Dr. Pham has contributed to
16
publications.

Title 
Using 96well Tissue Culture Polystyrene Plates and a Fluorescence Plate Reader As Tools to Study the Survival and Inactivation of Viruses on Surfaces. 
Date 
November 2011 
Journal 
Cytotechnology 
Excerpt 
A method for studying the behavior of viruses on surfaces has been developed and is illustrated by determining the temperatures that inactivate adsorbed viral hemorrhagic septicemia virus (VHSV) and the concentration of 1propanol that disinfected surfaces with adsorbed VHSV and chum salmon virus (CSV). VHSV is a rhabdovirus; CSV, a reovirus, and they were detected with two fish cell lines, EPC and CHSE214, respectively. When polystyrene tissue culture surfaces were incubated with virus, rinsed, and left to dry, they still supported the attachment and spreading of cell lines and after 7 days these cells showed the characteristic CPE of the viruses. Thus cells appeared to be infected directly from surfaces on which viruses had been adsorbed. Applying this property to 96well plates allowed duplicate surfaces to be examined for their infectiousness or support of CPE. For each treatment 80 replicate surfaces in a 96well plate were tested at one time and the results expressed as the number of wells showing CPE. VHSV adsorbed to polystyrene was inactivated by drying in the dark at temperatures above 14 °C, but remained infectious for at least 15 days of drying at 4 °C. For chemical sterilization of polystyrene surfaces with adsorbed virus, disinfection was achieved with 1propanol at 40% for VHSV and at 60% for CSV. As CPE can be conveniently monitored in 96well plates with a fluorescence plate reader, this method can be used to rapidly evaluate a variety of treatments for their ability to inactivate surfacebound viruses.


Title 
Lactate Threshold at the Same Fatfree Mass and Age is Larger in Men Than Women. 
Date 
December 2008 
Journal 
European Journal of Applied Physiology 
Excerpt 
The lactate threshold (LT) represents the onset of a metabolic acidosis during graded exercise testing (GXT). It is typically measured as an oxygen uptake (VO(2)) but then ratio scaled by body mass or VO(2) peak to make comparisons among subjects. Theoretical considerations and empirical evidence suggest that this type of ratio scaling is not valid. A method that allows a dependent variable to be compared between groups at that same value of one or more covariates is analysis of covariance (ANCOVA). Our purpose was to compare the LT, estimated noninvasively by gas exchange (LT(GE)), at the same fatfree mass (FFM) and age in 203 sedentary subjects (102 men) aged 2070 years. Each subject underwent cycle ergometer GXT with LT(GE) measured by the Vslope method. In model development, we discovered an interaction term between sex and age. As dimensional analysis predicts a loglinear relationship between LT(GE) and FFM, two of the model terms were ln LT(GE) and ln FFM. The ANCOVA model was then as follows: dependent variable = ln LT(GE), fixed factor = sex, covariates = ln FFM, age, and sex x age. Sex made a significant contribution to the model (F = 30.7, P < 0.001). At the mean FFM (56.32 kg) and age (44.01 years) of both sexes combined, the LT(GE) was 29% larger in males (1,307 ml min(1) versus 1,011 ml min(1)). The model's interaction term resulted in larger differences at younger ages and smaller differences at older ages. We conclude that LT(GE) at the same FFM and age is larger in sedentary men compared to sedentary women.


Title 
Complete Genomes of Three Subtype 6t Isolates and Analysis of Many Novel Hepatitis C Virus Variants Within Genotype 6. 
Date 
April 2008 
Journal 
The Journal of General Virology 
Excerpt 
In this study, the complete genomic sequence was determined for three hepatitis C virus variants (VT21, TV241 and TV249) of genotype 6 that do not classify within the established subtypes. All three genomes were isolated from patients in Vietnam and sequenced using 100 microl of serum. They showed 91.493.6% nucleotide similarities to each other but only 71.779.4% similarities to 17 reference sequences representing subtypes 6a6q and to isolates km41 and gz52557. VT21, TV241 and TV249 displayed genome lengths of 9407, 9460 and 9445 nt, respectively. All three isolates contained a single open reading frame of 9051 nt while the 5'UTRs and 3'UTRs were 324338 nt and 3271 nt, respectively. They shared common sizes with QC227/6o and QC216/6p isolates in all ten protein regions. Phylogenetic analyses demonstrated that VT21, TV241 and TV249 clustered independently and were assigned subtype 6t, following the recent designations of 6r and 6s. Analysis of partial genomic sequences available for genotype 6 variants revealed five additional subtype 6t isolates, all originating from Vietnam. This analysis revealed two additional groups of isolates, and at least seven novel variants analogous to km41 and gz52557 that group independently and do not classify within the subtypes 6a6t. This suggests the existence of at least 11 additional subtypes for genotype 6. In addition, the existence of isolates showing genetic distances greater than those within subtypes, but lesser than those between subtypes, raises interesting questions regarding the classification of HCV.


Title 
Comparison of Three Methods for Detection of the Lactate Threshold. 
Date 
December 2007 
Journal 
Clinical Physiology and Functional Imaging 
Excerpt 
The lactate threshold (LT) represents the onset of a metabolic acidosis during graded exercise testing (GXT). It is a valuable measurement in clinical exercise testing and correlates well with endurance performance. Our purpose was to compare three LT detection methods, namely, Inspection (work rate at onset of a systematic increase in blood lactate concentration determined by inspection of blood lactate versus work rate plot), 0.5 mM (work rate which just precedes a rise in blood lactate concentration of >0.5 mM) and loglog (work rate at the intersection of two linear lines in plot of log lactate versus log work rate where the residual sum of squares for both lines added together is minimized). Fourteen subjects underwent cycle ergometer GXT with blood samples obtained at the end of each 3min work rate increment and analysed for lactate concentration. The mean +/ 95% confidence limits of work rates at LT for the Inspection, 0.5 mM and loglog methods were 104.5 +/ 28.0, 103.2 +/ 28.1 and 105.1 +/ 27.3 W, respectively. Repeatedmeasures analysis of variance yielded a nonsignificant F ratio. The BlandAltman bias +/ 95% limits of agreement for Inspection versus 0.5 mM, Inspection versus loglog and 0.5 mM versus loglog were 1.3 +/ 20.6, 0.6 +/ 12.5 and 1.9 +/ 20.5 W, respectively. The intraclass correlation coefficients for Inspection versus 0.5 mM, Inspection versus loglog and 0.5 mM versus loglog were 0.978, 0.992 and 0.977, respectively. The results of this study suggest that all three methods detect the LT at the same work rate.


Title 
Effect of Plasma Volume Loss During Graded Exercise Testing on Blood Lactate Concentration. 
Date 
August 2007 
Journal 
The Journal of Physiological Sciences : Jps 
Excerpt 
Previous studies have shown that plasma volume (PV) loss can be a confounding variable in the interpretation of changes in blood constituents. We examined the effect of PV loss on three features of the blood lactate versus workrate relationship, namely, slight blood lactate increase during the early stages of graded exercise testing (GXT); work rate at the onset of a systematic increase in blood lactate, i.e., lactate threshold (LT); and work rate at a blood lactate concentration of 4 mM, i.e., onset of blood lactate accumulation (OBLA). Fourteen subjects underwent cycle ergometer GXT. Blood samples were obtained at rest and at the end of each 3min workrate increment and analyzed for hematocrit and lactate concentration. For exercise levels up to and including LT, PV loss was relatively stable at approximately 2.8%. Beyond LT, PV loss accelerated. From the first work rate to LT, blood lactate concentration uncorrected for PV loss increased 0.24 +/ 0.07 mM (P < 0.05). After correction for PV loss, the increase was 0.21 +/ 0.08 mM (P < 0.05). These mean increases were not significantly different from each other. For the four exercise levels above LT common to most subjects, PVcorrected lactate values were significantly lower than uncorrected values. Correction of lactate values for PV loss did not alter LT for any subject, but it did result in a significant increase in OBLA. Thus, PV loss has the potential to be a confounding variable for the interpretation of blood lactate parameters that are determined at exercise levels above LT.


Title 
Scaling of Lactate Threshold by Peak Oxygen Uptake and by Fatfree Mass 0 X 67. 
Date 
June 2007 
Journal 
Clinical Physiology and Functional Imaging 
Excerpt 
The lactate threshold (LT) represents the onset of metabolic acidosis during cardiopulmonary exercise testing (CPET). It is measured as a O(2) in the units of ml min(1). In order to make comparisons among subjects, LT is often scaled or normalized by O(2) peak resulting in the LT/O(2) peak ratio. Ratio variables have underlying assumptions. One assumption is that the relationship between the numerator and denominator is linear with a zero yintercept. If the relationship has a positive yintercept, then the ratio will decrease with increasing values of the scaling variable thereby penalizing subjects with larger values of the scaling variable. Our purpose was to examine the validity of scaling LT by O(2) peak and by fatfree mass raised to 0 x 67 power (FFM(0 x 67)) as dimensional analysis predicts that LT is proportional to FFM(0 x 67). Cycle ergometer CPET was administered to 204 healthy, sedentary subjects (103 males) to the limit of tolerance. Lactate threshold was estimated noninvasively using the Vslope technique. Fatfree mass was assessed by skinfolds. The relationship of LT versus O(2) peak was linear with a positive yintercept for both sexes. Consequently, the LT/O(2) peak ratio decreased as O(2) peak increased for both sexes. The relationship of LT versus FFM(0 x 67)was linear with a zero yintercept for both sexes. Consequently, the plot of the LT/FFM(0 x 67) ratio versus FFM resulted in a straight line with a slope of zero for both sexes. The results of this study support the conclusion that FFM(0 x 67), but not O(2) peak, is a valid scaling variable for LT.


Title 
Reliability and Validity of the Lung Volume Measurement Made by the Bod Pod Body Composition System. 
Date 
March 2007 
Journal 
Clinical Physiology and Functional Imaging 
Excerpt 
The BOD POD Body Composition System uses airdisplacement plethysmography to measure body volume. To correct the body volume measurement for the subject's lung volume, the BOD POD utilizes pulmonary plethysmography to measure functional residual capacity (FRC) at midexhalation as that is the subject's lung volume during the body volume measurement. Normally, FRC is measured at endexhalation. The BOD POD FRC measurement can be corrected to an endexhalation volume by subtracting approximately onehalf of the measured tidal volume. Our purpose was to determine the reliability and validity of the BOD POD FRC measurement at endexhalation. Ninetytwo healthy adults (half female) underwent duplicate FRC measurements by the BOD POD and one FRC measurement by a traditional gas dilution technique. The latter method was used as the reference method for the validity component of the study. The order of the FRC measurements by the two methods was randomized. The testretest correlation coefficients for the duplicate BOD POD FRC measurements for the male and female subjects were 0.966 and 0.948, respectively. The mean differences between the BOD POD FRC trial #1 measurement and gas dilution FRC measurement for the male and female subjects were 32 and 23 ml, respectively. Neither difference was statistically significant. The correlation coefficients for these two measurements in the male and female subjects were 0.925 and 0.917, respectively. Based on these results, we conclude that the BOD POD FRC measurement in healthy males and females is both reliable and valid.


Title 
Computed Tomography Calcium Quantification As a Measure of Atherosclerotic Plaque Morphology and Stability. 
Date 
December 2006 
Journal 
Investigative Radiology 
Excerpt 
OBJECTIVES: We examined the relationship between computed tomography (CT)quantified calcium and histopathologic atherosclerotic plaque morphology and rupture. MATERIALS AND METHODS: Seven aortae were harvested from autopsy cases. All were scanned, ex vivo, on a 16slice CT scanner and CT calcium scores (CTCS) were calculated using a Siemens Calcium Scoring package. The aorta segments were physically cross sectioned at 3mm intervals corresponding to CT reconstructions. Two pathologists evaluated the cross sections for histology calcium score (HCS), plaque fibrous cap disruption, overlying thrombus, internal hemorrhage, size, lipid content, and inflammation. CT and histology data were subsequently paired using predetermined quadrant and slice conventions. RESULTS: Three hundred fortynine aorta cross sections yielded 41 atherosclerotic plaques. Eleven plaques demonstrated plaque disruption and thrombosis and all contained calcium. CTCS was not significantly different between atherosclerotic plaques with and without evidence of disruption/thrombosis (F[1,30] = 1.525, P = 0.227). CT was 100% sensitive for nodular calcification, but only 56% (5 of 9 plaques) sensitive for nonnodular calcification. There was no significant relationship between CTCS and intraplaque hemorrhage, lipid content, inflammation, and plaque size (P = 0.179, P = 0.230, P = 0.314, and P = 0.054). There was significant correlation between CTCS and HCS (Pearson coefficient = 0.535; P < 0.01). CONCLUSIONS: Calcium quantity does not appear to predict plaque morphology or likelihood of rupture. CT has lower sensitivity for nonnodular compared with nodular calcification.


Title 
Testretest Reliability for Two Indices of Ventilatory Efficiency Measured During Cardiopulmonary Exercise Testing in Healthy Men and Women. 
Date 
July 2006 
Journal 
Clinical Physiology and Functional Imaging 
Excerpt 
The level of ventilation (VE)) at a given carbon dioxide output (CO2) determines ventilatory efficiency. During cardiopulmonary exercise testing (CPET), ventilatory efficiency can be measured as the slope of the (VE) versus VCO2 relationship or the lowest VE/VCO2. We evaluated the testretest reliability of these two ventilatory efficiency indices in 29 healthy subjects (14 males). Each subject performed duplicate cycle ergometer tests on different days. Ventilation and the gas fractions for oxygen and CO2 were measured with a Vacumed metabolic cart. Linear regression analysis of the VE versus VCO2 slope for the duplicate tests in the males, females, and both sexes combined yielded correlation coefficients of 0.822, 0.942, and 0.910, respectively. The corresponding correlation coefficients for the lowest VE/VCO2 were 0.745, 0.929, and 0.884. A comparison of the testretest correlation coefficients between the two ventilatory efficiency measures for the men, women, and both sexes combined revealed that they were not significantly different and, for a given index, there were no sex differences. The bias (mean of difference scores between tests) and 95% limits of agreement for the VE versus VCO2 slope in the males, females, and both sexes combined were 0.05 +/ 2.41, 0.57 +/ 1.92, and 0.32 +/ 2.20, respectively. The bias and 95% limits of agreement for the lowest VE/VCO2 were very similar with values of 0.06 +/ 2.45, 0.22 +/ 2.03, and 0.10 +/ 2.27. We conclude that the testretest reliability for the VE versus VCO2 slope and the lowest VE/VCO2 is the same and that there is no sex difference in reliability for either index of ventilatory efficiency.


Title 
Is Ventilatory Efficiency Dependent on the Speed of the Exercise Test Protocol in Healthy Men and Women? 
Date 
July 2006 
Journal 
Clinical Physiology and Functional Imaging 
Excerpt 
Indices of ventilatory efficiency have proven useful in assessing patients with heart and lung disease. One of these indices is the slope of the ventilation (V(E)) versus carbon dioxide output (VCO(2)) relationship during cardiopulmonary exercise testing (CPET) for work rates where the relationship is linear. However, this relationship is defined not only by the slope but also by the yintercept. To examine whether this relationship is dependent on the speed of the CPET protocol, 30 healthy subjects (16 males) were administered a rapid CPET with 1min increment duration (1min CPET) to the limit of tolerance and a slow CPET with 4min increment duration (4min CPET) to the lactate threshold. Ventilation and the gas fractions for oxygen and CO(2) were measured with a Vacumed metabolic cart. The average increment size of both protocols for both sexes was not significantly different (P>0.05). For the males, the mean (SD) slope for the 1 and 4min CPET was 20.12 (2.61) and 20.37 (2.41), respectively. The corresponding values for the yintercept were 4..89 (2.08) and 5..10 (2.00) l min(1). For the females, the mean (SD) slope for the 1 and 4min CPET was 23.90 (2.38) and 24.16 (2.55), respectively. The corresponding values for the yintercept were 3.93 (0.39) and 3.77 (0.71) l min(1). Paired ttest analysis demonstrated for both sexes that the slopes and yintercepts were not different for the two protocols (P>0.05). The results of this study demonstrate that the V(E) versus VCO(2) relationship is not dependent on the speed of the CPET protocol.


Title 
Exercise Test Mode Dependency for Ventilatory Efficiency in Women but Not Men. 
Date 
July 2006 
Journal 
Clinical Physiology and Functional Imaging 
Excerpt 
Ventilatory efficiency is commonly defined as the level of ventilation V(E) at a given carbon dioxide output (V(CO(2) )). The slope of the V(E) versus V(CO(2) ) relationship and the lowest V(E)/V(CO(2) ) are two ventilatory efficiency indices that can be measured during cardiopulmonary exercise testing (CPET). A possible CPET mode dependency for these indices was evaluated in healthy men and women. Also evaluated was the relationship between these two indices as, in theory, V(E)/V(CO(2) ) falls hyperbolically towards an asymptote that numerically equals the V(E) versus V(CO(2) ) slope at exercise levels below the ones that cause respiratory compensation for metabolic acidosis. Twentyeight healthy subjects (14 men) underwent treadmill and cycle ergometer CPET on different days. Ventilation and the gas fractions for oxygen and CO(2) were measured with a vacumed metabolic cart. In men, paired ttest analysis failed to find a mode difference for either ventilatory efficiency index but the opposite was true in the women as each woman had higher values for both indices on the treadmill. For men, the lowest V(E)/V(CO(2) ) was larger than the V(E) versus V(CO(2) ) slope by 1.3 on the treadmill and 0.8 on the cycle ergometer. The corresponding values for women were 1.7 and 1.4. We conclude that in healthy subjects, women, but not men, demonstrate a mode dependency for the two ventilatory efficiency indices investigated in this study. Furthermore, our results are consistent with the theoretical expectation that the lowest V(E)/V(CO(2) ) has a numerical value just above the asymptote of the V(E)/V(CO(2) ) versus V(CO(2) ) relationship.


Title 
Maximal Oxygen Uptake at the Same Fatfree Mass is Greater in Men Than Women. 
Date 
May 2006 
Journal 
Clinical Physiology and Functional Imaging 
Excerpt 
Maximal oxygen uptake (VO(2max)) is commonly divided by body mass or fatfree mass (body mass minus fat mass) in order to make it size independent so that comparisons among persons of different size can be made. However, numerous studies have shown that the ratio created is not sizeindependent. Analysis of covariance (ANCOVA) allows a dependent variable to be compared between groups at a common value of a covariate. The purpose of this study was to compare VO(2max) at the same fatfree mass (FFM) in 230 sedentary subjects (half men) who ranged in age from 20 to 70 years. The subjects underwent maximal cardiopulmonary exercise testing on a cycle ergometer as ventilation and the expired gas fractions were being measured. Two ANCOVA models were evaluated. The dependent variable, fixed factor and covariate(s) in the linear model were VO(2max), sex and FFM, respectively. The corresponding terms in the loglinear model were ln VO(2max), sex, and ln FFM and age. Sex made a significant contribution to both models. In the linear model, the mean VO(2max) at the same FFM was 27% higher in men (2,444 versus 1,929 ml min(1); P<0.001). In the loglinear model, the corresponding value at the same FFM and age was 32% higher in men (2,368 versus 1,794 ml min(1); P<0.001). The goodness of fit indices of squared multiple correlation coefficient and standard error of estimate were significantly better for the loglinear model. We conclude that VO(2max) at the same FFM is considerably higher in men than in women who have a sedentary lifestyle.


Title 
Comparison of Stroke Volume Estimation for Nonsteadystate and Steadystate Graded Exercise Testing. 
Date 
July 2005 
Journal 
Clinical Physiology and Functional Imaging 
Excerpt 
The stroke volume (SV) during exercise is an important index of the heart's functional capacity. A new method has been developed for the noninvasive estimation of exercise SV (SVex). It requires the determination of the slope for the oxygen uptake versus heart rate relationship in the steady state of graded exercise testing (GXT). The product of the slope and a constant (reciprocal of an assumed value of the arterial oxygen content) equals an estimated value for SVex. It was validated in a previous study using invasive measurements while subjects were performing steadystate GXT. However, currently the most commonly used GXT protocols are nonsteady state, e.g. protocols with 1min increment durations. We tested the hypothesis that SVex is the same for steadystate and nonsteadystate GXT. A total of 30 subjects (15 males and 15 females) served as subjects for the study. Each subject performed two GXTs on different days with different increment durations  1 and 4 min. Ventilation and gas exchange were measured with the Vacumed metabolic cart. For the male subjects, the mean (SD) SVex values for the 1 and 4min GXTs were 155.4 (39.5) and 134.6 (27.5) ml, respectively. The corresponding values for the female subjects were 151.6 (37.6) and 134.3 (36.4) ml. Paired ttest analysis demonstrated that for both genders the mean SVex for the 1min GXT was significantly larger than the 4min GXT mean value (P<0.05). Hence, the commonly used 1min GXT does not yield the same values for SVex as the steadystate GXT.


Title 
Lower Reference Limit for Maximal Oxygen Uptake in Men and Women. 
Date 
March 2003 
Journal 
Clinical Physiology and Functional Imaging 
Excerpt 
Maximal oxygen uptake (VO2max) is an important measure of exercise tolerance and low values may have clinical significance. Our purpose was to develop the necessary statisticsprediction equations and standard errors of estimate (SEE)so that the lower reference limit for VO2max can be predicted for men and women. The subjects were healthy, nonsmoking, sedentary men (n = 115) and women (n = 115) aged 2070 years who performed 15 W min1 cycle ergometer exercise tests. Three equations were developed for each gender using multiple linear regression with the nonexercise predictor variables of age and height, age and mass, and age and fatfree mass (FFM). The assumptions of regression analysis were examined and the predicted residual sum of squares (PRESS) method was used to crossvalidate each equation. Healthy and diseased individual subject data from the literature were used to externally validate our lower reference limit statistics. The equations developed meet the assumptions of regression analysis and have an accuracy similar to the nonexercise prediction equations in the literature with R2 values of approximately 0.581. The PRESS method revealed that the equations are generalizable, i.e. may be used in future studies without a significant loss of accuracy. The lower reference limit predictions for the healthy and diseased individual subject data from the literature produced few miscategorizations unless the subjects were obese and mass was used as a predictor variable. In conclusion, the equations and SEE generated in this study can be used to predict an accurate and valid VO2max lower reference limit for a given subject.


Title 
Lack of Relationship Between Toxicity and Bone Marrow Cell Colony Stimulating Activity of Endotoxin Preparations. 
Date 
March 1980 
Journal 
Proceedings of the Society for Experimental Biology and Medicine. Society for Experimental Biology and Medicine (new York, N.y.) 

Title 
Largescale Arrays of Singlelayer Graphene Resonators. 
Date 

Journal 
Nano Letters 
Excerpt 
We fabricated large arrays of suspended, singlelayer graphene membrane resonators using chemical vapor deposition (CVD) growth followed by patterning and transfer. We measure the resonators using both optical and electrical actuation and detection techniques. We find that the resonators can be modeled as flat membranes under tension, and that clamping the membranes on all sides improves agreement with our model and reduces the variation in frequency between identical resonators. The resonance frequency is tunable with both electrostatic gate voltage and temperature, and quality factors improve dramatically with cooling, reaching values up to 9000 at 10 K. These measurements show that it is possible to produce large arrays of CVDgrown graphene resonators with reproducible properties and the same excellent electrical and mechanical properties previously reported for exfoliated graphene.

