SELENBP1 overexpression in the prefrontal cortex underlies negative symptoms of schizophrenia.

The selenium-binding protein 1 (SELENBP1) has been reported to be up-regulated in the prefrontal cortex (PFC) of schizophrenia patients in postmortem reports. However, no causative link between SELENBP1 and schizophrenia has yet been established. Here, we provide evidence linking the upregulation of SELENBP1 in the PFC of mice with the negative symptoms of schizophrenia. We verified the levels of SELENBP1 transcripts in postmortem PFC brain tissues from patients with schizophrenia and matched healthy controls. We also generated transgenic mice expressing human SELENBP1 (hSELENBP1 Tg) and examined their neuropathological features, intrinsic firing properties of PFC 2/3-layer pyramidal neurons, and frontal cortex (FC) electroencephalographic (EEG) responses to auditory stimuli. Schizophrenia-like behaviors in hSELENBP1 Tg mice and mice expressing Selenbp1 in the FC were assessed. SELENBP1 transcript levels were higher in the brains of patients with schizophrenia than in those of matched healthy controls. The hSELENBP1 Tg mice displayed negative endophenotype behaviors, including heterotopias- and ectopias-like anatomical deformities in upper-layer cortical neurons and social withdrawal, deficits in nesting, and anhedonia-like behavior. Additionally, hSELENBP1 Tg mice exhibited reduced excitabilities of PFC 2/3-layer pyramidal neurons and abnormalities in EEG biomarkers observed in schizophrenia. Furthermore, mice overexpressing Selenbp1 in FC showed deficits in sociability. These results suggest that upregulation of SELENBP1 in the PFC causes asociality, a negative symptom of schizophrenia.

Clinicogenomic characteristics and synthetic lethal implications of germline homologous recombination-deficient hepatocellular carcinoma.

BACKGROUNDS AND AIMS: We performed an in-depth examination of pathogenic germline variants (PGVs) and somatic variants in DNA damage response (DDR) genes in hepatocellular carcinoma (HCC) to explore their clinical and genomic impacts. APPROACH AND RESULTS: We used a merged whole-exome or RNA sequencing data set derived from in-house ( n = 230) and The Cancer Genome Atlas ( n = 362) databases of multiethnic HCC samples. We also evaluated synthetic lethal approaches targeting mutations in homologous recombination (HR) genes using HCC cells selected from five genomic databases of cancer cell lines. A total of 110 PGVs in DDR pathways in 96 patients were selected. Of the PGV carriers, 44 were HR-altered and found to be independently associated with poorer disease-free survival after hepatectomy. The most frequently altered HR gene in both germline and somatic tissues was POLQ , and this variant was detected in 22.7% (10/44) and 23.8% (5/21) of all the corresponding carriers, respectively. PGVs in HR were significantly associated with upregulation of proliferation and replication-related genes and familial risk of HCC. Samples harboring PGVs in HR with loss of heterozygosity were most strongly correlated with the genomic footprints of deficient HR, such as mutation burden and denovoSig2 (analogous to Catalogue of Somatic Mutations in Cancer [COSMIC] 3), and poor outcome. Pharmacologic experiments with HCC cells defective in BRCA2 or POLQ suggested that tumors with this phenotype are synthetic lethal with poly(ADP-ribose) polymerase inhibitors. CONCLUSIONS: Our findings suggest that germline HR defects in HCC tend to confer a poor prognosis and result in distinctive genomic scarring. Tests of the clinical benefits of HR-directed treatments in the affected patients are needed.

Myelination defects in the medial prefrontal cortex of Fkbp5 knockout mice.

The hypothalamic-pituitary-adrenal (HPA) axis plays a principal role in stress response regulation and has been implicated in the etiology of stress-related disorders. The HPA axis regulates the normal synthesis and release of glucocorticoids; dysregulation of the HPA axis causes abnormal responses to stress. FK506-binding protein 5 (FKBP5), a co-chaperone of heat shock protein 90 in the glucocorticoid receptor (GR) molecular complex, is a key GR sensitivity regulator. FKBP5 single nucleotide polymorphisms are associated with dysregulated HPA axis and increased risk of stress-related disorders, including posttraumatic stress disorder (PTSD) and depression. In this study, we profiled the microRNAs (miRNAs) in the medial prefrontal cortex of Fkbp5 knockout (Fkbp5-/- ) mice and identified the target genes of differentially expressed miRNAs using sequence-based miRNA target prediction. Gene ontology analysis revealed that the differentially expressed miRNAs were involved in nervous system development, regulation of cell migration, and intracellular signal transduction. The validation of the expression of predicted target genes using quantitative polymerase chain reaction revealed that the expression of axon development-related genes, specifically actin-binding LIM protein 1 (Ablim1), lemur tyrosine kinase 2 (Lmtk2), kinesin family member 5c (Kif5c), neurofascin (Nfasc), and ephrin type-A receptor 4 (Epha4), was significantly decreased, while that of brain-derived neurotrophic factor (Bdnf) was significantly increased in the brain of Fkbp5-/- mice. These results suggest that axonal development-related genes can serve as potential targets in future studies focused on understanding the pathophysiology of PTSD.

Takotsubo syndrome as an overlooked and elusive cause of a single episode of dyspnea in young women: a case report.

BACKGROUND: Dyspnea is a common symptom in patients presenting to the emergency department. It has a variety of causes that range from non-urgent to life-threatening. One episode of dyspnea in a healthy young person is easy to overlook. However, if the symptoms occur after physically or emotionally stressful events, careful evaluation needs to be undertaken because it may be associated with Takotsubo syndrome, which is rarely expected but can be fatal. Herein, we report the case of Takotsubo syndrome in a healthy young woman who arrived at the emergency department after experiencing a short single episode of dyspnea following a minor surgery. CASE PRESENTATION: A 23-year old woman with no underlying chronic disease underwent closed reduction surgery for a nasal bone fracture under general anesthesia (with sevoflurane as the anesthetic). Approximately 5 h later, she presented to the emergency department with dyspnea, which improved soon upon arrival at the emergency department. There were no other symptoms. The dyspnea occurred about 5 h after being discharged on observation, with an uneventful postoperative course. Her electrocardiogram and chest X-ray findings were unremarkable. On testing, troponin I and creatine kinase myocardial band levels were elevated at 6.122 ng/mL and 11.2 µg/L (reference ranges: 0.000-0.046 ng/mL and 0.0-5.0 µg/L), respectively. Bedside echocardiography revealed an ejection fraction of 25%, with mid-ventricular and apical akinesia and basal hyperkinesia. The pulmonary and coronary angiographic computed tomographic scans were unremarkable. Hence, apical Takotsubo syndrome was suspected. A follow-up echocardiogram taken 5 days after admission showed full recovery with a normalized ejection fraction (60%) and no regional wall motion abnormality. The patient was discharged on the sixth day with no other complications. CONCLUSION: When atypical symptoms, such as transient dyspnea, manifest, it becomes necessary to suspect and diagnose Takotsubo syndrome to ensure timely and appropriate medical management, especially when a preceding stressful event, such as minor surgery has occurred. It might be helpful to perform bedside point-of-care echocardiography to check for regional wall motion abnormalities that are typically associated with Takotsubo syndrome.

ZHO-1, an intrinsic MBL from the environmental Gram-negative species Zhongshania aliphaticivorans.

OBJECTIVES: Our aim was to characterize the putative MBL of the environmental strain Zhongshania aliphaticivorans isolated from a marine environment. METHODS: The putative MBL was identified in silico using the NCBI database. The ß-lactamase gene was cloned into different Escherichia coli backgrounds. Kinetic parameters were determined using the purified enzyme. RESULTS: The enzyme named ZHO-1 shared 51% amino acid identity with the acquired class B carbapenemases IMP-1, KHM-1 and DIM-1. Expression of the blaZHO-1 gene in a susceptible E. coli resulted in a carbapenemase phenotype. Kinetic parameters determined from purified ZHO-1 enzyme showed that it had significant hydrolytic activity against most ß-lactams including penicillins, cephalosporins and carbapenems, with the exception of aztreonam and cefepime. CONCLUSIONS: This study adds to the knowledge regarding environmental species as a reservoir of possible clinically relevant MBLs.

Decreased SGK1 Expression and Function Contributes to Behavioral Deficits Induced by Traumatic Stress.

Exposure to extreme stress can trigger the development of major depressive disorder (MDD) as well as post-traumatic stress disorder (PTSD). The molecular mechanisms underlying the structural and functional alterations within corticolimbic brain regions, including the prefrontal cortex (PFC) and amygdala of individuals subjected to traumatic stress, remain unknown. In this study, we show that serum and glucocorticoid regulated kinase 1 (SGK1) expression is down-regulated in the postmortem PFC of PTSD subjects. Furthermore, we demonstrate that inhibition of SGK1 in the rat medial PFC results in helplessness- and anhedonic-like behaviors in rodent models. These behavioral changes are accompanied by abnormal dendritic spine morphology and synaptic dysfunction. Together, the results are consistent with the possibility that altered SGK1 signaling contributes to the behavioral and morphological phenotypes associated with traumatic stress pathophysiology.

Neuronal carbonic anhydrase VII provides GABAergic excitatory drive to exacerbate febrile seizures.

Brain carbonic anhydrases (CAs) are known to modulate neuronal signalling. Using a novel CA VII (Car7) knockout (KO) mouse as well as a CA II (Car2) KO and a CA II/VII double KO, we show that mature hippocampal pyramidal neurons are endowed with two cytosolic isoforms. CA VII is predominantly expressed by neurons starting around postnatal day 10 (P10). The ubiquitous isoform II is expressed in neurons at P20. Both isoforms enhance bicarbonate-driven GABAergic excitation during intense GABAA-receptor activation. P13-14 CA VII KO mice show behavioural manifestations atypical of experimental febrile seizures (eFS) and a complete absence of electrographic seizures. A low dose of diazepam promotes eFS in P13-P14 rat pups, whereas seizures are blocked at higher concentrations that suppress breathing. Thus, the respiratory alkalosis-dependent eFS are exacerbated by GABAergic excitation. We found that CA VII mRNA is expressed in the human cerebral cortex before the age when febrile seizures (FS) occur in children. Our data indicate that CA VII is a key molecule in age-dependent neuronal pH regulation with consequent effects on generation of FS.

Spatio-temporal transcriptome of the human brain.

Brain development and function depend on the precise regulation of gene expression. However, our understanding of the complexity and dynamics of the transcriptome of the human brain is incomplete. Here we report the generation and analysis of exon-level transcriptome and associated genotyping data, representing males and females of different ethnicities, from multiple brain regions and neocortical areas of developing and adult post-mortem human brains. We found that 86 per cent of the genes analysed were expressed, and that 90 per cent of these were differentially regulated at the whole-transcript or exon level across brain regions and/or time. The majority of these spatio-temporal differences were detected before birth, with subsequent increases in the similarity among regional transcriptomes. The transcriptome is organized into distinct co-expression networks, and shows sex-biased gene expression and exon usage. We also profiled trajectories of genes associated with neurobiological categories and diseases, and identified associations between single nucleotide polymorphisms and gene expression. This study provides a comprehensive data set on the human brain transcriptome and insights into the transcriptional foundations of human neurodevelopment.

Effect of finger motion on transverse median nerve movement in the carpal tunnel.

INTRODUCTION: We used ultrasonography (US) to investigate the effects of finger motion on movement of the median nerve in patients with carpal tunnel syndrome (CTS) and the correlation between these US parameters and CTS severity. METHODS: Ultrasonographic measures were performed in 23 control wrists and 22 CTS wrists in women. During first through third finger flexion and grip motion, median nerve movements were obtained using US and a tracing program. RESULTS: Nerve movements during third finger flexion in the dorsopalmar axis and grip motion in both axes, and during second finger flexion in the radioulnar axis, differed significantly between the control and CTS groups. US parameters correlated negatively with cross-sectional area. CONCLUSIONS: This study shows that transverse median nerve movements decreased during grip using US and correlated negatively with CTS severity. Muscle Nerve, 2016 Muscle Nerve 54: -, 2016 Muscle Nerve 54: 738-742, 2016.

The Different Effect of VEGF Polymorphisms on the Prognosis of Non-Small Cell Lung Cancer according to Tumor Histology.

Vascular endothelial growth factor (VEGF) contributes to tumor angiogenesis. The role of VEGF single nucleotide polymorphisms (SNPs) in lung cancer susceptibility and its prognosis remains inconclusive and controversial. This study was performed to investigate whether VEGF polymorphisms affect survival outcomes of patients with early stage non-small cell lung cancer (NSCLC) after surgery. Three potentially functional VEGF SNPs (rs833061T>C, rs2010963G>C, and rs3025039C>T) were genotyped. A total of 782 NSCLC patients who were treated with surgical resection were enrolled. The association of the SNPs with overall survival (OS) and disease free survival (DFS) was analyzed. In overall population, none of the three polymorphisms were significantly associated with OS or DFS. However, when the patients were stratified by tumor histology, squamous cell carcinoma (SCC) and adenocarcinoma (AC) had significantly different OS (Adjusted hazard ratio [aHR] = 0.76, 95% CI = 0.56-1.03 in SCC; aHR = 1.33, 95% CI = 0.98-1.82 in AC; P for heterogeneity = 0.01) and DFS (aHR = 0.75, 95% CI = 0.58-0.97 in SCC; aHR = 1.26, 95% CI = 1.00-1.60 in AC; P for heterogeneity = 0.004) according to the rs833061T>C genotypes. Our results suggest that the prognostic role of VEGF rs833061T>C may differ depending on tumor histology.

Zhongshania aliphaticivorans sp. nov., an aliphatic hydrocarbon-degrading bacterium isolated from marine sediment, and transfer of Spongiibacter borealis Jang et al. 2011 to the genus Zhongshania as Zhongshania borealis comb. nov.

A Gram-staining-negative, facultatively aerobic bacterium, designated SM-2(T), was isolated from a sea-tidal flat of Yellow Sea, South Korea. Cells were catalase- and oxidase-positive motile rods with a single polar flagellum. Growth of strain SM-2(T) was observed at 10-37 °C (optimum, 25-30 °C), at pH 5.5-8.5 (optimum, pH 7.0-7.5) and in the presence of 0-11% (w/v) NaCl (optimum, 2%). Strain SM-2(T) contained ubiquinone-8 (Q-8) as the sole isoprenoid quinone and C(17:1)ω8c, summed feature 3 (comprising C(16:1)ω7c and/or iso-C(15:0) 2-OH), C(17:0) and C(18:1)ω7c as the major fatty acids. Phosphatidylethanolamine, phosphatidylglycerol, diphosphatidylglycerol and an unidentified lipid were identified as the major cellular polar lipids. The G+C content of the genomic DNA was 52.2 mol%. Phylogenetic analysis based on 16S rRNA gene sequences showed that strain SM-2(T) formed a tight phyletic lineage with Zhongshania antarctica ZS5-23(T), Zhongshania guokunii ZS6-22(T) and Spongiibacter borealis CL-AS9(T), but that S. borealis CL-AS9(T) was distinct from other species of the genus Spongiibacter. Based on 16S rRNA gene sequence similarities, strain SM-2(T) was most closely related to S. borealis CL-AS9(T), Z. antarctica ZS5-23(T) and Z. guokunii ZS6-22(T), with similarities of 99.5%, 98.9% and 98.7%, respectively, but the DNA-DNA hybridization values among these species were clearly lower than 70%. On the basis of chemotaxonomic data and molecular properties, we propose strain SM-2(T) represents a novel species of the genus Zhongshania with the name Zhongshania aliphaticivorans sp. nov. (type strain SM-2(T) =KACC 18120(T) =JCM 30138(T)). We also propose the transfer of Spongiibacter borealis Jang et al. 2011 to the genus Zhongshania as Zhongshania borealis comb. nov. (type strain CL-AS9(T) =KCCM 90094(T) =JCM 17304(T)).

Recovery of synaptic loss and depressive-like behavior induced by GATA1 through blocking of the neuroinflammatory response.

GATA1, a member of the GATA transcription factor family, is a critical factor in hematopoietic system development. In a previous study, we demonstrated the increased expression of GATA1 in the dorsolateral prefrontal cortex (dlPFC) of patients suffering from depression and described its role as a transcriptional repressor of synapse-related genes. In this study, we investigated how GATA1 globally altered gene expression using multi-omics approaches. Through the combined analyses of ChIPseq, mRNAseq, and small RNAseq, we profiled genes that are potentially affected by GATA1 in cultured cortical neurons, and Gene Ontology (GO) analysis revealed that GATA1 might be associated with immune-related functions. We hypothesized that GATA1 induces immune activation, which has detrimental effects including synapse loss and depressive-like behavior. To test this hypothesis, we first performed a microglial morphometric analysis of a brain having overexpression of GATA1 because microglia are the resident immune cells of the central nervous system. Fractal analysis showed that the ramification and process length of microglia decreased in brains having GATA1 overexpression compared to the control, suggesting that GATA1 overexpression increases the activation of microglia. Through flow cytometry and immunohistochemical analysis, we found that activated microglia showed pro-inflammatory phenotypes characterized by the expression of CD86 and CD68. Finally, we demonstrated that the effects of GATA1 overexpression including synapse loss and depressive-like behavior could be blocked by inhibiting microglial activation using minocycline. These results will elucidate the regulatory mechanisms of GATA1 that affect pathophysiological conditions such as depression and provide a potential target for the treatment of depression.

Synaptotagmin-4 induces anhedonic responses to chronic stress via BDNF signaling in the medial prefrontal cortex.

Stressful circumstances are significant contributors to mental illnesses such as major depressive disorder. Anhedonia, defined as loss of the ability to enjoy pleasure in pleasurable situations, including rewarding activities or social contexts, is considered a key symptom of depression. Although stress-induced depression is associated with anhedonia in humans and animals, the underlying molecular mechanisms of anhedonic responses remain poorly understood. In this study, we demonstrated that synaptotagmin-4 (SYT4), which is involved in the release of neurotransmitters and neurotrophic factors, is implicated in chronic stress-induced anhedonia. Employing chronic unpredictable stress (CUS), we evaluated two subpopulations of mice, susceptible (SUS, anhedonic) and resilient (RES, nonanhedonic), based on sucrose preference, which was strongly correlated with social reward. The FosTRAP (targeted recombination in active populations) system and optogenetic approach revealed that neural activity in the medial prefrontal cortex (mPFC) was significantly associated with CUS-induced anhedonic behavioral phenotypes. By conducting weighted gene coexpression network analysis of RNA sequencing data from the mPFC of SUS and RES mice, we identified Syt4 as a hub gene in a gene network that was unique to anhedonia. We also confirmed that Syt4 overexpression in the mPFC was pro-susceptible, while Syt4 knockdown was pro-resilient; the pro-susceptible effects of SYT4 were mediated through a reduction in brain-derived neurotrophic factor (BDNF)-tropomyosin receptor kinase B (TrkB) signaling in the mPFC. These findings suggest that SYT4-BDNF interactions in the mPFC represent a crucial regulatory mechanism of anhedonic susceptibility to chronic stress.

Importance of Bilateral Hip Assessments in Unilateral Lower-Limb Amputees: A Retrospective Review Involving Older Veterans.

Background/Objectives: This study aimed to evaluate bone mineral density (BMD) discordance and its implications in veterans with unilateral lower-limb amputation, emphasizing the need for comprehensive hip assessments. Methods: Data were collected from 84 male veterans, and BMD was measured using dual-energy X-ray absorptiometry (DXA) at the lumbar spine, intact hip, and amputated hip. Results: The T-scores for the lumbar spine, intact hip, and amputated hip were -0.27 ± 1.69, -0.25 ± 1.20, and -1.07 ± 1.33, respectively. Osteoporosis and osteopenia were present in 19% and 34.6% of patients, respectively. Osteopenia and osteoporosis were most prevalent in the hips on the amputated side (32.1% and 13.1%, respectively), followed by the lumbar spines (22.6% and 8.3%) and the hips on the intact side (17.9% and 2.4%). BMD discordance between the lumbar spine and hip was found in 47.6% of participants, while discordance between both hips was observed in 39.3%. Transfemoral amputees had significantly lower BMD at the amputated hip compared to transtibial amputees (-2.38 ± 1.72 vs. -0.87 ± 1.16, p < 0.001). Conclusions: Veterans with unilateral lower-limb amputation exhibit a high prevalence of osteoporosis and significant BMD discordance, particularly between both hips. These findings underscore the necessity for bilateral hip assessments to ensure the accurate diagnosis and effective management of osteoporosis in this population.

Movement of the ulnar nerve at the elbow: a sonographic study.

OBJECTIVES: The aim of this study was to measure the degree of movement of the ulnar nerve in the cubital tunnel using sonography in patients with ulnar neuropathy at the elbow compared to a healthy control group. METHODS: We examined the ulnar nerve in the cubital tunnel using sonography with the elbow extended and then flexed in 26 patients with ulnar neuropathy and 13 control participants. The distance from the ulnar nerve to the skin, medial epicondyle, and tip of olecranon at the inlet of the cubital tunnel and the distance from the ulnar nerve to the skin and olecranon recess at the outlet of the cubital tunnel were measured when the elbow was extended and flexed, respectively. Displacement of the ulnar nerve during elbow extension and flexion between patients with ulnar neuropathy and controls was compared. RESULTS: There was significantly greater displacement of the ulnar nerve to the medial epicondyle at the inlet of the cubital tunnel in the patients with ulnar neuropathy (mean ± SD, 4.22 ± 3.79 mm) compared to the controls (1.76 ± 1.51 mm) during elbow extension and flexion (P = .008). CONCLUSIONS: A significantly greater degree of movement of the ulnar nerve occurs in patients with ulnar neuropathy at the elbow compared to healthy people, as determined by sonography.

Cell type characterization of spatiotemporal gene co-expression modules in Down syndrome brain.

Down syndrome (DS) is the most common genetic cause of intellectual disability and increases the risk of other brain-related dysfunctions, like seizures, early-onset Alzheimer's disease, and autism. To reveal the molecular profiles of DS-associated brain phenotypes, we performed a meta-data analysis of the developmental DS brain transcriptome at cell type and co-expression module levels. In the DS brain, astrocyte-, microglia-, and endothelial cell-associated genes show upregulated patterns, whereas neuron- and oligodendrocyte-associated genes show downregulated patterns. Weighted gene co-expression network analysis identified cell type-enriched co-expressed gene modules. We present eight representative cell-type modules for neurons, astrocytes, oligodendrocytes, and microglia. We classified the neuron modules into glutamatergic and GABAergic neurons and associated them with detailed subtypes. Cell type modules were interpreted by analyzing spatiotemporal expression patterns, functional annotations, and co-expression networks of the modules. This study provides insight into the mechanisms underlying brain abnormalities in DS and related disorders.

Distribution of CRISPR in Escherichia coli Isolated from Bulk Tank Milk and Its Potential Relationship with Virulence.

Escherichia coli is one of the most common causes of mastitis on dairy farms around the world, but its clinical severity is determined by a combination of virulence factors. Recently, clustered regularly interspaced short palindromic repeat (CRISPR) arrays have been reported as a novel typing method because of their usefulness in discriminating pathogenic bacterial isolates. Therefore, this study aimed to investigate the virulence potential of E. coli isolated from bulk tank milk, not from mastitis, and to analyze its pathogenic characterization using the CRISPR typing method. In total, 164 (89.6%) out of 183 E. coli isolated from the bulk tank milk of 290 farms carried one or more of eighteen virulence genes. The most prevalent virulence gene was fimH (80.9%), followed by iss (38.3%), traT (26.8%), ompT (25.7%), afa/draBC (24.0%), and univcnf (21.9%). Moreover, the phylogenetic group with the highest prevalence was B1 (64.0%), followed by A (20.1%), D (8.5%), and C (7.3%) (p < 0.05). Among the four CRISPR loci, only two, CRISPR 1 and CRISPR 2, were found. Interestingly, the distribution of CRISPR 1 was significantly higher in groups A and B1 compared to that of CRISPR 2 (p < 0.05), but there were no significant differences in groups C and D. The prevalence of CRISPR 1 by virulence gene ranged from 91.8% to 100%, whereas that of CRISPR 2 ranged from 57.5% to 93.9%. The distribution of CRISPR 1 was significantly higher in fimH, ompT, afa/draBC, and univcnf genes than that of CRISPR 2 (p < 0.05). The most prevalent E. coli sequence types (EST) among 26 ESTs was EST 22 (45.1%), followed by EST 4 (23.2%), EST 16 (20.1%), EST 25 (19.5%), and EST 24 (18.3%). Interestingly, four genes, fimH, ompT, afa/draBC, and univcnf, had a significantly higher prevalence in both EST 4 and EST 22 (p < 0.05). Among the seven protospacers derived from CRISPR 1, protospacer 163 had the highest prevalence (20.4%), and it only existed in EST 4 and EST 22. This study suggests that the CRISPR sequence-typing approach can help to clarify and trace virulence potential, although the E. coli isolates were from normal bulk tank milk and not from mastitis.

Genetic characterization of third- or fourth-generation cephalosporin-resistant avian pathogenic Escherichia coli isolated from broilers.

The third- or fourth-generation cephalosporins (3GC or 4 GC) are classified as "critically important antimicrobials for human medicine" by WHO, but resistance to these drugs is increasing rapidly in avian pathogenic E. coli (APEC). This study investigated the distribution and genetic characteristics of 3GC- or 4 GC-resistant APEC isolates from five major integrated broiler operations in Korea. The prevalence of 3GC- or 4GC-resistant APEC isolates in 1-week-old broilers was the highest in farms of operation C (53.3%); however, the highest prevalence of these isolates in 4-week-old broilers was the highest on the farms of operation A (60.0%), followed by operations E (50.0%) and C (35.7%). All 49 3GC- or 4GC-resistant APEC isolates had at least one ß-lactamase-encoding gene. The most common ß-lactamase-encoding genes was extended-spectrum ß-lactamase gene, bla CTX-M-15, detected in 24 isolates (49.0%), followed by bla TEM-1 (32.7%). Sixteen isolates (32.7%) harbored class 1 integrons, and four isolates (8.2%) showed different gene cassette-arrangements. However, only 1 of 26 isolates harboring class 2 integrons carried a gene cassette. Furthermore, both CRISPR 1 and 2 arrays were detected in most isolates (36 isolates; 73.5%), followed by CRISPR 2 (18.4%) and CRISPR 1 (4.1%). Interestingly, CRISPR 2 was significantly more prevalent in multidrug resistant (MDR)-APEC isolates than in non-MDR APEC isolates, whereas CRISPR 3 and 4 were significantly more prevalent in non-MDR APEC isolates (each 11.1%; p < 0.05). None of the protospacers of CRISPR arrays were directly associated with antimicrobial resistance. Our findings indicate that the distribution and characteristics of 3GC or 4GC-resistant APEC isolates differed among the integrated broiler operations; moreover, improved management protocols are needed to control the horizontal transmission of 3GC or 4GC-resistant APEC isolates.

Comparison of antimicrobial resistance and molecular characterization of Escherichia coli isolates from layer breeder farms in Korea.

In Korea, 4 big layer companies that possess one grandparent and 3 parent stocks are in charge of 100% of the layer chicken industry. In this study, we investigated the antimicrobial resistance of commensal 578 E. coli isolated from 20 flocks of 4-layer breeder farms (A, B, C, and D), moreover, compared the characteristics of their resistance and virulence genes. Isolates from farms B and D showed significantly higher resistance to the ß-lactam antimicrobials (amoxicillin, ampicillin, and 1st-, 2nd-, and 3rd-generation cephalosporins). However, resistance to ciprofloxacin, nalidixic acid, and tetracycline was significantly higher in the isolates from farm A (P < 0.05). Interestingly, the isolates from farm C showed significantly lower resistance to most antimicrobials tested in this study. The isolates from farms B, C, and D showed the high multiple resistance to the 3 antimicrobial classes. Furthermore, the isolates from farm A showed the highest multiple resistance against the 5 classes. Among the 412 ß-lactam-resistant isolates, 123 (29.9%) carried blaTEM-1, but the distribution was significantly different among the farms from 17.5% to 51.4% (P < 0.05). Similarly, the most prevalent tetracycline resistance gene in the isolates from farms B, C, and D was tetA (50.0-77.0%); however, the isolates from farm A showed the highest prevalence in tetB (70.6%). The distribution of quinolone (qnrB, qnrD, and qnrS) and sulfonamide (su12)-resistant genes were also significantly different among the farms but that of chloramphenicol (catA1)- and aminoglycoside (aac [3]-II, and aac [6']-Ib)-resistant genes possessed no significant difference among the farms. Moreover, the isolates from farm C showed significantly higher prevalence in virulence genes (iroN, ompT, hlyF, and iss) than the other 3 farms (P < 0.05). Furthermore, the phenotypic and genotypic characteristics of E. coli isolates were significantly different among the farms, and improved management protocols are required to control of horizontal and vertical transmission of avian disease, including the dissemination of resistant bacteria in breeder flocks.

Continuous centrifugal microfluidics (CCM) isolates heterogeneous circulating tumor cells via full automation.

Understanding cancer heterogeneity is essential to finding diverse genetic mutations in metastatic cancers. Thus, it is critical to isolate all types of CTCs to identify accurate cancer information from patients. Moreover, full automation robustly capturing the full spectrum of CTCs is an urgent need for CTC diagnosis to be routine clinical practice. Methods: Here we report the full capture of heterogeneous CTC populations using fully automated, negative depletion-based continuous centrifugal microfluidics (CCM). Results: The CCM system demonstrated high performance (recovery rates exceeding 90% and WBC depletion rate of 99.9%) across a wide range of phenotypes (EpCAM(+), EpCAM(-), small-, large-sized, and cluster) and cancers (lung, breast, and bladder). Applied in 30 lung adenocarcinoma patients harboring epidermal growth factor receptor (EGFR) mutations, the system isolated diverse phenotypes of CTCs in marker expression and size, implying the importance of unbiased isolation. Genetic analyses of intra-patient samples comparing cell-free DNA with CCM-isolated CTCs yielded perfect concordance, and CTC enumeration using our technique was correlated with clinical progression as well as response to EGFR inhibitors. Conclusion: Our system also introduces technical advances which assure rapid, reliable, and reproducible results, thus enabling a more comprehensive application of robust CTC analysis in clinical practice.