Multielemental analyses of Lonomia obliqua (Lepidoptera, Saturniidae) caterpillar using EDXRF and INAA techniques

In this study, multielemental analysis of Lonomia obliqua (Lepidoptera, Saturniidae) caterpillar was performed using energy dispersive X-ray fluorescence and instrumental neutron activation analysis techniques. This caterpillar is poisonous and has the ability to cause fatal hemorrhagic effects in humans after contact. The need of this study is related to morphological changes (mainly size and color) observed in some caterpillars used for preparation of antilonomic serum (antivenom). The samples were classified as healthy (caterpillars of control) and unhealthy (caterpillars visibly modified). The XRF measurements were performed in an energy dispersive X-ray fluorescence spectrometer and the instrumental neutron activation analysis using the IEA-R1 nuclear reactor at IPEN. The results show significant differences for several elements (mainly, P, Cl, Ca, Ti, Mn, Fe, Ni, and Zn) in unhealthy caterpillars that can affect the development of this species as well as the quality and yield of the antivenom. Furthermore, its elemental characterization contributes for the understanding the potential pharmacological (procoagulant and antithrombotic) in the prevention of life-threatening blood clots

Multielemental analyses of Lonomia obliqua (Lepidoptera, Saturniidae) caterpillar using EDXRF and INAA techniques

In this study, multielemental analysis of Lonomia obliqua (Lepidoptera, Saturniidae) caterpillar was performed using energy dispersive X-ray fluorescence and instrumental neutron activation analysis techniques. This caterpillar is poisonous and has the ability to cause fatal hemorrhagic effects in humans after contact. The need of this study is related to morphological changes (mainly size and color) observed in some caterpillars used for preparation of antilonomic serum (antivenom). The samples were classified as healthy (caterpillars of control) and unhealthy (caterpillars visibly modified). The XRF measurements were performed in an energy dispersive X-ray fluorescence spectrometer and the instrumental neutron activation analysis using the IEA-R1 nuclear reactor at IPEN. The results show significant differences for several elements (mainly, P, Cl, Ca, Ti, Mn, Fe, Ni, and Zn) in unhealthy caterpillars that can affect the development of this species as well as the quality and yield of the antivenom. Furthermore, its elemental characterization contributes for the understanding the potential pharmacological (procoagulant and antithrombotic) in the prevention of life-threatening blood clots

Antiviral activity of hemolymph of Podalia against rubella virus.

Many active principles produced by animals, plants and microorganisms have been employed in the development of new drugs for the treatment of human diseases. Among animals known to produce pharmacologically active molecules that interfere in human cell physiology. Rubella virus (genus Rubivirus, family Togaviridae) is a single stranded RNA virus of positive genome polarity. Rubella virus infection of susceptible women during the first trimester of pregnancy often results in long-term virus persistence in the fetus causing multiple organ abnormalities. Potent antiviral activity against rubella virus (RV) has been observed in the hemolymph of Podalia sp. (Lepidoptera: Megalopygidae). This study evaluated the effect of hemolymph on RV infected Statens Serum Institute Rabbit Cornea (SIRC) cells. Results of cell viability and cell proliferation assays indicated that hemolymph was not toxic to cultured SIRC cells. Viral binding assay, antiviral assay, PCR, real-time PCR, and transmission electron microscopy were used to demonstrate that hemolymph in post-treatment could inhibit the production of infectious RV particles. Specifically, hemolymph was found to inhibit RV adsorption to the SIRC cells.

Bovine papillomavirus isolation by ultracentrifugation.

The bovine papillomavirus (BPV) is the etiological agent of bovine papillomatosis, which causes significant economic losses to livestock, characterized by the presence of papillomas that regress spontaneously or persist and progress to malignancy. Currently, there are 13 types of BPVs described in the literature as well as 32 putative new types. This study aimed to isolate viral particles of BPV from skin papillomas, using a novel viral isolation method. The virus types were previously identified with new primers designed. 77 cutaneous papilloma samples of 27 animals, Simmental breed, were surgically removed. The DNA was extracted and subjected to PCR using Delta-Epsilon and Xi primers. The bands were purified and sequenced. The sequences were analyzed using software and compared to the GenBank database, by BLAST tool. The viral typing showed a prevalence of BPV-2 in 81.81% of samples. It was also detected the presence of the putative new virus type BR/UEL2 in one sample. Virus isolation was performed by ultracentrifugation in a single density of cesium chloride. The method of virus isolation is less laborious than those previously described, allowing the isolation of complete virus particles of BPV-2.

Bovine papillomavirus in Brazil: detection of coinfection of unusual types by a PCR-RFLP method.

Bovine papillomavirus (BPV) is recognized as a causal agent of benign and malignant tumors in cattle. Thirteen types of BPV are currently characterized and classified into three distinct genera, associated with different pathological outcomes. The described BPV types as well as other putative ones have been demonstrated by molecular biology methods, mainly by the employment of degenerated PCR primers. Specifically, divergences in the nucleotide sequence of the L1 gene are useful for the identification and classification of new papillomavirus types. On the present work, a method based on the PCR-RFLP technique and DNA sequencing was evaluated as a screening tool, allowing for the detection of two relatively rare types of BPV in lesions samples from a six-year-old Holstein dairy cow, chronically affected with cutaneous papillomatosis. These findings point to the dissemination of BPVs with unclear pathogenic potential, since two relatively rare, new described BPV types, which were first characterized in Japan, were also detected in Brazil.

Expression of an antiviral protein from Lonomia obliqua hemolymph in baculovirus/insect cell system.

The control of viral infections, mainly those caused by influenza viruses, is of great interest in Public Health. Several studies have shown the presence of active properties in the hemolymph of arthropods, some of which are of interest for the development of new pharmacological drugs. Recently, we have demonstrated the existence of a potent antiviral property in the hemolymph of Lonomia obliqua caterpillars. The aim of this study was to produce an antiviral protein in a baculovirus/Sf9 cell system. The resulting bacmid contains the sequence coding for the antiviral protein previously described by our group. Total RNA from L. obliqua caterpillars was extracted with Trizol and used in the reverse transcription assay with oligo(d)T primer followed by polymerase chain reactions (RT-PCR) with specific primers for the cDNA coding for the antiviral protein, based on the sequence deposited in the GenBank database. Restriction sites were inserted in the cDNA for ligation in the donor plasmid pFastBac1™. The recombinant plasmid was selected in Escherichia coli DH5α and subsequently used in the transformation of E. coli DH10Bac for the construction of the recombinant bacmid. This bacmid was used for the expression of the antiviral protein in the baculovirus/Sf9 cell system. After identifying the protein by western blot, activity tests were performed, showing that the purified recombinant protein was able to significantly reduce viral replication (about 4 logs). Studies on the optimization of the expression system for the production of this antiviral protein in insect cells are in progress.

Can established cultured papilloma cells harbor bovine papillomavirus?

Papillomaviruses have been reported to be very difficult to grow in cell culture. Also, there are no descriptions of cell cultures from lesions of bovine cutaneous papillomatosis, with identification of different bovine papilloma virus (BPV) DNA sequences. In the present report, we describe primary cell cultures from samples of cutaneous lesions (warts). We investigated the simultaneous presence of different BPV DNA sequences, comparing the original lesion to different passages of the cell cultures and to peripheral blood. BPV 1, 2 and 4 DNA sequences were found in lesion samples, and respective cell cultures and peripheral blood, supporting our previous hypothesis of the possible activity of these sequences in different samples and now also showing how they can be maintained in different passages of cell cultures.

Can established cultured papilloma cells harbor bovine papillomavirus?

Papillomaviruses have been reported to be very difficult to grow in cell culture. Also, there are no descriptions of cell cultures from lesions of bovine cutaneous papillomatosis, with identification of different bovine papilloma virus (BPV) DNA sequences. In the present report, we describe primary cell cultures from samples of cutaneous lesions (warts). We investigated the simultaneous presence of different BPV DNA sequences, comparing the original lesion to different passages of the cell cultures and to peripheral blood. BPV 1, 2 and 4 DNA sequences were found in lesion samples, and respective cell cultures and peripheral blood, supporting our previous hypothesis of the possible activity of these sequences in different samples and now also showing how they can be maintained in different passages of cell cultures.