Toxins from spider venom Crossopriza lyoni were subjected to purify on a Sepharose CL-6B 200
column. These were investigated for its antibacterial and antifungal activity
against 13 infectious microbial pathogenic strains. Antimicrobial
susceptibility was determined by using paper disc diffusion and serial micro-dilution
assays. Triton X-100 (0.1%) proved to be a good solubilizing agent for
toxin/proteins. Higher protein solubilization was observed in the supernatant
than in the residue, except TCA. The elution pattern of purified and
homogenized sting glands displayed two major peaks at 280 nm. First one was
eluted in fraction no. 43 - 51 while second one after fraction no. 61 - 90.
From gel filtration chromatography total yield of protein obtained was 67.3%.
From comparison of gel chromatographs eluted toxins peptide molecular weight
was ranging from 6.2 - 64 kD. Toxin peptides have shown lower MIC values i.e. 7.5 - 15 μg/ml against K.
pneumoniae, E. coli, L. acidophilus, B. cereus;
against S. aureus and M.
luteus that the broad spectrum antibiotics i.e. tetracycline and ampicillin. In tests, larger inhibition zone
diameter was obtained in comparison to control. Diameter of inhibition zones
obtained in spider toxins at a concentration range of197.12 - 0.96 g/mlfor E. coli was 17.86 ± 0.21, Bacillus cereus 19.13 ± 0.21, L. acidophilus 16.83 ± 0.25, Micrococcus luteus 18.46 ± 0.17, S. aeurus 16.23 ± 0.19, Klebsiella pneumoniae 21.83 ± 0.16, Salmonella typhi 16.16 ± 0.21, Vibrio cholera 18.66 ± 0.21, Pseudomonas aeruginosa 18.66 ± 0.21, Aspergillus niger 22.9 ± 0.24, Candida albicans 24.66 ± 0.28, Rhizopus stolonifer 21.1 ± 0.16. Spider
toxins generate cytotoxic effect on bacterial cells that results in heavy cell
death. No doubt spider toxins can be used as alternate of broad spectrum
antibiotics.
References
[1]
Coddington, J.A. and Levi, H.W. (1991) Systematics and Evolution of spider (Araneae). Annual Review of Ecology and Systematics, 22, 565-592. http://dx.doi.org/10.1146/annurev.es.22.110191.003025
[2]
Luo, J., Zhang, Y., Gong, M., Lu, S., Ma, Y., Zeng, X. and Liang, S. (2014) Molecular Surface of JZTX-V (β-Thera- photoxin-Cj2a) Interacting with Voltage-Gated Sodium Channel Subtype NaV1.4. Toxins (Basel), 6, 2177-2193. http://dx.doi.org/10.3390/toxins6072177
[3]
Anand, P., Grigoryan, A., Bhuiyan, M.H., Ueberheide, B., Russell, V., Quinonez, J., Moy, P., Chait, B.T., Poget, S.F. and Holford, M. (2014) Sample Limited Characterization of a Novel Disulfide-Rich Venom Peptide Toxin from Terebrid Marine Snail Terebra variegata. PLoS One, 9, e94122. http://dx.doi.org/10.1371/journal.pone.0094122
[4]
Sapag, A., Salinas-Luypaert, C. and Constenla-Monoz, C. (2014) First Report of the Vitro Selection of the RNA Aptamers Targated to Recombinant Loxosceles Laeta Spider Toxins. Biological Research, 47, 0717-6287.
[5]
Liu, H., Yu, H., Yan, Y., Duan, Z. and Wang, X. (2014) Detection and Identification of Huwentoxin-IV Interacting Proteins by Biotin-Avidin Chemistry Combined with Mass Spectrometry. Journal of Venomous Animals and Toxins including Tropical Diseases, 20, 18.
[6]
Ahmed, A. and Beg, A.Z. (2001) Antimicrobial and Phytochemical Studies on 45 Indian Medicinal Plants against Multi-Drug Resistant Human Pathogens. Journal of Ethopharmocology, 74, 113-123. http://dx.doi.org/10.1016/S0378-8741(00)00335-4
[7]
Hu, Z., Zhou, X., Chen, J., Tang, C., Xiao, Z., Ying, D., Liu, Z. and Liang, S. (2014) The Venom of the Spider Selenocosmia jiafu Contains Various Neurotoxins Acting on Voltage-Gated Ion Channels in Rat Dorsal Root Ganglion neurons. Toxins (Basel), 6, 988-1001. http://dx.doi.org/10.3390/toxins6030988
[8]
Lowry, O.H., Rosenbrough, N.J., Farr, A.L. and Randall, R.J. (1951) Protein Measurement with Phenol Reagent. The Journal of Biological Chemistry, 193, 265-275.
[9]
Spier, R.E. (1982) Gel Filtration Column: Animal Cell Technology: An Overview. Journal of Chemical Technology and Biotechnology, 32, 304-312. http://dx.doi.org/10.1002/jctb.5030320134
[10]
NCCLS (National Committee for Clinical Laboratory Standards) (1993) Performance Standard for Antimicrobial Disc Susceptibility Tests. Approved Standard. National Committee for Clinical Laboratory Standards, Villanova, P.A. Publication M2-A5. USA.
[11]
Amsterdam, D. (1996) Susceptibility Testing of Antimicrobials in Liquid Medium. In: Baltimore, L.V., Williams, M.D. and Wilikins, Eds., Antibiotics in Laboratoratory Medicine, 4th Edition, 52-111.
[12]
Wang, X., Yu, H., Liu, H., Yan, Y. and Duan, Z. (2014) Detection and Identification of Huwentoxin-IV Interacting Proteins by Biotin-Avidin Chemistry Combined with Mass Spectrometry. Journal of Venomous Animals and Toxins including Tropical Diseases, 20, 18.
[13]
Dubovskii, P.V., Vassilevski, A.A., Kozlov, S.A., Feofanov, A.V., Grishin, E.V. and Efremov, R.G. (2015) Latarcins: Versatile Spider Venom Peptides. Cellular and Molecular Life Sciences, 72, 4501-4522. http://dx.doi.org/10.1007/s00018-015-2016-x
[14]
Lei, Q., Yu, H., Peng, X., Yan, S., Wang, J., Yan, Y. and Wang, X. (2015) Isolation and Preliminary Characterization of Proteinaceous Toxins with Insecticidal and Antibacterial Activities from Black Widow Spider (L. tredecimguttatus) Eggs. Toxins (Basel), 16, 886-899.
[15]
Kuznetsov, A.S., Dubovskii, P.V., Vorontsova, O.V., Feofanov, A.V. and Efremov, R.G. (2014) Interaction of Linear Cationic Peptides with Phospholipid Membranes and Polymers of Sialic Acid. Biochemistry (Mosc), 79, 459-468. http://dx.doi.org/10.1134/S0006297914050101
[16]
Johnson, E.T., Dowd, P.F. and Hughes, S.R. (2014) Expression of a Wolf Spider Toxin in Tobacco Inhibits the Growth of Microbes and Insects. Biotechnology Letters, 36, 1735-1742. http://dx.doi.org/10.1007/s10529-014-1536-z
[17]
Tan, H., Ding, X., Meng, S., Liu, C., Wang, H., Xia. L., Liu, Z. and Liang, S. (2013) Antimicrobial Potential of Lycosin-I, a Cationic and Amphiphilic Peptide from the Venom of the Spider Lycosa singorensis. Current Molecular Medicine, 13, 900-910. http://dx.doi.org/10.2174/15665240113139990045
[18]
Chen, R. and Chung, S.H. (2013) Effect of Gating Modifier Toxins on Membrane Thickness: Implications for Toxin Effect on Gramicidin and Mechanosensitive Channels. Toxins (Basel), 5, 456-471.
[19]
Garcia, F., Villegas, E., Espino-Solis, G.P., Rodriguez, A., Paniagua-Solis, J.F., Sandoval-Lopez, G., Possani, L.D. and Corzo, G. (2013) Antimicrobial Peptides from Arachnid Venoms and Their Microbicidal Activity in the Presence of Commercial Antibiotics. The Journal of Antibiotics (Tokyo), 66, 3-10. http://dx.doi.org/10.1038/ja.2012.87
[20]
Idiong, G., Won, A., Ruscito, A., Leung, B.O., Hitchcock, A.P. and Ianoul, A. (2011) Investigating the Effect of a Single Glycine to Alanine Substitution on Interactions of Antimicrobial Peptide Latarcin 2a with a Lipid Membrane. European Biophysics Journal, 40, 1087-1100. http://dx.doi.org/10.1007/s00249-011-0726-z
[21]
Won, A., Khan, M., Gustin, S., Akpawu, A., Seebun, D., Avis, T.J. and Hitchcock, A.P. (2011) Investigating the Effects of L- to D-Amino Acid Substitution and Deamidation on the Activity and Membrane Interactions of Antimicrobial Peptide Anoplin. Biochimica et Biophysica Acta (BBA)—Biomembranes, 1808, 1592-1600. http://dx.doi.org/10.1016/j.bbamem.2010.11.010
[22]
Vassilevski, A.A., Kozlov, S.A., Samsonova, O.V., Egorova, N.S., Karpunin, D.V., Pluzhnikov, K.A., Feofanov, A.V. and Grishin, E.V. (2008) Cyto-Insectotoxins, a Novel Class of Cytolytic and Insecticidal Peptides from Spider Venom. Biochemical Journal, 411, 687-696. http://dx.doi.org/10.1042/BJ20071123
[23]
Vorontsova, O.V., Egorova, N.S., Arseniev, A.S. and Feofanov, A.V. (2011) Haemolytic and Cytotoxic Action of Latarcin Ltc2a. Biochimie, 93, 227-241. http://dx.doi.org/10.1016/j.biochi.2010.09.016
[24]
Ayroza, G., Ferreira, I.L., Sayegh, R.S., Tashima, A.K. and da Silva Junior, P.I. (2012) Juruin: An Antifungal Peptide from the Venom of the Amazonian Pink Toe Spider, Avicularia juruensis, which Contains the Inhibitory Cystine Knot Motif. Frontiers in Microbiology, 3, Article 324. http://dx.doi.org/10.3389/fmicb.2012.00324
[25]
Pukala, T.L., Doyle, J.R., Llewellyn, L.E., Kuhn-Nentwig, L., Apponyi, M.A., Separovic, F. and Bowie, J.H. (2007) Cupiennin 1a, an Antimicrobial Peptide from the Venom of the Neotropical Wandering Spider Cupiennius salei, Also Inhibits the Formation of Nitric Oxide by Neuronal Nitric Oxide Synthase. The FEBS Journal, 274, 1778-1784. http://dx.doi.org/10.1111/j.1742-4658.2007.05726.x
[26]
Kuhn-Nentwig, L., Willems, J., Seebeck, T., Shalaby, T., Kaiser, M. and Nentwig, W. (2011) Cupiennin 1a Exhibits a Remarkably Broad, Non-Stereospecific Cytolytic Activity on Bacteria, Protozoan Parasites, Insects, and Human Cancer cells. Amino Acids, 40, 69-76. http://dx.doi.org/10.1007/s00726-009-0471-0
[27]
Santos, D.M., Verly, R.M., Piló-Veloso, D., de Maria, M., de Carvalho, M.A., Cisalpino, P.S. and Soares, B.M. (2010) LyeTx I, a Potent Antimicrobial Peptide from the Venom of the Spider Lycosa erythrognatha. Amino Acids, 39, 135-144. http://dx.doi.org/10.1007/s00726-009-0385-x
[28]
Shlyapnikov, Y.M., Andreev, Y.A., Kozlov, S.A., Vassilevski, A.A. and Grishin, E.V. (2008) Bacterial Production of Latarcin 2a, a Potent Antimicrobial Peptide from Spider Venom. Protein Expression and Purification, 60, 89-95. http://dx.doi.org/10.1016/j.pep.2008.03.011
[29]
Ali, M.P., Yoshimatsu, K., Suzuki, T., Kato, T. and Park, E.Y. (2014) Expression and Purification of Cyto-Insectotoxin (Cit1a) Using Silkworm Larvae Targeting for an Antimicrobial Therapeutic Agent. Applied Microbiology and Biotechnology, 98, 6973-6982. http://dx.doi.org/10.1007/s00253-014-5728-1
