摘要: 目的：旨在為明確番石榴黑斑病優(yōu)勢病原菌的種類及其特性提供科學(xué)依據(jù)。方法：從受黑斑病病害侵染的番石榴果實上分離純化出一株優(yōu)勢病原菌，命名為S1，并對其進(jìn)行形態(tài)學(xué)觀察、致病性測定、生長特性研究及聚合酶鏈?zhǔn)椒磻?yīng)（Polymerase Chain Reaction，PCR）測序比對分析，較為全面的對分離出的優(yōu)勢病原菌S1進(jìn)行鑒定。結(jié)果：由形態(tài)學(xué)觀察可知，S1菌落的氣生菌絲蓬松且致密，顏色呈墨綠色或黑色，其孢子形態(tài)為單孢且透明，大小均勻，約為(6~10) μm×(6~7) μm，呈圓形或橢圓形，與《真菌鑒定手冊》比對，初步判定其屬葉點霉屬菌，將其反接于健康番石榴果實上進(jìn)行致病性測定。結(jié)果表明，S1為致使番石榴產(chǎn)生黑斑病的主導(dǎo)優(yōu)勢病原菌。通過生長特性研究表明，S1在偏堿性、黑暗、常溫條件下較為適宜生長，結(jié)合PCR測序結(jié)果，確定出致使番石榴黑斑病的優(yōu)勢病原菌S1為首都葉點霉（Phyllosticta capitalensis）。結(jié)論：引起番石榴黑斑病的優(yōu)勢病原菌為首都葉點霉。

關(guān)鍵詞: 番石榴  /  病原菌  /  鑒定  /  首都葉點霉  

Abstract: Objective: In order to provide scientific basis for further clarifying the species and characteristics of dominant pathogens causing guava black spot. Method: A dominant pathogen, named S1, was isolated and purified from the guava fruit infected with black spot disease, and subjected to morphology observation, pathogenicity determination, growth characteristics study and polymerase chain reaction (PCR) sequencing comparison analysis were conducted for comprehensive identification of the dominant pathogen S1. Results: According to the morphological observation, the aerial hyphae of the S1 colony were fluffy and dense, and the color was dark green or black. The spore morphology was monospore and transparent, and the size was uniform, about (6~10) μm×(6~7) μm, round or elliptical, compared with the "Fungus Identification Manual", it was preliminarily determined to belong to Phyllanthus spp., and once again inoculated to healthy guava fruits for pathogenicity determination. The results showed that S1 was the dominant pathogen causing guava black spot disease. Research on growth characteristics showed that S1 was more suitable for growth under alkaline, dark and normal temperature conditions. Moreover, S1 was identified as Phyllosticta capitalensis by the comparison of the PCR sequencing results. Conclusion: The dominant pathogen causing guava black spot disease was Phyllosticta capitalensis.

圖  1   分離純化菌落的形態(tài)

注：a：分離出的菌落；b：純化后菌落（正面）；c：純化后菌落（背面）。

Figure  1.   Morphology of isolated and purified colonies

圖  2   病原菌的顯微形態(tài)

注：a：菌絲體（100×）；b：分生孢子（200×）；c：分生孢子（400×）。

Figure  2.   Microscopic morphology of pathogens

圖  3   分離純化菌株對番石榴的致病性

注：a：致病性測定結(jié)果（5 d）；b：致病性測定結(jié)果（8 d）；c：致病性測定結(jié)果（14 d）；d：自然發(fā)病癥狀；e：致病性測定結(jié)果（空白組）。

Figure  3.   Pathogenicity of isolated and purified strains to guava

圖  4   pH對病原菌菌絲的影響

Figure  4.   Effect of pH on the hyphae of pathogenic bacteria

圖  5   光照對病原菌菌絲的影響

Figure  5.   Effect of light on the hyphae of pathogenic bacteria

圖  6   溫度對病原菌菌絲的影響

Figure  6.   Influence of temperature on the hyphae of pathogenic bacteria

圖  7   S1菌株P(guān)CR擴(kuò)增產(chǎn)物的瓊脂糖凝膠電泳圖

注：M：標(biāo)準(zhǔn)DNA分子質(zhì)量；1：菌株S1。

Figure  7.   Agarose gel electrophoresis of PCR amplification products of strain S1

圖  8   S1菌株基于rDNA-ITS序列構(gòu)建的系統(tǒng)發(fā)育樹

Figure  8.   Phylogenetic tree constructed by S1 strain based on rDNA-ITS sequence

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