Ontogenetic variation in metabolic rate-temperature relationships during larval development.

Predictive models of ectotherm responses to environmental change often rely on thermal performance data from the literature. For insects, the majority of these data focus on two traits, development rate and thermal tolerance limits. Data are also often limited to the adult stage. Consequently, predictions based on these data generally ignore other measures of thermal performance and do not account for the role of ontogenetic variation in thermal physiology across the complex insect life cycle. Theoretical syntheses for predicting metabolic rate also make similar assumptions despite the strong influence of body size as well as temperature on metabolic rate. The aim of this study was to understand the influence of ontogenetic variation on ectotherm physiology and its potential impact on predictive modeling. To do this we examined metabolic rate-temperature (MR-T) relationships across the larval stage in a laboratory strain of the Spongy moth (Lymantria dispar dispar). Routine metabolic rates (RMR) of larvae were assayed at eight temperatures across the first five instars of the larval stage. After accounting for differences in body mass, larval instars showed significant variation in MR-T. Both the temperature sensitivity and allometry of RMR increased and peaked during the third instar, then declined in the fourth and fifth instar. Generally, these results show that insect thermal physiology does not remain static during larval ontogeny and suggest that ontogenetic variation should be an important consideration when modeling thermal performance.

Microencapsulation of Probiotics with Soy Protein Isolate and Alginate for the Poultry Industry.

Many probiotic products, with properly selected microorganisms, may not be effective for the intended purpose due to the low tolerance of microorganisms to gastrointestinal digestion. The microencapsulation seems to be one of the most promising techniques to protect probiotics against adverse environmental conditions. Therefore, the aim of this work was the design of soy protein isolate-alginate microcapsules for the encapsulation of probiotics for the poultry industry by the water-in-oil emulsion technique. To this end, the strain Ligilactobacillus salivarius CRL2217, with the ability to bind wheat germ agglutinin (WGA) on its surface and protect intestinal epithelial cells from the cytotoxicity of the glycoprotein, was used as model microorganism. Several parameters were varied in order to find the better conditions for microencapsulation: oil source and nature, SPI and sodium alginate concentration, stirring equipment and time for emulsion formation, CaCl2 concentration, and absence or presence of stirring after the addition of the CaCl2 solution. The survival of entrapped cells to a simulated gastric digestion and their survival and release during simulated intestinal digestion were also investigated. The obtained particles effectively protected L. salivarius CRL2217 from the proteolytic activity and low pH present in the gastric environment. Besides, their content was released in contact with a simulated intestinal juice, as viable counts and binding of WGA after a simulated intestinal digestion revealed. This work paves the way for the design of probiotic supplements for poultry including gastrointestinal digestion-susceptible bacteria.

Rapid Identification of new isolates of Acidipropionibacterium acidipropionici by fluorescence in situ hybridization (FISH)

Abstract Acidipropionibacterium acidipropionici is widely used for many applications, suchas propionic acid production, cereal silage, and also as probiotic. Due to this plethora of appli-cations, new isolates of A. acidipropionici with improved features are being searched for. Thesenew isolates must be accurately identified, however, most approaches become expensive andtime-consuming when the number of isolates is high. On the contrary, fluorescence in situhybridization allows the affordable, reliable, and rapid identification of microorganisms in purecultures and environmental and medical samples. Therefore, the aim of this work was to applya fluorescent in situ hybridization probe for the reliable identification of new A. acidipropioniciisolates. To this end, probe Pap446, specific for A. acidipropionici, was validated by hybridiza-tion assays with strains of this species from different origins, other species of the same genusor family, and unrelated genera. Eight isolates with propionibacterium characteristics wereobtained from milk and feces of cows. Probe Pap446, hybridized only with isolates III and VI.The identity of these isolates was further confirmed by PCR using group and species-specificprimers for propionibacteria and 16S rDNA sequencing.

Resumen Acidipropionibacterium acidipropionici es ampliamente usada para diversas aplicaciones, como producción de ácido propiónico, ensilado de cereales y probiótico. Debido a esta variedad de aplicaciones, continuamente se buscan nuevos aislamientos de A. acidipropionici con características nuevas. Estos nuevos aislamientos deben ser identificados correctamente, pero la mayoría de las técnicas disponibles resultan costosas e insumen mucho tiempo cuando el número de aislamientos es elevado. Por el contrario, la hibridación fluorescente in situ permite una identificación barata, confiable y rápida de microorganismos en cultivos puros y en muestras ambientales y médicas. Por lo tanto, el objetivo de este trabajo fue la aplicación de una sonda oligonucleotídica en un protocolo de hibridación fluorescente in situ para la identificación confiable de nuevos aislamientos de A. acidipropionici. Con este fin, se validó la sonda Pap446, específica de A. acidipropionici mediante ensayos de hibridación con cepas de esta especie de diferente origen, otras especies del mismo género o familia, y géneros no relacionados. Se obtuvieron ocho aislamientos con características de propionibacterias a partir de leche y heces de vacas. La sonda Pap446, hibridó únicamente con los aislamientos III y VI. La identidad de estos aislamientos fue confirmada a través de PCR con cebadores específicos para propionibacterias y para A. acidipropionici, y mediante secuenciación del ADNr 16S.

Growth studies of dominant lactic acid bacteria in orange juice and selection of strains to ferment citric fruit juices with probiotic potential.

The study aimed to evaluate the ability of dominant lactic acid bacteria (LAB) in orange juice to growth on N-depleted MRS medium supplemented or not with cysteine (mMRS), then to select the most nutritionally promising strains for growth assays in the food matrix and evaluation of beneficial attributes for fruit juice fermentation. Levilactobacillus brevis and Lactiplantibacillus plantarum were dominant species among the total of 103 LAB isolates as confirmed by multiplex PCR and/or 16 s rDNA sequence analysis. Based on growing lower than 20% and higher than 70% in mMRS (1.0 g/l meat extract, without peptone and yeast extract) with and without cysteine requirement, one L. brevis (JNB23) and two L. plantarum (JNB21 and JNB25) were selected. These bacteria and the L. plantarum strains N4 and N8 (previously isolated from oranges peel) when inoculated in orange juice grew up to 1.0 log cfu/ml for 24 h incubation at 30 °C and mainly produced lactic acid, with strains JNB25 and JNB23 reaching the highest and lowest cell densities in agreement with their nutritional exigency. In addition, all L. plantarum strains exhibited antagonistic activity against the majority of tested bacterial pathogens (in opposition to L. brevis), ability to grow or survive to pH 3.0 for 3 h, to grow with 0.5% sodium taurocholate, and a decrease after simulated gastrointestinal digestion assay which did not exceed 1.0 or 2.0 log units, depending on the strain. Thus, autochthonous L. plantarum strains with ability for overcoming nutritional limitations and beneficial attributes are promising candidates for further investigations as novel probiotic and/or preservative starters to ferment citric fruit juices.

Rapid identification of new isolates of Acidipropionibacterium acidipropionici by fluorescence in situ hybridization (FISH).

Acidipropionibacterium acidipropionici is widely used for many applications, such as propionic acid production, cereal silage, and also as probiotic. Due to this plethora of applications, new isolates of A. acidipropionici with improved features are being searched for. These new isolates must be accurately identified, however, most approaches become expensive and time-consuming when the number of isolates is high. On the contrary, fluorescence in situ hybridization allows the affordable, reliable, and rapid identification of microorganisms in pure cultures and environmental and medical samples. Therefore, the aim of this work was to apply a fluorescent in situ hybridization probe for the reliable identification of new A. acidipropionici isolates. To this end, probe Pap446, specific for A. acidipropionici, was validated by hybridization assays with strains of this species from different origins, other species of the same genus or family, and unrelated genera. Eight isolates with propionibacterium characteristics were obtained from milk and feces of cows. Probe Pap446, hybridized only with isolates III and VI. The identity of these isolates was further confirmed by PCR using group and species-specific primers for propionibacteria and 16S rDNA sequencing.

Seasonal changes in mitochondrial bioenergetics and physiological performance of the bluegill sunfish, Lepomis macrochirus, from a shallow, Midwest river.

As global temperature shifts due to anthropogenic impacts, seasonal temperatures in shallow aquatic ecosystems are expected to increase. Previous studies on freshwater fishes that experience significant temperature changes during the annual seasons found pronounced physiological restructuring not observed in animals inhabiting more thermally stable environments. Studies evaluating mitochondrial bioenergetics in fish are often performed on animals acclimated to constant temperatures in the laboratory. However, natural habitats are much more complex. Fishes may experience substantial daily and seasonal variation in temperature, energy requirements and resource availability, which are impossible to emulate on acclimation studies. Our study explores the effects of these more complex natural environments on whole-organism thermal tolerance and mitochondrial bioenergetics in bluegill sunfish (Lepomis macrochirus), a native fish to the temperate zone of North America. Compensatory mechanisms and variations in physiological thresholds were observed in specimens acclimatized to the fall season compared to specimens acclimatized to spring and summer seasons. Somatic indices, such as relative weights and hepatosomatic indices, showed significant differences across seasons and critical thermal maxima significantly decreased in the cold acclimatized specimens. Liver mitochondria from L. macrochirus also showed significantly higher uncoupled proton conductance, cytochrome c oxidase (COX) activity, and reduced respiratory control ratios in individuals sampled in the colder season. These findings suggest that mechanisms regulating proton conductance and COX activity modulate mitochondrial function across seasons to sustain physiological fitness in ectotherms inhabiting shallow, inland aquatic habitats.

Protection of the intestinal epithelium of poultry against deleterious effects of dietary lectins by a multi-strain bacterial supplement.

The intake of antinutritional factors produce impairment on the intestinal digestive function, impeding the efficient use of nutrients. Probiotics could be useful in poultry breeding to prevent negative effects of antinutritional factors, like the dietary lectins soybean agglutinin (SBA) and wheat germ agglutinin (WGA). Therefore, this investigation aimed to verify that SBA and wheat, which contains WGA, exert harmful effects on the intestinal mucosa and the digestive system of young poultry, and determine if the administration of probiotics able to capture lectins could counteract their effects. The trials performed demonstrated that a mixture of Bifidobacterium infantis CRL 1395, Enterococcus faecium LET 301, Lactobacillus salivarius LET 201, L. reuteri LET 210, and Propionibacterium acidipropionici LET 103, strains with ex vivo ability to interfere with the interaction of lectins and epithelial cells, has no negative effect on young chickens health. Middle levels of SBA, as well as wheat as a source of WGA, resulted in lower activities of intestinal and brush border enzymes and alterations in the integrity and morphological parameters of the chicks jejunal mucosa. The bacteria blend increased the activity of several digestive enzymes and the intestinal maturation marker alkaline phosphatase in birds fed with a conventional diet. Besides, it partially countered the deleterious effects of increased content of SBA, as well as the negative effect of a dietary source of WGA, on digestive enzymes activity and intestinal mucosa integrity. The results highlight the capability of multifunctional bacterial mixtures to protect the digestive system of avian against residual dietary lectins.

Prevalence of Cyclomodulin-Positive E. coli and Klebsiella spp. Strains in Mexican Patients with Colon Diseases and Antimicrobial Resistance.

Colon diseases, such as colorectal cancer (CRC), are multifactor diseases that affect more than one million people per year; recently, the microbiota has been associated with an etiologic factor, specifically bacterial cyclomodulin positivity (CM+). Unfortunately, there are no studies from Mexico that detail the presence of bacterial CM+ in patients with colon diseases. We therefore performed a comprehensive study to investigate the associations and prevalence of cyclomodulin-positive Diarrheagenic E. coli (DEC), non-DEC, and Klebsiella spp. strains isolated from Mexican subjects with colon diseases. In this work, we analyzed 43 biopsies, 87 different bacteria were isolated, and E. coli was the most frequently noted, followed by Klebsiella spp., and Enterococcus spp. E. coli, non-DEC, and EPEC belonging to phylogroup B2 were the most prevalent. More than 80% of E. coli and Klebsiella were CM+. pks, cdt, cnf, and cif were identified. cdt was associated with non-DEC, cif and its combinations with EPEC, as well as cdt and psk with Klebsiella. Lastly, all the CM+ bacteria were resistant to at least one antibiotic (34% were MDR, and 48% XDR). In conclusion, the high prevalence of bacterial CM+ in colon disease patients suggests that these bacteria play an important role in the genesis of these diseases.

Long-Term Effectiveness of Polymerized-Type I Collagen Intra-Articular Injections in Patients with Symptomatic Knee Osteoarthritis: Clinical and Radiographic Evaluation in a Cohort Study.

OBJECTIVE: Polymerized-type I collagen (polymerized-collagen) is a downregulator of inflammation and a tissue regenerator. The aim was to evaluate the effect of intra-articular injections (IAIs) of polymerized-collagen among patients with symptomatic knee osteoarthritis (OA) in delaying or preventing joint replacement surgery. Patients and Methods. This was a cohort study of 309 patients with knee OA. Patients with mild-to-moderate disease were treated weekly with IAIs of 2 mL of polymerized-collagen for six weeks (n = 309). Follow-up was for 6-60 months. The primary endpoints included the following determinations: (1) therapeutic effect; (2) survival from total knee replacement surgery (TKR); (3) Western Ontario and McMaster University Osteoarthritis Index (WOMAC) and pain (visual analogue scale, VAS). Clinical improvement was defined as a decrease in pain exceeding 20 mm on the VAS and the achievement of at least 20% improvement from baseline with respect to the WOMAC score. Radiographic analysis was performed at baseline and 60 months. The joint space width in the medial, lateral, and patellofemoral compartments was calculated. RESULTS: Patients who received IAIs of polymerized-collagen had a statistically significant improvement in the primary criteria (p < 0.05). Kaplan-Meier survival analysis of the therapeutic effect demonstrated 98.8% survival at 60 months with TKR as the endpoint. There was no significant reduction in joint space in any compartment based on the analyzed radiographs. No serious adverse events were recorded. CONCLUSION: Polymerized-collagen increased the time to TKR by at least 60 months, modifying the disease course, improving functional disability, and decreasing pain.

The impact of urgent intervention on the neurologic recovery in patients with thoracolumbar fractures.

BACKGROUND: We carried out an observational longitudinal retrospective study between 2000 and 2009 in 28 patients who underwent surgery for unstable vertebral fractures with neurologic deficits. METHODS: For the statistical analysis, we used the Chi2-test to compare proportions in independent groups and the exact Fisher test and the Wilcoxon test for repeated measures, and we compared the mean values using the Mann-Whitney U test at a significance level of P<0.05. Timing to surgical intervention (urgent ≤8 vs. >8 h), and neurologic status using the American Spinal Injury Association (ASIA) Impairment Scale at baseline and at the end of follow-up were assessed. We tested the ASIA score improvement at the end of follow-up using multiple regression analysis, adjusted by variables such as ISS, timing of intervention, location, approach and type of fracture. RESULTS: Twenty-eight patients were included in the analysis. Of the total, 11 (39.2%) underwent surgery urgently (≤8 h) and 17 (60.8%) in >8 h. The mean difference in the neurologic improvement in all patients was 0.97 (95% CI, 0.51-1.42) and was statistically significant (P=0.001). The mean difference in the neurologic improvement in patients with incomplete lesions was 1.59 (95% CI, 1.01-2.17, P=0.001). In these patients, the mean improvement for those intervened in less than 8 h was 1.73 compared to those operated on after more than 8 h (mean improve 0.47) with a difference statistically significant (P=0.007). CONCLUSIONS: Urgent surgery was associated with neurologic improvement in patients with spinal cord injury (SCI). This improvement was mainly observed in patients with an incomplete lesion.

Geographic Variation in Larval Metabolic Rate Between Northern and Southern Populations of the Invasive Gypsy Moth.

Thermal regimes can diverge considerably across the geographic range of a species, and accordingly, populations can vary in their response to changing environmental conditions. Both local adaptation and acclimatization are important mechanisms for ectotherms to maintain homeostasis as environments become thermally stressful, which organisms often experience at their geographic range limits. The spatial spread of the gypsy moth (Lymantria dispar L.) (Lepidoptera: Erebidae) after introduction to North America provides an exemplary system for studying population variation in physiological traits given the gradient of climates encompassed by its current invasive range. This study quantifies differences in resting metabolic rate (RMR) across temperature for four populations of gypsy moth, two from the northern and two from southern regions of their introduced range in North America. Gypsy moth larvae were reared at high and low thermal regimes, and then metabolic activity was monitored at four temperatures using stop-flow respirometry to test for an acclimation response. For all populations, there was a significant increase in RMR as respirometry test temperature increased. Contrary to our expectations, we did not find evidence for metabolic adaptation to colder environments based on our comparisons between northern and southern populations. We also found no evidence for an acclimation response of RMR to rearing temperature for three of the four pairwise comparisons examined. Understanding the thermal sensitivity of metabolic rate in gypsy moth, and understanding the potential for changes in physiology at range extremes, is critical for estimating continued spatial spread of this invasive species both under current and potential future climatic constraints.

Reduced Mitochondrial Efficiency Explains Mismatched Growth and Metabolic Rate at Supraoptimal Temperatures.

The relationship between whole-organism growth and metabolism is generally assumed to be positive and causative; higher metabolic rates support higher growth rates. In Manduca sexta, existing data demonstrate a deviation from this simple prediction: at supraoptimal temperatures for larval growth, metabolic rate keeps increasing while growth rate is decreasing. This mismatch presumably reflects the rising "cost of maintenance" with temperature. Precisely what constitutes this cost is not clear, but we suspect the efficiency with which mitochondria harness oxygen and organic substrates into cellular energy (ATP) is key. We tested this by integrating existing data on M. sexta growth and metabolism with new data on mitochondrial bioenergetics across the temperature range 14°-42°C. Across this range, our measure of mitochondrial efficiency closely paralleled larval growth rates. At supraoptimal temperatures for growth, mitochondrial efficiency was reduced, which could explain the mismatch between growth and metabolism observed at the whole-organism level.

Cytotoxic damage of soybean agglutinin on intestinal epithelial cells of broiler chicks: in vitro protection by Bifidobacterium infantis CRL1395.

Plant lectins, which are proteins/glycoproteins present in a wide range of vegetables, fruits, cereals and beans, are resistant to digestive enzymes and food cooking temperatures. They bind reversibly to specific glycosidic residues expressed on the membrane of intestinal epithelial cells (IEC) and cause anti-nutritional effects in humans and animals. Soybean lectin (SBA) has been detected in poultry diets, and its ability to bind to the intestinal epithelium has been reported. The development of new methods for removing SBA from feeds or to prevent interaction with the intestinal mucosa is of interest. In this study, the in vitro cytotoxicity of SBA on IEC of chicks was demonstrated for the first time. The LD50, assessed after 2 h exposure of IEC to SBA, was 6.13 µg mL(-1) The ability of Bifidobacterium infantis CRL1395 to bind SBA on the bacterial envelope was confirmed, and prevention of IEC cytotoxicity by lectin removal was demonstrated. Safety of B. infantis CRL1395, resistance to gastrointestinal stress and adhesion were also determined. It was concluded that the early administration of B. infantis CRL1395 to chicks would effectively reduce the toxicity of SBA. Besides, it would favour the colonization of the gut with a beneficial microbiota.

Budget-limited thermal biology: Design, construction and performance of a large, walk-in style temperature-controlled chamber.

We describe a partial redesign of the conventional air-conditioning system and apply it to the construction of a relatively large (1.87m(3) air mass), walk-in style temperature-controlled chamber (TCC) using parts easily obtained in most countries. We conducted several tests to demonstrate the performance of the TCC. Across the physiologically relevant range of 5-37°C, the TCC took 26.5-50.0min to reach the desired set point temperature. Once at set point, temperature inside the chamber was controlled with an accuracy of ±1.0°C. User-entry effects on deviations from and return times to set point temperature were minimal. Overall, performance of the TCC was sufficient to make precise physiological measurements of insect metabolic rate while controlling assay temperature. Major advantages of the TCC include its simplicity, flexibility, and low cost.

Physiological performance of warm-adapted marine ectotherms: Thermal limits of mitochondrial energy transduction efficiency.

Thermal regimes in aquatic systems have profound implications for the physiology of ectotherms. In particular, the effect of elevated temperatures on mitochondrial energy transduction in tropical and subtropical teleosts may have profound consequences on organismal performance and population viability. Upper and lower whole-organism critical temperatures for teleosts suggest that subtropical and tropical species are not susceptible to the warming trends associated with climate change, but sub-lethal effects on energy transduction efficiency and population dynamics remain unclear. The goal of the present study was to compare the thermal sensitivity of processes associated with mitochondrial energy transduction in liver mitochondria from the striped mojarra (Eugerres plumieri), the whitemouth croaker (Micropogonias furnieri) and the palometa (Trachinotus goodei), to those of the subtropical pinfish (Lagodon rhomboides) and the blue runner (Caranx crysos). Mitochondrial function was assayed at temperatures ranging from 10 to 40°C and results obtained for both tropical and subtropical species showed a reduction in the energy transduction efficiency of the oxidative phosphorylation (OXPHOS) system in most species studied at temperatures below whole-organism critical temperature thresholds. Our results show a loss of coupling between O2 consumption and ATP production before the onset of the critical thermal maxima, indicating that elevated temperature may severely impact the yield of ATP production per carbon unit oxidized. As warming trends are projected for tropical regions, increasing water temperatures in tropical estuaries and coral reefs could impact long-term growth and reproductive performance in tropical organisms, which are already close to their upper thermal limit.

Acute toxicity assessment of N,N-diethyl-m-toluamide (DEET) on the oxygen flux of the dinoflagellate Gymnodinium instriatum.

Despite the ubiquitous occurrence of N,N-diethyl-m-toluamide (DEET) in aquatic systems, assessments evaluating the toxicity of DEET on phytoplankton species are summed to a single study on a unicellular green alga. In particular, the toxicological effects of DEET in dinoflagellates are unknown. In this study, we employed the mixotrophic dinoflagellate Gymnodinium instriatum as a study system to evaluate acute effects of DEET on the oxygen flux of laboratory cultures. This study reports an inhibitory reaction model of DEET described by the equation y = 4.99x(0.54), where y represents the percent inhibition of oxygen flux and x represents DEET concentration in mg L(−1) (r(2) = 0.98). Based on this model, the effective concentration of DEET needed to reduce O2 flux by 50% (EC50) for this species was found to be at 72.9 mg L(−1). The reported EC50 is more than five times lower than the EC50 reported previously for the unicellular green algae Chlorella protothecoides. This study raises the question of the potential toxicological effects of DEET in dinoflagellates, in particular those populations inhabiting systems characterized by low water circulation such as enclosed bays and lagoons.

Selection of indigenous lactic acid bacteria to reinforce the intestinal microbiota of newly hatched chicken: relevance of in vitro and ex vivo methods for strains characterization.

Based on the natural benefits of the indigenous microbiota, lactic acid bacteria (LAB) from poultry origin were isolated from hens and broilers intestine, and their probiotic potential was further studied. The tolerance to digestion, adhesion, capture of a mannose-binding lectin, absence of virulent factors and antibiotic resistances were studied. Different in vitro and ex vivo assays were performed to select tolerant and adherent strains because standardized protocols have not been defined. Fourteen strains highly tolerant to gastrointestinal digestion were genetically identified. Hydrophobic surfaces were not required for the bacterial adhesion and only nine strains adhered ex vivo to the intestinal mucosa. Three strains captured a lectin of the same specificity of Type-1 fimbriae. Virulence factors were absent but some strains evidenced multiple antibiotic resistances. These results provide bases for a future standardization of methods for the selection of probiotic strains intended to reinforce the microbiota of newly hatched chickens.

Physiological and functional characteristics of Propionibacterium strains of the poultry microbiota and relevance for the development of probiotic products.

The prevention and control of pathogens colonization through probiotics administration in poultry feeding is of increasing interest. The genus Propionibacterium is an attractive candidate for the development of probiotic cultures as they produce short chain fatty acids (SCFA) by carbohydrates fermentation. The presence of strains of this genus in hens of conventional production systems and backyard hens was investigated. Propionibacteria were isolated from the intestine and identified by physiological and biochemical tests. PCR amplification of the 16S rRNA gene of the isolates was performed and products were compared with sequences from databases. The presence of the genus Propionibacterium was demonstrated in 26% of hens and Propionibacterium acidipropionici and Propionibacterium avidum were the identified species. A comparative study of their physiological and functional characteristics was performed. P. acidipropionici strains were the most resistant to in vitro gastrointestinal digestion, but the adhesion to intestinal tissue was strain dependent. Some differences were found between both species with respect to their growth and SCFA production in an in vitro cecal water model, but all the strains were metabolically active. The production of SCFA in cecal slurries inoculated with the strain P. acidipropionici LET 105 was 30% higher than in non-inoculated samples. SCFA concentrations obtained were high enough to inhibit Salmonella enterica serovar Enteritidis when assayed in a cecal water model. P. acidipropionici LET 105 was also able to compete with Salmonella for adhesion sites on the intestinal mucosa in ex vivo assays. Results contribute to the knowledge of the species diversity of the genus Propionibacterium in the intestine of poultry and provide evidence of their potential for probiotics products development.

Evaluación de la morbilidad y mortalidad de la colecistectomía laparoscópica con técnica de tres puertos en el Hospital Regional de Alta Especialidad de Oaxaca / Assessment of morbidity and mortality due to laparoscopic cholecystectomy with three ports technique at the Regional High Specialty Hospital of Oaxaca

Objetivo: Evaluar la morbilidad y mortalidad de la colecistectomía laparoscópica (CL) en el Hospital regional de Alta Especialidad de Oaxaca (HRAEO), y compararlo con lo reportado en la literatura. Sede: Hospital Regional de Alta Especialidad de Oaxaca. Diseño: Estudio retrospectivo, transversal, descriptivo y comparativo. Análisis estadístico: Análisis estadístico bivariado con medidas de tendencia central y χ². Pacientes y métodos: Pacientes sometidos a CL de enero del 2010 a diciembre del 2011. Se evaluó el tiempo quirúrgico, sangrado transoperatorio, porcentaje de conversión, días de estancia intrahospitalaria, complicaciones transoperatorias y postoperatorias. Resultados: Se incluyeron 386 pacientes, 68 (17.6%) hombres, edad de 43.51 ± 16.8 años. Se identificó un paciente con lesión de vía biliar (0.3%), un paciente con fuga biliar (0.3%), dos pacientes con bilomas residuales (0.5%) y un porcentaje de conversión del 2.8% (11 casos). Un paciente falleció en el grupo de tres puertos (mortalidad global = 0.3%) por colangitis aguda. El porcentaje de complicaciones fue del 4.9%; en 355 (92%) pacientes se usaron tres puertos y cuatro puertos en 31 (8%). Al comparar ambas técnicas, la estancia intrahospitalaria fue menor en el grupo de tres puertos, 1.92 ± 1.22 días versus 2.87 ± 2.84 días (p = 0.0001), el sangrado fue menor 55.23 ± 123.48 ml versus 114.52 ± 193.04 ml (p = 0.0001), así como el tiempo operatorio 71.05 ± 41.87 min versus 110.26 ± 61.25 min (p = 0.0001). Sin diferencia en la morbilidad (frecuencia de fístulas y lesiones de vía biliar). Conclusiones: La morbilidad y mortalidad de la CL en el HRAEO es similar a la reportada en la literatura mundial. El uso de tres puertos sobre cuatro puertos demostró un menor tiempo quirúrgico, estancia hospitalaria y sangrado.

Objective: To assess morbidity and mortality of laparoscopic cholecystectomy (LC) at the Regional Hospital of High Specialty (HRAEO, for its initials in Spanish). Setting: Regional Hospital of High Specialty of Oaxaca (third level health care center). Design: Retrospective, cross-sectional, descriptive, comparative study. Statistical analysis: Bivariate statistical analysis with central tendency measures and chi square. Patients and methods: Patients subjected to LC from January 2010 to December 2011. We assessed surgical time, transoperative bleeding, percentage of conversion, days of in-hospital stay, transoperative and postoperative complications. Results: A total of 386 patients were included, 68 (17.6%) were men of 43.51 ± 16.8 years of age. We identified: one patient with biliary tract lesion (0.3%), one patient with biliary leakage (0.3%), two patients with residual bilomas (0.5%), and a conversion percentage of 2.8% (11 cases). One patient died in the group of three ports due to acute cholangitis (global mortality = 0.3%). Percentage of complications was 4.9%; three ports were used in 355 (92%) patients and four ports in 31 (8%). When comparing both techniques, in-hospital stay was lower in the three ports group, 1.92 ± 1.22 days versus 2.87 ± 2.84 days (p = 0.0001); bleeding was lower, 55.23 + 123.48 ml versus 114.52 ± 193.04 ml (p = 0.0001); as well as surgical time, 71.05 ± 41.87 min versus 110.26 ± 61.25 min (p = 0.0001). There were no differences in morbidity (frequency of fistulae and injuries to the biliary tract). Conclusions: Morbidity and mortality of LC at the HRAEO is similar to that reported in the world literature. The use of three ports, as compared to four ports, demonstrated a lower surgical time, in-hospital stay, and bleeding.

Fluorescence in situ hybridization for detection of classical propionibacteria with specific 16S rRNA-targeted probes and its application to enumeration in Gruyère cheese.

The classical or dairy propionibacteria have well-documented industrial applications and have been proposed for probiotic applications. Given their industrial importance it is necessary to employ fast and reliable techniques to monitor the growth during products elaboration, industrial fermentations or the intestinal transit. Therefore, the aim of this investigation was to design oligonucleotide probes targeting the 16S rRNA of dairy propionibacteria and optimise the fluorescence in situ hybridization (FISH) protocol to detect these bacteria. Two specific probes were in silico designed to detect Propionibacterium freudenreichii and P. jensenii, named Pfr435 and Pj446 respectively. The FISH protocol was optimised for the hybridisation of propionibacteria cells with the universal probe Eub338 and the designed probes. These probes were assayed in situ for their specificity to hybridise species of propionibacteria by observation using fluorescence microscopy and results were compared with the probe Pap446 previously designed for P. acidipropionici. Probes Pap446, Pfr435 and Pj446 were also evaluated by fluorescence spectrophotometry to assess the influence of cells physiological state during growth in batch culture in the fluorescence intensity. The maximum fluorescence intensity was observed at the onset of the stationary phase of growth and was then reduced. However, changes on the cells permeability did not reduce the efficiency of 16S rRNA hybridisation with the fluorescence-labelled probes. Propionibacteria counts obtained by FISH and plate count methods were compared in a commercial Gruyère cheese. The results showed that this method can be used as a rapid technique for the enumeration of these bacteria in cheese samples.