- Email: [email protected]
257. Comparative Analysis of Transcriptomes of Phoneutria pertyi and P. nigriventer Venom Glands
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Abstracts Toxins 2012 / Toxicon 60 (2012) 95–248
Methods: An analytical study was designed to compare the physicochemical properties of 3 separate batches of CroFabÒ when reconstituted using the standard procedure (10 mL WFI with gentle swirling) and a modified rapid procedure using 18 mL 0.9% saline and manual inversion. The physical and chemical characteristics of the same 3 batches were assessed using various analytic methodologies associated with routine quality control release testing. In addition further analytical methodologies were applied in order to elucidate possible structural changes that may be induced by the changed reconstitution procedure. Results: Batches A, B, and C required mean reconstitution times of 25 min 51 secs using the label method and 3 mins 07 secs (88.0% mean decrease) using the modified method. Physicochemical characteristics (color and clarity, pH, purity, protein content, potency) were found to be highly comparable. Characterization assays (dynamic light scattering, analytical ultracentrifugation, LC-MS, SDS-PAGE and circular dichroism spectroscopy were also all found to be comparable between methods. Discussion: When comparing CroFabÒ batches that were reconstituted using the labeled and modified methods, the physicochemical and biological (potency) characteristics of CroFabÒ were not significantly changed when challenged by the various standard analytical methodologies applied in routine quality control analysis. Additionally, no changes in the CroFabÒ molecule regarding degradation, aggregation, purity, structure, or mass were observed. Conclusion: The analyses performed support the use of the more rapid reconstitution method using 18 mL 0.9% saline in order to allow a significantly reduced time to administration of CroFabÒ to patients in need. Keywords: Antivenoms, polyvalent immune fab, crotalid envenomation 10.1016/j.toxicon.2012.04.257
O. Spiders 257. Comparative Analysis of Transcriptomes of Phoneutria pertyi and P. nigriventer Venom Glands Marcelo R.V. Diniz 1, Camilla R.L. Machado 1, Mauricio A. Mudado 2, Ana Luiza B. Paiva 1 1 Centro de Pesquisa e Desenvolvimento Carlos Ribeiro Diniz, Fundação Ezequiel Dias, Belo Horizonte, Brazil 2 Empresa Brasileira de Pesquisa Agropecuaria – Embrapa, São Carlos, Brazil E-mail address: [email protected] (M.R.V. Diniz).
Background: Species of the genus Phoneutria known as "aranha-armadeira" or armed spiders are responsible for a large number of spider bites in Brazil. Until recently, all studies on the venom of the genus have been restricted to the species P. nigriventer. However, some recent proteomic studies revealed that other species venoms also contains a wide variety of proteins and peptides, including neurotoxins which act on the ion channels and chemical receptors of the neuro-muscular systems of insects and mammals. Thus, these venoms have emerged as invaluable tools for research, drug discovery and drug development with application in medicine and agriculture.
Methods: In order to find novel venom components with biological activity and to provide a database for comparative study with the previously described P. nigriventer venom gland transcriptome, we constructed a plasmidial cDNA library from the species Phoneutria pertyi mRNA venom gland to generate Expressed Sequence Tags (ESTs) data. Results: After editing, 710 good quality reads were clustered and 295 unique sequences were obtained (106 contigs and 189 singlets). Of these, 197 (67%) had a high degree of homology to spiders toxins deposited in the Uniprot database, most are P.nigriventer toxins isoforms. We observed that P. pertyi venom gland transcriptome were more abundant in insecticidal toxin sequences than P. nigriventer one. We also found new sequences for putative toxins in P. pertyi transcriptome, indicating that they can be novel toxins. Conclusions: These results show that although these spider venoms contain a similar range of toxins isoforms, the expression levels of each type of toxins are different, and also they contain toxins with unique sequences, what can suggest adaptation to different environments. Keywords: Phoneutria venom gland, transcriptome, neurotoxin 10.1016/j.toxicon.2012.04.258
258. Unusual Cases of the Spider Cheiracanthium Punctorium Biting in Volgograd Region, Russia Vasiliy I. Emtsov 1, Yury N. Ostapenko 2, Sergey S. Larionov 1 1 Volgograd Regional Poisoning Treatment Centre, Volgograd Regional Narcological Hospital, Volgograd, Russian Federation 2 Research and Applied Toxicology Centre, Federal Medical and Biological Agency, Moscow, Russian Federation E-mail address: [email protected] (Y.N. Ostapenko).
Background: Beginning in 2009 cases of bites by the spider Cheiracanthium punctorium, were reported in the Volgograd region of Russia. There had been no previous reports of bites by this species in that geographic area. The aim of this study was characterization of the epidemiology and clinical picture of such envenomations. Methods: Retrospective analysis of 19 case reports of inpatients in Volgograd Regional Poisoning Treatment Center in 2010-2011. Only cases of confirmed bites by Cheiracanthium punctorium were included. Identification of the spider was made by qualified specialists in the Volgograd State University. Results: Cases of biting of poisonous animals in Russia are responsible for 1,9% of all poisoning cases in patients admitted to toxicological treatment centers. According to data from 29 centers in 2010 549 (43,4%) patients were admitted with snake bites, 693 (54,8%) with poisonous insect and spider bites, and 23 (1,8%) after contact with poisonous sea animals. Bites of the common viper, Vipera Berus, occur over the entire country. In addition, in territories bordering Central Asian countries, including Volgograd region, the bites by tarantulas and black widow spiders are regularly reported. The number of patients admitted to the Volgograd Regional Poisoning Treatment Centre with poisonous spider biting for the last two years was 111,
Abstracts Toxins 2012 / Toxicon 60 (2012) 95–248
Methods: An analytical study was designed to compare the physicochemical properties of 3 separate batches of CroFabÒ when reconstituted using the standard procedure (10 mL WFI with gentle swirling) and a modified rapid procedure using 18 mL 0.9% saline and manual inversion. The physical and chemical characteristics of the same 3 batches were assessed using various analytic methodologies associated with routine quality control release testing. In addition further analytical methodologies were applied in order to elucidate possible structural changes that may be induced by the changed reconstitution procedure. Results: Batches A, B, and C required mean reconstitution times of 25 min 51 secs using the label method and 3 mins 07 secs (88.0% mean decrease) using the modified method. Physicochemical characteristics (color and clarity, pH, purity, protein content, potency) were found to be highly comparable. Characterization assays (dynamic light scattering, analytical ultracentrifugation, LC-MS, SDS-PAGE and circular dichroism spectroscopy were also all found to be comparable between methods. Discussion: When comparing CroFabÒ batches that were reconstituted using the labeled and modified methods, the physicochemical and biological (potency) characteristics of CroFabÒ were not significantly changed when challenged by the various standard analytical methodologies applied in routine quality control analysis. Additionally, no changes in the CroFabÒ molecule regarding degradation, aggregation, purity, structure, or mass were observed. Conclusion: The analyses performed support the use of the more rapid reconstitution method using 18 mL 0.9% saline in order to allow a significantly reduced time to administration of CroFabÒ to patients in need. Keywords: Antivenoms, polyvalent immune fab, crotalid envenomation 10.1016/j.toxicon.2012.04.257
O. Spiders 257. Comparative Analysis of Transcriptomes of Phoneutria pertyi and P. nigriventer Venom Glands Marcelo R.V. Diniz 1, Camilla R.L. Machado 1, Mauricio A. Mudado 2, Ana Luiza B. Paiva 1 1 Centro de Pesquisa e Desenvolvimento Carlos Ribeiro Diniz, Fundação Ezequiel Dias, Belo Horizonte, Brazil 2 Empresa Brasileira de Pesquisa Agropecuaria – Embrapa, São Carlos, Brazil E-mail address: [email protected] (M.R.V. Diniz).
Background: Species of the genus Phoneutria known as "aranha-armadeira" or armed spiders are responsible for a large number of spider bites in Brazil. Until recently, all studies on the venom of the genus have been restricted to the species P. nigriventer. However, some recent proteomic studies revealed that other species venoms also contains a wide variety of proteins and peptides, including neurotoxins which act on the ion channels and chemical receptors of the neuro-muscular systems of insects and mammals. Thus, these venoms have emerged as invaluable tools for research, drug discovery and drug development with application in medicine and agriculture.
Methods: In order to find novel venom components with biological activity and to provide a database for comparative study with the previously described P. nigriventer venom gland transcriptome, we constructed a plasmidial cDNA library from the species Phoneutria pertyi mRNA venom gland to generate Expressed Sequence Tags (ESTs) data. Results: After editing, 710 good quality reads were clustered and 295 unique sequences were obtained (106 contigs and 189 singlets). Of these, 197 (67%) had a high degree of homology to spiders toxins deposited in the Uniprot database, most are P.nigriventer toxins isoforms. We observed that P. pertyi venom gland transcriptome were more abundant in insecticidal toxin sequences than P. nigriventer one. We also found new sequences for putative toxins in P. pertyi transcriptome, indicating that they can be novel toxins. Conclusions: These results show that although these spider venoms contain a similar range of toxins isoforms, the expression levels of each type of toxins are different, and also they contain toxins with unique sequences, what can suggest adaptation to different environments. Keywords: Phoneutria venom gland, transcriptome, neurotoxin 10.1016/j.toxicon.2012.04.258
258. Unusual Cases of the Spider Cheiracanthium Punctorium Biting in Volgograd Region, Russia Vasiliy I. Emtsov 1, Yury N. Ostapenko 2, Sergey S. Larionov 1 1 Volgograd Regional Poisoning Treatment Centre, Volgograd Regional Narcological Hospital, Volgograd, Russian Federation 2 Research and Applied Toxicology Centre, Federal Medical and Biological Agency, Moscow, Russian Federation E-mail address: [email protected] (Y.N. Ostapenko).
Background: Beginning in 2009 cases of bites by the spider Cheiracanthium punctorium, were reported in the Volgograd region of Russia. There had been no previous reports of bites by this species in that geographic area. The aim of this study was characterization of the epidemiology and clinical picture of such envenomations. Methods: Retrospective analysis of 19 case reports of inpatients in Volgograd Regional Poisoning Treatment Center in 2010-2011. Only cases of confirmed bites by Cheiracanthium punctorium were included. Identification of the spider was made by qualified specialists in the Volgograd State University. Results: Cases of biting of poisonous animals in Russia are responsible for 1,9% of all poisoning cases in patients admitted to toxicological treatment centers. According to data from 29 centers in 2010 549 (43,4%) patients were admitted with snake bites, 693 (54,8%) with poisonous insect and spider bites, and 23 (1,8%) after contact with poisonous sea animals. Bites of the common viper, Vipera Berus, occur over the entire country. In addition, in territories bordering Central Asian countries, including Volgograd region, the bites by tarantulas and black widow spiders are regularly reported. The number of patients admitted to the Volgograd Regional Poisoning Treatment Centre with poisonous spider biting for the last two years was 111,