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The aim of this study is to investigate the occurrence of plasmid mediated quinolone resistance (PMQR) determinants in Campylobacter coli isolates collected from broilers, laying hens and poultry farm environments. One hundred and thirty-nine C. coli isolates were isolated from broilers (n = 41), laying hens (n = 53), eggs (n = 4) and the environment (n = 41) of 23 poultry farms located in northeastern of Tunisia. Antimicrobial susceptibility testing was performed on all isolates according to the recommendation of the European Committee on Antimicrobial Susceptibility Testing guidelines. The detection of PMQR genes: qnrA, qnrB, qnrC, qnrD, qnrS, qepA, and aac(6)-Ib gene was performed using polymerase chain reaction (PCR) and specific primers. aac(6′)-Ib amplicons were further analyzed by digestion with BtsCI to identify the aac(6′)-Ib-cr variant. Mutations in GyrA and the occurrence of RE-CmeABC efflux pump were determined by mismatch amplification mutation assay (MAMA) PCR and PCR, respectively. In addition, eleven isolates were selected to determine their clonal lineage by MLST. The 139 C. coli isolates were resistant to ciprofloxacin, and 86 (61.8%) were resistant to nalidixic acid. High rates of resistance were also observed toward erythromycin (100%), azithromycin (96.4%), tetracycline (100%), chloramphenicol (98.56%), ampicillin (66.1%), amoxicillin-clavulanic acid (55.39%), and kanamycin (57.55%). However, moderate resistance rates were observed for gentamicin (9.35%) and streptomycin (22.3%). All quinolone-resistant isolates harbored the Thr-86-Ile amino acid substitution in GyrA, and the RE-CmeABC efflux pump was detected in 40.28% of isolates. Interestingly, the qnrB, qnrS, qepA, and aac(6′)-Ib-cr were detected in 57.7%, 61.15%, 21.58%, and 10% of isolates, respectively. The eleven isolates studied by MLST belonged to a new sequence type ST13450. This study described for the first time the occurrence of PMQR genes in C. coli isolates in Tunisia and globally.

Campylobacter species, especially C. coli and C. jejuni, have been associated with a range of human gastrointestinal diseases. During the last two decades, due to the irrational use of antibiotics in poultry farms, high rates of antimicrobial resistance have been globally reported in C. coli and C. jejuni isolates. Recently, acquired linezolid-resistance mechanisms have been reported in Campylobacter spp. isolates, which is a cause of concern to human health. In this study, we performed a retrospective analysis of 139 C. coli isolates previously collected from broilers (n = 41), laying hens (n = 53), eggs (n = 4), and environment (n = 41) to detect acquired genes implicated in linezolid resistance. Isolates were tested for their susceptibility to antimicrobial agents using the Kirby–Bauer disk diffusion assay. Chloramphenicol- and linezolid-resistant isolates were subjected to PCR screening for the following genes: fexA, fexB, floR, RE-cmeABC, cfrA, and optrA. The genetic relatedness of eight multidrug-resistant isolates was determined by multilocus sequence typing (MLST). Among the 139 C. coli isolates, high rates of resistance (57.55%–100%) were detected toward nalidixic acid, ciprofloxacin, erythromycin, azithromycin, ampicillin, chloramphenicol, linezolid, and kanamycin. Among 135 chloramphenicol-resistant isolates, the optrA, cfr, fexA floR, RE-cmeABC, and fexB genes were detected in 124 (124/135, 91.85%), 108 (80%), 105 (77.7%), 64 (47.4%), 56 (41, 48%), and 27 (20%) isolates, respectively. In addition, the majority of isolates harbored more than one of these genes. The selected eight isolates belonged to the same sequence type ST13450, which is a new sequence type (ST), not belonging to ST828 and ST1150 complexes. In conclusion, the emergence of optrA gene in Campylobacter spp. isolates makes this genus an optrA reservoir and vector to other pathogens such as Staphylococcus aureus and Enterococcus spp., which is a cause of concern for human and animal health.

Abstract Background Diabetes like many diseases and biological processes is not mono-causal. On the one hand multi-factorial studies with complex experimental design are required for its comprehensive analysis. On the other hand, the data from these studies often include a substantial amount of redundancy such as proteins that are typically represented by a multitude of peptides. Coping simultaneously with both complexities (experimental and technological) makes data analysis a challenge for Bioinformatics. Results We present a comprehensive work-flow tailored for analyzing complex data including data from multi-factorial studies. The developed approach aims at revealing effects caused by a distinct combination of experimental factors, in our case genotype and diet. Applying the developed work-flow to the analysis of an established polygenic mouse model for diet-induced type 2 diabetes, we found peptides with significant fold changes exclusively for the combination of a particular strain and diet. Exploitation of redundancy enables the visualization of peptide correlation and provides a natural way of feature selection for classification and prediction. Classification based on the features selected using our approach performs similar to classifications based on more complex feature selection methods. Conclusions The combination of ANOVA and redundancy exploitation allows for identification of biomarker candidates in multi-dimensional MALDI-TOF MS profiling studies with complex experimental design. With respect to feature selection our method provides a fast and intuitive alternative to global optimization strategies with comparable performance. The method is implemented in R and the scripts are available by contacting the corresponding author.

Abstract The neuropeptide urocortin 3 (UCN3) has a beneficial effect on metabolic disorders, such as obesity, diabetes, and cardiovascular disease. It has been reported that UCN3 regulates insulin secretion and is dysregulated with increasing severity of obesity and diabetes. However, its function in the adipose tissue is unclear. We investigated the overexpression of UCN3 in 3T3-L1 preadipocytes and differentiated adipocytes and its effects on heat shock response, ER stress, inflammatory markers, and glucose uptake in the presence of stress-inducing concentrations of palmitic acid (PA). UCN3 overexpression significantly downregulated heat shock proteins (HSP60, HSP72 and HSP90) and ER stress response markers (GRP78, PERK, ATF6, and IRE1α) and attenuated inflammation (TNFα) and apoptosis (CHOP). Moreover, enhanced glucose uptake was observed in both preadipocytes and mature adipocytes, which is associated with upregulated phosphorylation of AKT and ERK but reduced p-JNK. Moderate effects of UCN3 overexpression were also observed in the presence of 400 μM of PA, and macrophage conditioned medium dramatically decreased the UCN3 mRNA levels in differentiated 3T3-L1 cells. In conclusion, the beneficial effects of UCN3 in adipocytes are reflected, at least partially, by the improvement in cellular stress response and glucose uptake and attenuation of inflammation and apoptosis.

In vertebrates, the octopeptide angiotensin II (AngII) is an important in vivo regulator of the cardiovascular system. It acts mainly through two G protein-coupled receptors, AT1 and AT2. To better understand distinctive features of these receptors, we carried out a phylogenetic analysis that revealed a mirror evolution of AT1 and AT2, each one split into two clades, separating fish from terrestrial receptors. It also revealed that hallmark mutations occurred at, or near, the sodium binding site in both AT1 and AT2. Electrostatics computations and molecular dynamics simulations support maintained sodium binding to human AT1 with slow ingress from the extracellular side and an electrostatic component of the binding free energy around -3kT, to be compared to around -2kT for human AT2 and the δ opioid receptor. Comparison of the sodium binding modes in wild type and mutated AT1 and AT2 from humans and eels indicates that the allosteric control by sodium in both AT1 and AT2 evolved during the transition from fish to amniota. The unusual S7.46N mutation in AT1 is mirrored by a L3.36M mutation in AT2. In the presence of sodium, the N7.46 pattern in amniota AT1 stabilizes the inward orientation of N3.35 in the apo receptor, which should contribute to efficient N3.35 driven biased signaling. The M3.36 pattern in amniota AT2 favours the outward orientation of N3.35 and the receptor promiscuity. Both mutations have physiological consequences for the regulation of the renin-angiotensin system. Author summary The analysis of protein sequences from different species can reveal interesting trends in the structural and functional evolution of a protein family. Here, we analyze the evolution of two G protein-coupled receptors, AT1 and AT2, which bind the angiotensin II peptide and are important regulators of the cardiovascular system. We show that these receptors underwent a mirror evolution. Specific mutations at, or near, the sodium binding pocket occurred in both AT1 and AT2 during the transition to terrestrial life. We carried out electrostatics computations and molecular dynamics simulations to decipher the details of the sodium binding mode in eel and human receptors, as prototypes of fish and amniota receptors. Our results indicate that sodium binding is kinetically slow but thermodynamically stable. Comparison of the sodium binding modes in eel and human receptors reveals that an unusual mutation in the sodium binding pocket of AT1 is critical for biased signaling of amniota AT1 whereas a mutation in AT2 promotes promiscuity of amniota AT2. In turn, these data indicate that a few mutations at a strategic position (here the sodium binding pocket) are an efficient way to gain functional evolution.

Methanol and methanol/water extracts of olive stones and seeds from Olea europaea var. meski were analyzed by reversed-phase high-performance liquid chromatography (HPLC) with diode array detection and mass spectrometry (LC-MS/MS). A total of 28 metabolites were identified; among them are hydroxycinnamic acid derivatives, phenolic alcohols, flavonoids and flavonoid glucosides, secoiridoids, and terpenes. All the extracts were screened for the inhibitory effect of key enzymes related to diabetes and obesity, such as α-amylase and lipase. An in vitro study revealed that Olea meski stone ethanol (MSE) and methanol (MSM) extracts and Olea meski seed ethanol (MSE1) and methanol (MSM1) extracts exert an inhibitory action against lipase and α-amylase. The most potent activity was observed in the StM extract with IC50 equal to 0.19 mg/ml against DPPH oxidation, 1.04 mg/ml against α-amylase, and 2.13 mg/ml against lipase. In HFFD rats, the findings indicated that the increase of body weight, LDL, TC, and glucose levels and then the decrease in HDL-C were significantly suppressed in the MSM-treated group than those in HFFD rats. Moreover, the MSM extract exhibited a prominent selective inhibitory effect against intestinal lipase and α-amylase activities. The MSM extract was also able to protect the liver-kidney functions efficiently, which was evidenced by biochemicals and histological studies.