Koch’s postulates

科赫的假设
  • 文章类型: Journal Article
    刺梨是一种起源于中国西南部的玫瑰灌木(Fu等人。2020)。2022年9月至10月,贵州省发生刺梨腐病,中国,在5公顷至50公顷的主要刺梨生产区内,发病率为10%至15%。症状表现为果实表面的黑色和棕色病变,是凹的,软,臭味,周围是灰棕色组织.从每个家庭随机收集三株受感染的刺梨灌木,放在透明的塑料袋中,病原体分离是在实验室进行的。用0.5%NaOCl对感染的刺梨果实进行表面灭菌2分钟,用无菌水冲洗五次,干了。将坏死组织和健康组织之间的边缘的症状组织切成5×5mm的碎片,置于马铃薯葡萄糖琼脂(PDA)上,并在黑暗中在28ºC下孵育5天。将从样品生长的真菌的菌丝尖端转移到新的PDA平板上并孵育直到它们产生分生孢子。总共获得了5种具有相似形态特征的真菌分离株。通过单孢子纯化获得的菌落为浅紫色至深紫色,具有丰富的气生菌丝体。大分生孢子相对细长,呈曲线状,为零至三个间隔。微分生孢子出现卵形至梨形,尺寸为5.2至17.2×2.1至3.3μm(n=50)。形态特征与环状镰刀菌一致(Yilmaz等人。2021)。使用DN14十六烷基三甲基溴化铵植物基因组快速提取试剂盒提取了两个代表性分离株(赵昌林1621和1622)的基因组DNA(AidlabBiotechnologiesCo.,Ltd,北京)。使用引物EF1-983F/EF1-2218R通过聚合酶链反应扩增TEF1和RPB2基因(Rehner等,2005),bRPB2-6F/bRPB2-7.1R(Matheny等人,2002),分别。所有序列都保存在GenBank(TEF1,PP236860,PP236861;RPB2,PP767864,PP767865)中。对TEF1和RPB2序列的BLAST搜索显示,TEF1序列与环生F.分离株CBS258.54具有99.89%(937/938bp)的同一性;RPB2序列具有99.86%(737/738bp)的同一性,以分离CBS267.93。在系统发育树中,分离株(赵昌林1621和1622)与环生F.形态学和多基因系统发育分析表明,该分离株为环生F.为了完成科赫的假设,五个成熟,健康的刺梨果实用1%NaClO溶液表面消毒1分钟,用无菌水冲洗,25℃干燥30分钟。将从两个分离株(赵昌林1621和1622)收集的分生孢子悬浮液(106孢子/ml)喷洒到刺梨果实上,和对照处理用无菌蒸馏水喷雾。所有刺梨果实在25ºC和80%相对湿度下孵育。实验有五个重复。经过7天的孵化,所有接种的水果都表现出与最初在最初感染的植物上观察到的相似的症状。通过形态学特征和分子分析对同一病原体进行了重新分离和鉴定,履行科赫的假设。因此,导致刺梨腐烂的病原体被确定为环叶F.(H.张等人,2024).据我们所知,这是在中国首次报道引起刺梨腐烂病。环孢菌具有广泛的宿主,据报道可以感染多种作物,水果,和蔬菜(培根和纳尔逊1994)。本研究为进一步研究和开发刺梨的病害防治方法及提高刺梨的经济效益奠定了基础。
    Rosa roxburghii Tratt is a rosaceous shrub originating from southwest China (Fu et al. 2020). From September to October 2022, R. roxburghii rot occurred in Guizhou Province, China, within a major R. roxburghii production area covering from 5 ha to 50 ha, with an incidence rate of 10 to 15%. Symptoms manifested as black and brown lesions on the fruit surface, which were concave, soft, foul-smelling, and surrounded by grayish-brown tissue. Three infected R. roxburghii shrubs were randomly collected from each household, placed in transparent plastic bags, and pathogen isolation was conducted in a laboratory. Infected R. roxburghii fruits were surface-sterilized with 0.5% NaOCl for 2 min, rinsed five times with sterile water, and dried. Symptomatic tissues from the margin between necrotic and healthy tissues were cut into 5 × 5 mm pieces, placed onto potato dextrose agar (PDA), and incubated at 28ºC in the dark for 5 days. Hyphal tips of fungi growing from the samples were transferred onto new PDA plates and incubated until they produced conidia. A total of five fungal isolates with similar morphological characteristics were obtained. The colony obtained by single-spore purification was light purple to dark purple with abundant aerial mycelium. Macroconidia were relatively slender with a curve and zero to three septate. Microconidia appeared obovoid to pyriform, with sizes of 5.2 to 17.2 × 2.1 to 3.3 μm (n = 50). The morphological characteristics were consistent with Fusarium annulatum (Yilmaz et al. 2021). Genomic DNA of two representative isolates (Zhaochanglin 1621 and 1622) was extracted using the DN14 cetyltrimethylammonium bromide rapid plant genome extraction kit (Aidlab Biotechnologies Co., Ltd, Beijing). The TEF1 and RPB2 gene were amplified by Polymerase Chain Reaction using primers EF1-983F/EF1-2218R (Rehner et al, 2005), bRPB2-6F/bRPB2-7.1R (Matheny et al, 2002), respectively. All sequences were deposited in GenBank (TEF1, PP236860, PP236861; RPB2, PP767864, PP767865). BLAST searches of the TEF1, and RPB2 sequences revealed the TEF1 sequences had 99.89% (937/938 bp) identity with F. annulatum isolate CBS 258.54; and the RPB2 sequences had 99.86% (737/738 bp) identity to isolate CBS 267.93. In the phylogenetic tree, the isolates (Zhaochanglin 1621 and 1622) clustered with the representative strains of F. annulatum. The morphology and multi-gene phylogenetic analysis indicated that is the isolates were F. annulatum. To complete Koch\'s postulates, five mature, healthy R. roxburghii fruits were surface disinfected with 1% NaClO solution for 1 min, rinsed with sterile water, and dried at 25℃ for 30 min. A conidial suspension (106 spores/ml) collected from two isolates (Zhaochanglin 1621 and 1622) was sprayed onto R. roxburghii fruits, and the control treatments were sprayed with sterile distilled water. All R. roxburghii fruits were incubated at 25 ºC with 80% relative humidity. The experiment had five replicates. After 7 days of incubation, all the inoculated fruits showed similar symptoms to those initially observed on the originally infected plants. The same pathogen was reisolated and identified by morphological character ization and molecular analysis, fulfilling Koch\'s postulates. Thus, the pathogen causing rot of R. roxburghii was determined to be F. annulatum (H. Zhang et al, 2024). To our knowledge, this is the first report of F. annulatum causing R. roxburghii rot disease in China. F.annulatum has a wide range of hosts and has been reported to infect a wide range of crops, fruits, and vegetables (Bacon and Nelson 1994). This study lays a foundation for further study and developing disease control methods and the improvement of the economic benefits of R. roxburghii.
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  • 文章类型: Journal Article
    maliVideira和Crous是苹果上一种新兴的采后病原体(Malus×domesticaBorkh。)在意大利和其他苹果生产国(Prencipe等人。2023年)。在1-2°C和0.5-2%的低氧水平下冷藏3至6个月后,皮孔显示直径为1毫米-5毫米的黑褐色斑点干腐病,没有定植于下面的组织。受影响最大的品种(简历。)在南蒂罗尔(意大利北部)是金色的,由于特征性的皮孔斑点,采后损失范围从10%到50%以上。四种有症状的水果,起源于两个果园(Latsch/Laces和Bozen/Bolzano;南蒂罗尔,意大利),分别,在冷藏后采样(=超低氧;0.5%O2和1°C)。表面用70%EtOH消毒1min后,将来自皮孔斑点的16个外植体在25°C的马铃薯葡萄糖琼脂(PDA)上培养。两个分离株,在形态上鉴定为Ramulariasp.,进行了测序,并显示出与R.mali型培养物CBS129581的高同一性:ITS区域的100%和99.31%同一性(MH865432);TEF-1α的94.66%和91.41%(KJ504693);RpbII的97.22%和97.40%(KJ504649)。将分离物在25°C培养2周,并用3.0mL0.05%Tween®20收获分生孢子。对5个月的冷藏水果cv一式三份进行接种。金色美味。用棉签表面消毒1分钟后,浸入70%的乙醇中,用注射器(Hamilton®型号710N)将每种分离物的10μL孢子悬浮液(在0.05%Tween®20中的8.50×107孢子mL-1)水平注射到表皮下方。此外,使用两种分离物的混合物。仅用0.05%Tween®20进行对照。将苹果在9°C下在黑暗中或在1°C和0.5%氧气下储存4个月。在9°C下2周后观察到两种孢子浓度的最初症状。注射途径变成了褐色,而对照组没有显示任何变化(图。1).4个月后进行最终评估,但是水果没有显示出进一步的症状发展。同时评估在1°C下储存5个月的水果,确认病原体侵入注射部位周围的组织,没有深入果肉。(Fig.2).从人工感染的苹果中成功分离,并对获得的分离物的DNA提取物进行序列分析。ITS的串联序列(以登录号存放于GenBank:PP439643-PP439647),TEF-1α(PP480231-PP480235),和RbpII(PP480226-PP480230)进行多位点序列分析。R.nyssicolaCBS127665,R.collo-cygniCBS101181,R.vizellaeCBS115981,R.eucalyptiCBS120726,R.hyrangeae-巨噬细胞CBS122272,R.glenniiCBS129441和R.maliCBS129581的参考序列包括并通过软件Geneiatters®新西兰)。用MEGAX(10.2.6版)重建系统发育(Kumar等人。2018)基于最大似然(ML)算法(图。3).来自人工感染的果实的分离株与R.mali型培养物成簇。虽然Gianetti等人。(2012)和林德纳(2013),分别,首先描述了Ramulariasp.作为苹果的采后病原体,本研究证明了马氏毛虫对皮孔干腐病症状的再现。
    Ramularia mali Videira & Crous is an emerging postharvest pathogen on apple (Malus × domestica Borkh.) in Italy and other apple producing countries (Prencipe et al. 2023). After 3 to 6 months of cold storage at 1 - 2 °C and low oxygen levels of 0.5 - 2 %, lenticels show black-brown speckled dry rot of 1 mm - 5 mm in diameter, without colonizing underlying tissue. The most affected cultivar (cv.) in South Tyrol (northern Italy) is Golden Delicious and postharvest losses due to characteristic lenticel spots range from 10 % to above 50 %. Four symptomatic fruits, originating from two orchards (Latsch/Laces and Bozen/Bolzano; South Tyrol, Italy), respectively, were sampled after cold storage (= ultra-low oxygen; 0.5 % O2 and 1 °C). After surface disinfection with 70 % EtOH for 1 min, sixteen explants from lenticel spots were cultivated on potato dextrose agar (PDA) at 25 °C. Two isolates, morphologically identified as Ramularia sp., were sequenced and showed high identities to R. mali type culture CBS 129581: 100 % and 99.31 % identity for ITS region (MH865432); 94.66 % and 91.41 % for TEF-1α (KJ504693); 97.22% and 97.40% for RpbII (KJ504649). Isolates were cultivated at 25 °C for 2 weeks and conidia were harvested with 3.0 mL 0.05 % Tween®20. Inoculation was performed in triplicate on 5-month cold stored fruits cv. Golden Delicious. After surface disinfection for 1 min with cotton swabs, which were immersed in 70 % EtOH, 10 µL spore suspension of each isolate (8.50 × 107 spores mL-1 in 0.05 % Tween®20) were injected horizontally beneath the epidermis with a syringe (Hamilton® model 710N). Also, a mixture of both isolates was used. Controls were carried out with 0.05 % Tween®20 only. Apples were stored either at 9 °C in the dark or at 1°C and 0.5 % oxygen for 4 months. First symptoms were observed for both spore concentrations after 2 weeks at 9 °C. The injection pathway changed to a brownish color, whereas the control did not show any change (Fig. 1). Final evaluation was carried out after 4 months, but the fruits did not show further symptom development. Fruits stored at 1°C for 5 months were simultaneously evaluated, confirming that the pathogen invaded the tissue surrounding the injection site, without penetrating deeper into the fruit flesh. (Fig. 2). Reisolation from artificially infected apples was successfully achieved, and sequence analysis was performed on the DNA extracts from the obtained isolates. Concatenated sequences of ITS (deposited to GenBank under the accession numbers: PP439643 -PP439647), TEF-1α (PP480231-PP480235), and RbpII (PP480226-PP480230) were subjected to multi-locus sequence analysis. References sequences of R. nyssicola CBS 127665, R. collo-cygni CBS 101181, R. vizellae CBS 115981, R. eucalypti CBS 120726, R. hydrangeae-macrophyllae CBS 122272, R. glennii CBS 129441 and R. mali CBS 129581 included and aligned by the CLUSTALW algorithm within the software Geneious® 11.1.5 (Biomatters Inc., New Zealand). Phylogeny was reconstructed with MEGAX (Version 10.2.6) (Kumar et al. 2018) based on the Maximum Likelihood (ML) algorithm (Fig. 3). Isolates from artificially infected fruit clustered with the R. mali type culture. Although Gianetti et al. (2012) and Lindner (2013), respectively, first described Ramularia sp. as a postharvest pathogen on apple, the present study demonstrated the reproduction of lenticel dry rot symptoms by R. mali.
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  • 文章类型: Journal Article
    背景:蜘蛛网病是一种真菌病,通常会影响食用蘑菇的种植和生产,导致严重的产量和经济损失。它被认为是食用蘑菇领域的主要真菌疾病。在石竹酱的种植过程中发现了蛛网病的症状。本研究旨在鉴定蜘蛛网病的病原并评估有效的杀菌剂,为L.dastes蛛网病的现场控制和管理提供有价值的见解。
    结果:从感染的样品中分离出蛛网病的病因,并根据形态和文化特征鉴定为嗜菌枝虫,以及多位点系统发育分析(ITS,RPB1、RPB2和TEF1-α)。致病性测试进一步证实了嗜霉菌梭菌是这种状况的主要病原体。在选定的杀菌剂中,丙氯胺-氯化锰络合物,Trifloxystrobin,戊唑醇,苯醚环唑对病原菌菌丝体有明显的抑制作用,EC50值为0.076µg/mL,0.173微克/毫升,和0.364µg/mL,分别。这些杀真菌剂可作为今后在L.debastes中防治蜘蛛网病的参考。
    结论:本研究是中国首次报道的嗜霉菌梭菌是乳糜泻蜘蛛网病的病因。值得注意的是,丙氯胺-氯化锰复合物对嗜霉菌梭菌表现出最强的抑制作用。
    BACKGROUND: Cobweb disease is a fungal disease that commonly affects the cultivation and production of edible mushrooms, leading to serious yield and economic losses. It is considered a major fungal disease in the realm of edible mushrooms. The symptoms of cobweb disease were found during the cultivation of Lyophyllum decastes. This study aimed to identify the causative pathogen of cobweb disease and evaluate effective fungicides, providing valuable insights for field control and management of L. decastes cobweb disease.
    RESULTS: The causal agent of cobweb disease was isolated from samples infected and identified as Cladobotryum mycophilum based on morphological and cultural characteristics, as well as multi-locus phylogeny analysis (ITS, RPB1, RPB2, and TEF1-α). Pathogenicity tests further confirmed C. mycophilum as the responsible pathogen for this condition. Among the selected fungicides, Prochloraz-manganese chloride complex, Trifloxystrobin, tebuconazole, and Difenoconazole exhibited significant inhibitory effects on the pathogen\'s mycelium, with EC50 values of 0.076 µg/mL, 0.173 µg/mL, and 0.364 µg/mL, respectively. These fungicides can serve as references for future field control of cobweb disease in L. decastes.
    CONCLUSIONS: This study is the first report of C. mycophilum as the causing agent of cobweb disease in L. decastes in China. Notably, Prochloraz-manganese chloride complex demonstrated the strongest inhibitory efficacy against C. mycophilum.
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  • 文章类型: Journal Article
    细菌理论指出,病原微生物是引起传染病的原因。该理论本质上是以微生物为中心的,没有考虑到个体之间疾病严重程度的变异性和无症状的携带者-两种现象表明宿主变异性在感染结果中的重要作用。细菌理论的基本原则最近受到挑战,并提出了一种激进的以主机为中心的范式,称为“全面的主机理论”。根据这个观点,病原体被还原成被动的环境触发因素,相反,疾病的发展是由于宿主预先存在的免疫缺陷。这里,我们使用有关进化生物学的既定知识来考虑确定疾病严重程度的因素,微生物发病机理,和宿主-病原体相互作用。我们注意到,现有数据支持非中心观点,该观点认识到致病微生物和宿主在决定感染结果方面的关键作用。
    The germ theory states that pathogenic microorganisms are responsible for causing infectious diseases. The theory is inherently microbe-centric and does not account for variability in disease severity among individuals and asymptomatic carriership-two phenomena indicating an important role for host variability in infection outcome. The basic tenet of the germ theory was recently challenged, and a radically host-centric paradigm referred to as the \"full-blown host theory\" was proposed. According to this view, the pathogen is reduced to a passive environmental trigger, and the development of disease is instead due to pre-existing immunodeficiencies of the host. Here, we consider the factors that determine disease severity using established knowledge concerning evolutionary biology, microbial pathogenesis, and host-pathogen interactions. We note that the available data support a noncentric view that recognizes key roles for both the causative microbe and the host in dictating infection outcome.
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  • 文章类型: Journal Article
    尖孢镰刀菌引起的叶枯病是中国铁皮石斛生产中的新兴问题。在瑞丽市苗圃种植的D.chrysothoum幼苗上观察到叶枯病的症状,云南省,中国。在这项研究中,我们分离了镰刀菌。与病苗中D.chrysotoxum的叶枯病有关。进行致病性测试以符合Koch的假设,以确认分离菌株的致病性,并使用形态学和分子技术进行鉴定。结果表明,所有四个分离的镰刀菌。分离株(DHRL-01〜04)产生典型的枯萎病症状,随后在D.chrysotoxum植物上叶片边缘坏死。在PDA介质上,真菌菌落呈白色至紫色,带有棉质菌丝生长。微分生孢子是椭圆形的,而大分生孢子是镰刀形的,在2-4隔的两端逐渐变细。系统发育树基于内部转录间隔区(ITS)进行解释,平移延伸因子(EF-1α),和RNA聚合酶亚基基因RPB1和RPB2基因,分别,并对NCBI数据库进行了爆炸以进行物种确认。根据NCBI数据库的爆炸结果,分离株显示,超过99%的人鉴定为尖孢镰刀菌。据我们所知,这是云南省首份关于尖孢镰刀菌为铁皮石斛叶枯病致病因子的综合鉴定报告,中国,基于形态和分子特征。
    Leaf-blight disease caused by the Fusarium oxysporum is an emerging problem in Dendrobium chrysotoxum production in China. Symptoms of leaf blight were observed on seedlings of D. chrysotoxum cultivated in a nursery in Ruili City, Yunnan Province, China. In this study, we isolated the Fusarium sp. associated with leaf-blight disease of D. chrysotoxum from the diseased seedlings. A pathogenicity test was performed to fulfill Koch\'s postulates to confirm the pathogenicity of isolated strains and identified using morphological and molecular techniques. The results revealed that all four isolated Fusarium sp. isolates (DHRL-01~04) produced typical blight symptoms followed by marginal necrosis of leaves on the D. chrysotoxum plants. On the PDA medium, the fungal colony appeared as a white to purplish color with cottony mycelium growth. Microconidia are oval-shaped, whereas macroconidia are sickle-shaped, tapering at both ends with 2-4 septations. The phylogenetic trees were construed based on internal transcribed spacer (ITS), translation elongation factor (EF-1α), and RNA polymerase subunit genes RPB1 and RPB2 genes, respectively, and blasted against the NCBI database for species confirmation. Based on the NCBI database\'s blast results, the isolates showed that more than 99% identify with Fusarium oxysporum. To our knowledge, this is the first comprehensive report on the identification of Fusarium oxysporum as the causal agent of Dendrobium chrysotoxum leaf blight in Yunnan Province, China, based on morphological and molecular characteristics.
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  • 文章类型: Journal Article
    在2015年和2016年,新加坡的两个Barramundi(Latescalcarifer)农场报告了以嗜睡行为为特征的疾病爆发,明显食欲不振,全身性皮肤损伤,鳍和尾巴的侵蚀,和最终高死亡率的鱼。下一代测序和PCR证实了一种属于同种疱疹病毒科的新病毒的存在。Latescalcarifer疱疹病毒(LCHV),随后被分离和培养。我们表征,第一次,两个培养的LCHV分离株的完整基因组。基因组包含一个约105,000bp的长独特区域,侧翼为约24,800bp的末端重复序列,其中前8.2kb与异源疱疹病毒科中描述的基因组没有任何相似性。两个培养的分离株共有89%的核苷酸同一性,它们的近亲是属于Ictalurivirus属的病毒。使用培养的LCHV分离株之一的实验性感染导致与索引农场最初描述的相同的临床症状。在腹膜内注射感染的鱼和同居的鱼中,两组都有死亡率。组织病理学分析显示the明显异常。病毒培养和PCR分析证实了LCHV在被感染的鱼中的复制,因此,科赫的假设得到了满足。
    In 2015 and 2016, two Barramundi (Lates calcarifer) farms in Singapore reported a disease outbreak characterized by lethargic behavior, pronounced inappetence, generalized skin lesions, erosions of the fins and tail, and ultimately high mortality in their fish. Next-generation sequencing and PCR confirmed presence of a novel virus belonging to the Alloherpesviridae family, Lates calcarifer herpesvirus (LCHV), which was subsequently isolated and cultured. We characterize, for the first time, the complete genome of two cultured LCHV isolates. The genome contains a long unique region of approximately 105,000 bp flanked by terminal repeats of approximately 24,800 bp, of which the first 8.2 kb do not show any similarity to described genomes in the Alloherpesviridae family. The two cultured isolates share 89% nucleotide identity, and their closest relatives are the viruses belonging to the genus Ictalurivirus. Experimental infections using one of the cultured LCHV isolates resulted in identical clinical signs as originally described in the index farm, both in intraperitoneal-injection infected fish and cohabitant fish, with mortality in both groups. Histopathological analysis showed pronounced abnormalities in the gills. Virus culture and PCR analysis confirmed the replication of LCHV in the infected fish, and thus Koch\'s postulates were fulfilled.
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  • 文章类型: Journal Article
    UNASSIGNED: Leaf spot disease severely impacts Ginkgo biloba (G. biloba) yield and quality. While microbial agents offer effective and non-toxic biological control for plant diseases, research on controlling leaf spot disease in G. biloba is notably scarce.
    UNASSIGNED: The pathogenic fungi were isolated and purified from diseased and healthy leaves of G. biloba, Subsequent examinations included morphological observations and molecular identification via PCR techniques. A phylogenetic tree was constructed to facilitate the analysis of these pathogenic fungi, and Koch\'s postulates were subsequently employed to reaffirm their pathogenic nature. The antagonistic experiment was employed to select biocontrol bacteria, and subsequently, the isolated biocontrol bacteria and pathogenic fungi were inoculated onto healthy leaves to assess the inhibitory effects of the biocontrol bacteria.
    UNASSIGNED: Two pathologies responsible for the leaf spot disease on G. biloba were identified as Botryosphaeria dothidea and Neofusicoccum parvum via the analysis of phylogenetic tree and the application of Koch\'s Postulates. Additionally, we isolated two strains of biocontrol bacteria, namely Bacillus velezensis and Bacillus amyloliquefaciens. Their average inhibitory zones were measured at 4.78 cm and 3.46 cm, respectively. The inhibition zone of B. velezensis against N. parvum was 4 cm. B. velezensis showed a stronger inhibitory effect compared to B. amyloliquefaciens on the development of lesions caused by B. dothidea via leaf culture experiment.
    UNASSIGNED: This research reports, for the first time, the presence of B. dothidea as a pathogenic fungus affecting G. biloba. Moreover, the biocontrol bacteria, B. velezensis and B. amyloliquefaciens, exhibited the capability to effectively inhibit the growth and reproduction of B. dothidea, indicating their promising potential as environmentally friendly biocontrol resources.
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  • 文章类型: Journal Article
    茶(山茶(L.)昆茨)是中国最重要的经济工厂之一,对人类健康有很多好处。炭疽病是我国茶病中最严重的一种,由于大多数中国品种对真菌敏感,因此控制真菌很重要。茶炭疽病的试剂最初被描述为日本的五宅一世,后来被转移到Discula,但是这个分类位置仍然存在问题。为了阐明这些分类和系统发育问题,对茶叶炭疽病病原菌进行了重新研究。结合nrITS的形态特征和多基因系统发育分析,nrLSU,rpb2和tef1序列数据,提出了一种新属Sinodiscula,以适应茶炭疽病的致病真菌,包括一个新物种山羊科和一个新组合。此外,根据Koch的假设确定病原体的致病性。本研究彻底解决了茶炭疽病病原体长期存在的分类学和系统发育问题。
    Tea (Camellia sinensis (L.) Kuntze) is one of the most important economic plants in China, and has many benefits for human health. Anthracnose is one of the most serious diseases of tea in China, and control of the fungus is important since most Chinese cultivars are susceptible to it. The agent of tea anthracnose was initially described as Gloeosporium theae-sinensis I. Miyake in Japan, which was later transferred to Discula, but this taxonomic position remains problematic. To shed light on these taxonomic and phylogenetic issues, the tea anthracnose pathogens were re-studied. Combining the morphological characteristics and a multigene phylogenetic analysis of nrITS, nrLSU, rpb2, and tef1 sequence data, a new genus Sinodiscula was proposed to accommodate the causal fungi of tea anthracnose, including a new species Sinodiscula camellicola and a new combination Sinodiscula theae-sinensis. Furthermore, the pathogenicity of the pathogens was determined according to Koch\'s postulates. This study thoroughly resolves the long-standing taxonomic and phylogenetic problems of the tea anthracnose pathogens.
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  • 文章类型: Journal Article
    木耳角膜是我国重要的食用菌作物,但蜘蛛网病的发生对其生产造成了重要的经济损失。全国疾病发生率为16.65%。在本研究中,一种新的病原体角膜Hypomycessp.11月。被发现导致蜘蛛网病。2021年7月,根据形态学研究和核核糖体DNA内部转录间隔区(ITS)的分子系统发育分析,从感染的子实体中分离出三株真菌病原体,并鉴定为H。rRNA的线粒体大亚基(LSU)和部分翻译延伸因子1-alpha基因。用于使用孢子悬浮液进行致病性测试的致病性高酵母物种的代表性分离株,其引起的症状与在耕种田地中观察到的相似,同样的病原体可以重新分离,这符合科赫的假设。检查了严重病原体的典型生物学特性,以确定其有利的生长条件,包括合适的温度,pH值,碳,氮源和光照条件。研究结果表明,最佳温度为25°C,pH为6,可溶性淀粉和蛋白胨为首选碳源和氮源,分别。采用菌丝生长抑制法对7种常用杀菌剂进行了初步的体外筛选试验,最合适的杀菌剂是丙氯胺氯化锰络合物,蜘蛛网病原体和蘑菇的EC50值分别为0.085μg/mL和2.452μg/mL,分别。我们的研究结果为有效预防和治疗角膜蛛网病提供了循证依据。
    Auricularia cornea is an important edible mushroom crop in China but the occurrence of cobweb disease has cause significance economic loss in its production. The rate of disease occurrence is 16.65% all over the country. In the present study, a new pathogen Hypomyces cornea sp. nov. was found to cause the cobweb disease. In July 2021, three strains of fungal pathogen were isolated from infected fruiting bodies and identified as H. cornea based on morphological studies and molecular phylogenetic analysis of internal transcribed spacer (ITS) of nuclear ribosomal DNA, mitochondrial large subunit (LSU) of rRNA and the partial translation elongation factor 1-alpha genes. The representative isolates of the pathogenic Hypomyces species used to perform pathogenicity test with spore suspension that caused similar symptoms as those observed in the cultivated field, and same pathogens could be re-isolated, which fulfill Koch\'s postulates. The typical biological characterization was examined of the serious pathogen to determine its favorable growth conditions, including suitable temperature, pH, carbon, nitrogen sources and light conditions. The findings revealed an optimum temperature of 25 °C, pH of 6, and soluble starch and peptone as the preferred carbon and nitrogen sources, respectively. The hyphal growth inhibition method was used for primary in vitro screening test of seven common fungicides, and the most suitable fungicide is Prochloraz manganese chloride complex, the EC50 values of cobweb pathogen and mushrooms were 0.085 μg/mL and 2.452 μg/mL, respectively. The results of our research provide an evidence-based basis for the effective prevention and treatment of A. cornea cobweb disease.
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  • 文章类型: Journal Article
    Artemisia capillaris (Asteraceae) is an annual herb found in ˃10 provinces in China. It is cultivated on ˃670 ha, with annual production around 2,500 tons. Its shoot is used in traditional Chinese medicine (Liu et al. 2021). From April to May 2023, Sclerotinia rot symptoms were seen at the Institute of Medicinal Plant Development (40.04°N, 116.28°E), Beijing, China. Disease incidence was up to 10% in the field through investigation of 300 plants. Initial symptoms were irregular tan-brown lesions (0.5 to 5.0 mm) that expended to circumferential necrosis on the roots and basal stem, aerial mycelia and sclerotia were developed on them. The leaves and stem tips were withered and droopy in severe cases. Twelve symptomatic primary roots of 12 plants from two sites were cut into 5 × 5 mm pieces, surface sterilized with 75% ethanol for 30 s and 5% NaClO for 60 s, rinsed with distilled water for three times, dried with sterile filter paper, put on potato dextrose agar (PDA), and incubated at 25°C in the dark for 2 days. Two Sclerotinia-like isolates were obtained using the hyphaltip method. White aerial mycelia were sparse and appressed for isolate YC1-3 and dense for isolate YC1-7. After incubated at 25°C in the dark for 15 days, 10 to 25 sclerotia were developed near the colony margin. Sclerotia of isolate YC1-3 were 1.0 to 3.9 × 1.2 to 4.5 (mean 1.8 × 2.2) mm (n = 60), ovoid or arc-shaped. Sclerotia of isolate YC1-7 were 1.5 to 3.4 × 2.7 to 9.2 (mean 2.3 × 4.3) mm (n = 60), ovoid, dumbbell shaped or curved. The isolates were identified as Sclerotinia sclerotiorum based on morphology (Maas 1998). To further identify the pathogens, molecular identification was performed with isolates YC1-3 and YC1-7. DNA of the two isolates were extracted by the cetyltrimethylammonium bromide (CTAB) method. Polymerase chain reaction was performed with primers ITS1/ITS4 for the internal transcribed spacer (ITS) region (Choi et al. 2020; White et al. 1990) and primers G3PDHfor/G3PDHrev for the glyceraldehyde 3-phosphate dehydrogenase (G3PDH) gene (Garfinkel. 2021). BLAST search analysis revealed that the ITS sequence (GenBank OR229758 and OR229762) was ≥99% similar to S. sclerotiorum (MN099281, MZ379265, KX781301, etc.), and the G3PDH sequence (OR778388 and OR761975) was too (MZ493894, JQ036048, OQ790148, etc.). Phylogenetic trees were computed with ITS and G3PDH sequences using the Maximum Likelihood in MEGA 11. Nine two-month-old seedlings of A. capillaris were used to test pathogenicity. The epidermis layer of each primary root was slightly wounded (2 × 2 mm, 1 mm deep) using a sterile dissecting blade. Three plants were inoculated with mycelial plugs (5 mm in diameter) of YC1-3 and YC1-7 that cultured on PDA for 7 days. Control plants were inoculated with sterile PDA plugs. All seedlings were then incubated at 25oC and 90% relative humidity. After isolate YC1-7 inoculation 3 days and isolate YC1-3 inoculation 5 days, inoculated roots had symptoms like those in the field, controls had no symptoms. S. sclerotiorum was consistently re-isolated from diseased roots, fulfilling Koch\'s postulates. Diseases caused by S. sclerotiorum have been reported threatens several important economical crops (Marin and Peres 2020; Guan et al. 2022). To our knowledge, this is the first report of S. sclerotiorum causes Sclerotinia rot on A. capillaris. To avoid of significant economic losses, it is urgent to establish an effective disease-management strategy.
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