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Background: Butyrate is a histone deacetylase inhibitor that induces apoptosis and inhibits cell proliferation of colorectal cancer cells. To improve its anticancer activity, butyrate has been evaluated mixed with drugs and different molecules. Plant antimicrobial peptides are attractive anticancer alternative molecules because they show selective cytotoxic activity against different cancer cell lines. In this work, we explore if the plant defensin γ-thionin ( Capsicum chinense ) can improve butyrate activity on Caco-2 cell line and we also determined the mechanism of death activated. Results: The combined treatment of γ-thionin (3.5 µM) and butyrate (50 mM) showed higher cytotoxicity on Caco-2 cells with respect to single treatments. Also, the combined treatment reduced cell proliferation and exhibited a higher rate of apoptosis than single treatments. Combined treatment induced caspases 8 and 9 activation to an extent comparable with that of butyrate while γ-thionin did not activate caspases. Additionally, reactive oxygen species generation preceded the onset of apoptosis, and superoxide anion production was higher in cells treated with the combined treatment. Conclusions: The γ-thionin from Habanero chili pepper improved the butyrate cytotoxicity on Caco-2 cells. This effect occurred through apoptosis induction associated with reactive oxygen species production. Therefore, the combination of butyrate with cytotoxic antimicrobial peptides could be an attractive strategy for cancer therapy. Normal 0 false false false EN-US X-NONE X-NONE /* Style Definitions */ table.MsoNormalTable {mso-style-name:"Tabla normal"; mso-tstyle-rowband-size:0; mso-tstyle-colband-size:0; mso-style-noshow:yes; mso-style-priority:99; mso-style-parent:""; mso-padding-alt:0cm 5.4pt 0cm 5.4pt; mso-para-margin-top:0cm; mso-para-margin-right:0cm; mso-para-margin-bottom:8.0pt; mso-para-margin-left:0cm; line-height:107%; mso-pagination:widow-orphan; font-size:11.0pt; font-family:"Calibri",sans-serif; mso-ascii-font-family:Calibri; mso-ascii-theme-font:minor-latin; mso-hansi-font-family:Calibri; mso-hansi-theme-font:minor-latin; mso-bidi-font-family:"Times New Roman"; mso-bidi-theme-font:minor-bidi;}

Desde el discurso gubernamental y la literatura especializada, se ha postulado que el Cártel Jalisco Nueva Generación (CJNG) es una de las redes criminales más poderosas en México, tanto en términos territoriales como en económicos. Con el objetivo de conocer cómo opera en el territorio la economía del Cártel y sus alcances regionales, desarrollamos una metodología mixta. Ésta incluye trabajo de campo etnográfico, aplicación de entrevistas a usuarios y vendedores pertenecientes al CJNG, así como una exhaustiva revisión documental. En este trabajo analizamos la manera en que el CJNG se ha consolidado particularmente en la zona occidental del país, a partir de forjar una economía basada en la producción de cristal de metanfetamina. Dicha economía está sustentada en una reconfiguración del mercado debido al desplazamiento que el propio CJNG ha realizado de otras sustancias como la cocaína, o la marihuana, para dar prioridad a tejer toda una red sustentada en este estimulante.

Genome editing methods based on group II introns (known as targetron technology) have long been used as a gene knockout strategy in a wide range of organisms, in a fashion independent of homologous recombination. Yet, their utility as delivery systems has typically been suboptimal due to the reduced efficiency of insertion when carrying exogenous sequences. We show that this limitation can be tackled and targetrons can be adapted as a general tool in Gram-negative bacteria. To this end, a set of broad-host-range standardized vectors were designed for the conditional expression of the Ll.LtrB intron. After establishing the correct functionality of these plasmids in Escherichia coli and Pseudomonas putida, we created a library of Ll.LtrB variants carrying cargo DNA sequences of different lengths, to benchmark the capacity of intron-mediated delivery in these bacteria. Next, we combined CRISPR/Cas9-facilitated counterselection to increase the chances of finding genomic sites inserted with the thereby engineered introns. With these novel tools, we were able to insert exogenous sequences of up to 600 bp at specific genomic locations in wild-type P. putida KT2440 and its ΔrecA derivative. Finally, we applied this technology to successfully tag P. putida with an orthogonal short sequence barcode that acts as a unique identifier for tracking this microorganism in biotechnological settings. These results show the value of the targetron approach for the unrestricted delivery of small DNA fragments to precise locations in the genomes of Gram-negative bacteria, which will be useful for a suite of genome editing endeavors.

Group II introns were developed some time ago as tools for the construction of knockout mutants in a wide range of organisms, ranging from Gram-positive and Gram-negative bacteria to human cells. Utilizing these introns is advantageous because they are independent of the host’s DNA recombination machinery, they can carry heterologous sequences (and thus be used as vehicles for gene delivery), and they can be easily retargeted for subsequent insertions of additional genes at the user’s will. Alas, the use of this platform has been limited, as insertion efficiencies greatly change depending on the target sites and cannot be predicted a priori. Moreover, the ability of introns to perform their own splicing and integration is compromised when they carry foreign sequences. To overcome these limitations, we merged the group II intron-based TargeTron system with CRISPR/Cas9 counterselection. To this end, we first engineered a new group-II intron by replacing the retrotransposition-activated selectable marker (RAM) with ura3 and retargeting it to a new site in the lacZ gene of E. coli. Then, we showed that directing CRISPR/Cas9 toward the wild-type sequences dramatically increased the chances of finding clones that integrated the retrointron into the target lacZ sequence. The CRISPR-Cas9 counterselection strategy presented herein thus overcomes a major limitation that has prevented the use of group II introns as devices for gene delivery and genome editing at large in a recombination-independent fashion., This work was funded by the SETH Project of the Spanish Ministry of Science RTI 2018-095584−B-C42, MADONNA (H2020-FET-OPEN-RIA-2017-1-766975), BioRoboost (H2020-NMBP-BIO-CSA-2018), and SYNBIO4FLAV (H2020-NMBP/0500). Contracts of the European Union and the S2017/BMD-3691 InGEMICS-CM funded by the Comunidad de Madrid (European Structural and Investment Funds). E.V. was the recipient of a Fellowship from the Education Ministry, Madrid, Spanish Government (FPU15/04315).

Genome editing methods based on Group II introns (known as Targetron technology) have been long used as a gene knock-out strategy in a wide range of organisms in a fashion independent of homologous recombination. Yet, their utility as delivery systems has been typically suboptimal because of their reduced efficiency of insertion when they carry exogenous sequences. We show that this limitation can be tackled and Targetron adapted as a general tool in Gram-negative bacteria. To this end, a set of broad host range standardized vectors were designed for conditional expression of the Ll.LtrB intron. After testing the correct functionality of these plasmids inEscherichia coliandPseudomonas putida, we created a library of Ll.LtrB variants carrying cargo DNA sequences of different lengths to benchmark the capacity of intron-mediated delivery in these bacteria. Next, we combined CRISPR/Cas9-facilitated counterselection to increase the chances of finding genomic sites inserted with the thereby engineered introns. By following this pipeline, we were able to insert exogenous sequences of up to 600 bp at specific genomic locations in wild-typeP. putidaKT2440 and its ΔrecAderivative. Finally, we were able to apply this technology to successfully tag this strain with an orthogonal short sequence (barcode) that acts as a unique identifier for tracking this microorganism in biotechnological settings. The results withP. putidaexemplified the value of the Targetron approach for unrestricted delivery of small DNA fragments to the genomes of Gram-negative bacteria for a suite of genome editing endeavours.

“Full-stack”biotechnology platforms for cell line (re)programming are on the horizon, due mostly to (a) advances in gene synthesis and editing techniques as well as (b) the growing integration with informatics, the internet of things and automation. These emerging platforms will accelerate the production and consumption of biological products. Hence, transparency, traceability and -ultimately-trustworthiness is required -from cradle to grave- for engineered cell lines and their engineering processes. We report here the first version control system for cell engineering that integrates a new cloud-based version control software for cell lines’ digital footprint with molecular barcoding of living samples. We argue that version control for cell engineering marks a significant step towards more open, reproducible, easier to trace and share, and more trustworthy engineering biology.One Sentence SummaryWe demonstrate a transparent and open way of engineering and sharing cell lines.

Microalgae are photosynthetic microorganisms that represent an important source of valuable compounds, including lipids, proteins, carbohydrates, minerals, vitamins, antioxidants, and a variety of pigments. Therefore these microorganisms denote a potential ecological and economical option for applications in various industries. There are more than 30,000 species of microalgae, of which only six are used safely in food. This chapter describes the benefits of microalgae incorporation into food, as well as the main strains used in food processing and various products that are currently in the market. Special emphasis is placed on the species that have been studied for nonfood purposes, like biofuels, bioremediation, pharmaceutics, or cosmetics. Also, the safety of their consumption and nutritional properties are described in order to demonstrate their potential applications as food additives.

Objective: To describe the distribution of pneumococcal serotypes causing infectious diseases in patients with hematological malignancies and solid tumors and their antimicrobial susceptibility before and after introduction of pneumococcal conjugate vaccine (PCV7) in Mexico. Materials and methods: Consecutive pneumococcal isolates from hospitalized patients from the SIREVA-network were serotyped using the Quellung reaction and antimicrobial susceptibility was performed using the broth microdilution method. Results: A total of 175 pneumococcal isolates were recovered, 105 from patients with hematological malignancies and 70 with solid tumors. Serotypes 19A (22.7%), 19F (20.4%), and 35B (17.7%) were the most frequent isolates in the first group and serotypes 3 (27.2%) and 19A (28.6%) in the second group. No decreased susceptibility to beta-lactams or TMP/SMX was observed after introduction of PCV7. Conclusions: An increase in non-vaccine types is observed without significate changes in antimicrobial susceptibility after introduction of PCV7.

Microalgae are eukaryotic microorganisms that photosynthesize to produce biomass and many other useful products. Certain microalgae are used as human food supplements, feed, biofuel, cosmetics, and pharmaceuticals. In order to grow and tap the potentials of microalgae, four distinct modes of growth are recognized: photoautotrophy, heterotrophy, mixotrophy, and photoheterotrophy. There are two basic systems for microalgal culture: open systems where the culture is exposed to the atmosphere and closed systems, commonly called photobioreactors, in which the microalga has little or no exposition to the atmosphere. Once the biotechnological interest of many of these autotrophic microorganisms has been recognized, the next step is the development of bioprocesses that link scientific discoveries to business needs. The design and optimization of culture strategies and bioreactors suitable for cultivating these microorganisms is an essential part of this approach. In this chapter, the main characteristics of microalgae, their classification, the factors that influence their growth, the types of cultivation, the different cultivation systems and the potential applications of these photosynthetic microorganisms are described.

espanolEn esta revision se describen los principales factores que afectan la produccion de pigmentos por Monascus en medio solido, asi como los diferentes residuos utilizados y aquellos potenciales remanentes regionales en Mexico que pueden ser aprovechados en la produccion de pigmentos. Los colorantes producidos por el hongo Monascus han mostrado potencial aplicacion en las industrias alimentaria, cosmetica, farmaceutica y textil. La produccion de pigmentos por Monascus se ha llevado a cabo principalmente en cultivos en estado solido, para lo cual el uso de residuos agroindustriales como fuente de sustrato es una alternativa real que incrementa la factibilidad economica del proceso, aunado a ser una alternativa de bajo impacto ambiental. Los factores de temperatura, pH, humedad relativa y tamano de particula se identificaron como los factores de mayor influencia en la produccion de pigmentos. Los residuos regionales de maiz, arroz y Aloe vera son los sustratos con mayor rendimiento. EnglishIn this review the main factors affecting pigment production by Monascus in solid medium are described, as well as the different agro-industrial waste used and those potential regional wastes in Mexico that can be exploited in the production of pigments. The pigments produced by the fungus Monascus have shown potential application in food, textile, cosmetics and pharmaceutical industries. Pigment production by Monascus has been carried out mainly in solid-state fermentation, for which the use of agro-industrial waste as a source of substrate is a real alternative that increases the economic feasibility of the process with low-impact environmental. The factors of temperature, pH, humidity and particle size were identified as the most influential factors in pigment production and the regional agro-industrial waste of corn, rice and Aloe vera the substrates with higher yield.

Metastatic tumors located in nasal and paranasal sinuses are rare. Metastases to the head and neck region of primary tumors located below the clavicles are uncommon; the vast majority are presented in lymph nodes. We present a clinical case of a 46-year-old woman with facial asymmetry, proptosis, and frontal soreness; the tomography showed a hyperdense lesion in the frontal sinus. The biopsy specimen revealed a clear cell carcinoma; therefore, it was decided to start the search for an unknown primary tumor and subsequent immunohistochemistry complementary tests. A computed tomography scan could identify a tumor of the lower pole of the kidney compatible with clear cell renal carcinoma. The differential diagnosis of clear cell tumors is difficult because they are in almost every malignant neoplasm of salivary glands, thyroid carcinoma, or melanoma metastasis. We are reporting the first case in Mexico of metastasis in paranasal sinus of renal carcinoma.

Las infecciones nosocomiales ocasionadas por cepas de Staphylococcus aureus meticilinorresistentes (SAMR) son un problema de salud importante en todo el mundo. Este microorganismo produce una gran variedad de infecciones incluyendo osteomielitis, endocarditis invasora, artritis séptica y septicemia. La multirresistencia es un factor que influye en la persistencia de los SAMR dentro del ámbito hospitalario. La introducción de técnicas de tipificación molecular dentro de las investigaciones epidemiológicas ha provisto nuevas herramientas para conocer el origen y las vías de diseminación de este microorganismo. Una de las conclusiones importantes que han surgido de este tipo de estudios es que un número pequeño de clonas son las responsables de las infecciones estafilocócicas en todo el mundo.Nosocomial infections due to methicillin-resistant Staphylococcus aureus (MRSA) is an important health problem worldwide. This microorganism causes a variety of clinical infections, including osteomyelitis, invasive endocarditis, septic arthritis and septicemia. Antimicrobial resistance is a factor that influences the persistence of MRSA in the hospital environment. The introduction of molecular typing techniques in epidemiological investigations has provided new tools for identifying the microorganism's origin and routes of dissemination. One of the most important conclusions that have resulted from these types of studies is that a small number of clones are responsible for most of the staphylococcal infections throughout the world.

We determined the seroprevalence of protective antibodies against Hib in Mexican children under the age of five using a standardized enzyme-linked immunosorbent assay. Hib antibodies (≥0.15 μg/ml) were present in 95.34% (±1.14% [seroprevalence ± standard error]) of samples. Fewer children aged 30 to 47 months had protective Hib antibody levels (91.45% ± 2.60%) than children from 12 to 29 and 48 to 59 months (97.3% ± 1.34% and 97.44% ± 1.80%, respectively).

OBJETIVO:Determinar la evolucion de la resistencia a la eritromicina, el cloranfenicol, el trimetoprim-sulfametozaxol (SXT) y la vancomicina de aislamientos invasores de Streptococcus pneumoniae obtenidos de ninos de 10 paises de America Latina y del Caribe en seis anos de vigilancia. METODOS: Se analizaron 8 993 aislamientos de S. pneumoniae recuperados entre 2000 y 2005 de ninos menores de 6 anos con infecciones invasoras, procedentes de Argentina, Brasil, Chile, Colombia, Cuba, Mexico, Paraguay, Republica Dominicana, Uruguay y Venezuela. La sensibilidad a los antibioticos se determino mediante los metodos establecidos y estandarizados en el proyecto SIREVA. La resistencia a multiples antibioticos se definio como la resistencia a tres o mas familias de antibioticos, de los no betalactamicos analizados en este estudio o de los betalactamicos evaluados en un estudio previo en el que 37,8% de estos aislamientos presentaron sensibilidad disminuida a la penicilina. RESULTADOS: Se encontro algun grado de resistencia al SXT y la eritromicina (56,4% y 15,4% de los aislamientos estudiados, respectivamente) y 4,6% presento alta resistencia al cloranfenicol. Todos los aislamientos fueron sensibles a la vancomicina. Se observo la mayor frecuencia de resistencia al SXT en los aislamientos de neumonia y a la eritromicina en los casos de sepsis (61,6% y 25,5%, respectivamente; P < 0,01). La mayor frecuencia de resistencia al SXT se observo en Brasil (71,9%) y a la eritromicina en Mexico (38,2%) y Venezuela (32,9%). Los serotipos 14, 6B, 19F y 23F fueron los que mas frecuentemente se asociaron con la resistencia a los antibioticos estudiados. CONCLUSIONES: Se observo una elevada y creciente frecuencia de aislamientos resistentes al SXT y la eritromicina, y una disminucion en la proporcion de aislamientos resistentes al cloranfenicol. Estas tendencias mostraron diferencias entre los paises estudiados.

Nosocomial infections due to methicillin-resistant Staphylococcus aureus (MRSA) is an important health problem worldwide. This microorganism causes a variety of clinical infections, including osteomyelitis, invasive endocarditis, septic arthritis and septicemia. Antimicrobial resistance is a factor that influences the persistence of MRSA in the hospital environment. The introduction of molecular typing techniques in epidemiological investigations has provided new tools for identifying the microorganism's origin and routes of dissemination. One of the most important conclusions that have resulted from these types of studies is that a small number of clones are responsible for most of the staphylococcal infections throughout the world.

Targetron technology, a gene-editing approach based on the use of mobile group II introns, is particularly useful for bacterial strains deficient in homologous recombination. Specifically, the Ll.LtrB intron from Lactococcus lactis can be used in a wide range of species and can be easily retargeted, that is, modified for integration into any locus of interest. Targetron technology is thus a powerful tool for generating genomic insertions in a broad range of genetic backgrounds, mainly when no other techniques can be efficiently employed. Notably, the approach can be coupled to CRISPR/Cas9 counterselection of wildtype DNA sequences to decrease the population of unmodified cells and ultimately improve Ll.LtrB insertion efficiency. Here, we describe a step-by-step protocol for delivering exogenous sequences into the genome of Gram-negative bacteria by means of targetron technology and CRISPR/Cas9 counterselection using Pseudomonas putida as a model. We describe the retargeting of the Ll.LtrB intron to the locus selected for insertion, the design of specific spacers for eliminating unmutated cells through CRISPR/Cas9 counterselection, and the cloning of exogenous sequences into Ll.LtrB. We also provide a protocol for delivering a specific cargo to the locus of choice once all necessary components of the system are ready. Lastly, we describe a general protocol for curing the engineered strain of all plasmids. CRISPR/Cas9-enhanced Ll.LtrB insertion can be an efficient alternative for overcoming low recombination-based editing efficiency and can be used in numerous bacterial species. © 2022 The Authors. Current Protocols published by Wiley Periodicals LLC. Basic Protocol 1: Retargeting the Ll.LtrB intron to the target locus Support Protocol 1: Preparation of competent E. coli Basic Protocol 2: Design and cloning of CRISPR spacers to counterselect Ll.LtrB insertions Support Protocol 2: Interference assay to check efficiency of selected spacers Basic Protocol 3: Cloning cargos into Ll.LtrB Basic Protocol 4: Ll.LtrB/CRISPR/Cas9-mediated insertion Basic Protocol 5: Curing the engineered strain of plasmids.