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In-depth proteomic analysis of Varroa destructor: Detection of DWV-complex, ABPV, VdMLV and honeybee proteins in the mite.

Erban T, Harant K, Hubalek M, Vitamvas P, Kamler M, Poltronieri P, Tyl J, Markovic M, Titera D - Sci Rep (2015)

Bottom Line: Isoforms of viral structural proteins of highest abundance were localized via 2D-E.The absence/scarce detection of non-structural proteins compared with high-abundance structural proteins suggest that the viruses did not replicate in the mite; hence, virions accumulate in the Varroa gut via hemolymph feeding.Hemolymph feeding also resulted in the detection of a variety of honeybee proteins.

View Article: PubMed Central - PubMed

Affiliation: Crop Research Institute, Prague 6, Czechia.

ABSTRACT
We investigated pathogens in the parasitic honeybee mite Varroa destructor using nanoLC-MS/MS (TripleTOF) and 2D-E-MS/MS proteomics approaches supplemented with affinity-chromatography to concentrate trace target proteins. Peptides were detected from the currently uncharacterized Varroa destructor Macula-like virus (VdMLV), the deformed wing virus (DWV)-complex and the acute bee paralysis virus (ABPV). Peptide alignments revealed detection of complete structural DWV-complex block VP2-VP1-VP3, VDV-1 helicase and single-amino-acid substitution A/K/Q in VP1, the ABPV structural block VP1-VP4-VP2-VP3 including uncleaved VP4/VP2, and VdMLV coat protein. Isoforms of viral structural proteins of highest abundance were localized via 2D-E. The presence of all types of capsid/coat proteins of a particular virus suggested the presence of virions in Varroa. Also, matches between the MWs of viral structural proteins on 2D-E and their theoretical MWs indicated that viruses were not digested. The absence/scarce detection of non-structural proteins compared with high-abundance structural proteins suggest that the viruses did not replicate in the mite; hence, virions accumulate in the Varroa gut via hemolymph feeding. Hemolymph feeding also resulted in the detection of a variety of honeybee proteins. The advantages of MS-based proteomics for pathogen detection, false-positive pathogen detection, virus replication, posttranslational modifications, and the presence of honeybee proteins in Varroa are discussed.

No MeSH data available.


Related in: MedlinePlus

2D-E 14% pI 3-10 analysis of protein fractions from Varroa destructor with localized virus structural proteins.Legend: (A) p-ABA-unbound fraction, (B) p-ABA-purified fraction, (C) supernatant-before-purification (e.g., total soluble proteome). Lowercase marked figures (a–c) show the details of ~23–24 kDa viral spots in the corresponding fractions. For details on the viral spots identified using MALDI TOF/TOF, see Table 2; and for details on MS/MS identification, see Table Supplement 2.
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f2: 2D-E 14% pI 3-10 analysis of protein fractions from Varroa destructor with localized virus structural proteins.Legend: (A) p-ABA-unbound fraction, (B) p-ABA-purified fraction, (C) supernatant-before-purification (e.g., total soluble proteome). Lowercase marked figures (a–c) show the details of ~23–24 kDa viral spots in the corresponding fractions. For details on the viral spots identified using MALDI TOF/TOF, see Table 2; and for details on MS/MS identification, see Table Supplement 2.

Mentions: Normalized total spectral counts (Table Supplement 1C and 3C) from Scaffold provided quantitative values in each 1D-E slice (BioSample) analyzed. The source for p-ABA affinity fractions C and D was fraction B (Fig. 1); three 1D-E slices were analyzed from each of these fractions. The spectral counts were found to differ between the three fractions. The highest spectral counts for DWV polyprotein (Cluster 1) and coat protein VdMLV (Clusters 3–5) were found in the p-ABA-unbound fraction (D). Stronger spots for the DWV and VdMLV structural proteins were also detected in the 2D-E of fraction D (Fig. 2A) than in the 2D-E of the p-ABA-purified fraction C (Fig. 2B). However, the sums of the spectral counts were similar for ABPV capsid protein in all three fractions.


In-depth proteomic analysis of Varroa destructor: Detection of DWV-complex, ABPV, VdMLV and honeybee proteins in the mite.

Erban T, Harant K, Hubalek M, Vitamvas P, Kamler M, Poltronieri P, Tyl J, Markovic M, Titera D - Sci Rep (2015)

2D-E 14% pI 3-10 analysis of protein fractions from Varroa destructor with localized virus structural proteins.Legend: (A) p-ABA-unbound fraction, (B) p-ABA-purified fraction, (C) supernatant-before-purification (e.g., total soluble proteome). Lowercase marked figures (a–c) show the details of ~23–24 kDa viral spots in the corresponding fractions. For details on the viral spots identified using MALDI TOF/TOF, see Table 2; and for details on MS/MS identification, see Table Supplement 2.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4566121&req=5

f2: 2D-E 14% pI 3-10 analysis of protein fractions from Varroa destructor with localized virus structural proteins.Legend: (A) p-ABA-unbound fraction, (B) p-ABA-purified fraction, (C) supernatant-before-purification (e.g., total soluble proteome). Lowercase marked figures (a–c) show the details of ~23–24 kDa viral spots in the corresponding fractions. For details on the viral spots identified using MALDI TOF/TOF, see Table 2; and for details on MS/MS identification, see Table Supplement 2.
Mentions: Normalized total spectral counts (Table Supplement 1C and 3C) from Scaffold provided quantitative values in each 1D-E slice (BioSample) analyzed. The source for p-ABA affinity fractions C and D was fraction B (Fig. 1); three 1D-E slices were analyzed from each of these fractions. The spectral counts were found to differ between the three fractions. The highest spectral counts for DWV polyprotein (Cluster 1) and coat protein VdMLV (Clusters 3–5) were found in the p-ABA-unbound fraction (D). Stronger spots for the DWV and VdMLV structural proteins were also detected in the 2D-E of fraction D (Fig. 2A) than in the 2D-E of the p-ABA-purified fraction C (Fig. 2B). However, the sums of the spectral counts were similar for ABPV capsid protein in all three fractions.

Bottom Line: Isoforms of viral structural proteins of highest abundance were localized via 2D-E.The absence/scarce detection of non-structural proteins compared with high-abundance structural proteins suggest that the viruses did not replicate in the mite; hence, virions accumulate in the Varroa gut via hemolymph feeding.Hemolymph feeding also resulted in the detection of a variety of honeybee proteins.

View Article: PubMed Central - PubMed

Affiliation: Crop Research Institute, Prague 6, Czechia.

ABSTRACT
We investigated pathogens in the parasitic honeybee mite Varroa destructor using nanoLC-MS/MS (TripleTOF) and 2D-E-MS/MS proteomics approaches supplemented with affinity-chromatography to concentrate trace target proteins. Peptides were detected from the currently uncharacterized Varroa destructor Macula-like virus (VdMLV), the deformed wing virus (DWV)-complex and the acute bee paralysis virus (ABPV). Peptide alignments revealed detection of complete structural DWV-complex block VP2-VP1-VP3, VDV-1 helicase and single-amino-acid substitution A/K/Q in VP1, the ABPV structural block VP1-VP4-VP2-VP3 including uncleaved VP4/VP2, and VdMLV coat protein. Isoforms of viral structural proteins of highest abundance were localized via 2D-E. The presence of all types of capsid/coat proteins of a particular virus suggested the presence of virions in Varroa. Also, matches between the MWs of viral structural proteins on 2D-E and their theoretical MWs indicated that viruses were not digested. The absence/scarce detection of non-structural proteins compared with high-abundance structural proteins suggest that the viruses did not replicate in the mite; hence, virions accumulate in the Varroa gut via hemolymph feeding. Hemolymph feeding also resulted in the detection of a variety of honeybee proteins. The advantages of MS-based proteomics for pathogen detection, false-positive pathogen detection, virus replication, posttranslational modifications, and the presence of honeybee proteins in Varroa are discussed.

No MeSH data available.


Related in: MedlinePlus