Thermal stability and inactivation of hepatitis C virus grown in cell culture

<p>Abstract</p> <p>Background</p> <p>Hepatitis C virus (HCV) is a blood-borne flavivirus that infects many millions of people worldwide. Relatively little is known, however, concerning the stability of HCV and reliable procedures for inactivating this virus.</p> <p>Methods</p> <p>In the current study, the thermostability of cell culture-derived HCV (HCVcc, JFH-1 strain) under different environmental temperatures (37°C, room temperature, and 4°C) and the ability of heat, UVC light irradiation, and aldehyde and detergent treatments to inactivate HCVcc were evaluated. The infectious titers of treated viral samples were determined by focus-forming unit (FFU) assay using an indirect immunofluorescence assay for HCV NS3 in hepatoma Huh7-25-CD81 cells highly permissive for HCVcc infection. MTT cytotoxicity assay was performed to determine the concentrations of aldehydes or detergents at which they were no longer cytotoxic.</p> <p>Results</p> <p>HCVcc in culture medium was found to survive 37°C and room temperature (RT, 25 ± 2°C) for 2 and 16 days, respectively, while the virus was relatively stable at 4°C without drastic loss of infectivity for at least 6 weeks. HCVcc in culture medium was sensitive to heat and could be inactivated in 8 and 4 min when incubated at 60°C and 65°C, respectively. However, at 56°C, 40 min were required to eliminate HCVcc infectivity. Addition of normal human serum to HCVcc did not significantly alter viral stability at RT or its susceptibility to heat. UVC light irradiation (wavelength = 253.7 nm) with an intensity of 450 μW/cm²efficiently inactivated HCVcc within 2 min. Exposures to formaldehyde, glutaraldehyde, ionic or nonionic detergents all destroyed HCVcc infectivity effectively, regardless of whether the treatments were conducted in the presence of cell culture medium or human serum.</p> <p>Conclusions</p> <p>The results provide quantitative evidence for the potential use of a variety of approaches for inactivating HCV. The ability of HCVcc to survive ambient temperatures warrants precautions in handling and disposing of objects and materials that may have been contaminated with HCV.</p

Recombination-mediated escape from primary CD8+ T cells in acute HIV-1 infection

Abstract Background A major immune evasion mechanism of HIV-1 is the accumulation of non-synonymous mutations in and around T cell epitopes, resulting in loss of T cell recognition and virus escape. Results Here we analyze primary CD8+ T cell responses and virus escape in a HLA B*81 expressing subject who was infected with two T/F viruses from a single donor. In addition to classic escape through non-synonymous mutation/s, we also observed rapid selection of multiple recombinant viruses that conferred escape from T cells specific for two epitopes in Nef. Conclusions Our study shows that recombination between multiple T/F viruses provide greater options for acute escape from CD8+ T cell responses than seen in cases of single T/F virus infection. This process may contribute to the rapid disease progression in patients infected by multiple T/F viruses

Cooperation of B Cell Lineages in Induction of HIV-1-Broadly Neutralizing Antibodies

Development of strategies for induction of HIV-1 broadly neutralizing antibodies (bnAbs) by vaccines is a priority. Determining the steps of bnAb induction in HIV-1-infected individuals who make bnAbs is a key strategy for immunogen design. Here we study the B cell response in a bnAb-producing individual, and report cooperation between two B cell lineages to drive bnAb development. We isolated an autologous virus-neutralizing antibody lineage that targeted an envelope region (loop D) and selected virus escape mutants that resulted in both enhanced bnAb lineage envelope binding and escape mutant neutralization—traits associated with increased B cell antigen drive. Thus, in this individual, two B cell lineages cooperated to induce the development of bnAbs. Design of vaccine immunogens that simultaneously drive both autologous and broadly neutralizing B cell lineages may be important for vaccine-induced recapitulation of events that transpire during the maturation of neutralizing antibodies in HIV-1-infected individuals

Fatigue Performance Prediction of RC Beams Based on Optimized Machine Learning Technology

The development of fatigue damage in reinforced concrete (RC) beams is affected by various factors such as repetitive loads and material properties, and there exists a complex nonlinear mapping relationship between their fatigue performance and each factor. To this end, a fatigue performance prediction model for RC beams was proposed based on the deep belief network (DBN) optimized by particle swarm optimization (PSO). The original database of fatigue loading tests was established by conducting fatigue loading tests on RC beams. The mid-span deflection, reinforcement strain, and concrete strain during fatigue loading of RC beams were predicted and evaluated. The fatigue performance prediction results of the RC beam based on the PSO-DBN model were compared with those of the single DBN model and the BP model. The models were evaluated using the R2 coefficient, mean absolute percentage error, mean absolute error, and root mean square error. The results showed that the fatigue performance prediction model of RC beams based on PSO-DBN is more accurate and efficient

Design, Fabrication, and Testing of a Monolithically Integrated Tri-Axis High-Shock Accelerometer in Single (111)-Silicon Wafer

In this paper, a monolithic tri-axis piezoresistive high-shock accelerometer has been proposed that has been single-sided fabricated in a single (111)-silicon wafer. A single-cantilever structure and two dual-cantilever structures are designed and micromachined in one (111)-silicon chip to detect Z-axis and X-/Y-axis high-shock accelerations, respectively. Unlike the previous tri-axis sensors where the X-/Y-axis structure was different from the Z-axis one, the herein used similar cantilever sensing structures for tri-axis sensing facilitates design of uniform performance among the three elements for different sensing axes and simplifies micro-fabrication for the multi-axis sensing structure. Attributed to the tri-axis sensors formed by using the single-wafer single-sided fabrication process, the sensor is mechanically robust enough to endure the harsh high-g shocking environment and can be compatibly batch-fabricated in standard semiconductor foundries. After the single-sided process to form the sensor, the untouched chip backside facilitates simple and reliable die-bond packaging. The high-shock testing results of the fabricated sensor show linear sensing outputs along X-/Y-axis and Z-axis, with the sensitivities (under DC 5 V supply) as about 0.80&#8211;0.88 &#956;V/g and 1.36 &#956;V/g, respectively. Being advantageous in single-chip compact integration of the tri-axis accelerometers, the proposed monolithic tri-axis sensors are promising to be embedded into detection micro-systems for high-shock measurement applications

Application of PVC pipes as an adjustable bilateral traction device in lower limb fractures

Abstract Objective To introduce a new type of simple adjustable bilateral bidirectional polyvinyl chloride (PVC) tube traction device and discuss the value of using this device before surgery in patients with lower limb fractures. Methods To introduce the manufacturing process of an adjustable bilateral traction device made of PVC pipes. From August 2018 to November 2019, the data of 36 patients with lower limb fractures who were treated with this traction device were retrospectively analysed. The treatment outcomes were analysed, including length of both lower limbs, fracture reduction, lower limb mobility, visual analogue scale (VAS) score, incidence of complications, and patient satisfaction. Results All patients were able to move the affected limb immediately after using the device. The patient's pain was significantly reduced, they were able to turn over freely during bed rest, and the length of the affected limb was restored to that of the healthy limb. Thirty-four (94.5%) patients were satisfied with the reduction of the fracture end, 2 (5.5%) patients with tibiofibular fractures showed angular displacement of the fractured end and satisfactory reduction after the position of the bone traction needle was adjusted; 7 (19.5%) patients developed deep vein thrombosis of the affected lower limb during traction; there was no decubitus or vascular nerve injury, and the overall complication rate was 25% (9/36). All the patients and their families were satisfied with the results of this treatment. Conclusion The aim of this study is to introduce a new type of traction device. It is advantageous in that it is light weight, low cost, easy to assemble, promotes immediate movement of the affected limb after assembly, improves patient comfort and can be used with a titanium steel needle for MRI examination under traction. In the clinical setting, it has been shown to be suitable for the temporary treatment of patients with lower leg fractures prior to surgery, particularly patients who, for various reasons, require nonsurgical treatment in the short term

Recombination frequencies at different conditions during PCR.

<p>(A) Recombination frequencies were determined at different thermal cycles. Equal amount of NL4-3 and 89.6 plasmids (10⁷ copies per template) were mixed together and co-amplified. The PCR was carried with 5, 10, 15, 20, 25 or 30 thermal cycles. (B) Recombination frequencies were determined with different numbers of templates. Equal amount of NL4-3 and 89.6 plasmids (10¹, 10³, 10⁵ or 10⁷ copies each) was mixed together and co-amplified by 30 cycles of PCR. (C) Recombination frequencies were determined with different extension time. Equal amount of NL4-3 and 89.6 plasmids (10⁷ copies per template) were mixed together and co-amplified. The PCR was carried with different extension time (1, 2, 4 or 8 minutes). The PCR products were analyzed by the PASS assay and the recombination frequency at each condition was determined by linkage analysis of six bases.</p

Recombination analysis of two low genetic diversity templates during PCR.

<p>A partial <i>pol</i> gene (870 bp) was amplified from two genetic variants (1B7 and 1D1) of WEAU. Nucleotides that are distinct at six positions in 1B7 and 1D1 are shown. The regions between two neighbor nucleotides are named as A through E and the genetic distances between them are indicated.</p