Activity of Bacteriophage D29 Loaded on Nanoliposomes against Macrophages Infected with Mycobacterium tuberculosis.

The search for new antimicrobial agents is a continuous struggle, mainly because more and more cases of resistant strains are being reported. Mycobacterium tuberculosis (MTB) is the main microorganism responsible for millions of deaths worldwide. The development of new antimicrobial agents is generally aimed at finding strong interactions with one or more bacterial receptors. It has been proven that bacteriophages have the ability to adhere to specific and selective regions. However, their transport and administration must be carefully evaluated as an excess could prevent a positive response and the bacteriophages may be eliminated during their journey. With this in mind, the mycobacteriophage D29 was encapsulated in nanoliposomes, which made it possible to determine its antimicrobial activity during transport and its stability in the treatment of active and latent Mycobacterium tuberculosis. The antimicrobial activity, the cytotoxicity in macrophages and fibroblasts, as well as their infection and time-kill were evaluated. Phage nanoencapsulation showed efficient cell internalization to induce MTB clearance with values greater than 90%. Therefore, it was shown that nanotechnology is capable of assisting in the activity of degradation-sensitive compounds to achieve better therapy and evade the immune response against phages during treatment.

Changes in clinical characteristics and outcomes of patients hospitalized with COVID-19 during two years of the pandemic: experience in a venezuelan hospital. / Cambios en las características clínicas y desenlaces de pacientes hospitalizados por COVID-19 durante dos años de pandemia: experiencia en un hospital venezolano.

OBJECTIVES.: To determine changes in the clinical characteristics and in-hospital outcomes of patients hospitalized for COVID-19 in a private hospital in Caracas during two years of the pandemic. MATERIALS AND METHODS.: Retrospective, observational study of patients hospitalized for COVID-19. We evaluated the correspondence between waves of hospital admissions and circulating variants of SARS-CoV-2 in the general population of the Capital District and Miranda state. RESULTS.: A total of 1025 patients (569 men and 456 women) were included, with a mean age of 62.9 SD: 16.2 years. Four waves of hospital admissions were identified: first (March-November 2020) 150/1025 (14.6%) cases; second (December 2020 to May 2021) 415/1025 (40.5%) cases; third (June-December 2021) 344/1025 (33.6%) cases; fourth (January-February 2022) 116/1025 (11.3%) cases. The mean age was higher in the fourth wave (first: 64.0±15.7, second: 61.4±15.8, third: 62.1±16.5, and fourth wave: 68.5±16.4), while the proportion of male patients (first: 66.7%, second: 58.8%, third: 50.3%, and fourth wave: 44.8%), patients with severe-critical illness (first: 65.3%, second: 57%, third: 51.7%, and fourth wave: 44.8%), in-hospital stay (first: 9.1±6.0, second: 9.0±7.3, third: 8.8±7.7, and fourth wave: 6.9±5.0 days), ICU admissions (first: 23.3%, second: 15.7%, third: 14.0%, and fourth wave: 11.2%; p=0.027) and mortality (first: 21. 8%, second: 10.7%, third: 9.1%, and fourth wave: 7.1%; p<0.001) progressively decreased over time. CONCLUSIONS.: The results show lower frequency of severe cases and improvement of in-hospital outcomes in two years of the pandemic. Changes in circulating variants, improvements in disease management and vaccination are likely to have influenced these results.

OBJETIVOS.: Determinar los cambios en las características clínicas y desenlaces intrahospitalarios de los pacientes hospitalizados por COVID-19 en un hospital privado de Caracas durante dos años de pandemia. MATERIALES Y MÉTODOS.: Estudio retrospectivo, observacional, de pacientes hospitalizados por COVID-19. Se investigó la correspondencia entre las olas de ingresos hospitalarios con las variantes circulantes del SARS-CoV-2 en la población general del Distrito Capital y estado Miranda. RESULTADOS.: Se incluyeron 1025 pacientes (569 hombres y 456 mujeres), con edad promedio de 62,9 DE: 16,2 años. Cuatro olas de ingresos hospitalarios fueron identificadas: primera (marzo-noviembre 2020) 150/1025 (14,6%) casos; segunda (diciembre-2020 a mayo-2021) 415/1025 (40,5%) casos; tercera (junio-diciembre 2021) 344/1025 (33,6%) casos; cuarta (enero-febrero 2022) 116/1025 (11,3%) casos. La edad promedio fue mayor en la cuarta ola (primera 64,0±15,7, segunda 61,4±15,8, tercera 62,1±16,5, y cuarta ola 68,5±16,4), mientras que la proporción de pacientes masculinos (primera 66,7%, segunda 58,8%, tercera 50,3%, y cuarta 44,8%), los pacientes con enfermedad grave-crítica (primera 65,3%, segunda 57%, tercera 51,7% y cuarta 44,8%), la estadía intrahospitalaria (primera 9,1±6,0, segunda 9,0±7,3, tercera 8,8±7,7, y cuarta 6,9±5,0 días), los ingresos a la UCI (primera 23,3%, segunda 15,7%, tercera 14,0%, y cuarta 11,2%; p=0,027) y la mortalidad (primera 21.8%, segunda 10,7%, tercera 9,1%, y cuarta 7,1%; p<0,001) disminuyeron progresivamente con el tiempo. CONCLUSIONES.: Los resultados muestran menor frecuencia de casos severos y mejoría de los desenlaces intrahospitalarios en dos años de pandemia. Es probable que los cambios en las variantes circulantes, las mejoras del manejo de la enfermedad y la vacunación hayan influido sobre estos resultados.

Antibacterial activity of a new monocarbonyl analog of curcumin MAC 4 is associated with divisome disruption.

Curcumin (CUR) is a symmetrical dicarbonyl compound with antibacterial activity. On the other hand, pharmacokinetic and chemical stability limitations hinder its therapeutic application. Monocarbonyl analogs of curcumin (MACs) have been shown to overcome these barriers. We synthesized and investigated the antibacterial activity of a series of unsymmetrical MACs derived from acetone against Mycobacterium tuberculosis and Gram-negative and Gram-positive species. Phenolic MACs 4, 6 and 8 showed a broad spectrum and potent activity, mainly against M. tuberculosis, Acinetobacter baumannii and methicillin-resistant Staphylococcus aureus (MRSA), with MIC (minimum inhibitory concentration) values ranging from 0.9 to 15.6 µg/mL. The investigation regarding toxicity on human lung cells (MRC-5 and A549 lines) revealed MAC 4 was more selective than MACs 6 and 8, with SI (selectivity index) values ranging from 5.4 to 15.6. In addition, MAC 4 did not demonstrate genotoxic effects on A549 cells and it was more stable than CUR in phosphate buffer (pH 7.4) for 24 h at 37 °C. Fluorescence and phase contrast microscopies indicated that MAC 4 has the ability to disrupt the divisome of Bacillus subtilis without damaging its cytoplasmic membrane. However, biochemical investigations demonstrated that MAC 4 did not affect the GTPase activity of B. subtilis FtsZ, which is the main constituent of the bacterial divisome. These results corroborated that MAC 4 is a promising antitubercular and antibacterial agent.

Benzofuroxan Derivatives as Potent Agents against Multidrug-Resistant Mycobacterium tuberculosis.

Tuberculosis (TB) is currently the leading cause of death related to infectious diseases worldwide, as reported by the World Health Organization. Moreover, the increasing number of multidrug-resistant tuberculosis (MDR-TB) cases has alarmed health agencies, warranting extensive efforts to discover novel drugs that are effective and also safe. In this study, 23 new compounds were synthesized and evaluated in vitro against the drug-resistant strains of M. tuberculosis. The compound 6-((3-fluoro-4-thiomorpholinophenyl)carbamoyl)benzo[c][1,2,5]oxadiazole 1-N-oxide (5 b) was particularly remarkable in this regard as it demonstrated MIC90 values below 0.28 µM against all the MDR strains evaluated, thus suggesting that this compound might have a different mechanism of action. Benzofuroxans are an attractive new class of anti-TB agents, exemplified by compound 5 b, with excellent potency against the replicating and drug-resistant strains of M. tuberculosis.

Antibacterial activity of 3,3'-dihydroxycurcumin (DHC) is associated with membrane perturbation.

Curcumin is a plant diphenylheptanoid and has been investigated for its antibacterial activity. However, the therapeutic uses of this compound are limited due to its chemical instability. In this work, we evaluated the antimicrobial activity of diphenylheptanoids derived from curcumin against Gram-positive and Gram-negative bacteria, and also against Mycobacterium tuberculosis in terms of MIC (Minimum Inhibitory Concentration) and MBC (Minimum Bactericidal Concentration) values. 3,3'-Dihydroxycurcumin (DHC) displayed activity against Enterococcus faecalis, Staphylococcus aureus and M. tuberculosis, demonstrating MIC values of 78 and 156 µg/mL. In addition, DHC was more stable than curcumin in acetate buffer (pH 5.0) and phosphate buffer (pH 7.4) for 24 h at 37 °C. We proposed that membrane and the cell division protein FtsZ could be the targets for DHC due to that fact that curcumin exhibits this mode of antibacterial action. Fluorescence microscopy of Bacillus subtilis stained with SYTO9 and propidium iodide fluorophores indicated that DHC has the ability to perturb the bacterial membrane. On the other hand, DHC showed a weak inhibition of the GTPase activity of B. subtilis FtsZ. Toxicity assay using human cells indicated that DHC has moderate capacity to reduce viability of liver cells (HepG2 line) and lung cells (MRC-5 and A549 lines) when compared with doxorubicin. Alkaline comet assay indicated that DHC was not able to induce DNA damage in A549 cell line. These results indicated that DHC is promising compound with antibacterial and antitubercular activities.

Apoptosis Caused by Triterpenes and Phytosterols and Antioxidant Activity of an Enriched Flavonoid Extract from Passiflora mucronata.

BACKGROUND: P. mucronata (Pm) comes from South America, Brazil and is characterized as "Maracujá de Restinga". It is used in folk medicine for its soothing properties and in treating insomnia. OBJECTIVE: The present study for the first time analyzed the antioxidant and cytotoxicity of the hydroalcoholic leaves extract and fractions from Pm. METHOD: The cytotoxicity test will be evaluated by different assays (MTT and CV) against human prostate cancer (PC3) and mouse malignant melanoma (B16F10) cell lines, and the antioxidant test by DPPH method. RESULTS: ß-Amyrin, oleanolic acid, ß-sitosterol and stigmasterol were isolated of the most active, hexane fraction. These substances were tested against the tumor cell lines: ß-sitosterol and stigmasterol showed the most relevant activity to PC3 in CV assay and, oleanolic acid to B16F10 by the MTT assay. In addition, it was possible to indicate that the mode of cell death for stigmasterol, presumably is apoptosis. In terms of antioxidant activity, the hydroalcoholic leaves extract presented higher activity (EC50 133.3 µg/mL) compared to the flower (EC50 152.3 µg/mL) and fruit (EC50 207.9 µg/mL) extracts. By the HPLC-MS, it was possible to identify the presence of flavones in the leaf extract (isoschaftoside, schaftoside, isovitexin, vitexin, isoorientin, orientin). CONCLUSIONS: P. mucronata hexane fraction showed promising cytotoxic effect against cancer cell lines, and stigmasterol contributes to this activity, inducing apoptosis of these cells. Furthermore, as other Passiflora species, Pm extract showed antioxidant activity and flavones are its major phenolic compounds.

Biological activities and phytochemical profile of Passiflora mucronata from the Brazilian restinga

ABSTRACT In general, Passiflora species have been reported for their folk medicinal use as sedative and anti-inflammatory. However, P. caerulea has already been reported to treat pulmonary diseases. Severe pulmonary tuberculosis, generally caused by Mycobacterium tuberculosis strains resistant to multiple drugs, can lead to deleterious inflammation and high mortality, encouraging new approaches in drug discovery. Thus, the aim of this work was to evaluate the Passiflora mucronata Lam., Passifloraceae, potential for tuberculosis treatment. Specifically, related to antimycobacterial activity and anti-inflammatory related effects (based on inhibition of nitric oxide, tumor necrosis factor-alpha production and antioxidant potential), as well as the chemical profile of P. mucronata. High performance liquid chromatography coupled with diode-array ultraviolet and mass spectrometer analyses of crude hydroalcoholic extract and ethyl acetate fraction showed the presence of flavonoids. Ethyl acetate fraction showed to be as antioxidant as Ginkgo biloba standard extract with EC50 of 14.61 ± 1.25 µg/ml. One major flavonoid isolated from ethyl acetate fraction was characterized as isoorientin. The hexane fraction and its main isolated compound, the triterpene β-amyrin, exhibited significant growth inhibitory activity against Mycobacterium bovis BCG (MIC50 1.61 ± 1.43 and 3.93 ± 1.05 µg/ml, respectively). In addition, Passiflora mucronata samples, specially hexane and dichloromethane fractions, as well as pure β-amyrin, showed a dose-related inhibition of lipopolysaccharide (LPS)-induced nitric oxide production. In conclusion, Passiflora mucronata presented relevant biological potential and should be considered for further studies using in vivo pulmonary tuberculosis model.

Antibacterial and Antitubercular Activities of Cinnamylideneacetophenones.

Cinnamaldehyde is a natural product with broad spectrum of antibacterial activity. In this work, it was used as a template for design and synthesis of a series of 17 cinnamylideneacetophenones. Phenolic compounds 3 and 4 exhibited MIC (minimum inhibitory concentration) and MBC (minimum bactericidal concentration) values of 77.9 to 312 µM against Staphylococcus aureus, Streptococcus mutans, and Streptococcus sanguinis. Compounds 2, 7, 10, and 18 presented potent effects against Mycobacterium tuberculosis (57.2 µM ≤ MIC ≤ 70.9 µM). Hydrophilic effects caused by substituents on ring B increased antibacterial activity against Gram-positive species. Thus, log Po/w were calculated by using high-performance liquid chromatography-photodiode array detection (HPLC-PDA) analyses, and cinnamylideneacetophenones presented values ranging from 2.5 to 4.1. In addition, the effects of 3 and 4 were evaluated on pulmonary cells, indicating their moderate toxicity (46.3 µM ≤ IC50 ≤ 96.7 µM) when compared with doxorubicin. Bioactive compounds were subjected to in silico prediction of pharmacokinetic properties, and did not violate Lipinski's and Veber's rules, corroborating their potential bioavailability by an oral route.

Bis(diphenylphosphino)amines-containing ruthenium cymene complexes as potential anti-Mycobacterium tuberculosis agents.

Several ruthenium complexes have been investigated regarding anti-Mycobacterium tuberculosis (anti-MTb) activity, with some diphosphine-containing ruthenium complexes comparable to first and second line drugs. However, to the best of our knowledge, there is no PNP-containing ruthenium complexes applied as metallodrugs. Thus, this study focused on the synthesis, characterization and anti-MTb activity of a new series of coordination compounds with general formula [RuCl(η6-p-cymene)(PNRP)]X (R=CH2Py (Py=pyridine)-[1a], CH2Ph (Ph=phenyl)-[1b], Ph-[1c] and p-tol (p-tol=p-tolyl)-[1d]; X=PF6- or BF4-). The complexes were fully characterized by NMR (1H, 31P{1H}), vibrational spectroscopy (FTIR), ESI-MS, molar conductance, elemental analysis and X-ray diffraction studies. The molecular structures of [1a]PF6, [1c]BF4 and [1d]PF6 were determined and confirm the spectroscopic and ESI-MS data. The complexes were used in anti-MTb trials, and the preliminary results are presented. The complexes are promising anti-MTb agents with MIC90 (Minimum Inhibitory Concentration of compounds required to inhibit the growth of 90% of MTb) values comparable with the Ethambutol, the reference drug used in this work, and complex [1a]BF4 presented the highest selectivity index.

Spatialization of Brazilian pig production: relationship between productive, physical, environmental, and socio-economic variables.

Brazilian pig production spans over a large territory encompassing regions of different climatic and socio-economic realities. Production, physical, socio-economic, and environmental data were used to characterize pig production in the country. Multivariate analysis evaluated indices including number productivity, production levels, and income from pigs, together with the average area of pig farm and socio-economic variables such as municipal human development index, technical guidance received from agricultural cooperatives and industrial companies, number of family farms, and offtake; and finally, environmental variables: latitude, longitude, annual temperature range, solar radiation index, as well as temperature and humidity index. The Southern region has the largest herd, number of pigs sold/sow, and offtake rate (p < 0.05), followed by the Midwest and Southeast. No significant correlations were seen between production rates and productivity with the socio-economic and environmental variables in the regions of Brazil. Production indexes, productivity, and offtake rate discriminated Northeast and Midwest and Northeast and Southeast regions. The Northern region, with a large area, has few and far-between farms that rear pigs for subsistence. The Northeast region has large herds, but low productivity. Number of slaughtered pigs has been variable over the past three decades, with few states responsible for maintaining high production in Brazil. However, the activity can be effective in any region of the country with technology and technical assistance adapted to regional characteristics.

Role of the acidic tail of high mobility group protein B1 (HMGB1) in protein stability and DNA bending.

High mobility group box (HMGB) proteins are abundant nonhistone proteins found in all eukaryotic nuclei and are capable of binding/bending DNA. The human HMGB1 is composed of two binding motifs, known as Boxes A and B, are L-shaped alpha-helix structures, followed by a random-coil acidic tail that consists of 30 Asp and Glu residues. This work aimed at evaluating the role of the acidic tail of human HMGB1 in protein stability and DNA interactions. For this purpose, we cloned, expressed and purified HMGB1 and its tailless form, HMGB1ΔC, in E. coli strain. Tryptophan fluorescence spectroscopy and circular dichroism (CD) experiments clearly showed an increase in protein stability promoted by the acidic tail under different conditions, such as the presence of the chemical denaturant guanidine hydrochloride (Gdn.HCl), high temperature and low pH. Folding intermediates found at low pH for both proteins were denatured only in the presence of chemical denaturant, thus showing a relatively high stability. The acidic tail did not alter the DNA-binding properties of the protein, although it enhanced the DNA bending capability from 76° (HMGB1ΔC) to 91° (HMGB1), as measured using the fluorescence resonance energy transfer technique. A model of DNA bending in vivo was proposed, which might help to explain the interaction of HMGB1 with DNA and other proteins, i.e., histones, and the role of that protein in chromatin remodeling.

Ethanolic extract of Casearia sylvestris and its clerodane diterpen (caseargrewiin F) protect against DNA damage at low concentrations and cause DNA damage at high concentrations in mice's blood cells.

Casearia sylvestris is used in Brazil as a popular medicine to treat ulcer, inflammation and tumour. Caseargrewiin F is a clerodane diterpene isolated from the ethanolic leaf extract of C.sylvestris. The aim of the study was to assess the capacity of the ethanolic extract of C.sylvestris leaves and caseargrewiin F to protect DNA and verify if both the compounds cause some DNA damage, using the micronucleus (MN) test and comet assay in mice. Balb-C mice were treated with the extract [3.13, 6.25, 12.5, 25, 50 and 75 mg/kg body weight (b.w.)] and caseargrewiin F (0.16, 0.32, 0.63, 1.3, 2.5 and 3.8 mg/kg b.w.) for 14 days. On day 15, DNA damage was induced by intra-peritoneal (i.p.) injection of cyclophosphamide (CP) (i.p.) at 50 mg/kg b.w. after the MN test and comet assay were performed. A protective effect of ethanolic extract was observed in MN test (6.25 and 12.5 mg/kg b.w.) and the comet assay (3.13 and 6.25, 12.5 and 25 mg/kg b.w.). Caseargrewiin F showed protective effect at 0.63, 1.3 and 2.5 mg/kg b.w. only in comet assay. We also tested the ability of compounds of C.sylvestris to induce MN and to increase the comet assay tail moment. The experimental design was similar to the DNA protection assay except that in test groups we omitted the CP challenge. We observed increased damage at 50 and 75 mg/kg b.w. of ethanolic extract of C.sylvestris and caseargrewiin F at 3.18 mg/kg b.w. in both the MN test and comet assay. We conclude that ethanolic extract of C. sylvestris and caseargrewiin F can protect cells against DNA damage induced by CP at low concentrations, but at high concentrations these compounds also induce DNA damage.

Enfermedad periodontal y preeclampsia: revisión de la literatura / Periodontal disease and preeclampsia: literature review

Introducción: La preeclampsia es una complicación del embarazo que puede comprometer tanto la vida de la madre, como la del bebé. Existen diferentes factores etiológicos para el desarrollo de la preeclampsia, incluidos las infecciones crónicas. La Enfermedad Periodontal, como infección crónica, recientemente ha sido asociada con la aparición de la preeclampsia. Objetivo: Realizar una revisión de la literatura, para constatar la posible asociación entre Enfermedad Periodontal (EP) y Preeclampsia. Métodos y Materiales: Búsqueda en las bases de datos de la red de publicaciones científicas con el fin de revisar los estudios reportados a partir de 1996 hasta 2007, para evaluar la posible asociación entre EP y Preeclampsia. Resultados: Se encontraron 11 estudios y se evaluaron sus diseños y sus resultados. Conclusiones: De esta revisión se concluye que existen muy pocos estudios dirigidos a evaluar la presencia de EP y la incidencia de Preeclampsia. Los estudios evaluados muestran resultados contradictorios lo que se puede deber a los diferentes diseños de los estudios ya que se encontró falta de unificación en los criterios de inclusión en los estudios, falta de unificación en los criterios de diagnóstico para la EP y el tamaño de la muestra es muy diferente según el tipo de estudio. Se necesitan más estudios de intervención, en los cuales se demuestre que al realizar un control o eliminación de la EP, las complicaciones del embarazo descritas anteriormente disminuyen, para reafirmar o descartar esas posibles asociaciones.

Introduction: Preeclampsia is a complication of pregnancy that can threaten both the life of the mother as the baby. There are different etiological factors for the development of preeclampsia, including chronic infections. Periodontal disease as a chronic infection has recently been associated with the onset of preeclampsia. Objective: To review the literature with the purpose of determine the possible association between periodontal disease (PD) and Preeclampsia. Methods and Materials: Database search of scientific publications in order to review studies reported from 1996 to 2007 to assess the possible association between PD and Preeclampsia. Results: There were evaluated designs and results of 11 scientific articles. Conclusions: This review concluded that there are very few studies to evaluate the presence of PD and the incidence of preeclampsia. The evaluated studies show conflicting results which may be due to the different designs of the studies, lack of unification among inclusion criteria between them, lack of standardization in diagnostics criteria for PD, and the sample size is very different depending on the type of study. More intervention studies are needed to demonstrate that controlling or eliminating PD, the previously described complications of pregnancy are minimized, in order to affirm or refute these possible associations.