Identification and validation of reference genes for RT‐qPCR analysis in banana (Musa spp.) under Fusarium wilt resistance induction conditions

Banana (Musa spp.) is severely damaged by Fusarium wilt caused by Fusarium oxysporum f. sp. cubense (Foc). Biocontrol by inducing systemic resistance has been considered as one of the most important strategies to improve plant health. Very few studies have investigated appropriate reference gene selection for RT‐qPCR (quantitative real‐time polymerase chain reaction) analysis suitable for conditions of systemic activated resistance. In this study, we assessed over a time‐course the expression of seven candidate reference genes (EF1, TUB, ACT1, ACT2, L2, RPS2 and RAN) for Cavendish cultivar Brazilian (Musa spp. AAA) and dwarf banana cultivar Guangfen No. 1 (Musa spp. ABB) that were inoculated by Bacillus subtilis strain TR21 and Foc. We choose these plants because they are commonly planted in Southern China. Expression stability of the candidate genes was evaluated using various software packages (GeNorm, NormFinder and BestKeeper). L2 and TUB genes displayed maximum stability in Guangfen No. 1. In Brazilian, ACT1 and TUB were the most stable genes. To further validate the suitability of the reference genes identified in this study, the expression of pathogenesis‐related 1 (PR1) gene under TR21 and Foc strains Foc004/Foc009 treatments was also studied. Identified reference genes in this work that are most suitable for normalizing gene expression data in banana under Fusarium wilt resistance induction conditions will contribute to the understanding of disease resistance mechanisms induced by biocontrol strains in banana.     

香蕉植株内生细菌群落多态性研究

采用平板法对香蕉(Musa nana)植株的内生细菌进行分离纯化,并采用细菌脂肪酸法进行鉴定。结果表明,从香蕉的健康植株和感病植株中共分离得到内生细菌21属24种。从健株分离得到9种内生细菌,其中根、茎和叶分别分离到6种、2种和8种内生细菌。从病株分离得到15属17种内生细菌,其中根、茎和叶分别分离到3种、11种和6种。香蕉健株根部的内生细菌含量最高,达5.195×106cfu g-1,下部叶片内生细菌的含量最低,仅为30 cfu g-1;香蕉病株茎部内生细菌的数量显著高于其他部位,达1.05×107cfu g-1。这说明香蕉在不同生长状态下,其内生细菌的种类和数量存在多样性。     

Patterns of variability in the diameter of lateral roots in the banana root system

The relative importance of root system structure, plant carbon status and soil environment in the determination of lateral root diameter remains unclear, and was investigated in this study. Banana (Musa acuminata) plants were grown at various moderate levels of soil compaction in two distinct experiments, in a field experiment (FE) and in a glasshouse experiment (GE). Radiant flux density was 5 times lower in GE. The distribution of root diameter was measured for several root branching orders. Root diameters ranged between 0.09 and 0.52 mm for secondary roots and between 0.06 and 0.27 mm for tertiary roots. A relationship was found between the diameter of the parent bearing root and the median diameter of its laterals, which appears to be valid for a wide range of species. Mean lateral root diameter increased with distance to the base of the root and decreased with branching density [number of lateral roots per unit length of bearing root (cm(-1))]. Typical symptoms of low light availability were observed in GE. In this case, lateral root diameter variability was reduced. Although primary root growth was affected by soil compaction, no effects on lateral root diameter were observed.     

Assessment of a species-specific element (Brep 1) in banana

 The nuclear genome of wild-type banana accessions was investigated for repetitive elements. We report here the occurrence, in the banana genome, of a sequence family of species-specific repetitive elements: Brep 1. This sequence family is distributed throughout the Musaceae with various copy numbers. The two species Musa acuminata and M. schizocarpa carry the highest copy numbers in contrast to M. balbisiana and tested representatives of different other sections. PCR primers were defined in the core consensus sequence for specific amplifications, which allow representatives of this sequence family to be easily detected in wild and cultivated banana clones. Sequence data were analysed and hypotheses on the evolution of banana cultivars from the wild-type banana clones are discussed. Received: 17 January 1997 / Accepted : 7 March 1997     

A Review of: “HPLC of Biological Macromolecules (Methods and Applications) K.M. Goodring F.E. Regnier,Eds Marcel Dekker,New York, 1990; hardbound, 676 pages, $150”

An ionically unbound and thermostable polyphenol oxidase (PPO) was extracted from the leaf of Musa paradisiaca. The enzyme was purified 2.54-fold with a total yield of 9.5% by ammonium sulfate precipitation followed by Sephadex G-100 gel filtration chromatography. The purified enzyme exhibited a clear single band on native polyacrylamide gel electrophoresis (PAGE) and sodium dodecyl sulfate (SDS) PAGE. It was found to be monomeric protein with molecular mass of about 40 kD. The zymographic study using crude extract as enzyme source showed a very clear band around 40 kD and a faint band at around 15 kD, which might be isozymes. The enzyme was optimally active at pH 7.0 and 50°C temperature. The enzyme was active in wide range of pH (4.0–9.0) and temperature (30–90°C). From the thermal inactivation studies in the range 60–75°C, the half-life (t_1/2) values of the enzyme ranged from 17 to 77 min. The inactivation energy (Ea) value of PPO was estimated to be 91.3 kJ mol^?1. It showed higher specificity with catechol (K_m = 8 mM) as compared to 4-methylcatechol (K_m = 10 mM). Among metal ions and reagents tested, Cu²⁺, Fe²⁺, Hg²⁺, Mn²⁺, Ni²⁺, protocatechuic acid, and ferrulic acid enhanced the enzyme activity, while K⁺, Na⁺, Co²⁺, kojic acid, ascorbic acid, ethylenediamine tetraacetic acid (EDTA), sodium azide, β-mercaptoethanol, and L-cysteine inhibited the activity of the enzyme.     

Isolation of Endophytic Actinomycetes From Roots and Leaves of Banana (Musa Acuminata) Plants and Their Activities Against Fusarium oxysporumf. sp. cubense

Two hundred and forty-two actinomycete strains were isolated from the interior of leaves and roots of healthy and wilting banana plants. Most of them were streptomycetes, Streptomyces griseorubiginosus-like strains were the most frequently isolated strains. Community analysis demonstrated increased actinomycete diversity in wilting leaves compared to that in healthy leaves, similar actinomycete communities were found in wilting and healthy roots. Screening of the isolates for antagonistic activity against Fusarium oxysporumf. sp. cubenserevealed that the proportion of antagonistic streptomycetes in healthy roots was higher than that in wilting roots (P < 0.01), but no difference was found between antagonistic strains isolated from healthy and wilting leaves. The potential biological control of Panama disease of banana by endophytic streptomycetes, especially Streptomyces griseorubiginosus-like strains was discussed.     

香蕉试管苗混倍性变异的研究

首次报道了香蕉试管苗变异中一种重要的类型－混倍性变异体,并且以形态变异、染色体数量变异、同工酶（ＭＤＨ,ＥＳＴ,ＰＯＸ）变异和内源激素（ｉＰＡｓ和ＩＡＡ）变异等项检测为主要手段,研究了这种混倍性变异体在试管繁殖过程中的变异性质。用混倍性变异作外植体培养时,细胞水平染色体数量畸变具有起点高的前１５代增长快的特点,同工酶变异率比正常有显著增加。检测了用混倍性变异体作外植体培养所得的大棚苗的染色体数量畸     

Evaluation of an organic treatment for post-harvest control of crown rot of banana

An organic treatment for control of crown rot disease of banana was developed and evaluated at EARTH University in Costa Rica. Studies were conducted to evaluate the efficacy of Biocto 6 (seed extract from citrus) in combination with the wax-based adjuvant Verdiol for control of post-harvest crown rot of banana. The standard commercial fungicide treatment (thiabendazol, imazalil and ammonium sulfate) and an untreated control were included for comparison. Bananas with the various treatments were processed using standard commercial procedures and stored in a refrigerated chamber that was modified to simulate commercial transport, distribution and controlled ripening for exported bananas. Fruit clusters were evaluated for percent weight loss, ripening in storage and crown rot disease severity. At the end of the 28-day storage period, there were no significant differences in percent weight loss between any of the treatments. There was no significant difference in ripening (maturity level) between the organic treatment and the commercial fungicide standard in 2 years of testing. In 2003, the untreated control had a significantly higher maturity rating than the organic or standard fungicide treatment. However, there were no significant differences in any of the treatments in maturity level in 2005. There was no significant difference between the organic and standard fungicide treatment for crown rot control and both treatments had significantly less crown rot than the untreated control. Results indicate that Biocto 6 in combination with Verdiol wax provides a new organic alternative for control of post-harvest crown rot of banana.     

香蕉基因组SRAP反应体系的建立和优化

为建立并优化适于香蕉（Musa spp.）SRAP分析的扩增体系,对影响香蕉SRAP反应的dNTP、Mg²⁺、模板DNA、引物浓度和Taq酶用量等因素进行优化。确定的优化扩增体系为Mg²⁺ 2.5 mmol·L^-1,dNTP 250 μmol·L^-1,Taq酶1.0 U,引物0.5 μmol·L^-1,模板DNA 20 ng,10×PCR buffer 2.5 μL,在此条件下SRAP扩增香蕉基因组DNA条带清晰,多态性丰富。该体系在29个香蕉基因组中获得较好的扩增结果,可望在香蕉植物起源和进化研究中应用。     

Field Evaluation of Selected Formulations of Beauveria bassiana for the Management of the Banana Weevil ( Cosmopolites sordidus ) on Plantain ( Musa spp., AAB Group)

Field experiments were undertaken to determine the efficacy of two formulations of Beauveria bassiana isolate IMI331094 against the banana weevil, Cosmopolites sordidus . Oil palm kernel cake-based formulation of conidia (OPKC-C) and conidial powder (CP) were applied to the planting holes and suckers. After artificial weevil release, OPKC-C and CP gave the same high level of weevil mortality (75%) compared with only 1% in the untreated control. Under natural levels of weevil infestation, 42% of weevils collected from suckers treated with OPKC-C died compared with 6% and 3% weevil mortality for CP and the untreated control, respectively. None of the suckers treated with OPKC-C died during the study period (60 days) while 17% and 19% of suckers from the CP treatment and the untreated control respectively, were killed. A study on the spread of fungal conidia using artificially infected and non-infected adult weevils showed a possible dissemination of B. bassiana conidia from infected weevils up to 18 m from the release point. On the basis of these results, the isolate IMI330194 of B. bassiana could clearly play a key role in the management of C. sordidus adults on plantain.