Purification and characterization of a novel defensin from the salivary glands of the black fly, Simulium bannaense.
Purification and characterization of a novel defensin from the salivary glands of the black fly, Simulium bannaense.
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黑蝇唾液腺中新型防御素的纯化和表征
DOI：
10.1186/s13071-015-0669-9
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发表时间：
2015-02-04
影响因子：
3.2
通讯作者：
Yang H
中科院分区：
文献类型：
--
作者：
Wei L;Mu L;Wang Y;Bian H;Li J;Lu Y;Han Y;Liu T;Lv J;Feng C;Wu J;Yang H
研究背景：蚋属双翅目：蚋科，是一种能引起过敏反应和传播病原体的吸血昆虫。虽然他们的唾液已被认为含有各种各样的生理活性分子，很少有信息是可用的黑飞唾液腺中的抗菌因子，特别是没有防御素已报道至今.MethodsA新的阳离子防御素指定SibaDef纯化使用反相高效液相色谱法（RP-HPLC）从黑飞的Simuelibannaense唾液腺。通过自动Edman降解和cDNA测序的组合方法确定SibaDef的氨基酸序列。通过扫描电子显微镜（SEM）评估SibaDef处理的革兰氏阳性菌金黄色葡萄球菌或枯草芽孢杆菌的形态变化。采用定量PCR（qPCR）技术分析了昆虫经口感染沙门氏菌后SibaDef mRNA在昆虫体内的表达。金黄色葡萄球菌或B。结果令人惊讶的是，防御素相关的氨基酸序列的系统发育分析表明，SibaDef是最密切相关的防御素从人体虱人体虱（Anoplura：Pediculidae），而不是其他双翅目防御素。SibaDef对革兰氏阳性菌具有强效抗菌活性，最小抑菌浓度（MIC）范围为0.83 μM至2.29 μM。SEM分析表明，SibaDef通过破坏细胞膜完整性杀死微生物。SibaDef在细菌免疫果蝇中的转录水平随时间的推移而增加，在36 h时达到最大值，然后从该时间点开始缓慢下降。bannaense，它可能在抵御糖和血餐中的微生物方面发挥重要作用。
BackgroundBlack flies (Diptera: Simuliidae) are haematophagous insects that can cause allergic reactions and act as vectors of pathogens. Although their saliva has been thought to contain a diverse array of physiologically active molecules, little information is available on antimicrobial factors in black fly salivary glands, especially no defensins have been reported so far.MethodsA novel cationic defensin designated SibaDef was purified using reverse phase high-performance liquid chromatography (RP-HPLC) from the salivary glands of the black fly Simulium bannaense. The amino acid sequence of SibaDef was determined by a combination method of automated Edman degradation and cDNA sequencing. The morphologic changes of Gram-positive bacteria Staphylococcus aureus or Bacillus subtilis treated with SibaDef were assessed by scanning electron microscopy (SEM). Quantitative PCR (qPCR) was performed to analyze the expression of SibaDef mRNA in whole bodies of insects after oral infection with the bacteria S. aureus or B. subtilis.ResultsSurprisingly, the phylogenetic analysis of defensin-related amino acid sequences demonstrated that SibaDef is most closely related to defensins from the human body louse Pediculus humanus corporis (Anoplura: Pediculidae), rather than to other dipteran defensins. SibaDef showed potent antimicrobial activities against Gram-positive bacteria with minimal inhibitory concentrations (MICs) ranging from 0.83 μM to 2.29 μM. SEM analysis indicated that SibaDef killed microorganisms through the disruption of cell membrane integrity. The transcript levels of SibaDef in the bacteria-immunized flies increased with the time course, reaching maximum at 36 h and then slowly decreased from that time point.ConclusionsOur results indicate that SibaDef is involved in the innate humoral response of the black fly S. bannaense, and it might play a significant role in the defence against microorganisms in both sugar and blood meals.
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影响因子：
1.8
作者：
Kuntal BK;Aparoy P;Reddanna P
通讯作者：
Reddanna P
影响因子：
3.2
作者：
Nasar F;Haddow AD;Tesh RB;Weaver SC
通讯作者：
Weaver SC
影响因子：
2.9
作者：
BJELLQVIST, B;HUGHES, GJ;HOCHSTRASSER, D
通讯作者：
HOCHSTRASSER, D
影响因子：
0.7
作者：
Bulet, P
通讯作者：
Bulet, P
影响因子：
14.9
作者：
Chenna, R;Sugawara, H;Thompson, JD
通讯作者：
Thompson, JD