Effect of the addition of antifreeze protein type I on the quality of post-thawed domestic cat epididymal sperm.

Cryopreservation of domestic cat semen is mainly performed as a model for the establishment of endangered wild feline protocols. The supplementation of antifreeze protein type I (AFP I) to cryopreservation medium has shown improvement in frozen-thawed sperm quality in other species, but its effect on cat semen has not yet been tested. This study aimed to assess the addition of AFP I to cryopreservation medium in domestic cats. Sperm was obtained from the cauda epididymis of orchiectomized cats; sperm was then pooled in Tris buffer and allocated into three treatments, according to AFP I final concentration: 0 (control), 0.1, and 0.5 µg/ml. Nine replicates were cryopreserved in a two-step protocol and subsequently thawed at 37°C for 30 s. There was no difference (P > 0.05) among the control, 0.1 and 0.5 µg/ml groups for parameters such as motility, vitality, functional membrane integrity, mature chromatin, normal morphology, and sperm binding to egg perivitelline membrane. In the 0.5 µg/ml group only, percentages of live sperm with intact acrosome and of sperm with most inactive mitochondria (DAB III) showed a significant reduction, along with a tendency (P = 0.053) to an increase in the percentage of sperm with most active mitochondria (DAB II). In conclusion, the supplementation of 0.1 and 0.5 µg/ml of AFP I did not promote consistent beneficial effects on the overall sperm cryotolerance in domestic cats.

Evaluation of cytotoxicity and genotoxicity of Acacia aroma leaf extracts.

Acacia aroma, native plant from San Luis, Argentina, is commonly used as antiseptic and for healing of wounds. The present study was conducted to investigate the in vitro cytotoxicity and genotoxicity of hot aqueous extract (HAE) and ethanolic extract (EE) of A. aroma. The cytotoxic activity was assayed by neutral red uptake assay on Vero cell. Cell treatment with a range from 100 to 5000 µg/mL of HAE and EE showed that 500 µg/mL and 100 µg/mL were the maximum noncytotoxic concentrations, respectively. The CC50 was 658 µg/mL for EE and 1020 µg/mL for HAE. The genotoxicity was tested by the single-cell gel electrophoresis comet assay. The results obtained in the evaluation of DNA cellular damage exposed to varied concentrations of the HAE showed no significant genotoxic effect at range of 1-20 mg/mL. The EE at 20 mg/mL showed moderate genotoxic effect related to the increase of the DNA percentage contained in tail of the comet; DNA was classified in category 2. At concentrations below 5 mg/mL, the results of cytotoxicity and genotoxicity of aqueous and ethanolic extracts of Acacia aroma guarantee the safety at cell and genomic level. However further studies are needed for longer periods including animal models to confirm the findings.

A tissue-dependent hypothesis of dental caries.

Current understanding of dental caries considers this disease a demineralization of the tooth tissues due to the acid produced by sugar-fermenting microorganisms. Thus, caries is considered a diet- and pH-dependent process. We present here the first metagenomic analysis of the bacterial communities present at different stages of caries development, with the aim of determining whether the bacterial composition and biochemical profile are specific to the tissue affected. The data show that microbial composition at the initial, enamel-affecting stage of caries is significantly different from that found at subsequent stages, as well as from dental plaque of sound tooth surfaces. Although the relative proportion of Streptococcus mutans increased from 0.12% in dental plaque to 0.72% in enamel caries, Streptococcus mitis and Streptococcus sanguinis were the dominant streptococci in these lesions. The functional profile of caries-associated bacterial communities indicates that genes involved in acid stress tolerance and dietary sugar fermentation are overrepresented only at the initial stage (enamel caries), whereas other genes coding for osmotic stress tolerance as well as collagenases and other proteases enabling dentin degradation are significantly overrepresented in dentin cavities. The results support a scenario in which pH and diet are determinants of the disease during the degradation of enamel, but in dentin caries lesions not only acidogenic but also proteolytic bacteria are involved. We propose that caries disease is a process of varying etiology, in which acid-producing bacteria are the vehicle to penetrate enamel and allow dentin degrading microorganisms to expand the cavity.

Occurrence and risk assessment of pesticides in a Mediterranean Basin with strong agricultural pressure (Guadiana Basin: Southern of Portugal).

The study aimed to assess the occurrence and the environmental risk of a group of 51 selected pesticides in the Guadiana Basin (a biodiversity hotspot, in the Mediterranean). The most abundant pesticides were bentazone and 2,4-D, while terbuthylazine together with terbutryn constituted the most ubiquitous pesticides. Eighteen out of the 38 pesticides detected are no longer approved in Europe, and 5 of them are included in the list of priority substances. The risk assessment showed that azinphos ethyl, diflufenican, irganol, imidacloprid, and oxadiazon occurred occasionally, but always in concentrations above their respective ecotoxicological threshold value. Contrary, bentazone, terbuthylazine, and terbutryn presented a high risk in most of the sampled locations and periods. The site-specific risk assessment showed a spatial and temporal pattern, with a higher risk occurring mainly in intermittent streams, in the drought period. The presence of pesticides banned from the EU market since 2009 showed the importance of improving the monitoring process, to identify the main sources of pollution and the fate of these emerging compounds. The results showed the need of implementing actions to improve the sustainable use of pesticides in agricultural areas, working with farmers and management entities to reduce the contamination of aquatic ecosystems. Transboundary water governance is also required to solve potential transboundary contamination problems.

Characterization of a selective and potent antagonist of human P2X(7) receptors, AZ11645373.

BACKGROUND AND PURPOSE: The ATP-gated P2X(7) receptor has been shown to play a role in several inflammatory processes, making it an attractive target for anti-inflammatory drug discovery. We have recently identified a novel set of cyclic imide compounds that inhibited P2X(7) receptor-mediated dye uptake in human macrophage THP-1 cells. In this study the actions and selectivity of one of these compounds, AZ11645373, were characterized. EXPERIMENTAL APPROACH: We measured membrane currents, calcium influx, and YOPRO-1 uptake from HEK cells expressing individual P2X receptors, and YOPRO1 uptake and interleukin-1beta release from THP-1 cells in response to ATP and the ATP analogue benzoylbenzoyl ATP (BzATP). KEY RESULTS: AZ11645373 up to 10 microM, had no agonist or antagonist actions on membrane currents due to P2X receptor activation at human P2X(1), rat P2X(2), human P2X(3), rat P2X(2/3), human P2X(4), or human P2X(5) receptors expressed in HEK cells. AZ11645373 inhibited human P2X(7) receptor responses in HEK cells in a non-surmountable manner with K (B) values ranging from 5 - 20 nM, with mean values not significantly different between assays. K (B) values were not altered by removing extracellular calcium and magnesium. ATP-evoked IL-1beta release from lipopolysaccharide-activated THP-1 cells was inhibited by AZ11645373, IC(50) = 90 nM. AZ11645373 was > 500-fold less effective at inhibiting rat P2X(7) receptor-mediated currents with less than 50% inhibition occurring at 10 microM. CONCLUSIONS AND IMPLICATIONS: AZ11645373 is a highly selective and potent antagonist at human but not rat P2X(7) receptors and will have much practical value in studies of human cells.

Repercusiones en la salud mental del paciente anciano tras padecer COVID-19: trastorno de estrés postraumático. A propósito de un caso / Impacts on the mental health of the elderly patient after COVID-19: Post-traumatic stress disorder. About clinical case

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Transcriptional profiles of Haloferax mediterranei based on nitrogen availability.

The haloarchaeon Haloferax mediterranei is able to grow in the presence of different inorganic and organic nitrogen sources by means of the assimilatory pathway under aerobic conditions. In order to identify genes of potential importance in nitrogen metabolism and its regulation in the halophilic microorganism, we have analysed its global gene expression in three culture media with different nitrogen sources: (a) cells were grown stationary and exponentially in ammonium, (b) cells were grown exponentially in nitrate, and (c) cells were shifted to nitrogen starvation conditions. The main differences in the transcriptional profiles have been identified between the cultures with ammonium as nitrogen source and the cultures with nitrate or nitrogen starvation, supporting previous results which indicate the absence of ammonium as the factor responsible for the expression of genes involved in nitrate assimilation pathway. The results have also permitted the identification of transcriptional regulators and changes in metabolic pathways related to the catabolism and anabolism of amino acids or nucleotides. The microarray data was validated by real-time quantitative PCR on 4 selected genes involved in nitrogen metabolism. This work represents the first transcriptional profiles study related to nitrogen assimilation metabolism in extreme halophilic microorganisms using microarray technology.

Novel conversion of 1,2-disubstituted cis-epoxides to one-carbon homologated allylic alcohols using dimethylsulfonium methylide.

[reaction: see text] The treatment of cis-epoxides with an excess of dimethylsulfonium methylide affords one-carbon homologated allylic alcohols in good to excellent yields.

Epidemiology of burns in Malaga, Spain.

The incidence of burns in the province of Malaga, Spain, was determined by means of a descriptive, cross-sectional, population-based study, and the individual, social and environmental conditions of the patients were analysed. Five hundred families (1846 persons), selected by a three-stage, stratified sampling, were interviewed in their homes. Five hundred and six burns were found in 406 persons (1.25 burns/person); 89.5 per cent of these were in an urban environment and 10.5 per cent in a rural environment. Eighteen and a half per cent of the sample had burnt themselves only once and 4.7 per cent more than once. The burns affected 23.3 per cent of the population, although the majority were of little clinical importance. The risk of burns is greater in the urban environment than in the rural environment, with burns occurring most often in the home (65.8 per cent), and especially in the kitchen. The most frequent burns involve hot liquids with special risk from cooking oil. The other burns (in the strict sense of the word, proper burns or true burns), were primarily caused by contact. The incidence was higher in women (33.0 vs. 21.1 per cent), with burns occurring mostly on the hands. Only 21.9 per cent of the burns received the correct first aid after the accident.

[Role of high-dose chemotherapy with hemopoietic stem-cell support in the treatment of adult patients with high-grade glioma]. / Place de la chimiothérapie intensive avec autogreffe de cellules souches hématopoïétiques dans le traitement des tumeurs gliales de haut grade chez l'adulte.

Despite surgery, post-operative irradiation and adjuvant conventional chemotherapy, prognosis of high-grade gliomas remains poor. Carmustine (BCNU) has been shown to have limited activity at conventional dosage but is still the standard chemotherapy. Activity of chemotherapy is limited by the blood-brain barrier impermeability and high levels of expression of multidrug resistance proteins on tumor and/or endothelial cells. Despite high response rates, development of intra-arterial chemotherapy remains limited because of frequent acute brain toxicity related to drug administration. High-dose intravenous chemotherapy rescued by autologous hemopoietic stem cell transplantation is an alternative that might increase drug delivery through the blood-brain barrier and tumor control. Several phase I-II trials using high-dose BCNU were published. The maximum tolerated dose seems to be 800 mg/m2 and interstitial pneumonitis and hepatitis are dose-limiting toxicities. Few phase I-II trials of high-dose therapy were published using drug combinations. High response rates in patients with progressive tumor were observed and in adjuvant setting, encouraging results in terms of median survival time and long survivors were published. No phase III trial was reported to date. Future investigations should include randomized trials comparing high-dose and conventional-dose chemotherapy and development of new high-dose regimens that incorporate new drugs such as temozolomide.

[Radiotherapy of stage T1-T2 M0 prostatic adenocarcinoma. Analysis of the carcinologic results of a multicenter study of 610 patients. Groupe Radiothérapie de la Commission de Coopération Médicale Intercentres (CCMI)]. / Radiothérapie de l'adénocarcinome prostatique de stade T1-T2 M0. Analyse des résultats carcinologiques d'une étude multicentrique de 610 patients. Groupe Radiothérapie de la Commission de Coopération Médicale Intercentres (CCMI).

PURPOSE: Retrospective analysis of the results of radiotherapy in localized prostatic adenocarcinoma. Complications were excluded. PATIENTS AND METHODS: Six-hundred-and-ten T1-T2 adenocarcinomas of the prostate were treated with continuous courses of external beam radiation therapy in 19 participating Institutes between January 1983 and January 1988. The mean follow-up was 10.4 years; the mean age of patients at the beginning of radiotherapy was 68.5 years. RESULTS: A 10-year, local control had been achieved in 86% of T1-T2 (81.4% for T2). The 5- and 10-year metastatic relapse rates were 25.3% and 30% (29% and 38.1% for T2), respectively. At 10 years, 62.4% of T1-T2 were recurrence-free; overall survival rate was 45.8% and cause-specific survival rate was 70.5%; 29.9% of T1-T2 patients were alive and disease-free. T category (TNM), pathologic grade, pelvic lymph node status, local tumor control, and obstructive ureteral symptoms were correlated with survival. The influence of pelvic nodes radiation, dose, overall treatment time, previous endocrine treatment, and transuretral resection was not significant for disease-free survival (alive and disease-free) and other endpoints. CONCLUSION: There was no difference between the French series (1975-1982 and 1983-1988). The results of the literature are comparable to ours. As far as prognostic factors are concerned, this report provides evidence that the explainable variables which influence survival depend on the tumor and patient status.

[Bacteriologic analysis and detection of Clostridium botulinum spores in honey]. / Análisis bacteriológico e investigación de esporas de Clostridium botulinum en mieles.

The bacteriological analysis and, particularly, the detection of Clostridium botulinum spores from 42 honey samples collected in apiaries of the province of San Luis as well as neighbouring areas of La Pampa, Córdoba and Mendoza, were carried out. Samples were processed by the dilution and centrifugation procedures. For spores detection, culture of the pellets were performed in 2 tubes with cooked meat medium (MCC), one of them warmed up to 80 degrees C for 15 min, and both incubated at 30 degrees C during 7 days. Mice were used to search for toxin in the supernatant. Sediments were also searched for anaerobic bacteria detection in yolk agar plates and in nutritive agar plates for the aerobics. Botulinum toxin type A production was found in one of the MCC cultures. No anaerobic bacteria were isolated. All samples contained Bacillus spp.; 21.4% of the strains, were tentatively classified as B. alvei. A working model for the bacteriological analysis of honey and guides that could be enclosed in publications of official institutions (Figure 1) is proposed.

[Detection of Clostridium botulinum spores in honey]. / Detección de esporas de Clostridium botulinum en mieles.

A total of 177 honey samples were examined for Clostridium botulinum, 68 of which were from commercial origin, 8 from small rural producers for family consumption, and the remaining 6 from fractionizing centers in Mendoza and San Luis provinces in Argentina. C. botulinum type A was detected in two samples of rural producer origin (1.1%) by the centrifugation-dilution method. The strain was recovered from one of the samples, obtaining a spore count of 55/g of honey. Even though the positive percentage was lower than that found in other countries, honey consumption by children under one year old should be avoided in order to prevent infant botulism.

[1st isolation of strains of Staphylococcus aureus producing toxic shock syndrome toxin 1 in food handlers in Argentina]. / Primer aislamiento de cepas de Staphylococcus aureus productores de toxina del síndrome de shock tóxico-1 en manipuladores de alimentos en Argentina.

Thirty nine milk handlers from a factory of dairy products in the Province of Buenos Aires were examined for their nasal carriage of S. aureus strains capable of producing toxic-shock-syndrome toxin-1 (TSST-1). In addition, chance samples of handled foods, crude milk and milky fermented derivates (MFD) were studied. Strain isolation was made on Mannitol Salt Agar and on Baird-Parker Agar. Typical colonies were identified by their biochemical properties. Cultures that were found to be S. aureus were selected for analysis of the TSST-1 production. Eight milk handlers (20.5%) were carriers of S. aureus strains. Seven isolates (87.5%) were classified as biotype A (human ecovar) and 1(12.5%) was classified as biotype B (swine and poultry ecovar). Three out of 8 S. aureus biotype A isolates (37.5%), produced TSST-1. Taking into account the number of milk food handlers sampled (39), the carried rate of toxigenic strains was 7.6%. Three S. aureus strains were isolated from crude milk; 1(33.3%) was classified as biotype B and 2(66.6%) as biotype C (cattle and sheep ecovar). Thirteen S. aureus strains were isolated from MDF; 5(38.0%) were classified as biotype A, 1(7.7%) as belonging to biotype B and 7(53.8%) as belonging to biotype C. None of them had the ability to produce TSST-1.

[Prevalence of Staphylococcus aureus isolated from subclinical bovine mastitis in dairies of the city of San Luis]. / Prevalencia de Staphylococcus aureus aislados de mastitis subclínica bovina en tambos de la cuenca lechera de la ciudad de San Luis.

In order to detect subclinical mastitis by means of California Mastitis Test and recounting of somatic cells, 163 cows from the dairies of San Luis city, Argentina, were examined. Seventy six individuals (46.6%) exhibited an inflammatory response ranging > or = 2+ grade and a cellular recounting value of > or = 5 x 10(5), data compatible with those of subclinical mastitis. Staphylococcus aureus was isolated from 39 (51.3%) cultures as estimated by the sum of the two last values listed in Table 1. Organisms were isolated by plating on brain heart infusion agar with 5% of sheep blood and on Baird-Parker media. One hundred and three S. aureus isolates recovered from 51 of 63 cows were characterized by coagulase activity by the tube method using human and bovine plasma; clumping factor; glucose and mannitol fermentation; thermonuclease (TNase), pigment, gelatinase, fibrinolysin, acetoin, hemolysin production; egg yolk, tellurite and catalase reaction and crystal violet types. All isolates were susceptible to cephalothin, clindamycin, methicillin, gentamycin and vancomycin; 94.1% were susceptible to chloramphenicol and 53.8% to G penicillin. Sixty three isolates (61.1%) were classified according to Hájek and Marsálek scheme as biotype C (bovine and ovine ecovar), 33 isolates (32.0%) were classified as biotype B (swine and poultry ecovar); 1 isolated (0.9%) as intermediate between B and D; 5 isolates (4.8%) as biotype A (human ecovar) and 1 isolated (0.9%) as biotype D (ecovar silvestres spp) (Table 2). Production of enterotoxins A to E and toxic shock syndrome toxin-1 (TSST-1) was determined by the optimal susceptibility plate method on 27 isolates (26.2%) which were coagulase 3+ to 4+ and TNase highly positive. None of them produced enterotoxins including TSST-1. The subclinical mastitis data and the prevalence of S. aureus coincide with those of other authors, both from Argentina and from other countries.

[Determination of the sanitary quality and detection of Salmonella spp and Yersinia enterocolitica in ice cream]. / Determinación de la calidad higiénica e investigación de Salmonella spp y Yersinia enterocolítica en helados.

In order to determine the sanitary quality of ice-creams and the presence of pathogenic or potentially pathogenic species of Salmonella and Yersinia enterocolitica, 50 samples from 5 different industrial and semi-industrial producers in San Luis (Argentine) were examined. The enumeration of coliforms was positive for all the samples with values less than or equal to 20/g. Fourteen per cent of the samples were positive for the investigation of Staphylococcus aureus in 1 g. For the plates enumeration 12.0% of the samples gave less than 10 u.f.c./g, 4.0% between 101 and 1000 and 4.0% between 1001 and 10,000. Fifteen strains were isolated, 26.6% biotype A (human ecovar) and the others biotype C (bovine ecovar). All of them were susceptible to chloramphenicol, cephalosporin and erythromycin; 46.6% to penicillin G and ampicillin; 93.3% to kanamycin (6.6% intermediate ones = I); 73.3% to methicillin (26.6% I); 86.6% to tetracycline (13.3% I). Six per cent of the samples over came the acceptability limit for S. aureus. Salmonella spp was not isolated. In 4.0% of the samples Y. enterocolitica were isolated, one of them typified as B1; 0:3, 50, 51, Lis Xz. The latter, isolated in samples with values of coliforms inferior to the limit fixed by some legislations, suggests a post elaboration contamination.

Single-layer centrifugation through PureSperm® 80 selects improved quality spermatozoa from frozen-thawed dog semen.

The aim of this study was to investigate whether single-layer centrifugation (SLC) with PureSperm® 80 could select good quality spermatozoa, including those with specific motility patterns, from doses of frozen dog semen. Semen from 5 dogs was collected and cryopreserved following a standard protocol. After thawing, semen samples were divided into two aliquots: one of them was used as control and the other one processed by SLC. Assessment of sperm motility (assessed by computer-assisted semen analysis), morphology (Diff-Quick staining) and viability (triple fluorescent stain of propidium iodine/isothiocyanate-labeled peanut (Arachis hypogaea) agglutinin/Rhodamine 123), were performed on aliquots of fresh semen, frozen-thawed control and frozen-thawed SLC treated samples. A multivariate clustering procedure separated 26,051 motile spermatozoa into three subpopulations (sP): sP1 consisting of highly active but non-progressive spermatozoa (40.3%), sP2 consisting of spermatozoa with high velocity and progressive motility (30.0%), and sP3 consisting of poorly active and non-progressive spermatozoa (29.7%). SLC with PureSperm® 80 yielded sperm suspensions with improved motility, morphology, viability and acrosome integrity (P<0.001). The frozen-thawed SLC treated samples were enriched in sP2, reaching a proportion of 44.1% of the present spermatozoa. From these results, we concluded that SLC with PureSperm® 80 may be an alternative and successful method for improving the quality of frozen-thawed dog spermatozoa. Moreover, sP2 (high-speed and progressive spermatozoa) was more frequently observed after SLC. Finally, this study also demonstrated that the general motile sperm structure present in dogs remained constant despite the effect caused by either cryopreservation or separation by SLC through PureSperm® 80.