Design of a New Vaccine Prototype against Porcine Circovirus Type 2 (PCV2), M. hyopneumoniae and M. hyorhinis Based on Multiple Antigens Microencapsulation with Sulfated Chitosan.

This work evaluated in vivo an experimental-multivalent-vaccine (EMV) based on three Porcine Respiratory Complex (PRC)-associated antigens: Porcine Circovirus Type 2 (PCV2), M. hyopneumoniae (Mhyop) and M. hyorhinis (Mhyor), microencapsulated with sulfated chitosan (M- ChS + PRC-antigens), postulating chitosan sulphate (ChS) as a mimetic of the heparan sulfate receptor used by these pathogens for cell invasion. The EMV was evaluated physicochemically by SEM (Scanning-Electron-Microscopy), EDS (Energy-Dispersive-Spectroscopy), Pdi (Polydispersity-Index) and zeta potential. Twenty weaned pigs, distributed in four groups, were evaluated for 12 weeks. The groups 1 through 4 were as follows: 1-EMV intramuscular-route (IM), 2-EMV oral-nasal-route (O/N), 3-Placebo O/N (M-ChS without antigens), 4-Commercial-vaccine PCV2-Mhyop. qPCR was used to evaluate viral/bacterial load from serum, nasal and bronchial swab and from inguinal lymphoid samples. Specific humoral immunity was evaluated by ELISA. M-ChS + PRC-antigens measured between 1.3-10 µm and presented low Pdi and negative zeta potential, probably due to S (4.26%). Importantly, the 1-EMV protected 90% of challenged animals against PCV2 and Mhyop and 100% against Mhyor. A significant increase in antibody was observed for Mhyor (1-EMV and 2-EMV) and Mhyop (2-EMV), compared with 4-Commercial-vaccine. No difference in antibody levels between 1-EMV and 4-Commercial-vaccine for PCV2-Mhyop was observed. Conclusion: The results demonstrated the effectiveness of the first EMV with M-ChS + PRC-antigens in pigs, which were challenged with Mhyor, PCV2 and Mhyop, evidencing high protection for Mhyor, which has no commercial vaccine available.

Antimicrobial Activity and Phytochemical Characterization of Baccharis concava Pers., a Native Plant of the Central Chilean Coast.

Few sclerophyllous plants from the central coast of Chile have been systematically studied. This work describes the phytochemical composition and antimicrobial properties of Baccharis concava Pers. (sin. B. macraei), a shrub found in the first line and near the Pacific coast. B. concava has been traditionally used by indigenous inhabitants of today's central Chile for its medicinal properties. Few reports exist regarding the phytochemistry characterization and biological activities of B. concava. A hydroalcoholic extract of B. concava was prepared from leaves and small branches. Qualitative phytochemical characterization indicated the presence of alkaloids, steroids, terpenoids, flavonoids, phenolic, and tannin compounds. The antimicrobial activity of this extract was assessed in a panel of microorganisms including Gram-positive bacteria, Gram-negative bacteria, and pathogenic yeasts. The extract displayed an important antimicrobial effect against Gram-positive bacteria, Candida albicans, and Cryptococcus neoformans but not against Gram-negatives, for which an intact Lipopolysaccharide is apparently the determinant of resistance to B. concava extracts. The hydroalcoholic extract was then fractionated through a Sephadex LH-20/methanol-ethyl acetate column. Afterward, the fractions were pooled according to a similar pattern visualized by TLC/UV analysis. Fractions obtained by this criterion were assessed for their antimicrobial activity against Staphylococcus aureus. The fraction presenting the most antimicrobial activity was HPLC-ESI-MS/MS, obtaining molecules related to caffeoylquinic acid, dicaffeoylquinic acid, and quercetin, among others. In conclusion, the extracts of B. concava showed strong antimicrobial activity, probably due to the presence of metabolites derived from phenolic acids, such as caffeoylquinic acid, and flavonoids, such as quercetin, which in turn could be responsible for helping with wound healing. In addition, the development of antimicrobial therapies based on the molecules found in B. concava could help to combat infection caused by pathogenic yeasts and Gram-positive bacteria, without affecting the Gram-negative microbiota.

Parasitic findings on threatened pudu deer from Central Chile accounts first genetic characterization of lice parasitizing P. puda in Chile and the first molecular report of Taenia hydatigena metacestodes in this species.

Southern pudu (Pudu puda) is a threatened endemic deer of the temperate forests of Chile. In recent years pudu populations rates have decreased mainly due to anthropogenic causes including forest loss and landscape fragmentation. In this context, the parasitic fauna of Chilean pudu has been scarcely investigated. The aim of this study was to determine the parasitic status of rescued pudu n = 13 from its natural habitat in Central Chile (Maule region) during March 2022 and June 2023 by applying morphological, histopathological, and molecular analyses. As result, we report the presence of transmission of parasites from dogs to pudus as showed by the presence of metacestodes of the parasite Taenia hydatigena on omentum, liver, and pleura of pudus during postmortem examinations, being the first molecular report on the presence of this parasite on Chilean pudu. Meanwhile, ectoparasite examinations determined the presence of chewing and sucking lice on pudu exemplars here analysed. Molecular and phylogenetic analysis of lice revealed new insights on Bovicola and Anoplura lice parasitizing P. puda in Chile, equally being the first genetic characterization of lice parasitizing pudu exemplars in Chile. In addition, parasite loads of lice and metacestodes were analysed. However, no statistically significance was observed when comparing environmental and individual traits influence on parasite load variation. Overall, the study area is the northern limit of habitat distribution of this specie in Chile and we here provide novel information on pudu deer parasites, thus making a useful and valuable contribution to the parasitological knowledge on this threatened species.

Comparative transcriptomic analysis of chemoreceptors in two sympatric scarab beetles, Hylamorpha elegans and Brachysternus prasinus.

Chemoreception through odorant receptors (ORs), ionotropic receptors (IRs) and gustatory receptors (GRs) represents the functions of key proteins in the chemical ecology of insects. Recent studies have identified chemoreceptors in coleopterans, facilitating the evolutionary analysis of not only ORs but also IRs and GRs. Thus, Cerambycidae, Tenebrionidae and Curculionidae have received increased attention. However, knowledge of the chemoreceptors from Scarabaeidae is still limited, particularly for those that are sympatric. Considering the roles of chemoreceptors, this analysis could shed light on evolutionary processes in the context of sympatry. Therefore, the aim of this study was to identify and compare the repertoires of ORs, GRs and IRs between two sympatric scarab beetles, Hylamorpha elegans and Brachysternus prasinus. Here, construction of the antennal transcriptomes of both scarab beetle species and analyses of their phylogeny, molecular evolution and relative expression were performed. Thus, 119 new candidate chemoreceptors were identified for the first time, including 17 transcripts for B. prasinus (1 GR, 3 IRs and 13 ORs) and 102 for H. elegans (22 GRs, 14 IRs and 66 ORs). Orthologs between the two scarab beetle species were found, revealing specific expansions as well as absence in some clades. Purifying selection appears to have occurred on H. elegans and B. prasinus ORs. Further efforts will be focused on target identification to characterize kairomone and/or pheromone receptors.

Biological evolution and human cognition are analogous information processing systems.

The mechanisms that govern biological evolution and human cognition are analogous, as both follow the same principles of natural information processing systems. In this article, we describe the following five principles that provide an analogy between biological evolution and human cognition: (a) Randomness as Genesis Principle and (b) Borrowing and Reorganizing Principle, which indicate how natural information processing systems obtain information; (c) Narrow Limits of Change Principle and (d) Information Store Principle, which indicate how information is processed and stored; and (e) Environmental Organizing and Linking Principle, which indicate how stored information is used to generate actions appropriate to an environment. In human cognition, these analogs only apply to cognitive processes associated with biologically secondary knowledge, the knowledge typically taught in educational institutions. Based on these five principles, cognitive load theory researchers have provided diverse prescriptions to optimize instructional activities and materials. We conclude by discussing general instructional implications and future research directions based on this analogy.

Human pluripotent stem cell-derived macrophages host Mycobacterium abscessus infection.

Human macrophages are a natural host of many mycobacterium species, including Mycobacterium abscessus (M. abscessus), an emerging pathogen affecting immunocompromised and cystic fibrosis patients with few available treatments. The search for an effective treatment is hindered by the lack of a tractable in vitro intracellular infection model. Here, we established a reliable model for M. abscessus infection using human pluripotent stem cell-derived macrophages (hPSC-macrophages). hPSC differentiation permitted reproducible generation of functional macrophages that were highly susceptible to M. abscessus infection. Electron microscopy demonstrated that M. abscessus was present in the hPSC-macrophage vacuoles. RNA sequencing analysis revealed a time-dependent host cell response, with differing gene and protein expression patterns post-infection. Engineered tdTOMATO-expressing hPSC-macrophages with GFP-expressing mycobacteria enabled rapid image-based high-throughput analysis of intracellular infection and quantitative assessment of antibiotic efficacy. Our study describes the first to our knowledge hPSC-based model for M. abscessus infection, representing a novel and accessible system for studying pathogen-host interaction and drug discovery.

Blocking phospholamban with VHH intrabodies enhances contractility and relaxation in heart failure.

The dysregulated physical interaction between two intracellular membrane proteins, the sarco/endoplasmic reticulum Ca2+ ATPase and its reversible inhibitor phospholamban, induces heart failure by inhibiting calcium cycling. While phospholamban is a bona-fide therapeutic target, approaches to selectively inhibit this protein remain elusive. Here, we report the in vivo application of intracellular acting antibodies (intrabodies), derived from the variable domain of camelid heavy-chain antibodies, to modulate the function of phospholamban. Using a synthetic VHH phage-display library, we identify intrabodies with high affinity and specificity for different conformational states of phospholamban. Rapid phenotypic screening, via modified mRNA transfection of primary cells and tissue, efficiently identifies the intrabody with most desirable features. Adeno-associated virus mediated delivery of this intrabody results in improvement of cardiac performance in a murine heart failure model. Our strategy for generating intrabodies to investigate cardiac disease combined with modified mRNA and adeno-associated virus screening could reveal unique future therapeutic opportunities.

Impact of dental cone-beam computed tomography on diagnostic evaluation and treatment plan for the maxillary sinus in dental implant planning / Impacto de la tomografía computarizada de haz cónico en el diagnóstico y plan detratamiento preimplantologico de senos maxilares

Objective: To determine the impact of CBCT on diagnostic evaluation and treatment plan for the maxillary sinus in dental implant planning. Material and Methods: Diagnostic evaluation and treatment plan for the maxillary sinus were evaluated by eight specialists experienced in dental implant placement. Eight panoramic radiographs (PAN) and CBCT examinations were obtained from five adult patients with a specific clinical need for dental implants. Evaluation was performed first on PAN then, at least 2 weeks later, on CBCT. Residual alveolar ridge height, mucosal thickening, radiographic findings and treatment plan were recorded. The confidence level was evaluated for both diagnostic evaluation and treatment plan. The kappa statistic for intra-observer reproducibility and McNemar test were performed. Results: In the diagnostic evaluation, CBCT showed significant impact on the diagnosis of radiographic findings. Availability of CBCT significantly changed the treatment plan, for less invasive treatment, or no treatment need. Observers had significantly greater confidence when using CBCT than PAN, when indicating presence of mucosal thickening and radiographic findings in the maxillary sinus. In addition, CBCT increased confidence in the treatment plan. Conclusion: The present study suggests that CBCT has an impact on the diagnostic evaluation of radiographic findings in the maxillary sinus and on the decision to place implants, owing to misdiagnosis of pathology and planning of more invasive treatments when using PAN. Availability of CBCT also improves clinician confidence. Further studies at higher levels of diagnostic efficacy should be performed, to justify the use of CBCT, by evaluating the actual treatment performed and its outcome.

Objetivo: Determinar el impacto de la tomografía computarizada de haz cónico (CBCT) en la evaluación diagnóstica y el plan de tratamiento del seno maxilar en la planificación de implantes dentales. Material y Métodos: La evaluación diagnóstica y el plan de tratamiento del seno maxilar fueron evaluados por ocho especialistas con experiencia en la colocación de implantes dentales. Se obtuvieron ocho radiografías panorámicas (PAN) y exámenes CBCT de cinco pacientes adultos con una necesidad clínica específica de implantes dentales. La evaluación se realizó primero en PAN y luego, al menos dos semanas después, en CBCT. Se registraron la altura del reborde alveolar residual, el engrosamiento de la mucosa, los hallazgos radiográficos y el plan de tratamiento. Se evaluó el nivel de confianza tanto para la evaluación diagnóstica como para el plan de tratamiento. Se realizó el estadístico kappa para la reproducibilidad intraobservador y la prueba de McNemar. Resultados: En la evaluación diagnóstica, CBCT mostró un impacto significativo en el diagnóstico de los hallazgos radiográficos. La disponibilidad de CBCT cambió significativamente el plan de tratamiento, para un tratamiento menos invasivo o sin necesidad de tratamiento. Los observadores tuvieron una confianza significativamente mayor al usar CBCT que PAN, al indicar la presencia de engrosamiento de la mucosa y hallazgos radiográficos en el seno maxilar. Además, CBCT aumentó la confianza en el plan de tratamiento. Conclusión: El presente estudio sugiere que la CBCT tiene un impacto en la evaluación diagnóstica de los hallazgos radiográficos en el seno maxilar y en la decisión de colocar implantes, debido al diagnóstico erróneo de la patología y la planificación de tratamientos más invasivos al usar PAN. La disponibilidad de CBCT también mejora la confianza del clínico. Se deben realizar más estudios a niveles más altos de eficacia diagnóstica para justificar el uso de CBCT, evaluando el tratamiento real realizado y su resultado.

Low molecular weight sulfated chitosan efficiently reduces infection capacity of porcine circovirus type 2 (PCV2) in PK15 cells.

BACKGROUND: Porcine circovirus type 2 (PCV2)-associated diseases are a major problem for the swine industry worldwide. In addition to vaccines, the availability of antiviral polymers provides an efficient and safe option for reducing the impact of these diseases. By virtue of their molecular weight and repetitious structure, polymers possess properties not found in small-molecule drugs. In this perspective, we focus on chitosan, a ubiquitous biopolymer, that adjusts the molecular weight and sulfated-mediated functionality can act as an efficient antiviral polymer by mimicking PCV2-cell receptor interactions. METHODS: Sulfated chitosan (Chi-S) polymers of two molecular weights were synthesized and characterized by FTIR, SEM-EDS and elemental analysis. The Chi-S solutions were tested against PCV2 infection in PK15 cells in vitro and antiviral activity was evaluated by measuring the PCV2 DNA copy number, TCID50 and capsid protein expression, upon application of different molecular weights, sulfate functionalization, and concentrations of polymer. In addition, to explore the mode of action of the Chi-S against PCV2 infection, experiments were designed to elucidate whether the antiviral activity of the Chi-S would be influenced by when it was added to the cells, relative to the time and stage of viral infection. RESULTS: Chi-S significantly reduced genomic copies, TCID50 titers and capsid protein of PCV2, showing specific antiviral effects depending on its molecular weight, concentration, and chemical functionalization. Assays designed to explore the mode of action of the low molecular weight Chi-S revealed that it exerted antiviral activity through impeding viral attachment and penetration into cells. CONCLUSIONS: These findings help better understanding the interactions of PCV2 and porcine cells and reinforce the idea that sulfated polymers, such as Chi-S, represent a promising candidates for use in antiviral therapies against PCV2-associated diseases. Further studies in swine are warranted.

Biological Effects of Quinolones: A Family of Broad-Spectrum Antimicrobial Agents.

Broad antibacterial spectrum, high oral bioavailability and excellent tissue penetration combined with safety and few, yet rare, unwanted effects, have made the quinolones class of antimicrobials one of the most used in inpatients and outpatients. Initially discovered during the search for improved chloroquine-derivative molecules with increased anti-malarial activity, today the quinolones, intended as antimicrobials, comprehend four generations that progressively have been extending antimicrobial spectrum and clinical use. The quinolone class of antimicrobials exerts its antimicrobial actions through inhibiting DNA gyrase and Topoisomerase IV that in turn inhibits synthesis of DNA and RNA. Good distribution through different tissues and organs to treat Gram-positive and Gram-negative bacteria have made quinolones a good choice to treat disease in both humans and animals. The extensive use of quinolones, in both human health and in the veterinary field, has induced a rise of resistance and menace with leaving the quinolones family ineffective to treat infections. This review revises the evolution of quinolones structures, biological activity, and the clinical importance of this evolving family. Next, updated information regarding the mechanism of antimicrobial activity is revised. The veterinary use of quinolones in animal productions is also considered for its environmental role in spreading resistance. Finally, considerations for the use of quinolones in human and veterinary medicine are discussed.

Genetic Characterization of Salmonella Infantis with Multiple Drug Resistance Profiles Isolated from a Poultry-Farm in Chile.

Salmonella comprises over 2500 serotypes and foodborne contamination associated with this pathogen remains an important health concern worldwide. During the last decade, a shift in serotype prevalence has occurred as traditionally less prevalent serotypes are increasing in frequency of infections, especially those related to poultry meat contamination. S. Infantis is one of the major emerging serotypes, and these strains commonly display antimicrobial resistance and can persist despite cleaning protocols. Thus, this work aimed to isolate S. Infantis strains from a poultry meat farm in Santiago, Chile and to characterize genetic variations present in them. We determined their genomic and phenotypic profiles at different points along the production line. The results indicate that the strains encompass 853 polymorphic sites (core-SNPs) with isolates differing from one another by 0-347 core SNPs, suggesting variation among them; however, we found discrete correlations with the source of the sample in the production line. Furthermore, the pan-genome was composed of 4854 total gene clusters of which 2618 (53.9%) corresponds to the core-genome and only 181 (3.7%) are unique genes (those present in one particular strain). This preliminary analysis will enrich the surveillance of Salmonella, yet further studies are required to assess their evolution and phylogeny.

Phospholamban antisense oligonucleotides improve cardiac function in murine cardiomyopathy.

Heart failure (HF) is a major cause of morbidity and mortality worldwide, highlighting an urgent need for novel treatment options, despite recent improvements. Aberrant Ca2+ handling is a key feature of HF pathophysiology. Restoring the Ca2+ regulating machinery is an attractive therapeutic strategy supported by genetic and pharmacological proof of concept studies. Here, we study antisense oligonucleotides (ASOs) as a therapeutic modality, interfering with the PLN/SERCA2a interaction by targeting Pln mRNA for downregulation in the heart of murine HF models. Mice harboring the PLN R14del pathogenic variant recapitulate the human dilated cardiomyopathy (DCM) phenotype; subcutaneous administration of PLN-ASO prevents PLN protein aggregation, cardiac dysfunction, and leads to a 3-fold increase in survival rate. In another genetic DCM mouse model, unrelated to PLN (Cspr3/Mlp-/-), PLN-ASO also reverses the HF phenotype. Finally, in rats with myocardial infarction, PLN-ASO treatment prevents progression of left ventricular dilatation and improves left ventricular contractility. Thus, our data establish that antisense inhibition of PLN is an effective strategy in preclinical models of genetic cardiomyopathy as well as ischemia driven HF.

Tratamiento con HeberFERON® del carcinoma basocelular en la Atención Primaria de Salud en Cuba / HeberFERON® Treatment of Basal Cell Carcinoma in Primary Healthcare in Cuba

Introducción: El cáncer de piel es el tipo de cáncer más frecuente en el ser humano, el carcinoma basocelular es el más común de todos los cánceres de piel (80-90 por ciento). Excepcionalmente producen metástasis, pero pueden causar significativa morbilidad e involucran a edades más jóvenes, se tratan con éxito mediante cirugía, radioterapia, quimioterapia y crioterapia, generalmente en el nivel secundario de salud, sin embargo, estos tratamientos no siempre son posibles o deseables. El HeberFERON® es una combinación de interferones alfa y gamma humanos recombinantes, que ha mostrado producir efectos sinérgicos en la reducción de la proliferación de varias líneas de células cancerosas, esta formulación ha sido aprobada en Cuba para el tratamiento del carcinoma basocelular. Presentación de casos: Se presentaron tres casos con diagnóstico de carcinoma basocelular, localizados en la cara, tratados con HeberFERON®, en dos casos fue observada la desaparición de la lesión al finalizar la tercera semana de tratamiento. En el tercer caso, una mujer de 84 años de edad, al finalizar el primer ciclo de tratamiento, fue reducido el tamaño de la lesión tratada y desapareció otra lesión adyacente que no recibió directamente tratamiento, la lesión residual, en esta paciente, fue valorada por ultrasonido para determinar su extensión y profundidad, en los tres casos las reacciones adversas fueron leves y transitorias. Conclusiones: El HeberFERON® es una opción efectiva y segura para el tratamiento del carcinoma basocelular en la atención primaria de salud en Cuba(AU)

Introduction: Skin cancer is the commonest type of cancer in humans. Basal cell carcinoma is the commonest of all skin cancers, accounting for 80 percent to 90 percent of all cases. Exceptionally, they metastasize, but can cause significant morbidity and involve younger ages. They are successfully treated by surgery, radiotherapy, chemotherapy and cryotherapy, mostly at the secondary level of health. However, these treatments are not always possible or desirable. HeberFERON® is a combination of recombinant human alpha and gamma interferons, which has been shown to produce synergistic effects in reducing the proliferation of several lines of cancer cells. This formulation has been approved in Cuba for treating basal cell carcinoma. Case presentation: Three cases are presented with a diagnosis of basal cell carcinoma, located on the face, treated with HeberFERON®. In two cases, the lesion was observed to disappear at the end of the third week of treatment. In the third case, an 84-year-old woman, at the end of the first cycle of treatment, the size of the treated lesion was reduced and another adjacent lesion disappeared, which did not receive direct treatment. In this patient, the residual lesion was assessed by ultrasound to determine its extension and depth. In the three cases, the adverse reactions were mild and transitory. Conclusions: HeberFERON® is an effective and safe option for treating basal cell carcinoma in primary health care in Cuba(AU)

Sex-Specific Control of Human Heart Maturation by the Progesterone Receptor.

BACKGROUND: Despite in-depth knowledge of the molecular mechanisms controlling embryonic heart development, little is known about the signals governing postnatal maturation of the human heart. METHODS: Single-nucleus RNA sequencing of 54 140 nuclei from 9 human donors was used to profile transcriptional changes in diverse cardiac cell types during maturation from fetal stages to adulthood. Bulk RNA sequencing and the Assay for Transposase-Accessible Chromatin using sequencing were used to further validate transcriptional changes and to profile alterations in the chromatin accessibility landscape in purified cardiomyocyte nuclei from 21 human donors. Functional validation studies of sex steroids implicated in cardiac maturation were performed in human pluripotent stem cell-derived cardiac organoids and mice. RESULTS: Our data identify the progesterone receptor as a key mediator of sex-dependent transcriptional programs during cardiomyocyte maturation. Functional validation studies in human cardiac organoids and mice demonstrate that the progesterone receptor drives sex-specific metabolic programs and maturation of cardiac contractile properties. CONCLUSIONS: These data provide a blueprint for understanding human heart maturation in both sexes and reveal an important role for the progesterone receptor in human heart development.

Tomografía computarizada de haz cónico como examen concluyente de hiperplasia de procesos coronoides bilateral. Reporte de tres casos / Cone beam computed tomography as a conclusive examination of bilateral coronoid process hyperplasia. Report of three cases

RESUMEN La hiperplasia del proceso coronoides (HPC) es un crecimiento exagerado del proceso coronoides. La HPC es poco frecuente y genera limitación asintomática de la apertura bucal debido a la impactación del proceso coronoides con el segmento posterior del hueso cigomático. La HPC es un posible diagnóstico en pacientes con limitación progresiva tanto de la apertura bucal como del movimiento protrusivo mandibular. En el presente artículo, se reportan tres casos de pacientes con limitación de apertura bucal. Mediante examen de tomografía computarizada de haz cónico (TCHC), se observó un aumento de tamaño uniforme de los procesos coronoides, confirmando el diagnóstico de HPC bilateral. La TCHC con campo de visión grande determinó el diagnóstico final, pues mostró la impactación del proceso coronoides con el segmento posterior del hueso cigomático en apertura bucal. Además, la TCHC permitió distinguir entre una HPC y una neoplasia del proceso coronoides y/o estructuras anatómicas vecinas.

SUMMARY Coronoid process hyperplasia (CPH) is an excessive coronoid process growing. CPH is infrequent and produces an asymptomatic mouth opening limitation due to the impaction of the coronoid process in the posterior portion of the zygomatic bone. CPH is a possible diagnosis in patients with both progressive mouth opening and protrusive movement limitations. In the present article, cases of three patients with mouth opening limitation are reported. In cone-beam computed tomography (CBCT) examination, a uniform growing of coronoid processes was observed, which confirmed the diagnosis of bilateral CPH. CBCT with large field of view allowed the determination of the final diagnosis, because the impaction of the coronoid process in the posterior portion of the zygomatic bone during mouth opening could be observed. Furthermore, CBCT allowed to distinguish between CPH and neoplasia of coronoid process and/or neighboring anatomical structures.

First Molecular Characterization of Trichostrongylus colubriformis Infection in Rural Patients from Chile.

PURPOSE: The aim was to characterize the infection by Trichostrongylus spp. in patients from Chile using a combination of molecular detection techniques and phylogenetic analysis relating the findings to clinical and epidemiological reports of the patients METHODS: Strongylid eggs were detected in seven patients by coproparasitological techniques. From each sample a fragment of the ITS-2 ribosomal gene was amplified by PCR, sequenced and analyzed by the Neighbor-Joining method. RESULTS: All the sequences and phylogenetic clusters corresponded to T. colubriformis. Two samples presented a single nucleotide polymorphism showing two possible haplotypes. Six patients presented gastrointestinal symptoms. All of them lived on farms and used sheep manure as fertilizer. CONCLUSION: T. colubriformis was the strongylid involved in the infections of these Chilean patients associated with the presence of livestock and agricultural practices that favor infection by this type of nematode.

Inactivation of Glutamine Synthetase-Coding Gene glnA Increases Susceptibility to Quinolones Through Increasing Outer Membrane Protein F in Salmonella enterica Serovar Typhi.

Ciprofloxacin is the choice treatment for infections caused by Salmonella Typhi, however, reduced susceptibility to ciprofloxacin has been reported for this pathogen. Considering the decreased approbation of new antimicrobials and the crisis of resistance, one strategy to combat this problem is to find new targets that enhances the antimicrobial activity for approved antimicrobials. In search of mutants with increased susceptibility to ciprofloxacin; 3,216 EZ-Tn5 transposon mutants of S. Typhi were screened. S. Typhi zxx::EZ-Tn5 mutants susceptible to ciprofloxacin were confirmed by agar diffusion and MIC assays. The genes carrying EZ-Tn5 transposon insertions were sequenced. Null mutants of interrupted genes, as well as inducible genetic constructs, were produced using site-directed mutagenesis, to corroborate phenotypes. SDS-PAGE and Real-time PCR were used to evaluate the expression of proteins and genes, respectively. Five mutants with increased ciprofloxacin susceptibility were found in the screening. The first confirmed mutant was the glutamine synthetase-coding gene glnA. Analysis of outer membrane proteins revealed increased OmpF, a channel for the influx of ciprofloxacin and nalidixic acid, in the glnA mutant. Expression of ompF increased four times in the glnA null mutant compared to WT strain. To understand the relationship between the expression of glnA and ompF, a strain with the glnA gene under control of the tetracycline-inducible Ptet promoter was created, to modulate glnA expression. Induction of glnA decreased expression of ompF, at the same time that reduced susceptibility to ciprofloxacin. Expression of sRNA MicF, a negative regulator of OmpF was reduced to one-fourth in the glnA mutant, compared to WT strain. In addition, expression of glnL and glnG genes (encoding the two-component system NtrC/B that may positively regulate OmpF) were increased in the glnA mutant. Further studies indicate that deletion of glnG decreases susceptibility to CIP, while deletion of micF gene increases susceptibility CIP. Our findings indicate that glnA inactivation promotes ompF expression, that translates into increased OmpF protein, facilitating the entry of ciprofloxacin, thus increasing susceptibility to ciprofloxacin through 2 possible mechanisms.

Multivariate patterning of human pluripotent cells under perfusion reveals critical roles of induced paracrine factors in kidney organoid development.

Creating complex multicellular kidney organoids from pluripotent stem cells shows great promise. Further improvements in differentiation outcomes, patterning, and maturation of specific cell types are, however, intrinsically limited by standard tissue culture approaches. We describe a novel full factorial microbioreactor array-based methodology to achieve rapid interrogation and optimization of this complex multicellular differentiation process in a facile manner. We successfully recapitulate early kidney tissue patterning events, exploring more than 1000 unique conditions in an unbiased and quantitative manner, and define new media combinations that achieve near-pure renal cell type specification. Single-cell resolution identification of distinct renal cell types within multilayered kidney organoids, coupled with multivariate analysis, defined the definitive roles of Wnt, fibroblast growth factor, and bone morphogenetic protein signaling in their specification, exposed retinoic acid as a minimal effector of nephron patterning, and highlighted critical contributions of induced paracrine factors on cell specification and patterning.

The Transcription Factor ArcA Modulates Salmonella's Metabolism in Response to Neutrophil Hypochlorous Acid-Mediated Stress.

Salmonella Typhimurium, a bacterial pathogen with high metabolic plasticity, can adapt to different environmental conditions; these traits enhance its virulence by enabling bacterial survival. Neutrophils play important roles in the innate immune response, including the production of microbicidal reactive oxygen species (ROS). In addition, the myeloperoxidase in neutrophils catalyzes the formation of hypochlorous acid (HOCl), a highly toxic molecule that reacts with essential biomolecules, causing oxidative damage including lipid peroxidation and protein carbonylation. The bacterial response regulator ArcA regulates adaptive responses to oxygen levels and influences the survival of Salmonella inside phagocytic cells. Here, we demonstrate by whole transcriptomic analyses that ArcA regulates genes related to various metabolic pathways, enabling bacterial survival during HOCl-stress in vitro. Also, inside neutrophils, ArcA controls the transcription of several metabolic pathways by downregulating the expression of genes related to fatty acid degradation, lysine degradation, and arginine, proline, pyruvate, and propanoate metabolism. ArcA also upregulates genes encoding components of the oxidative pathway. These results underscore the importance of ArcA in ATP generation inside the neutrophil phagosome and its participation in bacterial metabolic adaptations during HOCl stress.

Cysteine auxotrophy drives reduced susceptibility to quinolones and paraquat by inducing the expression of efflux-pump systems and detoxifying enzymes in S. Typhimurium.

Currently, Salmonella enterica serovar Typhimurium (S. Typhimurium), is a major global public health problem, which has caused food-borne illnesses in many countries. Today, with the extensive use of antimicrobials, antimicrobial resistance is increasing at a serious rate in S. Typhimurium isolates. The present study sought the role of cysteine (Cys) auxotrophy on the resistance to quinolones and paraquat in S. Typhimurium. Cys auxotrophy was achieved by deleting either the cysDNC, cysJIH or cysQ loci. Deletion of these loci resulted in loss of susceptibility against nalidixic acid, levofloxacin, ciprofloxacin (CIP) and paraquat. Further studies with cysJIH mutant indicated increased expression of multi-antibiotic resistance genes marA and ramA, and consequently increased expression of efflux-pump systems. The cysJIH mutant presented a smaller increase of reactive oxygen species (ROS) in presence of paraquat or CIP. Expression of katG and sodA (expressing for a catalase and a superoxide dismutase, respectively) genes was increased in presence of paraquat in the cysJIH mutant; while expression of the superoxide dismutase gene sodB was decreased. These results indicate that deletion of cysDNC, cysJIH or cysQ genes of S. Typhimurium renders Cys auxotrophy along with decreased susceptibility in response to quinolone and paraquat. Overexpression of efflux-pump systems AcrB-TolC and SmvA-OmpD and antioxidant enzymes KatG and SodA could explain the mechanisms of antimicrobial resistance in the Cys auxotrophic mutants.