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Reported cases of tick-borne diseases have steadily increased for more than a decade. In the United States, a majority of tick-borne infections are caused by bacteria. Clinical diagnosis may be challenging, as tick-borne diseases can present with similar symptoms. Laboratory diagnosis has historically relied on serologic methods, which have limited utility during the acute phase of disease. Pathogen-specific molecular methods have improved early diagnosis, but can be expensive when bundled together and may miss unexpected or novel pathogens. To address these shortcomings, we developed a 16S rRNA gene PCR with a next-generation sequencing (NGS) approach to detect tick-borne bacteria in whole blood. A workflow was optimized by comparing combinations of two extraction platforms and two primer sets, ultimately pursuing DNA extraction from blood with the MagNA Pure 96 and PCR amplification using dual-priming oligonucleotide primers specific to the V1-V3 region of the 16S rRNA gene. The amplified product underwent modified Illumina 16S metagenomics sequencing library preparation and sequencing on a MiSeq V2 Nano flow cell, with data analysis using Pathogenomix RipSeq NGS software. Results with the developed method were compared to those from a V1-V2 16S rRNA gene primer set described by the Centers for Disease Control and Prevention (CDC). The V1-V3 assay demonstrated equivalent performance to the CDC assay, with each method showing concordance with targeted PCR results in 31 of 32 samples, and detecting 22 of 23 expected organisms. These data demonstrate the potential for using a broad-range bacterial detection approach for diagnosis of tick-borne bacterial infection from blood.Multilocus sequence typing (MLST) provides allele-based characterization of bacterial pathogens in a standardized framework. However, classical MLST schemes for Bordetella pertussis, the causative agent of whooping cough, seldom reveal diversity among the small number of gene targets and thereby fail to delineate population structure. To improve the discriminatory power of allele-based molecular typing of B. pertussis, we have developed a whole-genome MLST (wgMLST) scheme from 225 reference-quality genome assemblies. see more Iterative refinement and allele curation resulted in a scheme of 3,506 coding sequences and covering 81.4% of the B. pertussis genome. This wgMLST scheme was further evaluated with data from a convenience sample of 2,389 B. pertussis isolates sequenced on Illumina instruments, including isolates from known outbreaks and epidemics previously characterized by existing molecular assays, as well as replicates collected from individual patients. wgMLST demonstrated concordance with whole-genome single nucleotide polymorphism (SNP) profiles, accurately resolved outbreak and sporadic cases in a retrospective comparison, and clustered replicate isolates collected from individual patients during diagnostic confirmation. Additionally, a reanalysis of isolates from two statewide epidemics using wgMLST reconstructed the population structures of circulating strains with increased resolution, revealing new clusters of related cases. Comparison with an existing core genome (cgMLST) scheme highlights the stable gene content of this bacterium and forms the initial foundation for necessary standardization. These results demonstrate the utility of wgMLST for improving B. pertussis characterization and genomic surveillance during the current pertussis disease resurgence.
Serum calcification propensity is emerging as an independent predictor for cardiovascular outcomes in high-risk populations. Calcification propensity can be monitored by the maturation time of calciprotein particles in serum (T
test). A low T
value is an independent determinant of cardiovascular morbidity and mortality in various populations. Aim was to investigate the T
and its relationship to type 2 diabetes mellitus.

Using nephelometry, serum T
was cross-sectionally measured in 932 stable patients with type 2 diabetes mellitus (55% male) with a median age of 66 (62-75) years, diabetes duration of 6.5 (3.0-10.2) years and hemoglobin A1c (HbA1c) of 49 (44-54) mmol/mol.

Serum T
was normally distributed with a mean value of 261±66 min. In linear regression, serum T
was lower in women and current smokers. A lower T
value was found in patients with a higher HbA1c or higher systolic blood pressure, insulin users and patients with a longer history of diabetes. The association with HbA1c was independent of other determinants in multivariable analysis. There was no association between T
and previous macrovascular events or the presence of microvascular disease.

Serum calcification propensity is independently associated with glycemic control, suggesting that a lower HbA1c may be associated with better cardiovascular outcomes. Retrospective analysis could not establish an association between a history of macrovascular events and T
, and prospective studies will have to be performed to address this hypothesis.

NCT01570140.
NCT01570140.
The concept of glucolipotoxicity refers to the combined, deleterious effects of elevated glucose and/or fatty acid levels.

To investigate the effects of chronic glucolipotoxicity on glucagon-like peptide-1-(7-36) amide (GLP-1) secretion, we generated glucolipotoxic conditions in human NCI-H716 enteroendocrine cells using either 5 or 25 mM glucose with or without 500 µM palmitate for 72 hours. For in vivo study, we have established a chronic nutrient infusion model in the rat. Serial blood samples were collected for 2 hours after the consumption of a mixed meal to evaluate insulin sensitivity and β-cell function.

Chronic glucolipotoxic conditions decreased GLP-1 secretion and the expressions of pCREB, pGSK3β, β-catenin, and TCF7L2 in NCI-H716 cells. Glucolipotoxicity conditions reduced glucose transporter expression, glucose uptake, and nicotinamide-adenine dinucleotide phosphate (NADPH) levels in L-cells, and increased triglyceride accumulation. In contrast, PPARα and ATP levels were reduced, which corr GLP-1 secretion through changes in glucose and lipid metabolism, gene expressions, and proglucagon biosynthesis and suggest the interrelationship between glucolipotoxicities of L-cells and β-cells which develops earlier than that of L-cells.
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