Specific rolling circle amplification of low-copy human polyomaviruses BKV, HPyV6, HPyV7, TSPyV, and STLPyV

Eleven new human polyomaviruses have been recently discovered, yet for most of these viruses, little is known of their biology and clinical impact. Rolling circle amplification (RCA) is an ideal method for the amplification of the circular polyomavirus genome due to its high fidelity amplification of circular DNA. In this study, a modified RCA method was developed to selectively amplify a range of polyomavirus genomes. Initial evaluation showed a multiplexed temperature-graded reaction profile gave the best yield and sensitivity in amplifying BK polyomavirus in a background of human DNA, with up to 1 × 10(8)-fold increases in viral genomes from as little as 10 genome copies per reaction. Furthermore, the method proved to be more sensitive and provided a 200-fold greater yield than that of random hexamers based standard RCA. Application of the method to other novel human polyomaviruses showed successful amplification of TSPyV, HPyV6, HPyV7, and STLPyV from low-viral load positive clinical samples, with viral genome enrichment ranging from 1 × 10(8) up to 1 × 10(10). This directed RCA method can be applied to selectively amplify other low-copy polyomaviral genomes from a background of competing non-specific DNA, and is a useful tool in further research into the rapidly expanding Polyomaviridae family.

Dark-energy constraints and correlations with systematics from CFHTLS weak lensing, SNLS supernovae Ia and WMAP5

Aims We combine measurements of weak gravitational lensing from the CFHTLS-Wide survey, supernovae Ia from CFHT SNLS and CMB anisotropies from WMAP5 to obtain joint constraints on cosmological parameters, in particular, the dark-energy equation-of-state parameter w. We assess the influence of systematics in the data on the results and look for possible correlations with cosmological parameters. Methods We implemented an MCMC algorithm to sample the parameter space of a flat CDM model with a dark-energy component of constant w. Systematics in the data are parametrised and included in the analysis. We determine the influence of photometric calibration of SNIa data on cosmological results by calculating the response of the distance modulus to photometric zero-point variations. The weak lensing data set is tested for anomalous field-to-field variations and a systematic shape measurement bias for high-redshift galaxies. Results Ignoring photometric uncertainties for SNLS biases cosmological parameters by at most 20% of the statistical errors, using supernovae alone; the parameter uncertainties are underestimated by 10%. The weak-lensing field-to-field variance between 1 deg2-MegaCam pointings is 5-15% higher than predicted from N-body simulations. We find no bias in the lensing signal at high redshift, within the framework of a simple model, and marginalising over cosmological parameters. Assuming a systematic underestimation of the lensing signal, the normalisation increases by up to 8%. Combining all three probes we obtain -0.10 < 1 + w < 0.06 at 68% confidence ( -0.18 < 1 + w < 0.12 at 95%), including systematic errors. Our results are therefore consistent with the cosmological constant . Systematics in the data increase the error bars by up to 35%; the best-fit values change by less than 0.15.

Detection of recently discovered human polyomaviruses in a longitudinal kidney transplant cohort

A large number of human polyomaviruses have been discovered in the last 7 years. However, little is known about the clinical impact on vulnerable immunosuppressed patient populations. Blood, urine, and respiratory swabs collected from a prospective, longitudinal adult kidney transplant cohort (n = 167) generally pre-operatively, at day 4, months 1, 3, and 6 posttransplant, and at BK viremic episodes within the first year were screened for 12 human polyomaviruses using real-time polymerase chain reaction. Newly discovered polyomaviruses were most commonly detected in the respiratory tract, with persistent shedding seen for up to 6 months posttransplant. Merkel cell polyomavirus was the most common detection, but was not associated with clinical symptoms or subsequent development of skin cancer or other skin abnormalities. In contrast, KI polyomavirus was associated with respiratory disease in a subset of patients. Human polyomavirus 9, Malawi polyomavirus, and human polyomavirus 12 were not detected in any patient samples.