Bean Common Mosaic Virus Infection Research Articles

Integrated transcriptome and microRNA sequencing analyses reveal gene responses in poplar leaves infected by the novel pathogen bean common mosaic virus (BCMV).

Recently, a novel poplar mosaic disease caused by bean common mosaic virus (BCMV) was investigated in Populus alba var. pyramidalis in China. Symptom characteristics, physiological performance of the host, histopathology, genome sequences and vectors, and gene regulation at the transcriptional and posttranscriptional levels were analyzed and RT-qPCR (quantitative reverse transcription PCR) validation of expression was performed in our experiments. In this work, the mechanisms by which the BCMV pathogen impacts physiological performance and the molecular mechanisms of the poplar response to viral infection were reported. The results showed that BCMV infection decreased the chlorophyll content, inhibited the net photosynthesis rate (Pn) and stomatal conductance (Gs), and significantly changed chlorophyll fluorescence parameters in diseased leaves. Transcriptome analysis revealed that the expression of the majority of DEGs (differentially expressed genes) involved in the flavonoid biosynthesis pathway was promoted, but the expression of all or almost all DEGs associated with photosynthesis-antenna proteins and the photosynthesis pathway was inhibited in poplar leaves, suggesting that BCMV infection increased the accumulation of flavonoids but decreased photosynthesis in hosts. Gene set enrichment analysis (GSEA) illustrated that viral infection promoted the expression of genes involved in the defense response or plant-pathogen interaction. MicroRNA-seq analysis illustrated that 10 miRNA families were upregulated while 6 families were downregulated in diseased poplar leaves; moreover, miR156, the largest family with the most miRNA members and target genes, was only differentially upregulated in long-period disease (LD) poplar leaves. Integrated transcriptome and miRNA-seq analyses revealed 29 and 145 candidate miRNA-target gene pairs; however, only 17 and 76 pairs, accounting for 2.2% and 3.2% of all DEGs, were authentically negatively regulated in short-period disease (SD) and LD leaves, respectively. Interestingly, 4 miR156/SPL (squamosa promoter-binding-like protein) miRNA-target gene pairs were identified in LD leaves: the miR156 molecules were upregulated, but SPL genes were downregulated. In conclusion, BCMV infection significantly changed transcriptional and posttranscriptional gene expression in poplar leaves, inhibited photosynthesis, increased the accumulation of flavonoids, induced systematic mosaic symptoms, and decreased physiological performance in diseased poplar leaves. This study elucidated the fine-tuned regulation of poplar gene expression by BCMV; moreover, the results also suggested that miR156/SPL modules played important roles in the virus response and development of viral systematic symptoms in plant virus disease.

First report of bean common mosaic virus naturally infecting yam bean (Pachyrhizus erosus) in China.

Yam bean (Pachyrhizus erosus), a high-yielding leguminous root crop with good nutritional value, is widely cultivated in southern China. In 2020, P. erosus (cv. Mumashan) plants exhibiting irregular yellow leaves and malformed seed pods (Supplementary Fig S1) were observed at Ningbo city, Zhejiang Province, China. To determine the causal agent(s) of the disease, symptomatic leaves (n=4) were collected for electron microscopy negative staining. Virus particles with a length of about 700nm, similar to viruses in the genus Potyvirus, were observed via transmission electron microscope (TEM), suggesting the presence a potyvirus(es). To further confirm which potyvirus(es) infected yam bean, total RNA was extracted from leaf samples of a total of six plants, including four symptomatic plants and two asymptomatic plants using TRIzol reagent (Invitrogen Carlsbad, CA, USA) according to the manufacturer's instructions. RNA was reverse-transcribed into cDNA with M4-T as the 3'-anchoring primer by ReverTra Ace® kit (Toyobo, Japan). Sprimer/M4 Potyviridae specific primers (Chen et al., 2001) were used for PCR analysis. A ~1,700-bp-long product was amplified from four symptomatic plants using KOD FX enzyme (Toyobo, Japan). No such band was amplified from the two asymptomatic plants. The PCR product (~1.7kb) amplified from a single symptomatic plant was ligated into the pEASY®-Blunt Zero vector (TransGen Bio, Beijing, China) and sequenced (Sangon Bio, Shanghai, China). The amplicon showed 99% nucleotide sequence identities with bean common mosaic virus (BCMV) isolate NKY021 (KJ807819). Subsequently, the complete nucleotide sequences of this BCMV isolate (referred as BCMV-NB) was amplified by overlapping RT-PCR and rapid amplification of cDNA ends with primers (Supplementary Table S1) designed from the sequence of BCMV isolate NKY021. The BCMV-NB full genome (Accession No. OL871237) consists of 10,053 nucleotides excluding the poly(A) tail and contains a large open reading frame encoding a polyprotein of 3222 amino acids. BLASTn analysis showed that BCMV-NB shared a sequence identity of 96.4% with BCMV isolate HZZB011 (KJ807815). Phylogenetic tree generated by Neighbour-Joining method revealing the BCMV-NB isolate was grouped together with Chinese isolates from Glycine max (Supplementary Fig S1). To test the infectivity of BCMV-NB, virus-free yam bean (cv. Mumashan) and Nicotiana benthamiana seedlings were mechanically inoculated with sap extracted from the symptomatic leaves of a BCMV-NB-infected yam bean plant. The inoculated yam bean plants developed typical BCMV mosaic and chlorotic symptoms at 16 days post inoculation (dpi), while Nicotiana benthamiana had no obvious symptoms at 10 or 20 dpi (Supplementary Fig S1). BCMV infections were confirmed in yam bean plants (infection rate 6/6) and N. benthamiana plants (infection rate 8/8) by RT-PCR at 16 dpi and 10 dpi, respectively. Twelve further P. erosus plants (cv. Mumashan) were collected from a field in Ningbo city and tested by RT-PCR with BCMV-specific primer pair BCMV CP (+)/(-) (Supplementary Table 1). Eight out of the 12 samples tested positive for BCMV by PCR-gel electrophoresis (Supplementary Fig S1) and Sanger sequencing, suggesting a high incidence of BCMV infection in this field. BCMV infection in yam bean has been reported from Indonesia (Damayanti et al., 2008) and Peru (Fuentes et al., 2012). To the best of our knowledge, this is the first report of BCMV naturally infecting yam bean in China. Thus, special attention and appropriate management strategies are needed to minimize the damage caused by BCMV to yam bean crops in China.

Development of a recombinase polymerase amplification combined with lateral flow dipstick assay for rapid and sensitive detection of bean common mosaic virus

Bean common mosaic virus (BCMV) is one of the most widespread and damaging viruses of cultivated legumes in the world. In addition to serious yield reduction and germplasm decline, BCMV infection also makes legumes more vulnerable to other pathogens. Early diagnosis of the virus is particularly important in limiting its spread. Recombinase polymerase amplification (RPA) is a novel isothermic amplification technology. The whole reaction can be done outside the laboratory environment after the nucleic acid sample is obtained. In this study, we established a rapid and sensitive RPA combined with the lateral flow dipstick (LFD) assay for detection of BCMV, based on the conserved BCMV coat protein (CP) gene sequence. Specific primers and a probe were designed, which amplify ~ 150 bp CP fragments from BCMV-infected samples under a constant temperature of 37 °C for 20 min. The end-labeled amplification products were detected by high-affinity LFD within 5 min. Sensitivity of this RPA-LFD assay was 1000 times greater than that of the conventional polymerase chain reaction (PCR) assay. Furthermore, when the primers/probe were used against related potyviruses including soybean mosaic virus (SMV), bean yellow mosaic virus (BYMV), and turnip mosaic virus (TuMV), the three potyviruses were not detected, indicating that the assay was BCMV species-specific. The RPA-LFD assay was also successfully applied for the detection of seed-borne BCMV in beans. The RPA-LFD assay has great potential application in the rapid diagnosis of BCMV in the field.

Identification of stable resistant donors against necrosis inducing strain of bean common mosaic virus in common bean (Phaseolus vulgaris L.)

Most of the commercially approved cultivars of Phaseolus vulgaris were susceptible to bean common mosaic virus (BCMV) which results in severe reduction in yield. In the present study, a total of 350 common bean genotypes were screened under natural epiphytotic conditions and artificial inoculation during 2016–2017, 2017–2018 and 2018–2019. Only eleven genotypes viz., EC400445, EC400414, ET4515B, EC540173, IC360831, BD9116291, EC31084, EC541703, ET8409, IC340947 and IC356024 were found stable resistant. The BCMV infection was confirmed through reverse-transcription polymerase chain reaction using coat protein gene amplification. The amplified region was sequenced and analyzed. Percent identity of BCMV-K with reported BCMV strains from different parts of world and based on nucleotide sequence ranged between 94% and 99.70% and maximum identity (99.70%) is observed with the strains of Uttar Pradesh (MT265069) and Himachal Pradesh (FJ387162, EU492546). The identified resistant sources from this study could be utilized as donors to develop BCMV resistant common bean cultivars.

Enhanced tolerance to seed-borne infection of bean common mosaic virus in salicylic acid treated bean plant

Bean common mosaic virus (BCMV)-infected bean seeds (P. vulgaris L. cv. Sadri) were subjected to low doses of gamma irradiation and the obtained seedlings were treated with 100 µm salicylic acid (SA). Bean seedlings treated with just the low doses of gamma rays, SA alone, or those treated with gamma rays and SA, were screened for seed-borne infection of BCMV by the indirect plate-trapped antigen-enzyme-linked immunosorbant assay. The results indicated that BCMV infection and its symptom severity in bean seedlings was decreased in a dose-dependent manner either by gamma irradiations alone or in associated with SA, compared to the samples treated with just SA. The specific activity of various antioxidant enzymes, as well as the total protein content of the treated beans leaves, were also measured in virus-free healthy and infected bean seedlings. The results showed that all three treatments significantly increased the protein content in healthy and infected bean leaves compared to the non-irradiated control beans. The 20 and 30 Gy doses of gamma irradiation, alone or in association with the SA were also effective treatments to induce the antioxidant enzymes. These preliminary findings may indicate that the low doses of gamma irradiation may have increased the tolerance level of bean seedlings to BCMV infection.

Göller Bölgesi Fasulye Üretim Alanlarında Fasulye Adi Mozayik ve Fasulye Adi Mozayik Nekroz Virüs’ü Enfeksiyonları

Common bean, belonging to family Leguminosae (Fabaceae), has an important place in people nutrition for thousands of years. Although many viruses infect bean plants, economically most important ones are potyviruses. Among them, BCMV (Bean common mosaic virus) and BCMNV (Bean common mosaic necrosis virus) are the most common viruses in common bean production areas all over the world. During 2014-2015 growing season, total of 361 bean samples (275 leaf and 86 seed) were collected from several localities in the Lakes Region. According to the results of DAS-ELISA (Double Antibody Sandwich Enzyme Linked Immunusorbent Assay) test, percentages of the infections of BCMV and BCMNV in the leaf samples were 26.18% and 15.27%, respectively. In seed samples, BCMV and BCMNV infection rates were 2.32% and 6.97%, respectively. Percentages of the mixed infections of the viruses in the leaf samples were 25.45%, while that in the seed samples were. 55.81%. In the mechanically inoculation studies where two isolates (BCMV-I and BCMN-BT) were used, typical symptoms of the viruses were observed. In molecular studies, a 850 bp fragment of the CP (Coat Protein) gene was amplified by IC-RT-PCR (Immunocapture Reverse Transcription Polimerase Chain Reaction) using BCMV specific primer pairs, but amplification of the CP region of RNA genome of BCMNV could not be succeeded. This is first report of biological and serological methods in Lakes region.

Potential of Pseudomonas putida F1 to manage Bean common mosaic virus of bean

The ability of the isolated rhizobacterial strain F1 of Pseudomonas putida to promote bean growth and resistance against Bean common mosaic virus (BCMV) was evaluated. Moreover, defense enzymes and the temporal expression profile of defense genes were assessed using quantitative real-time polymerase chain reaction (RT-qPCR) in bean plants. Bean plants treated with P. putida exhibited increased shoot and root dry weight relative to the control plants under greenhouse conditions. Similarly, the yield and yield components were increased in rhizobacterial treated bean plants under field conditions. Plants inoculated with P. putida showed the best inhibition effect of BCMV under greenhouse and field conditions. BCMV titer was significantly reduced in P. putida inoculated plants. Treatment with P. putida recorded the highest values of peroxidase and polyphenol oxidase enzymes after BCMV inoculation. The transcriptional profiles of PR1, PR2, PR3 and LOX genes were highly increased in P. putida treated plants. The obtained results in this study elucidate the potential of P. putida in controlling BCMV infection in bean plants as well as the involved mechanisms in disease resistance.

Exogenous Application of RNAi-Inducing Double-Stranded RNA Inhibits Aphid-Mediated Transmission of a Plant Virus.

Plant viruses are difficult to control, and they decrease both the quality and yield of crops, thus threatening global food security. A new approach that uses topical application of double-stranded RNA (dsRNA) to induce antiviral RNA-interference has been shown to be effective at preventing virus infection in a range of plants following mechanical inoculation. In this study, topical application of dsRNA was effective against mechanical inoculation and aphid-mediated inoculation with the potyvirus bean common mosaic virus (BCMV). Topical application of dsRNAs targeting either the coding region of the potyviral nuclear inclusion b (NIb) protein (BCMVNIb-dsRNA) or the coat protein (CP) coding region (BCMVCP-dsRNA) protected Nicotiana benthamiana and cowpea (Vigna unguiculata) plants against mechanical inoculation with BCMV. BCMVCP-dsRNA was selected for subsequent aphid transmission experiments. BCMVCP-dsRNA was loaded onto layered double hydroxide nanoparticles to form BCMVCP-BioClay which is a more stable formulation for delivering dsRNA than naked dsRNA. BCMVCP-BioClay was shown to be successful in protecting plants against BCMV transmission by the aphid Myzus persicae. Spraying detached N. benthamiana leaves with BCMVCP-BioClay 5 days prior to exposure to viruliferous aphids protected the leaves from infection by BCMV. Importantly, spraying of intact N. benthamiana and cowpea plants with BCMVCP-BioClay 5 days prior to exposure to viruliferous aphids protected plants of both species from BCMV infection. This study demonstrates that topical application of dsRNA using BioClay protects plants from aphid-mediated virus transmission, which is an important first step toward developing practical application of this approach in crop protection.

Dynamic transcriptome profiling of Bean Common Mosaic Virus (BCMV) infection in Common Bean (Phaseolus vulgaris L.).

BackgroundBean common mosaic virus (BCMV) is widespread, with Phaseolus species as the primary host plants. Numerous BCMV strains have been identified on the basis of a panel of bean varieties that distinguish the pathogenicity types with respect to the viral strains. The molecular responses in Phaseolus to BCMV infection have not yet been well characterized.ResultsWe report the transcriptional responses of a widely susceptible variety of common bean (Phaseolus vulgaris L., cultivar ‘Stringless green refugee’) to two BCMV strains, in a time-course experiment. We also report the genome sequence of a previously unreported BCMV strain. The interaction with the known strain NL1-Iowa causes moderate symptoms and large transcriptional responses, and the newly identified strain (Strain 2 or S2) causes severe symptoms and moderate transcriptional responses. The transcriptional profiles of host plants infected with the two isolates are distinct, and involve numerous differences in splice forms in particular genes, and pathway specific expression patterns.ConclusionsWe identified differential host transcriptome response after infection of two different strains of Bean common mosaic virus (BCMV) in common bean (Phaseolus vulgaris L.). Virus infection initiated a suite of changes in gene expression level and patterns in the host plants. Pathways related to defense, gene regulation, metabolic processes, photosynthesis were specifically altered after virus infection. Results presented in this study can increase the understanding of host-pathogen interactions and provide resources for further investigations of the biological mechanisms in BCMV infection and defense.Electronic supplementary materialThe online version of this article (doi:10.1186/s12864-016-2976-8) contains supplementary material, which is available to authorized users.

Bahasa Indonesia

Recently, a mosaic disease was observed on jack bean cultivation in Dramaga, Bogor,West Java. The symptom on infected plants, i.e. combination of pale and dark green area was more obvious on young leaves. Vein banding, leaf malformation, and growth inhibition was also observed in infected plants. The symptom was not very obvious in older leaves. Serological detection gave positive reaction to Bean common mosaic virus (BCMV) antiserum, but reaction was negative when detected using specific primer to coat protein (CP) of BCMV strain black eye by reverse transcription polymerase chain reaction. DNA fragments were successfully amplified by RT-PCR using universal primer of CP gene and degenerated primer for cylindrical inclusion gen (CI) of Potyvirus. DNA analysis showed the highest homology (86.4–91.5%) to several isolates of BCMV-infecting different crops from several countries based on partial sequence of CP, whereas based on partial sequence of CI the highest homology (87.3%) was to BCMV-RU1-P. This is the first report of BCMV infection on jack bean in Bogor, West Java, Indonesia.

Control of Bean common mosaic virus by plant extracts in bean plants

The present study aimed to evaluate some plant extracts from Plectranthus tenuiflorus, Azadirachta indica, Clerodendrum inerme, Schinus terebinthifolius and Mirabilis jalapa as antiviral materials against Bean common mosaic virus (BCMV) in bean plants. The results showed that all the tested plant extracts were effective in reducing the number of local lesions formed by BCMV. The mixing of P. tenuiflorus extracts with BCMV inoculum achieved the highest reduction in BCMV infection (92%), while the mixing of S. terebinthifolius extracts with BCMV inoculum recorded the lowest reduction in BCMV infection (68%). Seed treatment with 1% (w/v) of P. tenuiflorus showed the highest reduction in disease incidence (93% and 85%) under greenhouse and field conditions, respectively. Spray treatments of P. tenuiflorus and M. jalapa reduced disease incidence to 12% and 17% under greenhouse conditions, whereas under field conditions they reduced disease incidence to 17% and 23%, respectively. Protein analysis revealed different effects of the tested treatments on bean plants indicating different gene(s) expression. This study suggests the possibility of using plant extracts for controlling BCMV infection.

Bahasa Indonesia

Incidence of yellow mosaic disease on yard long bean occurred widely in some areas in West and Central Java in 2008–2009. The causal agent of this disease is Bean common mosaic virus (BCMV) which can be transmitted through seed and aphids. Research was conducted to determine response of five varieties of yard long bean (Vigna sinensis), i.e. Parade, New Jaliteng, Long Silk, Super Sainan, and Pilar to BCMV infection. Yard long bean plants was inoculated by BCMV mechanically, followed by observation of disease incidence and severity, incubation period, flowering time, and yield. All five varieties of yard long bean showed susceptible response to BCMV infection. Incubation period was 6–16 days, disease incidence was 90–100%, and disease severity was 49.1–69.7%. BCMV infection caused severe symptoms of leaf malformation and plant stunting. Flowering time was delayed 2–5 days on infected plants and yield was decreased up to 46.6%. Breeding program to develop resistance varieties to BCMV is necessary.

AKTIVITAS ANTIVIRUS BEBERAPA EKSTRAK TANAMAN TERHADAP BEAN COMMON MOSAIC VIRUS STRAIN BLACK EYE COWPEA (BCMV-BIC) PADA KACANG PANJANG

ABSTRACTAntivirus actitivity of several plant extracts against Bean common mosaic virus strain Black eye cowpea (BCMV-BlC) on Yard long bean. Bean common mosaic virus (BCMV) is an important virus on yard long bean and it is difficult to control. One of control effort way by utilizing antiviral substances of plant origin. The research was done to select and test the effectiveness of plant extracts in suppressing BCMV infection on yard long bean. Twenty two plant extracts were selected by (1) spraying the crude extract to Chenopodium amaranticolor leaves, then plant inoculated by BCMV 1 hour after spraying, and (2) mixturing the crude extract with sap containing BCMV, then inoculated mechanically to C. amaranticolor. Local necrotic lesion number and inhibition percentage are measured. All plant extract treatments were able to reduce Necrotic lokal lesion formation significantly compared to untreatment control. Further, fifteen plant extracts were selected to test their effectiveness in controlling BCMV on yard long bean in green house trial. The results showed that except geranium and red ginger treatment, other extract treatments were able to reduce significantly the disease incidence and severity, symptoms, and BCMV titer, respectively. Among tested extracts, Bougainvillea spectabilis, Mirabilis jalapa, and Celosia cristata are the most effective crude extracts in suppressing BCMV infection.

Strobilurins Seed Treatment Enhances Resistance of Common Bean Against Bean common mosaic virus

AbstractCommercial formulations of strobilurins (azoxystrobin, kresoxim‐methyl, trifloxystrobin and pyraclostrobin) were evaluated for their efficacy against Bean common mosaic virus (BCMV) in screenhouse and field conditions. Highest seed germination and seedling vigour were recorded with 20 μg/ml pyraclostrobin seed treatment in comparison with the control. In screenhouse studies, 76% protection against BCMV was recorded with pyraclostrobin seed treatment at 10 μg/ml. Under field conditions with natural BCMV inoculum, pyraclostrobin seed treatment resulted in 65% protection against BCMV. The protection offered by strobilurins against BCMV was evaluated by ELISA, with lowest immunoreactive values recorded in common bean seedlings raised from seeds treated with pyraclostrobin and kresoxim‐methyl. Strobilurins in addition to exerting a direct positive physiological effect on common bean plants also protect bean plants against BCMV infection in screen house and field conditions. Thus, it is proposed that these reduced‐risk pesticides are potential inducers against BCMV and growth enhancers and could be a beneficial component of integrated disease management of common bean.

First Report of Bean common mosaic virus Infecting Lablab purpureus in India

Lablab bean (Lablab purpureus L. Sweet) is a widely cultivated, highly drought tolerant legume vegetable crop grown in diverse environmental conditions worldwide. In India and elsewhere, the young pods are consumed as a fresh vegetable and mature dry seeds are important in the diet of people preferring vegetarian food (2). Small-holding farmers use their own saved seeds for sowing. During October 2008, L. purpureus exhibiting symptoms of stunting, mosaic, vein-banding, vein-clearing, mottling, and blisters suggestive of a viral infection were observed in and around the Mysore District of Karnataka State, India. Incidence of the disease ranged from 1 to 10% in different fields. Symptomatic leaves were collected from fields of Daripura Village, Mysore District, Karnataka. Viruses that were tested by indirect ELISA included Cucumber mosaic virus, Tobacco mosaic virus, Cowpea aphid-borne mosaic virus, Cowpea mosaic virus, Cowpea mottle virus, Southern bean mosaic virus, and Bean common mosaic virus (BCMV). Results of the ELISA tests indicated that all 28 samples collected from different fields were infected with BCMV. Examination of tissue sap from symptomatic plants by electron microscopy revealed flexuous rod-shaped particles (~750 nm long). An immunocapture-reverse transcription (IC-RT)-PCR assay employing degenerate primers for amplifying partial coat protein (CP) and 3'-UTR of potyviruses (1) yielded a ~700-bp product that was cloned and sequenced (GenBank Accession No. HM776637). Sequence identity at the nucleotide level was 96% with BCMV strain NL-7n (GenBank Accession No. GQ456169) infecting common bean from Himachal Pradesh, India. RTPCR was performed with a virus-specific primer pair (FW3-5'-GCAGTAGCACAGATGAAGGCA-3': Rv3-5'-GGTTCTTCCGGCTTACTCATAAACAT-3') designed to amplify 340 bp, the partial coat protein gene of BCMV. All symptomatic L. purpureus field samples and screenhouse-grown seedlings manually inoculated with infected sap were positive for BCMV infection in RT-PCR assay employing specific primers with amplification of a 340-bp product. To our knowledge, this is the first report of BCMV infecting L. purpureus in India. BCMV has also been reported in L. purpureus in Uganda (4) and Nigeria (3). Plants that were confirmed by ELISA to be infected were tagged, and from these plants, seeds were collected and pooled. Four hundred seeds were germinated and a rate of 6.5% seed transmission was determined based on symptoms, ELISA, and PCR. From December 2008 to December 2010, different L. purpureus plantings were monitored for BCMV incidence. Plants infected at different growth stages were tagged and pods were harvested from infected and healthy plants. Data from at least 100 BCMV-infected L. purpureus plants from each of 12 different fields were recorded for yield loss analysis. In terms of number of pods per plant, number of seeds per pod, and seed weight, an average as much as 40% yield loss was recorded from 12 different fields. Because seeds collected from these plants are used for subsequent plantings, these plants may act as virus reservoirs or foci of infection.

Elimination of Bean common mosaic virus using an electrotherapy technique

Bean common mosaic virus (BCMV) is a major seed transmitted virus of common bean (Phaseolus vulgaris). The use of virus-free germplasm is a prerequisite for production of certified seeds, which is an important disease control measure. In this study, the efficiency of electrotherapy in eliminating BCMV and producing virus-free plants in two common bean cultivars, Khomein and Capsouli, was evaluated. Nodal cuttings of infected bean plants were exposed to electric currents of 5, 10 and 15 mA for 10 minutes and planted in vitro. One month old regenerated plantlets were tested by a Double Antibody Sandwich (DAS)-ELISA method for BCMV infection. The three intensities of electric current resulted in mean virus elimination percentages of 37.2%, 45.5% and 71.9% and mean plant regeneration of 79.5%, 68.6% and 58.4%, respectively. The Therapy Efficiency Index for the three electrotherapy treatments was estimated as 28.9, 29.2 and 39.1 in cv Khomein and 26.9, 33.3 and 44.8 in cv Capsouli, respectively. Based on the Therapy Efficiency Indices, an electric current of 15 mA for 10 min was the most effective treatment in both cultivars. The technique developed in this study can be a useful procedure for the production of virus-free germplasm and/or nucleus seed stock in crop plants infected with seed-borne viruses.

Chemical and Morphological Expression of the B and Asp Seedcoat Genes in Phaseolus vulgaris

Three common bean (Phaseolus vulgaris L.) seedcoat color (or glossiness) genotypes, differing from each other by a single substitution at a seedcoat locus, were analyzed for presence and concentration of three anthocyanins: delphinidin 3-O-glucoside, petunidin 3-O-glucoside, and malvidin 3-O-glucoside. The three anthocyanins were present in Florida common bean breeding line 5-593 (P C J G B V Asp), matte black (P C J G B V asp), and dark brown violet (P C J G b V Asp), but the amounts varied greatly depending on the genotype. Dark brown violet had 19% of the total anthocyanin content when compared to 5-593, whereas matte black had amounts intermediate between the two other genotypes. The B gene acts to regulate the production of precursors of anthocyanins in the seedcoat color pathway above the level of dihydrokaempferol formation, perhaps at the chalcone synthase or chalcone isomerase steps in the biosynthetic pathway. We hypothesize that B regulates simultaneously the flavonoid (color) and isoflavonoid (resistance) pathways. The I gene for resistance to bean common mosaic virus (BCMV) is known to be linked closely to B. It is therefore hypothesized that the I gene function may be to respond to BCMV infection by dramatically increasing (over a low constituitive level) production in the 5-dehydroxy isoflavonoid pathway, which leads to synthesis of the major phytoalexin, phaseollin, for resistance to BCMV. Alternatively, the B and I genes may be allelic. The Asp gene affects seedcoat glossiness by means of a structural change to the seedcoat. We demonstrate that Asp in the recessive condition (asp/asp) changes the size and shape of the palisade cells of the seedcoat epidermis, making them significantly smaller than either 5-593 or dark brown violet. Asp, therefore, limits the amounts of anthocyanins in the seedcoat by reducing the size of palisade cells.

Bean common mosaic virus reduces ozone sensitivity of pinto bean

Bean common mosaic virus infection reduced the sensitivity of pinto bean to ozone (O 3) damage. Immature primary leaves inoculated with the virus five days prior to exposure with 0·25 ppm (0·49 mg/m 3) O 3 for 4 h showed less O 3 injury than the non-infected control plants. Protection was influenced by the initial inoculum dilution and by the length of time between inoculation and exposure to O 3.