Faculty Bibliography

OBJECTIVES/OBJECTIVE:Long-non-coding RNAs (lncRNAs) have been involved in central nervous system recently. A number of studies have reported that lncRNA NEAT1 exerts critical roles in neurodegenerative disorder. Beta-site amyloid precursor protein cleaving enzyme 1 (BACE1) has been reported to exert function in the accumulation of amyloid-β (Aβ). Moreover, BACE1 acts as a target of miR-124 in the progression of AD. So far, the biological role and underlying mechanisms of NEAT1 and miR-124 in AD remains elusive. METHODS:The relative NEAT1 and miR-124 expression was examined by qRT-PCR in the tissues and cells line of AD. Cell apoptosis was examined by FACS. Luciferase reporter assay was performed to verify that miR-124 is a direct target of NEAT1, and BACE1 is a downstream target of miR-124. qRT-PCR and western blot analysis were also performed to determinate the BACE1 and the phosphorylation of tau protein. RESULTS:NEAT1 was notably up-regulated and miR-124 was remarkably down-regulated in AD mouse model. Knockdown of NEAT1 or overexpression of miR-124 showed the protective effects on cellular AD model induced by Aβ. Moreover, miR-124 expression could be up- and down-regulated by suppression or overexpression of NEAT1, respectively. In addition, the expression of BACE1 was the potential functional target of miR-124. These findings suggested that NEAT1 might play a vital role in the development of AD by regulating miR-124/BACE1 axis. DISCUSSION/CONCLUSIONS:The present study showed that NEAT1 worked as a regulating factor to promote the development of AD via modulating miR-124/BACE1 axis, which might be considered as a novel target in AD treatment.

Whole-genome sequencing is increasingly adopted in clinical settings to identify pathogen transmissions, though largely as a retrospective tool. Prospective monitoring, in which samples are continuously added and compared to previous samples, can generate more actionable information. To enable prospective pathogen comparison, genomic relatedness metrics based on single-nucleotide differences must be consistent across time, efficient to compute and reliable for a large variety of samples. The choice of genomic regions to compare, i.e., the core genome, is critical to obtain a good metric. We propose a novel core genome method that selects conserved sequences in the reference genome by comparing its k-mer content to that of publicly available genome assemblies. The conserved-sequence genome is sample set-independent, which enables prospective pathogen monitoring. Based on clinical data sets of 3436 S. aureus, 1362 K. pneumoniae and 348 E. faecium samples, ROC curves demonstrate that the conserved-sequence genome disambiguates same-patient samples better than a core genome consisting of conserved genes. The conserved-sequence genome confirms outbreak samples with high sensitivity: in a set of 2335 S. aureus samples, it correctly identifies 44 out of 44 known outbreak samples, whereas the conserved-gene method confirms 38 known outbreak samples.

In certain regions of New York state, USA, Ixodes scapularis ticks can potentially transmit 4 pathogens in addition to Borrelia burgdorferi: Anaplasma phagocytophilum, Babesia microti, Borrelia miyamotoi, and the deer tick virus subtype of Powassan virus. In a prospective study, we systematically evaluated 52 adult patients with erythema migrans, the most common clinical manifestation of B. burgdorferi infection (Lyme disease), who had not received treatment for Lyme disease. We used serologic testing to evaluate these patients for evidence of co-infection with any of the 4 other tickborne pathogens. Evidence of co-infection was found for B. microti only; 4-6 patients were co-infected with Babesia microti. Nearly 90% of the patients evaluated had no evidence of co-infection. Our finding of B. microti co-infection documents the increasing clinical relevance of this emerging infection.

Compared with conventional serologic, culture-based, and molecular-based diagnostic tests, next-generation sequencing (NGS) provides sequence-evidenced detection of various microbes, without prior knowledge, and thus is becoming a novel diagnostic approach. Herein we describe an RNA-based metatranscriptomic NGS (mtNGS) protocol for culture-independent detection of potential infectious pathogens, using clinical bronchoalveolar lavage specimens as an example. We present both an optimized workflow for experimental sequence data collection and a simplified pipeline for bioinformatics sequence data processing. As shown, the whole protocol takes approximately 24 to 36 hours to detect a wide range of Gram-positive and -negative bacteria and possibly other viral and/or fungal pathogens. In particular, we introduce a spike-in RNA mix as an internal control, which plays a critical role in mitigating false-positive and false-negative results of clinical diagnostic tests. Moreover, our mtNGS method can detect antibiotic resistance genes and virulence factors; although it may not be comprehensive, such information is imperative and helpful for the clinician to make better treatment decisions. Results from our preliminary testing suggest that the mtNGS approach is a useful alterative in diagnostic detection of emerging infectious pathogens in clinical laboratories. However, further improvements are needed to achieve better sensitivity and accuracy.

The purpose of the present study was to evaluate intra-articular ozone injection following arthroscopic surgery for knee osteoarthritis (OA) with regard to its efficacy in pain reduction, joint function and quality of life improvement. The present study retrospectively evaluated 80 patients with symptomatic knee OA (Kellgren-Lawrence grade II or III), who either did or did not receive 20 ml of 20 µg/ml ozone as an intra-articular injection after arthroscopic surgery. The minimum follow-up period was 12 months. The outcomes evaluated for knee OA were pain on the Visual Analogue Scale (VAS), Lequesne Index, Western Ontario and McMaster Universities Osteoarthritis Index (WOMAC) and Clinical Global Impression (CGI). The VAS score in the ozone group was significantly better than that in the control group at all post-operative follow-up time-points (P<0.05). The ozone group also exhibited a significantly greater improvement in Lequesne Index scores (P<0.05). In the ozone group, the score on the WOMAC-pain, WOMAC-stiffness and WOMAC-function subscales, as well as the total WOMAC score decreased significantly (P<0.05). Furthermore, in the ozone group a significantly higher number of patients (P<0.05) with better CGI grades was encountered compared with that in the control group at the 12-month follow-up assessment, despite comparable baseline values in all aforementioned clinical measures between the two groups of patients. The present study suggests that intra-articular ozone injections after arthroscopic surgery may effectively improve the outcomes of arthroscopic surgery in terms of pain relief, functional improvement and quality of life in patients with knee OA of Kellgren-Lawrence grade II or III.

OBJECTIVE:To compare the characteristics and severity of respiratory disease in children testing positive for enterovirus D68 (EV-D68) and for human rhinovirus (RhV). STUDY DESIGN:A retrospective single center study of children presenting with acute respiratory symptoms and positive polymerase chain reaction for RhV/EV from September 1, 2014 through October 31, 2014 was performed. Specimens were subsequently tested specifically for EV-D68 and specimens identified as RhV were subtyped when possible into RhV-A, RhV-B, and RhV-C species. Clinical manifestations in patients with EV-D68 were compared with those with non-EV-D68, RhV, and RhV-C. RESULTS:Of the 173 patients included in the analysis, 72 tested positive for EV-D68, 61 for RhV, and 30 for RhV-C. There were significantly fewer infants in the EV-D68 group. Patients with EV-D68 were more likely than those without EV-D68, and specifically with RhV-C, to have fever and wheezing. Patients with EV-D68 received more magnesium sulfate for respiratory distress not responding adequately to repeated doses of inhaled albuterol. Hospitalized patients with EV-D68 received more bronchodilator therapy than patients with RhV. Patients with EV-D68 were more likely to be admitted to the intensive care unit and were older than patients without EV-D68. There was no difference in length of overall hospitalization or time in the pediatric intensive care unit. CONCLUSIONS:Children with EV-D68 appeared to have more severe respiratory disease on admission than children with RhV as evidenced by higher rates of fever, wheezing, bronchodilator use and pediatric intensive care unit admission. Despite the initial difference in severity, no significant difference in length of stay was found suggesting that patients with EV-D68 recovered as quickly as other groups.

R2R3 MYB transcription factors play key functions in the regulation of secondary metabolites. In the present study, a R2R3 MYB transcriptional factor NtMYB2 was identified from Chinese narcissus (Narcissus tazetta L. var. Chinensis Roem) and functionally characterized. NtMYB2 belongs to subgroup 4 of the R2R3 MYB transcription factor family that are related to repressor MYBs involved in the regulation of anthocyanin and flavonoids. Transient expression confirmed that NtMYB2 strongly reduced the red pigmentation induced by MYB- anthocyanin activators in agro-infiltrated tobacco leaves. Ectopic expression of NtMYB2 in tobacco significantly reduced the pigmentation and altered the floral phenotypes in transgenic tobacco flowers. Gene expression analysis suggested that NtMYB2 repressed the transcript levels of structural genes involved in anthocyanin biosynthesis pathway, especially the UFGT gene. NtMYB2 gene is expressed in all examined narcissus tissues; the levels of transcription in petals and corona is higher than other tissues and the transcription level at the bud stage was highest. These results show that NtMYB2 is involved in the regulation of anthocyanin biosynthesis pathway and may act as a repressor by down regulating the transcripts of key enzyme genes in Chinese narcissus.

We describe the cases of 2 infants with congenital babesiosis born to mothers with prepartum Lyme disease and subclinical Babesia microti infection. The infants both developed anemia, neutropenia, and thrombocytopenia, and 1 infant required red blood cell transfusion. Both infants recovered with treatment. Additional studies are warranted to define the optimal management strategy for pregnant women with early Lyme disease in geographic areas in which B microti infection is endemic.

Complete genome sequences of four toxigenic Clostridium difficile isolates from patients in the lower Hudson Valley, New York, USA, were achieved. These isolates represent four common sequence types (ST1, ST2, ST8, and ST42) belonging to two distinct phylogenetic clades. All isolates have a 4.0- to 4.2-Mb circular chromosome, and one carries a phage.

We recently identified a novel vancomycin-resistant Enterococcus faecium (VREfm) clone ST736 with reduced daptomycin susceptibility. The objectives of this study were to assess the population dynamics of local VREfm strains and genetic alterations predisposing to daptomycin resistance in VREfm ST736 strains. Multilocus sequence typing and single nucleotide variant data were derived from whole-genome sequencing of 250 E. faecium isolates from 1994-1995 (n = 43), 2009-2012 (n = 115) and 2013 (n = 92). A remarkable change was noticed in the clonality and antimicrobial resistance profiles of E. faecium strains between 1994-1995 and 2013. VREfm sequence type 17 (ST17), the prototype strain of clade A1, was the dominant clone (76.7%) recognized in 1994-1995. By contrast, clone ST736 accounted for 46.7% of VREfm isolates, followed by ST18 (26.1%) and ST412 (20.7%) in 2013. Bayesian evolutionary analysis suggested that clone ST736 emerged between 1996 and 2009. Co-mutations (liaR.W73C and liaS.T120A) of the liaFSR system were identified in all ST736 isolates (n = 111, 100%) examined. Thirty-eight (34.2%) ST736 isolates exhibited daptomycin-resistant phenotype, of which 13 isolates had mutations in both the liaFSR and cardiolipin synthase (cls) genes and showed high level of resistance with a daptomycin MIC50 of 32 μg/mL. The emergence of ST736 strains with mutations predisposing to daptomycin resistance and subsequent clonal spread among inpatients contributed to the observed high occurrence of daptomycin resistance in VREfm at our institution. The expanding geographic distribution of ST736 strains in other states and countries raises concerns about its global dissemination.