Chitinase Gene Expression Research Articles

Effect of cell wall integrity stress and RlmA transcription factor on asexual development and autolysis in Aspergillus nidulans

The cell wall integrity (CWI) signaling pathway is responsible for cell wall remodeling and reinforcement upon cell wall stress, which is proposed to be universal in fungal cultures. In Aspergillus nidulans, both the deletion of rlmA encoding the RlmA transcription factor in CWI signaling and low concentrations of the cell wall polymer intercalating agent Congo Red caused significant physiological changes. The gene deletion mutant ΔrlmA strain showed decreased CWI and oxidative stress resistances, which indicated the connection between the CWI pathway and the oxidative stress response system. The Congo Red stress resulted in alterations in the cell wall polymer composition in submerged cultures due to the induction of the biosynthesis of the alkali soluble fraction as well as the hydrolysis of cell wall biopolymers. Both RlmA and RlmA-independent factors induced by Congo Red stress regulated the expression of glucanase (ANID_00245, engA) and chitinase (chiB, chiA) genes, which promoted the autolysis of the cultures and also modulated the pellet sizes. CWI stress and rlmA deletion affected the expression of brlA encoding the early conidiophore development regulator transcription factor BrlA and, as a consequence, the formation of conidiophores was significantly changed in submerged cultures. Interestingly, the number of conidiospores increased in surface cultures of the ΔrlmA strain. The in silico analysis of genes putatively regulated by RlmA and the CWI transcription factors AnSwi4/AnSwi6 in the SBF complex revealed only a few jointly regulated genes, including ugmA and srrA coding for UgmA UDP-galactopyranose mutase and SrrA stress response regulator, respectively.

CT gene modulate differential expression of chitinase gene under variant habitats in Vibrio cholerae

ObjectiveTo investigate the interrelation of cholera toxin gene (CT gene) in expression of chitinase gene under different pH conditions among pathogenic and Non-pathogenic strains of Vibrio cholera (V. cholera). MethodsThe chitinase assay well diffusion method and calorimetric chitinase assay were performed. Further, time depended chitinase activity among pathogenic and nonpathogenic strain was evaluated with control as Escherichia coli. The expressed protein in variant environment was purified by cascade of chromatographic techniques. The partially purified protein was analyzed by SDS-PAGE in both the strain of V. cholera. ResultsThe results have shown differential expression of chitinase gene among vibrio in time depended chitinase activity, purification of expressed protein and SDS-PAGE analysis. ConclusionsFrom the current study, two conclusions came in picture, habitat is prime factor that regulation of chitin gene expression among many bacterial strains, second, moreover among the vibrio pathogenic strains (CT+) expression of chitinase gene is more precisely regulated by CT gene rather than external environments while in non-pathogenic strain (CT-) completely absent.

Chitinase Expression Due to Reduction in Fusaric Acid Level in an Antagonistic Trichoderma harzianum S17TH

To study the effect of reduction in phytotoxin level on fungal chitinases, antagonistic Trichoderma spp. were screened for their ability to reduce the level of fusaric acid (FA), the phytotoxin produced by Fusarium spp. A T. harzianum isolate S17TH was able to tolerate high levels of FA (up to 500ppm) but was unable to reduce the toxin to a significant level (non-toxic) added to minimal synthetic broth (MSB). However, the isolate was able to reduce 400ppm FA in the liquid medium after 7days to a non-toxic level and displayed similar level of antagonism over the control (without FA). In studies of the effect of the reduction in FA (400ppm) level on chitinase gene expression in PCR assays, nag1 was significantly repressed but ech42 expression was only slightly repressed. Chitinase activity was either reduced or absent in the extracellular proteins of MSB supplemented with 400ppm FA, which could be attributed to the effect of residual FA or its breakdown products through unknown mechanisms. Selection of S17TH as a toxin insensitive isolate that could commensurate the negative effect on chitinase activity makes it a potential antagonist against Fusarium spp.

Selection of a compatible biocontrol strain mixture based on co-cultivation to control rhizome rot of ginger

Rhizobacteria and Trichoderma spp. that were antagonistic against Fusarium oxysporum f.sp. zingiberi and Pythium splendens inciting rhizome rot disease of ginger were identified. In vitro assays identified positive interactions for growth, antifungal and chitinase gene expression between antagonistic TEPF-Sungal (Burkholderia cepacia) and S2BC-1 (Bacillus subtilis) with S17TH (Trichoderma harzianum), and vice versa. In comparison with the other strains and strain mixtures, one strain mixture, TEPF-Sungal + S17TH, recorded a maximum rhizome production of 84% efficiency with less incidences of yellows and rhizome rot at an 84.2 and 79.7% reduction over the pathogen control, respectively, in a polyhouse with a challenge inoculation with the pathogens. Inoculation of the mixture of biocontrol agents was associated with an increase in known defence gene products such as chitinase etc., so we speculate that these are involved in the mechanism of disease suppression. In field experiments, the strain mixture reduced yellows and rhizome rot to 45.9 and 49.3% over the untreated control, respectively, which is comparatively better than that produced with fungicide (44.3 and 45.3%, respectively). The treatment also produced an increased rhizome yield with an average increase of 60.0% relative to the control.

Sequence analysis and gene expression of putative oil palm chitinase and chitinase-like proteins in response to colonization of Ganoderma boninense and Trichoderma harzianum

Chitinases are glycosyl hydrolases that cleave the β-1,4-glycosidic linkages between N-acetylglucosamine residues in chitin which is a major component of fungal cell wall. Plant chitinases hydrolyze fungal chitin to chitin oligosaccharides that serve as elicitors of plant defense system against fungal pathogens. However, plants synthesize many chitinase isozymes and some of them are not pathogenesis-related. In this study, three full-length cDNA sequences encoding a putative chitinase (EgChit3-1) and two chitinase-like proteins (EgChit1-1 and EgChit5-1) have been cloned from oil palm (Elaeis guineensis) by polymerase chain reaction (PCR). The abundance of these transcripts in the roots and leaves of oil palm seedlings treated with Ganoderma boninense (a fungal pathogen) or Trichoderma harzianum (an avirulent symbiont), and a combination of both fungi at 3, 6 and 12 weeks post infection were profiled by real time quantitative reverse-transcription (qRT)-PCR. Our findings showed that the gene expression of EgChit3-1 increased significantly in the roots of oil palm seedlings treated with either G. boninense or T. harzianum and a combination of both; whereas the gene expression of EgChit1-1 in the treated roots of oil palm seedlings was not significantly higher compared to those of the untreated oil palm roots. The gene expression of EgChit5-1 was only higher in the roots of oil palm seedlings treated with T. harzianum compared to those of the untreated oil palm roots. In addition, the gene expression of EgChit1-1 and EgChit3-1 showed a significantly higher gene expression in the leaf samples of oil palm seedlings treated with either G. boninense or T. harzianum.

Expression of a rice chitinase gene in transgenic banana (‘Gros Michel’, AAA genome group) confers resistance to black leaf streak disease

Transgenic banana (Musa acuminata ‘Gros Michel’) integrating either of two rice chitinase genes was generated and its resistance to Black Leaf Streak disease caused by the fungus Mycosphaerella fijiensis was tested using a leaf disk bioassay. PCR screening indicated the presence of the hpt selectable marker gene in more than 90 % of the lines tested, whereas more than three quarters of the lines contained the linked rice chitinase gene resulting in a co-transformation frequency of at least 71.4 %. Further, a unique stable integration of the transgenes in each line revealed some false negative PCR results and the expected co-transformation frequency of 100 %. The transgene insert number per line ranged from 1 to 5 and single transgene insert lines (25 % of all) were identified. Considerable delay in disease development (up to 63 days post-incoculation) over a monitoring period of 108 days occurred in nine lines with extracellularly targeted chitinase out of 17 transgenic lines tested and their necrotic leaf area decreased by 73–94 % compared to the untransformed susceptible control line. Finally, correlation between symptom development and rice chitinase expression was confirmed in two lines by Western analysis. The potential of rice chitinase genes to enhance resistance against M. fijiensis in banana was demonstrated as well as the usefulness of the leaf disk bioassay for early disease screening in transgenic banana lines.Electronic supplementary materialThe online version of this article (doi:10.1007/s11248-012-9631-1) contains supplementary material, which is available to authorized users.

Expression of chitinase genes of Metarhizium anisopliae isolates in lepidopteran pests and on synthetic media

Pathogenecity of the well characterized entomopathogenic fungus Metarhizium anisopliae used for biocontrol of a wide range of insect pests secretes hydrolytic enzymes that degrade the host cuticle. The chitinolytic activity of high and low virulent isolates of M. anisopliae was assayed on minimal medium (MM) + colloidal chitin and MM supplemented with insect cuticles. Ex- pression pattern of four chitinase genes (chitinase (chi), chi 1, chi 2, chi 3) was profiled during pathogenic stages of the entomopathogen under in vitro and in vivo conditions. Reverse-transcription polymerase chain reaction (RT-PCR) analysis confirmed that chitinase cDNAs were expressed during the germination of fungus under nutrient-deprived conditions. RT-PCR analysis performed for the four chitinase genes on the two insect hosts Spodoptera litura and Helicoverpa armigera at six developmental stages of the pathogen displayed up-regulation in S. litura at mycosed and conidiated condition while with H. armigera there was expression only after 48 h of incubation. Differential expression of chi, chi 1 and chi 2 genes in vitro (nitrogen rich and nitrogen limiting media) and in vivo (live insect hosts S. litura and H. armigera) implicate the role of substrate differences in pathogenesis.

RETRACTED ARTICLE: Expression of a rice chitinase gene enhances antifungal response in transgenic litchi (cv. Bedana)

To enhance the antifungal response of litchi (Litchi chinensis Sonn.), transgenic plants were generated by transferring rice chitinase gene driven by a maize-ubiquitin promoter along with its first intron into the zygotic embryos via Agrobacterium tumefaciens-mediated transformation. After co-cultivation for 2 days, zygotic embryos were transferred onto Murashige and Skoog (MS) modified media supplemented with 25 mgl−1 hygromycin and 400 mgl−1 cefotaxime. Consequently embryos were selected and the antibiotic resistant transgenic plantlets were regenerated. The culture time from zygotic embryo to transgenic plants was 14 months. The integration of the transgene was confirmed by PCR, RT-PCR, Southern and western blot analyses. The transgenic plants exhibited higher chitinase activity than the non-transformed plants. The chitinase activity when examined using the native polyacrylamide in-gel assay, indicated that the foreign gene expression resulting in the protein of expected molecular weight that showed chitinase activity. The transgenic plants showed delayed onset of the disease and smaller lesions following in vitro inoculation of die-back, leaf spots and blight pathogen (Phomopsis sp.). The transgenic plants were adapted to the greenhouse and did not show any phenotypic alterations.

Construction of a Promoter-probe Vector for Bacillus thuringiensis: the Identification of cis-acting Elements of the chiA Locus

The expression and application of Bacillus thuringiensis (Bt) chitinase genes have been extensively investigated. However, little information is available regarding the regulation of chitinase gene expression in Bt. In this study, a shuttle promoter-probe vector was constructed incorporating the thermostable β-galactosidase gene bgaB of B. stearothermophilus as the reporter for the study of Bt promoters. Using this plasmid, the activity of the chiA gene promoter in Bt was investigated. Deletion analysis of the putative chiA promoter region revealed that the sequence located ~75 bp DNA from positions -116 to -42, with respect to the translation start site, is the core promoter of chiA gene. Furthermore, a site for chitin induction was identified near position -36. This site for negative regulation was indicated downstream of the RNA polymerase binding sites of the promoter of chiA. The expression of chiA started in cell grown for about 6 h and reached the maximum after 60 h of incubation. Induction of chiA expression by chitin was demonstrated by an increase in β-galactosidase activity of ~2.5-fold.

Development of transgenic finger millet (Eleusine coracana (L.) Gaertn.) resistant to leaf blast disease

Finger millet plants conferring resistance to leaf blast disease have been developed by inserting a rice chitinase (chi11) gene through Agrobacterium-mediated transformation. Plasmid pHyg-Chi.11 harbouring the rice chitinase gene under the control of maize ubiquitin promoter was introduced into finger millet using Agrobacterium strain LBA4404 (pSB1). Transformed plants were selected and regenerated on hygromycin-supplemented medium. Transient expression of transgene was confirmed by GUS histochemical staining. The incorporation of rice chitinase gene in R0 and R1 progenies was confirmed by PCR and Southern blot analyses. Expression of chitinase gene in finger millet was confirmed by Western blot analysis with a barley chitinase antibody. A leaf blast assay was also performed by challenging the transgenic plants with spores of Pyricularia grisea. The frequency of transient expression was 16.3% to 19.3%. Stable frequency was 3.5% to 3.9%. Southern blot analysis confirmed the integration of 3.1 kb chitinase gene. Western blot analysis detected the presence of 35 kDa chitinase enzyme. Chitinase activity ranged from 19.4 to 24.8. In segregation analysis, the transgenic R1 lines produced three resistant and one sensitive for hygromycin, confirming the normal Mendelian pattern of transgene segregation. Transgenic plants showed high level of resistance to leaf blast disease compared to control plants. This is the first study reporting the introduction of rice chitinase gene into finger millet for leaf blast resistance.

Over Expression of Chitinase and Chitosanase Genes from <i>Trichoderma harzianum</i> under Constitutive and Inducible Promoters in order to Increase Disease Resistance in Wheat (<i>Triticum aestivum</i> L)

Powdery mildew of wheat, caused by Erysiphe graminis f.sp. tritici is a serious threat to wheat grain production in many parts of the world. Efforts have been made to engineer resistance into wheat using single gene technology which culminated in the improved resistance efficiency but never a complete control. Some efforts of expressing genes for disease resistance under the control of constitutive promoter resulted in the production of abnormal plants. We used double gene technology and over expressed a couple of antifungal genes i.e., chitinase and chitosanase from Trichoderma harzianum simultaneously into winter wheat genotype Florida . They were expressed separately under the constitutive promoter ( Ubiquitin- 1) from Maize and stress cum disease inducible promoter ( Vst -1) from Vitis vinefera . Six lines achieved under constitutive promoter and five lines under inducible promoter showed the co-integration and expression of chitinase and chitosanase genes. All these lines showed multiple copy integration of both these genes except one line which showed single copy integration of chitinase under ubiquitin promoter. The copy number varied for Chitinase (1-3) and Chitosanase (2-10). Different promoters did not seem to have any impact on transformation efficiency. Pathological analysis done with E.graminis showed a decrease in the susceptibility for both types of transgenic plants. A decrease in susceptibility was seen upto 75% when the transgenics were under inducible promoter. While in transgenics under constitutive promoter the decrease in susceptibility of upto 60% was seen. All the primary transforments with the exception of a couple showed normal growth.

Regulation of chitinase genes expression in bacteria

Chitinases, which can hydrolyze chitin, occur in a wide range of microorganisms including viruses, bacteria, and fungi. The derivatives of chitin are potentially useful in several areas such as food processing, medicines, and biological control in agriculture. Some bacteria can uptake and utilize chitin as carbon source by secreting chitinase. The chitin is degraded into chito-oligosaccharides [(GlcNAc)n] or N-acetylglucosamine (GlcNAc) by chitinases, and then the chitin derivatives are transferred into cells by specific transport systems of bacteria. The intracellular chitin derivatives activate or suppress the transcription of a series of chi genes and affect the amount of chitinase. The expression of chitinase genes are strictly regulated by various regulatory factors and responsive cis-acting elements. The present review will focus on the transport system and the regulation of chitinase genes expression in bacteria.

Differential Regulation of Orthologous Chitinase Genes in Mycoparasitic Trichoderma Species

Mycoparasitic Trichoderma species have expanded numbers of fungal subgroup C chitinases that contain multiple carbohydrate binding modules and could thus be important for fungal cell wall degradation during the mycoparasitic attack. In this study, we analyzed the gene regulation of subgroup C chitinases in the mycoparasite Trichoderma virens. In addition to regulation by nutritional stimuli, we found complex expression patterns in different parts of the fungal colony, and also, the mode of cultivation strongly influenced subgroup C chitinase transcript levels. Thus, the regulation of these genes is governed by a combination of colony-internal and -external signals. Our results showed completely different expression profiles of subgroup C chitinase genes in T. virens than in a previous study with T. atroviride, although both fungi are potent mycoparasites. Only a few subgroup C chitinase orthologues were found in T. atroviride and T. virens, and even those showed substantially divergent gene expression patterns. Microscopic analysis revealed morphogenetic differences between T. atroviride and T. virens, which could be connected to differential subgroup C chitinase gene expression. The biological function of fungal subgroup C chitinases therefore might not be as clear-cut as previously anticipated. They could have pleiotropic roles and might be involved in both degradation of exogenous chitinous carbon sources, including other fungal cell walls, and recycling of their own cell walls during hyphal development and colony formation.

High expression levels of chitinase genes inStreptomyces coelicolorA3(2) grown in soil

Although Streptomyces species are major chitin-degraders in soil ecosystems, the expression of the diverse chitinase genes within Streptomyces coelicolor grown in soil has not been assessed. As a first step, the induction pattern of nine chitinase genes in S. coelicolor growing in autoclaved soil was compared with those in liquid cultures. The relative expression levels of nine chitinase genes were measured using real-time reverse transcription PCR. The expression of all chitinase genes was induced by chitin in both autoclaved soil and liquid cultures, but to different levels. The expression levels of five chitinase genes in autoclaved soil were significantly higher than those in the liquid cultures. In particular, a putative chitinase gene, chitinase H, showed the highest induction in autoclaved soil. The same induction pattern was confirmed in nonautoclaved soil, indicating that soil contains some factors affecting the expression of chitinase genes. The chiH gene product, ChiH, cloned in Streptomycetes lividans was secreted and exhibited chitin degradation activity that was stable within a wide range of acidic pHs. The disruption of dasR, a transcriptional regulator for the uptake of N-acetylglucosamine, abolished the expression of chiH, demonstrating that DasR is required for the regulation of ChiH expression.

Quantitative analysis of chitinase gene expression in chickpea

The synthesis of chitinases is one of the strategies evolved in higher plants to resist stresses. To investigate the role of chitinases in the interaction between chickpea (Cicer arietinum L.) and pathogens, the expression of two classes of chitinase, I and III, was monitored in infected and non-infected plants with different disease resistance (resistant MCC10 and sensitive MCC724 genotypes). The expression levels of these genes in chickpea were studied at different times after inoculation with Fusarium oxysporum f. sp. ciceri isolate 85. Semi-quantitative reverse transcription PCR was used to study the differential induction of the chitinase genes. Expression levels of chitinase I in non-infected and infected MCC724 plants were higher than those in MCC10. Since chitinase III may play an effective role in plant defense, its expression levels were quantified by reverse transcription PCR. Chitinase III was induced in MCC724 within 6 h and in MCC10 in 48 h after inoculation. However, the expression levels of chitinase III in MCC10 non-infected plants were significantly higher than in MCC724. The results showed that chitinase III can be effective to enhance resistance to F. oxysporum f. sp. ciceri isolate 85.

Over Expression of Rice chitinase Gene in Transgenic Peanut (Arachis hypogaea L.) Improves Resistance Against Leaf Spot

A Rice chitinase-3 under enhance version of CaMV 35S was introduced into peanut (Arachis hypogaea L.) through Agrobacterium mediation. Agrobacterium tumefaciens strain LB4404 was used harboring the binary vector (pB1333-EN4-RCG3) containing the chitinase (chit) and hygromycin resistance (hpt) gene as selectable marker. Putative transgenic shoots were regenerated and grown on MS medium supplemented with 5 mg/l BAP, 1 mg/l kinetin, and 30 mg/l hygromycin. Elongated shoots were examined for the presence of the integrated rice chitinase gene along with hygromycin gene as selectable. The integration pattern of transgene in the nuclear genome of the putative transformed plants (T(0)) was confirmed through Southern hybridization analysis of the genomic DNA. Survival rate of the in vitro regenerated plantlets was over 60% while healthy putatively transgenic (T(0)) plants with over 42% transformation frequency were produced through Agrobacterium mediated gene transfer of the rice chitinase gene and all the plants flowered and set seed normally. T1 plants were tested for resistance against Cercospora arachidicola by infection with the microspores. Transgenic strains exhibited a higher resistance than the control (non-transgenic plants). chitinase gene expression in highly resistant transgenic strains was compared to that of a susceptible control. A good correlation was observed between chitinase activity and fungal pathogen resistance.

Analysis of subgroup C of fungal chitinases containing chitin-binding and LysM modules in the mycoparasite Trichoderma atroviride

Fungi have a plethora of chitinases, which can be phylogenetically divided into three subgroups (A, B and C). Subgroup C (sgC) chitinases are especially interesting due to their multiple carbohydrate-binding modules, but they have not been investigated in detail yet. In this study, we analyzed sgC chitinases in the mycoparasites Trichoderma atroviride and Trichoderma virens. The expression of sgC chitinase genes in T. atroviride was induced during mycoparasitism of the fungal prey Botrytis cinerea, but not Rhizoctonia solani and correspondingly only by fungal cell walls of the former. Interestingly, only few sgC chitinase genes were inducible by chitin, suggesting that non-chitinous cell wall components can act as inducers. In contrast, the transcriptional profile of the most abundantly expressed sgC chitinase gene tac6 indicated a role of the protein in hyphal network formation. This shows that sgC chitinases have diverse functions and are not only involved in the mycoparasitic attack. However, sequence analysis and 3D modelling revealed that TAC6 and also its ortholog in T. virens have potentially detrimental deletions in the substrate-binding site and are thus probably not catalytically active enzymes. Genomic analysis showed that the genes neighboring sgC chitinases often encode proteins that are solely composed of multiple LysM modules, which were induced by similar stimuli as their neighboring sgC chitinase genes. This study provides first insights into fungal sgC chitinases and their associated LysM proteins.

Regeneration and Transformation of Taro (Colocasia esculenta) with a Rice Chitinase Gene Enhances Resistance to Sclerotium rolfsii

Genetic engineering has the potential to improve disease resistance in taro [Colocasia esculenta (L.) Schott]. To develop a method to produce highly regenerable calluses of taro, more than 40 combinations of Murashige and Skoog (MS) media at full- or half-strength with varying concentrations of auxin [α-naphthaleneacetic acid (NAA) or 2, 4-dichlorophenoxyacetic acid (2, 4-D)], cytokinin [benzyladenine (BA) or kinetin], and taro extract were tested for callus initiation and plant regeneration. The best combination, MS medium with 2 mg·L−1 BA and 1 mg·L−1 NAA (M5 medium), was used to produce regenerable calluses from taro cv. Bun Long initiated from shoot tip explants. After 8 weeks of growth, multiple shoots from these calluses could be induced on MS medium with 4 mg·L−1 BA (M15 medium). The rice chitinase gene (ricchi11) along with the neomycin phosphotransferase (npt II) selectable marker and β-glucuronidase (gus) genes were introduced into these taro calluses through particle bombardment. Transformed calluses were selected on M5 medium containing 50 mg·L−1 geneticin (G418). Histochemical assays for beta-glucuronidase (GUS), polymerase chain reaction (PCR), reverse transcription–PCR, and Southern blot analyses confirmed the presence, integration, and expression of the rice chitinase gene in one transgenic line (efficiency less than 0.1%). Growth and morphology of the transgenic plants appeared normal and similar to non-transformed controls. In pathogenicity tests, the transgenic line exhibited improved resistance to the fungal pathogen, Sclerotium rolfsii, but not to the oomycete pathogen, Phytophthora colocasiae.

Differential expressions of PR1 and chitinase genes in harvested bananas during ripening, and in response to ethephon, benzothiadizole and methyl jasmonate

Ripening of banana ( Musa acuminate L.) fruit is concomitant with weakening of disease resistance. The aim of this research was to determine the patterns of expression of PR1 and chitinase genes during banana ripening. Northern blots show that the MaPR1 did not have expression in peel of young growing banana fruit, nor in harvested bananas until the onset of ripening. The chitinase gene ( MaChit) had constitutive expression in bananas during development and after harvest. MaChit expression increased with time after harvest and reached the highest level at the onset of ripening when MaPR1 transcripts began to give detectable signals. Infection of the banana pathogen Colletotrichum musae did not induce MaPR1 but increased MaChit expression. The ethylene producer ethephon immediately induced MaPR1, but benzothiadiazole (BTH) and methyl jasmonate (MeJA), when applied alone, did not. The combination of ethephon and BTH or MeJA treatment increased intensity of PR1 signals 12 h after treatment compared with ethephon alone, and in the combination treatments, the inoculation-related lesions were significantly ( P < 0.05) smaller. In bananas treated with ethephon alone, the levels of MaPR1 and MaChit transcripts decreased in distal uninfected areas, but in a combination treatment, it did not decrease. The triple combination of ethephon, BTH and diphenylene iodonium (DPI) reduced MaPR1 and MaChit expression compared with a double combination of ethephon and BTH, whereas when MeJA replaced BTH in the triple treatment, the expression of the two genes increased compared with the double treatment. This research suggests that MaPR1 could be involved in defense mechanisms of ripe bananas, while MaChit could more be involved in banana resistance to disease before ripening. The results also suggest that BTH and MeJA-induced systemic defense in banana fruit is ethylene-dependent, and H 2O 2 might be necessary in BTH-induced SAR, but not in MeJA-induced systemic resistance. In the banana industry, MaPR1 could be used as a molecular indicator for monitoring banana ripening.

Differential expression of chitinase and stilbene synthase genes in Florida hybrid bunch grapes to Elsinoë ampelina infection

Florida hybrid bunch grape (Vitis spp.) cultivars with commercial value are susceptible to anthracnose disease caused by the fungi, Elsinoe ampelina. The Florida grape industry is keen on improving anthracnose tolerance characteristics of these grape cultivars as the cultivation of European grapes is limited in Southeastern United States due to Pierce’s disease incidence. In this connection, a molecular approach was taken to identify and characterize the gene/s differentially expressed upon infestation with E. ampelina in selected Florida hybrid bunch grape cultivars differing in their disease tolerance level. Variation in gene expressions was monitored by employing Differential Display RT-PCR. The results showed significant up-regulation of transcripts ranging between 200 and 800 bp in anthracnose-tolerant (cvs. Blue Lake and Lake Emerald) grape cultivars compared to anthracnose-susceptible (cvs. Suwannee and Blanc du Bois) cultivars. The transcripts that were uniquely induced in tolerant cultivars were identified as chitinase, stilbene synthase, protein/sugar kinases and transcriptional factor. Expression of chitinase and stilbene synthase genes was validated using real time PCR analysis. The results showed that these genes were more rapidly expressed in tolerant cultivars within a day after inoculating with E. ampelina and peaking by day 3, compared to susceptible cultivars. Induction of these anti-fungal genes appears to enable tolerant cultivars to withstand fungal infestation compared to susceptible cultivars. Rapid expression of chitinase and stilbene synthase genes along with others genes may be partly responsible for anthracnose-tolerance in Florida hybrid bunch grapes. Further analysis may shed light on their specific role in anthracnose tolerance.