Genetic basis of antibiotic resistance in bovine mastitis and its possible implications for human and ecological health.

Bovine mastitis is a mammary gland inflammation that can occur due to infectious pathogens, Staphylococcus aureus and Escherichia coli, which are, respectively, the most prevalent Gram-positive and Gram-negative bacteria associated with this disease. Currently, antibiotic treatment has become more complicated due to the presence of resistant pathogens. This review, therefore, aims to identify the most common resistance genes reported for these strains in the last four years. During the review, it was noted that blaZ, blaSHV, blaTEM, and blaampC are the most reported genes for S. aureus and E. coli, associated with drug inactivation, mainly ß-lactamases. They are characterized by generating bacterial resistance to ß-lactam antibiotics, the most common treatment in animal and human bacterial treatments (penicillins and cephalosporins, among others). Genes associated with efflux systems were also present in the two strains and included norA, tetA, tetC, and tetK, which generate resistance to macrolide and tetracycline antibiotics. Additionally, the effects of spreading resistance between animals and humans through direct contact (such as consumption of contaminated milk) or indirect contact (through environmental contamination) has been deeply discussed, emphasizing the importance of having adequate sanitation and antibiotic control and administration protocols.

Computer-made peptide RQ18 acts as a dual antifungal and antibiofilm peptide though membrane-associated mechanisms of action.

The spread of fungi resistant to conventional drugs has become a threatening problem. In this context, antimicrobial peptides (AMPs) have been considered as one of the main alternatives for controlling fungal infections. Here, we report the antifungal and antibiofilm activity and some clues about peptide RQ18's mechanism of action against Candida and Cryptococcus. This peptide inhibited yeast growth from 2.5 µM and killed all Candida tropicalis cells within 2 h incubation. Moreover, it showed a synergistic effect with antifungal agent the amphotericin b. RQ18 reduced biofilm formation and promoted C. tropicalis mature biofilms eradication. RQ18's mechanism of action involves fungal cell membrane damage, which was confirmed by the results of RQ18 in the presence of free ergosterol in the medium and fluorescence microscopy by Sytox green. No toxic effects were observed in murine macrophage cell lines and Galleria mellonella larvae, suggesting fungal target selectivity. Therefore, peptide RQ18 represents a promising strategy as a dual antifungal and antibiofilm agent that contributes to infection control without damaging mammalian cells.

Unwrapping the structural and functional features of antimicrobial peptides from wasp venoms.

The study of wasp venoms has captured attention due to the presence of a wide variety of active compounds, revealing a diverse array of biological effects. Among these compounds, certain antimicrobial peptides (AMPs) such as mastoparans and chemotactic peptides have emerged as significant players, characterized by their unique amphipathic short linear alpha-helical structure. These peptides exhibit not only antibiotic properties but also a range of other biological activities, which are related to their ability to interact with biological membranes to varying degrees. This review article aims to provide updated insights into the structure/function relationships of AMPs derived from wasp venoms, linking this knowledge to the potential development of innovative treatments against infections.

Evaluating virulence features of Acinetobacter baumannii resistant to polymyxin B.

The increasing resistance to polymyxins in Acinetobacter baumannii has made it even more urgent to develop new treatments. Anti-virulence compounds have been researched as a new solution. Here, we evaluated the modification of virulence features of A. baumannii after acquiring resistance to polymyxin B. The results showed lineages attaining unstable resistance to polymyxin B, except for Ab7 (A. baumannii polymyxin B resistant lineage), which showed stable resistance without an associated fitness cost. Analysis of virulence by a murine sepsis model indicated diminished virulence in Ab7 (A. baumannii polymyxin B resistant lineage) compared with Ab0 (A. baumannii polymyxin B susceptible lineage). Similarly, downregulation of virulence genes was observed by qPCR at 1 and 3 h of growth. However, an increase in bauE, abaI, and pgAB expression was observed after 6 h of growth. Comparison analysis of Ab0, Ab7, and Pseudomonas aeruginosa suggested no biofilm formation by Ab7. In general, although a decrease in virulence was observed in Ab7 when compared to Ab0, some virulence feature that enables infection could be maintained. In light of this, virulence genes bauE, abaI, and pgAB showed a potential relevance in the maintenance of virulence in polymyxin B-resistant strains, making them promising anti-virulence targets.

Repurposing streptomycin and chloramphenicol against bacterial pathogens by combination with diminazene aceturate.

Bacterial resistance is a threat to health worldwide, mainly due to reduced effective treatment. In this context, the search for strategies to control such infections and suppress antimicrobial resistance is necessary. One of the strategies that has been used is combination therapy. In the present work, we investigated the in vitro efficacy of the antimicrobials diminazene aceturate (DA), chloramphenicol (CHL), and streptomycin (STP) alone and in combination against Escherichia coli, Klebsiella pneumoniae, and Staphylococcus aureus clinical isolates. DA was capable of inhibiting all strains with MIC of 25-400 µg mL-1, while STP and CHL showed antibacterial activity with minimum inhibitory concentration (MICs) of ≤3.12-400 µg mL-1. The combination of aceturate with STP showed synergism toward almost all Gram-negative bacteria, with fractional inhibitory concentration index (FICIs) of 0.09-0.37. In addition, for CHL and aceturate, synergisms for Gram-negative and -positive strains were observed. A time-kill assay against E. coli revealed that the aceturate and STP combination can inhibit bacterial growth in a shorter time when compared with single antibiotics. In addition, antimicrobials did not show hemolytic activity even at the highest concentrations used. Therefore, the antimicrobial combinations presented in this work showed important results, demonstrating that combined therapy can be used as an alternative strategy for pathogen control.

Anti-Protozoan Activities of Polar Fish-Derived Polyalanine Synthetic Peptides.

Chagas disease, sleeping sickness and malaria are infectious diseases caused by protozoan parasites that kill millions of people worldwide. Here, we performed in vitro assays of Pa-MAP, Pa-MAP1.9, and Pa-MAP2 synthetic polyalanine peptides derived from the polar fish Pleuronectes americanus toward Trypanosoma cruzi, T. brucei gambiense and Plasmodium falciparum activities. We demonstrated that the peptides Pa-MAP1.9 and Pa-MAP2 were effective to inhibit T. brucei growth. In addition, structural analyses using molecular dynamics (MD) studies showed that Pa-MAP2 penetrates deeper into the membrane and interacts more with phospholipids than Pa-MAP1.9, corroborating the previous in vitro results showing that Pa-MAP1.9 acts within the cell, while Pa-MAP2 acts via membrane lysis. In conclusion, polyalanine Pa-MAP1.9 and Pa-MAP2 presented activity against bloodstream forms of T. b. gambiense, thus encouraging further studies on the application of these peptides as a treatment for sleeping sickness.

CRISPR-Cas engineering in food science and sustainable agriculture: recent advancements and applications.

The developments in the food supply chain to support the growing population of the world is one of today's most pressing issues, and to achieve this goal improvements should be performed in both crops and microbes. For this purpose, novel approaches such as genome editing (GE) methods have upgraded the biological sciences for genome manipulation and, among such methods, clustered regularly interspaced short palindromic repeats (CRISPR) and CRISPR-associated proteins (Cas) are the main exciting innovations since the Green Revolution. CRISPR/Cas systems can be a potent tool for the food industry, improvement of agricultural crops and even for protecting food-grade bacteria from foreign genetic invasive elements. This review introduces the history and mechanism of the CRISPR-Cas system as a genome editing tool and its applications in the vaccination of starter cultures, production of antimicrobials and bioactive compounds, and genome editing of microorganisms.

Purification of PaTx-II from the Venom of the Australian King Brown Snake and Characterization of Its Antimicrobial and Wound Healing Activities.

Infections caused by multi-drug-resistant (MDR) bacteria are a global threat to human health. As venoms are the source of biochemically diverse bioactive proteins and peptides, we investigated the antimicrobial activity and murine skin infection model-based wound healing efficacy of a 13 kDa protein. The active component PaTx-II was isolated from the venom of Pseudechis australis (Australian King Brown or Mulga Snake). PaTx-II inhibited the growth of Gram-positive bacteria in vitro, with moderate potency (MICs of 25 µM) observed against S. aureus, E. aerogenes, and P. vulgaris. The antibiotic activity of PaTx-II was associated with the disruption of membrane integrity, pore formation, and lysis of bacterial cells, as evidenced by scanning and transmission microscopy. However, these effects were not observed with mammalian cells, and PaTx-II exhibited minimal cytotoxicity (CC50 > 1000 µM) toward skin/lung cells. Antimicrobial efficacy was then determined using a murine model of S. aureus skin infection. Topical application of PaTx-II (0.5 mg/kg) cleared S. aureus with concomitant increased vascularization and re-epithelialization, promoting wound healing. As small proteins and peptides can possess immunomodulatory effects to enhance microbial clearance, cytokines and collagen from the wound tissue samples were analyzed by immunoblots and immunoassays. The amounts of type I collagen in PaTx-II-treated sites were elevated compared to the vehicle controls, suggesting a potential role for collagen in facilitating the maturation of the dermal matrix during wound healing. Levels of the proinflammatory cytokines interleukin-1ß (IL-1ß), interleukin-6 (IL-6) and tumor necrosis factor-α (TNF-α), cyclooxygenase-2 (COX-2) and interleukin-10 (IL-10), factors known to promote neovascularization, were substantially reduced by PaTx-II treatment. Further studies that characterize the contributions towards efficacy imparted by in vitro antimicrobial and immunomodulatory activity with PaTx-II are warranted.

Relationship between intestinal microbiota, diet and biological systems: an integrated view.

The health-disease process can be influenced by the intestinal microbiota. As this plays a fundamental role in protecting the organism, the importance of studying the composition and diversity of this community becomes increasingly evident. Changes in the composition of the intestinal bacterial community may result in dysbiosis, and this process may contribute to triggering various diseases in all biological systems. This imbalance of intestinal microbiota homeostasis may alter commensal bacteria and the host metabolism, as well as immune function. Dysbiosis also causes an increase in intestinal permeability due to exposure to molecular patterns associated with the pathogen and lipopolysaccharides, leading to a chronic inflammatory process that can result in diseases for all biological systems. In this context, dietary intervention through the use of probiotics, prebiotics and antioxidant foods can be considered a contribution to the modulation of intestinal microbiota. Probiotics have been used to provide up to 10 billion colony forming units, and probiotic foods, Kefir and fermented natural yogurt are also used. Prebiotics, in turn, are found in supplemental formulations of processed foods and in functional foods that are also sources of phenolic compounds, such as flavonoids, antioxidant and anti-inflammatory substances, polyunsaturated fatty acids, vitamins, and minerals. In this review, we will discuss the relationship between an imbalance in the intestinal microbiota with the development of diseases, besides indicating the need for future studies that can establish bacterial parameters for the gastrointestinal tract by modulating the intestinal microbiota, associated with the adoption of healthy habits during all life cycles.

Antimicrobial peptides used as growth promoters in livestock production.

Antibiotic growth promoters (AGPs) have been administered in livestock for decades to improve food digestion in growing animals, while also contributing to the control of microbial pathogens. The long-term and indiscrimate use of AGPs has generated genetic modifications in bacteria, leading to antimicrobial resistance (AMR), which can be disseminated to commensal and pathogenic bacteria. Thus, antimicrobial peptides (AMPs) are used to replaced AGPs. AMPs are found in all domains of life, and their cationic characteristics can establish electrostatic interactions with the bacterial membrane. These molecules used as growth promoters can present benefits for nutrient digestibility, intestinal microbiota, intestinal morphology, and immune function activities. Therefore, this review focuses on the application of AMPs with growth promoting potential in livestock, as an alternative to conventional antibiotic growth promoters, in an attempt to control AMR. KEY POINTS: • The long-term and indiscriminate use of AGPs in animal food can cause AMR. • AMPs can be used as substitute of antibiotics in animal food suplementation. • Animal food suplementated with AMPs can provied economic efficiency and sustainable livestock production.

Concentrated MTA Repair HP reduced biofilm and can cause reparative action at a distance.

AIM: To evaluate in vitro whether MTA Repair HP can induce repair processes at a distance, including its effects on biofilm, cell viability, migration, production of TGF-ß, phosphate and ALP, evaluated through MTA diluted extracts. METHODOLOGY: Initially, antibacterial tests were performed with the bacterium Streptococcus mutans (ATCC 25175) in the presence of MTA extracts (dilutions of 1:1, 1:2 and 1:4). Growth inhibition assay by microdilution in broth, antibiofilm plate assay of young biofilm and antibiofilm assay in confocal microscopy of mature biofilm were carried out. Then, pulp cells were stimulated in the presence of several MTA dilutions, and cell viability (MTT assay), proliferation and migration capacity (scratch assay) were evaluated. To evaluate the capacity of 1:1, 1:2 and 1:4 dilutions of MTA Repair HP to promote the production of important agents of odontogenic differentiation and mineralization, ALP activity, TGF-ß secretion and phosphate quantification were measured. Statistical differences were verified using one-way and two-way anova and Tukey's post-tests. RESULTS: The test dilutions of MTA Repair HP did not inhibit planktonic S. mutans growth but were able to reduce young and mature S. mutans biofilm (p < 0.001). In addition, none of the MTA Repair HP dilutions was cytotoxic for pulp cells. The 1:2 and 1:4 dilutions of MTA Repair HP induced migration and proliferation of pulp cells (p < 0.05). ALP activity and TGF-ß secretion were independent of the tested dilution (p < 0.001). Diluted 1:4 MTA Repair HP produced less phosphate than the more concentrated 1:1 and 1:2 MTA dilutions (p < 0.001). CONCLUSIONS: Undiluted MTA Repair HP reduced S. mutans biofilm, when compared to 1:2 and 1:4 MTA dilutions. Furthermore, none of the tested dilutions was cytotoxic to pulp cells. MTA Repair HP promoted cell migration and proliferation at a distance, assessed through the dilution of the MTA. Even from a distance, MTA Repair HP has the ability to participate in some events related to repair, such as migration, proliferation and TGF production.

Antimicrobial and immunomodulatory in vitro profile of double antibiotic paste.

AIM: To evaluate the antimicrobial and immunomodulatory activity of double antibiotic paste (DAP) in an in vitro infection model. METHODOLOGY: The minimum inhibitory and bactericidal concentrations (MIC and MBC) and the antibiofilm activities (TTC assay) of DAP and its components (ciprofloxacin and metronidazole) were evaluated against Staphylococcus aureus and Enterococcus faecalis compared with triple antibiotic paste (TAP). The cellular viability of RAW 264.7 macrophages (24 and 72 h) and L929 fibroblasts (48 and 72 h) was evaluated by MTT. Furthermore, the production of TNF-α, IL-12, IL-6, IL-1α, IL-10 and NO (on RAW 264.7), besides IL-6, TGF-ß and NO (on L929), stimulated with DAP in baseline and associated with heat-killed microbial-antigen conditions was measured by ELISA and Griess reaction. Data were analysed using the one-way ANOVA test with Bonferroni's corrections. RESULTS: The MBC of pharmacopoeia DAP was similar to TAP for E. faecalis (0.25 µg.  mL-1 ) and lower for S. aureus (DAP 1 µg. mL-1 and TAP 2 µg. mL-1 ; p < .001). Ciprofloxacin was the most effective antibiofilm drug from the pastes (35% of reduction for E. faecalis and S. aureus; p < .0001), and both pastes had a similar antibiofilm eradication against both biofilm species (29% and 35% for S. aureus and 76% and 85% for E. faecalis; p < .0001). DAP was cytotoxic against the tested cells. DAP significantly upregulated IL-1α (p < .001), IL-6 (p < .0001), TNF-α (p < .01) and IL-12 (p < .05; in the absence of antigens) and significantly reduced IL-6 (p < .0001; in the presence of HK-S. aureus) and IL-10 (p < .05; in the presence of both antigens) on macrophages. Furthermore, DAP upregulated IL-6 (p < .001) and NO (p < .05; in the absence of antigens), IL-6 (p < .001; in the presence of HK-S. aureus) and reduced NO (p < .001; in the presence of HK-S. aureus). CONCLUSIONS: Double antibiotic paste and TAP had similar antimicrobial activity against S. aureus and E. faecalis. DAP upregulated pro-inflammatory cytokines mainly in the absence of antigens and had pro- and anti-inflammatory activity in RAW 264.7 macrophages and L929 fibroblasts in the presence of antigens involved in pulp infections.

Proteomic analysis of human dental pulp in different clinical diagnosis.

OBJECTIVES: The present study aimed to identify proteins obtained from pulp tissue and correlate with each clinical diagnosis (healthy pulp, inflamed pulp, and necrotic pulp). MATERIALS AND METHODS: A total of forty-five molars were used. Three biological replicas were evaluated. Lysis and sonication were used for protein extraction. Protein quantification was assessed by using the Bradford technique, and shotgun proteome analysis was performed by nanoUPLC-MSE using a Synapt G2 mass spectrometer. Mass spectra data were processed using the Waters PLGS software, and protein identification was done using the human Uniprot database appended to the PLGS search engine. RESULTS: A total of 123 different proteins were identified in all evaluated pulp conditions. Among these, 66 proteins were observed for healthy pulp, 66 for inflamed pulp, and 91 for necrotic pulp. Most protein identification was related to immune response, multi-organism process, platelet activation, and stress in inflamed pulp samples compared to healthy pulp. Proteins related to cellular component organization or biogenesis, developmental process, growth, immune response, multi-organism process, response to stimulus, signaling, stress, and transport were identified in cases of apical periodontitis compared to inflamed pulp. CONCLUSIONS: The progression of the disease to inflamed pulp promoted a high abundance of proteins related to the immune system and stress. Comparing the necrotic pulp with inflamed pulp conditions, a high abundance of proteins was noticed related to metabolism, transport, and response between organisms. CLINICAL RELEVANCE: This finding may assist in future studies of new markers, understanding of tissue engineering, and development of future products.

Identification of the Active Principle Conferring Anti-Inflammatory and Antinociceptive Properties in Bamboo Plant.

Early plants began colonizing earth about 450 million years ago. During the process of coevolution, their metabolic cellular pathways produced a myriad of natural chemicals, many of which remain uncharacterized biologically. Popular preparations containing some of these molecules have been used medicinally for thousands of years. In Brazilian folk medicine, plant extracts from the bamboo plant Guadua paniculata Munro have been used for the treatment of infections and pain. However, the chemical basis of these therapeutic effects has not yet been identified. Here, we performed protein biochemistry and downstream pharmacological assays to determine the mechanisms underlying the anti-inflammatory and antinociceptive effects of an aqueous extract of the G. paniculata rhizome, which we termed AqGP. The anti-inflammatory and antinociceptive effects of AqGP were assessed in mice. We identified and purified a protein (AgGP), with an amino acid sequence similar to that of thaumatins (~20 kDa), capable of repressing inflammation through downregulation of neutrophil recruitment and of decreasing hyperalgesia in mice. In conclusion, we have identified the molecule and the molecular mechanism responsible for the anti-inflammatory and antinociceptive properties of a plant commonly used in Brazilian folk medicine.

Adepamycin: design, synthesis and biological properties of a new peptide with antimicrobial properties.

Antimicrobial peptides (AMP) are molecules with a broad spectrum of activities that have been identified in most living organisms. In addition, synthetic AMPs designed from natural polypeptides have been largely investigated. Here, we designed a novel AMP using the amino acid sequence of a plant trypsin inhibitor from Adenanthera pavonina seeds (ApTI) as a template. The 176 amino acid residues ApTI sequence was cleaved in silico using the Collection of Antimicrobial Peptides (CAMPR3), through the sliding-window method. Further improvements in AMP structure were carried out, resulting in adepamycin, an AMP designed from ApTI. Adepamycin showed antimicrobial activity from 0.9 to 3.6 µM against Escherichia coli, Klebsiella oxytoca, Klebsiella pneumoniae, Pseudomonas aeruginosa, and Staphylococcus aureus strains. Moreover, this peptide also displayed activity against Candida albicans and Candida tropicalis. No toxic effects were observed on healthy human cells. Studies on the mechanism of action of adepamycin were carried out using an E. coli and C. tropicalis. Adepamycin triggers membrane disturbances, leading to intracellular nucleic acids release in E. coli. For C. tropicalis, an initial interference with the plasma membrane integrity is followed by the formation of intracellular reactive oxygen species (ROS), leading to apoptosis. Structurally, adepamycin was submitted to circular dichroism spectroscopy, molecular modeling and molecular dynamics simulations, revealing an environment-dependent α-helical structure in the presence of 2,2,2- trifluoroethanol (TFE) and in contact with mimetic vesicles/membranes. Therefore, adepamycin represents a novel lytic AMP with dual antibacterial and antifungal properties.

Proteomic changes in skeletal muscle of aged rats in response to resistance training.

Sarcopenia is a multifactorial process defined by loss of strength and skeletal muscle mass, which leads to a reduction in muscle cross-sectional area (CSA). Although resistance training (RT) has been indicated as a tool to counteract sarcopenia, the protein profile associated with skeletal muscle adaptations remains to be determined. We investigated the effects of 12 weeks of RT on the skeletal muscle proteome profile and CSA of young and older rats. Twenty-four animals were divided into four groups: young sedentary or trained and older sedentary or trained (six animals per group). A 12-week RT protocol was performed, which consisted of climbing a vertical ladder. The proteins from the gastrocnemius were analysed by LC-ESI-MS/MS. One-hundred and thirty-one proteins were identified, of which 28 were assessed between the groups. Ageing induced an increase in proteins associated with the glycolytic pathway, transport and stress response, which represent crucial mechanisms for muscle adaptation. RT upregulated metabolic enzymes, anti-oxidant activity and transport proteins, besides increasing hypertrophy, regardless of age, suggesting a beneficial adaptation to mitigate age-related sarcopenia. RT reduced muscle atrophy through the regulation of stress response and by increasing proteins related to energy production and transport, which in turn might protect tissue damage arising from exercise and ageing. SIGNIFICANCE OF THE STUDY: Protein abundance levels related to the metabolic process and stress response were increased in the aged muscle. RT proved to be an important intervention capable of inducing significant effects on muscle proteome regardless of ageing, due to upregulation of glycolytic enzymes, and anti-oxidant and transport proteins. This effect could lead to a beneficial adaptation in muscle structure, cellular function and overall homeostasis maintenance. This study contributes to better understanding of the basic biology of ageing and clarifies more profoundly the molecular networks behind physiological adaptations promoted by exercise training. Therefore, the results open new perspectives and insights for studies based on transcriptomics, metabolomics and functional assays.

Antiviral peptides as promising therapeutic drugs.

While scientific advances have led to large-scale production and widespread distribution of vaccines and antiviral drugs, viruses still remain a major cause of human diseases today. The ever-increasing reports of viral resistance and the emergence and re-emergence of viral epidemics pressure the health and scientific community to constantly find novel molecules with antiviral potential. This search involves numerous different approaches, and the use of antimicrobial peptides has presented itself as an interesting alternative. Even though the number of antimicrobial peptides with antiviral activity is still low, they already show immense potential to become pharmaceutically available antiviral drugs. Such peptides can originate from natural sources, such as those isolated from mammals and from animal venoms, or from artificial sources, when bioinformatics tools are used. This review aims to shed some light on antimicrobial peptides with antiviral activities against human viruses and update the data about the already well-known peptides that are still undergoing studies, emphasizing the most promising ones that may become medicines for clinical use.

Development of a novel anti-biofilm peptide derived from profilin of Spodoptera frugiperda.

Candida yeast infections are the fourth leading cause of death worldwide. Peptides with antimicrobial activity are a promising alternative treatment for such infections. Here, the antifungal activity of a new antimicrobial peptide-PEP-IA18-was evaluated against Candida species. PEP-IA18 was designed from the primary sequence of profilin, a protein from Spodoptera frugiperda, and displayed potent activity against Candida albicans and Candida tropicalis, showing a minimum inhibitory concentration (MIC) of 2.5 µM. Furthermore, the mechanism of action of PEP-IA18 involved interaction with the cell membrane (ergosterol complexation). Treatment at MIC and/or 10 × MIC significantly reduced biofilm formation and viability. PEP-IA18 showed low toxicity toward human fibroblasts and only revealed hemolytic activity at high concentrations. Thus, PEP-IA18 exhibited antifungal and anti-biofilm properties with potential applicability in the treatment of infections caused by Candida species.

High-intensity aerobic training lowers blood pressure and modulates the renal renin-angiotensin system in spontaneously hypertensive rats.

Background: This study aimed to verify the effects of high-intensity aerobic training (HIAT) on BP control and renin-angiotensin system (RAS) components in renal tissue of SHR. Ten SHRs received HIAT or control for 8-weeks. At the end of the training, the SBP showed a reduction of ~ 30mmHg (p < .01) in HIAT and increased by ~ 15 mmHg in the  control group. HIAT resulted in a higher release of nitrite, IL-6, ACE2 and ATR2. These results indicated an association between BP, NO and renal RAS.Abbreviations: JAA: writing, carried out all experimental procedures, performed statistical analysis, original draft and revised manuscript DMS: data interpretation, formal analysis, writing, editing and revised manuscript BAP: carried all experimental procedures, revised manuscritpt CPCG: carried all experimental procedures, revised manuscritpt MEN: experimental procedures, revised manuscript and data interpretation RWP: drafted and revised manuscript RCA: writing, experimental procedures, revised manuscript JP: writing, data interpretation and revised manuscript OLF: writing, original draft and revised manuscript.

Effects of a Reserve Protein on Spodoptera frugiperda Development: A Biochemical and Molecular Approach to the Entomotoxic Mechanism.

Talisin is a storage protein from Talisia esculenta seeds that presents lectin-like and peptidase inhibitor properties. These characteristics suggest that talisin plays a role in the plant defense process, making it a multifunctional protein. This work aimed to investigate the effects of chronic intake of talisin on fifth instar larvae of Spodoptera frugiperda, considered the main insect pest of maize and the cause of substantial economic losses in several other crops. The chronic intake of talisin presented antinutritional effects on the larvae, reducing their weight and prolonging the total development time of the insects. In addition, talisin-fed larvae also showed a significant reduction in the activity of trypsin-like enzymes. Midgut histology analysis of talisin-fed larvae showed alterations in the intestinal epithelium and rupture of the peritrophic membrane, possibly causing an increase of aminopeptidase activity in the midgut lumen. Talisin also proved to be resistant to degradation by the digestive enzymes of S. frugiperda. The transcription profile of trypsin, chymotrypsin and aminopeptidase genes was also analyzed through qPCR technique. Talisin intake resulted in differential expression of at least two genes from each of these classes of enzymes. Molecular docking studies indicated a higher affinity of talisin for the less expressed enzymes.