Bioinformatics Analysis of Antifungal Peptide 25a2 of Cotton Verticillium Wilt

Protein 25a2 is the antifungal peptide of cotton Verticillium wilt which was isolated from Bacillus amyloliquefaciens. The amino acid sequence of antifungal peptide 25a2 was analyzed using bioinformatics tools, and the characters of signal peptides, transmembrane topological structura, physicochemical signatures, protein domain, secondary and tertiary structure of protein were predicted. The results showed that 25a2 was a secreted protein, the sequence of which included a signal peptide in N end and a transmembrane domain in C end. The predicted secondary structure showed that the antibacterial peptide was mainly free random coils, belonging to mixed protein, three-dimensional model of 25a2 was a compact ball. These results showed that the most possible action mechanism of antifungal peptide 25a2 might be ＂carpet＂ model.

Efficacy evaluation and mechanism of Bacillus subtilis EBS03 against cotton Verticillium wilt

Background:In our previous study,a strain EBS03 with good biocontrol potential was screened out of 48 strains of cotton endophyte Bacillus subtilis by evaluating the controlling effect against cotton Verticillium wilt.However,its mechanism for controlling Verticillium wilt remains unclear.The objective of this study was to further clarify its con-trolling effect and mechanism against cotton Verticillium wilt.Results:The results of confrontation culture test and double buckle culture test showed that the inhibitory effects of EBS03 volatile and nonvolatile metabolite on mycelium growth of Verticillium dahliae were 70.03%and 59.00%,respectively;the inhibitory effects of sporulation and microsclerotia germination were 47.16%and 70.06%,respec-tively.In the greenhouse test,the EBS03 fermentation broth root irrigation had the highest controlling effect at 87.11%on cotton Verticillium wilt,and significantly promoted the growth of cotton seedlings.In the field experi-ment,the controlling effect of EBS03 fermentation broth to cotton Verticillium wilt was 42.54%at 60 days after cotton sowing,and the boll number per plant and boll weight in EBS03 fermentation broth seed soaking,root irrigation,and spraying treatments significantly increased by 19.48%and 7.42%,30.90%and 2.62%,15.99%and 9.20%,respec-tively.Furthermore,EBS03 improved the resistance of cotton leaves against the infection of V.dahliae,and induced the outbreak of reactive oxygen species and accumulation of callose.In addition,the results of real time fluorescent quantitative polymerase chain reaction(RT-qPCR)detection showed that EBS03 significantly induced upregulation expression level of defense-related genes PAL,POD,PPO,and PR10 in cotton leaves,enhanced cotton plant resistance to V.dahliae,and inhibited colonization level of this fungal pathogen in cotton.Conclusion:Bacillus subtilis EBS03 has a good biological defense capability,which can inhibit the growth and coloni-zation level of V.dahliae,and activate the resistance of cotton to Verticillium wilt,thus increase cotton yield.

Study on Cotton Verticillium Wilt

The pathogen, characteristics, pathogenic mechanism, infection cycle, occurrence regularity and damage symptom of cotton Verticillium wilt are expounded in the paper. The paper puts forward the strategy of protecting disease-free area first, planting resistant varieties and rational rotation in severe area and strengthening the field management, as well as combining with biological control and chemical control methods to effectively control infection and spread of pathogen, so as to provide certain theoretical basis for reducing the damage of cotton Verticillium wilt and improving yield and quality of cotton.

Assessing the severity of cotton Verticillium wilt disease from in situ canopy images and spectra using convolutional neural networks

Verticillium wilt(VW)is a common soilborne disease of cotton.It occurs mainly in the seedling and bollopening stages and severely impairs the yield and quality of the fiber.Rapid and accurate identification and evaluation of VW severity(VWS)forms the basis of field cotton VW control,which has great significance to cotton production.Cotton VWS values are conventionally measured using in-field observations and laboratory test diagnoses,which require abundant time and professional expertise.Remote and proximal sensing using imagery and spectrometry have great potential for this purpose.In this study,we performed in situ investigations at three experimental sites in 2019 and 2021 and collected VWS values,in situ images,and spectra of 361 cotton canopies.To estimate cotton VWS values at the canopy scale,we developed two deep learning approaches that use in situ images and spectra,respectively.For the imagery-based method,given the high complexity of the in situ environment,we first transformed the task of healthy and diseased leaf recognition to the task of cotton field scene classification and then built a cotton field scenes(CFS)dataset with over 1000 images for each scene-unit type.We performed pretrained convolutional neural networks(CNNs)training and validation using the CFS dataset and then used the networks after training to classify scene units for each canopy.The results showed that the Dark Net-19 model achieved satisfactory performance in CFS classification and VWS values estimation(R^(2)=0.91,root-mean-square error(RMSE)=6.35%).For the spectroscopy-based method,we first designed a one-dimensional regression network(1D CNN)with four convolutional layers.After dimensionality reduction by sensitive-band selection and principal component analysis,we fitted the 1D CNN with varying numbers of principal components(PCs).The 1D CNN model with the top 20 PCs performed best(R^(2)=0.93,RMSE=5.77%).These deep learning-driven approaches offer the potential of assessing crop disease severity from spatial and spectral perspectives.

Physiological mechanisms involved in resistance to cotton verticillium wilt induced by AM fungi

It was proved that arbuscular mycorrhizal (AM) fungi played an important role in increasing plant resistance to soilborne pathogens, especially when plants were pre-inoculated with AM fungi. Mechanisms involved in this phenomenon are not yet well understood. On the basis of the former experiment results in our lab, effects of AM fungi on cotton Verticillium wilt and the mechanisms of increasing disease resisitance by the tested fungi were studied in pot culture under greenhouse conditions. Two cotton cutivars Litai 8 and 86-1 which are susceptible to Verticillium dahliae were pre-inoculated with Glomus fasiculatum, and Gigaspora margarita, then inoculated with the strain of Verticillium dahliae, namely “An-Yang” (belong to intermediate virulent type) 30 days after the former inoculation. Results showed that AM fungi could improve the growth and development of cotton plants, increase plants dry mass, decrease incidence and disease index of Verticillium wilt of cotton plants, inhibit the infection and development of V. dahliae to different extent in the rhizosphere of cotton pre-inoculated with AM fungi, while the colonization and spore numbers of AM fungi were not reduced significantly by this pathogen. The defence enzymes, such as phenylalanine ammonia-lyase (PAL), chitinase, β-1,3-glucanase, peroxidase, polyphenoloxidase (PPO) were induced, and their activities and peak increased by AM fungi in roots and leaves, and the increasing speed and peak of the enzyme activity were higher in treatment with AM fungus preinoculation than the inoculation with only V. dahliae, which suggested that defense response was activated by AM fungi, and then made the cotton to react strongly and rapidly to the infection of V. dahliae. In addition, AM fungi decreased the content of malondiadehyde (MDA) in cotton roots and leaves, protected membrane system and alleviated the damage caused by the pathogen. The AM fungus, Glomus fasiculatum showed the superior effects of biological control. It was concluded that AM fungi could provide the biological control to cotton diseases and there were application prospects of this biological agents.

Development and identification of Verticillium wilt-resistant upland cotton accessions by pyramiding QTL related to resistance

Cotton Verticillium wilt is a serious soil-borne disease that leads to significant losses in fiber yield and quality worldwide. Currently, the most effective way to increase Verticillium wilt resistance is to develop new resistant cotton varieties. Lines 5026 and 60182 are two Verticillium wilt-resistant upland cotton accessions. We previously identified a total of 25 quantitative trait loci（QTLs） related to Verticillium wilt resistance from 5026 and 60182 by assembling segregating populations from hybridization with susceptible parents. In the current study, using 13 microsatellite markers flanking QTLs related to Verticillium wilt resistance, we developed 155 cotton inbred lines by pyramiding different QTLs related to Verticillium wilt resistance from a filial generation produced by crossing 5026 and 60182. By examining each allele＇s effect and performing multiple comparison analysis, we detected four elite QTLs/alleles（q-5/NAU905-2, q-6/NAU2754-2, q-8/NAU3053-1 and q-13/NAU6598-1） significant for Verticillium wilt resistance, pyramiding these elite alleles increased the disease resistance of inbred lines. Furthermore, we selected 34 elite inbred lines, including five lines simultaneously performing elite fiber quality, high yield and resistance to V. dahliae, 14 lines with elite fiber quality and disease resistance, three lines with high yield and disease resistance, and 12 lines with resistance to V. dahliae. No correlation between Verticillium wilt resistance and fiber quality traits/yield and its components was detected in the 155 developed inbred lines. Our results provide candidate markers for disease resistance for use in marker-assisted breeding（MAS）, as well as elite germplasms for improving important agronomic traits via modern cotton breeding.

Advances and prospects of genetic mapping of Verticillium wilt resistance in cotton

Verticillium wilt is one of the most important diseases affecting cotton production in China.The fungus,Verticillium dahliae,has a wide host range and a high degree of genetic variability.No resistance resources have been found in the available planting resources,thus presenting difficulties and challenges for our study.The long-term production practice shows that selection of disease-resistant varieties is the most economical and effective measure to control Verticillium wilt of cotton to reduce the yield loss and quality decline of cotton.In this paper,we summarized the genetic mapping population,the analysis method of genetic localization,the discovery,mining and cloning of disease-resistant quantitative trait loci/markers,and the analysis of their genetic functions,so as to provide information for the molecular breeding approach of disease-resistant cotton.

Present Status, Problems and Prospects of Molecular Breeding of Cotton (Gossypium) with Resistance against Verticillium dahliae Kleb.

VerticiUium wilt is an important disease in cotton production, which seriously affects the production of cotton. The classification and pathogenesis of Yer-ticillium dahliae Kleb. , achievements in QTL mapping and molecular breeding were reviewed in the paper. Meanwhile, the current problems and future direction of resistance breeding against V. dahliae Kleb. were discussed so as to provide reference information for further research on resistance breeding against the pathogen.

The Influence of the Verticillium dahliae Kleb Infection on the Anti-Enzyme Inside the Body of the Cotton with Different Root Injured Degree

This study was to explore the influence of the Verticillium dahliae Kleb inflection on the anti-enzyme inside the body of the cotton with a different root injured degree. When the cotton seedling was long, with four leaves, it was flushed with water carefully, and then the following were obtained： （1） complete root seedling; （2） cut root seedling - by cutting off the lower part, 3-5 cm of the root, with a disinfected knife; （3） injured root seedling - by cutting off most of the side roots, but keeping the main root. Three kinds of cotton seedlings with different roots were immersed separately in different concentrations of the germ liquid （V. dahliae） of 20 mL each. Through 0- 48 h, the wilt degree of the seedling was recorded, and the related anti-enzyme of the variety was measured. After being immersed in the germ liquid, there was a significant difference in the wilt degree of the three kinds of injured root. When the germ liquid was in the ratio of 1：10, the complete root seedling was the lightest with no wilt; the injured root seedling was the second with a 2-degree wilt; but the cut root seedling was the most serious with a 3- degree wilt. At the same time, the changes in the peroxidase and malondialdehvde activities were determined. Peroxidase （POD） activities in the cut root seedling were 38.2 U mg^-1 min^-1, in the injured root seedling were 42.96 U mg^-1 min^-1, and in the complete root seedling were the highest at 49.2 U mg^-1 min^-1. The malondialdenvde （MDA） content in cut root seedling was 39.483 mmol g^-1, injured root seedling was 27.12 mmol g^-1, and the complete root seedling was only 3.845 mmol g^-1 The activity of the related anti-enzymes, such as POD was high or low, the quantity of the MDA was more or less, which they met the order of the harm of the seedlings. The change of SOD activities in cut root seedling was the most obvious as well. After injuring and inflecting the young roots, the exterior pathological reaction of the seedling and the dynamic state biochemical reaction of the related enzymes inside the plant body were studied. It showed that the plant exterior pathology responded to the test, with the internal biochemical reaction fitting together mutually.

Spectrum Characteristics of Cotton Canopy Infected with Verticillium Wilt and Applications

Hyper spectrum remote sensing with fine spectrum information is an efficient method to estimate the verticillium wilt of cotton. The research was conducted in Xinjiang, the largest cotton plant region of China, by using the data which were collected both by canopy spectrum infected with verticillium wilt and severity level （SL） in the year 2005-2006. The quantitative correlation was analyzed between SL and canopy of reflectance spectrum or derivative spectrum reflectance. The results indicated that spectrum characteristics of cotton canopy infected with verticillium wilt changed regularly with the increase of SL in different periods and varieties, Spectrum reflectance increased in the visible light region （620-700 nm） with the increase of the SL, which inverted in near-infrared region and was extremely significant in the region of （780-1 300 nm）. When SL attained b2 （DI = 25）, cotton canopy infected with verticillium wilt was used as a watershed and diagnosed index in the beginning stages of the disease. The results also indicated that there were marked different characteristics of the first derivative spectrum in these SL, it changed significantly in the red edge ranges （680-760 nm） with different SL, i.e., red edge swing decreased, and red edge position equally moved to the blue. In this study 1 001-1 110 nm and 1 205- 1 320 nm were selected out as sensitive bands for SL of canopy. Inversion models established for estimating cotton canopy infected with verticillium wilt reached the most significant level. Finally, the different spectrum characteristics of cotton canopy infected with verticillium wilt were marked, some inversion models were established, which could estimate SL of canopy infected with verticillium wilt. The best recognized model was the first derivative spectra at （FD 731 nm- FD 1317 nm）, and it might be used to forecast the position of cotton canopy infected with verticillium wilt quantitatively.

Advances in Cotton Breeding for Resistance to Fusarium and Verticillium Wilt in the Last Fifty Years in China

This review summarizes the main advances in cotton breeding for resistance to fusarium and verticillium wilt in the last fifty years in China. The topics discussed include main achievements, experiences, problems and countermeasures for solving.

Down regulation of cotton GbTRP1 leads to accumulation of anthranilates and confers resistance to Verticillium dahliae

Background:Verticillium wilt,caused by Verticillium dahliae,is called a "cancer" disease of cotton.The discovery and identification of defense-related genes is essential for the breeding of Verticillium wilt-resistant varieties.In previous research we identified some possible broad-spectrum resistance genes.Here,we report a tryptophan synthesis-related gene GbTRP1 and its functional analysis in relation to the resistance of cotton to V.dahliae.Results:Expression analysis shows that GbTRP1 is suppressed at 1 h and 6 h post V.dahliae infection,but activated at 12 h and 24 h,and the expression of GbTRP1 is highly induced by treatment with salicylic acid and jasmonic acid.Sub-cellular localization studies show that GbTRP1 is localized in the chloroplast.Suppression of GbTRP1 expression leads to lesion-mimic phenotypes and activates the immune response in cotton by showing enhanced resistance to V.dahliae and B.cinerea.Metabolomic analysis shows that anthranilic compounds significantly accumulated in GbTRP1-silenced plants,and these metabolites can inhibit the growth of V.dahliae and B.cinerea in vitro.Conclusions:Our results show that suppression of GbTRPI expression dramatically activates the immune response and increases resistance of cotton to V.dahliae and B.cinerea,possibly due to the accumulation of anthranilate compounds.This study not only provides genetic resources for disease resistance breeding,but also may provide a basis for new chemical control methods for combatting of fungal disease in cotton.

Isolation and characterization of the GbVIP1 gene and response to Verticillium wilt in cotton and tobacco

Background: Verticillium wilt is a serious soil-borne vascular disease that causes major losses to upland cotton(Gossypium hirutum L.) worldwidely every year. The protein VIP1(VirE2 interaction protein 1), a bZIP transcription factor, is involved in plant response to many stress conditions, especially pathogenic bacteria. However, its roles in cotton response to Verticillium wilt are poorly understood.Results: The GbVIP1 gene was cloned from resistant sea-island cotton(G. barbadense) cv. Hai 7124. Expression of GbVIP1 was up-regulated by inoculation with Verticillium dahliae and exogenous treatment with ethylene. Results of virus-induced gene silencing suggested that silencing of GbVIP1 weakened cotton resistance to Verticillium wilt. The heterologous expression of GbVIP1 in tobacco showed enhanced resistance to Verticillium wilt. The PR1, PR1-like and HSP70 genes were up-regulated in GbVIP1 transgenic tobacco after Verticillium wilt infection.Conclusion: Our results suggested that GbVIP1 increased plant resistance to Verticillium wilt through up-regulating expressions of PR1, PR1-like, and HSP70. These results provide new approaches to improving resistance to Verticillium wilt in upland cotton and also have great potential for disease-resistance breeding of cotton.

Studies on Cotton Breeding Resistant to Fusarium and Verticillium wilt Diseases

Both Fusarium and Verticillium wilts are important soil-borne diseases,which can not be effectively controlled by chemical fungicides.The two diseases,especially Verticillium wilt,have

Test on Wilt-Resistance of Simple Hybrides F7 and Beckross Hybrid Lines F6 B1 of <i>G. hirsutum</i>L. of Cotton to New Virulent Isolates of Fungus <i>Fusarium verticillioides</i>and <i>Verticillium dahliae</i>

The article presents the results of studies on the resistance of hybrid cotton lines to a new virulent isolate (strain) of the fungus Fusarium verticillioides upon inoculation of the host plant. Based on the studies, it was found that the complex genotypic resistance of the studied lines, when the host plants are inoculated with isolates of -100 V. dahliae Kleb fungus and 103 Fusarium verticillioides fungi, depends on the degree of resistance of the parental forms and their combination ability.

Genotypic Resistance of F_1 Cotton Hybrids by Inoculation with Different Virulent Isolates of the Fungus Verticillium Dahliae Klebahn

The plant pathogen Verticillium dahliae causes severe cotton losses in Uzbekistan. To create cotton varieties that are resistant to the more virulent races of V.dahliae we wanted to determine

黄萎病菌诱导下陆地棉酵母双杂交文库的构建及GhMAPKKK2互作蛋白的筛选

为了探究棉花响应黄萎病菌侵染的分子机制并筛选GhMAPKKK2的互作蛋白,以陆地棉XLZ35为材料,基于Gateway克隆技术构建了棉花黄萎病菌诱导的陆地棉酵母双杂交文库,其中初级文库和次级文库的库容分别为1.2×10^(7)和2.0×10^(7)CFU,插入片段重组率均为100%且平均长度大于1000 bp。次级文库质粒浓度为953 ng·μL^(-1),符合构建酵母文库的标准,可用于酵母双杂交筛选互作蛋白。构建了pGBKT7-GhMAPKKK2表达载体,并验证了该载体对酵母细胞无毒性且不存在自激活活性。以GhMAPKKK2为诱饵蛋白,利用酵母文库初步筛选,获得81个阳性互作蛋白,经测序对比选取6个候选蛋白,通过酵母双杂交及双分子荧光互补试验(BiFC)验证,明确了候选蛋白GhFLA与GhMAPKKK2存在互作关系。本研究结果为解析GhMAPKKK2调控棉花抗黄萎病反应分子机制及调控网络提供了重要参考。

有关棉花黄萎病（Verticillium dahliae Kleb．）抗性遗传研究的几个问题

棉花黄萎病抗性遗传问题迄今尚未获得肯定的结论。据作者等估计，这可能和寄主（棉株）和寄生物（黄萎病镰刀菌）之间的关系、黄萎病菌不同菌系间的互作以及影响抗性鉴定的正确性等问题有关。为此，必须创设有利条件，使寄主和病原菌间的关系得以充分表现；必须了解病原菌菌系间的互作，寄主必须有较大的群体。这一切都是为了寄主和病原物问的关系得以充分体现，为抗性遗传研究奠定基础。抗性遗传研究宜用单菌系接种，并且宜按数量性状和质量性状遗传方式同时进行研究和分析，以便得到比较确切可靠的结果。

大丽轮枝菌(Verticillium dahliae)微菌核缺陷突变体的筛选及其侧端序列分析

为了获得大丽轮枝菌(Verticillium dahliae)微菌核缺陷突变体,利用农杆菌介导转化大丽轮枝菌,共获得270个T-DNA随机插入突变体,从中筛选出了4个气生菌丝发达、不产微菌核的突变体,利用高效TAIL-PCR技术,获得了其侧端序列,结合生物信息学方法,获得4个突变体T-DNA在大丽轮枝菌基因组插入位点的基因信息。

大丽轮枝菌(Verticillium dahliae)酯酶同工酶的测定

对棉花黄萎病菌不同致病力及致病类型菌系，不同寄主上的大力轮枝菌（Verticilliumdahliae）共14个菌系的酯酶，进行了聚丙烯酰胺凝胶平板电泳。结果表明棉花黄萎病不同致病类型菌系有其独特的酯酶同工酶谱。E6酶带的有无与致病力类型有关，E3酶带活性与病菌致病力有关，E3酶带活性与棉花发病病指关联度为0．85。不同寄主大丽轮枝菌酯酶同工酶总酶带数在7～12条之间，主酶带数在2～4条之间，按E3酶带活性强、中、弱划分为3个类型。E3酶带活性与病菌致病力有关，其活性大小与棉花、番茄、茄子病指的关联度均为0．774以上。说明酯酶同工酶技术可和于鉴定大丽轮枝菌同一种内不同致病类型致病力菌系。