The association between physical activity and cognition in men with and without HIV infection.

OBJECTIVES: HIV-associated neurocognitive disorders are highly prevalent, and physical activity (PA) is a modifiable behaviour that may affect neurocognitive function. Our objective was to determine the association between PA and neurocognitive function and the effect of HIV on this association. METHODS: PA was assessed in the Multicenter AIDS Cohort Study with the International Physical Activity Questionnaire. A neuropsychological test battery assessed global impairment and domain-specific impairment (executive function, speed of processing, working memory, learning, memory, and motor function) every 2 years. Semiannually, the Symbol Digit Modalities Test and Trail Making Test Parts A and B were performed. Adjusted logistic regression models were used to assess the PA-neurocognitive function association. Using longitudinal data, we also assessed the PA category-decline of neurocognitive function association with multivariate simple regression. RESULTS: Of 601 men, 44% were HIV-infected. Low, moderate, and high PA was reported in 27%, 25%, and 48% of the HIV-infected men vs. 19%, 32% and 49% of the HIV-uninfected men, respectively. High PA was associated with lower odds of impairment of learning, memory, and motor function [odds ratio (OR) ranging from 0.52 to 0.57; P < 0.05 for all]. The high PA-global impairment association OR was 0.63 [95% confidence interval (CI) 0.39, 1.02]. Among HIV-infected men only, across multiple domains, the high PA-impairment association was even more pronounced (OR from 0.27 to 0.49). Baseline high/moderate PA was not associated with decline of any domain score over time. HIV infection was marginally associated with a higher speed of decline in motor function. CONCLUSIONS: A protective effect of high PA on impairment in neurocognitive domains was observed cross-sectionally. Longitudinal PA measurements are needed to elucidate the PA-neurocognitive function relationship over time.

Neural correlates of working memory training in HIV patients: study protocol for a randomized controlled trial.

BACKGROUND: Potent combined antiretroviral therapy decreased the incidence and severity of HIV-associated neurocognitive disorders (HAND); however, no specific effective pharmacotherapy exists for HAND. Patients with HIV commonly have deficits in working memory and attention, which may negatively impact many other cognitive domains, leading to HAND. Since HAND may lead to loss of independence in activities of daily living and negative emotional well-being, and incur a high economic burden, effective treatments for HAND are urgently needed. This study aims to determine whether adaptive working memory training might improve cognitive functions and neural network efficiency and possibly decrease neuroinflammation. This study also aims to assess whether subjects with the LMX1A-rs4657412 TT(AA) genotype show greater training effects from working memory training than TC(AG) or CC(GG)-carriers. METHODS/DESIGN: 60 HIV-infected and 60 seronegative control participants will be randomized to a double-blind active-controlled study, using adaptive versus non-adaptive Cogmed Working Memory Training® (CWMT), 20-25 sessions over 5-8 weeks. Each subject will be assessed with near- and far-transfer cognitive tasks, self-reported mood and executive function questionnaires, and blood-oxygenation level-dependent functional MRI during working memory (n-back) and visual attention (ball tracking) tasks, at baseline, 1-month, and 6-months after CWMT. Furthermore, genotyping for LMX1A-rs4657412 will be performed to identify whether subjects with the TT(AA)-genotype show greater gain or neural efficiency after CWMT than those with other genotypes. Lastly, cerebrospinal fluid will be obtained before and after CWMT to explore changes in levels of inflammatory proteins (cytokines and chemokines) and monoamines. DISCUSSION: Improving working memory in HIV patients, using CWMT, might slow the progression or delay the onset of HAND. Observation of decreased brain activation or normalized neural networks, using fMRI, after CWMT would lead to a better understanding of how neural networks are modulated by CWMT. Moreover, validating the greater training gain in subjects with the LMX1A-TT(AA) genotype could lead to a personalized approach for future working memory training studies. Demonstrating and understanding the neural correlates of the efficacy of CWMT in HIV patients could lead to a safe adjunctive therapy for HAND, and possibly other brain disorders. TRIAL REGISTRATION: ClinicalTrial.gov, NCT02602418.

FLI1 polymorphism affects susceptibility to cutaneous leishmaniasis in Brazil.

Mapping murine genes controlling cutaneous leishmaniasis (CL) identified Fli1 as a candidate influencing resistance to L. major and enhanced wound healing. We examine FLI1 as a gene controlling CL and mucosal leishmaniasis (ML) caused by L. braziliensis in humans. Intron 1 single nucleotide polymorphisms tagging promoter and enhancer elements were analysed in 168 nuclear families (250 CL; 87 ML cases) and replicated in 157 families (402 CL; 39 ML cases). Robust case-pseudocontrol logistic regression analysis showed association between allele C (odds ratio (OR) 1.65; 95% confidence interval 1.18-2.29; P=0.003) of FLI1_rs7930515 and CL in the primary sample that was confirmed (OR 1.60; 95% confidence interval 1.10-2.33; P=0.014) in the replication set (combined P=1.8 × 10(-4)). FLI1_rs7930515 is in linkage disequilibrium with the functional GAn microsatellite in the proximal promoter. Haplotype associations extended across the enhancer, which was not polymorphic. ML associated with inverse haplotypes compared with CL. Wound healing is therefore important in CL, providing potential for therapies modulating FLI1.

Increased furfural tolerance due to overexpression of NADH-dependent oxidoreductase FucO in Escherichia coli strains engineered for the production of ethanol and lactate.

Furfural is an important fermentation inhibitor in hemicellulose sugar syrups derived from woody biomass. The metabolism of furfural by NADPH-dependent oxidoreductases, such as YqhD (low K(m) for NADPH), is proposed to inhibit the growth and fermentation of xylose in Escherichia coli by competing with biosynthesis for NADPH. The discovery that the NADH-dependent propanediol oxidoreductase (FucO) can reduce furfural provided a new approach to improve furfural tolerance. Strains that produced ethanol or lactate efficiently as primary products from xylose were developed. These strains included chromosomal mutations in yqhD expression that permitted the fermentation of xylose broths containing up to 10 mM furfural. Expression of fucO from plasmids was shown to increase furfural tolerance by 50% and to permit the fermentation of 15 mM furfural. Product yields with 15 mM furfural were equivalent to those of control strains without added furfural (85% to 90% of the theoretical maximum). These two defined genetic traits can be readily transferred to enteric biocatalysts designed to produce other products. A similar strategy that minimizes the depletion of NADPH pools by native detoxification enzymes may be generally useful for other inhibitory compounds in lignocellulosic sugar streams and with other organisms.

Effect of reduced sulfur compounds on the fermentation of phosphoric acid pretreated sugarcane bagasse by ethanologenic Escherichia coli.

The addition of reduced sulfur compounds (thiosulfate, cysteine, sodium hydrosulfite, and sodium metabisulfite) increased growth and fermentation of dilute acid hydrolysate of sugarcane bagasse by ethanologenic Escherichia coli (strains LY180, EMFR9, and MM160). With sodium metabisulfite (0.5mM), toxicity was sufficiently reduced that slurries of pretreated biomass (10% dry weight including fiber and solubles) could be fermented by E. coli strain MM160 without solid-liquid separation or cleanup of sugars. A 6-h liquefaction step was added to improve mixing. Sodium metabisulfite also caused spectral changes at wavelengths corresponding to furfural and soluble products from lignin. Glucose and cellobiose were rapidly metabolized. Xylose utilization was improved by sodium metabisulfite but remained incomplete after 144 h. The overall ethanol yield for this liquefaction plus simultaneous saccharification and co-fermentation process was 0.20 g ethanol/g bagasse dry weight, 250 L/tonne (61 gal/US ton).

Simplified process for ethanol production from sugarcane bagasse using hydrolysate-resistant Escherichia coli strain MM160.

Hexose and pentose sugars from phosphoric acid pretreated sugarcane bagasse were co-fermented to ethanol in a single vessel (SScF), eliminating process steps for solid-liquid separation and sugar cleanup. An initial liquefaction step (L) with cellulase was included to improve mixing and saccharification (L+SScF), analogous to a corn ethanol process. Fermentation was enabled by the development of a hydrolysate-resistant mutant of Escherichia coli LY180, designated MM160. Strain MM160 was more resistant than the parent to inhibitors (furfural, 5-hydroxymethylfurfural, and acetate) formed during pretreatment. Bagasse slurries containing 10% and 14% dry weight (fiber plus solubles) were tested using pretreatment temperatures of 160-190°C (1% phosphoric acid, 10 min). Enzymatic saccharification and inhibitor production both increased with pretreatment temperature. The highest titer (30 g/L ethanol) and yield (0.21 g ethanol/g bagasse dry weight) were obtained after incubation for 122 h using 14% dry weight slurries of pretreated bagasse (180°C).

Evidence for associations between the purinergic receptor P2X(7) (P2RX7) and toxoplasmosis.

Congenital Toxoplasma gondii infection can result in intracranial calcification, hydrocephalus and retinochoroiditis. Acquired infection is commonly associated with ocular disease. Pathology is characterized by strong proinflammatory responses. Ligation of ATP by purinergic receptor P2X(7), encoded by P2RX7, stimulates proinflammatory cytokines and can lead directly to killing of intracellular pathogens. To determine whether P2X(7) has a role in susceptibility to congenital toxoplasmosis, we examined polymorphisms at P2RX7 in 149 child/parent trios from North America. We found association (FBAT Z-scores +/-2.429; P=0.015) between the derived C(+)G(-) allele (f=0.68; OR=2.06; 95% CI: 1.14-3.75) at single-nucleotide polymorphism (SNP) rs1718119 (1068T>C; Thr-348-Ala), and a second synonymous variant rs1621388 in linkage disequilibrium with it, and clinical signs of disease per se. Analysis of clinical subgroups showed no association with hydrocephalus, with effect sizes for associations with retinal disease and brain calcifications enhanced (OR=3.0-4.25; 0.004

Genetic changes that increase 5-hydroxymethyl furfural resistance in ethanol-producing Escherichia coli LY180.

The ability of a biocatalyst to tolerate furan inhibitors present in hemicellulose hydrolysates is important for the production of renewable chemicals. This study shows EMFR9, a furfural-tolerant mutant of ethanologenic E. coli LY180, has also acquired tolerance to 5-hydroxymethyl furfural (5-HMF). The mechanism of action of 5-HMF and furfural appear similar. Furan tolerance results primarily from lower expression of yqhD and dkgA, two furan reductases with a low K(m) for NADPH. Furan tolerance was also increased by adding plasmids encoding a NADPH/NADH transhydrogenase (pntAB). Together, these results support the hypothesis that the NADPH-dependent reduction of furans by YqhD and DkgA inhibits growth by competing with biosynthesis for this limiting cofactor.

Interleukin 10 gene polymorphisms and development of post kala-azar dermal leishmaniasis in a selected sudanese population.

BACKGROUND: Post kala-azar dermal leishmaniasis (PKDL) is a cutaneous form of disease that develops at variable times after individuals have received treatment for clinical visceral leishmaniasis (VL). The study aimed to investigate the possible role of interleukin 10 (IL-10) and development of PKDL. METHODS: 77 families composed of 41 complete case-parent trios and 36 case-parent pairs from the Masalit ethnic group were genotyped for 3 IL10 promoter polymorphisms: -1082A/G, -819C/T and -592C/A. RESULTS: Single point analysis using the transmission disequilibrium test showed no evidence of association between any of these IL10 promoter single nucleotide polymorphisms (SNPs) and development of PKDL. Haplotype analysis performed using TRANSMIT showed borderline significance between PKDL and the haplotype AA across -592C/A and -1082A/G (p = 0.053). Haplotypes GCC (0.33) and ATA (0.30) were the common haplotypes in this Sudanese population. Allele frequencies for the 3 SNPs differed significantly in Sudan compared to other African (Gambian, Malawian, YRI) populations. CONCLUSION: There is no evidence for an association between 3 SNPs in the IL10 gene promoter and susceptibility to PKDL in the Masalit ethnic group in Sudan, although some evidence for haplotype association was observed.

Vascular risk factors, HIV serostatus, and cognitive dysfunction in gay and bisexual men.

BACKGROUND: The purpose of this study was to evaluate the relationship between cognitive performance, risk factors for cardiovascular and cerebrovascular disease (CVD), and HIV infection in the era of highly active antiretroviral therapy. METHODS: We evaluated the cognitive functions of men enrolled in the cardiovascular disease substudy of the Multicenter AIDS Cohort Study who were aged > or =40 years, with no self-reported history of heart disease or cerebrovascular disease. Results from comprehensive neuropsychological evaluations were used to construct composite scores of psychomotor speed and memory performance. Subclinical CVD was assessed by measuring coronary artery calcium and carotid artery intima-media thickness (IMT), as well as laboratory measures, including total cholesterol, fasting glucose, glycosylated hemoglobin, glomerular filtration rate (estimated), and standardized blood pressure and heart rate measures. RESULTS: After accounting for education, depression, and race, carotid IMT and glomerular filtration rate were significantly associated with psychomotor speed, whereas IMT was associated with memory test performance. HIV serostatus was not significantly associated with poorer cognitive test performance. However, among the HIV-infected individuals, the presence of detectable HIV RNA in plasma was linked to lower memory performance. CONCLUSIONS: These findings suggest that HIV infection may not be the most important predictor of cognitive performance among older gay and bisexual men in the post-highly active antiretroviral therapy era, at least among those with access to medical care and to appropriate medications. Medical factors associated with normal aging are significantly associated with performance on neuropsychological tests, and good clinical management of these factors both in HIV-infected individuals and those at risk for infection may have beneficial effects in the short term and could reduce the risk of subsequent cognitive decline.

Silencing of NADPH-dependent oxidoreductase genes (yqhD and dkgA) in furfural-resistant ethanologenic Escherichia coli.

Low concentrations of furfural are formed as a side product during the dilute acid hydrolysis of hemicellulose. Growth is inhibited by exposure to furfural but resumes after the complete reduction of furfural to the less toxic furfuryl alcohol. Growth-based selection was used to isolate a furfural-resistant mutant of ethanologenic Escherichia coli LY180, designated strain EMFR9. Based on mRNA expression levels in the parent and mutant in response to furfural challenge, genes encoding 12 oxidoreductases were found to vary by more than twofold (eight were higher in EMFR9; four were higher in the parent). All 12 genes were cloned. When expressed from plasmids, none of the eight genes in the first group increased furfural tolerance in the parent (LY180). Expression of three of the silenced genes (yqhD, dkgA, and yqfA) in EMFR9 was found to decrease furfural tolerance compared to that in the parent. Purified enzymes encoded by yqhD and dkgA were shown to have NADPH-dependent furfural reductase activity. Both exhibited low K(m) values for NADPH (8 microM and 23 microM, respectively), similar to those of biosynthetic reactions. Furfural reductase activity was not associated with yqfA. Deleting yqhD and dkgA in the parent (LY180) increased furfural tolerance, but not to the same extent observed in the mutant EMFR9. Together, these results suggest that the process of reducing furfural by using an enzyme with a low K(m) for NADPH rather than a direct inhibitory action is the primary cause for growth inhibition by low concentrations of furfural.

Genetics and visceral leishmaniasis: of mice and man.

Ninety per cent of the 500,000 annual new cases of visceral leishmaniasis (VL) occur in India/Bangladesh/Nepal, Sudan and Brazil. Importantly, 80-90% of human infections are sub-clinical or asymptomatic, usually associated with strong cell-mediated immunity. Understanding the environmental and genetic risk factors that determine why two people with the same exposure to infection differ in susceptibility could provide important leads for improved therapies. Recent research using candidate gene association analysis and genome-wide linkage studies (GWLS) in collections of families from Sudan, Brazil and India have identified a number of genes/regions related both to environmental risk factors (e.g. iron), as well as genes that determine type 1 vs. type 2 cellular immune responses. However, until now all of the allelic association studies carried out have been underpowered to find genes of small effect sizes (odds ratios or OR < 2), and GWLS using multicase pedigrees have only been powered to find single major genes, or at best oligogenic control. The accumulation of large DNA banks from India and Brazil now makes it possible to undertake genome-wide association studies (GWAS), which are ongoing as part of phase 2 of the Wellcome Trust Case Control Consortium. Data from this analysis should seed research into novel genes and mechanisms that influence susceptibility to VL.

Sucrose and overexpression of trehalose biosynthetic genes (otsBA) increase desiccation tolerance of recombinant Escherichia coli.

Survival after desiccation was highest for recombinant strains of E. coli engineered to produce ethanol (KO11 and LY163) and lactate (TG106) when sucrose was provided as the fermentable sugar. Desiccation tolerance was lower with glucose and xylose. Further improvements in desiccation tolerance with sucrose were obtained by combining this with increased expression of otsBA genes encoding trehalose biosynthesis, removal of products from metabolism by resuspending in fresh medium, and harvesting cells prior to the end of log phase. With sucrose and otsBA expression, survivals of 20%-80% were readily achieved. Fermentation tests with EM2L, a derivative of LY163 expressing ostBA, demonstrated that ethanol production from seed fermentations begun with desiccated cells is equivalent to that of an undesiccated control.

Longitudinally preserved psychomotor performance in long-term asymptomatic HIV-infected individuals.

BACKGROUND: Recent case reports have suggested that some asymptomatic HIV-infected individuals can develop CNS disturbances despite intact immunologic functioning and long-term suppression of plasma HIV concentrations to undetectable levels. This possibility has not yet been systematically studied longitudinally. METHODS: Using longitudinal data from the Multicenter AIDS Cohort Study, we investigated neuropsychological performance in long-term asymptomatic HIV-infected men who have sex with men. Performance over a 5-year period on the Symbol Digit Modalities test and the Trail Making Tests were compared in three HIV-positive asymptomatic groups [defined as 1) highly active antiretroviral therapy (HAART) treated with undetectable viral loads (n = 83), 2) AIDS-free for more than 15 years without HAART (n = 29), and 3) absence of clinical AIDS or CD4(+) lymphocyte count below 200 cells/muL at the beginning and end of the study period (n = 233)] and in HIV-negative controls (n = 237). Data were analyzed using linear mixed models and proportional odds logistic regression modeling with generalized estimating equations. RESULTS: There was no evidence of performance differences or performance declines over the 5-year period of study in any of the three long-term asymptomatic groups as compared with the HIV-negative group in the Symbol Digit Modalities test or the Trail Making Tests. Performance decrements were, however, observed with increasing age in each of the tests administered, demonstrating that performance declines could be detected by these methods. CONCLUSIONS: Regardless of how long-term asymptomatic status was defined immunologically or virologically, neuropsychological test performances remained stable. These findings suggest that psychomotor speed is preserved over many years in HIV-infected individuals with controlled HIV viremia.

Genes at human chromosome 5q31.1 regulate delayed-type hypersensitivity responses associated with Leishmania chagasi infection.

Visceral leishmaniasis (VL) caused by Leishmania chagasi is endemic to northeast Brazil. A positive delayed-type hypersensitivity skin test response (DTH+) is a marker for acquired resistance to disease, clusters in families and may be genetically controlled. Twenty-three single nucleotide polymorphisms (SNPs) were genotyped in the cytokine 5q23.3-q31.1 region IRF1-IL5-IL13-IL4-IL9-LECT2-TGFBI in 102 families (323 DTH+; 190 DTH-; 123 VL individuals) from a VL endemic region in northeast Brazil. Data from 20 SNPs were analyzed for association with DTH+/- status and VL using family-based, stepwise conditional logistic regression analysis. Independent associations were observed between the DTH+ phenotype and markers in separate linkage disequilibrium blocks in LECT2 (OR 2.25; P=0.005; 95% CI=1.28-3.97) and TGFBI (OR 1.94; P=0.003; 95% CI=1.24-3.03). VL child/parent trios gave no evidence of association, but the DTH- phenotype was associated with SNP rs2070874 at IL4 (OR 3.14; P=0.006; 95% CI=1.38-7.14), and SNP rs30740 between LECT2 and TGFBI (OR 3.00; P=0.042; 95% CI=1.04-8.65). These results indicate several genes in the immune response gene cluster at 5q23.3-q31.1 influence outcomes of L. chagasi infection in this region of Brazil.

A multicenter trial of selegiline transdermal system for HIV-associated cognitive impairment.

BACKGROUND: Cognitive impairment continues to be a significant neurologic complication of HIV infection and has been associated with oxidative stress-induced neuronal injury. Selegiline is an MAO-B inhibitor with antioxidant and neurotrophic properties. This rationale has led to the design and implementation of this Selegiline Transdermal System (STS) study with the primary aims of assessing safety and tolerability of STS as well as improvement in cognitive performance. METHODS: HIV-1 infected individuals with impaired cognitive functioning were enrolled in this placebo-controlled, three-arm study of STS across 17 sites. Cognitive impairment was determined using a standard battery of neuropsychological tests. Subjects were randomized to receive STS 3 mg/24 hours, STS 6 mg/24 hours, or matching placebo patches daily. The primary efficacy endpoint was defined as the change in neuropsychological composite Z-score (NPZ-6) from baseline to week 24. Measures of safety included frequencies of adverse experiences and abnormal results on laboratory tests. RESULTS: A total of 128 subjects (88% men, 51% white) were enrolled, median age 45 years. Most subjects (62%) had mild to moderate AIDS dementia complex. The 24-week NPZ-6 median (interquartile range) changes were 0.22 (-0.28, 0.55) for the selegiline 3 mg/24 hours arm, 0.21 (-0.18, 0.62) for the selegiline 6 mg/24 hours arm, and 0.28 (-0.16, 0.64) for the placebo arm (a positive score indicates improvement from baseline) (p = 0.914). Severe laboratory abnormalities were few and occurred in similar proportion among the three treatment arms. CONCLUSION: Selegiline was safe and well tolerated by HIV-infected individuals with cognitive impairment and mild to moderate immune suppression; however, no cognitive or functional improvement was observed in this phase II study.

Combined effect of betaine and trehalose on osmotic tolerance of Escherichia coli in mineral salts medium.

In mineral salts medium, supplementing with betaine in combination with increased production of endogenous osmoprotectant from a second copy of the trehalose biosynthetic genes (otsBA) improved growth of E. coli and increased the MIC for xylose, glucose, sodium lactate and NaCl. With these compounds, this combination was more effective than either betaine or trehalose alone. With succinate, this combination was no more effective than betaine alone. Neither approach improved tolerance to ethanol. A combination of betaine and increased trehalose may improve strain productivity for many bioproducts by promoting growth in the presence of high sugar concentrations.

IFNG and IFNGR1 gene polymorphisms and susceptibility to post-kala-azar dermal leishmaniasis in Sudan.

Post-kala-azar dermal leishmanaisis (PKDL) in Sudan is associated with elevated interferon-gamma (IFN-gamma). To study interferon-gamma pathways in PKDL, we genotyped 80 trios from the Masalit ethnic group for polymorphisms at -470 ins/delTT, -270T/C, -56T/C and +95T/C in IFNGR1 and at -179G/A and +874T/A in IFNG. No associations occurred at IFNG. Global association with haplotypes comprising all four markers at IFNGR1 (chi(2)(10df)=21.97, P=0.015) was observed, associated with a significant (chi(2)(1df)=4.54, P=0.033) bias in transmission of the haplotype insTT T T T and less (chi(2)(1df)=5.59, P=0.018) than expected transmission of insTT C C C. When compared with data on malaria associations from Gambia, the results suggest a complex pattern of haplotypic variation at the IFNGR1 promoter locus associated with different infectious disease in African populations that reflect the complex roles of IFN-gamma in parasite killing versus inflammation and pathogenesis.

Genome-wide scan for visceral leishmaniasis susceptibility genes in Brazil.

A genome-wide scan was conducted for visceral leishmaniasis (VL) in Brazil. Initially, 405 markers were typed in 22 multicase pedigrees (28 nuclear families; 174 individuals; 66 affected). Non-parametric multipoint analysis detected nine chromosomal regions with provisional evidence (logarithm of the odds (LOD) scores 0.95-1.66; 0.003

Genome-wide scan for loci influencing quantitative immune response traits in the Belém family study: comparison of methods and summary of results.

Here we report the results from a genome-wide linkage scan to identify genes and chromosomal regions that influence quantitative immune response traits, using multi-case leprosy and tuberculosis families from north-eastern Brazil. Total plasma IgE, antigen-specific IgG to Mycobacterium leprae soluble antigen (MLSA), M. tuberculosis soluble antigen (MTSA) and M. tuberculosis purified protein derivative (PPD), and antigen-specific lymphocyte proliferation (stimulation index or SI) and interferon-gamma (IFN-gamma) release to MLSA and PPD, were measured in 16 tuberculosis (184 individuals) and 21 leprosy (177 individuals) families. The individuals were genotyped at 382 autosomal microsatellite markers across the genome. The adjusted immune-response phenotypes were analysed using a variety of variance components and regression-based methods. These analyses highlighted a number of practical issues and problems with regard to implementation of the methods and, interestingly, differences were observed between several standard statistical and genetic analysis packages used. From this we determined that, for this set of traits in these pedigrees, significant p values for linkage using variance components analysis, supported by significance using the Visscher-Hopper modification of the Haseman-Elston method, provided the most compelling evidence for linkage. Using these criteria, linkage (5.8 x 10(-5) < p < 0.008) was seen for: total plasma IgE on chromosome 2; IgG to MLSA on chromosomes 8, 17 and 21; IgG to PPD on chromosome 12; SI to PPD on chromosome 1; IFN-gamma to MLSA on chromosomes 6, 7, 10, 12 and 14; and IFN-gamma to PPD on chromosomes 1, 16 and 19.