Preliminary phospholipase, Hemolytic and Bactericidal activity of the venom of the Snake of Cascabel Tolima
Abstract
In the department of Tolima, there are no studies that allow us to specify with certainty the magnitude of the ophidic accident caused by Crotalus durissus; Therefore, there is a need to generate toxicological information, as well as knowledge of the potential antimicrobial potential of molecules such as proteins and peptides that make up the venom. In this work, we analyzed the electrophoretic profile by SDS-PAGE of crude venom extracted from individuals collected in the municipality of Natagaima (Tolima) and the association of its components with direct and indirect hemolytic activity on blood agar and its bactericidal activity. The potential of the crude venom against Escherichia coli, Staphylococcus aureus, and Pseudomonas aeuroginosa was evaluated. The venom showed molecular weight bands 26.6 kDa., 17, 14.2, 6.5, 3.5 and 1.06 kDa., corresponding to reports of the venom of this species. The direct and indirect hemolytic activity was dose-dependent, with a minimum hemolytic dose of 200 mg / ml. The antimicrobial effect of the crude venom was differential on the microorganisms evaluated, presenting moderate activity on E. coli. The results constitute valuable data to preliminarily recognize the antimicrobial potential of Crotalus durissus venom (rattle) from the Natagaima-Tolima area.
Keywords
Crotalus durissus, Bactericidal activity, Hemolysis, Phospholipases, Rattlesnake, Toxinology, Venom., Bactericidal activity, Rattlesnake, Crotalus durissus, Phospholipases, Hemolysis, Toxinology, Venom
References
[1] M. J. Sevilla-Sánchez, D. Mora-Obando, J. J. Calderón, J. A. Guerrero-Vargas, and S. Ayerbe-González, “Accidente ofídico en el departamento de Nariño, Colombia: análisis retrospectivo (2008-2017),” Biomédica, vol. 39, no. 4, May 2019, doi: 10.7705/biomedica.v39i4.4830.
[2] M. A. Bárcenas Rojas, “INFORME DE EVENTO ACCIDENTE OFÍDICO, COLOMBIA, 2017,” 2018.
[3] M. Acero and J. Usaquen, “INFORME DEL EVENTO ACCIDENTE OFIDICO HASTA EL PERIODO EPIDEMOLGICO XII, Colombia,2014,” vol. 2. Instituto Nacional de Salud, pp. 1–29, 2014.
[4] J. J. Calvete, L. Sanz, Y. Angulo, B. Lomonte, and J. M. Gutiérrez, “Venoms, venomics, antivenomics,” FEBS Lett., vol. 583, no. 11, pp. 1736–1743, Jun. 2009, doi: 10.1016/j.febslet.2009.03.029.
[5] M. S. R. Gomes et al., “BthMP: a new weakly hemorrhagic metalloproteinase from Bothrops moojeni snake venom,” Toxicon, vol. 53, no. 1, pp. 24–32, Jan. 2009, doi: 10.1016/J.TOXICON.2008.10.007.
[6] O. H. P. Ramos and H. S. Selistre-de-Araujo, “Snake venom metalloproteases — structure and function of catalytic and disintegrin domains,” Comp. Biochem. Physiol. Part C Toxicol. Pharmacol., vol. 142, no. 3–4, pp. 328–346, Mar. 2006, doi: 10.1016/J.CBPC.2005.11.005.
[7] R. Zouari-Kessentini et al., “Two purified and characterized phospholipases A2 from Cerastes cerastes venom, that inhibit cancerous cell adhesion and migration,” Toxicon, vol. 53, no. 4, pp. 444–453, Mar. 2009, doi: 10.1016/J.TOXICON.2009.01.003.
[8] A. C. Patiño, J. A. Pereañez, J. M. Gutiérrez, and A. Rucavado, “Biochemical and biological characterization of two serine proteinases from Colombian Crotalus durissus cumanensis snake venom,” Toxicon, vol. 63, pp. 32–43, Mar. 2013, doi: 10.1016/J.TOXICON.2012.11.010.
[9] J. Quintana-Castillo et al., “Characterization of the Venom of C. d. cumanesis of Colombia: Proteomic Analysis and Antivenomic Study,” Toxins (Basel)., vol. 10, no. 2, p. 85, Feb. 2018, doi: 10.3390/toxins10020085.
[10] I. M. . Francischetti, M. E. . Gombarovits, J. G. Valenzuela, C. R. Carlini, and J. A. Guimarães, “Intraspecific variation in the venoms of the South American rattlesnake (Crotalus durissus terrificus),” Comp. Biochem. Physiol. Part C Pharmacol. Toxicol. Endocrinol., vol. 127, no. 1, pp. 23–36, Aug. 2000, doi: 10.1016/S0742-8413(00)00129-8.
[11] F. E. Lozano Manrique, “purificacion, caracterizacion y actividad biologica de una l-aminoacido oxidasa presente en el veneno de la serpiente Botrops atrox (jergon),” 2005.
[12] F. Lazo, O. Málaga, A. Yarlequé, R. Severino, and S. Gutiérrez, “Actividad antimicrobiana de una flavoproteína aislada del veneno de la serpiente peruana Bothrops atrox (‘jergón’),” Rev. la Soc. Química del Perú, vol. 73, no. 4, pp. 197–207, 2007.
[13] J. Do Carmo Dietz, D. A. De Almeida, L. C. Cintra, B. F. R. De Oliveira, M. R. Magalhães, and R. S. A. Jesuíno, “Evaluation of the antibacterial activity of Crotalus durissus terrificus crude venom,” Cienc. Anim. Bras., no. 19, pp. 1–12, 2018, doi: 10.1590/1809-6891v19e-51322.
[14] J. L. Rheubert, M. F. Meyer, R. M. Strobel, M. A. Pasternak, and R. A. Charvat, “Predicting antibacterial activity from snake venom proteomes,” PLoS One, vol. 15, no. 1, p. e0226807, Jan. 2020, doi: 10.1371/journal.pone.0226807.
[15] M. M. Bradford, “A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding,” Anal. Biochem., vol. 72, no. 1–2, pp. 248–254, May 1976, doi: 10.1016/0003-2697(76)90527-3.
[16] C. A. Yábar Varas, Manual de procedimientos de electroforesis para proteínas y ADN, 1era. Lima, Perú., 2003.
[17] F. Lazo, E. Rodríguez, and A. Yarlequé, “Evaluación comparativa de dos métodos para determinar la actividad de fosfolipasa en veneno de serpientes.,” Rev. Biol. Trop., vol. 5, pp. 98–102, 1998, doi: https://doi.org/10.15381/rpb.v5i2.8325.
[18] J. A. Pereañez J., S. L. Jiménez., J. C. Quintana., V. Nuñez., M. Fernández., and Y. Restrepo., “Inhibición de las actividades proteolítica, coagulante y hemolítica indirecta inducidas por el veneno de Bothrops asper por extractos etanólicos de tres especies de heliconias,” Vitae (Medellín), pp. 157–164, 2008.
[19] J. M. Gutiérrez, F. Chaves, E. Rojas, J. Elizondo, C. Avila, and L. Cerdas, “Production of monovalent anti-Bothrops asper antivenom: development of immune response in horses and neutralizing ability.,” Rev. Biol. Trop., vol. 36, no. 2B, pp. 511–7, Nov. 1988.
[20] E. Habermann and K. L. Hardt, “A sensitive and specific plate test for the quantitation of phospholipases,” Anal. Biochem., vol. 50, no. 1, pp. 163–173, 1972, doi: 10.1016/0003-2697(72)90495-2.
[21] J. B. Patel et al., Methods for Dilution Antimicrobial Susceptibility Tests for Bacteria That Grow Aerobically, 10th ed., vol. 35, no. 2. Clinical and Laboratory Standarts Institute, 2015.
[22] S. R. Pritchard, M. Phillips, and K. Kailasapathy, “Identification of bioactive peptides in commercial Cheddar cheese,” 2010.
[23] C. Rivera, L. Flores, C. Pantigoso, and E. Escobar, “Aislamiento y caracterización de un péptido antibacteriano del veneno de Centruroides margaritatus,” Rev. Peru. Biol., vol. 17, no. 1, pp. 129–132, 2014, doi: 10.15381/rpb.v17i1.61.
[24] J. do C. Dietz, D. A. de Almeida, L. C. Cintra, B. F. R. de Oliveira, M. R. Magalhães, and R. S. A. Jesuíno, “EVALUATION OF THE ANTIBACTERIAL ACTIVITY OF Crotalus durissus terrificus CRUDE VENOM,” Ciência Anim. Bras., vol. 19, no. 0, Nov. 2018, doi: 10.1590/1809-6891v19e-51322.
[25] J. C. Quintana-Castillo, I. C. Ávila-Gómez, J. F. Ceballos-Ruiz, L. J. Vargas-Muñoz, and S. Estrada-Gómez, “Efecto citotóxico de fosfolipasas A2 del veneno de Crotalus durissus cumanensis de Colombia,” Rev. Investig. en Salud Univ. Boyacá, vol. 4, no. 1, p. 16, Jul. 2017, doi: 10.24267/23897325.194.
[26] T.-W. Wu and D. O. Tinker, “Phospholipase A2 fromo Crotalus atrox Venom.,” Biochemistry, vol. 8, pp. 1558–1568, 1969.
[27] E. Coles, D. L. Mcilwain, and M. M. Rapport, “The activity of pure phospholipase a2 from crotalus venom on myelin and on pure phospholipids atrox,” Biochimic, vol. 337, pp. 68–78, 1974.
[28] C. Remuzgo, M. P. Alvarez, F. Lazo, and A. Yarleque, “Caracterizacion parcial del veneno de la serpiente cascabel peruana crotalus durissus terrificus,” Rev. Peru. Biol., vol. 7, no. 1, pp. 67–73, Jun. 2000, doi: 10.15381/rpb.v7i1.6729.
[29] C. Ortiz, F. Lazo, C. Bellido, E. Gonzales, and A. Yarlequé, “Variaciones en las actividades enzimáticas del veneno de la serpiente Bothrops atrox "jergón", de tres zonas geográficas del Perú,” Rev. Peru. Med. Exp. Salud Publica, vol. 29, pp. 198–205, 2012, doi: 10.1590/S1726-46342012000200005.
[30] M. Holzer and S. P. Mackessy, “An aqueous endpoint assay of snake venom phospholipase A2,” Toxicon, vol. 34, no. 10, pp. 1149–1155, 1996, doi: 10.1016/0041-0101(96)00057-8.
[31] J. A. Pereañez et al., “Correlation of the inhibitory activity of phospholipase A2 snake venom and the antioxidant activity of Colombian plant extracts,” Brazilian J. Pharmacogn., vol. 20, no. 6, pp. 910–916, Dec. 2010, doi: 10.1590/S0102-695X2010005000030.
[32] C. Pirela, D. Salas, J. Carlos, L. Jim, and L. Hernández, “CARACTERIZACIÓN TOXINOLÓGICA DEL VENENO TOTAL DE LA SERPIENTE DE CASCABEL Crotalus durissus cumanensis ( VIPERIDAE ), PRESENTE EN LA LOCALIDAD DE PORSHOURE , Toxinological Characterization of the Whole Venom of the Rattlesnake Crotalus durissus cumanensis,” vol. XVI, pp. 232–238, 2006.
[33] A. Lourenço et al., “Individual venom profiling of Crotalus durissus terrificus specimens from a geographically limited region: crotamine assessment and captivity evaluation on the biological activities.,” Toxicon, vol. 69, pp. 75–81, Jul. 2013.
[34] J. G. Soto, J. C. Perez, and S. A. Minton, “Proteolytic , hemorrhagic and hemolytic activities of snake venoms,” Toxicon, vol. 26, pp. 875–882, 1988, doi: https://doi.org/10.1016/0041-0101(88)90328-5.
[35] M. C. Dos Santos, L. C. L. Ferreira, W. D. Da Silva, and M. de F. D. Furtado, “Caracterizacion de las actividades biologicas de los venenos ‘amarillo’ y ‘blanco’ de Crotalus durissus ruruima comparados con el veneno de Crotalus durissus terrificus. Poder neutralizante de los antivenenos frente a los venenos de Crotalus durissus ruruima,” Toxicon, vol. 31, no. 11, pp. 1459–1469, Nov. 1993, doi: 10.1016/0041-0101(93)90211-Z.
[36] L. J. Vargas et al., “An acidic phospholipase A2 with antibacterial activity from Porthidium nasutum snake venom,” Comp. Biochem. Physiol. - B Biochem. Mol. Biol., vol. 161, no. 4, pp. 341–347, Apr. 2012, doi: 10.1016/j.cbpb.2011.12.010.
[37] L. J. Vargas, J. C. Quintana, J. A. Pereañez, V. Núñez, L. Sanz, and J. Calvete, “Cloning and characterization of an antibacterial l-amino acid oxidase from Crotalus durissus cumanensis venom,” Toxicon, vol. 64, pp. 1–11, Mar. 2013, doi: 10.1016/J.TOXICON.2012.11.027.
[38] E. S. Yamane et al., “Unraveling the antifungal activity of a South American rattlesnake toxin crotamine,” Biochimie, vol. 95, no. 2, pp. 231–240, Feb. 2013, doi: 10.1016/J.BIOCHI.2012.09.019.
[39] N. Oguiura, M. Boni-Mitake, R. Affonso, and G. Zhang, “In vitro antibacterial and hemolytic activities of crotamine, a small basic myotoxin from rattlesnake Crotalus durissus,” J. Antibiot. (Tokyo)., vol. 64, no. 4, pp. 327–331, Apr. 2011, doi: 10.1038/ja.2011.10.
[40] A. J. Magro, R. J. DA Silva, P. R. R. Ramos, A. L. Cheruboni, and M. R. Hataide, “Intraspecific variation in the venom electrophoretic profile of recently captured Crotalus durissus terrificus (Laurenti, 1768) snakes,” J. Venom. Anim. Toxins, vol. 7, no. 2, pp. 276–301, Dec. 2001, doi: 10.1590/S0104-79302001000200010.