Proof of concept for a nonadditive stochastic model of supercooled liquids.

The recently proposed nonadditive stochastic model (NSM) offers a coherent physical interpretation for diffusive phenomena in glass-forming systems. This model presents nonexponential relationships between viscosity, activation energy, and temperature, characterizing the non-Arrhenius behavior observed in supercooled liquids. In this work, we fit the NSM viscosity equation to experimental temperature-dependent viscosity data corresponding to 25 glass-forming liquids and compare the fit parameters with those obtained using the Vogel-Fulcher-Tammann (VFT), Avramov-Milchev (AM), and Mauro-Yue-Ellison-Gupta-Allan (MYEGA) models. The results demonstrate that the NSM provides an effective fitting equation for modeling viscosity experimental data in comparison with other established models (VFT, AM, and MYEGA), characterizing the activation energy in fragile liquids, presenting a reliable indicator of the degree of fragility of the glass-forming liquids.

Non-Arrhenius behavior and fragile-to-strong transition of glass-forming liquids.

Characterization of the non-Arrhenius behavior of glass-forming liquids is a broad avenue for research toward the understanding of the formation mechanisms of noncrystalline materials. In this context, this paper explores the main properties of the viscosity of glass-forming systems, considering super-Arrhenius diffusive processes. We establish the viscous activation energy as a function of the temperature, measure the degree of fragility of the system, and characterize the fragile-to-strong transition through the standard Angell's plot. Our results show that the non-Arrhenius behavior observed in fragile liquids can be understood through the non-Markovian dynamics that characterize the diffusive processes of these systems. Moreover, the fragile-to-strong transition corresponds to a change in the spatiotemporal range of correlations during the glass transition process.

Melatonin and Caffeine Supplementation Used, Respectively, as Protective and Stimulating Agents in the Cryopreservation of Human Sperm Improves Survival, Viability, and Motility after Thawing compared to Traditional TEST-Yolk Buffer.

Cryopreservation processes can damage spermatozoa and impair structural and functional cell characteristics. Plasma, nuclear membranes, and cellular organelles can suffer from the freeze and thaw process. This study evaluates the protective and stimulant effect of melatonin and caffeine supplementation on the functional characteristics of human spermatozoa before and after freezing. Thirty seminal samples from normozoospermic men aged 19-45 years old collected between October 2012 and May 2017 were included. Semen samples were supplemented with either 2 mM melatonin (MEL) prior to cryopreservation, 2 mM caffeine (CAF) in postthaw, or CAF and MEL (CM) in precryopreservation and postthaw, respectively. Kinetics and seminal parameters, mitochondrial activity, DNA fragmentation, and reactive oxygen species (ROS) levels were analyzed before and after cryopreservation. A significant reduction in sperm concentration, total and progressive motility, sperm kinetics, and mitochondrial activity, as well as a significant increase in DNA fragmentation and ROS production in postthaw samples compared to fresh samples, was identified. After administration of a caffeine and/or melatonin supplement, there was a significant increase in progressive motility in the CAF (p = 0.005) and CM (p = 0.048) groups, as well as mitochondrial activity in the CM group (p < 0.05). Cryopreservation has negative effects on overall sperm quality and increases ROS production. A combination of caffeine and melatonin in prefreeze and postthaw sperm samples has proven to be a very effective and simple way to improve semen quality. This will be particularly useful for initial low-quality semen samples, those which suffer the most from the freezing/thawing process.

Visual and automatic classification of the cyclic alternating pattern in electroencephalography during sleep.

Cyclic alternating pattern (CAP) is a neurophysiological pattern that can be visually scored by international criteria. The aim of this study was to verify the feasibility of visual CAP scoring using only one channel of sleep electroencephalogram (EEG) to evaluate the inter-scorer agreement in a variety of recordings, and to compare agreement between visual scoring and automatic scoring systems. Sixteen hours of single-channel European data format recordings from four different sleep laboratories with either C4-A1 or C3-A2 channels and with different sampling frequencies were used in this study. Seven independent scorers applied visual scoring according to international criteria. Two automatic blind scorings were also evaluated. Event-based inter-scorer agreement analysis was performed. The pairwise inter-scorer agreement (PWISA) was between 55.5 and 84.3%. The average PWISA was above 60% for all scorers and the global average was 69.9%. Automatic scoring systems showed similar results to those of visual scoring. The study showed that CAP could be scored using only one EEG channel. Therefore, CAP scoring might also be integrated in sleep scoring features and automatic scoring systems having similar performances to visual sleep scoring systems.

Visual and automatic classification of the cyclic alternating pattern in electroencephalography during sleep

Cyclic alternating pattern (CAP) is a neurophysiological pattern that can be visually scored by international criteria. The aim of this study was to verify the feasibility of visual CAP scoring using only one channel of sleep electroencephalogram (EEG) to evaluate the inter-scorer agreement in a variety of recordings, and to compare agreement between visual scoring and automatic scoring systems. Sixteen hours of single-channel European data format recordings from four different sleep laboratories with either C4-A1 or C3-A2 channels and with different sampling frequencies were used in this study. Seven independent scorers applied visual scoring according to international criteria. Two automatic blind scorings were also evaluated. Event-based inter-scorer agreement analysis was performed. The pairwise inter-scorer agreement (PWISA) was between 55.5 and 84.3%. The average PWISA was above 60% for all scorers and the global average was 69.9%. Automatic scoring systems showed similar results to those of visual scoring. The study showed that CAP could be scored using only one EEG channel. Therefore, CAP scoring might also be integrated in sleep scoring features and automatic scoring systems having similar performances to visual sleep scoring systems.

Sedative and cardiorespiratory effects of low doses of xylazine with and without acepromazine in Nordestino donkeys.

BACKGROUND: Information on appropriate protocols for sedation of Nordestino donkeys is scarce. OBJECTIVES: To evaluate the sedative and cardiorespiratory effects of low doses of intravenous (i.v.) xylazine with and without acepromazine in 'Nordestino' donkeys. STUDY DESIGN: Seven healthy female Nordestino donkeys (150 ± 18 kg) were included in this blinded, randomised, crossover experiment. METHODS: Four treatments were administered, consisting of two i.v. injections, at baseline (T0, 1st injection) and 15 min later (T15, 2nd injection). Treatments included acepromazine 0.05 mg/kg bwt + saline (AS), saline + xylazine 0.5 mg/kg bwt (SX0.5), acepromazine + xylazine 0.25 mg/kg bwt (AX0.25) or acepromazine + xylazine 0.5 mg/kg bwt (AX0.5). Sedative and cardiorespiratory parameters were evaluated before T0 and 15, 20, 30, 45, 60, 75 and 90 min after treatment. Degree [height of head above ground (HHAG)] and quality of sedation [ataxia, responses to stimuli and visual analogue scale (VAS) scoring] and respiratory rate were evaluated by the main investigator in situ, and heart rate was measured by an assistant investigator. Three experienced evaluators assessed vídeos for ataxia and responses to stimuli. Normal data were analysed by repeated measures ANOVA, and non-normal by Kruskal-Wallis (P<0.05). RESULTS: HHAG was lower than baseline for 15 min after xylazine administration in AX0.25 and for 30 min in SX0.5 and AX0.5 groups. All treatments with xylazine increased VAS and ataxia scores in situ for 15 min after xylazine administration, with no differences between groups. Ataxia scores in situ were higher in SX0.5 and AX0.5 groups than AS for 15 and 30 min after xylazine administration, respectively. MAIN LIMITATIONS: Absence of a negative control group (saline-saline). CONCLUSION: Acepromazine added to xylazine at 0.25 mg/kg bwt produced briefer and milder sedation than xylazine at 0.5 mg/kg bwt.

Antinociceptive effects of methadone combined with detomidine or acepromazine in horses.

REASONS FOR PERFORMING STUDY: To investigate two protocols to provide antinociception in horses. OBJECTIVES: To evaluate the antinociceptive effects of intravenous methadone combined with detomidine or acepromazine in adult horses. STUDY DESIGN: Randomised, blinded, crossover study. METHODS: Mechanical, thermal and electrical stimuli were applied to the dorsal left and right metacarpus and coronary band of the left thoracic limb, respectively. A thermal stimulus was applied caudal to the withers. The horses were treated with saline (C), a combination of methadone (0.2 mg/kg bwt) and detomidine (10 µg/kg bwt) (MD) or methadone (0.2 mg/kg bwt) and acepromazine (0.05 mg/kg bwt) (MA) at 1 week intervals. Nociceptive thresholds were measured before and at 15 min intervals until 150 min after treatment. Wilcoxon rank-sum and Wilcoxon signed rank tests were used to compare data between groups at each time point and over time within each group, followed by the Bonferroni method to adjust the P value. RESULTS: The mechanical stimulus was the most sensitive test to differentiate the antinociceptive effects of the treatments. Mechanical thresholds were greater after MD than MA between 15 and 30 min and with both MD and MA these thresholds were greater than C from 15 to 60 min. Electrical and thermal limb thresholds were greater after MD than C at 15 and 45 min and at 15, 30, 45, 75 and 105 min, respectively. Thermal limb thresholds were greater with MA than C at 30 min. Thoracic thermal threshold in MD and MA were higher than C at 45, 75, 90 and 120 min and from 30 to 75 min, respectively. CONCLUSIONS: Methadone and acepromazine produced less pronounced mechanical antinociception than MD.

Validation of mechanical, electrical and thermal nociceptive stimulation methods in horses.

REASONS FOR PERFORMING STUDY: To validate a model for investigating the effects of analgesic drugs on mechanical, thermal and electrical stimulation testing. OBJECTIVES: To investigate repeatability, sensitivity and specificity of nociceptive tests. STUDY DESIGN: Randomised experiment with 2 observers in 2 phases. METHODS: Mechanical (M), thermal (TL) and electrical (E) stimuli were applied to the dorsal metacarpus (M-left and TL-right) and coronary band of the left thoracic limb (E) and a thoracic thermal stimulus (TT) was applied caudal to the withers in 8 horses (405 ± 43 kg). Stimuli intensities were increased until a clear avoidance response was detected without exceeding 20 N (M), 60°C (TL and TT) and 15 V (E). For each set of tests, 3 real stimuli and one sham stimulus were applied (32 per animal) using a blinded, randomised, crossover design repeated after 6 months. A distribution frequency and, for each stimulus, Chi-square and McNemar tests compared both the proportion of positive responses detected by 2 observers and the 2 study phases. The κ coefficients estimated interobserver agreement in determining endpoints. Sensitivity (384 tests) and specificity (128 tests) were evaluated for each nociceptive stimulus to assess the evaluators' accuracy in detecting real and sham stimuli. RESULTS: Nociceptive thresholds were 3.1 ± 2 N (M), 8.1 ± 3.8 V (E), 51.4 ± 5.5°C (TL) and 55.2 ± 5.3°C (TT). The level of agreement after all tests, M, E, TL and TT, was 90, 100, 84, 98 and 75%, respectively. Sensitivity was 89, 100, 89, 98 and 70% and specificity 92, 97, 88, 91 and 94%, respectively. CONCLUSIONS: The high interobserver agreement, sensitivity and specificity suggest that M, E and TL tests are valid for pain studies in horses and are suitable tools for investigating antinociceptive effects of analgesics in horses.

Pathology and tissue distribution of turkey coronavirus in experimentally infected chicks and turkey poults.

Twenty 1-day-old specific pathogen free chicks and 20 1-day-old commercially derived turkey poults were inoculated with a Brazilian strain of turkey coronavirus (TCoV) to study the pathogenicity and virus distribution up to 14 days post-inoculation by histopathology, immunohistochemistry, reverse transcriptase polymerase chain reaction and sequencing. At 2-14 dpi, TCoV antigens were detected in the paranasal sinus and lachrymal accessory gland (Harderian gland) of infected chicks and in the ileum, ileocaecal junction and caecum of infected poults. Lymphocytic inflammation was present in these tissues. TCoV was re-isolated from pooled tissue suspensions of nasal concha, Harderian gland and paranasal sinus from chicks, as well as from the ileum, ileocaecal junction and caecum of poults, after three consecutive passages in 28-day-old embryonated turkey eggs. Viral RNA corresponding to the spike gene region (1178-2073 genome position) was amplified from the upper respiratory tract of chickens and from the intestinal tract of poults and phylogenetic analysis confirmed the identity as TCoV. This is the first description of TCoV antigens and mRNA in upper respiratory tissues in experimentally infected chickens.

Iron-limited condition modulates biofilm formation and interaction with human epithelial cells of enteroaggregative Escherichia coli (EAEC).

AIMS: The aim of this study was to investigate the influence of low iron availability on biofilm formation and adherence to HEp-2 cells of enteroaggregative Escherichia coli (EAEC) strains isolated from diarrhoea cases. METHODS AND RESULTS: The ability of EAEC to form biofilm on a plastic surface was evaluated quantitatively and qualitatively after 3 and 18 h of incubation of strains with or without the iron chelator 2,2-dipyridyl. When submitted to low iron conditions, prototype EAEC 042 strain showed a decrease in biofilm formation. Conversely, an increase in biofilm formation was observed for the clinical EAEC strains cultured in restricted iron condition. Moreover, the reduction of iron concentration inhibited the aggregative adherence to HEp-2 cells of all EAEC strains tested. However, all effects promoted by iron chelation were suppressed by thiourea. CONCLUSIONS: Low iron availability may modulate biofilm formation and adhesive properties of EAEC strains to HEp-2 cells. SIGNIFICANCE AND IMPACT OF THE STUDY: The data obtained in this study provide useful insights on the influence of low iron conditions possibly associated with redox stress on the pathogenesis of EAEC strains.

A Bayesian approach to estimate the accuracy of "in-house" ELISA assay to measure rabies antibodies from compulsory vaccinated dogs and cattle

Rabies is a vaccine-preventable disease that causes acute encephalitis in mammals, and it is still a significant public health problem in numerous countries. lnfected dogs represent the main vectors involved in human rabies. Additionally, cattle rearing close to geographic areas where vampire bats are found presents an important connection with rural epidemiology. We applied two "in-house" enzyme-linked immunosorbent assay (ELISA) methodologies, considered alternatives to measure antibodies from vaccinated dogs and cattle, without employing the gold standard approach. The ELISA assays were performed on individual serum samples taken from domestic adult dogs and cows compulsory vaccinated against rabies (147 urban dogs and 64 cows; n equal 211). The sandwich and liquid-phase competitive ELISA (scELlSA and ipcELlSA). considered "in-house" assays. were performed according to previous works. The only statistical methodology that allows this study is the Bayesian approach, developed to replace the conventional Hui-Walter paradigm. For conditional independent Bayesian model (one population, two tests and no gold standard) the prior information for sensitivity and specificity of each test, mode, prevalence and transformed (alpha, beta) were submitted to Bayesian inference. The "in-house" IpcELISA revealed 16 - out of 261 serum samples - negative results, whereas in scELISA all results were positive. The Bayesian approach showed that prior information was specified for all parameters; posterior medians were SescELISA 89%, SpscELISA 88%, SPipcELISA 95% SeipcELISA 98%, and prevalence (pi) of 99%, without the use of gold standard analysis to measure specific anti-rabies antibodies

Potential pathogenic role of aggregative-adhering Corynebacterium diphtheriae of different clonal groups in endocarditis.

Invasive diseases caused by Corynebacterium diphtheriae have been described increasingly. Several reports indicate the destructive feature of endocarditis attributable to nontoxigenic strains. However, few reports have dealt with the pathogenicity of invasive strains. The present investigation demonstrates a phenotypic trait that may be used to identify potentially invasive strains. The study also draws attention to clinical and microbiological aspects observed in 5 cases of endocarditis due to C. diphtheriae that occurred outside Europe. Four cases occurred in female school-age children (7-14 years) treated at different hospitals in Rio de Janeiro, Brazil. All patients developed other complications including septicemia, renal failure and/or arthritis. Surgical treatment was performed on 2 patients for valve replacement. Lethality was observed in 40% of the cases. Microorganisms isolated from 5 blood samples and identified as C. diphtheriae subsp mitis (N = 4) and C. diphtheriae subsp gravis (N = 1) displayed an aggregative adherence pattern to HEp-2 cells and identical one-dimensional SDS-PAGE protein profiles. Aggregative-adhering invasive strains of C. diphtheriae showed 5 distinct RAPD profiles. Despite the clonal diversity, all 5 C. diphtheriae invasive isolates seemed to display special bacterial adhesive properties that may favor blood-barrier disruption and systemic dissemination of bacteria. In conclusion, blood isolates from patients with endocarditis exhibited a unique adhering pattern, suggesting a pathogenic role of aggregative-adhering C. diphtheriae of different clones in endocarditis. Accordingly, the aggregative-adherence pattern may be used as an indication of some invasive potential of C. diphtheriae strains.

Potential pathogenic role of aggregative-adhering Corynebacterium diphtheriae of different clonal groups in endocarditis

Invasive diseases caused by Corynebacterium diphtheriae have been described increasingly. Several reports indicate the destructive feature of endocarditis attributable to nontoxigenic strains. However, few reports have dealt with the pathogenicity of invasive strains. The present investigation demonstrates a phenotypic trait that may be used to identify potentially invasive strains. The study also draws attention to clinical and microbiological aspects observed in 5 cases of endocarditis due to C. diphtheriae that occurred outside Europe. Four cases occurred in female school-age children (7-14 years) treated at different hospitals in Rio de Janeiro, Brazil. All patients developed other complications including septicemia, renal failure and/or arthritis. Surgical treatment was performed on 2 patients for valve replacement. Lethality was observed in 40 percent of the cases. Microorganisms isolated from 5 blood samples and identified as C. diphtheriae subsp mitis (N = 4) and C. diphtheriae subsp gravis (N = 1) displayed an aggregative adherence pattern to HEp-2 cells and identical one-dimensional SDS-PAGE protein profiles. Aggregative-adhering invasive strains of C. diphtheriae showed 5 distinct RAPD profiles. Despite the clonal diversity, all 5 C. diphtheriae invasive isolates seemed to display special bacterial adhesive properties that may favor blood-barrier disruption and systemic dissemination of bacteria. In conclusion, blood isolates from patients with endocarditis exhibited a unique adhering pattern, suggesting a pathogenic role of aggregative-adhering C. diphtheriae of different clones in endocarditis. Accordingly, the aggregative-adherence pattern may be used as an indication of some invasive potential of C. diphtheriae strains.

Phenotypic and genotypic identification of Candida dubliniensis from subgingival sites in immunocompetent subjects in Argentina.

INTRODUCTION: It is generally recognized that Candida dubliniensis is commonly found in immunocompromised patients, such as those with advanced human immunodeficiency virus infection, at sites of periodontal disease. Since there are no data available for Argentina, the aim of this study was to determine the prevalence of and to identify C. dubliniensis in periodontal pockets from immunocompetent subjects living in Buenos Aires, Argentina, through a comparison of phenotypic and molecular assays. METHODS: Yeasts recovered from subgingival plaque samples were studied for 180 immunocompetent non-smoking patients with periodontal disease. Yeasts were identified by conventional mycological methods and by specific polymerase chain reaction (PCR) assay. Fluconazole and voriconazole susceptibility studies were performed in keeping with the Clinical and Laboratory Standards Institute. RESULTS: Among 76 yeasts isolated, C. dubliniensis comprised 10.5% (n = 8; 95% confidence interval 4.7-19.7), which corresponded to 4.4% of patients studied (8/180). C. albicans was the most frequently isolated species of yeast. A great majority of C. dubliniensis isolates was susceptible with only one isolate resistant to both antifungals. CONCLUSION: Micromorphology on Staib agar was the phenotypic method that was most concordant with PCR and it was useful for selecting presumptive C. dubliniensis. This is the first report to use PCR to identify C. dubliniensis in subgingival fluid from immunocompetent individuals with periodontal disease in Argentina. On the basis of the findings presented here, we confirm that C. dubliniensis can colonize periodontal pockets of immunocompetent patients with periodontal disease.

Validation of an immunohistochemistry assay to detect turkey coronavirus: a rapid and simple screening tool for limited resource settings.

The objective of the present study was to develop and apply the direct immunohistochemistry (D-IHC) assay to search for turkey coronavirus (TCoV) antigens in formalin-fixed embedded-paraffin tissues by the use of biotin-labeled polyclonal antibody. Twenty-eight-day-old embryonated turkey eggs (n = 50) were inoculated with TCoV-purified virus, and 3 d after inoculation, sections from ileum, ileum-cecal junction, and ceca were harvested, fixed in neutral formalin, and embedded in paraffin blocks and used as positive control. In addition, a total of 100 field samples from ileum, ileum-cecal junction, and ceca, collected from 30 to 45-d-old turkeys poults experiencing an outbreak of acute enteritis, were used to search for TCoV by the same D-IHC. All results were compared with those obtained by conventional RT-PCR and indirect fluorescent antibody assay (IFA) for all tested samples. Turkey coronavirus was detected in experimentally infected embryo tissues and also in field samples in 100% of ileum-cecal junction and ceca by the 3 detection procedures. With IFA as a reference assay, sensitivity and specificity of D-IHC were 98 and 58%, whereas sensitivity and specificity of reverse transcription-PCR were 96 and 66%, calculated from the total of tested samples from experimental infection. Each of the examined procedures was highly specific (D-IHC, 93%; RT-PCR, 90%), sensitive (D-IHC, 85%; RT-PCR, 86%), and agreement of both D-IHC and RT-PCR was 99 and 100%, respectively, compared with IFA results obtained from all the field samples. These findings demonstrated the utility of D-IHC for direct detection of TCoV from field samples and considering the sensitivity and specificity found here, can be used as an alternative technique.

Influence of different orthodontic brackets on adherence of microorganisms in vitro.

OBJECTIVE: To define the capacity of different bracket materials to modify the growth and adherence of microorganisms. METHODS: Three types of brackets from the right upper central incisor were used: metallic, ceramic, and composite. Streptococcus mutans and Candida albicans were studied. The association of both species was also evaluated. The brackets were placed in flat-bottomed vials containing basal medium with 20% sucrose added; the flasks were inoculated with each of the microbial suspensions. The samples were incubated at 37 degrees C for 48 hours, after which the brackets were removed. The supernatant was removed from the flasks, the cells adhering to the glass were counted, and the brackets were studied with electron microscopy. RESULTS: The adherence of Streptococcus mutans was not modified by the different brackets. The adherence of Candida albicans was increased by the composite bracket, whereas the use of metallic brackets decreased the number of colony-forming units (CFUs). By electron microscopy we demonstrated that the adherence of Streptococcus mutans plus Candida albicans together varied according to the bracket materials with composite > ceramic > metallic. CONCLUSIONS: Orthodontic appliances serve as different impact zones and modify microbial adherence and colonization, acting as foreign reserves and possible sources of infection.

Improving access to the control region and tRNA gene clusters of dipteran mitochondrial DNA.

Despite the successful use of universal primers for amplifying insect mtDNA, specific regions remain difficult to recover and demand the use of taxon-specific primers. In this work, we describe a new set of primers for efficiently amplifying and sequencing the mtDNA control region and three tRNA gene clusters of dipterans of medical and veterinary importance, including Muscidae, Calliphoridae, and Oestridae species. These new primers were useful for investigating the nucleotide information and the structural organization of dipteran mtDNA.

Phenotypic and genotypic characterisation of Escherichia coli strains serogrouped as enteropathogenic E. coli (EPEC) isolated from pasteurised milk.

Fifty-six Escherichia coli strains, serogrouped as EPEC, isolated from three different brands of pasteurised milk commercialised in Rio de Janeiro, Brazil, were tested for enteropathogenicity markers. Most of the strains (71.4%) were adherent to HEp-2 cells. The adherent strains were distributed among 7 EPEC serogroups (O26, O55, O111, O114, O125, O127, O128, O158). Although almost half of these strains (33.9%) presented unrecognisable adherence phenotypes, classical adherence patterns (localised-like, aggregative and diffuse adherence) described for E. coli and epidemiologically associated with diarrheagenic strains were observed. None of the strains showed typical localised adherence, usually associated with EPEC strains, but 4 of them displayed a localised-like adherence (LAL) phenotype, characterised by fewer and less compact microcolonies but that is still associated with diarrheagenic strains as well as strains of non-human origin. Indeed, 3 of these 4 strains were able to elicit the attaching-effacing lesion (FAS-positive), the central feature of EPEC pathogenesis, and hybridised with bfpA and eae DNA probes. The other LAL-positive strain hybridised with the bfpA probe but gave negative results for the eae probe and FAS assays. Interestingly, all LAL-positive strains produced amplicons of 200 bp in the PCR for bfpA, instead of the expected 326 bp fragment. PCR reactions for stx1 and stx2, two shiga-toxin-encoding genes, gave negative results. Typing of LEE-associated genes by PCR showed the profile eae (beta), tir (beta), espA (alpha) and espB (alpha) for one of the LAL-positive strain. The most prevalent adherence phenotype was the aggregative pattern which is observed in strains epidemiologically associated with persistent diarrhea. Additionally, one strain promoted complete detachment of the Hep-2 cell monolayer after 3 h of infection which might be related to the production of citotoxins, a feature that has been increasingly observed in clinical strains. The possession of EPEC-related O and H antigens is no longer deemed an essential characteristic of true pathogenic EPEC strains, emphasising the importance of routinely screen for virulence markers in E. coli strains isolated from foods. Our results are in accordance with data from the literature that demonstrate that environmental strains display atypical features but yet are capable of eliciting the classical A/E lesion and thus must be considered as potentially pathogenic. Further, our results demonstrate the potential of pasteurised milk as a vehicle for transmission of diarrheagenic E. coli in Brazil.

Interactions of Escherichia coli strains of non-EPEC serogroups that carry eae and lack the EAF and stx gene sequences with undifferentiated and differentiated intestinal human Caco-2 cells.

Escherichia coli strains of non-EPEC serotypes that carry eae and lack the EAF and the Shiga toxin (stx) gene sequences have been found in acute diarrhea. Both the cell association and the cell entry of these strains in human intestinal epithelial cells were studied as a function of cell differentiation and polarization. The eae+/EAF-/stx- non-EPEC E. coli strains invaded undifferentiated Caco-2 cells more efficiently than differentiated cells. In contrast, prototype EPEC strain E2348/69 did not show significative differences from invasion rates of undifferentiated and differentiated cells. The uptake of these strains was greatly enhanced by pretreatment of differentiated Caco-2 cells with EGTA. These results suggest that the eae+/EAF-/stx- non-EPEC E. coli invasion of intestinal cells may be dependent on receptors expressed on the surface of undifferentiated cells and the basolateral pole of differentiated cells.

Effects of handling and storage on sterile dental instruments.

The microbial contamination post-sterilization of dental instruments has been the object of permanent study. The aim of the present study was to evaluate factors affecting long-term sterility of dental instruments sterilized in the dry-oven or autoclave at the Central Sterilizing Service of the School of Dentistry, University of Buenos Aires stored under room temperature and humidity conditions. Half of the 192 samples were placed in standard closed metal containers and sterilized in a dry-oven (D.O), and the remaining half were placed in perforated metal containers and sterilized in an autoclave (A). All the samples were placed in sterilizing paper bags for medical use. Post sterilization, each group (DO and A) was divided into: Group I: minimal handling (control); Group II: wrapping torn mechanically (1 cm); Group III: wrapping torn manually (1 cm). All the samples were stored a closed cabinet. Contamination was evaluated at 30 and 180 days, by seeding under aerobic and anaerobic conditions. Temperature was monitored throughout the experiment, and ranged between 20 degrees C and 31 degrees C (x: 24 degrees C +/- 3.9). Humidity was measured with a digital hygrometer, and ranged between 40% and 60% (x: 54% +/- 10). Group I evidenced no microbial contamination, unlike Groups II and III. Our results evidence that 1) dry oven or autoclave sterilized material that is handled properly during storage remains sterile regardless of variations in temperature and humidity; 2) improper handling affects sterility, and contamination is time-dependent.