Cargando…

Purification and characterization of a novel defensin from the salivary glands of the black fly, Simulium bannaense

BACKGROUND: Black 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...

Descripción completa

Detalles Bibliográficos
Autores principales: Wei, Lin, Mu, Lixian, Wang, Yipeng, Bian, Hui, Li, Jun, Lu, Yiling, Han, Yi, Liu, Tong, Lv, Jing, Feng, Cuiping, Wu, Jing, Yang, Hailong
Formato: Online Artículo Texto
Lenguaje:English
Publicado: BioMed Central 2015
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4324660/
https://www.ncbi.nlm.nih.gov/pubmed/25649358
http://dx.doi.org/10.1186/s13071-015-0669-9
_version_ 1782356707231399936
author Wei, Lin
Mu, Lixian
Wang, Yipeng
Bian, Hui
Li, Jun
Lu, Yiling
Han, Yi
Liu, Tong
Lv, Jing
Feng, Cuiping
Wu, Jing
Yang, Hailong
author_facet Wei, Lin
Mu, Lixian
Wang, Yipeng
Bian, Hui
Li, Jun
Lu, Yiling
Han, Yi
Liu, Tong
Lv, Jing
Feng, Cuiping
Wu, Jing
Yang, Hailong
author_sort Wei, Lin
collection PubMed
description BACKGROUND: Black 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. METHODS: A 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. RESULTS: Surprisingly, 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. CONCLUSIONS: Our 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.
format Online
Article
Text
id pubmed-4324660
institution National Center for Biotechnology Information
language English
publishDate 2015
publisher BioMed Central
record_format MEDLINE/PubMed
spelling pubmed-43246602015-02-12 Purification and characterization of a novel defensin from the salivary glands of the black fly, Simulium bannaense Wei, Lin Mu, Lixian Wang, Yipeng Bian, Hui Li, Jun Lu, Yiling Han, Yi Liu, Tong Lv, Jing Feng, Cuiping Wu, Jing Yang, Hailong Parasit Vectors Research BACKGROUND: Black 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. METHODS: A 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. RESULTS: Surprisingly, 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. CONCLUSIONS: Our 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. BioMed Central 2015-02-04 /pmc/articles/PMC4324660/ /pubmed/25649358 http://dx.doi.org/10.1186/s13071-015-0669-9 Text en © Wei et al.; licensee BioMed Central. 2015 This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/4.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly credited. The Creative Commons Public Domain Dedication waiver (http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated.
spellingShingle Research
Wei, Lin
Mu, Lixian
Wang, Yipeng
Bian, Hui
Li, Jun
Lu, Yiling
Han, Yi
Liu, Tong
Lv, Jing
Feng, Cuiping
Wu, Jing
Yang, Hailong
Purification and characterization of a novel defensin from the salivary glands of the black fly, Simulium bannaense
title Purification and characterization of a novel defensin from the salivary glands of the black fly, Simulium bannaense
title_full Purification and characterization of a novel defensin from the salivary glands of the black fly, Simulium bannaense
title_fullStr Purification and characterization of a novel defensin from the salivary glands of the black fly, Simulium bannaense
title_full_unstemmed Purification and characterization of a novel defensin from the salivary glands of the black fly, Simulium bannaense
title_short Purification and characterization of a novel defensin from the salivary glands of the black fly, Simulium bannaense
title_sort purification and characterization of a novel defensin from the salivary glands of the black fly, simulium bannaense
topic Research
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4324660/
https://www.ncbi.nlm.nih.gov/pubmed/25649358
http://dx.doi.org/10.1186/s13071-015-0669-9
work_keys_str_mv AT weilin purificationandcharacterizationofanoveldefensinfromthesalivaryglandsoftheblackflysimuliumbannaense
AT mulixian purificationandcharacterizationofanoveldefensinfromthesalivaryglandsoftheblackflysimuliumbannaense
AT wangyipeng purificationandcharacterizationofanoveldefensinfromthesalivaryglandsoftheblackflysimuliumbannaense
AT bianhui purificationandcharacterizationofanoveldefensinfromthesalivaryglandsoftheblackflysimuliumbannaense
AT lijun purificationandcharacterizationofanoveldefensinfromthesalivaryglandsoftheblackflysimuliumbannaense
AT luyiling purificationandcharacterizationofanoveldefensinfromthesalivaryglandsoftheblackflysimuliumbannaense
AT hanyi purificationandcharacterizationofanoveldefensinfromthesalivaryglandsoftheblackflysimuliumbannaense
AT liutong purificationandcharacterizationofanoveldefensinfromthesalivaryglandsoftheblackflysimuliumbannaense
AT lvjing purificationandcharacterizationofanoveldefensinfromthesalivaryglandsoftheblackflysimuliumbannaense
AT fengcuiping purificationandcharacterizationofanoveldefensinfromthesalivaryglandsoftheblackflysimuliumbannaense
AT wujing purificationandcharacterizationofanoveldefensinfromthesalivaryglandsoftheblackflysimuliumbannaense
AT yanghailong purificationandcharacterizationofanoveldefensinfromthesalivaryglandsoftheblackflysimuliumbannaense