microbiota-analysis-using-two-step-pcr-next-generation-16s-rrna-gene

Microbiota Analysis Using Two-step PCR and Next-generation 16S rRNA Gene Sequencing

The human gut is colonized by trillions of bacteria that support physiologic functions such as food metabolism, energy harvesting, and regulation of the ...

University of Iowa

Background.  We determined the feasibility of using computer vision and depth sensing to detect healthcare worker (HCW)-patient contacts to estimate both hand hygiene (HH) opportunities and personal protective equipment (PPE) adherence. Methods.  We used multiple Microsoft Kinects to track the 3-dimensional movement of HCWs and their hands within hospital rooms. We applied computer vision techniques to recognize and determine the position of fiducial markers attached to the patient's bed to determine the location of the HCW's hands with respect to the bed. To measure our system's ability to detect HCW-patient contacts, we counted each time a HCW's hands entered a virtual rectangular box aligned with a patient bed. To measure PPE adherence, we identified the hands, torso, and face of each HCW on room entry, determined the color of each body area, and compared it with the color of gloves, gowns, and face masks. We independently examined a ground truth video recording and compared it with our system's results. Results.  Overall, for touch detection, the sensitivity was 99.7%, with a positive predictive value of 98.7%. For gowned entrances, sensitivity was 100.0% and specificity was 98.15%. For masked entrances, sensitivity was 100.0% and specificity was 98.75%; for gloved entrances, the sensitivity was 86.21% and specificity was 98.28%. Conclusions.  Using computer vision and depth sensing, we can estimate potential HH opportunities at the bedside and also estimate adherence to PPE. Our fine-grained estimates of how and how often HCWs interact directly with patients can inform a wide range of patient-safety research. 

Using Computer Vision and Depth Sensing to Measure Healthcare Worker-Patient Contacts and Personal Protective Equipment Adherence Within Hospital Rooms.

We have developed a publicly available tool, AxonJ, which quantifies the axons in optic nerve sections of rodents stained with paraphenylenediamine (PPD). In this study, we compare AxonJ's performance to human experts on 100x and 40x images of optic nerve sections obtained from multiple strains of mice, including mice with defects relevant to glaucoma. AxonJ produced reliable axon counts with high sensitivity of 0.959 and high precision of 0.907, high repeatability of 0.95 when compared to a gold-standard of manual assessments and high correlation of 0.882 to the glaucoma damage staging of a previously published dataset. AxonJ allows analyses that are quantitative, consistent, fully-automated, parameter-free, and rapid on whole optic nerve sections at 40x. As a freely available ImageJ plugin that requires no highly specialized equipment to utilize, AxonJ represents a powerful new community resource augmenting studies of the optic nerve using mice.

Automated Axon Counting in Rodent Optic Nerve Sections with AxonJ.

Prolonged space flight, specifically microgravity, presents a problem for space exploration. Animal models with altered connections of the vestibular ear, and thus altered gravity sensation, would allow the examination of the effects of microgravity and how various countermeasures can establish normal function. We describe an experimental apparatus to monitor the effects of ear manipulations to generate asymmetric gravity input on the tadpole escape response. To perform the movement pattern analysis, an imaging apparatus was developed that uses a high-speed camera to obtain time-resolved, high-resolution images of tadpole movements. Movements were recorded in a temperature-controlled test chamber following mechanical stimulation with a solenoid actuator, to elicit a C-start response. Temperature within the test cell was controlled with a recirculating water bath. Xenopus laevis embryos were obtained using a standard fertilization technique. Tadpole response to a controlled perturbation was recorded in unprecedented detail and the approach was validated by describing the distinct differences in response between normal and one-eared tadpoles. The experimental apparatus and methods form an important element of a rigorous investigation into the response of the tadpole vestibular system to mechanical and biochemical manipulations, and can ultimately contribute to improved understanding of the effects of altered gravity perception on humans.

A method for detailed movement pattern analysis of tadpole startle response.

Sonic hedgehog (Shh) signaling plays a major role in vertebrate development, from regulation of proliferation to the patterning of various organs. In amniotes, Shh affects dorsoventral patterning in the inner ear but affects anteroposterior patterning in teleost ears. It remains unknown how altered function of Shh relates to morphogenetic changes that coincide with the evolution of limbs and novel auditory organs in the ear. In this study, we used the tetrapod, Xenopus laevis, to test how increasing concentrations of the Shh signal pathway antagonist, Vismodegib, affects ear development. Vismodegib treatment dose dependently alters the development of the ear, hypaxial muscle, and indirectly the Mauthner cell through its interaction with the inner ear afferents. Together, these phenotypes have an effect on escape response. The altered Mauthner cell likely contributes to the increased time to respond to a stimulus. In addition, the increased hypaxial muscle in the trunk likely contributes to the subtle change in animal C-start flexion angle. In the ear, Vismodegib treatment results in decreasing segregation between the gravistatic sensory epithelia as the concentration of Vismodegib increases. Furthermore, at higher doses, there is a loss of the horizontal canal but no enantiomorphic transformation, as in bony fish lacking Shh. Like in amniotes, Shh signaling in frogs affects dorsoventral patterning in the ear, suggesting that auditory sensory evolution in sarcopterygians/tetrapods evolved with a shift of Shh function in axis specification. © 2017 Wiley Periodicals, Inc. Develop Neurobiol 77: 1385-1400, 2017.

Sonic hedgehog antagonists reduce size and alter patterning of the frog inner ear.

Delirium is common and dangerous among elderly inpatients; yet, it is underdiagnosed and thus undertreated. This study aimed to test the diagnostic characteristics of a noninvasive point-of-care device with two-channel (bispectral) electroencephalography (EEG) for the screening of delirium in the hospital.

Delirium detection by a novel bispectral electroencephalography device in general hospital.

Multiple sclerosis (MS) is a demyelinating disease of the central nervous system. We and others have shown that there is enrichment or depletion of some gut bacteria in MS patients compared to healthy controls (HC), suggesting an important role of the gut bacteria in disease pathogenesis. Thus, specific gut bacteria that are lower in abundance in MS patients could be used as a potential treatment option for this disease. In particular, we and others have shown that MS patients have a lower abundance of  compared to HC, whereas the abundance of  is increased in patients that receive disease-modifying therapies such as Copaxone® (Glatiramer acetate-GA). This inverse correlation between the severity of MS disease and the abundance of  suggests its potential for use as a therapeutic option to treat MS. Notably we have previously identified a specific strain (), that suppresses disease in the animal model of MS, experimental autoimmune encephalomyelitis (EAE) compared with sham treatment. In the present study we analyzed whether the disease suppressing effects of  synergize with those of the disease-modifying drug Copaxone® to more effectively suppress disease compared to either treatment alone. Treatment with  was as effective in suppressing disease as treatment with Copaxone®, whereas the combination of  plus Copaxone® was not more effective than either individual treatment. -treated mice had an increased frequency and number of CD4FoxP3 regulatory T cells in periphery as well as gut and a decreased frequency of pro-inflammatory IFN-γ and IL17-producing CD4 T cells in the CNS, suggesting  suppresses disease by boosting anti-inflammatory immune responses and inhibiting pro-inflammatory immune responses. In conclusion, our study indicates that the human gut commensal  can suppress disease as efficiently as Copaxone® and may provide an alternative treatment option for MS patients.

, A Human Gut Commensal, Is as Potent as COPAXONE® in an Animal Model of Multiple Sclerosis.

Delirium is common and dangerous, yet underdetected and undertreated. Current screening questionnaires are subjective and ineffectively implemented in busy hospital workflows. Electroencephalography (EEG) can objectively detect the diffuse slowing characteristic of delirium, but it is not suitable for high-throughput screening due to size, cost, and the expertise required for lead placement and interpretation. This study hypothesized that an efficient and reliable point-of-care EEG device for high-throughput screening could be developed.

Kasra Zarei

Medical Scientist Training Program

University of Iowa

Microbiota Analysis Using Two-step PCR and Next-generation 16S rRNA Gene Sequencing