Severe malaria in Cameroonian children: correlation between plasma levels of three soluble inducible adhesion molecules and TNF-alpha.

Plasma levels of three soluble inducible adhesion molecules, namely: intercellular adhesion molecule-1 (sICAM-1), vascular cell adhesion molecule-1 (sVCAM-1) and endothelial leucocyte adhesion molecule-1 (sELAM-1) or sE-selectin and the pro-inflammatory cytokine, tumour necrosis factor-alpha (TNF-alpha) were measured in well-defined clinical groups of children with severe and uncomplicated malaria. The goal of the study was to investigate the role of these molecules in immunopathogenic processes associated with severe malaria in Cameroonian children. Results showed significantly increased plasma concentrations of sICAM-1, sVCAM-1 and sE-selectin in children with severe malaria compared to those with uncomplicated malaria and healthy children (P<0.001). TNF-alpha levels increased significantly in children with severe malaria, approximately 2-folds compared to those with uncomplicated malaria and about 3-folds compared to healthy children (P<0.001). More importantly, levels of TNF-alpha strongly correlated with those of the three adhesion molecules and were significantly associated with increased risk of death (P=0.03). In addition, children who died from severe malaria showed higher mean levels of all measured factors compared to those who recovered, with significant differences observed with sICAM-1 (P<0.001) and sE-selectin (P=0.002). Furthermore, children with severe malarial anemia relative to those without, showed significantly elevated levels of the three soluble molecules; and sICAM-1 was significantly associated with increased risk of severe anemia. Taken together, these results confirm the role of TNF-alpha and the three adhesion molecules in pathogenic processes associated with severe malaria in children, and suggest an association between sICAM-1 and severe malarial anemia.

Multiplex assay for simultaneous measurement of antibodies to multiple Plasmodium falciparum antigens.

Antibodies to Plasmodium falciparum are classically measured using the enzyme-linked immunosorbent assay (ELISA). Although highly sensitive, this technique is labor-intensive when large numbers of samples must be screened against multiple antigens. The suspension array technology (SAT) might be an alterative to ELISA, as it allows measurement of antibodies against multiple antigens simultaneously with a small volume of sample. This study sought to adapt the new SAT multiplex system for measuring antibodies against nine malarial vaccine candidate antigens, including recombinant proteins from two variants of merozoite surface protein 1, two variants of apical merozoite antigen 1, erythrocyte binding antigen 175, merozoite surface protein 3, and peptides from the circumsporozoite protein, ring erythrocyte surface antigen, and liver-stage antigen 1. Various concentrations of the antigens were coupled to microspheres with different spectral addresses, and plasma samples from Cameroonian adults were screened by SAT in mono- and multiplex formats and by ELISA. Optimal amounts of protein required to perform the SAT assay were 10- to 100-fold less than that needed for ELISA. Excellent agreement was found between the single and multiplex formats (R > or = 0.96), even when two variants of the same antigen were used. The multiplex assay was rapid, reproducible, required less than 1 mul of plasma, and had a good correlation with ELISA. Thus, SAT provides an important new tool for studying the immune response to malaria rapidly and efficiently in large populations, even when the amount of plasma available is limited, e.g., in studies of neonates or finger-prick blood.

Malaria in pregnant Cameroonian women: the effect of age and gravidity on submicroscopic and mixed-species infections and multiple parasite genotypes.

Polymerase chain reaction (PCR)-based methods were used to investigate malaria in pregnant women residing in Yaounde, Cameroon. Microscopy and species-specific PCR-based diagnosis show that at delivery 82.4% of the women were infected with Plasmodium falciparum (27.5% blood-smear positive and 54.9% submicroscopic infections). The prevalence of P. malariae and P. ovale was 7.6% and 2.5%, respectively, with 9.4% infected with more than one species. Based on genotyping of the merozoite surface protein 1 (msp-1) and msp-2 alleles, the mean number of genetically different P. falciparum parasites in peripheral blood was 3.4 (range = 1-9) and 3.5 (range 1-8) in the placenta. Plasmodium falciparum detected by microscopy and PCR as well as mixed-species infections were significantly higher in women < or = 20 years old and paucigravidae, but maternal anemia was associated only with microscopic detection of parasites. Neither submicroscopic infections nor number of parasite genotypes decreased significantly with age or gravidity. Thus, pregnancy-associated immunity helps reduce malaria to submicroscopic levels, but does not reduce the number of circulating parasite genotypes.

Risk factors for placental malaria and its effect on pregnancy outcome in Yaounde, Cameroon.

Between 1996 and 2001, the prevalence of placental malaria in pregnant women living in Yaounde, Cameroon and its effect on pregnancy outcome were evaluated with respect to gravidity and maternal age. Results showed that 19.9% of the women had placental malaria at delivery. After adjusting for relevant covariates, the major risk factor for placental malaria was an age < 25 years old. Placental malaria significantly increased the prevalence of anemia in women regardless of gravidity or age. In addition, the mean infant birth weight was lower and the percentage of pre-term deliveries (PTDs) and low birth weight (LBW) babies were higher in primigravidae and women < 20 years of age who had placental malaria. However, in a multivariate regression model taking relevant covariates into consideration, the major risk factor for PTDs was maternal anemia, and maternal anemia as well as first and second pregnancies were important risk factors for LBW babies.

Parental cultural perspectives in relation to weight-related behaviors and concerns of African-American girls.

OBJECTIVE: To determine whether cultural perspectives of parents may influence children's eating and physical activity behaviors and patterns of weight gain. RESEARCH METHODS AND PROCEDURES: African-American girls (ages 8 to 10 years) and their parents (or caregivers) (n = 210) participated at one of four Girls Health Enrichment Multisite Studies Phase 1 Field Centers. At baseline, parents completed questionnaires adapted from the African-American Acculturation Scale (AAAS), the Multiethnic Identity Scale (MEIS), and an original question on Global Cultural Identity. Girls' baseline measures included physical activity assessment by accelerometer, 24-hour dietary recalls, and questionnaires about body image and weight concerns. RESULTS: Principal components analysis indicated the expected AAAS and MEIS factor structures, with moderate to good internal consistency (Cronbach's alpha = 0.61 to 0.82) and some intercorrelation among these measures (r = 0.17 to 0.57). Overall mean (SD) AAAS subscale scores of 4.1 (2.1) and 5.5 (1.8) of a possible 7 and 3.0 (0.9) of a possible 4 on the MEIS indicated, respectively, moderate to high levels of parental African-American cultural orientation and identity with moderate variability. Parental AAAS and MEIS scores were inversely correlated with girls' body image discrepancy and weight concern. One AAAS subscale was positively associated with total energy intake and percentage energy from fat. Overall, however, parental AAAS and MEIS scores were unrelated or inconsistently related to girls' physical activity and diet measures. DISCUSSION: The AAAS and MEIS measures had acceptable psychometric properties, except for weight concern, but did not give a consistent picture of how parental perspectives related to the girls' baseline attitudes and behaviors.

Lack of an association between antibodies to Plasmodium falciparum glycosylphosphatidylinositols and malaria-associated placental changes in Cameroonian women with preterm and full-term deliveries.

Sequestration of Plasmodium falciparum parasites within the placenta often leads to an accumulation of macrophages within the intervillous space and increased production of tumor necrosis factor alpha (TNF-alpha), a cytokine associated with placental pathology and poor pregnancy outcomes. P. falciparum glycosylphosphatidylinositol (GPI) anchors have been shown to be the major parasite component that induces TNF-alpha production by monocytes and macrophages. Antibodies against P. falciparum GPI (anti-PfGPI), however, can inhibit the induction of TNF-alpha and inflammation. Thus, the study was undertaken to determine whether anti-PfGPI antibodies down-regulate inflammatory-type changes in the placentas of women with malaria. Anti-PfGPI immunoglobulin M (IgM) and IgG levels were measured in 380 pregnant women with or without placental malaria, including those who delivered prematurely and at term. Results showed that anti-PfGPI antibody levels increased with gravidity and age and that malaria infection boosted anti-PfGPI antibodies in pregnant women. However, no association was found between anti-PfGPI antibodies and placental TNF-alpha levels or the presence of acute or chronic placental malaria. Furthermore, anti-PfGPI antibody levels were similar in women with preterm and full-term deliveries and were not associated with an increase in infant birth weight. Thus, these results fail to support a strong role for anti-PfGPI antibodies in the prevention of chronic placental malaria infections and malaria-associated poor birth outcomes.

Measurement characteristics of diet-related psychosocial questionnaires among African-American parents and their 8- to 10-year-old daughters: results from the Girls' health Enrichment Multi-site Studies.

OBJECTIVE: This paper presents the reliability and validity of several diet-related psychosocial questionnaires. METHODS: At baseline and 12 weeks follow-up, parents/caregivers of one hundred fifty 8- to 10-year-old African-American completed questionnaires on food preparation habits for their daughter, perceived home barriers to healthy eating, and fruit, juice, vegetable (FJV), low-fat and high-fat food availability. Girls completed a sweetened beverage preferences questionnaire and two 24-h dietary recalls to assess intake. Principal components analyses were conducted for two newly designed measures. Internal consistency was calculated and construct validity was assessed between the psychosocial scales and obesity-related dietary variables. RESULTS: Low-fat and high-fat food preparation for daughters, and perceived home barriers to eating low-fat food and FJV subscales were derived from the new questionnaires. Internal consistency reliabilities were moderate (0.58) to substantial (0.80) across all new and existing scales. Test-retest reliabilities were moderate (0.44) to substantial (0.79). Girls' intake of fat as a percentage of energy was positively related to parental high-fat food preparation for daughters (P < 0.01) and negatively related to parental low-fat food preparation practices for daughters (P < 0.05). CONCLUSIONS: Measures of family influences on FJV, fat, and sweetened beverage consumption were internally consistent with moderate to substantial stability. Scales for low-fat and high-fat food preparation practices for daughters achieved construct validity with fat consumption in the hypothesized direction. Family food preparation habits appear to be important targets for future interventions.

Physical activity self-report and accelerometry measures from the Girls health Enrichment Multi-site Studies.

BACKGROUND: Valid and reliable physical activity checklists are needed to assess effectiveness of interventions. This study tested the validity and reliability of the Girls health Enrichment Multi-site Studies (GEMS) Activity Questionnaire. METHODS: Two-hundred and ten African-American girls completed the GEMS Activity Questionnaire (GAQ), a checklist of 28 physical and 7 sedentary activities, including TV viewing, inquiring whether performed on the previous day ("yesterday"), and whether usually performed ("usual"). The girls wore an accelerometer (used as the criterion for validity) for three consecutive days at baseline and after a 12-week pilot intervention. Data from 172 girls at baseline and follow-up were usable for this report. RESULTS: Girls were (mean +/- SD) 8.8 +/- 0.8 years old with a BMI of 22.3 +/- 5.9 kg/m2. The intraclass correlation (ICC) examining reliability for the accelerometer across 3 days at baseline for the combined group was fair (ICC = 0.33, P < 0.21). The test-retest reliability coefficient for the 18-item MET-weighted GAQ yesterday scores for the comparison group of girls was 0.57 (P < 0.001). At baseline and follow-up, nonsignificant correlations were observed between 3-day accelerometer counts/minute and GAQ 18-item usual score for both comparison and intervention groups. A significant correlation was found between change in accelerometer minutes of moderate-to-vigorous activity (MVPA) between 12 noon and 6 PM and change in GAQ physical activities in the comparison girls (R = 0.35, P < 0.01). The TV-usual score was correlated with 3-day accelerometer counts/minute (R = -0.19, P = 0.02) at baseline for the total sample. CONCLUSIONS: Correlations between the GAQ and accelerometer were low, indicating low validity. Although the GAQ may be helpful in describing types of physical activities performed, it needs further development to improve its psychometric properties.

Measurement characteristics of weight concern and dieting measures in 8-10-year-old African-American girls from GEMS pilot studies.

BACKGROUND: Reliability and validity were established for weight concern measures completed by 8-10-year-old African-American girls participating in a pilot obesity prevention program. METHODS: Two hundred ten girls and parents participated in the program. Girls completed subscales of the McKnight Risk Factor Survey (MRFS) and body silhouette ratings, had height, weight, and body fat measured, wore accelerometers for 3 days, and completed two dietary recalls. Principal components analysis, internal consistency, and test-retest reliability were computed for weight concerns and body image measures along with convergent validity with body mass index (BMI), percent body fat (PBF), physical activity, and dietary intake. RESULTS: A Moderate Weight Control Behaviors (MWCB) subscale was derived from the MRFS. Overconcern with Weight and Shape (OWS) was a stand-alone scale. Internal consistency estimates for the scales were substantial ranging from 0.71 to 0.84. Test-retest reliabilities were moderate (0.45-0.58). OWS, MWCB, body silhouette rating, and body size discrepancy were positively associated with BMI and PBF. The "like to look" silhouette rating was negatively associated with PBF. CONCLUSIONS: Weight concern measures had reasonable levels of internal consistency and promising validity, but only moderate test-retest reliability among preadolescent African-American girls. Refinement and further validation of weight concern measures in this population are warranted.

Measurement characteristics of activity-related psychosocial measures in 8- to 10-year-old African-American girls in the Girls Health Enrichment Multisite Study (GEMS).

BACKGROUND: This paper presents reliability and validity analyses of physical activity-related psychosocial questionnaires completed by 8- to 10-year-old African-American girls at baseline and follow-up assessments of pilot intervention studies in the Girls health Enrichment Multi-site Study (GEMS). METHODS: Two hundred ten girls participated in the GEMS 12-week pilot studies and had their height and weight measured, wore an accelerometer for 3 days and completed a measure of their usual physical activity (PA) at baseline and after the 12-week intervention. Subgroups of girls also completed physical activity-related psychosocial measures at these two time points including: (a) self-concept; (b) self-efficacy; (c) outcome expectancies; and d) preferences. Principal components analysis was conducted on the psychosocial measures obtained at baseline. Cronbach's alpha and test-retest reliability were computed. Convergent validity was assessed by correlating the baseline psychosocial measures with baseline physical activity measures and body mass index (BMI). RESULTS: The following sub-scales were derived: Activity Preference, Positive Expectancies and Negative Expectancies for physical activity. Physical Performance Self-Concept and Self-Efficacy for physical activity were kept as single dimensional scales. Sub-scales, derived from principal components analyses, were Activity Preference, Positive Expectancies, and Negative Expectancies for physical activity. Internal consistency estimates for the various scales were substantial to excellent (0.67-0.85), while test-retest reliability estimates were fair to moderate (0.22-0.56). Correlations between the PA psychosocial sub-scales and measured levels of activity measures showed evidence of convergent validity for the Activity Preference sub-scale, although social desirability may have influenced the significant associations observed. CONCLUSIONS: The Activity Preference was a fairly reliable and valid measure. Further studies are needed to examine the utility of activity-related psychosocial measures in interventions to increase physical activity among preadolescent African-American girls.

Psychometric properties of optimism and pessimism: results from the Girls' Health Enrichment Multisite Studies.

UNLABELLED: Background. This study investigated the relationships among optimism, pessimism, physical activity, and dietary behaviors among 8- to 10-year-old African-American girls in the Girls' Health Enrichment Multisite Studies (GEMS). Methods. Ninety-two girls were randomly assigned to a 12-week physical activity and diet intervention or comparison group and completed psychosocial assessments including the Youth Life Orientation Test (YLOT) of optimism and pessimism, physical performance self-concept, physical activity self-efficacy, physical activity outcome expectancies, and physical activity and sedentary preferences. Also, preferences for bottled water and sweetened beverages were assessed. Physical activity and dietary intake were assessed by self-report and parent-report at baseline and follow-up. Also, physical activity was objectively assessed by wearing an accelerometer for 3 days. The psychometric properties of the optimism-pessimism subscales were analyzed. Results. The measures of optimism and pessimism in children were reliable (r = 0.75-0.82). In the multiple regression analyses without the intervention interaction terms, pessimism was positively and significantly related to increases in MET-adjusted usual activity (P = 0.008) and sedentary behaviors (P = 0.0004). Additionally, a negative (P = 0.026) pessimism by intervention interaction term for MET-adjusted usual activity was found such that the intervention group had a lower change in physical activity per unit increase in pessimism compared to the control group. CONCLUSIONS: Among 8- to 10-year-old African-American girls, pessimism was related to increased sedentary behaviors and usual activity. Previous studies have reported relationships between optimism and health-compromising behaviors. This study found that pessimism may positively or negatively influence efforts to increase health-promoting behaviors. Future research should confirm and clarify the meaning of these findings.

Human leukocyte antigen class II alleles influence levels of antibodies to the Plasmodium falciparum asexual-stage apical membrane antigen 1 but not to merozoite surface antigen 2 and merozoite surface protein 1.

The apical membrane antigen 1 (AMA1), merozoite surface antigen 2 (MSA2), and merozoite surface protein 1 (MSP1) are asexual-stage proteins currently being evaluated for inclusion in a vaccine for Plasmodium falciparum. Accordingly, it is important to understand factors that control antibody responses to these antigens. Antibody levels in plasma from residents of Etoa, Cameroon, between the ages of 5 and 70 years, were determined using recombinant AMA1, MSA2, and the N-terminal region of MSP1 (MSP1-190L). In addition, antibody responses to four variants of the C-terminal region of MSP1 (MSP1(19)) were assessed. Results showed that all individuals produced antibodies to AMA1, MSA2, and MSP1-190L; however, a proportion of individuals never produced antibodies to the MSP1(19) variants, although the percentage of nonresponders decreased with age. The influence of age and human leukocyte antigen (HLA)-DRB1/DQB1 alleles on antibody levels was evaluated using two-way analysis of variance. Age was correlated with levels of antibodies to AMA1 and MSP1(19) but not with levels of antibodies to MSA2 and MSP1-190L. No association was found between a single HLA allele and levels of antibodies to MSA2, MSP1-190L, or any of the MSP1(19) variants. However, individuals positive for DRB1*1201 had higher levels of antibodies to the variant of recombinant AMA1 tested than did individuals of all other HLA types. Since the effect was seen across all age groups, HLA influenced the level but not the rate of antibody acquisition. This association for AMA1, combined with the previously reported association between HLA class II alleles and levels of antibodies to rhoptry-associated protein 1 (RAP1) and RAP2, indicates that HLA influences the levels of antibodies to three of the five vaccine candidate antigens that we have evaluated.

Changes in the levels of chemokines and cytokines in the placentas of women with Plasmodium falciparum malaria.

Plasmodium falciparum-infected erythrocytes often are sequestered in the placenta and stimulate the accumulation of maternal mononuclear cells. In this study, the role that chemokines and cytokines play in mediating the inflammatory response was investigated. Placental parasites elicited a statistically significant increase in the levels of interferon (IFN)-gamma, tumor necrosis factor (TNF)-alpha, and interleukin (IL)-10, in plasma collected from the intervillous space. Explants of fetal tissue from malaria-positive placentas also secreted significantly enhanced amounts of IFN-gamma. Culture supernatant of maternal intervillous leukocytes obtained from infected placentas contained significantly higher levels of TNF-alpha, IL-10, monocyte chemotactic protein-1, macrophage inflammatory protein (MIP)-1alpha, MIP-1beta, and IFN-gamma inducible protein-10 than did cultures of white blood cells obtained from uninfected placentas. Taken together, these results show that both fetal and maternal cells secrete inflammatory and immunoregulatory cytokines in response to P. falciparum and suggest that beta-chemokines produced by maternal cells contribute to the accumulation of macrophages in the intervillous space.

Validity and reliability of activity measures in African-American girls for GEMS.

PURPOSE: To determine the reliability and validity of physical activity monitors and self-report instruments suitable for young African-American girls. METHODS: A validation study was conducted by the Girls health Enrichment Multi-site Studies (GEMS) research team to compare an accelerometer with a pedometer and two self-report instruments for assessing physical activity in African-American girls, age 8-9 yr. Girls (N= 68) attended two clinic visits spaced 4 d apart. Each girl wore a MTI/CSA accelerometer (used as the criterion standard for validity) and a pedometer simultaneously for four consecutive days. Girls completed on two occasions a 24-h physical activity checklist of yesterday and usual activities, including sedentary activities (GEMS Activity Questionnaire, GAQ), and a 3-d computerized self-report instrument (Activitygram). RESULTS: Girls were (mean +/- SD) 9.0 +/- 0.6 yr old and had a body mass index of 19.4 kg x m. Reliability measured by intraclass correlations (ICC) and Pearson correlation coefficients (r) were calculated for the MTI/CSA (ICC = 0.37, P< 0.0001), pedometer (ICC = 0.08, = 0.094), Activitygram (ICC = 0.24) (P = 0.005), and GAQ for physical (r = 0.80, P< 0.0001) and sedentary (r = 0.3-0.5, P< 0.005) activities. Significant Pearson correlations between the MTI/CSA and the other instruments, as a measure of validity, were observed for the 4-d average pedometer score (r = 0.47, P< 0.0001), 3-d average Activitygram score (r = 0.37, P= 0.002), and the average of the two yesterday and two usual GAQ activity scores for a subset of 18 physical activities questions (r = 0.27, = 0.03; and r = 0.29,P = 0.02, respectively). The MTI/CSA was uncorrelated with single day scores from the three other instruments. CONCLUSION: The reliability of the instruments tested was acceptable, except the pedometer. Validity correlations were significant when more than one day was used. Self-report instruments need further development for improved reliability and validity.

Congenital exposure to Plasmodium falciparum antigens: prevalence and antigenic specificity of in utero-produced antimalarial immunoglobulin M antibodies.

Congenital Plasmodium falciparum malaria in newborns is uncommon in sub-Saharan Africa. A significant number of infants, however, become infected or exposed to malarial antigens either in utero or at delivery and have the potential to produce antimalarial antibodies and memory cells before their first natural infection. In Yaounde, Cameroon, parasite-specific immunoglobulin M (IgM) was detected in 14% of cord blood samples. The IgM antibodies reacted with a wide range of asexual-stage antigens, with each newborn having its own unique pattern of IgM reactivity. PCR-based detection and genotyping of cord blood parasites found that the prevalence, total number of parasite genotypes, and complexity of infection were higher in newborns who had produced antimalarial IgM than those who had not. Maternal placental malaria and anemia were associated with the production of P. falciparum-specific IgM by the fetus. The effect of early immune priming on acquisition of immunity by infants is unknown and merits further investigation, since a significant proportion of Cameroonian newborns developed a humoral response to malaria before birth.

Malaria-associated cytokine changes in the placenta of women with pre-term deliveries in Yaounde, Cameroon.

The prevalence of pre-term deliveries (PTDs) is increased in women who become infected with Plasmodium falciparum during pregnancy. Because prematurity is a risk factor for newborns, it is important to identify conditions that contribute to malaria-associated PTDs. Plasmodium falciparum-infected erythrocytes sequester in the placenta and attract activated mononuclear cells that secrete pro-inflammatory cytokines. Increased inflammatory cytokine levels in other microbial infections are associated with PTDs. To determine if such is the case in women with placental malaria, concentrations of interferon-gamma (IFN-gamma), tumor necrosis factor-alpha (TNF-alpha), interleukin-4 (IL-4), and IL-10 were measured in placental plasma of 391 malaria-infected and -uninfected Cameroonian women with premature and full-term deliveries. Risk factors for malaria-associated PTDs included peripheral and placental parasitemias greater than 1%, maternal anemia, elevated IL-10 levels, and low TNF-alpha:IL-10 ratios due to over-expression of IL-10. Alterations in cytokine levels may contribute to PTDs through the induction of anemia and/or altering cellular immune responses required for eliminating placental parasites.

Prevalence of Plasmodium falciparum infection in pregnant Cameroonian women.

Between 1995 and 1998, a longitudinal study was conducted at 2 health centers in Cameroon to determine the prevalence of Plasmodium falciparum infection in pregnant women. There were 520 pregnant women enrolled at a rural clinic in the village of Etoa and 199 enrolled at the Biyem Assi Clinic in the city of Yaounde. In Etoa, pregnant women were younger, fewer took prophylaxis, and malaria prevalence was higher compared with pregnant women in the Biyem Assi Clinic. The prevalence of malaria infection peaked during the fourth month of pregnancy at both sites. Age was identified as a major risk factor because women < or = 20 years old were 1.8 and 3.4 times more likely to have malaria infection than women > 20 years old at Etoa and the Biyem Assi Clinic after adjusting for gravidae and other factors. The use of chemoprophylaxis and seasonality did not have a major effect.