Data_Sheet_1.DOCX

<p>The spread of carbapenem-resistant Enterobacteriaceae (CRE) mediated by New Delhi metallo-β-lactamase (NDM) poses a serious challenge to clinicians and has become a major public health concern. NDM has been evolving into variants that possess different hydrolysis activity toward antibiotics, so as to affect treatment strategy. In addition, very few studies on NDM variants have focused on animal-derived bacterial isolates. Our study reports a novel NDM variant, NDM-20, in an isolate of Escherichia coli CCD1 recovered from the food animal swine in China. The isolate that was assigned to ST1114, exhibited high level resistance to all β-lactams tested, including aztreonam and carbapenems. The gene of bla_NDM-20 was located on an IncX3-type plasmid, surrounded by multiple insertion sequences. Sequencing analysis demonstrated that bla_NDM-20 contained three point mutations at positions 262 (G→T), 460 (A→C), and 809 (G→A), compared with bla_NDM-1, and just one point mutation at position 809 (G→A), relative to bla_NDM-5. Functional analysis revealed that the bla_NDM-20 transformant, DH5α+pHSG398/NDM-20, exhibited a higher resistance to ertapenem than that of bla_NDM-1 transformant DH5α+pHSG398/NDM-1. Kinetic parameter analysis showed that NDM-20 had increased enzymatic activity against some penicillins and cephalosporins but decreased carbapenemase activity relative to NDM-5. The identification of NDM-20 further confirms the evolution and prevalence of NDM variants in bacteria of food-animal origin.</p

Image_2.TIF

<p>The spread of carbapenem-resistant Enterobacteriaceae (CRE) mediated by New Delhi metallo-β-lactamase (NDM) poses a serious challenge to clinicians and has become a major public health concern. NDM has been evolving into variants that possess different hydrolysis activity toward antibiotics, so as to affect treatment strategy. In addition, very few studies on NDM variants have focused on animal-derived bacterial isolates. Our study reports a novel NDM variant, NDM-20, in an isolate of Escherichia coli CCD1 recovered from the food animal swine in China. The isolate that was assigned to ST1114, exhibited high level resistance to all β-lactams tested, including aztreonam and carbapenems. The gene of bla_NDM-20 was located on an IncX3-type plasmid, surrounded by multiple insertion sequences. Sequencing analysis demonstrated that bla_NDM-20 contained three point mutations at positions 262 (G→T), 460 (A→C), and 809 (G→A), compared with bla_NDM-1, and just one point mutation at position 809 (G→A), relative to bla_NDM-5. Functional analysis revealed that the bla_NDM-20 transformant, DH5α+pHSG398/NDM-20, exhibited a higher resistance to ertapenem than that of bla_NDM-1 transformant DH5α+pHSG398/NDM-1. Kinetic parameter analysis showed that NDM-20 had increased enzymatic activity against some penicillins and cephalosporins but decreased carbapenemase activity relative to NDM-5. The identification of NDM-20 further confirms the evolution and prevalence of NDM variants in bacteria of food-animal origin.</p

Table_1.DOCX

<p>The spread of carbapenem-resistant Enterobacteriaceae (CRE) mediated by New Delhi metallo-β-lactamase (NDM) poses a serious challenge to clinicians and has become a major public health concern. NDM has been evolving into variants that possess different hydrolysis activity toward antibiotics, so as to affect treatment strategy. In addition, very few studies on NDM variants have focused on animal-derived bacterial isolates. Our study reports a novel NDM variant, NDM-20, in an isolate of Escherichia coli CCD1 recovered from the food animal swine in China. The isolate that was assigned to ST1114, exhibited high level resistance to all β-lactams tested, including aztreonam and carbapenems. The gene of bla_NDM-20 was located on an IncX3-type plasmid, surrounded by multiple insertion sequences. Sequencing analysis demonstrated that bla_NDM-20 contained three point mutations at positions 262 (G→T), 460 (A→C), and 809 (G→A), compared with bla_NDM-1, and just one point mutation at position 809 (G→A), relative to bla_NDM-5. Functional analysis revealed that the bla_NDM-20 transformant, DH5α+pHSG398/NDM-20, exhibited a higher resistance to ertapenem than that of bla_NDM-1 transformant DH5α+pHSG398/NDM-1. Kinetic parameter analysis showed that NDM-20 had increased enzymatic activity against some penicillins and cephalosporins but decreased carbapenemase activity relative to NDM-5. The identification of NDM-20 further confirms the evolution and prevalence of NDM variants in bacteria of food-animal origin.</p

Image_1.TIF

<p>The spread of carbapenem-resistant Enterobacteriaceae (CRE) mediated by New Delhi metallo-β-lactamase (NDM) poses a serious challenge to clinicians and has become a major public health concern. NDM has been evolving into variants that possess different hydrolysis activity toward antibiotics, so as to affect treatment strategy. In addition, very few studies on NDM variants have focused on animal-derived bacterial isolates. Our study reports a novel NDM variant, NDM-20, in an isolate of Escherichia coli CCD1 recovered from the food animal swine in China. The isolate that was assigned to ST1114, exhibited high level resistance to all β-lactams tested, including aztreonam and carbapenems. The gene of bla_NDM-20 was located on an IncX3-type plasmid, surrounded by multiple insertion sequences. Sequencing analysis demonstrated that bla_NDM-20 contained three point mutations at positions 262 (G→T), 460 (A→C), and 809 (G→A), compared with bla_NDM-1, and just one point mutation at position 809 (G→A), relative to bla_NDM-5. Functional analysis revealed that the bla_NDM-20 transformant, DH5α+pHSG398/NDM-20, exhibited a higher resistance to ertapenem than that of bla_NDM-1 transformant DH5α+pHSG398/NDM-1. Kinetic parameter analysis showed that NDM-20 had increased enzymatic activity against some penicillins and cephalosporins but decreased carbapenemase activity relative to NDM-5. The identification of NDM-20 further confirms the evolution and prevalence of NDM variants in bacteria of food-animal origin.</p

Data_Sheet_2.DOCX

<p>The spread of carbapenem-resistant Enterobacteriaceae (CRE) mediated by New Delhi metallo-β-lactamase (NDM) poses a serious challenge to clinicians and has become a major public health concern. NDM has been evolving into variants that possess different hydrolysis activity toward antibiotics, so as to affect treatment strategy. In addition, very few studies on NDM variants have focused on animal-derived bacterial isolates. Our study reports a novel NDM variant, NDM-20, in an isolate of Escherichia coli CCD1 recovered from the food animal swine in China. The isolate that was assigned to ST1114, exhibited high level resistance to all β-lactams tested, including aztreonam and carbapenems. The gene of bla_NDM-20 was located on an IncX3-type plasmid, surrounded by multiple insertion sequences. Sequencing analysis demonstrated that bla_NDM-20 contained three point mutations at positions 262 (G→T), 460 (A→C), and 809 (G→A), compared with bla_NDM-1, and just one point mutation at position 809 (G→A), relative to bla_NDM-5. Functional analysis revealed that the bla_NDM-20 transformant, DH5α+pHSG398/NDM-20, exhibited a higher resistance to ertapenem than that of bla_NDM-1 transformant DH5α+pHSG398/NDM-1. Kinetic parameter analysis showed that NDM-20 had increased enzymatic activity against some penicillins and cephalosporins but decreased carbapenemase activity relative to NDM-5. The identification of NDM-20 further confirms the evolution and prevalence of NDM variants in bacteria of food-animal origin.</p

Fluorescence Polarization Immunoassay Based on a New Monoclonal Antibody for the Detection of the Zearalenone Class of Mycotoxins in Maize

To develop a sensitive fluorescence polarization immunoassay (FPIA) for screening the zearalenone class of mycotoxins in maize, two new monoclonal antibodies with uniform affinity to the zearalenone class and four fluorescein-labeled tracers were prepared. After careful selection of appropriate tracer–antibody pairs in terms of sensitivity and specificity, a FPIA that could simultaneously detect the zearalenone class with similar sensitivity was developed. Under optimum conditions, the half maximal inhibitory concentrations of the FPIA in buffer were 1.89, 1.97, 2.43, 1.99, 2.27, and 2.44 μg/L for zearalenone, α-zearalenol, β-zearalenol, α-zearalanol, β-zearalanol, and zearalanone, respectively. The limit of detection of FPIA for the zearalenone class was around 12 μg/kg in maize, and the recoveries ranged from 84.6 to 113.8%, with coefficients of variation below 15.3% in spiked samples. Finally, the FPIA was applied for screening naturally contaminated maize samples, and the results indicated a good correlation with that of high-performance liquid chromatography–tandem mass spectrometry

Association of the Genetic Polymorphisms in Pre-MicroRNAs with Risk of Ischemic Stroke in a Chinese Population

<div><p>microRNA (miRNA) plays a role in the pathogenesis of ischemic stroke, and single nucleotide polymorphisms in miRNA genes may contribute to disease susceptibility. However, the effect of miR-146a, miR-196a2, and miR-499 polymorphisms on ischemic stroke susceptibility has been rarely reported. Using the TaqMan assay, we evaluated the association of hsa-miR-146a/rs2910164, hsa-miR-196a2/rs11614913, and hsa-miR-499/rs3746444 polymorphisms with the risk of ischemic stroke in a Chinese population with 531 ischemic stroke patients and 531 control subjects. Rs2910164 C/G genotypes were significantly associated with increased risk of ischemic stroke in different genetic model (homozygote comparison: OR = 2.00, 95% CI, 1.29–3.12, <i>P</i> = 0.002; additive model: OR = 1.35, 95% CI, 1.10–1.65, <i>P</i> = 0.004;dominant model: OR = 1.33, 95% CI, 1.00–1.75, <i>P</i> = 0.049; recessive model: OR = 1.82, 95% CI, 1.20–2.74, <i>P</i> = 0.004). Subjects with allele G of hsa-miR-146a/ rs2910164 also showed increased risk of ischemic stroke (OR = 1.33, 95% CI, 1.09–1.62, <i>P</i> = 0.005). Stratification analysis showed that the association between rs2910164 and the risk of ischemic stroke was more pronounced in subjects over 60 years old, females, non-drinkers, subjects without hypertension or diabetes mellitus. There were significant combined effects between miR-146a/rs2910164 and fasting glucose/low-density lipoprotein cholesterol levels on ischemic stroke susceptibility. However, we failed to find any association between the alleles/genotypes of rs11614913 T/C and ischemic stroke, respectively (<i>P</i>> 0.05). In summary, this study provides evidence that miR-146a/rs2910164 might be associated with a significantly increased risk of ischemic stroke in a Chinese population, and the combined effects between miRNA polymorphism and fasting glucose /blood lipid levels may contribute to stroke pathogenesis.</p></div

General characteristics of the study population.

<p>General characteristics of the study population.</p

Genotype frequency of miRNA polymorphisms between ischemic stroke patients and control subjects.

<p>Genotype frequency of miRNA polymorphisms between ischemic stroke patients and control subjects.</p