A novel strategy to construct multi-strain starter cultures: an insight to evolve from natural to directed fermentation.

Some biotechnological strategies have succeeded in the attempt to imitate natural fermentation, and bioprocesses have been efficiently designed when the product is the result of a unique biological reaction. However, when the process requires more than one biological reaction, few bioprocesses have been successfully designed because the available tools to construct multi-strain starter cultures are not yet well defined. In this work, a novel experimental strategy to construct multi-strain starter cultures with selected native microorganisms from natural fermentation is proposed. The strategy analyses, selects, and defines the number and proportion of each strain that should form a starter culture to be used in directed fermentations. It was applied to evolve natural fermentation to directed fermentation in distilled agave production. The results showed that a starter culture integrated by Kluyveromyces marxianus, Clavispora lusitaniae, and Kluyveromyces marxianus var. drosophilarum in proportions of 35, 32, and 33%, respectively, allows obtaining fermented agave juice containing a 2.1% alcohol yield and a distilled product with a broad profile of aromatic compounds. Hence, the results show, for the first time, a tool that addresses the technical challenge for multi-strain starter culture construction, offering the possibility of preserving the typicity and genuineness of the original traditional product.

A validated strategy to design efficient fermentation-industrial processes: agave spirit production.

An industrial process is profitable when its individual unit operations are efficient and thus, this work shows a guideline for designing efficient fermentation-industrial processes for agave distilled production based on a sequential approach of optimization, beginning in the laboratory and followed by the adjustment of the variable values using the evolutionary operation method for successful process scaling. The results at the laboratory showed that a starter inoculum containing a 5 × 106 cells/mL mixture of Kluyveromyces marxianus, Clavispora lusitaniae, and Kluyveromyces marxianus var. drosophilarum strains in a bioreactor containing agave syrup with 120 g/L fermented sugar, processed at a constant temperature of 33 °C and 1.0 VVM aeration for 1.6 h, led to a fermented product with a 4.18% (v/v) alcohol content after 72 h of processing time. The scale-up process results showed that the best operating conditions at the pilot-plant level were a temperature of 35 °C and aeration at 1.0 VVM for 1.2 h, which led to a fermented product with a 4.22% (v/v) total alcohol content after 72 h of processing time. These represent similar performance values for both production processes, but each one worked with their specific values of process variables, which demonstrates that each level of production had its own specific values for process variables. The volatile compound analysis shows that both distilled products contained a similar profile of volatile components that provide fruity and ethereal aromatic notes pleasant to the palate. Therefore, the process design for agave spirit production at the semi-industrial level was successfully achieved.

Assurance for Endurance? Introducing a Novel Ergonomics Curriculum to Reduce Pain and Enhance Physical Well-Being Among GI Fellows.

Musculoskeletal pain and injuries (MSKPI) are common among gastroenterologists (GI) and GI fellows. Common areas of pain include the back, neck, hands/fingers, shoulders, and elbows. Although the prevalence of career-related pain and injuries among GIs is high, few endoscopists receive training in how to prevent MSKPI. We developed an ergonomics curriculum for our GI fellows that consisted of two modules that were led by physical therapists. Twelve out of 15 GI fellows, and one out of two hepatology fellows, participated in Module 1. Prior to the first module, 77% of participants reported pain in one or more body parts. Of those who reported pain, 100% of the fellows stated that this pain occurred during procedures, and 50% indicated this pain was performance-limiting. After completing Module 1, 100% of fellows reported that this was a valuable topic and 100% of the participants felt that this information would both help them feel and perform better. All fellows stated they had an immediate decrease in physical discomfort after engaging in the exercises that were included in Module 1. Eight fellows participated in Module 2. At the end of this module, 100% of fellows reported that this ergonomics training would likely help them to "physically perform better during procedures" and 100% of fellows indicated a reduction of physical discomfort (pain, aching) immediately after completing these exercises. Preliminary data indicated that this novel curriculum was perceived as valuable by GI fellows and that practicing these exercises reduced pain, particularly in the neck and the lower back.

An experimental strategy validated to design cost-effective culture media based on response surface methodology.

For any fermentation process, the production cost depends on several factors, such as the genetics of the microorganism, the process condition, and the culture medium composition. In this work, a guideline for the design of cost-efficient culture media using a sequential approach based on response surface methodology is described. The procedure was applied to analyze and optimize a culture medium of registered trademark and a base culture medium obtained as a result of the screening analysis from different culture media used to grow the same strain according to the literature. During the experiments, the procedure quantitatively identified an appropriate array of micronutrients to obtain a significant yield and find a minimum number of culture medium ingredients without limiting the process efficiency. The resultant culture medium showed an efficiency that compares favorably with the registered trademark medium at a 95% lower cost as well as reduced the number of ingredients in the base culture medium by 60% without limiting the process efficiency. These results demonstrated that, aside from satisfying the qualitative requirements, an optimum quantity of each constituent is needed to obtain a cost-effective culture medium. Study process variables for optimized culture medium and scaling-up production for the optimal values are desirable.

Mitochondrial ATP is required for the maintenance of membrane integrity in stallion spermatozoa, whereas motility requires both glycolysis and oxidative phosphorylation.

To investigate the hypothesis that oxidative phosphorylation is a major source of ATP to fuel stallion sperm motility, oxidative phosphorylation was suppressed using the mitochondrial uncouplers CCCP and 2,4,-dinitrophenol (DNP) and by inhibiting mitochondrial respiration at complex IV using sodium cyanide or at the level of ATP synthase using oligomycin-A. As mitochondrial dysfunction may also lead to oxidative stress, production of reactive oxygen species was monitored simultaneously. All inhibitors reduced ATP content, but oligomycin-A did so most profoundly. Oligomycin-A and CCCP also significantly reduced mitochondrial membrane potential. Sperm motility almost completely ceased after the inhibition of mitochondrial respiration and both percentage of motile sperm and sperm velocity were reduced in the presence of mitochondrial uncouplers. Inhibition of ATP synthesis resulted in the loss of sperm membrane integrity and increased the production of reactive oxygen species by degenerating sperm. Inhibition of glycolysis by deoxyglucose led to reduced sperm velocities and reduced ATP content, but not to loss of membrane integrity. These results suggest that, in contrast to many other mammalian species, stallion spermatozoa rely primarily on oxidative phosphorylation to generate the energy required for instance to maintain a functional Na+/K+ gradient, which is dependent on an Na+-K+ antiporter ATPase, which relates directly to the noted membrane integrity loss. Under aerobic conditions, however, glycolysis also provides the energy required for sperm motility.

Effect of overnight staining on the quality of flow cytometric sorted stallion sperm: comparison with tradtitional protocols.

Flow cytometry is considered the only reliable method for the separation of X and Y chromosome bearing spermatozoa in equines. The MoFlo SX DP sorter is highly efficient, allowing the production of foals of the desired sex. However, to achieve acceptable pregnancy rates the currently used protocol requires working with fresh semen obtained close to, or at, the sorting facility. An alternative protocol was tested during two consecutive breeding seasons. Fresh stallion semen was cooled for 20 h, during which staining with Hoechst 33342 took place. On the following day, this sample was flow sorted and compared with spermatozoa from the same ejaculate that had been sexed on the previous day. All sperm parameters evaluated remained unchanged when fresh sorted and refrigerated sorted semen were compared. Pre-sorting storage at 5°C did not alter sperm velocities nor kinetics, viability or membrane permeability, production of reactive oxygen species, mitochondrial membrane potential or DNA fragmentation index of the sorted sample. The findings open for the possibility of using semen from stallions housed far from the sorting facilities. Processed and stained sperm could be shipped refrigerated on the previous day, sorted and inseminated on the next day.

Caspase activation, hydrogen peroxide production and Akt dephosphorylation occur during stallion sperm senescence.

To investigate the mechanisms inducing sperm death after ejaculation, stallion ejaculates were incubated in BWW media during 6 h at 37°C. At the beginning of the incubation period and after 1, 2, 4 and 6 h sperm motility and kinematics (CASA), mitochondrial membrane potential and membrane permeability and integrity were evaluated (flow cytometry). Also, at the same time intervals, active caspase 3, hydrogen peroxide, superoxide anion (flow cytometry) and Akt phosphorylation (flow cytometry) were evaluated. Major decreases in sperm function occurred after 6 h of incubation, although after 1 h decrease in the percentages of motile and progressive motile sperm occurred. The decrease observed in sperm functionality after 6 h of incubation was accompanied by a significant increase in the production of hydrogen peroxide and the greatest increase in caspase 3 activity. Additionally, the percentage of phosphorylated Akt reached a minimum after 6 h of incubation. These results provide evidences that sperm death during in vitro incubation is largely an apoptotic phenomena, probably stimulated by endogenous production of hydrogen peroxide and the lack of prosurvival factors maintaining Akt in a phosphorylated status. Disclosing molecular mechanisms leading to sperm death may help to develop new strategies for stallion sperm conservation.

Identification of apoptotic bodies in equine semen.

Apoptosis in the testis is required to ensure an efficient spermatogenesis. However, sometimes, defective germ cells that are marked for elimination during this process escape elimination in the testes, giving rise to ejaculates with increased percentages of abnormal and apoptotic spermatozoa and a high percentage of apoptotic bodies. Apoptosis markers in the ejaculate have been associated with low fertility, either in animals or humans. Therefore, the goal of this study was to investigate whether fresh equine semen contains apoptotic bodies [initially named Merocyanine 540 (M540) bodies] and to study the relationship between the quantity of these bodies and cell concentration, the volume of ejaculate, viability and motility. Moreover, we also studied whether the presence apoptotic bodies in fresh semen was related to the resistance of the stallion spermatozoa to being incubated at 37 °C or being frozen and thawed. Fresh equine semen was stained with fluorescent dyes such as M540 and Annexin-V. Active Caspase 3 was studied in fresh semen through Western blotting and immunofluorescence with a specific antibody. Sperm kinematics was assessed in fresh, incubated and thawed samples using computer-assisted semen analysis, and viability was evaluated with the LIVE/DEAD Sperm Viability Kit. Overall, our results demonstrate for the first time the presence of apoptotic bodies in equine semen. The quantity of apoptotic bodies was highly variable among stallions and was positively correlated with Caspase 3 activity in fresh samples and negatively correlated with the viability and motility of stallion spermatozoa after the cryopreservation process.

The Asociación Mexicana de Gastroenterología consensus on the diagnosis and treatment of acute pancreatitis in children and adolescents.

Acute pancreatitis (AP) and recurrent acute pancreatitis (RAP) are conditions, whose incidence is apparently on the rise. Despite the ever-increasing evidence regarding the management of AP in children and adults, therapeutic actions that could potentially affect having a poor prognosis in those patients, especially in the pediatric population, continue to be carried out. Therefore, the Asociación Mexicana de Gastroenterología convened a group of 24 expert pediatric gastroenterologists from different institutions and areas of Mexico, as well as 2 pediatric nutritionists and 2 specialists in pediatric surgery, to discuss different aspects of the epidemiology, diagnosis, and treatment of AP and RAP in the pediatric population. The aim of this document is to present the consensus results. Different AP topics were addressed by 6 working groups, each of which reviewed the information and formulated statements considered pertinent for each module, on themes involving recommendations and points of debate, concerning diagnostic or therapeutic approaches. All the statements were presented and discussed. They were then evaluated through a Delphi process, with electronic and anonymous voting, to determine the level of agreement on the statements. A total of 29 statements were formulated, all of which reached above 75% agreement in the first round of voting.

Dimethylformamide improves the in vitro characteristics of thawed stallion spermatozoa reducing sublethal damage.

A total of 42 ejaculates were used in the experiment; six ejaculates per stallion, obtained from seven Pure Spanish stallions (PRE), were split and frozen in freezing media with different concentrations and combinations of cryoprotectant (CPA): (i) Cáceres (skim milk based extender) containing 2.5% glycerol (2.5GL), (ii) Cáceres containing 1.5% glycerol and 1.5% dimethylformamide (1.5%GL-1.5%DMFA), (iii) Cáceres extender supplemented with 1.5% glycerol and 2.5% dimethylformamide (1.5%GL-2.5%DMFA) and (iv) Cáceres extender supplemented with 4% dimethylformamide (4%DMFA). After at least 4 weeks of storage in liquid nitrogen (LN), straws were thawed and semen analysed by computer-assisted sperm analysis and flow cytometry (membrane lipid architecture (Merocyanine 540), integrity and sublethal damage (YoPro-1) and mitochondrial membrane potential (JC-1)). After thawing, better results were observed in samples frozen in 4%DMFA or in combinations of 1.5%GL-2.5%DMFA, in fact total motility increased by 16% in the 4%DMFA group compared to 2.5%GL (P < 0.05). Also, there was an increment in the percentage of progressive motile sperm in the 1.5%GL-2.5%DMFA group (9.8% 2.5GL vs 19% in the 1.5%GL-2.5%DMFA group p < 0.05); also, samples frozen in the 4%DMFA group had more intact (YoPro-1 negative) sperm post-thawing, 29.3% in 2.5%GL vs 36.7% in 4%DMFA group (p < 0.05). Membrane lipid architecture was not affected by any of the cryoprotectants tested, while samples frozen in 4%DFMA had a lower percentage of mitochondria with lower membrane potential. It is concluded that DMFA improves the outcome of cryopreservation of stallion spermatozoa mainly reducing sublethal cryodamage.

Cadmium phytoextraction by Helianthus annuus (sunflower), Brassica napus cv Wichita (rapeseed), and Chyrsopogon zizanioides (vetiver).

The use of phytoextraction plant species to accumulate soil metals into harvestable plant parts is a method used for managing soils with high cadmium (Cd). We evaluated three Cd accumulating species recently recommended for such use in cacao farms where Cd removal is needed to maintain markets: Helianthus annuus (sunflower), Brassica napus (rapeseed), and Chyrsopogon zizanioides (vetiver). Plants were grown in two greenhouse pot experiments with different Cd-spiked growth media: (sand plus perlite) and a natural soil. Plant total Cd and Cd uptake in shoot biomass of all species, across both experiments, increased linearly with increasing amounts of added Cd. Rapeseed had the highest plant total Cd and sunflower had the highest Cd uptake in shoot biomass. The highest application of Cd corresponded to the highest plant total Cd and shoot biomass Cd uptake, regardless of species. The bioconcentration factor (BCF) for each species increased in a curvilinear manner with added Cd, with maximum BCF values for plants grown in the sand and perlite matrix at 2.5 mg kg-1 added Cd and in the natural soil at 5.0 mg kg-1 added Cd. We conclude that the Cd uptake (shoot biomass only) capability of the three species examined is greatest for sunflower given its increased uptake with Cd additions, its BCF value > 1, and lack of observed visual Cd toxicity symptoms, fungus and insect damage. Although these species had BCF >1, the potential annual removal of Cd would have been too small to support a meaningful phytoextraction practice.

Anti-Inflammatory Local Effect of Hydroxytyrosol Combined with Pectin-Alginate and Olive Oil on Trinitrobenzene Sulfonic Acid-Induced Colitis in Wistar Rats.

Purpose: Evaluate the efficacy of hydroxytyrosol in the local treatment of inflammatory colitis. Currently, the existing treatments for inflammatory bowel diseases does not cure the disease and it is associated with high rates of side effects and complications. Hydroxytyrosol is a phenyl-ethyl-alcohol derived from the hydrolysis of oleuropein and present in olive oil, previous studies have demonstrated the anti-inflammatory effect of dietary hydroxytyrosol supplement, with no toxicity. Materials & Methods: Colitis has been induced by using Trinitrobenzene Sulfonic Acid at 40 rats. They were divided into four groups randomly: 10 rats without treatment; 10 rats with pectin/alginate mixture; 10 rats treated with pectin/alginate + olive oil; 10 rats treated with pectin/alginate + olive oil + hydroxytyrosol. Animals were sacrificed 10 days after induction of trinitrobenzene sulfonic acid, receiving 5 days of continuous treatment. Samples of the rectal area were studied and observed under a microscope to determine the damage by Hunter scoring modified, assessing inflammatory infiltration, number of intestinal walls involved, damage to the mucosal architecture, and edema. Results: When the rectum was analyzed in a global way, nonsignificant differences were observed; however, when performing an individualized analysis, statistically significant differences in the inflammatory infiltrate are present in the samples, which were evaluated using the ANOVA and Student-T statistics. Conclusions: Local treatment with the natural antioxidant hydroxytyrosol combined with pectin/alginate and olive oil of inflammatory bowel disease has been shown to be effective against inflammatory infiltration of TNBS-induced colitis.

Identification of sperm subpopulations in stallion ejaculates: changes after cryopreservation and comparison with traditional statistics.

In an attempt to improve the information obtained after computer-assisted sperm analysis (CASA), data from five stallions (three ejaculates from each) were analysed before (fresh, extended semen) and after cryopreservation using traditional statistics as well as a cluster analysis. The data matrix consisted of 13 987 observations of individual spermatozoa for fresh, extended semen, and 8305 for frozen-thawed samples. As expected, freezing and thawing resulted in a marked decrease of CASA-derived variables of sperm kinematics. All sperm velocities were significantly lower in frozen-thawed samples than in samples before cooling. Using sperm velocities, six sperm subpopulations were identified in fresh semen (S1-S6). As such, subpopulations S1 and S2 were characterized by low sperm velocities, subpopulations S3 and S4 corresponded to spermatozoa depicting medium speed values, and finally, subpopulations S5 and S6 were those depicting the highest velocities. After freezing and thawing, four sperm subpopulations were identified, listed as nr FT1 to FT4. While subpopulations FT1-FT3 were characterized by low sperm velocities, and thus corresponded speed-wise to those listed as S1-S4 for fresh, extended semen, the one called number FT4 in frozen semen was characterized by high velocities, of the same range as that of the subpopulations S5 and S6 for fresh spermatozoa. The sperm subpopulation structure varied among stallions, but the cluster analysis hereby assayed was able to provide valuable information about the freezability of the samples that the customary statistics did not reveal.

Dietary oleuropein and its new acyl-derivate attenuate murine lupus nephritis through HO-1/Nrf2 activation and suppressing JAK/STAT, NF-κB, MAPK and NLRP3 inflammasome signaling pathways.

Systemic lupus erythemathosus (SLE) is a chronic inflammatory and autoimmune disease which can affect multiple organ systems, without an effective and safe treatment. Olive leaf extracts are of special interest for their therapeutic effects. Oleuropein (OL) is the most abundant constituents of olive leaf extract and possesses many beneficial properties. In this study, we evaluated the effects of dietary OL and its new derivate, peracetylated oleuropein (Per-OL), in a pristane-induced SLE model. Mice received an injection of pristane or saline solution and were fed with experimental diets: enriched with OL and Per-OL. The levels of proinflammatory cytokines and markers were evaluated by enzyme-linked immunosorbent assay. The protein expressions of inducible nitric oxide synthase, microsomal prostaglandin E synthase 1, heme oxygenase (HO-1), nuclear factor E2-related factor 2 (Nrf2), mitogen-activated protein kinases (MAPKs), Janus kinase/signal transducer and activator of transcription (JAK/STAT), nuclear transcription factor-kappa B (NF-κB) and inflammasome nucleotide-binding domain, leucine-rich repeats-containing family, pyrin domain-containing-3 (NLRP3) pathways activation were determined in kidneys by Western blot. OL and Per-OL significantly reduced renal damage and decreased serum matrix metalloproteinase 3 and prostaglandine E2 kidneys levels. Our findings indicate that Nrf2 and HO-1 antioxidant protein expressions were up-regulated in mice fed with OL and Per-OL diets, whereas the activation of JAK/STAT, MAPK, NF-κB and NLRP3 inflammasome pathways was significantly ameliorated. These results suggest that OL and Per-OL supplementation might provide a new alternative approach as a preventive/palliative treatment of nephritis in SLE management.

Poly(ADP-ribose) polymerase-1 protects neurons against apoptosis induced by oxidative stress.

In neurons, DNA is prone to free radical damage, although repair mechanisms preserve the genomic integrity. However, activation of the DNA repair system, poly(ADP-ribose) polymerase (PARP-1), is thought to cause neuronal death through NAD+ depletion and mitochondrial membrane potential (delta psi(m)) depolarization. Here, we show that abolishing PARP-1 activity in primary cortical neurons can either enhance or prevent apoptotic death, depending on the intensity of an oxidative stress. Only in severe oxidative stress does PARP-1 activation result in NAD+ and ATP depletion and neuronal death. To investigate the role of PARP-1 in an endogenous model of oxidative stress, we used an RNA interference (RNAi) strategy to specifically knock down glutamate-cysteine ligase (GCL), the rate-limiting enzyme of glutathione biosynthesis. GCL RNAi spontaneously elicited a mild type of oxidative stress that was enough to stimulate PARP-1 in a Ca2+-calmodulin kinase II-dependent manner. GCL RNAi-mediated PARP-1 activation facilitated DNA repair, although neurons underwent delta psi(m) loss followed by some apoptotic death. PARP-1 inhibition did not prevent delta psi(m) loss, but enhanced the vulnerability of neurons to apoptosis upon GCL silencing. Conversely, mild expression of PARP-1 partially prevented to GCL RNAi-dependent apoptosis. Thus, in the mild progressive damage likely occur in neurodegenerative diseases, PARP-1 activation plays a neuroprotective role that should be taken into account when considering therapeutic strategies.

Detection of "apoptosis-like" changes during the cryopreservation process in equine sperm.

The kinematics of the appearance of apoptotic markers was studied by flow cytometry and immunoblot assays in equine spermatozoa subjected to freezing and thawing. Caspase activity, low mitochondrial membrane potential, and increases in sperm membrane permeability were observed in all of the phases of the cryopreservation procedure. Freezing and thawing caused an increase in membrane permeability and changes in the pattern of caspase activity; decreases in mitochondrial membrane potential were observed after centrifugation and cooling to 4 degrees C and after freezing and thawing. It is proposed that sperm mitochondria may be directly involved in the subtle damage that is present in most spermatozoa surviving freezing and thawing.

Design of a fermentation process for agave fructooligosaccharides production using endo-inulinases produced in situ by Saccharomyces paradoxus.

Saccharomyces paradoxus, a native microorganism of the aguamiel, was used successfully for endoinulinase synthesis for agave fructooligasaccharide (FOS) production. We optimized the fermentation parameters to maximize the enzyme synthesis, and we performed enzyme kinetics studies to achieve agave fructans hydrolysis. The results showed that under constant operating conditions (pH 7.7, 40 °C, 175 rpm of agitation, and 0.005 VVM of aeration) results in the production of an enzymatic extract with 49.57 mg/L. This enzymatic extract, when mixed with an agave fructans solution containing 37.8 g/L, allowed us to obtain products with 18% more FOS content the original concentration. The mass spectrum plot shows that the hydrolyzed product contains FOS with a degree of polymerization from 5 to 9 hexose units. These results are promising because they show FOS production from agave and confirm that importance of using native strains in the design of directed fermentation processes.

Lack of evidence for Mycoplasma spp. in bulk tank milk of herds located in mid-western Colombia / Falta de evidencia para Mycoplasma spp. en leche de tanques de enfriamiento de hatos del centro-occidente colombiano

ABSTRACT Mycoplasma spp. is reported as a highly contagious mastitis-causing bacteria in dairy cattle, without successful or low response to most common antibiotic treatments due to the lack of cell wall. In Colombia it has been reported in the Central Andean region during 2014. The aim was to estimate the prevalence of Mycoplasma spp. in bulk tank milk using microbiological and molecular diagnosis. A random longitudinal study enrolling 220 commercial dairy farms located in four provinces of the mid-western region of Colombia from four pasteurizer companies was performed. Bulk tank milk samples were collected once monthly for three months period for determining somatic cell count (SCC) and microbiological and molecular diagnosis of Mycoplasma spp. cultures were done without pre-enrichment procedures directly in mycoplasma agar with cefoperazone to inhibit growth of opportunistic microorganisms, plates were incubated under 37° C and atmosphere of 10% CO2 and inspected during a 10d period. Molecular analysis was done by a multiplex PCR using specific primers targeting the 16S-23S rARN gene of Mycoplasma spp. and from non-pathogenic bacteria occasionally found in milk. LnSCC average of included dairy farms was 6.19 x103 cells/mL, Mycoplasma spp. was not isolated during microbiological cultures, and no DNA belonging to the species was detected by PCR in the 220 bulk tanks milk, with an estimated prevalence lower than 2.3%. This finding shows that there is not microbiological or molecular evidence that demonstrates the presence of the pathogen in the milk from the mid-western region of Colombia at herd level.

RESUMEN Mycoplasma spp. está descrito como una bacteria causante de mastitis altamente contagiosa en ganado lechero, sin o con baja respuesta a tratamientos antibióticos convencionales debido a que carece de pared celular. En Colombia ha sido reportado en la región Andina Central durante 2014. El objetivo fue estimar la prevalencia de Mycoplasma spp. en leche de tanques de enfriamiento empleando diagnósticos microbiológicos y moleculares. Se realizó un estudio aleatorio longitudinal que incluyó 220 lecherías comerciales en cuatro departamentos del centro-occidente colombiano acopiadas por cuatro compañías pasteurizadoras. Se recolectaron muestras de leche del tanque de enfriamiento mensualmente durante tres meses para determinar el recuento de células somáticas (SCC) y el diagnóstico microbiològico y molecular de Mycoplasma spp. Los cultivos se realizaron sin procedimientos de preenriquecimiento directamente en agar micoplasma con cefoperazona para inhibir crecimiento de microorganismos oportunistas, los agares se incubaron a 37° C con una atmosfera del 10% CO2 e inspeccionados durante 10d. Los análisis moleculares se realizaron por PCR multiplex usando cebadores específicos para los genes 16S-23S rRNA del Mycoplasma spp. y de algunas bacterias oportunistas ocasionales en la leche. El promedio del LnSCC fue de 6.19 x103 células/ mL, Mycoplasma spp. no fue aislado de los cultivos microbiológicos y no se encontró ADN de a esta especie mediante PCR en los 220 tanques de leche. Lo anterior indica una prevalencia estimada menor a 2,3%. Se concluye que no existe evidencia micro-biológica ni molecular para demostrar la presencia del patógeno en la leche de la región centro-occidente colombiana a nivel de hato.

Interrelationships between astrocyte function, oxidative stress and antioxidant status within the central nervous system.

Astrocytes have, until recently, been thought of as the passive supporting elements of the central nervous system. However, recent developments suggest that these cells actually play a crucial and vital role in the overall physiology of the brain. Astrocytes selectively express a host of cell membrane and nuclear receptors that are responsive to various neuroactive compounds. In addition, the cell membrane has a number of important transporters for these compounds. Direct evidence for the selective co-expression of neurotransmitters, transporters on both neurons and astrocytes, provides additional evidence for metabolic compartmentation within the central nervous system. Oxidative stress as defined by the excessive production of free radicals can alter dramatically the function of the cell. The free radical nitric oxide has attracted a considerable amount of attention recently, due to its role as a physiological second messenger but also because of its neurotoxic potential when produced in excess. We provide, therefore, an in-depth discussion on how this free radical and its metabolites affect the intra and intercellular physiology of the astrocyte(s) and surrounding neurons. Finally, we look at the ways in which astrocytes can counteract the production of free radicals in general by using their antioxidant pathways. The glutathione antioxidant system will be the focus of attention, since astrocytes have an enormous capacity for, and efficiency built into this particular system.