Differential regulation of MBP kinases by a glycoproptein elicitor and a polypeptide suppressor from Mycosphaerella pinodes in pea

A polypeptide fungal suppressor from a pea pathogen Mycosphaerella pinodes plays a key role in pathogenesis by suppressing elicitor-induced defense response(s) in pea (Pisum sativum L). In this study, we show that treatment of pea tissues with the polysaccharide elicitor secreted by M. pinodes results in rapid increased activation of two myelin basic protein (MBP)-dependent kinases p44 (≈44 kDa) and p48 (≈48 kDa) within 15–30 min upon elicitation. Interestingly, the suppressor inhibited the elicitor-induced activation of only p44 kinase. While the defense-inducing signalling molecules, chitosan and salicylic acid (SA) activated the p44 and p48 kinases, methyl jasmonate (MeJA) did not. The abiotic stress signals, abscisic acid (ABA), NaCl and wounding activated the p48 kinase alone. These results demonstrate that MAPKs are differentially activated in response to pathogen invasion and abiotic stress in pea. Furthermore, specific inhibition of elicitor-induced p44 kinase activation by a MAPKK inhibitor, PD098059 and protein kinase inhibitor, K252a correlated with the suppression of elicitor-induced phenylalanine ammonia lyase (PAL) gene expression, supporting a role for p44 in the elicitor-induced defense response(s) in pea. Inhibition of p44 by the phosphoinositide (PI) turnover inhibitor, neomycin (a fungal suppressor mimic), and potentiation of p44 by the diacylglycerol (DAG) kinase inhibitor, R59022 indicated that p44 may be acting downstream of (PI) metabolism. Taken together, our results indicate that suppressor of defense elicitation from M. pinodes acts through inhibition of a MAPK (p44), possibly through a PI signaling pathway, facilitating the establishment of basic compatibility during infection of pea.     

SHOOT ZONE UPTAKE OF SOIL-APPLIED HERBICIDES*

Summary. Studies were conducted to determine the effects of herbicide placement at different zones of maize (Zea mays L.) and pea (Pisum sativwm L.) shoots below the soil surface after emergence. Soil was removed from around the shoots and replaced with herbicide-treated soil. A wax barrier ensured separate exposure of the zones to treated soil. EPTC, chlorpropham, propham and sulfallate did not affect pea shoot growth, but in maize the shoot zone adjacent to the crown root node was extremely sensitive. Treatment in this area markedly reduced growth and severely inhibited the crown roots. The difference in susceptibility between these species may he due to the location of the growing point relative to the treated soil. Shoots of maize and pea were sensitive to diuron. In maize the shoot adjacent to the crown root node and the tissue of the first internode were the most susceptible. In pea the- uppermost shoot (beneath the soil surface) was the most sensitive. Trifluralin did not affect growth of maize and pea when placed in the shoot zone after emergence, although the crown roots of maize were severely inhibited. Naptalam, dalapon and 2,4-D did not affect growth of maize under similar conditions, and of these only 2,4-D reduced growth of pea. Zone d'abiorption des tiges pour les herbicides appliqués sur h sol     

不同秸秆发酵还田对制种玉米田土壤肥力质量和玉米品质的影响

在甘肃省张掖市甘州区甘浚镇巴吉村连续15 a种植制种玉米基地上,进行了小麦、玉米、蚕豆、豌豆、油菜籽和葵花秸秆发酵还田对制种玉米田土壤肥力质量和玉米品质影响的研究。结果表明：6种发酵秸秆还田后制种玉米田0～20 cm土层容重排序为：小麦秸秆＜玉米秸秆＜豌豆秸秆＜葵花秸秆＜油菜籽秸秆＜蚕豆秸秆,孔隙度、水稳性团聚体和总持水量排序为：小麦秸秆＞玉米秸秆＞豌豆秸秆＞葵花秸秆＞油菜籽秸秆＞蚕豆秸秆。制种玉米田pH值排序为：小麦秸秆＜玉米秸秆＜葵花秸秆＜油菜籽秸秆＜蚕豆秸秆＜豌豆秸秆,CEC、有机质、速效氮磷钾、微生物数量、酶活性和制种玉米产量排序为：小麦秸秆＞玉米秸秆＞葵花秸秆＞油菜籽秸秆＞蚕豆秸秆＞豌豆秸秆。小麦秸秆与玉米秸秆、葵花秸秆、油菜籽秸秆、蚕豆秸秆和豌豆秸秆比较,碱解氮分别增加0.87%、3.51%、6.23%、8.07%和9.93%,速效磷分别增加2.20%、2.51%、6.45%、6.67%和7.57% ,速效钾分别增加1.98%、3.12%、4.59%、6.12%和6.40%,细菌数量分别增加3.66%、4.70%、11.03%、14.29%和19.54%,放线菌数量分别增加7.28%、8.87%、17.55%、27.01%和36.42%,蔗糖酶活性分别提高0.74%、3.70%、6.56% 、12.16%和13.41%,脲酶活性分别提高6.59% 、10.23%、16.16%、25.97%和30.20%, 磷酸酶活性分别提高3.67% 、5.16% 、7.62% 、8.26%和10.88%, 多酚氧化酶活性分别提高4.35% 、9.09%、30.91%、41.18%和50.00%,制种玉米产量分别增加2.03% 、3.17%、5.33%、6.36%和8.49%。 制种玉米可溶性糖、淀粉和粗蛋白含量排序为：小麦秸秆＞玉米秸秆＞油菜籽秸秆＞葵花秸秆＞豌豆秸秆＞蚕豆秸秆。小麦秸秆与玉米秸秆、油菜籽秸秆、葵花秸秆、豌豆秸秆和蚕豆秸秆比较,可溶性糖含量分别增加2.02%、7.75%、13.25%、18.52%和23.10%,淀粉含量分别增加2.05%、3.11%、6.28%、10.70%和14.13%,粗蛋白含量分别增加1.01%、5.11%、12.56%、13.41%和16.77%。施肥利润和肥料投资效率排序为：小麦秸秆＞玉米秸秆＞葵花秸秆＞油菜籽秸秆＞蚕豆秸秆＞豌豆秸秆。小麦秸秆与玉米秸秆、葵花秸秆、油菜籽秸秆、蚕豆秸秆和豌豆秸秆比较,制种玉米施肥利润分别增加 0.09、0.16、0.25 、0.31、0.40万元·hm^-2,肥料投资效率分别增加0.17、0.40、0.58 、0.77、0.97元·元^-1。因此,在玉米制种基地上推广小麦秸秆发酵还田,有利于解决土壤肥力质量下降、制种玉米产量和品质低而不稳的问题。     

Weed communities respond to changes in the diversity of crop sequence composition and double cropping

Agricultural intensification, besides increasing land productivity, also affects weed communities. We studied weed shifts in cropping sequences differing in the identity and number of crops grown. We also evaluated whether dissimilar weed communities in different cropping systems converge towards more similar communities, when the same sequence is cropped during 2 years. In three locations in the Rolling Pampa, Argentina, field experiments were conducted including five cropping systems in the first year (winter cereal/soyabean, field pea/soyabean, and field pea/maize double crops, and maize and soyabean as single crops), while the same sequence was grown in the following 2 years (wheat/soyabean double crop and maize). Changes in weed community composition and structure were analysed through multivariate analyses and frequency–species ranking plots. Weed communities differed first among sites, while weed shifts within each site were mainly associated with growing season and crop type. Differences among crop sequences were higher in the first year, mostly related to specific crop grown, rather than to the number of crops in the sequences. Differences were reduced when the same sequence was grown during two consecutive seasons. Frequency of highly common weeds was negatively associated with the number of days with high crop cover. Our findings contribute to understand weed shifts in consecutive growing seasons, which may help readapting crop sequences to reduce the occurrence of abundant weed species.     

Complete nucleotide sequence and latency of a novel blueberry-infecting closterovirus

A novel latent closterovirus was detected from highbush blueberry in Japan and provisionally named blueberry virus A (BVA). The BVA genome (17,798 nucleotides) contains 10 open reading frames, but no minor coat protein could be identified in the virus genome. The BVA RNA-dependent RNA polymerase (RdRp), heat shock protein 70 homolog (HSP70h), and major coat protein (CP) shared the highest amino acid sequence identities with those of viruses in the genus Closterovirus (61.2, 27.6, and 20.9 %, respectively). In a phylogenetic analysis of the RdRp, HSP70h, and CP, BVA did not cluster with any genus in the family Closteroviridae.     

The fungicide captan reduces nuclear DNA content in maize seedlings

Captan, a major fungicide used by the US maize industry, was studied to determine its effect on the nuclear DNA content of maize (Zea mays L.) seedlings. Captan was applied to kernels at the recommended agronomic rates. The nuclear DNA content of the seedlings was determined by flow cytometry. In two maize hybrids examined, a reduction in nuclear DNA content was observed. Two different fluorochromes, with different staining mechanisms, were used to determine nuclear DNA content. Both of these fluorochromes revealed a reduction in nuclear DNA. Captan is reported to inhibit DNA synthesis in several animal systems. The reduction of maize nuclear DNA observed in this study may be a result of DNA synthesis inhibition induced by captan.     

The DNA damage signal transducer ortholog <Emphasis Type="Italic">Mop53BP1</Emphasis> is required for proper appressorium differentiation and pathogenicity in <Emphasis Type="Italic">Pyricularia oryzae</Emphasis>

Appressorium differentiation, one of the most important steps in pathogenesis by the rice blast fungus, Pyricularia oryzae, is strongly coordinated with the cell cycle. In this study, we identified an ortholog gene of 53BP1, which encodes a signal transducer protein that participates in G2-M cell cycle checkpoint in higher eukaryotes, in the genome of P. oryzae and characterized the phenotype of deletion and overexpression mutants. Deletion mutants showed no significant deficiency in vegetative growth compared to wild-type and complemented strains, even on the media containing DNA-damaging agents. However, these null mutants had abnormal appressoria and formed more appressoria per conidium than in the wild type and were unable to penetrate the epidermis of rice leaves. eGFP-fused Mop53BP1 and qRT-PCR analyses revealed that Mop53BP1 is expressed during the first hours of appressorium formation. In addition, in overexpression mutants, Mop53BP1 localized to nuclei during all stages of appressorium maturation and penetration, and the mutants were resistant to the microtubule inhibitor benomyl, suggesting that Mop53BP1 is nuclear protein and may have some role related to microtubules.     

根际分隔对玉米/豌豆间作种间竞争及豌豆结瘤固氮的影响

通过盆栽试验研究了玉米/豌豆间作条件下,两种施氮水平(0、0.15 g·kg~(-1))及三种分隔方式(不分隔、塑料膜分隔、尼龙网分隔)对玉米、豌豆生长及豌豆结瘤固氮的影响。结果表明:与单作相比,玉米/豌豆间作后对玉米的生长和养分的吸收有显著的促进作用,在整个生育期,玉米生物量的增加量随着豌豆生育期的推进而增加,豌豆苗期、豌豆结荚初期、豌豆收获期玉米的生物量在不施氮肥条件下分别增加:28.5%、32.8%、48.7%;在施氮0.15 g·kg~(-1)时分别增加-8.6%、8.1%、63.2%。在玉米豌豆间作体系中三种分隔方式玉米生物量的大小顺序为:间作不分隔间作尼龙网隔间作塑料膜分隔单作;在不施氮肥时,间作后豌豆苗期生物量与单作相比增加显著,增加幅度达35.8%;随着玉米生育期的推进,间作豌豆的生物量增加幅度随着降低,到豌豆收获期与单作豌豆相比差异不显著。收获时三种分隔方式豌豆生物量的大小顺序为:间作塑料膜分隔间作不分隔间作尼龙网隔单作;在施氮肥时,随着玉米的生长间作豌豆与单作豌豆相比显著减产,收获时三种分隔方式对豌豆生物量的大小顺序为:间作尼龙网隔间作塑料膜分隔单作间作不分隔。与单作相比,间作提高了豌豆的结瘤数;间作不分隔、间作尼龙网隔、间作塑料膜分隔条件下,碗豆的结瘤数分别增加120%、82.5%、22.5%。     

Lack of prevention of chlorsulfuron-induced inhibition by amino acids

The amino acids valine and isoleucine, which had been indicated by Ray (1984) as antidotes against growth inhibition of pea plants induced by chlorsulfuron, were ineffective in preventing the damage induced by chlorsulfuron on the plasma membrane of maize roots as measured by K⁺ uptake. In addition, in seedlings of two corn genotypes and three pea genotypes grown in culture solution containing chlorsulfuron, growth inhibition was not alleviated by valine and isoleucine. However, in agreement with Ray's results, acetolactate synthase, the key enzyme of the biosynthesis of valine and isoleucine, was inhibited in vitro by chlorsulfuron in both maize and pea. It is suggested that the chlorsulfuron mechanism of action involves several sites.     

Genetic and physiological evidence for the production of N-acyl homoserine lactones by Pseudomonas syringae pv. syringae and other fluorescent plant pathogenic Pseudomonas species

N-acyl homoserine lactones (AHLs) function as cell density (quorum) sensing signals and regulate diverse metabolic processes in several gram negative bacteria. We report that strains of Pseudomonas syringae pvs. syringae (Pss), tabaci and tomato as well as P. corrugata and P. savastanoi produce difussible AHLs that activate the lux operons of Vibrio fischeri or the tra::lacZ fusion of Agrobacterium tumefaciens. In Pss strain B3A, AHL production occurs in cell density dependent manner. Nucleotide sequence and genetic complementation data revealed the presence of ahlI_Pss, a luxI homolog within the Ahl⁺ DNA of Pss strain B3A. The DNA expresses in AHL-deficient strains of P. fluorescens and E. carotovora subsp. carotovora (Ecc), and restores extracellular enzyme production and pathogenicity in the Ecc strain. The derivatives of Pss strains B3A and 301D carrying chromosomal ahlI::lacZ do not produce AHL, but like their wild type parents, produce extracellular protease and the phytotoxin syringomycin as well as elicit the hypersensitive reaction in tobacco leaves. While these strains also produce a basal level of -galactosidase activity, the expression of ahlI::lacZ is substantially stimulated in the presence of multiple copies of the DNA or by the addition of cell-free spent cultures containing AHL. The activation of -galactosidase production occurs with spent cultures of some, but not all Pseudomonas strains which produce AHL as indicated by the Lux and tra::lacZ assays. Pss strains deficient in the global regulatory genes, gacA or lemA, produce very low levels of AHL. Since inactivation of ahlI_Pss eliminates AHL production and since Ahl⁺ Pseudomonas strains carry the homolog of ahlI_Pss, we conclude that ahlI_Pss specifies a key step in AHL biosynthesis and it has been conserved in many plant pathogenic pseudomonads.     

Partial Nucleotide Sequence and Genome Organization of a Canadian Isolate of Little cherry virus and Development of an Enzyme-Linked Immunosorbent Assay-Based Diagnostic Test

ABSTRACT Approximately 12.4 kb of the genome of a mealybug-transmissible, North American isolate of Little cherry virus (LChV-3, previously designated LChV-LC5) has been cloned and sequenced. The sequenced portion of the genome contains 10 open reading frames (ORFs) and, based on sequence comparisons, encodes a putative RNA helicase (HEL), RNA-dependent RNA polymerase (POL), two coat proteins (CPs), a homologue of HSP70, a 53K protein (p53) that is similar to an equivalent-size protein in other closteroviruses, and a 22K (p22) protein of unknown function. The genome also potentially encodes two small proteins (p5 and p6), one of which is similar to the small hydrophobic proteins of other closteroviruses. Phylogenetic analyses utilizing sequences of the HEL, POL, and HSP70 homologue suggest that LChV-3 is most similar to other mealybug-transmitted closteroviruses. Further comparisons between LChV-3 and a 4.7-kb region of the recently described Little cherry virus-2 (LChV-2) reveals 77% nucleotide sequence identity. Based on this low sequence identity, we propose that LChV-3 be considered a separate species, designated LChV-3. Unexpectedly, the LChV-3 CP duplicate ORF was found to lie upstream of the HSP70 ORF; therefore, the genome organization of LChV-3 is distinct from that of other closteroviruses. Polyclonal antiserum raised to bacterially expressed LChV-3 CP was useful for detection of LChV-diseased trees in the cherry-growing districts of British Columbia, Canada.     

Variation in the Fusarium section Liseola: pathogenicity and genetic studies of isolates of Fusarium moniliforme Sheldon from different hosts in Ghana

Fusarium moniliforme , the imperfect stage of the ascomycete Gibberella fujikuroi , is an economically important pathogen with a very wide host range. The genetic characteristics of isolates of the fungus collected from different regions of Ghana from maize, rice and sorghum were determined using the random amplified polymorphic DNA (RAPD) and restriction fragment length polymorphism (RFLP) techniques. The pathogenicity of the isolates was also compared on maize and rice. DNA fingerprints detected as RFLPs of ribosomal DNA and RAPDs separated the isolates into discrete groups which were generally host-related. The possibility of a sub-structuring of the maize population of the fungus into tissue-related subgroups was suggested by the results. A dendrogram of the relatedness of the isolates is presented. However, the pathogenicity of the isolates on rice, measured by their ability to cause 'bakanae' symptoms, did not resolve the isolates into the clearly defined groups suggested by the genetic studies, and maize isolates of the fungus could cause 'bakanae' symptoms to the same extent as rice isolates. Similarly, some isolates identified as rice-type isolates caused as much shoot stunting in maize as maize isolates. However, the effects of the isolates on root growth of maize seedlings showed a broad correlation with the defined genetic groups, with maize isolates of the fungus showing the greatest tendency to cause root stunting     

Studies on the mechanism of action of cymoxanil in Phytophthora infestans

Colony growth and germ tube emergence of sporangia and encysted zoospores of Phytophthora infestans were highly sensitive to cymoxanil (ED₅₀ 0.5–1.5 μg/ml), whereas differentiation of sporangia and zoospore release were insensitive at concentrations up to 100 μg/ml. Treated sporangia did not show distorted germ tubes. Oxygen consumption for glucose oxidation by germinating sporangia and zoospore motility were not inhibited at concentrations up to 100 μg/ml. Cymoxanil hardly affected the uptake of radiolabeled precursors of DNA, RNA, and protein at concentrations up to 100 μg/ml. Incorporation of [¹⁴C]phenylalanine into protein was completely insensitive. RNA synthesis as measured by [³H]uridine incorporation was differentially inhibited in the various developmental stages of the fungus. Inhibition did not occur at differentiation of sporangia, whereas at cyst and sporangial germination and mycelial growth this process was inhibited 20–45% at a concentration of 100 μg cymoxanil/ml. Endogenous RNA polymerase activity of isolated nuclei was not inhibited by cymoxanil. DNA synthesis as measured by [methyl-³H]thymidine incorporation was inhibited 20–80% at the various stages of development at cymoxanil concentrations higher than 10 μg/ml. Metalaxyl, a specific inhibitor of ribosomal RNA synthesis, inhibited [³H]uridine incorporation 40–60% at all developmental stages. The data suggest that although DNA synthesis is affected more than RNA synthesis, inhibition of both biosynthetic processes is a secondary effect. The primary mode of action of cymoxanil thus remains unknown.     

拟南芥BOI 基因在抗氧化胁迫和细胞死亡中的功能分析

 本文利用BOI(Botrytis Susceptible 1 Interactor)基因敲减(knock-down) 株系、过量表达株系以及烟草瞬时表达系统,研究了BOI 基因在细胞程序化死亡(PCD)中的作用以及BOI 蛋白中RING 和WRD 结构域的功能。结果表明,BOI 基因的表达下调导致抗氧化胁迫能力下降,弱化了对活性氧介导的PCD 抑制作用;BOI 过量表达或瞬间表达可以增强对活性氧介导和α-吡啶甲酸诱导的PCD 的抑制作用。进一步研究发现,BOI 蛋白中的RING 结构域是BOI 抑制PCD 所必需的,WRD 结构域与BOI 对PCD 的抑制作用关系不大。     

Effect of paraoxon-methyl and parathion-methyl on DNA in human lymphocytes and protective action of vitamin C

The molecular basis of the genotoxicity of the commonly used organophosphorus insecticide, parathion-methyl is poorly understood and there is a lack of information on the possible effects of its metabolic conversion products. In the present work the action of parathion-methyl and its immediate metabolite paraoxon-methyl on DNA in human lymphocytes was compared using the comet assay. Parathion-methyl at 25 and 75 µM did not cause any significant changes but at 200 µM a significant increase in the tail moment was observed as compared with the control. Paraoxon-methyl at 25, 75 and 200 µM evoked dose-dependent DNA damage measured as a significant increase in comet tail moment of lymphocytes. The change evoked by paraoxon-methyl at 200 µM was much more pronounced than that by parathion-methyl at the same concentration. To search for the mechanism underlying the observed effect, the action of a well-known antioxidant, vitamin C, along with parathion-methyl and paraoxon-methyl was studied. The vitamin at 10 and 50 µM reduced the DNA-damaging activity of paraoxon-methyl at all its concentrations. The results indicate that the reported genotoxic effects of parathion-methyl could be mainly attributed to its metabolite paraoxon-methyl. The protective action of vitamin C suggests that paraoxon-methyl may cause oxidative DNA damage. © 1999 Society of Chemical Industry     

Activation of systemic disease resistance in pea by an avirulent bacterium or a benzothiadiazole, but not by a fungal leaf spot pathogen

Inoculation of first expanded leaves of pea seedlings with an avirulent strain of Pseudomonas syringae pv. pisi , or treatment with sprays of a benzothiadiazole (20 or 100  μ g a.i. mL⁻¹), decreased the susceptibility of subsequent leaves 7 or 14 days later to challenge inoculation with Mycosphaerella pinodes . Inoculation of first leaves with a virulent strain of P. syringae pv. pisi or with M. pinodes did not decrease the susceptibility of plants to M. pinodes . Treatments effective in decreasing susceptibility to M. pinodes were similarly active against Uromyces viciae-fabae and virulent P. syringae pv. pisi . Effective treatments also enhanced the activities of the enzymes β-1,3-glucanase and chitinase in untreated upper leaves 6 days later. Ineffective treatments for decreased susceptibility had no effect on the activity of the enzymes. None of the treatments enhanced peroxidase activities. The results are discussed in relation to the reported signalling effects of the benzothiadiazole and in relation to a suggested high activity of the avirulent P. syringae pv. pisi strain and inactivity of M. pinodes in enhancing natural signalling.     

Identification of an RNA silencing suppressor encoded by Grapevine leafroll-associated virus 3

GLRaV-3, a member of the Closteroviridae family and type member of the genus Ampelovirus, is involved in the grapevine leafroll disease. Until now no RNA silencing suppressor has been found among viruses of this genus, contrary to what happens with a large number of other viral genera. In the sister genus Closterovirus, RNA silencing suppressors are present in the 3’ end of the genome and have molecular weights close to 20 KDa. To test for RNA suppressing activity screening of p21, p19.6 and p19.7 proteins, coded for in an analogous genomic location of the GLRaV-3 was undertaken. Only p19.7 revealed suppressor activity demonstrated in diverse silencing inducing systems. This suppressor is able to overcome strong silencing inducers and shares several properties with the BYV p21-like family of suppressors of the closteroviruses. This is the first report of an RNA silencing suppressor in the genus Ampelovirus.     

Tobacco rattle tobravirus: variation among strains and detection by cDNA probes1

Because of their extreme antigenic variability, only a small proportion of tobacco rattle tobravirus isolates will be detected in serological tests with a single antiserum. Moreover, serological tests will not detect NM-type isolates that do not produce coat protein, and would misidentify some natural recombinant isolates as pea early-browning tobravirus. In contrast, nucleic acid hybridization tests with a probe to any part of RNA1 can detect all these different variants of tobacco rattle tobravirus.     

Genetic diversity in the mango malformation pathogen and development of a PCR assay

Malformation is a destructive disease of mango, Mangifera indica . Its causal agent possesses the morphological features of Fusarium subglutinans , a species whose taxonomy and nomenclature has recently been in a state of flux. Genetic diversity was examined among 74 F. subglutinans -like isolates from malformed mango in Brazil, Egypt, Florida (USA), India, Israel and South Africa. With nitrate-nonutilizing ( nit ) auxotrophic mutants, seven vegetative compatibility groups (VCGs) were identified. Three of the VCGs were found in a single country, and VCG diversity was greatest in Egypt and the USA where, respectively, four and three different VCGs were found. RAPD profiles generated with arbitrary decamer primers were variable among isolates in different VCGs, but were generally uniform for isolates within a VCG. In PCR assays, a 20-mer primer pair that was developed previously to identify F. subglutinans from maize (mating population [MP]-E of the Gibberella fujikuroi complex) also amplified a specific 448 bp fragment for isolates of F. sacchari from sugarcane (MP-B) and what was probably F. circinatum (pine, MP-H). With the exception of three isolates from Brazil, it did not amplify the fragment from F. subglutinans -like isolates from mango. A second pair of 20-mer primers was developed from a unique fragment in the RAPD assays. It amplified a specific 608 bp fragment for 51 of 54 isolates from mango (all but the three Brazilian isolates). It also amplified a smaller, 550 bp fragment from isolates of F. nygamai (MP-G), but did not amplify DNA of isolates of any other taxon of Fusarium that was tested.