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Interactions between the microbiota and their colonized environments mediate critical pathways from biogeochemical cycles to homeostasis in human health. Here we report a soil-inspired chemical system that consists of nanostructured minerals, starch granules and liquid metals. Fabricated via a bottom-up synthesis, the soil-inspired chemical system can enable chemical redistribution and modulation of microbial communities. We characterize the composite, confirming its structural similarity to the soil, with three-dimensional X-ray fluorescence and ptychographic tomography and electron microscopy imaging. We also demonstrate that post-synthetic modifications formed by laser irradiation led to chemical heterogeneities from the atomic to the macroscopic level. The soil-inspired material possesses chemical, optical and mechanical responsiveness to yield write-erase functions in electrical performance. The composite can also enhance microbial culture/biofilm growth and biofuel production in vitro. Finally, we show that the soil-inspired system enriches gut bacteria diversity, rectifies tetracycline-induced gut microbiome dysbiosis and ameliorates dextran sulfate sodium-induced rodent colitis symptoms within in vivo rodent models.

Integrating stimuli-responsive materials into energy storage technologies opens a new approach to introduce novel functionalities as well as addressing some of the crucial and unresolved issues. As an example, current flexible lithium-ion batteries (LIBs) still suffer from a fragile or susceptible nature and limited electrochemical performance recovery against severe or repeated mechanical deformations. Thus, integrating a stimuli-responsive material with strong ability to recover from severe and repeated mechanical deformations into the flexible LIBs can be an effective approach to address the shape and performance recovery issue concerning the flexible LIBs [1-3]. Herein, we design and synthesize a solid polymer electrolyte (SPE) based on a shape memory polymer (SMP) to integrate into flexible lithium batteries for smart applications, for example, shape recovery from severe mechanical deformation. SMPs are attractive class of programmable, stimuli-responsive polymer materials demonstrating shape-memory behavior. A SMP has the ability to memorize its original or permanent shape; deform, fix into a temporary or secondary shape; and recover its original form via applying an external stimulus, for example, heat, magnetic or electrical field,light [3]. In fact, when mechanical deformation occurs, batteries typically suffer from reduced power, efficiency, capacity and recovery from the deformation without sacrificing the electrochemical performance would be highly desirable. The shape-memory SPE is made based on a cross-linkable polyethylene oxide (PEO) with controlled crystallinity and the ability to indue shape memory behavior above the melting point. PEO is chosen as a model shape-memory polymer due to its unique properties including low-cost, high dielectric constant and Li+-ion solvating ability, high electrochemical and mechanical stability, semi-crystalline nature, and ability to introduce cross-linkable functionalities. In response to a temperature exceeding the melting point of the polymer matrix (~70 oC), the engineered shape memory SPE can recover its original shape and size from mechanical deformations, for example, bending and folding. The all-solid-state shape-memory SPEs also show excellent electrochemical performance in Li/Li and Li/LFP cells at ambient temperature. The Li/LFP cell made using the shape memory SPE delivers a specific capacity ~140 mAh g‒1 with ~92% capacity retention after 100 cycles at 0.2C charge/discharge rate and ~99.85% Coulombic efficiency. Besides excellent electrochemical performance, we demonstrate that a flexible Li based battery made using the shape memory SPE can recover from severe mechanical deformations, for example, bending and folding, upon applying heat. This proof-of-concept study opens up a new approach to design and integrate smart functionalities into energy storage technologies. Reference [1] Z. Fang, J. Wang, H. Wu, Q. Li, S. Fan, J. Wang, J. Power Sources 454, 227932 (2020). [2] M. Koo, K. Park, S. H. Lee, M. Suh, D. Y. Jeon, J. W. Choi, K. Kang, K. J. Lee, Nano Lett. 12, 4810–4816 (2012). [3] V. Jabbari, V. Yurkiv, M. G. Rasul, M. Cheng, P. Griffin, F. Mashayek, R. Shahbazian-Yassar, Small 2021, 2102666, DOI: 10.1002/smll.202102666.

The simulation of the complex flow in a wind turbine wake is a challenging problem. To date, much of the research has been inhibited by both the time and computational costs associated with turbulence modelling. Additionally, the majority of numerical investigations focus on turbine performance and therefore neglect the near wake of a Horizontal Axis Wind Turbine (HAWT) entirely. This investigation focuses on experimentally and numerically quantifying the near wake structure of a model HAWT. The Shear Stress Transport (SST) k − ω , Elliptical-Blending Reynolds Stress Model (EB-RSM) and the Reynolds Stress Transport (RST) turbulence models were used to model a turbine wake in the current study, with the results verified against experimental hot-wire data. Near wake velocity and turbulence characteristics were investigated to determine if low-order models can accurately predict the magnitude and distribution of velocity and turbulence values in the near wake of a model HAWT. The HAWT model was operated at two TSR values of 2.54 and 3.87. All models predicted velocity deficit values to within 2–4% and 4–7% of experimental results for TSR values of 2.54 and 3.87 respectively. Results showed that all models were able to accurately predict the mean velocity deficit generated in the near wake. All models were able to predict the fluctuating u and v velocity components in the near wake to the correct order of magnitude with the fluctuating velocity components having an inverse Laplace distribution in the wake. However, all models under-estimated the magnitude of these velocity values with predictions as low as −43% of experimental results.

A review of wind turbine aerodynamics research is presented. The review is limited to Horizontal Axis Wind Turbine (HAWT) investigations. The focus is on recent near wake experiments, wake predictions by commercial CFD codes and current FSI and structural modelling attempts. For the near wake, the review considers experiments carried out in controlled conditions whereby the incoming freestream is perpendicular to the rotor plane. Additional anomalies such as wind shear, gusts and yaw transition are not considered. The survey of 3D commercial codes is also focused on HAWT models in parallel flow conditions. Finally, the structural models reviewed are divided into two separate categories: 1) blade deflection and performance under aerodynamic loads, and 2) the vibrational response of blades under aerodynamic loading. The aim is to highlight common trends within near wake experiments and investigate both CFD and FE modelling strategies – to identify current limitations and future opportunities within the sector.

l-lysine (Lys) is an essential amino acid that is added to foods and dietary supplements. Lys may interact with mineral nutrients and affect their metabolism. This study examined the effect of dietary Lys supplementation on the bioavailability of copper (Cu) and iron (Fe). Weanling male Sprague-Dawley rats were fed one of five diets (20% casein) for 4 weeks containing normal Cu and Fe (control) or low Cu or Fe without (LCu, LFe) or with (LCu+Lys, LFe+Lys) addition of 1.5% Lys. Final body weights, body weight gains and food consumption of the rats did not differ (P≥0.05) among diet groups. Rats fed the low Cu or Fe diets showed changes in nutritional biomarkers compared to control rats, demonstrating reduced Cu and Fe status, respectively. Hematological parameters, serum ceruloplasmin activity and Cu and Fe concentrations in serum, liver, kidney and intestinal mucosa were unaffected (P≥0.05) by Lys supplementation. These results indicate that in the context of an adequate protein diet, Lys supplementation at a relatively high level does not affect Cu or Fe bioavailability in rats.

In this study, the near-field (up to three chord lengths) development of a wing-tip vortex is numerically investigated at two angles of attack (five and ten degrees). The application of a vorticity confinement model to an Unsteady Reynolds averaged Navier–Stokes (URANS) model and Large Eddy Simulation (LES) is examined with the focus of preventing the rapid dissipation of vorticity in a wing-tip vortex. Vortex core size and trajectory were predicted well by the LES model, whereas the URANS model predicted a large vortex core, which remained constant with downstream distance. The LES model correctly predicted the jet-like axial velocity for an angle of attack of ten degrees and the LES and experimental axial velocity excess had the same value of 111% of free-stream velocity at two chord lengths downstream. The LES model predicted the turbulence in the vortex reasonably well as the maximum turbulent root mean square (rms) velocities were within 15% and 35% of experimental values at two and three chord lengths downstream for angles of attack of ten and five degrees respectively. The URANS model predicted the mean flow for an angle of attack of five degrees reasonably well but greatly under-predicted the mean flow for an angle of attack of ten degrees and the turbulence levels at all downstream locations.

The creation of an arteriovenous fistula offers a unique example of vascular remodelling and adaption. Yet, the specific factors which elicit remodelling events which determine successful maturation or failure have not been unambiguously determined. Computational fluid dynamic (CFD) simulations are increasingly been employed to investigate the interaction between local hemodynamics and remodelling and can potentially be used to assist in clinical risk assessment of maturation or failure. However, these simulations are inextricably linked to their prescribed boundary conditions and are reliant on in vivo measurements of flow and pressure to ensure their validity. The study compares in vivo measurements of the pressure distribution across arteriovenous fistulae against a representative numerical model. The results of the study indicate relative agreement (error ≈ 8-10%) between the in vivo and CFD prediction of the mean pressure drop across the AVFs. The large pressure drop across the AVFs coincided with a palpable thrill (perivascular vibration) in vivo and fluctuations were observed in the numerical pressure drop signal due to flow instabilities arising at the anastomosis. This study provides a benchmark of the pressure distribution within an AVF and validates that CFD solutions are capable of replicating the abnormal physiological flow conditions induced by fistula creation.

The near-field (up to three chord lengths) development of a wing-tip vortex is investigated both numerically and experimentally. The research was conducted in a medium speed wind tunnel on a NACA 0012 square tip half-wing at a Reynolds number of 3.2 × 105. A full Reynolds stress turbulence model with a hybrid unstructured grid was used to compute the wing-tip vortex in the near field while an x-wire anemometer and five-hole probe recorded the experimental results. The mean flow of the computed vortex was in good agreement with experiment as the circulation parameter was within 6% of the experimental value at x/ c = 0 for α = 10° and the crossflow velocity magnitude was within 1% of the experimental value at x/ c = 1 for α = 5°. The trajectory of the computed vortex was also in good agreement as it had moved inboard by the same amount (10% chord) as the experimental vortex at the last measurement location. The axial velocity excess is under predicted for α = 10°, whereas the velocity deficit is in relatively good agreement for α = 5°. The computed Reynolds shear stress component 〈 u′v′〉 is in good agreement with experiment at x/ c = 0 for α = 5°, but is greatly under predicted further downstream and at all locations for α = 10°. It is thought that a lack of local grid refinement in the vortex core and deficiencies in the Reynolds stress turbulence model may have led to errors in the mean flow and turbulence results respectively.

Calcium (Ca) intakes may affect cardiovascular disease risk by altering body composition (body weight and fat) and serum lipid profile, but results have been inconsistent and the underlying mechanisms are not well understood. The effects of dietary Ca on body composition and lipid metabolism were examined in rats. Male Sprague-Dawley rats were fed high-fat, high-energy diets containing (g/kg) low (0.75Ca, 0.86 ± 0.05; 2Ca, 2.26 ± 0.02), normal (5Ca, 5.55 ± 0.08) or high (10Ca, 11.03 ± 0.17; 20Ca, 21.79 ± 0.15) Ca for 10 weeks. Rats fed the lowest Ca diet (0.75Ca) had lower (p < 0.05) body weight and fat mass compared to other groups. Rats fed the high Ca diets had lower serum total and LDL cholesterol compared to rats fed normal or low Ca. Liver total cholesterol was lower in rats fed high compared to low Ca. In general, liver mRNA expression of genes involved in cholesterol uptake from the circulation (Ldlr), cholesterol synthesis (Hmgcr and Hmgcs1), fatty acid oxidation (Cpt2) and cholesterol esterification (Acat2) were higher in rats fed higher Ca. Apparent digestibility of total trans, saturated, monounsaturated and polyunsaturated fatty acids was lower in rats fed the high compared to the low Ca diets, with the largest effects seen on trans and saturated fatty acids. Fecal excretion of cholesterol and total bile acids was highest in rats fed the highest Ca diet (20Ca). The results suggest little effect of dietary Ca on body composition unless Ca intakes are very low. Decreased bile acid reabsorption and reduced absorption of neutral sterols and saturated and trans fatty acids may contribute to the better serum lipid profile in rats fed higher Ca.

A strong correlation between the localization of atherosclerotic lesions with abnormal wall shear stress (WSS) has long been recognized at the distal anastomosis of a peripheral bypass where disturbed flow occurs. Identification of a WSS variable that significantly contributes to disease formation at this site has been elusive to date, as endothelial cell (EC) response to the abnormal hemodynamics at this anastomotic junction has yet to be quantitatively characterized. In vitro experiments were performed exposing human aortic ECs to pulsatile flow variables appropriate to a distal bypass graft junction using a novel in vitro device. Computational fluid dynamics was employed to detail the hemodynamic variables of this flow chamber. These variables were then correlated to EC proliferation which was used as an indicator of intimal hyperplasia (IH) development. Under pulsatile flow, maximum absolute temporal WSS gradient was found to be the parameter that most significantly correlated to EC proliferation (p = 0.0001, r = 0.8947). This metric can be utilized as an indicator to detect arterial segments prone to the initiation and localization of IH, it may also be used to help optimize the surgical construction of coronary artery bypass, peripheral bypass and arteriovenous anastomotic junctions to improve patient outcomes.

Many viruses have evolved to exploit cell-surface glycosaminoglycans (GAG), particularly heparan sulfate, to facilitate their attachment and infection of host cells. Here, the case for the involvement of heparan sulfate GAG in cellular infection by human immunodeficiency virus Type 1 (HIV-1) compared with herpes simplex virus Type 1 (HSV-1) is re-examined. It is shown that HIV-1 infection is facilitated by heparan sulfate GAG in only one of three highly permissive cell lines tested, whereas HSV-1 infection is facilitated to varying extents in all three. To evaluate the physiological relevance of these findings, primary peripheral blood lymphocytes (PBL), the physiological host for HIV-1, were examined. It was found that treatment of PBL with heparitinase, to remove any traces of heparan sulfate GAG, did not alter their sensitivity to infection by either lymphocyte-tropic, X4-type strain HIV-1IIIB, nor the monocyte-tropic, R5-type strain, HIV-1Ba-L. It is concluded that heparan sulfate GAG has little physiological role in the infection of lymphocytes by HIV-1 and that evidence derived from studies on immortalized cell lines suggesting a significant role must be interpreted with caution.

Immune evasion by the human immunodeficiency virus (HIV) is unexplained but may involve the mutation of viral antigens. When cytotoxic T lymphocytes engaged CD4-positive cells that were acutely infected with HIV bearing natural variant epitopes in reverse transcriptase, substantial inhibition of specific antiviral lysis was observed. Mutant viruses capable of these transactive effects could facilitate the persistence of a broad range of HIV variants in the face of an active and specific immune response.

Sodium (Na) and potassium (K) are essential nutrients. Like people in many Western societies, Canadians consume too much Na and not enough K, both of which contribute to hypertension. We analysed the Na and K content of 154 food composites, broadly representative of the foods most commonly consumed in Canada, from the Canadian Total Diet Study collection of 2007. Foods were prepared as if for home consumption before compositing. No salt was added during food preparation. Samples prepared by aqueous extraction were analysed by atomic emission (Na) or atomic absorption spectrometry (K). Processed foods and soups contained large amounts of Na per reference amount (serving) of the food, with 17 samples containing over 1/3 of the Adequate Intake (AI) for adults, or >500 mg Na/serving. Fluid milk, unprocessed meats and several fruits and vegetables contained large amounts of K per reference amount, with 11 samples containing over 10% of the AI for adults, or >470 mg K/serving. Na:K molar ratios were typically either high or low, with few values near unity. Thus, with few exceptions, foods high in Na were lower in K, and vice versa. Through judicious food selection it may be possible for consumers to decrease Na intake while increasing K, with associated health benefits. Such choices would be consistent with common nutrition advice to decrease consumption of processed foods, while increasing intakes of fresh fruits and vegetables.

The records of 30 patients who had suffered from vertebral osteomyelitis were reviewed. They conformed to a constant pattern, though varying in terms of: (i) the severity of the disease, due to host-organism interrelationship; and (ii) age distribution. Causative organisms could not always be identified, though all lesions settled with conservative measures of rest and antibiotics. A high proportion of the patients who were followed up for more than one year were back at work. The anatomical distribution of the lesions can be explained by our knowledge of the vascular supply to the vertebral bodies.

Introduction Non-maturation and post-maturation venous stenosis are the primary causes of failure within arteriovenous fistulae (AVFs). Although the exact mechanisms triggering failure remain unclear, abnormal hemodynamic profiles are thought to mediate vascular remodelling and can adversely impact on fistula patency. Aim The review aims to clarify the role of shear stress on outward remodelling during maturation and evaluate the evidence supporting theories related to the localisation and development of intimal hyperplasia within AVFs. Methods A systematic review of studies comparing remodelling data with hemodynamic data obtained from computational fluid dynamics of AVFs during and after maturation was conducted. Results Outward remodelling occurred to reduce or normalise the level of shear stress over time in fistulae with a large radius of curvature (curved) whereas shear stress was found to augment over time in fistulae with a small radius of curvature (straight) coinciding with minimal to no increases in lumen area. Although this review highlighted that there is a growing body of evidence suggesting low and oscillating shear stress may stimulate the initiation and development of intimal medial thickening within AVFs. Further lines of evidence are needed to support the disturbed flow theory and outward remodelling findings before surgical configurations and treatment strategies are optimised to conform to them. This review highlighted that variation between the time of analysis, classification of IH, resolution of simulations, data processing techniques and omission of various shear stress metrics prevented forming pooling of data amongst studies. Conclusion Standardised measurements and data processing techniques are needed to comprehensively evaluate the relationship between shear stress and intimal medial thickening. Advances in image acquisition and flow quantifications coupled with the increasing prevalence of longitudinal studies commencing from fistula creation offer viable techniques and strategies to robustly evaluate the relationship between shear stress and remodelling during maturation and thereafter.

The creation of an arteriovenous fistula for hemodialysis has been reported to generate unstable to turbulent flow behaviour. On the other hand, the vast majority of computational fluid dynamic studies of an arteriovenous fistula use low spatial and temporal resolutions resolution in conjunction with laminar assumptions to investigate bulk flow and near wall parameters. The objective of the present study is to investigate if adequately resolved CFD can capture instabilities within an arteriovenous fistula. An experimental model of a representative fistula was created and the pressure distribution within the model was analysed for steady inlet conditions. Temporal CFD simulations with steady inflow conditions were computed for comparison. Following this verification a pulsatile simulation was employed to assess the role of pulsatility on bulk flow parameters. High frequency fluctuations beyond 100 Hz were found to occupy the venous segment of the arteriovenous fistula under pulsatile conditions and the flow within the venous segment exhibited unstable behaviour under both steady and pulsatile inlet conditions. The presence of high frequency fluctuations may be overlooked unless adequate spatial and temporal resolutions are employed. These fluctuations may impact endothelial cell function and contribute to the cascade of events leading to aggressive intimal hyperplasia and the loss of functionality of the vascular access.

High powered laser diodes are used in many applications that require stable optical output, such as materials processing, medical and military applications, and solid-state laser pumping [1,2]. Due to the low electrical to optical efficiencies of 25-35%, significant waste heat is generated which increases the junction temperature and alters the wavelength of the emitted beam. The industry standard for rejecting this waste heat is single-phase, copper microchannel cooler (MCC) using high-purity de-ionized water (DIW) coolant. However, the high-purity DIW coolant is pumped through the copper MCCs in presence of leakage current at high velocity, which causes erosion-corrosion damage to the micron sized heat transfer surface areas, and reduces the thermal performance of the cooler. Since the corrosion of copper is highly dependent on the pH and dissolved oxygen (DO) content of the DIW coolant, strict control on water conditioning system is employed. This strict control scheme leads to additional equipment, costs and reliability concerns. Extension of the cooler lifetime by protecting the MCCs against erosion-corrosion can be achieved by applying a uniform, pin-hole free and conformal coating to the internal features of the MCCs. However, many coating and plating techniques, such as nickel and gold plating, cannot meet these requirements. Thus, a vapor phase deposition technique that uniformly applies a nanometer thin, conformal, inert, hard, coating to the high-aspect ratio internal features of the MCC was developed to protect the copper MCC against erosioncorrosion in high powered laser diode applications. Corrosion rate measurements of baseline uncoated and coated copper samples exhibited a one to two order of magnitude reduction in corrosion rate when exposed to DIW with a pH of 6.0 – 9.0 and a DO concentration ranging from 0.5 ppm to 10.0 ppm. This study shows that the strict controls required to maintain the pH and DO can be severely relaxed by applying the coating resulting in reduced operational costs and increased reliability. Furthermore, evaluations of the coating thickness deposited throughout the microchannel region of the MCC demonstrate the uniform and conformal application of the coating in the high aspect ratio features. Lastly, thermal and hydraulic performance evaluations of coated MCCs revealed that the application does not impede the thermal or hydraulic performance of the MCC, thereby enabling lifetime extension without adding pumping po wer requirements.

Objective To determine HIV-1 reverse transcriptase (RT) and protease (PRO) mutations selected in isolates from antiretroviral therapy (ART)-experienced patients receiving an efavirenz/abacavir/amprenavir salvage regimen. Methods Open-label, single arm of abacavir, 300 mg twice daily, amprenavir, 1200 mg twice daily and efavirenz, 600 mg once daily, in ART-experienced patients of which 42% were non-nucleoside reverse transcriptase inhibitor-naive. The virology population examined consisted of all patients who took at least 16 weeks of study drugs (n=74). Plasma population sequencing was carried out at baseline and last time point at which patients were still taking the three study drugs ± other ART. The median follow-up was 48 weeks (range week 16–72). Results Baseline (n=73) and on-therapy (n=49) genotypes were obtained. By 48 weeks, 51% of isolates had ≥3 non-nucleoside reverse transcriptase inhibitor (NNRTI) mutations. NNRTI mutations selected on therapy were K103N (51%), substitutions at position 190 (17/49, 35%): G to A (n=11) / S (n=4) / E (n=1) and T (n=1); L100I (37%) and V108I (20%) mutations. P225H was not observed in this study. L100I and G190A/S/E/T mutations were rarely detected in the same viral population and baseline Y181C favoured the G190 mutations (OR=8.9, PConclusion Prior NNRTI and NRTI therapy influences the pathway of resistance to efavirenz. In this study, the prevalence of mutations selected by efavirenz were different from those described in less ART-experienced patients. Baseline Y181C was associated with the development of mutations at position 190, but not L100I or K103N. In this patient population, abacavir with efavirenz preferentially selected for L74V but not for thymidine analogue mutations. M184V was rarely selected and was maintained in only 77% of patients who did not add lamivudine or didanosine. Finally, amprenavir-specific mutations were selected in the background of other primary protease inhibitor mutations, confirming the distinct resistance profile of amprenavir.

Experimental data is presented for a flat plate test facility with augmented levels of freestream turbulence (FST). The turbulence decay downstream of two turbulence generating grids, in addition to the integral length scales, is provided and good comparison with established correlations is presented. Boundary layer measurements using a single normal hotwire probe were obtained at FST intensities of 7%, 6%, 5.5%, 1.55%, and 1.45%, and the results presented include the fifth and 95th percentile of the velocity fluctuations and the root mean squared (RMS) velocity profiles near the transition onset region. The transition onset Reynolds number for each of the turbulence levels considered is consistent with theoretical findings. In all cases analyzed, the maximum fifth and 95th percentile far exceeded the maximum RMS values, with the location of the maximum 95th percentile closer to the wall compared to the maximum fifth percentile. Using probability density function (PDF) analysis, it is demonstrated that there is a dominating positive velocity fluctuation in the near-wall region and a dominating negative velocity fluctuation further out in the boundary layer and that the fluctuations in the boundary layer are greater compared to the freestream. The effect of the FST on the boundary layer is discussed with comparison to the Blasius solution and the influence of the fluctuations on the deviation from the Blasius profile is presented and discussed. Through investigation of the energy spectrum of the fluctuating velocity component within the boundary layer, it is shown that there is a higher energy content at lower frequency in the boundary layer when compared to that of the freestream.

In common with all retroviruses, the human immunodeficiency virus type 1 (HIV-1) contains duplicated long terminal repeat (LTR) sequences flanking the proviral genome. These LTRs contain identical poly(A) signals, which are both transcribed into RNA. Therefore, to allow efficient viral expression, a mechanism must exist to either restrict promoter-proximal poly(A) site use or enhance the activity of the promoter-distal poly(A) site. We have examined the use of both poly(A) sites using proviral clones. Mutation of the previously defined upstream activatory sequences of the 3' LTR poly(A) site decreases the efficiency of polyadenylation when placed in competition with an efficient downstream processing signal. However, in the absence of competition, these mutations have no effect on HIV-1 polyadenylation. In addition, the 5' LTR poly(A) site is inactive, whereas a heterologous poly(A) site positioned in its place is utilized efficiently. Furthermore, transcription initiating from the 3' LTR promoter utilizes the 3' LTR poly(A) signal efficiently. Therefore, the main determinant of the differential poly(A) site use appears to be neither proximity to a promoter element in the 5' LTR nor the presence of upstream activating sequences at the 3' LTR. Instead, we show that the major splice donor site that is immediately downstream of the 5' LTR inhibits cleavage and polyadenylation at the promoter-proximal site. The fact that this poly(A) site is active in a proviral clone when the major splice donor site is mutated suggests that the selective use of poly(A) signals in HIV-1 is mediated by a direct inhibition of the HIV-1 poly(A) site by downstream splicing events or factors involved in splicing.

turbulence intensity of 9% just after the trailing edge. Maximum turbulence intensity after x/c = 0 is observed in the vortex core which decays with downstream distance. Five-hole probe mean velocity measurements revealed jet like and wake like axial velocity profiles depending on wing angle of attack, with values of 1.33U∞ and 0.83U∞ being measured for 10° and 5°. Axial vorticity is also observed to increase gradually with downstream distance. A full Reynolds stress turbulence model with a second order upwind differencing scheme is used to compute the vortex in the near-field. Vortex structure and trajectory correlate well with experiment. It is thought that the dissipative nature of the second order convection scheme led to discrepancies between numerical and experimental results.

Hemodialysis patients require a vascular access capable of accommodating the high blood flow rates required for effective dialysis treatment. The arteriovenous graft is one such access. However, this access type suffers from reduced one year primary & secondary patency rates of 59–90% and 50–82% respectively [1]. The main contributor to the failure of this access is stenosis via the development of intimal hyperplasia (IH) that predominately occurs at the venous anastomosis. It is hypothesized that the resulting transitional to turbulent flow regime within the venous anastomosis contributes to the development of IH. The aim of this study is to investigate the influence of this transitional to turbulent behavior on wall shear stress within the venous anastomosis via the use of large eddy simulation.Copyright © 2012 by ASME

The aim of the present study was to elucidate possible cholesterol-lowering mechanism(s) of high-dose supplemental Se in the form of selenite, a known hypocholesterolaemic agent. Male Syrian hamsters (four groups, ten per group) were fed semi-purified diets for 4 weeks containing 0·1 % cholesterol and 15 % saturated fat with selenite corresponding to varying levels of Se: (1) Se 0·15 parts per million (ppm), control diet; (2) Se 0·85 ppm; (3) Se 1·7 ppm; (4) Se 3·4 ppm. Lipids were measured in the bile, faeces, liver and plasma. The mRNA expression of several known regulators of cholesterol homeostasis (ATP-binding cassette transportersg5(Abcg5) andg8(Abcg8), 7-hydroxylase, 3-hydroxy-3-methylglutaryl-coenzyme A reductase, LDL receptor (LdLr)and Nieman-Pick C1-like 1 protein (Npc1l1)) were measured in the liver and/or jejunum. Oxysterols including 24-(S)-hydroxycholesterol, 25-hydroxycholesterol and 27-hydroxycholesterol (27-OHC) were measured in the liver. Significantly lower total plasma cholesterol concentrations were observed in hamsters consuming the low (0·85 ppm) and high (3·4 ppm) Se doses. The two highest doses of Se resulted in decreased plasma LDL-cholesterol concentrations and increased mRNA levels of hepaticAbcg8,Ldlrand jejunalLdlr.Higher hepatic 27-OHC and TAG concentrations and lower levels of jejunalNpc1l1mRNA expression were noted in the 1·7 and 3·4 ppm Se-treated hamsters. Overall, Se-induced tissue changes in mRNA expression including increased hepaticAbcg8andLdlr, increased jejunalLdlrand decreased jejunalNpc1l1, provide further elucidation regarding the hypocholesterolaemic mechanisms of action of Se in the form of selenite.

Hypercholesterolemia diets are associated with oxidative stress that may contribute to hypercholesterolemia by adversely affecting enzymatically-generated oxysterols involved in cholesterol homeostasis. An experiment was conducted to examine whether the cholesterol-lowering effects of the antioxidants selenium and α-tocopherol were related to hepatic oxysterol concentrations. Four groups of male Syrian hamsters (n = 7-8) were fed high cholesterol and saturated fat (0.46% cholesterol, 14.3% fat) hypercholesterolemic semi-purified diets: 1) Control; 2) Control + α-tocopherol (67 IU all-racemic-α-tocopheryl-acetate/kg diet); 3) Control + selenium (3.4 mg selenate/kg diet); and 4) Control + α-tocopherol + selenium. Antioxidant supplementation was associated with lowered plasma cholesterol concentrations, decreased tissue lipid peroxidation and higher hepatic oxysterol concentrations. A second experiment examined the effect of graded selenium doses (0.15, 0.85, 1.7 and 3.4 mg selenate/kg diet) on mRNA expression of the oxysterol-generating enzyme, hepatic 27-hydroxylase (CYP27A1, EC 1.14.13.15), in hamsters (n = 8-9) fed the hypercholesterolemic diets. Supplementation of selenium at 3.4 mg selenate/kg diet was not associated with increased hepatic 27-hydroxylase mRNA. In conclusion, the cholesterol lowering effects of selenium and α-tocopherol were associated with increased hepatic enzymatically generated oxysterol concentrations, which appears to be mediated via improved antioxidant status rather than increased enzymatic production.

Multifrequency susceptibility measurements on the perovskite relaxor ferroelectric ${({\text{PbMg}}_{1/3}{\text{Nb}}_{2/3}{\text{O}}_{3})}_{0.88}{({\text{PbTiO}}_{3})}_{0.12}$ were performed at various strengths of dc electric field applied along the [111] direction. The temperature-frequency dependences fit the Vogel-Fulcher form, allowing the extraction of a frequency-independent glassy freezing temperature. These Vogel-Fulcher temperatures showed significant reductions in applied fields, following an empirical Gabay-Toulouse form, similar to vector spin glasses. The magnitude of the sensitivity indicates that the glassy state is formed by interactions among the same entities which account for the susceptibility, i.e., the polar nanoregions. That interpretation is supported by other data showing a loss of Vogel-Fulcher behavior in a powder sample of ${\text{PbMg}}_{1/3}{\text{Nb}}_{2/3}{\text{O}}_{3}$, with grains too small to support large-scale internanoregion cooperativity.

Microscale fluid dynamics has played a significant role in the development of many applications in the medical diagnostic sector and in recent years many devices have implemented advances in this field. Capillary driven assays are commonly used in diagnostic areas, such as, cardiac risk, fertility, drug abuse and infectious diseases1. Typically a platform is used where bodily fluids or samples are taken, filtered and by means of small microchannels, transported and mixed with a variety of antibodies2. In order to perform correctly, these antibodies need to bind to the proteins in the fluid. It is therefore essential that the exposure of the proteins to the antibodies is maximized. To achieve this, capillary dimensions can be altered to obtain the required flow rates and exposure times3. This paper focuses on controlling these parameters. In this paper, a study was conducted in which the flows in four straight rectangular microchannels of varying cross sectional areas were assessed. The four microchannels were fabricated from an epoxy material. The microchannel widths varied from 100μm to 1000μm with each channel having a dept of 200μm. The four microchannels had aspect ratios of 0.5, 1, 2 and 10. The microchannels were sealed using a heat sealing hydrophilic tape. Fluid velocity rates were measured experimentally using an X-Stream XS-4 high speed camera at 500 frames per second. Preliminary contact angle results between water and the epoxy material gave a contact angle of 81.5 degrees +/− 6 degrees. Computational models of the four microchannels were performed using a Volume Of Fluid (VOF) model in Fluent 6.2.16, a commercially available CFD code. The computational models had four boundary types: pressure inlet, pressure outlet, epoxy wall and hydrophilic tape wall. The inlet boundary has an initial pressure applied to it, capillary pressure between the water and air interface in the microchannel. It was found that as the dimensions of the microchannels increased, the governing equations were less accurate in predicting the experimental fluid velocity in the microchannels.

Background Despite its widespread use to assess fibrosis, liver biopsy has several important drawbacks, including that is it semi-quantitative, invasive, and limited by sampling and observer variability. Non-invasive serum biomarkers may more accurately reflect the fibrogenetic process. To identify potential biomarkers of fibrosis, we compared serum protein expression profiles in patients with chronic hepatitis C (CHC) virus infection and fibrosis. Methods Twenty-one patients with no or mild fibrosis (METAVIR stage F0, F1) and 23 with advanced fibrosis (F3, F4) were retrospectively identified from a pedigreed database of 1600 CHC patients. All samples were carefully phenotyped and matched for age, gender, race, body mass index, genotype, duration of infection, alcohol use, and viral load. Expression profiling was performed in a blinded fashion using a 2D polyacrylamide gel electrophoresis/LC-MS/MS platform. Partial least squares discriminant analysis and likelihood ratio statistics were used to rank individual differences in protein expression between the 2 groups. Results Seven individual protein spots were identified as either significantly increased (α2-macroglobulin, haptoglobin, albumin) or decreased (complement C-4, serum retinol binding protein, apolipoprotein A-1, and two isoforms of apolipoprotein A-IV) with advanced fibrosis. Three individual proteins, haptoglobin, apolipoprotein A-1, and α2-macroglobulin, are included in existing non-invasive serum marker panels. Conclusion Biomarkers identified through expression profiling may facilitate the development of more accurate marker algorithms to better quantitate hepatic fibrosis and monitor disease progression.

Objectives Abacavir (ABC) selects for four mutations (K65R, L74V, Y115F and M184V) in HIV-1 reverse transcriptase (RT), both in vitro and during monotherapy in vivo. The aim of this analysis was to compare the selection of these and other nucleoside reverse transcriptase inhibitor (NRTI)-associated mutations by ABC-containing therapies in the presence and absence of concurrent lamivudine (3TC) and/or zidovudine (ZDV) and to assess the effect of these mutations on phenotypic susceptibility to the NRTIs. Design This study was a retrospective analysis of the patterns of NRTI-associated mutations selected following virological failure in six multicentre trials conducted during the development of ABC. Methods Virological failure was defined as confirmed vRNA above 400 HIV-1 RNA copies/mL. RT genotype and phenotype were determined using standard methods. Results K65R was selected infrequently by ABC-containing regimens in the absence of ZDV (13 of 127 patients), while L74V/I was selected more frequently (51 of 127 patients). Selection of both K65R and L74V/I was significantly reduced by co-administration of ZDV with ABC (one of 86 and two of 86 patients, respectively). Y115F was uncommon in the absence (seven of 127 patients) or presence (four of 86 patients) of ZDV. M184V was the most frequently selected mutation by ABC alone (24 of 70 patients) and by ABC plus 3TC (48 of 70 patients). Thymidine analogue mutations were associated with ZDV use. The K65R mutation conferred the broadest phenotypic cross-resistance of the mutations studied. Conclusions The resistance pathway selected upon virological failure of ABC-containing regimens is significantly altered by concurrent ZDV use, but not by concurrent 3TC use. These data may have important implications for the efficacy of subsequent lines of NRTI therapies.

Mutations selected or deselected during passage of human immunodeficiency virus strain HXB2 or resistant variants with tenofovir (TFV), abacavir (ABC), and lamivudine (3TC) differed depending on the drug combination and virus genotype. In the wild-type virus, TFV-ABC and TFV-3TC selected K65R (with reduced susceptibility to all three inhibitors) and then Y115F. TFV-containing regimens might increase K65R selection, which confers multiple nucleoside reverse transcriptase inhibitor resistance.

The final chapter of H. G. Wells's novel, Tono-Bungay (1909) has given rise to surprisingly wide differences of opinion among commentators. Geoffrey West, an early admirer of the novel, thought that 'in the last chapter, one of the most splendid passages Wells has ever written, is focused the whole spirit of the book';1 more recently J. R. Hammond referred to the chapter's 'series of brilliant images' and considered it 'one of the most carefully written ... in the whole corpus of [Wells's] fiction'.2 At the opposite extreme from these views is Mark Schorer's familiar dismissal of TonoBungay in his essay 'Technique as Discovery' (1948) and his condemnation of the final chapter as a 'significant failure' because it is merely 'a kind of meditative rhapsody which denies every value that the book has been aiming towards'.3 Even more severe than Schorer's criticism was Walter Allen's opinion that the chapter was 'one of Wells's shoddiest and most careless pieces of writing' in a generally muddled novel.4 In this final chapter George Ponderevo brings his story full circle by once more depicting himself in the act of writing it, and then expands on his new career of ship-designer by describing the seatrials of an experimental destroyer, the X2. This brief episode ends the novel. In addition to the stylistic and structural aspects of the chapter, the sudden appearance of the X2 has also puzzled critics. Although there are brief references to this vessel earlier in the novel, the X2 seems something of an after-thought, mainly because George gives no account of how he came to his new occupation of designing warships. Moreover, George himself seems uncertain what 'the value of a thing at once so essential and so immaterial' might be.5

Objective To examine changes in HIV-1 susceptibility (genotype and phenotype) during an initial abacavir monotherapy phase followed by the addition of zidovudine and lamivudine. Design Sixty HIV-1 infected, antiretroviral therapy-naive subjects were randomized to receive 100, 300 or 600 mg abacavir twice daily. Subjects completing 24 weeks of randomized therapy or meeting a protocol defined switch criterion could switch to open label abacavir/zidovudine/lamivudine. Methods Plasma HIV-1 reverse transcriptase was genotyped at baseline, week 12, and at the last time point on ABC monotherapy. Drug susceptibility was analysed at baseline and on subsequent samples with sufficient HIV-1 RNA levels using the recombinant virus assay. Virological responses (week 24) were correlated to week 24 genotypes. Results Mutant viruses were not detected before week 12 with the exception of one subject. At the latest time point on abacavir monotherapy (range, weeks 6-48), 21 out of 43 subjects harboured virus with resistance conferring mutations including single, double and triple combinations of K65R, L74V, Y115F and M184V. The most common mutational pattern was L74V + M184V (11/21 cases). Twenty of the 21 subjects with isolates containing abacavir-associated mutations reached week 48, and upon addition of lamivudine/zidovudiine, 16 out of 20 (80%) had week 48 plasma HIV-1 -RNA below 400 copies/ml. At week 48, 16 out of 46 genotypes were obtained; one of these was wild-type; 15 contained M184V either alone, in combination with K65R and/or L74V and/or Y115F or with thymidine analogue-associated mutations. Week 48 viral load levels for these 15 subjects was low (median 3.43 log10 copies/ml or -1.99 log10 copies reduction from baseline). Genotype correlated well with phenotypic resistance to ABC; four samples with three abacavir-associated mutations had high level abacavir resistance (> 8-fold) and six samples with two or three mutations showed intermediate (4-8-fold) resistance. All samples with single mutations retained full ABC susceptibility. Conclusions Resistance conferring mutations to abacavir were relatively slow to develop during the monotherapy phase, and did not preclude durable efficacy of abacavir/lamivudine/zidovudine up to 48 weeks.

Summary: Polydactylous squamous cell carcinoma (SCC) is rare and has been associated with human papillomavirus (HPV). Our recent case was HPV negative and provides greater evidence for chemical irritants being an alternative cause of subungual SCC. Our patient had spent a number of years with her hands in direct contact with undiluted cleaning chemicals including one containing ethanolamine. Ethanolamine has been shown to have carcinogen sensitizing role. Although HPV has a strong association with subungual SCCs, the accumulation and concentration of noxious substances around and under the nails must also be considered as a potential cause.

The bicyclams represent a new class of highly potent and selective HIV inhibitors. Time-of-addition experiments have previously shown that these compounds interfere with an early event in the viral replicative cycle. Additional experiments have now been carried out in order to investigate in more detail the mechanism of action of these promising compounds. As described in this paper, PCR experiments revealed that no viral DNA was formed following viral infection, thus confining the target(s) of action of the bicyclams to an early stage of HIV infection. An assay, using pseudotype virions containing the envelope of HIV-1 and the genome of a plaque-forming virus (Cocal Virus), pointed to viral entry as the main target of the bicyclams. HIV-1 strains resistant to two prototype bicyclams, JM2763 and SID791 (JM3100), were raised. Results obtained with SID791 with respect to syncytium formation induced by SID791-sensitive and -resistant HIV-1 strains and the cross-resistance observed for dextran sulfate, suggest inhibition of binding and/or fusion as a plausible target of SID791. Additional experiments enabled us to exclude SID791 and JM2763 as binding inhibitors and to conclude that bicyclams block the entry of cell-bound virus. Furthermore, a monoclonal antibody recognising the V3 loop of wild-type gp120 did not bind to this region in the two bicyclam-resistant strains. Our results point to gp120 as a possible target for the HIV-inhibitory effects of the bicyclams.

Interleukin-10 (IL-10), a product of T lymphocytes, B cells and macrophages, participates in Th-2 immune responses and modulates macrophage functions including possible interactions with pathogens. We have found that Chinese hamster ovary cell-derived human recombinant (hr) IL-10 inhibits human immunodeficiency virus type 1 strains Ada and Ba-L (HIV-1ADA and HIV-1Ba-L) replication in primary tissue culture-derived macrophages in a dose-dependent manner. Inhibition by IL-10 treatment (> 5 U/ml) was effective 72 h before or 24 h after infection and cytokine activity blocked by anti-hrIL-10 antibody (19F1), or lost after heat inactivation of IL-10. Viral production was measured by determining p24 and reverse transcriptase levels while reverse transcription kinetics for the long terminal repeat (LTR) and gag were assessed at timed intervals after infection and quantified by 32P end-labelling. IL-10 inhibited early steps of infection without modulating cell surface CD4+ levels. The onset of LTR reverse transcription was delayed by 4 to 8 h and the number of LTR transcripts was decreased by 77% at 24 h and by 87% 48 h after infection. IL-10 effects were reversible; after cytokine washout, cells treated before infection showed lower levels of virus compared with those treated after infection. IL-10 biological activity was confirmed in three virus-independent assays. These results demonstrate IL-10 decreases HIV-1 reverse transcription upon macrophage infection and subsequently mediates viral latency in vitro. Therefore, IL-10 may be involved in the effective control of HIV-1-infected macrophages in vivo.

A scheme for obtaining strain maps from a set of MRI scans of the heart is presented. Using a technique known as the Spatial Modulation of Magnetization a set of MRI scans are generated with tag-lines distributed over the walls of the heart. As the heart moves, these tag-lines will move accordingly. Active contours or snakes are then used to extract the tag-lines from each MRI image, and the motion of the tag-lines determined. Laplacian smoothing splines are then used to estimate the displacement of points between the tag-lines. The strain experienced through out the heart is then a simple function of the associated displacement. Regional strain analysis is of vital importance in diagnosing different forms of heart disease, such as myocardial infarction or ``heart attack''. The method presented in this paper makes the analysis of cardiac strain feasible.

We investigated HIV-1 reverse transcriptase (RT) polymorphisms of plasma isolates from 98 HIV-1-infected study subjects with >2 years of antiretroviral therapy who were failing their current protease inhibitor (PI)-containing regimen. In 1 patient, we detected a virus with a heavily mutated beta3-beta4 connecting loop of the HIV-1 RT fingers subdomain, consisting of a single aspartate codon insertion between positions 69 and 70 and five additional variations: 64N, K65, K66, 67G, 68Y, T69, Ins D, 70R, W71, R72, K73, 74I. Mutants with the recently described 2-aa insertions between codons 68 and 70 of RT were detected in another 3 patients. Among the four isolates with the 1- or 2-aa insertions, the novel genotype was the most refractory to therapy and displayed the highest level of phenotypic resistance to nucleoside reverse transcriptase inhibitors (NRTIs). Follow-up samples demonstrated that the novel mutant represents a stable genetic rearrangement and that the amino acid insertions can coexist with nonnucleoside analogue reverse transcriptase inhibitors (NNRTI) mutations resulting in phenotypic resistance to both NRTIs and NNRTIs. An increasing number of HIV-1 isolates containing various insertions in the beta3-beta4 hairpin of the HIV-1 RT fingers subdomain appear to emerge after prolonged therapy with different NRTIs, and these polymorphisms can confer multiple drug resistance against NRTIs.

To assess the comparative effectiveness of relaxation instructions and frontalis electromyographic (EMG) biofeedback in the treatment of muscle-contraction (tension) headaches, 21 volunteers were assigned to either a relaxation training group, a biofeedback group or a no-treatment control group. Each group met for six 1/2-hr sessions. The EMG biofeedback and the relaxation instructions resulted in significant decreases in reported headache activity. Both procedures were significantly more effective than the control procedure, but did not differ significantly from each other in effectiveness. The effectiveness of the two experimental procedures was maintained at follow-up. Frontalis EMG levels were higher during sessions in which a headache was reported than in sessions when no headache was reported. The importance of individual differences is emphasized.

Objective To evaluate HIV-1 reverse transcriptase (RT) drug resistance in patients receiving abacavir, lamivudine and zidovudine therapy. Methods In a randomized, double-blind study, 173 anti-retroviral treatment-naive HIV-1-infected adults received abacavir/lamivudine/zidovudine or lamivudine/zidovudine for up to 48 weeks. After week 16, patients could switch to open-label abacavir/lamivudine/zidovudine, and those with plasma HIV-1 RNA (vRNA) >400 copies/ml could add other antiretrovirals. From weeks 16 to 48, samples with vRNA >400 copies/ml were collected for genotyping and phenotyping. Results At baseline, 90% of isolates were wild-type (WT). At week 16, vRNA was >400 copies/ml in seven of 72 (10%) patients receiving abacavir/lamivudine/zidovudine and in 41 of 66 (62%) receiving lamivudine/ zidovudine. At week 16, the genotypes in isolates from the abacavir/lamivudine/zidovudine group were M184V alone ( n=3 cases), WT ( n=3) and M184V plus thymidine analogue mutations (TAMs) ( n=1). The genotypes in isolates from the lamivudine/zidovudine group were M184V alone ( n=37), WT ( n=1) and M184V plus TAMs ( n=3). In the four cases where M184V plus TAMs were detected some mutations were present at baseline. Despite detectable M184V in 74% of patients on lamivudine/zidovudine, addition of abacavir with or without another antiretroviral therapy resulted in a reduction in vRNA, with 42 of 65 (65%) patients having week 48 vRNA 10 below baseline), and M184V with or without TAMs in patients originally assigned to lamivudine/zidovudine. At week 48, phenotypic results were obtained for 11 isolates for patients from both arms, and all had reduced susceptibility to lamivudine but all remained sensitive to stavudine, all protease inhibitors and all non-nucleoside reverse transcriptase inhibitors. Three, three and two isolates had reduced susceptibility to abacavir, didanosine and zidovudine, respectively. Conclusions Abacavir retained efficacy against isolates with the M184V genotype alone. TAMs did not develop during 48 weeks of abacavir/lamivudine/zidovudine therapy and were uncommon when abacavir was added after 16 weeks of lamivudine/zidovudine therapy. Limited mutations upon rebound on this triple nucleoside combination allows for several subsequent treatment options.

This paper addresses the performance of various electronic conformal coatings specifically for the protection of the intemal components of liquid crystal display (LCD) modules from moisture. The sealant materials after curing on a glass substrate were aged according to standard cyclic and steadystate damp heat tests. At various stages during the tests the mechanical performance of the materials was assessed. In addition these data were complemented with further measurements of the chemical stability and tendency of the materials to absorb moisture. Overall, from the tests it can be inferred that cyclic damp heat is much more stressful on the materials than steady state. Selection of an appropriate sealant for LCD modules demands a balance of mechanical durability, residual chloride ion content, tendency to absorb moisture, and the volatility of the material at elcvated temperatures. The silicone elastomer was found to have the highest shear strength and a low tendency to absorb moisture, but it demonstrated appreciable chloride ion content and is not repairable. Thermoplastic polyurethane resins appear to be more suitable because, despite lower strength and higher water absorption, they have negligible chloride ion content and are repairable. finally, it was concluded from the strength measurements that cyclic damp heat is more severe than steady-state conditions as an aging test. Steady-state damp heat: temperature of 40 12°C and relative humidity of 93 +2, -3%RH Cyclic damp heat: temperatures varying cyclically from 25 +3"C to 55 *2OC on a 24 hour period, with constant relative humidity of 93 +3%RH. A number of samples were withdrawn at weekly intervals and subject to shear strength measurements. Two sets of additional tests were also conducted on unaged samples of the sealant materials: water absorption, and chloride ion content. In particular, moisture in the presence of ionic contaminants such as chlorides has been reported [I], [2] as potentially problematic for the indium tin oxide traces (ITO) within the LCD module as they are oxidized to chloride while IT0 reduces. Experimental Methods In this section, the entire testing procedure used in this investigation is described starting with the methods used for sample preparation and ageing of the sealant materials. Additionally techniques to assess the mechanical performance and chemical stability of the aged sealants are also explained along with thc procedures for determining their chloride ion content and water absorption ability.

We and others have identified the major determinant of cell tropism in human immunodeficiency virus type 1 (HIV-1) as the V3 loop of glycoprotein gp120. We have conducted a detailed study of two molecularly cloned isolates of HIV-1, HIVJR-CSF and HIVNL4-3, that differ in their tropism for immortalized CD4+ cell lines, by constructing a series of site-directed mutations within the V3 loop of HIVJR-CSF based on the sequence of HIVNL4-3. The phenotypes of these mutants fall into two classes, those which are viable and those which are not. A spontaneous mutant with significantly altered growth properties was also recovered and found to have an additional single amino acid change in the V3 loop sequence. The carboxy-terminal beta-strand part of the V3 loop is the major determinant of cell tropism. However, the results presented here indicate that the functional role of the V3 loop sequences can only be interpreted properly in the context of the original gp120 backbone from which they were derived. These findings show that over-simplistic interpretation of sequence data derived from unknown mixtures of HIV variants in infected persons may be highly misleading.