Formulative Study and Characterization of Novel Biomaterials Based on Chitosan/Hydrolyzed Collagen Films.

To date, the need for biomaterials capable of improving the treatment of chronic skin wounds remains a clinical challenge. The aim of the present work is to formulate and characterize chitosan (Cs)/hydrolyzed collagen (HC) films as potential biomaterials with improved mechanical and hydration performances compared to single component formulations. Films were made by the solvent casting method, with or without glycerin and/or PEG1500 as plasticizers, resulting in a total of eight formulations. All films were characterized by their physico-chemical characteristics and their mechanical and hydration features. A full factorial design was also used to statistically assess the effect of HC concentration, type and concentration of plasticizers and their possible interactions on mechanical and swelling behaviors. Solid state characterization confirmed the hybrid nature of the films, with suggested electrostatic interactions between Cs and HC. Mechanical and swelling properties, along with the analysis of the experimental design, allowed the identification of formulations containing high HC concentration (2% w/v) and glycerin or glycerin/PEG1500 as more suitable candidates for skin wound treatment. Finally, viability assay of immortalized human keratinocytes (HaCaT) showed no statistical differences in cell survival compared to the complete culture medium, suggesting their potential as a promising tool for biomedical applications.

Taxonomic and Functional Diversity of Flower-Visiting Insects in Coffee Crops.

Despite the important role that flower-visiting insects play in agricultural production, none of the previous studies of coffee pollinators in Colombia have incorporated functional diversity into their analysis. Therefore, this study aimed to quantify the abundance, richness, and functional diversity of insects that visit flowers in coffee crops. Twenty-eight plots were selected among five sites in the north, center, and south of Colombia. In each plot, coffee flower insect visitors were collected and recorded on 90 trees at eight-minute intervals per tree, at three different times over three days. All sampling was carried out during two flowering events per year, over three years, resulting in a total of 1240 h of observations. Subsequently, the insects were taxonomically identified, and the number of individuals and species, as well as the diversity of the order q, were estimated. Functional diversity was also characterized in the bee community. The results: (a) 23,735 individuals belonging to 566 species were recorded; of them, 90 were bees, with the native species being the most abundant during 10:30 and 13:00 h; (b) bees formed five functional groups, with corbiculate and long-tongued non-corbiculate bees being the most abundant and occupying the largest regions of functional space; (c) potential pollinators in coffee crops are Apis mellifera, Nannotrigona gaboi, Tetragonisca angustula, Geotrigona cf. tellurica, and Partamona cf. peckolti. Coffee crops host a wide diversity of flower visitors, especially bees, which could be beneficial for productivity and contribute to the maintenance of plant species that accompany coffee cultivation.

Pilot Testing of an Area-Wide Biological Control Strategy against the Coffee Berry Borer in Colombia Using African Parasitoids.

The coffee berry borer (CBB), Hypothenemus hampei (Ferrari, 1867) (Coleoptera: Curculionidae: Scolytinae), native to Africa, is a major global insect pest of coffee. It has invaded many coffee production areas around the world that do not have natural enemies. In this study, two African parasitoids, Prorops nasuta Waterston (Hymenoptera: Bethylidae) and Phymastichus coffea Waterston (Hymenoptera: Eulophidae), were mass-reared for field release against H. hampei in Chinchiná, Colombia. More than 1.5 million wasps of each species were released on a 61-hectare coffee farm in replicated plots, resulting in parasitism rates of up to 7.7% for P. nasuta and 56.3% for P. coffea. This led to a maximum reduction in H. hampei field populations of 81% from dispersal coffee plots (old coffee crops before plant stumping) and 64.3% in colonization coffee plots (new coffee crops with active growing and fruiting plants) within the farm. As a result of this area-wide strategy, the percentage of CBB-infested coffee berries in colonization coffee plots decreased from 51.1 to 77.5% compared to coffee plots without parasitoid releases. This approach offers a promising alternative to the use of chemical insecticides and could be integrated into current pest management programs to control H. hampei.

Antitumoral and Immunogenic Capacity of ß-D-Glucose-Reduced Silver Nanoparticles in Breast Cancer.

Immunogenic cell death (ICD) is a type of cell death capable of stimulating immunity against cancer through danger signals that lead to an adaptive immune response. Silver nanoparticles (AgNPs) have been shown to have a cytotoxic effect on cancer cells; however, their mechanism of action is not fully understood. The present study synthesized, characterized, and evaluated the cytotoxic effect of beta-D-glucose-reduced AgNPs (AgNPs-G) against breast cancer (BC) cells in vitro; and assess the immunogenicity of cell death in vitro and in vivo. The results showed that AgNPs-G induce cell death in a dose-dependent manner on BC cell lines. In addition, AgNPs show antiproliferative effects by interfering with the cell cycle. Regarding the detection of damage-associated molecular patterns (DAMPs), it was found that treatment with AgNPs-G induces calreticulin exposure and the release of HSP70, HSP90, HMGB1, and ATP. In vivo, prophylactic vaccination did not prevent tumor establishment; however, tumor weight was significantly lower in AgNPs-G vaccinated mice, while the survival rate increased. In conclusion, we have developed a new method for the synthesis of AgNPs-G, with in vitro antitumor cytotoxic activity on BC cells, accompanied by the release of DAMPs. In vivo, immunization with AgNPs-G failed to induce a complete immune response in mice. Consequently, additional studies are needed to elucidate the mechanism of cell death that leads to the design of strategies and combinations with clinical efficacy.

Putative Wound Healing Induction Functions of Exosomes Isolated from IMMUNEPOTENT CRP.

Chronic wounds in diabetic patients can take months or years to heal, representing a great cost for the healthcare sector and impacts on patients' lifestyles. Therefore, new effective treatment alternatives are needed to accelerate the healing process. Exosomes are nanovesicles involved in the modulation of signaling pathways that can be produced by any cell and can exert functions similar to the cell of origin. For this reason, IMMUNEPOTENT CRP, which is a bovine spleen leukocyte extract, was analyzed to identify the proteins present and is proposed as a source of exosomes. The exosomes were isolated through ultracentrifugation and shape-size, characterized by atomic force microscopy. The protein content in IMMUNEPOTENT CRP was characterized by EV-trap coupled to liquid chromatography. The in silico analyses for biological pathways, tissue specificity, and transcription factor inducement were performed in GOrilla ontology, Panther ontology, Metascape, and Reactome. It was observed that IMMUNEPOTENT CRP contains diverse peptides. The peptide-containing exosomes had an average size of 60 nm, and exomeres of 30 nm. They had biological activity capable of modulating the wound healing process, through inflammation modulation and the activation of signaling pathways such as PIP3-AKT, as well as other pathways activated by FOXE genes related to specificity in the skin tissue.

Characterization and Molecular Modelling of Non-Antibiotic Nanohybrids for Wound Healing Purposes.

The healing process of chronic wounds continues to be a current clinical challenge, worsened by the risk of microbial infections and bacterial resistance to the most frequent antibiotics. In this work, non-antibiotic nanohybrids based on chlorhexidine dihydrochloride and clay minerals have been developed in order to design advanced therapeutic systems aimed to enhance wound healing in chronic lesions. To prepare the nanohybrids, two methodologies have been compared: the intercalation solution procedure and the spray-drying technique, the latter as a one-step process able to reduce preparation times. Nanohybrids were then fully studied by solid state characterization techniques. Computational calculations were also performed to assess the interactions between the drug and the clays at the molecular level. In vitro human fibroblast biocompatibility and antimicrobial activity against Staphylococcus aureus and Pseudomonas aeruginosa were assessed to check biocompatibility and potential microbicidal effects of the obtained nanomaterials. The results demonstrated the effective organic/inorganic character of the nanohybrids with homogeneous drug distribution into the clayey structures, which had been confirmed by classical mechanics calculations. Good biocompatibility and microbicidal effects were also observed, especially for the spray-dried nanohybrids. It was suggested that it could be due to a greater contact area with target cells and bacterial suspensions.

Electromyographic and kinematic parameters of the shoulder in wheelchair rugby players: case reports.

Wheelchair rugby was created as part of the rehabilitation for patients with spinal cord injury. The biomechanical analysis of wheelchair propulsion (WP) in these athletes seems to be a key element to understand the reasons behind musculoskeletal injuries. This case reports study aimed to describe the electromyographic activity and kinematic parameters of the shoulder during the propulsion phases on the wheelchair in two Paralympic rugby players (A1 and A2) with spinal cord injury. Myoelectric activity (three portions of the deltoid, biceps and triceps brachii) and kinematics of the shoulder were assessed during the push (PP) and recovery (RP) phases. These variables were calculated considering ten propulsion cycles by each athlete. The results showed a different muscle activation between players, A1 described a high average amplitude of the anterior deltoid (PP = 58.44 ± 16.35%MVC; RP = 43.16 ± 13.48%MVC) in both propulsion phases, while A2 generated high average activity of triceps brachii (29.28 ± 10.63%MVC) and middle deltoid (46.53 ± 14.48%MVC), during PP and RP, respectively. At the same time, the player with a C7-T1 spinal cord injury (A2) showed a higher range of motion in the three plans, considering both propulsion phases.

Curcumin Sensitizes 4T1 Murine Breast Cancer Cells to Cisplatin Through PAR4 Secretion.

BACKGROUND/AIM: Prostate apoptosis response 4 (PAR4), a tumour-suppressor protein, selectively induces apoptosis of cancer cells without affecting normal cells. Its soluble form is induced by secretagogues (e.g., chloroquine), and it induces apoptosis by interacting with the receptor of glucose-regulated protein 78, which is overexpressed in cancer cells. In this study, curcumin was analyzed as an inducer of PAR4 expression in 4T1 murine breast cancer cell. and its ability to induce PAR4 secretion in Balb/c mice. In addition, the cisplatin sensitizing effect of soluble PAR4 was analyzed. MATERIAL AND METHODS: The 4T1 cell line was treated in vitro using different concentrations of curcumin; cell viability was analyzed using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay and PAR4 expression by western blotting. The expression of soluble PAR4 in the serum of mice treated with intraperitoneal curcumin was analyzed using the dot-blot method. Moreover, MTT assay was used to analyze the effects of serum from curcumin-treated mice on cell viability. Tumor size was analyzed in mice treated with curcumin alone and in combination with cisplatin. RESULTS: Curcumin showed a dose- and time-dependent effects on cell viability on 4T1 cells, as well as increasing PAR4 expression. Compared with the control group (phosphate-buffered saline), mice treated with curcumin showed an increase in plasma PAR4. In the Balb/C tumor model, mice treated with curcumin and cisplatin showed greater tumor shrinkage than the control group. CONCLUSION: These results indicate that curcumin induces expression of soluble PAR4 and sensitizes tumor cells to cisplatin.

Methodology to Test Control Agents and Insecticides Against the Coffee Berry Borer Hypothenemus hampei.

Prior to recommending insecticides to treat the coffee berry borer (CBB) Hypothenemus hampei, it is valuable to know the mortality and repellency of these insecticides against adult insects or their impact on reproductive output. However, currently available methods assess adult mortality only, limiting the selection of novel insecticides with a different mode of action. In this work, different experimental methods were examined to identify the diverse effects on the CBB under laboratory conditions. For this, green coffee fruits (GFs) were collected and disinfected by immersion in sodium hypochlorite solution followed by UV light irradiation. In parallel, CBB adults from a colony were disinfected by immersion in sodium hypochlorite solution. To assess fruit protection (preinfestation), the fruits were placed in plastic boxes, and the insecticides were applied. Then, the CBB adults were released at a rate of two CBBs per GF. The GFs were left under controlled conditions to evaluate CBB infestation and survival after 1, 7, 15, and 21 days. To evaluate insecticide efficacy after CBB infestation (postinfestation), CBB adults were released to the GFs in a 2:1 ratio for 3 h at 21 °C. Infested fruits showing CBB adults with their abdomens partially exposed were selected and placed in 96-well racks, and the CBBs boring into the fruits were treated directly. After 20 days, the fruits were dissected, and the CBB biological stages inside each fruit were recorded. The GFs served as substrates that mimic natural conditions to evaluate toxic, chemical, and biological insecticides against the CBB.

The Inflammatory Process Modulates the Expression and Localization of WT1 in Podocytes Leading to Kidney Damage.

BACKGROUND/AIM: Wilms' tumor 1 (WT1) is involved in the development of the urogenital system and is expressed in podocytes throughout life. Inflammation of renal glomeruli causes renal damage-induced nephrotic syndrome and steroid-resistant nephrotic syndrome have mutations in the WT1 gene. The aim of this work was to determine if the inflammatory process modulates the expression and localization of WT1 in podocytes that cause kidney damage using lipopolysaccharide (LPS)-treated mice as a sepsis model. MATERIALS AND METHODS: In investigation of renal damage, proteinuria and histology were analyzed. WT1 modulation was analyzed by indirect immunofluorescence, immunohistochemistry and western blot assays, and proinflammatory cytokines were analyzed by quantitative polymerase chain reaction assay. RESULTS: WT1 expression decreased most at 24 and 36 h after the induction of inflammation and phosphorylated WT1 was mainly localized in the cytoplasm, reduced nephrin mRNA expression and increased mRNA expression of tumor necrosis factor α and interleukin 1ß. CONCLUSION: These results indicate that the immune system plays an important role in the modulation of WT1, leading to kidney damage.

Structure and Dynamics of the Gut Bacterial Community Across the Developmental Stages of the Coffee Berry Borer, Hypothenemus hampei.

The coffee berry borer (CBB); Hypothenemus hampei (Coleoptera: Curculionidae), is widely recognized as the major insect pest of coffee crops. Like many other arthropods, CBB harbors numerous bacteria species that may have important physiological roles in host nutrition, detoxification, immunity and protection. To date, the structure and dynamics of the gut-associated bacterial community across the CBB life cycle is not yet well understood. A better understanding of the complex relationship between CBB and its bacterial companions may provide new opportunities for insect control. In the current investigation, we analyzed the diversity and abundance of gut microbiota across the CBB developmental stages under field conditions by using high-throughput Illumina sequencing of the 16S ribosomal RNA gene. Overall, 15 bacterial phyla, 38 classes, 61 orders, 101 families and 177 genera were identified across all life stages, including egg, larva 1, larva 2, pupa, and adults (female and male). Proteobacteria and Firmicutes phyla dominated the microbiota along the entire insect life cycle. Among the 177 genera, the 10 most abundant were members of Ochrobactrum (15.1%), Pantoea (6.6%), Erwinia (5.7%), Lactobacillus (4.3%), Acinetobacter (3.4%), Stenotrophomonas (3.1%), Akkermansia (3.0%), Agrobacterium (2.9%), Curtobacterium (2.7%), and Clostridium (2.7%). We found that the overall bacterial composition is diverse, variable within each life stage and appears to vary across development. About 20% of the identified OTUs were shared across all life stages, from which 28 OTUs were consistently found in all life stage replicates. Among these OTUs there are members of genera Pantoea, Erwinia, Agrobacterium, Ochrobactrum, Pseudomonas, Acinetobacter, Brachybacterium, Sphingomonas and Methylobacterium, which can be considered as the gut-associated core microbiota of H. hampei. Our findings bring additional data to enrich the understanding of gut microbiota in CBB and its possible use for development of insect control strategies.

Coffee Berry Borer (Hypothenemus hampei) Emergence from Ground Fruits Across Varying Altitudes and Climate Cycles, and the Effect on Coffee Tree Infestation.

During coffee harvest, picked berries fall to the ground where they serve as a reservoir for the coffee berry borer (CBB) which then infest coffee berries on the trees. This study tested the effect of fallen CBB-infested coffee berries on the infestation of coffee trees (Coffea arabica). Three-year-old trees were treated with either 0, 1, 5, 10, 15, or 20 CBB-infested berries placed on the root vicinity. The CBB infestation of coffee trees was sampled every 30 days during 6 months for four coffee productive cycles. The experiment was set up at four different locations comprising different altitudes (1,218; 1,381; 1,470; and 1,700 m.a.s.l.) and the measurements were taken during 4 years where the climatic events of El Niño, La Niña, Neutral, and transitions El Niño/La Niña were present. The results show that CBB-infested berries left on the ground are a reservoir of CBB for 140 ± 8.2 days and infest developing healthy coffee berries. In a climate Neutral year, one CBB-infested ground berry left on the ground infested on average 590.2 ± 142.2 berries in coffee trees grown at 1,218 m.a.s.l. At the same altitude, one CBB-infested ground berry resulted in 151.5 ± 29.1 infested tree berries during La Niña year and 959.0 ± 89.6 during El Niño year. The CBB infestation was positively correlated with temperature and negatively correlated with altitude (R2= 0.99 and R2= -0.96, respectively). This study highlights the importance of careful harvesting practices to prevent berries from falling to the ground, followed by ground sanitation to limit later infestation of the coffee crop.

A coffee berry borer (Hypothenemus hampei) genome assembly reveals a reduced chemosensory receptor gene repertoire and male-specific genome sequences.

Coffee berry borer-CBB (Hypothenemus hampei) is a globally important economic pest of coffee (Coffea spp.). Despite current insect control methods for managing CBB, development of future control strategies requires a better understanding of its biology and interaction with its host plant. Towards this objective, we performed de novo CBB genome and transcriptome sequencing, improved CBB genome assembly and predicted 18,765 protein-encoding genes. Using genome and transcriptome data, we annotated the genes associated with chemosensation and found a reduced gene repertoire composed by 67 odorant receptors (ORs), 62 gustatory receptors (GRs), 33 ionotropic receptors (IRs) and 29 odorant-binding proteins (OBPs). In silico transcript abundance analysis of these chemosensory genes revealed expression enrichment in CBB adults compared with larva. Detection of differentially expressed chemosensory genes between males and females is likely associated with differences in host-finding behavior between sexes. Additionally, we discovered male-specific genome content and identified candidate male-specific expressed genes on these scaffolds, suggesting that a Y-like chromosome may be involved in the CBB's functional haplodiploid mechanism of sex determination.

Expression of the Wilms' tumour gene and its association with PPARß/δ in healthy skin and melanoma of horses.

The Wilms' tumour gene (WT1) has previously been described as an oncogene in several neoplasms of humans, including melanoma, and its expression increases cancer cell proliferation. Recent reports associate the expression of the PPARß/δ gene (peroxisome proliferator-activated receptor beta/delta) with the downregulation of WT1 in human melanoma and murine melanoma cell lines. The aim of this work was to analyse the expression of WT1 and its association with PPARß/δ in samples of healthy and melanoma-affected skin of horses by immunohistochemistry. WT1 protein expression was detected in healthy skin, mainly in the epidermis, hair follicle, sebaceous gland and sweat gland, while no expression was observed in equine melanoma tissues. Moreover, it was observed that PPARß/δ has a basal expression in healthy skin and that it is overexpressed in melanoma. These results were confirmed by a densitometric analysis, where a significant increase of the WT1-positive area was observed in healthy skin (128.66 ± 19.84 pixels 106) compared with that observed in melanoma (1.94 ± 0.04 pixels 106). On the other hand, a positive area with an expression of PPARß/δ in healthy skin (214.94 ± 11.85 pixels 106) was significantly decreased compared to melanoma (624.86 ± 181.93 pixels 106). These data suggest that there could be a regulation between WT1 and PPARß/δ in this disease in horses.

DBD-FISH, an effective marker for detecting genotoxicity in buccal mucosa exfoliated cells of patients with oral cancer.

Oral squamous cell carcinoma (OSCC) is characterized by increased genetic instability as an essential variable of event of neoplastic transformation. The aim of this study was to evaluate genomic instability in exfoliated cells from the buccal mucosa of patients with OSCC vs. the control group, using DNA Breakage Detection/Fluorescence In Situ hybridization (DBD-FISH). Exfoliated cells from the buccal mucosa were obtained from 38 patients with oral cancer (case group) and from 10 individuals without oral lesions (control group). DNA damage was evaluated by DBD-FISH using the whole-genome DNA probe and digital imaging analysis. Collaterally, HPV infection was determined utilizing the INNO-LiPA HPV kit. Patients with OSCC showed an increase in the hybridization signal five times more intense than that of the baseline level of DNA damage detected in control individuals. The best cutoff value for predicting oral squamous cell carcinoma was 67.46, and an Odds Ratio (OR) value of 87. HPV detection analysis revealed than one patient with OSCC (2.6%) was positive for HPV. All controls were negative HPV. In conclusion, DBD-FISH permitted the clear visualization of level high of DNA damage in the buccal epithelial cells of patients with OSSC respect to control group. Chromosome instability in oral mucosa may be an individual marker of malignant transformation in OSCC.

Evaluation of Terpene-Volatile Compounds Repellent to the Coffee Berry Borer, Hypothenemus hampei (Ferrari) (Coleoptera: Curculionidae).

The coffee berry borer (CBB) is one of the main coffee pests in the world including Colombia. This pest is difficult to manage because of its cryptic habits and the continuous availability of coffee fruits. Among the new management strategies being tested is the use of volatile compounds as insect repellents. In this work, the behavioral response of female adult CBBs to terpenes previously identified in the CBB-repellent plant species Lantana camara was evaluated. α-Terpinene, (R)-limonene, farnesene and ß-caryophyllene terpenes were tested via a Y-tube olfactometer in which ripe coffee fruits were accompanied by terpenes at concentrations between 25 and 200 ppm. Only ß-caryophyllene induced a significant and consistent CBB repellent effect at all tested doses. The protective effect of microencapsulated ß-caryophyllene was then determined under laboratory conditions by incorporating the terpene in a colloidosome-gel system at 2.8 × 105 ng/h in the middle of coffee fruits with adult CBBs. The coffee fruits in turn presented a decrease in fruit infestation. Furthermore, the protection of coffee fruits when ß-caryophyllene gels were hung in coffee trees was evaluated in the field; infestations were artificially induced by the use of raisins (CBB-infested old coffee fruits) placed on the ground. Compared with unprotected trees, the trees treated with caryophyllene gels exhibited a 33 to 45% lower degree of infestation. Taken together, the results show that ß-caryophyllene is a promising compound for an integrated pest management (IPM) program in commercial coffee plantations.

IMMUNEPOTENT CRP plus doxorubicin/cyclophosphamide chemotherapy remodel the tumor microenvironment in an air pouch triple-negative breast cancer murine model.

In 1889, Steven Paget postulated the theory that cancer cells require a permissive environment to grow. This permissive environment is known as the tumor microenvironment (TME) and nowadays it is evident that the TME is involved in the progression and response to therapy of solid cancer tumors. Triple-negative breast cancer is one of the most lethal types of cancer for women worldwide and chemotherapy remains the standard treatment for these patients. IMMUNEPOTENT CRP is a bovine dialyzable leukocyte extract with immunomodulatory and antitumor properties. The combination of chemotherapy and IMMUNEPOTENT CRP improves clinical parameters of breast cancer patients. In the current study, we aimed to evaluate the antitumor effect of doxorubicin/cyclophosphamide chemotherapy plus IMMUNEPOTENT CRP and its impact over the tumor microenvironment in a triple-negative breast cancer murine model. We evaluated CD8+, CD4+, T regulatory cells, memory T cells, myeloid-derived suppressor cells, CD71+, innate effector cells and molecules such as α-SMA, VEGF, CTLA-4, PD-L1, Gal-3, IDO, IL-2, IFN-γ, IL-12, IL-6, MCP-1, and IL-10 as part of the components of the TME. Doxorubicin/cyclophosphamide + IMMUNEPOTENT CRP decreased tumor volume, prolonged survival, increased infiltrating and systemic CD8+ T cells and decreased tumor suppressor molecules (such as PD-L1, Gal-3, and IL-10 among others). In conclusion, we suggest that IMMUNEPOTENT CRP act as a modifier of the TME and the immune response, potentiating or prolonging anti-tumor effects of doxorubicin/cyclophosphamide in a triple-negative breast cancer murine model.

ABCB1/4 gallbladder cancer risk variants identified in India also show strong effects in Chileans.

BACKGROUND: The first large-scale genome-wide association study of gallbladder cancer (GBC) recently identified and validated three susceptibility variants in the ABCB1 and ABCB4 genes for individuals of Indian descent. We investigated whether these variants were also associated with GBC risk in Chileans, who show the highest incidence of GBC worldwide, and in Europeans with a low GBC incidence. METHODS: This population-based study analysed genotype data from retrospective Chilean case-control (255 cases, 2042 controls) and prospective European cohort (108 cases, 181 controls) samples consistently with the original publication. RESULTS: Our results confirmed the reported associations for Chileans with similar risk effects. Particularly strong associations (per-allele odds ratios close to 2) were observed for Chileans with high Native American (=Mapuche) ancestry. No associations were noticed for Europeans, but the statistical power was low. CONCLUSION: Taking full advantage of genetic and ethnic differences in GBC risk may improve the efficiency of current prevention programs.

Spatial Distribution of Mealybugs (Hemiptera: Coccomorpha: Coccoidea) in the Root System of Pruned and Non-pruned Coffea arabica Trees.

Root mealybugs are important pests of coffee in many parts of the world. Despite the importance of these insects very little is known about their biology, ecology, and coffee susceptibility at different developmental stages. This study determined the effects of coffee tree pruning on root biomass, the number and within-plant spatial distribution of root-associated mealybugs, and weed-mealybug associations near coffee root systems. We made destructive root sampling of pruned and non-pruned coffee trees every 3 mo during the first-year post-pruning. In each sampling, we quantified coffee root biomass, and the number and spatial distribution of mealybug genera. Mealybugs feeding on weed roots were sampled every 2 wk for the duration of 10 wk. We identified the mealybug genera Puto Signoret (Hemiptera: Putoidae), Dysmicoccus Ferris (Hemiptera: Pseudococcidae), Ripersiella Tinsley (Hemiptera: Rhizoecidae), Rhizoecus Künckel (Hemiptera: Rhizoecidae), and Geococcus Green (Hemiptera: Pseudococcidae) associated with coffee roots. Species from Puto and Ripersiella were the most abundant representing 91% of the total mealybugs found in non-pruned trees, and 75% of the ones found in pruned trees. Coffee tree pruning reduced root biomass by 60% and mealybug numbers during the first 9 mo post-pruning. Mealybugs associated with a variety of weeds growing near coffee roots increased their number during the first 6 wk after coffee pruning. Our results suggest that coffee tree pruning causes a high percentage of root death, which correlates with a decrease of root-associated mealybugs that probably migrate to weed roots. This study provides basic information for designing mealybug control strategies in pruned coffee trees. Las cochinillas de las raíces son plagas importantes del café en varias partes del mundo. A pesar de la importancia de estos insectos se conoce muy poco sobre su biologia, ecología y sobre la susceptibilidad de plantas de café en diferentes etapas de desarrrollo. Este estudio determinó el efecto de la poda de árboles de café en la biomasa de raíces, el número y la distribución espacial in-planta de cochinillas asociadas a las raíces, y asociaciones de cochinillas con arvenses en el plato de los árboles de café. Se realizaron muestreos destructivos de raíces en árboles de café podados y no podados cada tres meses durante el primer año después de la poda. Las cochinillas alimentándose de raíces de arvenses fueron muestreadas cada dos semanas durante diez semanas. En cada muestreo se cuantificó la biomasa de raíces de café, se identificó el género de cochinillas, su cantidad y su distribución espacial. Se identificaron los géneros de cochinillas Puto Signoret (Hemiptera: Putoidae), Dysmicoccus Ferris (Hemiptera: Pseudoccocidae), Ripersiella Tinsley (Hemiptera: Rhizoecidae), Rhizoecus Künckel (Hemiptera: Rhizoecidae), y Geococcus Green (Hemiptera: Pseudococcidae) asociados a raíces de café. Especies de Puto y Ripersiella fueron las más abundantes representando el 91% del total de cochinillas encontradas en árboles no podados y el 75% de aquellas encontradas en árboles podados. La poda de árboles de café redujo la biomasa de raíces en un 60% y el número de cochinillas durante los primeros nueve meses después de la poda. Las cochinillas asociadas a varias especies de arvenses presentes en los platos de los árboles de café incrementaron su número durante las primeras seis semanas después de la poda de los árboles. Los resultados de este estudio sugieren que la poda de árboles de café causa una alta muerte de raíces, lo cual se correlaciona con una reducción en las cochinillas quienes probablemente migran hacia las raíces de las arvenses. Este estudio proporciona información básica para el diseño de estrategias de control de cochinillas en árboles de café podados.

Inhibitory Effect of Cuphea aequipetala Extracts on Murine B16F10 Melanoma In Vitro and In Vivo.

Cuphea aequipetala (C. aequipetala) has been used in Mexican traditional medicine since prehispanic times to treat tumors. In this paper, we evaluated the antiproliferative and apoptotic effect of the methanolic and aqueous extracts of C. aequipetala on several cancer cell lines including the B16F10 cell line of murine melanoma and carried a murine model assay. In vitro assay analyzed the effect in the cellular cycle and several indicators of apoptosis, such as the caspase-3 activity, DNA fragmentation, phosphatidylserine exposure (Annexin-V), and induction of cell membrane permeabilization (propidium iodide) in the B16F10 cells. In vivo, groups of C57BL/6 female mice were subcutaneously injected with 5x105 B16F10 cells and treated with 25 mg/mL of C. aequipetala extracts via oral. Aqueous and methanolic extracts showed a cytotoxic effect in MCF-7, HepG2, and B16F10 cell lines. The methanolic extract showed more antiproliferative effect with less concentration, and for this reason, the in vitro experiments were only continued with it. This extract was able to induce accumulation of cells on G1 phase of the cell cycle; moreover, it was able to induce DNA fragmentation and increase the activity of caspase-3 in B16F10 cells. On the other hand, in the murine model of melanoma, the aqueous extract showed a greater reduction of tumor size in comparison with the methanolic extract, showing an 80% reduction versus one of around 31%, both compared with the untreated control, indicating a better antitumor effect of C. aequipetala aqueous extract via oral administration. In conclusion, the in vitro data showed that both C. aequipetala extracts were able to induce cytotoxicity through the apoptosis pathway in B16F10 cells, and in vivo, the oral administration of aqueous extract reduces the melanoma tumoral mass, suggesting an important antitumoral effect and the perspective to search for effector molecules involved in it.