacillus sp. on protection from Fusarium wilt by seed treatment with nonpathogenic Fusarium oxysporum

Seeds of kabuli chickpea cultivars ICCV 4 and PV 61 were treated with conidia of nonpathogenic Fusarium oxysporum isolate Fo 90105 suspended in methylcellulose (3 × 10⁶ conidia.seed^-1), or with methylcellulose alone, and sown in soil artificially infested with 500 or 1,000 chlamydospores.g^-1 of F. oxysporum f. sp. ciceris race 5. At an inoculum concentration of 500 chlamydospores.g^-1, seed treatment with Fo 90105 significantly increased the incubation period of the disease by 11 (ICCV 4) or 25 (PV 61) days, and reduced the final disease incidence, disease intensity and the standardized area under the curve of disease intensity over time. This protection from disease was higher and more consistent in PV 61 than in ICCV 4. However, it was annulled with an inoculum concentration of 1,000 chlamydospores.g^-1, except for the incubation period in PV 61 which was increased by 10 days. When ICCV 4 seeds were treated with Fo 90105 (3 × 10⁶ conidia.seed^-1) and/or Bacillus sp. isolate RGAF 51 (1 × 10⁷ cfu.seed^-1), then sown in infested soil, there was no influence by the Bacillus isolate on protection conferred by Fo 90105. However, the degree of protection by the nonpathogenic F. oxysporum was higher and more consistent when plants from treated seeds were grown in sterile sand for 6 days, then transplanted into infested soil.     

香蕉枯萎病菌RAPD分析及4号生理小种的快速检测

 用随机扩增多态性DNA(RAPD)技术,对采自广东、广西的香蕉和粉蕉上的30个香蕉枯萎病菌(Fusarium oxysporum f.sp.cubense)菌株和3个其它尖孢镰刀菌专化型的菌株进行比较及聚类分析。在遗传相似系数0.67时,可将供试菌株划分为3个RAPD群(RGs),其中香蕉枯萎病菌4号生理小种(FOC4)共15个菌株属于RGⅠ,1号生理小种(FOC1)共15个菌株属于RGⅡ,供试的其它尖孢镰刀菌专化型的3个菌株则属于RGⅢ。这说明香蕉枯萎病菌和供试3个其它专化型菌株与致病性间存在明显的相关性。1号生理小种内菌株间的遗传分化大于4号生理小种内菌株间的遗传分化。从90条RAPD随机引物中筛选出2条引物可产生4号生理小种的RAPD标记2个。将这2个RAPD标记电泳切胶回收、克隆及测序,并根据这2个特异片段序列设计SCAR上下游特异引物,通过对30个菌株的PCR扩增检验,其中一个RAPD标记成功地转化为SCAR标记,初步建立了以此为基础的4号生理小种快速检测技术,其检测灵敏度为2 ng新鲜菌丝。对采自不同地区的显症样品、吸芽、室内接种未显症的香蕉苗以及发病的香蕉植株不同部位进行检测,能够准确灵敏地鉴定出4号生理小种,从而为香蕉枯萎病菌的快速检测及防治奠定了基础。同时,快速检测结果发现,田间发病植株果柄的各部位及果实内并没有枯萎病菌的存在。     

Resistance sources to Fusarium oxysporum f. sp. cubense tropical race 4 in banana wild relatives

Target trait evaluation in crop wild relatives is an important prerequisite for efficiently using the potential useful genes located in this valuable germplasm. Over recent decades, Fusarium oxysporum f. sp. cubense tropical race 4 (Foc‐TR4) has seriously threatened worldwide banana plantations. Breeding new resistant cultivars from wild banana species is expected to provide invaluable additional resources. However, knowledge on resistance to Foc‐TR4 in wild Musa species is very limited. In this study, eight genotypes of wild banana relatives (Musa acuminata subsp. burmannica, M. balbisiana, M. basjoo, M. itinerans, M. nagensium, M. ruiliensis, M. velutina and M. yunnanensis) were characterized for resistance to Foc‐TR4 in both greenhouse and field conditions. Most wild bananas showed higher resistance levels to Foc‐TR4 than the reference cultivars ‘Brazilian’ (AAA, susceptible) and ‘Goldfinger’ (AAAB, moderate resistance). Among the wild species, M. balbisiana showed the highest levels of disease intensity followed by M. acuminata subsp. burmannica. Some individuals of M. yunnanensis, M. nagensium, M. ruiliensis and M. velutina showed low levels of rhizome discolouration in greenhouse conditions, but were resistant in the field. No symptoms were observed on M. basjoo and M. itinerans, suggesting higher levels of resistance to Foc‐TR4. The results revealed different sources of resistance to Foc‐TR4 in banana wild relatives, which constitute a valuable genetic resource for banana breeding programmes aiming to produce cultivars resistant to fusarium wilt.     

主要环境及营养因素对西瓜枯萎病菌生长的影响

采用菌丝生长速率法和产孢量测定研究了不同温度、光照、pH及碳氮源对来源于海南、河北和上海的3株西瓜枯萎病菌Fo-HN-46、Fo-HB-12和Fo-SH-1生长速率和产孢量的影响。结果表明:西瓜枯萎病菌菌株的适宜生长温度为20~30℃,Fo-HN-46最适生长温度为28℃,Fo-HB-12和Fo-SH-1最适生长温度为25℃,产孢的最适温度为28~30℃;光照对西瓜枯萎病菌生长速率无显著影响,对产孢影响显著,Fo-HN-46在黑暗条件下单位面积产孢量最高,Fo-HB-12和Fo-SH-1在半光照条件下产孢量最高;pH对西瓜枯萎病菌菌株的生长速率和产孢量有显著影响,pH 7~9时菌丝生长速率快,pH 8~11时产孢量高;葡萄糖、淀粉和乳糖作为碳源能够显著增加西瓜枯萎病菌的产孢量;蛋白胨作为氮源能显著促进菌丝生长,硝酸钠、酵母粉和蛋白胨均能显著增加西瓜枯萎病菌的产孢量。用最适碳源、氮源培养,Fo-HN-46生长速率大于Fo-HB-12和Fo-SH-1。Fo-SH-1产孢量高于Fo-HN-46和Fo-HB-12。     

Characterization of Fusarium oxysporum f. sp. radicis-cucumerinum attacking melon under natural conditions in Greece

A severe root and stem rot disease of melon was observed during the 2001 growing season on four glasshouse crops in Heraklio, Greece. A total of 43 isolates of F. oxysporum , obtained in Crete from glasshouse-grown melon and showing fusarium wilt or root and stem rot symptoms, were characterized by pathogenicity and vegetative compatibility. The majority of these isolates was also fingerprinted via amplified fragment length polymorphic (AFLP) analysis. Of the total number of isolates, 22 were identified by pathogenicity tests as F. oxysporum f. sp. melonis , 20 as F. oxysporum f. sp. radicis-cucumerinum , while one isolate was nonpathogenic on cucumber, melon, sponge gourd and pumpkin. All 22 isolates of F. oxysporum f. sp. melonis were assigned to vegetative compatibility group (VCG) 0134, and all 20 isolates of F. oxysporum f. sp. radicis-cucumerinum to VCG 0260. Isolates of F. oxysporum f. sp. radicis-cucumerinum were incompatible with isolates of F. oxysporum f. sp. melonis. AFLP fingerprinting allowed for the clustering of the isolates of the two formae speciales of F. oxysporum along two separate phenetic groups: f. sp. melonis to AFLP major haplotype I, and f. sp. radicis-cucumerinum to AFLP major haplotype II. Overall, pathogenicity, vegetative compatibility grouping and AFLP analysis were correlated and effectively distinguished isolates of F. oxysporum from melon. This appears to be the first report of natural infection of melon by F. oxysporum f. sp. radicis-cucumerinum worldwide.     

Interactions of Pratylenchus thornei and Fusarium oxysporum f. sp. ciceris on Chickpea

ABSTRACT Fusarium oxysporum f. sp. ciceris and the root-lesion nematode Pratylenchus thornei coinfect chickpeas in southern Spain. The influence of root infection by P. thornei on the reaction of Fusarium wilt-susceptible (CPS 1 and PV 61) and wilt-resistant (UC 27) chickpea cultivars to F. oxysporum f. sp. ciceris race 5 was investigated under controlled and field conditions. Severity of Fusarium wilt was not modified by coinfection of chickpeas by P. thornei and F. oxysporum f. sp. ciceris, in simultaneous or sequential inoculations with the pathogens. Root infection with five nematodes per cm(3) of soil and 5,000 chlamydospores per g of soil of the fungus resulted in significantly higher numbers of propagules of F. oxysporum f. sp. ciceris with the wilt-susceptible cultivar CPS 1, but not with the wilt-resistant one. However, infection with 10 nematodes per cm(3) of soil significantly increased root infection by F. oxysporum f. sp. ciceris in both cultivars, irrespective of fungal inoculum densities (250 to 2,000 chlamydospores per g of soil). Plant growth was significantly reduced by P. thornei infection on wilt-susceptible and wilt-resistant chickpeas in controlled and field conditions, except when shorter periods of incubation (45 days after inoculation) were used under controlled conditions. Severity of root necrosis was greater in wilt-susceptible and wilt-resistant cultivars when nematodes were present in the root, irrespective of length of incubation time (45 to 90 days), densities of nematodes (5 and 10 nematodes per cm(3) of soil), fungal inocula, and experimental conditions. Nematode reproduction on the wilt-susceptible cultivars, but not on the wilt-resistant one, was significantly increased by F. oxysporum f. sp. ciceris infections under controlled and field conditions.     

Vascular colonization patterns in susceptible and resistant tomato cultivars inoculated with Fusarium oxysporum f.sp. lycopersici races 0 and 1

The vascular colonization pattern of Fusarium oxysporum f.sp. lycopersici races 0 and 1 in tomato was studied in five susceptible and five resistant cultivar–fungus combinations during a 26-day period after inoculation by root immersion. Propagules spread discontinuously along the stems in all five cultivars 1 day after inoculation, irrespective of cultivar resistance. Five days later the fungus was limited to the stem bases in all cultivars. Between the fifth and 12th days, stem colonization by the fungus stopped in all cultivar–race combinations. Thereafter, the situation remained stable in resistant combinations, with inoculum distributed discontinuously, and no disease symptoms were apparent. By contrast, in the susceptible combinations a gradual upward colonization of the stems was seen such that fungal distribution was no longer discontinuous and disease symptoms appeared. These results suggest that a fungal 'incubation' period in the base of the vascular system is required before a secondary invasion of tissues occurs in susceptible genotypes. The slope of the regression line fitted between the height reached by the fungus up the stem ( y ) and the time after inoculation ( x ) provides a measure of the horizontal (polygenic) resistance in tomato cultivars     

香蕉枯萎病菌1号和4号生理小种细胞壁降解酶的比较

对香蕉枯萎病菌1号和4号生理小种的细胞壁降解酶进行比较。通过测定4号生理小种在寄主体内细胞壁降解酶的活性发现,能检测到多聚半乳糖醛酸酶(PG)、果胶甲基半乳糖醛酸酶(PMG)、多聚半乳糖醛酸反式消除酶(PGTE)、果胶甲基反式消除酶(PMTE)和纤维素酶(Cx)的活性。在不同碳源培养条件下,2个生理小种均有以上5种酶的活性,以1%柑桔果胶为碳源时产生的PMG和PG活性明显高于其他几种酶的活性,而以1%CMC为碳源时,所产生的Cx都比其他几种酶的活性高。细胞壁降解酶同工酶电泳后发现,4号生理小种在寄主体内和体外培养时都比1号生理小种多分泌一种PG。2个生理小种在体外培养时分泌的PMG、PGTE和PMTE没有差异。4号生理小种在寄主体内比1号生理小种多分泌一种PMG,却少分泌一种PGTE,2个生理小种在寄主体内的PMTE则没有差异。     

Phylogeny and origin of Fusarium oxysporum f. sp. vanillae in Indonesia

Vanilla stem rot, caused by Fusarium oxysporum f. sp. vanillae (Fov), is the main constraint to increasing vanilla production in the major vanilla‐producing countries, including Indonesia. The current study investigated the origin of Fov in Indonesia using a multigene phylogenetic approach. Nineteen Fov isolates were selected to represent Indonesia, the Comoros, Mexico and Réunion Island. The translation elongation factor 1 alpha gene and the mitochondrial small subunit ribosomal RNA gene phylogenies resolved the Fov isolates into three distinct clades in both phylogenetic species of the F. oxysporum species complex, indicating a polyphyletic pattern of evolution. In addition, Fov isolates from Indonesia were also polyphyletic. These results suggest that the vanilla stem rot pathogen in Indonesia has a complex origin. The implications for disease management are discussed.     

Development of a Fusarium oxysporum f. sp. melonis functional GFP fluorescence tool to assist melon resistance breeding programmes

Fusarium wilt, caused by Fusarium oxysporum f. sp. melonis (Fom), is one of the most widespread and devastating melon diseases. This vascular disease is caused by the colonization of melon xylem vessels by any of the four Fom races reported (r0, r1, r2 and r1,2, subdivided into r1,2w and r1,2y). The macroscopic evaluation of disease symptoms (disease rating, DR) at several days post‐inoculation (dpi) with Fom spores has been the traditional method to determine the resistance of melon accessions to this fungal pathogen. In this study, one isolate from each Fom race was transformed by Agrobacterium tumefaciens to constitutively express the green fluorescent protein (GFP). Fom‐GFP transformants, as virulent as the corresponding wildtype races, were selected to develop an inoculation assay based on the non‐invasive evaluation of the fluorescence emitted by Fom‐GFP. It was determined that melon root neck was the appropriate area to follow Fom‐GFP and a fluorescence signal rating (FSR) was established in parallel to DR determination. This method allowed the evaluation of GFP signal in the root neck of inoculated melon seedlings at 11–15 dpi. The GFP signal was scored in 62 melon accessions/breeding lines inoculated with different Fom‐GFP, followed by evaluation of the macroscopic DR in the aerial part of melon seedlings at 20–28 dpi. Correlation analysis demonstrated a direct and significant relationship between FSR and DR. This method has shown to be an effective and reliable tool that can assist Fom resistance breeding programmes in melon.     

Vegetative compatibility and heterokaryon stability in Fusarium oxysporum f.sp. radicis-lycopersici from Italy

Fusarium crown and root rot, caused by Fusarium oxysporum f.sp. radicis-lycopersici ( Forl ), is one of the most destructive soilborne diseases of tomato in Italy. Chlorate-resistant, nitrate-nonutilizing ( nit ) mutants were used to determine vegetative compatibility among 191 isolates of Forl collected in five geographic regions (Calabria, Emilia-Romagna, Liguria, Sardinia, Sicily) in Italy. The isolates were assigned to five vegetative compatibility groups (VCGs): 65 isolates to VCG 0090; 99 to VCG 0091; 23 to VCG 0092; two to VCG 0093; and two to VCG 0096. The population structure of Forl in Italy is similar to that reported for Israel, and differs from that found in North Atlantic European countries, where VCG 0094 is predominant. The stability of prototrophic heterokaryons originating from hyphal anastomosis between compatible complementary nit mutants was assessed through conidial analysis and mycelial mass transfer. Most monoconidial cultures (84%) recovered from 117 prototrophic heterokaryons were nit mutants, indicating that heterokaryons generally do not proliferate well through conidiation; most of the 177 prototrophic heterokaryons examined were unstable, and only 9% sustained prototrophic growth through the tenth mycelial transfer upon subculturing. The prototrophic growth is proposed to be maintained through restoration of the heterokaryotic state by continual anastomosis between adjacent homokaryotic hyphae. Since heterokaryosis is a prerequisite for parasexual recombination, we speculate that this mechanism is unlikely to play a major role in generating the VCG diversity found among Forl or other strains of F. oxysporum.