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The velvetbean caterpillar Anticarsia gemmatalis is one of the main soybean defoliators in Brazil. Currently, the main biopesticide used to control insect pests worldwide is the bacteria Bacillus thuringiensis (Bt), which produces entomopathogenic Crystal toxins (Cry) that act in the midgut of susceptible insects, leading them to death. The mode of action of Cry toxins in the midgut involves binding to specific receptors present on the brush border of epithelial cells such as aminopeptidase N (APN), alkaline phosphatase (ALP), cadherin, and others. Mutations in these receptors, among other factors, may be involved in the development of resistance; identification of functional Cry receptors in the midgut of A. gemmatalis is crucial to develop effective strategies to overcome this possible scenario. This study's goal is to characterize APNs of A. gemmatalis and identify a receptor for Cry1Ac in the midgut. The interaction of Bt spores with the midgut epithelium was observed in situ by immunohistochemistry and total aminopeptidase activity was estimated in brush border membrane vesicle (BBMV) samples, presenting higher activity in challenged individuals than in control ones. Ten APN sequences were found in a A. gemmatalis ' transcriptome and subjected to different in silico analysis, such as phylogenetic tree, multiple sequence alignment and identification of signal peptide, activity domains and GPI-anchor signal. BBMV proteins from 5th instar larvae were submitted to a ligand blotting using activated Cry1Ac toxin and a commercial anti-Cry polyclonal antibody; corresponding bands of proteins that showed binding to Cry toxin were excised from the SDS-PAGE gel and subjected to mass spectrometry analysis, which resulted in the identification of seven of those APNs. Quantitative PCR was realized to compare expression levels between individuals subjected to sublethal infection with Bt spores and control ones, presenting up- and downregulations upon Bt infection. From these results, we can infer that aminopeptidases N in A. gemmatalis could be involved in the mode of action of Cry toxins in its larval stage. [ABSTRACT FROM AUTHOR]

Aryel H Ferreira,1– 3,* Fábio LN Marques,4 Caroline C Real,5 Velaphi C Thipe,6,* Lucas F Freitas,1,2 Caroline SA Lima,1,2 Larissa Estessi de Souza,4 Mara S Junqueira,7 Daniele de Paula Faria,4 Gustavo HC Varca,1 Ademar B Lugão,1 Kattesh V Katti6,8,* 1Nuclear and Energy Research Institute, IPEN-CNEN/SP, Sao Paulo, 05508-000, Brazil; 2MackGraphe - Mackenzie Institute for Research in Graphene and Nanotechnologies, Mackenzie Presbyterian University, Sao Paulo, 01302-907, Brazil; 3Mackenzie Evangelical College of Paraná - Mackenzie Presbyterian University, Curitiba-PR, 80730-000, Brazil; 4Laboratory of Nuclear Medicine (LIM-43), Departamento de Radiologia e Oncologia, Faculdade de Medicina FMUSP, Universidade de Sao Paulo, Sao Paulo, 05403-911, Brazil; 5Aarhus University, Department of Nuclear Medicine & PET-Centre, Aarhus, Denmark; 6Department of Radiology, Institute of Green Nanotechnology, University of Missouri, Columbia, MO, 65212, USA; 7Center for Translational Research in Oncology, Cancer Institute of the State of Sao Paulo, Faculty of Medicine, University of Sao Paulo, Sao Paulo, Brazil; 8Department of Biotechnology & Food Technology, University of Johannesburg, Johannesburg, South Africa*These authors contributed equally to this workCorrespondence: Kattesh V Katti, Department of Radiology, Institute of Green Nanotechnology, University of Missouri, Columbia, MO, 65212, USA, Email kattik@health.missouri.edu Aryel H Ferreira, MackGraphe-Mackenzie Institute for Research in Graphene and Nanotechnologies, Mackenzie Presbyterian University, Consolação Street 930, São Paulo, 01302-907, Brazil, Email aryel.ferreira@mackenzie.brBackground: Recent advancements in nanomedicine and nanotechnology have expanded the scope of multifunctional nanostructures, offering innovative solutions for targeted drug delivery and diagnostic agents in oncology and nuclear medicine. Nanoparticles, particularly those derived from natural sources, hold immense potential in overcoming biological barriers to enhance therapeutic efficacy and diagnostic accuracy. Papain, a natural plant protease derived from Carica papaya, emerges as a promising candidate for green nanotechnology-based applications due to its diverse medicinal properties, including anticancer properties.Purpose: This study presents a novel approach in nanomedicine and oncology, exploring the potential of green nanotechnology by developing and evaluating technetium-99m radiolabeled papain nanoparticles (99mTc-P-NPs) for imaging breast tumors. The study aimed to investigate the efficacy and specificity of these nanoparticles in breast cancer models through preclinical in vitro and in vivo assessments.Methods: Papain nanoparticles (P-NPs) were synthesized using a radiation-driven method and underwent thorough characterization, including size, surface morphology, surface charge, and cytotoxicity assessment. Subsequently, P-NPs were radiolabeled with technetium-99m (99mTc), and in vitro and in vivo studies were conducted to evaluate cellular uptake at tumor sites, along with biodistribution, SPECT/CT imaging, autoradiography, and immunohistochemistry assays, using breast cancer models.Results: The synthesized P-NPs exhibited a size mean diameter of 9.3 ± 1.9 nm and a spherical shape. The in vitro cytotoxic activity of native papain and P-NPs showed low cytotoxicity in HUVEC, MDA-MB231, and 4T1 cells. The achieved radiochemical yield was 94.2 ± 3.1% that were sufficiently stable (≥ 90%) for 6 h. The tumor uptake achieved in the 4T1 model was 2.49 ± 0.32% IA/g at 2 h and 1.51 ± 0.20% IA/g at 6 h. In the spontaneous breast cancer model, 1.19 ± 0.20% IA/g at 2 h and 0.86 ± 0.31% IA/g at 6 h. SPECT/CT imaging has shown substantial tumor uptake of the new nanoradiopharmaceutical and clear tumor visualization. 99mTc-P-NPs exhibited a high affinity to tumoral cells confirmed by ex vivo autoradiography and immunohistochemistry assays.Conclusion: The findings underscore the potential of green nanotechnology-driven papain nanoparticles as promising agents for molecular imaging of breast and other tumors through SPECT/CT imaging. The results represent a substantial step forward in the application of papain nanoparticles as carriers of diagnostic and therapeutic radionuclides to deliver diagnostic/therapeutic payloads site-specifically to tumor sites for the development of a new generation of nanoradiopharmaceuticals. Keywords: papain nanoparticle, protein nanoparticle, enzymatic nanoparticle, imaging probe, nanoradiopharmaceuticals

O objetivo deste trabalho foi descrever a variação sazonal do fator de condição (K) correlacionado com os índices gonadossomático (IGS) e hepatossomático (IHS) de Leporinus obtusidens (Valenciennes, 1837), coletados a jusante da Usina Hidrelétrica do Funil, e avaliar se esses índices podem indicar o período reprodutivo dessa espécie. Os animais (n=110) foram capturados mensalmente entre setembro/2006 e agosto/2007 e agrupados por estação do ano. O comprimento e o peso dos animais foram obtidos para cálculo de K, o peso do fígado foi determinado para o cálculo do IHS e com o peso das gônadas foi obtido o IGS. O valor de K obtido para machos e fêmeas agrupados e somente para machos foi significativamente menor (p