Publications

Polymicrobial bronchopulmonary infections in cystic fibrosis (CF) cause progressive lung damage and death. Although the arrival of Pseudomonas aeruginosa often heralds a more rapid rate of pulmonary decline, there is significant inter-individual variation in the rate of decline, the causes of which remain poorly understood. By coupling culture-independent methods with ecological analyses, we discovered correlations between bacterial community profiles and clinical disease markers in respiratory tracts of 45 children with CF. Bacterial community complexity was inversely correlated with patient age, presence of P. aeruginosa and antibiotic exposure, and was related to CF genotype. Strikingly, bacterial communities lacking P. aeruginosa were much more similar to each other than were those containing P. aeruginosa, regardless of antibiotic exposure. This suggests that community composition might be a better predictor of disease progression than the presence of P. aeruginosa alone and deserves further study.

Staphylococcus saprophyticus, an obligate human pathogen, is the most common Gram-positive causative agent of urinary tract infection (UTI) in young, healthy women. Despite the clinical importance of S. saprophyticus, little is known about how it causes disease in the urinary tract or how the host responds to the infection. Here we established an in vivo model to study both host and bacterial factors contributing to S. saprophyticus UTI. Using this model, we show that S. saprophyticus preferentially infects C3H/HeN murine kidneys instead of the bladder, a trait observed for multiple clinical isolates. Bacterial persistence in the kidneys was observed in C3H/HeN mice but not in C57BL/6 mice, indicating that host factors strongly contribute to the ability of S. saprophyticus to cause UTI. Using C3H/HeN mice as a model, histologic and immunofluorescence analyses of infected tissues revealed that S. saprophyticus induced epithelial cell shedding in the bladder and an inflammatory response characterized by macrophage and neutrophil infiltration in the bladder and kidneys. The inflammatory response correlated with increased production of proinflammatory cytokines and chemokines in both the bladder and the kidneys. Finally, we observed that the putative S. saprophyticus virulence factors Ssp and SdrI were important for persistence, but not for initial colonization, in the murine urinary tract. Thus, we characterized both host and bacterial factors involved in progression of S. saprophyticus UTI, and we describe a useful model system for studying factors involved in the pathogenesis of this Gram-positive uropathogen.

BACKGROUND

Anaemia worsens as kidney function declines. Both conditions are associated with increased mortality. Serum cystatin C is purportedly a more sensitive marker of kidney disease and a better predictor of mortality than serum creatinine. However, studies suggest that extrarenal factors also influence cystatin C levels.

METHODS

We determined whether estimates of glomerular filtration rate [estimated glomerular filtration rate (eGFR)] based on serum cystatin C alone or in combination with serum creatinine were superior to those based on serum creatinine in recognizing impaired kidney function in the setting of anaemia in a sub-sample of the Third National Health and Nutrition Examination Survey of the USA consisting of 6734 participants, 20 years or older.

RESULTS

The prevalence of moderate to severe kidney disease (eGFR 15-59 mL/min/1.73 m(2)) among anaemic persons was 15-16% when based on serum creatinine alone (eGFR(SCR)) or combined with cystatin C (eGFR(SCR) (+) (CYSC)); this estimate increased to nearly 25% when kidney function was estimated by cystatin C (eGFR(CYSC)). The adjusted odds ratios of kidney disease in anaemic versus non-anaemic persons were slightly higher with eGFR(CYSC) than eGFR(SCR) and eGFR(SCR) (+) (CYSC) in younger adults [odds ratio (OR) = 5.22, 95% confidence interval (CI): 2.23, 12.17], women (OR = 5.34, 95% CI: 2.36, 12.06) and those with elevated C-reactive protein (CRP) (OR = 7.36, 95% CI: 1.98-27.36).

CONCLUSIONS

Impaired kidney function was common in individuals with anaemia. Among anaemic individuals, the prevalence estimate for kidney disease was notably higher when kidney function was estimated by cystatin C alone compared with the estimations by serum creatinine alone or in combination with serum cystatin C. eGFR(CYSC) may be particularly helpful in identifying kidney disease in the setting of anaemia among younger persons, women and those with elevated CRP. Regardless of which renal biomarker is used, our study suggests that an evaluation for underlying kidney disease should be considered in the standard workup of anaemia.

Multistate modeling methods are well-suited for analysis of some chronic diseases that move through distinct stages. The memoryless or Markov assumptions typically made, however, may be suspect for some diseases, such as hepatitis C, where there is interest in whether prognosis depends on history. This paper describes methods for multistate modeling where transition risk can depend on any property of past progression history, including time spent in the current stage and the time taken to reach the current stage. Analysis of 901 measurements of fibrosis in 401 patients following liver transplantation found decreasing risk of progression as time in the current stage increased, even when controlled for several fixed covariates. Longer time to reach the current stage did not appear associated with lower progression risk. Analysis of simulation scenarios based on the transplant study showed that greater misclassification of fibrosis produced more technical difficulties in fitting the models and poorer estimation of covariate effects than did less misclassification or error-free fibrosis measurement. The higher risk of progression when less time has been spent in the current stage could be due to varying disease activity over time, with recent progression indicating an "active" period and consequent higher risk of further progression.

BACKGROUND

The role of host genetics in the development of subclinical atherosclerosis in the context of HIV-infected persons who are being treated with highly active antiretroviral therapy (HAART) is not well understood.

METHODS

The present genome-wide association study (GWAS) is based on 177 HIV-positive Caucasian males receiving HAART who participated in the Fat Redistribution and Metabolic Change in HIV Infection (FRAM) Study. Common and internal carotid intima-media thicknesses (cIMT) measured by B-mode ultrasound were used as a subclinical measure of atherosclerosis. Single nucleotide polymorphisms (SNPs) were assayed using the Illumina HumanCNV370-quad beadchip. Copy Number Variants (CNV) were inferred using a hidden Markov Model (PennCNV). Regression analyses were used to assess the association of common and internal cIMT with individual SNPs and CNVs, adjusting for age, duration of antiretroviral treatment, and principal components to account for potential population stratification.

RESULTS

Two SNPs in tight linkage disequilibrium, rs2229116 (a missense, nonsynonymous polymorphism (IIe to Val)) and rs7177922, located in the ryanodine receptor (RYR3) gene on chromosome 15 were significantly associated with common cIMT (P-value < 1.61 x 10). The RYR gene family has been known to play a role in the etiology of cardiovascular disease and has been shown to be regulated by HIV TAT protein.

CONCLUSION

These results suggest that in the context of HIV infection and HAART, a functional SNP in a biologically plausible candidate gene, RYR3, is associated with increased common carotid IMT, which is a surrogate for atherosclerosis.

Chronic immune activation and inflammation (e.g., as manifest by production of type I interferons) are major determinants of disease progression in primate lentivirus infections. To investigate the impact of such activation on intrathymic T-cell production, we studied infection of the human thymus implants of SCID-hu Thy/Liv mice with X4 and R5 HIV. X4 HIV was observed to infect CD3(-)CD4(+)CD8(-)CXCR4(+)CCR5(-) intrathymic T-cell progenitors (ITTP) and to abrogate thymopoiesis. R5 HIV, by contrast, first established a nonpathogenic infection of thymic macrophages and then, after many weeks, began to replicate in ITTP. We demonstrate here that the tropism of R5 HIV is expanded and pathogenicity enhanced by upregulation of CCR5 on these key T-cell progenitors. Such CCR5 induction was mediated by interferon-alpha (IFN-alpha) in both thymic organ cultures and in SCID-hu mice, and antibody neutralization of IFN-alpha in R5 HIV-infected SCID-hu mice inhibited both CCR5 upregulation and infection of the T-cell progenitors. These observations suggest a mechanism by which IFN-alpha production may paradoxically expand the tropism of R5 HIV and, in so doing, accelerate disease progression.

Panton-Valentine leukocidin (PVL) is a cytolytic toxin associated with severe community-associated methicillin-resistant Staphylococcus aureus (CA-MRSA) infections. However, the relative contribution of PVL to host cell lysis during CA-MRSA infection remains unknown. Here we investigated the relative contribution of PVL to human polymorphonuclear leukocyte (PMN) plasma membrane permeability and lysis in vitro by using culture supernatants from wild-type and isogenic lukS/F-PV negative (Deltapvl) USA300 and USA400 strains. Using S. aureus culture conditions that favor selective high production of PVL (CCY medium), there was on average more PMN plasma membrane permeability and cell lysis caused by supernatants derived from wild-type strains compared with those from Deltapvl strains. Unexpectedly, plasma membrane permeability did not necessarily correlate with ultimate cell lysis. Moreover, the level of pore formation caused by culture supernatants varied dramatically (e.g., range was 0.32-99.09% for wild-type USA300 supernatants at 30 min) and was not attributable to differences in PMN susceptibility to PVL among human blood donors. We conclude that PMN pore formation assays utilizing S. aureus culture supernatants have limited ability to estimate the relative contribution of PVL to pathogenesis (or cytolysis in vitro or in vivo), especially when assayed using culture media that promote selective high production of PVL.