Condensin ATPase motifs contribute differentially to the maintenance of chromosome morphology and genome stability.

Effective transfer of genetic information during cell division requires a major reorganization of chromosome structure. This process is triggered by condensin, a conserved pentameric ATPase essential for chromosome condensation. How condensin harnesses the energy of ATP hydrolysis to promote chromatin reorganization is unknown. To address this issue, we performed a genetic screen specifically focused on the ATPase domain of Smc4, a core subunit of condensin. Our screen identified mutational hotspots that impair condensin's ability to condense chromosomes to various degrees. These mutations have distinct effects on viability, genome stability, and chromosome morphology, revealing unique thresholds for condensin enzymatic activity in the execution of its cellular functions. Biochemical analyses indicate that inactivation of Smc4 ATPase activity can result in cell lethality because it favors a specific configuration of condensin that locks ATP in the enzyme. Together, our results provide critical insights into the mechanism used by condensin to harness the energy of ATP hydrolysis for the compaction of chromatin.

Potential role of liver enzyme levels as predictive markers of glucose metabolism disorders in a Tunisian population.

The relationship between liver enzymes and T2D risk is inconclusive. We aimed to evaluate the association between liver markers and risk of carbohydrate metabolism disorders, as well as their discriminatory power, for T2D prediction. This cross-sectional study enrolled 216 participants classified as normoglycemic, prediabetic, newly diagnosed diabetics, and diagnosed diabetics. All participants underwent anthropometric and biochemical measurements. The relationship between hepatic enzymes and glucose metabolism markers was evaluated by analyses of covariance. The associations between liver enzymes and incident carbohydrate metabolism disorders were analyzed through logistic regression and their discriminatory capacity to predict T2D by ROC analysis. High AP, ALT, γGT, and AST levels were independently related to decreased insulin sensitivity. Interestingly, a higher AP level was significantly associated with an increased risk of prediabetes (p = 0.017), newly diagnosed diabetes (p = 0.004), and T2D (p = 0.007). An elevated γGT level was an independent risk factor for T2D (p = 0.032) and undiagnosed T2D (p = 0.010) in prediabetic and normoglycemic subjects, respectively. In ROC analysis, AP was a powerful predictor of incident diabetes and significantly improved T2D prediction. Liver enzymes within the normal range, specifically AP levels, are associated with increased risk of carbohydrate metabolism disorders and significantly improved T2D prediction.

Modulatory effects of quercetin on liver histopathological, biochemical, hematological, oxidative stress and DNA alterations in rats exposed to graded doses of score 250.

This study investigated the morphological, biochemical and molecular aspects of liver injury in rats after the exposure to difenoconazole and the protective effects of quercetin against hepatotoxicity and genotoxicity induced by this fungicide. Rats were given graded doses of difenoconazole associated or not to quercetin daily for 20 days. Our results showed a significant increase in PLT (platelets) and WBC (white blood cells) in rats treated with higher doses of difenoconazole (1/38 and 1/9 of LD50). However, a significant decrease in Hb (hemoglobin) rate and RBC (red blood cells) number in rats treated with higher doses of difenoconazole (1/38 and 1/9 of LD50) was obtained. Besides, difenoconazole treatment caused an increase in hepatic enzyme activities of alanine transaminase (ALT), aspartate transaminase (AST), alkaline phosphatase (ALP) and lactate dehydrogenase (LDH). Difenoconazole increased the levels of malondialdehyde (MDA) and advanced oxidation protein products (AOPPs), and decreased superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GPx) activities and vitamin C levels in liver tissues compared to the control group. We also noted a degradation of nucleic acids, testifying difenoconazole genotoxicity. Changes in hepatic tissues were confirmed by histological findings. Co-administration of quercetin (20 mg/kg) improved hematological and biochemical parameters and showed a significant liver protective effect by decreasing MDA levels and producing advanced oxidation protein, along with increased antioxidative enzyme activities and vitamin C levels. Results were confirmed by the improvement of histological impairments. Thus, it appears that quercetin was effective in preventing acute liver injury induced by exposure to difenoconazole.

Metabolic syndrome and physical activity measured by pedometer among adolescents.

AIM: To describe the prevalence of metabolic syndrome and to study the association of physical activity measured by pedometer with the metabolic syndrome components, in a sample of overweight and obese adolescents from Sfax City. METHODS: This study concerned 51 obese and overweight adolescents (28 girls and 23 boys), between the ages of 15 and 18 years, recruited by the unit of obesity and metabolic syndrome department of endocrinology, Hedi Chaker Hospital, University of Sfax, between december 2012 and october 2013. Metabolic syndrome was defined with the International Diabetes Federation (IDF) criteria. Physical activity was monitored with pedometer (Digi-Walker SW-200; Yamax Co, Tokyo, Japan). RESULTS: The frequency of metabolic syndrome was 21.6%. It was significantly higher in obese (25%) than in overweight (15,81%) adolescents (p=0.04). The most common component, associated with abdominal obesity, was hypoHDLemia observed in 58.8 % of the sample. The average steps / day measured by pedometer was significantly higher in subjects without metabolic syndrome than with (9648, 25±2297, 726 vs 7365, 91±1505, 65 steps/day; p=0, 03). Pedometer determined steps/day was inversely correlated with waist circumference (P<0.05), blood pressure (P<0.05) and triglycerides (P<0.05). CONCLUSION: Metabolic syndrome is prevalent in our young population. A more physically active lifestyle appears to be associated with lower probability of metabolic syndrome.

Effects of acrylamide graded doses on metallothioneins I and II induction and DNA fragmentation: Bochemical and histomorphological changes in the liver of adult rats.

The present study investigates the toxic effects of acrylamide (ACR) administered to rats at two doses on (i) oxidative stress and disruption of pro-oxidant/antioxidant balance in hepatic cells and (ii) its correlation with metallothioneins (MTs) genes expression, DNA damage and histomorphological changes. Treated rats with 20 and 40 mg/kg body weight of ACR led to an increase in malondialdehyde, hydrogen peroxide, advanced oxidation protein products, protein carbonyl levels as well as an alteration in the antioxidant status. Total MT content in the liver and MT I and MT II genes induction were increased. Plasma transaminases activities, albumin, total protein and glucose levels were also increased, while alkaline phosphatase activity was decreased. Moreover, total cholesterol (TC), triglyceride, low-density lipoprotein cholesterol (LDL-C) levels, TC/high-density lipoprotein cholesterol (HDL-C) and LDL-C/HDL-C ratios were increased, while HDL-C decreased in a dose-dependent manner. A random DNA degradation was observed only in the liver of ACR-treated rats with the highest dose. These changes were confirmed by histopathological observations.

A functional autophagy pathway is required for rapamycin-induced degradation of the Sgs1 helicase in Saccharomyces cerevisiae.

In yeast Saccharomyces cerevisiae, the immunosuppressant rapamycin mimics starvation by inhibiting the kinase Tor1. We recently documented that this treatment triggers a rapid degradation of Sgs1, a helicase involved in several biological processes such as the prevention of genomic instability. Herein, we show that yeast strains deleted for genes ATG2, ATG9, and PEP4, encoding components of the autophagy pathway, prevent rapamycin-induced degradation of Sgs1. We propose that defects in the autophagy pathway prevent degradation of key proteins in the rapamycin response pathway and as a consequence cause resistance to the drug.

Occurrence of IMP-8, IMP-10, and IMP-13 metallo-ß-lactamases located on class 1 integrons and other extended-spectrum ß-lactamases in bacterial isolates from Tunisian rivers.

BACKGROUND: Antibiotic-resistant bacteria have been surveyed widely in water bodies, but few studies have determined the diversity of antibiotic-resistant bacteria in river waters. This study was undertaken to investigate the origin of resistance among polluted river bacterial isolates in Tunisia. METHODS: In this study 128 isolates resistant to ß-lactam antibiotics were obtained from 2 polluted rivers in the north of Tunisia. Isolates were identified using Phoenix phenotyping criteria. The occurrence of bla(TEM), bla(SHV), bla(CTX-M), bla(CMY), bla(VIM), and bla(IMP) was studied by polymerase chain reaction (PCR) amplification and sequencing, and the genetic relatedness of the 16 IMP-producing Klebsiella pneumoniae isolates was analyzed by comparison of XbaI pulsed-field gel electrophoresis (PFGE) profiles. RESULTS: Using Phoenix phenotyping criteria, diverse genera of bacteria were identified with different rates of prevalence and with different minimum inhibitory concentrations against different antibiotics. The occurrence of bla(TEM), bla(SHV), bla(CTXM), bla(CMY), bla(VIM), and bla(IMP) genes was confirmed. The DNA sequences upstream and downstream of bla(IMP) genes were determined, revealing that all IMP-encoding genes constituted the first cassette of class 1 integrons, followed by aacA gene cassettes encoding aminoglycoside resistance. Comparison of PFGE profiles showed that only 2 of the isolates were clonal, the other 14 displaying unique profiles. The bla(CTX-M) gene was the most dominant of the extended-spectrum ß-lactamase (ESBL) genes, while the bla(TEM) gene was the second-most dominant. CONCLUSION: The discovery of highly diverse ESBL-producing bacteria and metallo-ß-lactamases, particularly bla(IMP), in polluted river water raises alarms with regard to the potential dissemination of antibiotic-resistant bacteria in communities through river environments.

Dyslipidemia in the Greater Tunis population: prevalence and determinants.

BACKGROUND: Economic development and socio-demographic changes have led to increased frequency of cardiovascular disease and other chronic diseases in Tunisia. OBJECTIVES: To assess the prevalence of different types of dyslipidemia and to examine their association with sociodemographic characteristics in the Greater Tunis population. METHODS: The study included 2712 subjects (1228 men and 1484 women) aged 35-70 years, recruited during the years 2004 and 2005 from the Greater Tunis population. Hypercholesterolemia, hypertriglyceridemia, and low HDL cholesterol were defined according to the National Cholesterol Education Program-Adult Treatment Panel III. RESULTS: The prevalence of hypercholesterolemia, hypertriglyceridemia, and low HDL cholesterol was 40.8% (34.9% in males and 45.8% in females; p < 0.001), 29.2% (31.1% in males and 27.6% in females; p < 0.05), and 21.2% (32.5% in males and 11.5% in females; p < 0.001), respectively. The prevalence was higher in urban than rural regions. Hypercholesterolemia was more frequent in illiterate women and in men with high education level. CONCLUSIONS: Dyslipidemias are common in Tunisians, mainly in urban areas, in illiterate women as well as in men with high levels of education. Profound changes of life style and dietary habits of Tunisians are needed to reduce the risk of cardiovascular diseases.

Occurrence of clinical isolates of Klebsiella pneumoniae harboring chromosomally mediated and plasmid-mediated CTX-M-15 ß-lactamase in a Tunisian hospital.

The spread of multidrug-resistant strains of Klebsiella pneumoniae in hospitals is of concern to clinical microbiologists, health care professionals, and physicians because of the impact infections caused by these bacteria have in causing morbidity and mortality. Clinical isolates of K. pneumoniae have been found to show resistance to third-generation cephalosporins as a result of acquiring extended-spectrum ß-lactamase-producing genes, such as bla(CTX-M). Since little is known about the mechanisms of antibiotic resistance observed in Kasserine hospital, Tunisia, this study was undertaken to investigate the mechanisms by which clinical isolates of K. pneumoniae resist ß-lactam antibiotics. Twelve strains of K. pneumoniae were collected from patients admitted to Kasserine hospital; these isolates showed multiresistance phenotypes. Molecular genetics investigations using polymerase chain reaction, S1 digestion, and pulsed-field gel electrophoresisshowed that bla(CTX-M-15) in association with ISEcp1 is responsible for the resistance of these strains to third-generation cephalosporins. It has been determined that bla(CTX-M-15) is chromosomally mediated and plasmid mediated, which alarming need for infection control to prevent the outbreak of such a resistance mechanism.

The isomerase Rrd1 mediates rapid loss of the Sgs1 helicase in response to rapamycin.

In Saccharomyces cerevisiae , rapamycin exposure inhibits the target of rapamycin (TOR) signaling pathway, causing a profound alteration in the transcription pattern of many genes, including those involved in ribosome biogenesis and nutritional changes. Deletion of the RRD1 gene encoding a peptidyl prolyl isomerase resulted in mutants that are resistant to rapamycin. These rrd1Δ mutants are unable to efficiently downregulate genes such as ribosomal protein genes, or to upregulate genes involved in diauxic shift. It is believed that the isomerase function of Rrd1 plays a role in changing the transcriptional profile upon rapamycin exposure. Herein, we set out to search for genes that when deleted in the rrd1Δ mutant would suppress the rapamycin-resistant phenotype. The analysis revealed that deletion of the SGS1 gene in the rrd1Δ mutant partially suppresses the rapamycin-resistant phenotype of the single rrd1Δ mutant. SGS1 encodes a helicase that functions in many biological processes, including transcriptional regulation. We further show, and for the first time, that Sgs1 is rapidly lost in the parent cells in response to rapamycin, but not by other agents. Interestingly, Sgs1 reduction was completely blocked in the rrd1Δ mutant, suggesting that Rrd1 is required to mediate this process. Genes such as PUT4 and HSP42, known to be upregulated in the parent in response to rapamycin, were not induced in the rrd1Δ mutant if the SGS1 gene was deleted. Since Sgs1 plays a role in transcriptional regulation, we propose that it acts as a repressor of a subset of rapamycin responsive genes. Thus, the observed Rrd1-dependent reduction in Sgs1 level may promote expression of specific classes of genes in response to rapamycin.

RNA polymerase II degradation in response to rapamycin is not mediated through ubiquitylation.

In Saccharomyces cerevisiae, the immunosuppressor rapamycin engenders the degradation of excessive RNA polymerase II leading to growth arrest but the regulation of this process is not known yet. Here, we show that this mechanism is dependent on the peptidyl prolyl cis/trans isomerase Rrd1. Strikingly this degradation is independent of RNA polymerase II polyubiquitylation and does not require the elongation factor Elc1. Our data reveal that there are at least two alternative pathways to degrade RNA polymerase II that depend on different type of stresses.

Evolution of ß-lactams resistance in Gram-negative bacteria in Tunisia.

Antimicrobial resistance is a major health problem worldwide, but marked variations in the resistance profiles of bacterial pathogens are found between countries and in different patient settings. In Tunisia, the strikingly high prevalence of resistance of bacteria to penicillins and cephalorosporins drugs including fourth generation in clinical isolates of Gram negative bacteria has been reported. During 30 years, the emerging problem of extended-spectrum ß-lactamase-producing Escherichia coli and Klebsiella pneumoniae isolates is substantial, and some unique enzymes have been found. Recently, evidence that Gram-negative bacteria are resistant to nearly all available antimicrobial agents, including carbapenems, have emerged.

Nocturnal Continuous Positive Airway Pressure (nCPAP) Decreases High-Sensitivity C-Reactive Protein (hs-CRP) in Obstructive Sleep Apnea-Hypopnea Syndrome.

BACKGROUND: Systemic and airway inflammation has recently been linked to obstructive sleep apnea-hypopnea syndrome (OSAHS) and is considered to be a probable risk factor for OSAHS-induced cardiovascular damage. High-sensitivity C-reactive protein (hs-CRP), as an inflammatory mediator, may be useful for the prediction of the risk of cardiovascular disease (CVD) and assessment of nocturnal continuous positive airway pressure (nCPAP) therapy effect in OSAHS patients. METHODS: A prospective, controlled, cross-sectional study was conducted on 64 consecutive adult subjects with suspected sleep-disordered breathing (SDB). RESULTS: OSAHS was confirmed in 43 patients (24 normotensive and 19 hypertensive patients) and ruled out in 21 normotensive subjects (controls). The median plasma level of hs-CRP did not differ significantly between OSAHS patients and controls. It showed an unmarked rise with the severity of OSAHS (p = 0.20) and was not correlated with AHI (p = 0.067; r = 0.28). After adjusting for cervical perimeter (CP), waist-to-hip ratio (WHR), and blood sugar level, hs-CRP level of 1 mg/dL or greater was significantly more often observed in OSAHS patients compared with controls (p = 0.032; OR = 5.60) and was also significantly associated with AHI (p = 0.021). A significant decrease in the median plasma hs-CRP level was observed in CPAP compliant patients (p = 0.006). Of those, only normotensive patients showed a significant decrease in plasma hs-CRP level. In hypertensive ones, however, the hs-CRP level dropped but not significantly. Using a linear regression model, the change in hs-CRP level (Δhs-CRP) following a 6-month-nCPAP therapy was found to positively correlate with the baseline hs-CRP level for both hypertensive (p = 0.02; r = 0.68), and even more normotensive OSAHS patients (p < 0.0001; r = 0.89). CONCLUSION: nCPAP therapy may have a cardiovascular protective effect in OSAHS patients. hs-CRP level would be useful as a valuable predictor of success in OSAHS treatment monitoring.

Affinity kinetics of leptin-reactive immunoglobulins are associated with plasma leptin and markers of obesity and diabetes.

Obese subjects display elevated plasma levels of leptin reflecting the phenomenon of leptin resistance. Here, we aimed to determine whether leptin-reactive immunoglobulins (Ig) are present in obese and type 2 diabetes (T2D) patients and whether their plasma levels and affinity kinetics may correlate with obesity and diabetes markers. We show that leptin levels are increased in obese patients with and without T2D. Although mean plasma levels of leptin-reactive IgG were similar between study groups, IgG in obese non-diabetic patients had increased dissociation rate and lower affinity (increased dissociation equilibrium constant value; KD). In controls and diabetic patients, the association rates of leptin IgG correlated negatively with obesity and diabetes markers, respectively. In contrast, KD values correlated positively with plasma leptin levels and obesity traits in our cohort, and with diabetes markers in both the total cohort and in the obese T2D group. Taken together, our data reveal that leptin-reactive IgG are present in healthy subjects, obese, and diabetic patients but display altered affinity kinetics in obesity. Increased IgG binding to leptin in healthy subjects associated with lower body mass index (BMI) suggests an enhancing role of IgG in leptin signaling. Accordingly, a decreased affinity of IgG for leptin, found in obese patients, can be relevant to leptin resistance.

Potential Predictive Role of Lipid Peroxidation Markers for Type 2 Diabetes in the Adult Tunisian Population.

OBJECTIVES: We evaluated the potential clinical relevance of malondialdehyde (MDA) and autoantibodies to copper oxidized low-density lipoprotein (CuOx-LDL) in type 2 diabetes occurrence. METHODS: This cross-sectional study enrolled 69 normoglycemic subjects, 18 prediabetic patients and 108 type 2 diabetes patients. MDA concentration was assessed spectrophotometrically. Plasma IgG, IgA and IgM levels to CuOx-LDL were determined by ELISA. RESULTS: Plasma MDA levels were considerably higher in obese, prediabetic and type 2 diabetes subjects compared to controls. In multiple linear regression analysis, both MDA and IgA to CuOx-LDL were significantly associated with glucose metabolism markers (p<0.05). Multiple logistic regression analyses showed that high plasma MDA and IgA to CuOx-LDL were independent risk factors for type 2 diabetes (OR 1.196, 95% CI: 1.058 to 1.353; p=0.004; OR 1.626, 95% CI: 1.066 to 2.481; p=0.024; respectively). Importantly, elevated IgA to CuOx-LDL predicted incident diabetes in patients with prediabetes (OR 2.321, 95% CI:1.063 to 5.066; p=0.035). From stratified analyses by body mass index (BMI), both MDA and IgA to CuOx-LDL remained independent predictors of type 2 diabetes occurrence in non-obese subjects (p<0.05). More interesting, elevated IgA to CuOx-LDL levels could be predictors of type 2 diabetes in obese prediabetic subjects (p=0.044). Conversely, neither IgG nor IgM to CuOx-LDL was associated with glucose metabolism markers, obesity or type 2 diabetes. CONCLUSIONS: Plasma MDA and IgA to CuOx-LDL were significantly associated with blood markers of glucose metabolism. High levels of MDA and IgA to CuOx-LDL could independently predict type 2 diabetes development in normoglycemia and prediabetic subjects.

Effects of vanillin on potassium bromate-induced neurotoxicity in adult mice: impact on behavior, oxidative stress, genes expression, inflammation and fatty acid composition.

CONTEXT: Vanillin is known to possess important antioxidant activity. OBJECTIVE: The current study was conducted to establish the therapeutic efficiency of vanillin against potassium bromate (KBrO3)-induced depression-like behavior and oxidative stress in mice. MATERIAL AND METHODS: Mice were exposed during 15 days either to potassium bromate (KBrO3), KBrO3+ vanillin or to only vanillin. RESULTS: Our results revealed a significant modification in the fatty acid composition of the KBrO3-treated mice. In addition, KBrO3 induced a significant reduction in enzymatic activities and gene expressions, Na+ -K+ and Mg2+-ATPases, acetylcholinesterase and butylcholinesterase activities. The gene expression of tumor necrosis factor-α, interleukin-1ß, interleukin-6 and COX2, significantly increased in the cerebrum of KBrO3-treated group. Histopathological observations were consistent with these effects. Co-treatment with vanillin significantly attenuated KBrO3-induced oxidative stress and inflammation. CONCLUSION: This work suggests that vanillin mitigates KBrO3-induced depression, and that this neuroprotective effect proceeds through anti-oxidant and anti-inflammatory activities.

How does serum brain natriuretic peptide level change under nasal continuous positive airway pressure in obstructive sleep apnea-hypopnea syndrome?

BACKGROUND: Obstructive sleep apnea-hypopnea syndrome (OSAHS) is associated with cardiovascular morbidity and mortality, which can be improved by using continuous positive airway pressure (CPAP) therapy. However, the pathophysiological links between the two kinds of disease and the mechanism of the CPAP effect remain incompletely understood. We aimed to inquire into the myocardial involvement in this relationship. We suggested that serum brain natriuretic peptide (BNP) is sensitive enough to detect myocardial stress caused by OSAHS. DESIGN AND METHODS: Sixty-four subjects without cardiovascular disease (21 controls, 24 normotensive OSAHS patients, and 19 hypertensive OSAHS patients) were analyzed for serum BNP at baseline and serially over 6 months. CPAP was applied to 23 patients with severe OSAHS. RESULTS: At baseline, the serum BNP levels were significantly higher (p=0.0001) in the OSAHS group (22.3±14.79 pg/ml) than in the control group (9.2±6.75 pg/ml). Increased serum BNP levels were significantly associated with mean transcutaneous oxygen saturation (SpO2) (p<0.0001), minimal SpO2 (p=0.002), oxygen desaturation index (p=0.001), and total sleep time spent with SpO2 lower than 90% (p=0.002). All patients with elevated BNP levels (≥37 pg/ml) had moderate or severe OSAHS (11/43 OSAHS patients). The more severe the OSAHS, the higher the BNP levels were. However, only the difference between severe and mild OSAHS was statistically significant (p=0.029). Hypertensive OSAHS patients had the highest baseline BNP levels (27.7±16.74 pg/ml). They were significantly higher (p=0.001) than in normotensive OSAHS patients (18±11.72 pg/ml) (p=0.039) and the controls (9.2±6.75 pg/ml). As compared with baseline, treatment with CPAP significantly decreased BNP levels in both hypertensive and normotensive OSAHS patients (respectively, from 36±16.10 to 29.7±14.29 pg/ml, p<0.001, and from 20±10.09 to 16±8.98 pg/ml, p<0.001). In contrast, the BNP levels slightly increased in the controls (from 9.2±6.75 to 9.5±7.02 pg/ml, p=0.029), but there was no statistically significant difference in comparison with the baseline value. The effect of CPAP on BNP levels was more marked in patients with higher baseline BNP levels and those with the most prolonged nocturnal desaturation (p=0.001, r=0.65). It was also more marked in hypertensive OSHAS patients (p=0.015, r=0.72) in comparison with normotensive OSAHS patients (p=0.03, r=0.62). CONCLUSION: BNP seems to be sensitive enough to detect myocardial stress caused by OSAHS. As such, it is a potential marker for screening of preclinical cardiovascular damage in patients with untreated OSAHS. Application of CPAP decreases levels significantly in normotensive and particularly in hypertensive OSAHS. These findings are consistent with previous results suggesting the potential benefits of CPAP on cardiovascular outcome in OSAHS patients.

First report of mefA and msrA/msrB multidrug efflux pumps associated with blaTEM-1 ß-lactamase in Enterococcus faecalis.

OBJECTIVES: Enterococcus faecalis is thought to possess a great deal of intrinsic resistance to several antimicrobial agents. In this study we identified ampicillin- and erythromycin-resistant clinical isolates of E. faecalis and sought to identify the resistance mechanisms among these isolates. METHODS: Twelve isolates of E. faecalis were collected from 12 different patients. Identification of the isolates and their susceptibility patterns were determined using the Phoenix automated phenotypic identification criteria. PCR amplification and sequencing were used to detect ß-lactamase production. Colony blotting was performed in order to screen for multidrug efflux pump production. Extraction and N-terminal sequencing of the multidrug efflux pumps was carried out. RESULTS: The E. faecalis isolates showed high resistance to erythromycin and ampicillin, with minimum inhibitory concentrations of >16 µg/ml. PCR amplification and sequencing showed that isolates produced TEM-1 ß-lactamase. Colony blotting showed that these isolates harbored multidrug efflux pump genes. Multidrug efflux pump extraction, purification, and sequencing showed the distribution of mefA and msrA/msrB efflux pumps. CONCLUSION: Two resistance mechanisms among E. faecalis are described - the production of TEM ß-lactamase and mefA and msrA/msrB efflux pumps. These results are of great interest because this is the first report of the co-existence of these resistance mechanisms among E. faecalis strains.

VIM and IMP metallo-ß-lactamases and other extended-spectrum ß-lactamases in Escherichia coli and Klebsiella pneumoniae from environmental samples in a Tunisian hospital.

An extremely drug-resistant Enterobacteriaceae species emerged in Kasserine Hospital, Tunisia between 2009 and 2010 causing a local outbreak. We aimed to characterize extended-spectrum ß-lactamase (ESBL) and metallo-ß-lactamase (MBL)-producing Enterobacteriaceae from the hospital environment. Swabs were collected from ten different wards from Kasserine Hospital, Tunisia. A total of 46 isolates were cultured onto MacConkey agar supplemented with ceftazidime to select for ESBL-producing Enterobacteriaceae. Identification and susceptibility patterns were performed using Phoenix-automated phenotypic identification criteria. Extended spectrum ß-lactamases (ESBLs) were detected using cefepime ESBL E-test. Colony blotting was first used to detect the occurrence of bla(SHV) , bla(CTX-M) , bla(CMY) , bla(IMP) , and bla(VIM) genes. PCR was used to amplify these genes, and the amplicons were sequenced and analyzed. Total DNA was digested with XbaI, and PFGE was used to type the major isolates that produced IMP-1. Among the 46 isolates, 63% were Klebsiella pneumoniae, 13% were Escherichia coli, 8.7% were Proteus mirabilis, 6% were Enterobacter cloaceae, 4.3% were Providencia rettgeri, 2.5% were Serratia marcescens, and 2.5% were Pantoea agglomerans. PCR amplification and DNA sequencing showed that hospital environment isolates produced SHV-125, CTX-M-15, CMY-2 ESBLs, and IMP-1 and VIM-2 MBLs. PFGE typing showed the emergence of IMP-1 MBL-producing K. pneumoniae isolates that were not clonal. In this study, we report the first characterization of IMP-1 and VIM-2 MBL-producing K. pneumoniae and E. coli isolates collected from Kasserine Hospital, Tunisia.