Muscle Transcriptome Sequencing Revealed Thermal Stress-Responsive Regulatory Genes in Farmed Rohu, Labeo rohita (Hamilton, 1822).

Rohu, Labeo rohita, is one of the most important aquaculture species in the Indian subcontinent. Understanding the molecular-level physiological responses to thermal stress or climate change is essential. In the present work, transcriptome sequencing was carried out in the muscle tissue of the rohu in response to heat stress (35 °C) in comparison with the control (28 °C). A total of 125 Gb of sequence data was generated, and the raw-reads were filtered and trimmed, which resulted in 484 million quality reads. Reference-based assembly of reads was performed using L. rohita genome, and a total of 90.17% of reads were successfully mapped. A total of 37,462 contigs were assembled with an N50 value of 1854. The differential expression analysis revealed a total of 107 differentially expressed genes (DEGs) (15 up-, 37 down-, and 55 neutrally regulated) as compared to the control group (Log2FC > 2, P < 0.05). Gene enrichment analysis of DEGs indicates that transcripts were associated with molecular, biological, and cellular activities. The randomly selected differentially expressed transcripts were validated by RT-qPCR and found consistent expression patterns in line with the RNA-seq data. Several transcripts such as SERPINE1(HSP47), HSP70, HSP90alpha, Rano class II histocompatibility A beta, PGC-1 and ERR-induced regulator, proto-oncogene c-Fos, myozenin2, alpha-crystallin B chain-like protein, angiopoietin-like protein 8, and acetyl-CoA carboxylases have been identified in muscle tissue of rohu that are associated with stress/immunity. This study identified the key biomarker SERPINE1 (HSP47), which showed significant upregulation (~ 2- to threefold) in muscle tissue of rohu exposed to high temperature. This study can pave a path for the identification of stress-responsive biomarkers linked with thermal adaptations in the farmed carps.

Approaches to the Maxillofacial Skeleton: Application of Standard and Minimally Invasive Techniques.

Management of maxillofacial trauma is complex and challenging and requires a clear understanding of facial anatomy and function. There are multiple approaches that can be used to access each anatomical region, each with specific indications and complication profiles. Open, "invasive" approaches are being replaced or augmented with minimally invasive and endoscopic approaches when possible. Thorough knowledge of indications, surgical techniques, and potential complications allows surgeons to make appropriate decisions for access and repair of fractures. This article is a comprehensive review of standard and minimally invasive approaches, with description of techniques and pros and cons for their use.

Modulation of heat shock protein 70 (HSP70) gene expression ex vivo in response to heat stress in chicken.

The present study was conducted to assess the effect of exposure to heat stress on the HSP70 gene expression pattern ex vivo in chickens. The adult healthy birds (n = 15) grouped into three replicates of n = 5 each were used to isolate peripheral blood mononuclear cells (PBMCs). The PBMCs were subjected to heat stress at 42 °C for 1 h, and cells without heat stress (NHS) were taken as control. The cells were seeded in 24 well plates and incubated in a humidified incubator at 37 °C under 5% CO2 for recovery. HSP70 expression kinetics were evaluated at 0, 2, 4, 6, and 8 h of the recovery period. Compared with NHS, the expression pattern of HSP70 was upregulated gradually from 0 to 4 h with peak (p < 0.05) expression recorded at 4 h of recovery time. mRNA expression of HSP70 escalated in a time-dependent manner from 0 to 4 h of heat exposure and thereafter exhibited a gradually decreasing pattern till 8 h of the recovery period. The findings from this study highlight the protective role of HSP70 against the deleterious effects of heat stress in chicken PBMCs. Further, the study demonstrates the possible use of PBMCs as a cellular system in assessing the heat stress effect in chickens ex vivo.

Virtual Surgery for the Nasal Airway: A Preliminary Report on Decision Support and Technology Acceptance.

IMPORTANCE: Nasal airway obstruction (NAO) is a common problem that affects patient quality of life. Surgical success for NAO correction is variable. Virtual surgery planning via computational fluid dynamics (CFD) has the potential to improve the success rates of NAO surgery. OBJECTIVE: To elicit surgeon feedback of a virtual surgery planning tool for NAO and to determine if this tool affects surgeon decision making. DESIGN, SETTING, AND PARTICIPANTS: For this cross-sectional study, 60-minute face-to-face interviews with board-certified otolaryngologists were conducted at a single academic otolaryngology department from September 16, 2016, through October 7, 2016. Virtual surgery methods were introduced, and surgeons were able to interact with the virtual surgery planning tool interface. Surgeons were provided with a patient case of NAO, and open feedback of the platform was obtained, with emphasis on surgical decision making. MAIN OUTCOMES AND MEASURES: Likert scale responses and qualitative feedback were collected for the virtual surgery planning tool and its influence on surgeon decision making. RESULTS: Our 9 study participants were all male, board-certified otolaryngologists with a mean (range) 15 (4-28) number of years in practice and a mean (range) number of nasal surgeries per month at 2.2 (0.0-6.0). When examined on a scale of 1 (not at all) to 5 (completely), surgeon mean (SD) score was 3.4 (0.5) for how realistic the virtual models were compared with actual surgery. On the same scale, when asked how much the virtual surgery planning tool changed surgeon decision making, mean (SD) score was 2.6 (1.6). On a scale of 1 (strongly disagree) to 7 (strongly agree), surgeon scores for perceived usefulness of the technology and attitude toward using it were 5.1 (1.1) and 5.7 (0.9), respectively. CONCLUSIONS AND RELEVANCE: Our study shows positive surgeon experience with a virtual surgery planning tool for NAO based on CFD simulations. Surgeons felt that future applications and areas of study of the virtual surgery planning tool include its potential role for patient counseling, selecting appropriate surgical candidates, and identifying which anatomical structures should be targeted for surgical correction. LEVEL OF EVIDENCE: NA.

Advances in Technology for Functional Rhinoplasty: The Next Frontier.

Advances in computer modeling and simulation technologies have the potential to provide facial plastic surgeons with information and tools that can aid in patient-specific surgical planning for rhinoplasty. Finite element modeling and computational fluid dynamics are modeling technologies that have been applied to the nose to study structural biomechanics and nasal airflow. Combining these technologies with patient-specific imaging data and symptom measures has the potential to alter the future landscape of nasal surgery.

EvaGreen-based Multiplex Real-time PCR Assay for Rapid Differentiation of Wild-Type and Glycoprotein E-Deleted Bovine Herpesvirus-1 Strains.

Bovine herpesvirus-1 (BoHV-1) is an important viral pathogen causing significant economic losses to the cattle industry. Glycoprotein E-deleted marker vaccines form the basis for BoHV-1 control programs widely, wherein detection and differentiation of wild-type and gE-deleted vaccine strains is of crucial importance for proper disease management. In the present study, we report an EvaGreen-based multiplex real-time polymerase chain reaction (EGRT-PCR) assay for rapid differentiation of wild-type and glycoprotein E-deleted strains of BoHV-1. The EGRT-PCR assay could simultaneously detect two viral genes (glycoprotein B and E) and an internal positive control gene (bovine growth hormone- bGH), in a single-tube reaction. The analytical sensitivity of the EGRT-PCR assay was as little as 10 copies of the BoHV-1 DNA per reaction. The modified real-time PCR assay could successfully differentiate wild-type and gE-deleted BoHV-1 strains based on gene specific melting temperatures (Tm) peaks. Our results have shown that the EGRT-PCR developed in this study might prove to be a promising tool in disease management by enabling rapid differentiation of wild-type and gE-deleted strains of BoHV-1.

Correlation of Nasal Mucosal Temperature With Subjective Nasal Patency in Healthy Individuals.

IMPORTANCE: Historically, otolaryngologists have focused on nasal resistance to airflow and minimum airspace cross-sectional area as objective measures of nasal obstruction using methods such as rhinomanometry and acoustic rhinometry. However, subjective sensation of nasal patency may be more associated with activation of cold receptors by inspired air than with respiratory effort. OBJECTIVE: To investigate whether subjective nasal patency correlates with nasal mucosal temperature in healthy individuals. DESIGN, SETTING, AND PARTICIPANTS: Healthy adult volunteers first completed the Nasal Obstruction Symptom Evaluation (NOSE) and a unilateral visual analog scale to quantify subjective nasal patency. A miniaturized thermocouple sensor was then used to record nasal mucosal temperature bilaterally in 2 locations along the nasal septum: at the vestibule and across from the inferior turbinate head. MAIN OUTCOMES AND MEASURES: Nasal mucosal temperature and subjective patency scores in healthy individuals. RESULTS: The 22 healthy adult volunteers (12 [55%] male; mean [SD] age, 28.3 [7.0] years) had a mean (SD) NOSE score of 5.9 (8.4) (range, 0-30) and unilateral VAS score of 1.2 (1.4) (range, 0-5). The range of temperature oscillations during the breathing cycle, defined as the difference between end-expiratory and end-inspiratory temperatures, was greater during deep breaths (mean [SD] change in temperature, 6.2°C [2.6°C]) than during resting breathing (mean [SD] change in temperature, 4.2°C [2.3°C]) in both locations (P < .001). Mucosal temperature measured at the right vestibule had a statistically significant correlation with both right-side visual analog scale score (Pearson r = -0.55; 95% CI, -0.79 to -0.17; P = .008) and NOSE score (Pearson r = -0.47; 95% CI, -0.74 to -0.06; P = .03). No other statistically significant correlations were found between mucosal temperature and subjective nasal patency scores. Nasal mucosal temperature was lower (mean of 1.5°C lower) in the first cavity to be measured, which was the right cavity in all participants. CONCLUSIONS AND RELEVANCE: The greater mucosal temperature oscillations during deep breathing are consistent with the common experience that airflow sensation is enhanced during deep breaths, thus supporting the hypothesis that mucosal cooling plays a central role in nasal airflow sensation. A possible correlation was found between subjective nasal patency scores and nasal mucosal temperature, but our results were inconsistent. The higher temperature in the left cavity suggests that the sensor irritated the nasal mucosa, affecting the correlation between patency scores and mucosal temperature. Future studies should consider noncontact temperature sensors to prevent mucosa irritation. LEVEL OF EVIDENCE: NA.

Loop-Mediated Isothermal Amplification for Rapid Detection and Differentiation of Wild-Type Bovine Herpesvirus-1 and Glycoprotein E-Deleted Marker Vaccine Strain.

Bovine herpesvirus-1 (BoHV-1) is an important viral pathogen affecting cattle and causing numerous reproductive disorders leading to significant economic losses to the cattle industry. The control programs for BoHV-1 are widely based on the use of glycoprotein E-deleted marker vaccines, wherein detection and differentiation of wild-type and gE-deleted vaccine strains is of crucial importance for proper disease management. In this study, we report rapid and simple loop-mediated isothermal amplification (LAMP) assays for detection and differentiation of gE-deleted BoHV-1 from wild-type virus under isothermal conditions. The assays could be completed in 90 mintes, including viral DNA isolation, target amplification and visual interpretation of results with naked eye. The analytical sensitivity of the assays was 10 times higher than conventional PCR and could detect as little as 100 fg of viral DNA per reaction. The applicability of LAMP for detection of BoHV-1 in bovine semen was assessed by testing semen samples collected from breeding bulls and compared with TaqMan real-time PCR (as gold standard). The LAMP assays had diagnostic specificity of 100%. The diagnostic sensitivity was 88.2% and 83.3% for gB- and gE-LAMP, respectively, when compared with TaqMan real-time PCR. Our results have shown that the LAMP method developed in this study is a potential tool for rapid, sensitive, specific, cost-effective, and user-friendly detection and differentiation of wild type BoHV-1 from gE-deleted marker vaccine.

Treatment of Prominent Ears and Otoplasty: A Contemporary Review.

Prominent ears affect approximately 5% of the population and can have a significant psychological impact on patients. A wide variety of otoplasty techniques have been described, all sharing the goal of re-creating the normal appearance of the ear and achieving symmetry between the 2 sides. Recent trends in otoplasty techniques have consistently moved toward less invasive options, ranging from nonsurgical newborn ear molding to cartilage-sparing surgical techniques and even incisionless, office-based procedures. Herein, we review anatomy of the external ear, patient evaluation, the evolution of nonsurgical and surgical otoplasty techniques, otoplasty outcomes, and future trends for treatment of prominent ears.

Reconstructive and rehabilitation challenges following a cranio-orbital gunshot wound.

We present a case of a 26-year-old man who sustained a close-range gunshot wound to the head. His injuries included significant left orbital injury resulting in a ruptured, blind eye and severely comminuted fractures of the left orbital roof, superior and inferior orbital rims, and orbital floor. Associated injuries included left frontal lobe injury, anterior and posterior table fractures of the left frontal sinus, and a comminuted left zygomaticomaxillary complex fracture. We employed an interdisciplinary surgical approach with collaboration among the Otolaryngology, Neurosurgery, and Oculoplastic Surgery services performed in two stages. Management of such extensive craniofacial injuries can be challenging and requires a coordinated, interdisciplinary approach.

Development of a SYBR Green I based duplex real-time PCR for detection of bovine herpesvirus-1 in semen.

Bovine herpesvirus-1 (BoHV-1) is a viral pathogen found in infected bull semen, which is transmitted to inseminated cows by artificial insemination. BoHV-1 infection can cause reproductive disorders leading to significant economic loss to cattle industry. To detect BoHV-1 in semen, in this study, a SYBR Green I based duplex real-time PCR was developed. The assay included primers from BoHV-1 glycoprotein C (gC) and bovine growth hormone (bGH) genes for simultaneous detection in single tube. The result was interpreted by analysing melting temperature (Tm) peaks obtained after melt curve analysis of the amplified products at the end of reaction. The Tm peaks for BoHV-1-gC indicated presence of BoHV-1 while the bGH peak indicated reaction without inhibition. The sensitivity of the assay was to detect ten BoHV-1 genome copies per reaction. The analytical sensitivity was to detect 0.21 TCID50 infectious BoHV-1 in spiked semen. The assay was validated with 80 semen samples collected from breeding bulls. The diagnostic sensitivity and specificity of the assay was 100% with OIE recommended TaqMan probe based real-time PCR.

Frontal sinus and naso-orbital-ethmoid fractures.

IMPORTANCE: Frontal sinus and naso-orbital-ethmoid (NOE) fractures are among the most challenging injuries in the treatment of maxillofacial trauma. OBJECTIVE: To summarize the current knowledge regarding frontal sinus and NOE fractures and to present some of the more recent, evidence-based literature to support current treatment recommendations. EVIDENCE REVIEW: A PubMed search of articles from 1990 through 2013 was performed. Search terms included frontal sinus fracture, NOE fracture, naso-orbito-ethmoid fracture, naso-ethmoid-orbital fracture, and nasoethmoid fracture. FINDINGS: Advances in sophisticated imaging and evolution in minimally invasive surgical techniques are introducing more conservative options that may provide better patient outcomes while minimizing the risks and morbidity associated with more traditional treatment approaches. CONCLUSIONS AND RELEVANCE: The treatment of frontal sinus and NOE fractures is challenging, given the complex anatomy and associated pattern of injuries. Traditional treatment paradigms are evolving and support the role of more conservative treatment algorithms in selected patients.

Rapid detection of bovine herpesvirus 1 in bovine semen by loop-mediated isothermal amplification (LAMP) assay.

Bovine herpesvirus 1 (BoHV-1) is the most common viral pathogen found in bovine semen, causing numerous reproductive disorders leading to economic losses to the cattle industry. For rapid detection of BoHV-1 in bovine semen, in this study, we applied a loop-mediated isothermal amplification (LAMP) assay. The assay could be completed within 90 min, including total DNA isolation, target amplification, and visual interpretation of positive or negative results with the naked eye. The assay detected as little as 10 fg of BoHV-1 DNA per reaction. The analytical sensitivity of the assay was 0.2 TCID50 BoHV-1 per reaction, which was 100 times more sensitive than conventional PCR and comparable to TaqMan real-time PCR. The applicability of the assay was assessed by analysing 118 semen samples collected from breeding bulls. On comparison with TaqMan real-time PCR, the LAMP assay had a diagnostic sensitivity of 97 %, specificity of 100 %, and accuracy of 99.2 % for detection of BoHV-1 in bovine semen. The LAMP assay developed in this study is a rapid, sensitive, and cost-effective alternative for detection of BoHV-1 in bovine semen.

Secondary cleft rhinoplasty.

The cleft nasal deformity seen in patients with unilateral and bilateral cleft lip presents a formidable challenge for the facial plastic surgeon. The underlying anatomic deformities combined with scarring from previous procedures make secondary cleft rhinoplasty a difficult procedure for even the most experienced surgeons. Numerous techniques for secondary cleft rhinoplasty have been described in the literature over the past several decades, yet the lack of wide adoption of any given technique highlights the great variability seen with this problem. Regardless, the fundamental goals of achieving nasal symmetry with definition of the nasal base and tip, correction of nasal airway obstruction, and repair of nasal scarring or webbing have driven the progressive evolution of techniques developed to correct various aspects of the cleft nasal deformity. Despite the number of techniques that have been published, very few studies have looked specifically at outcomes in secondary cleft rhinoplasty, and further work is needed in this area. In this article, we will review anatomy of the cleft nasal deformity, repair strategies and timing, surgical techniques for both unilateral and bilateral cleft nasal deformity, and outcomes for secondary cleft rhinoplasty.

Protection of mice against lethal rabies virus challenge using short interfering RNAs (siRNAs) delivered through lentiviral vector.

The antiviral potential of small interfering RNAs (siRNAs) targeting rabies virus (RV) polymerase (L) and nucleoprotein (N) genes delivered through lentiviral vector was investigated. For in vitro evaluation, siRNAs expressing BHK-21 cell lines (BHK-L and BHK-N) were developed using transduction with Lenti-L and Lenti-N lentiviruses encoding siRNAs against RV-L and N genes, respectively. When these cell lines were challenged in vitro with RV Pasteur virus-11 (PV-11) strain, there was reduction in number of RV-specific foci and target gene transcripts indicating inhibitory effect on RV multiplication. For in vivo evaluation, mice were treated intracerebrally with lentiviruses and challenged with 20 LD50 of RV challenge virus standard-11 (CVS-11) strain by intramuscular route in masseter muscle. Five out of eight mice treated with Lenti-N survived indicating 62.5 % protection. The control and Lenti-L-treated mice died within 7-10 days indicating lethal nature of challenge virus and no protection. These results demonstrated that siRNA targeting RV-N could not only inhibit RV multiplication, but also conferred protection in mice against lethal RV challenge. These findings have implication on therapeutic use of siRNA targeting RV-N against RV infection.

Evaluation of single and dual siRNAs targeting rabies virus glycoprotein and nucleoprotein genes for inhibition of virus multiplication in vitro.

Small interfering RNAs (siRNAs) targeting rabies virus (RV) glycoprotein (G) and nucleoprotein (N) genes were evaluated as antiviral agents against rabies virus in vitro in BHK-21 cells. To select effective siRNAs targeting RV-G, a plasmid-based transient co-transfection approach was used. In this, siRNAs were expressed as short hairpin RNAs (shRNAs), and their ability to inhibit RV-G gene expression was evaluated in cells transfected with a plasmid expressing RV-G. The nine different siRNAs designed to target RV-G exhibited varying degrees of knockdown of RV-G gene expression. One siRNA (si-G7) with considerable effect in knockdown of RV-G expression also demonstrated significant inhibition of RV multiplication in BHK-21 cells after in vitro challenge with the RV Pasteur virus-11 (PV-11) strain. A decrease in the number of fluorescent foci in siRNA-treated cells and a reduction (86.8 %) in the release of RV into infected cell culture supernatant indicated the anti-rabies potential of siRNA. Similarly, treatment with one siRNA targeting RV-N resulted in a decrease in the number of fluorescent foci and a reduction (85.9 %) in the release of RV. As a dual gene silencing approach where siRNAs targeting RV-G and RV-N genes were expressed from single construct, the anti-rabies-virus effect was observed as an 87.4 % reduction in the release of RV. These results demonstrate that siRNAs targeting RV-G and N, both in single and dual form, have potential as antiviral agent against rabies.

Updates in forehead flap reconstruction of facial defects.

PURPOSE OF REVIEW: The versatile and reliable paramedian forehead flap is the workhorse interpolated flap for nasal reconstruction and its use has been expanded to reconstruct other areas of the face. The goal of this article is to review the recent research describing the modern concepts and techniques in forehead flap reconstruction of facial defects. RECENT FINDINGS: A variety of forehead flap concepts, techniques, and applications have been described in the recent literature in both nasal and facial reconstruction. Recent concepts include the phenomenon of vascular delay as well as more detailed study of the perfusion of the forehead flap at its various stages. Novel techniques include various approaches in staged reconstruction, adoption of the forehead flap for reconstruction of internal lining defects, and flap thinning. SUMMARY: Although basic principles of the forehead flap have remained unchanged over the years, specific techniques and applications continue to be refined. The forehead flap continues to be a mainstay in the armamentarium of facial plastic and reconstructive surgeons.