PCR inhibition by nucleic acid extracts is a well known yet poorly described phenomenon. Inhibition assessment generally depends on the assumption that inhibitors affect all PCR reactions to the same extent; i.e. that the reaction of interest and the control reaction are equally susceptible to inhibition. To test this assumption we performed inhibition assessment on DNA extracts from human urine samples, fresh urine and EDTA using different PCR reactions. When copurified inhibitors were assessed using two different PCR reactions one reaction appeared to be inhibited whilst the other was not. Further experiments using various concentrations of unextracted urine to inhibit six different PCR reactions revealed that susceptibility to inhibition was highly variable between reactions. Similar results were obtained using EDTA as the PCR inhibitor. We could find no obvious explanation why one reaction should be more susceptible to inhibition than another, although a possible association with amplicon GC content was noted. These findings have serious implications for all PCR-based gene expression studies, including the relatively new PCR array method, and for both qualitative and quantitative PCR-based molecular diagnostic assays, suggesting that careful consideration should be given to inhibition compatibility when conducting PCR analyses. We have demonstrated unequivocally that it is not safe to assume that different PCR reactions are equally susceptible to inhibition by substances co-purified in nucleic acid extracts.

Background. Pneumocystis jirovecii pneumonia (PCP) remains the leading cause of opportunistic infection among human immunodeficiency virus (HIV)– inf

Background: Several studies have described improved outcomes for HIV-infected patients admitted to the intensive care unit (ICU) since the introduction of highly active antiretroviral therapy (HAART). A study was undertaken to examine the outcome from the ICU for HIV-infected patients and to identify prognostic factors. Methods: A retrospective study of HIV-infected adults admitted to a university affiliated hospital ICU between January 1999 and December 2005 was performed. Information was collected on patient demographics, receipt of HAART (no patient began HAART on the ICU), reason for ICU admission and hospital course. Outcomes were survival to ICU discharge and to hospital discharge. Results: 102 patients had 113 admissions to the ICU; HIV infection was newly diagnosed in 31 patients. Survival (first episode ICU discharge and hospital discharge) was 77% and 68%, respectively, compared with 74% and 65% for general medical patients. ICU and hospital survival was 78% and 67% in those receiving HAART, and 75% and 66% in those who were not. In univariate analysis, factors associated with survival were: haemoglobin (OR = 1.25, 95% CI 1.03 to 1.51, for an increase of 1 g/dl), CD4 count (OR = 1.59, 95% CI 0.98 to 2.58, for a 10-fold increase in cells/µl), APACHE II score (OR = 0.51, 95% CI 0.29 to 0.90, for a 10 unit increase) and mechanical ventilation (OR = 0.29, 95% CI 0.10 to 0.83). Conclusions: The outcome for HIV-infected patients admitted to the ICU was good and was comparable to that in general medical patients. More than a quarter of patients had newly diagnosed HIV infection. Patients receiving HAART did not have a better outcome.

Forty-seven bronchoalveolar lavage fluid samples from 16 human immunodeficiency virus (HIV)–infected patients were used to test the latency model of

Background: Despite a decline in incidence of Pneumocystis jirovecii pneumonia (PCP), severe PCP continues to be a common cause of admission to the intensive care unit (ICU) where mortality remains high. A study was undertaken to examine the outcome from intensive care for patients with PCP and to identify prognostic factors. Methods: A retrospective cohort study was conducted of HIV infected adults admitted to a university affiliated hospital ICU between November 1990 and October 2005. Case note review collected information on demographic variables, use of prophylaxis and highly active antiretroviral therapy (HAART), and hospital course. The main outcome was 1 month mortality, either on the ICU or in hospital. Results: Fifty nine patients were admitted to the ICU on 60 occasions. Thirty four patients (57%) required mechanical ventilation. Overall mortality was 53%. No patient received HAART before or during ICU admission. Multivariate analysis showed that the factors associated with mortality were the year of diagnosis (before mid 1996 (mortality 71%) compared with later (mortality 34%; p = 0.008)), age (p = 0.016), and the need for mechanical ventilation and/or development of pneumothorax (p = 0.031). Mortality was not associated with sex, ethnicity, prior receipt of sulpha prophylaxis, haemoglobin, serum albumin, CD4 count, Pao₂, A-ao₂ gradient, co-pathology in bronchoscopic lavage fluid, medical co-morbidity, APACHE II score, or duration of mechanical ventilation. Conclusions: Observed improved outcomes from severe PCP for patients admitted to the ICU occurred in the absence of intervention with HAART and probably reflect general improvements in ICU management of respiratory failure and ARDS rather than improvements in the management of PCP.

Background: Previous health economic studies recommend either a dual screening strategy [tuberculin skin test (TST) followed by interferon-γ-release assay (IGRA)] or a single one [IGRA only] for latent tuberculosis infection (LTBI), the former largely based on claims that it is more cost-effective. We sought to examine that conclusion through the use of a model that accounts for the additional costs of adverse drug reactions and directly compares two commercially available versions of the IGRA: the Quantiferon-TB-Gold-In-Tube (QFT-GIT) and T-SPOT.TB. Methods: A LTBI screening model directed at screening contacts was used to perform a cost-effectiveness analysis, from a UK healthcare perspective, taking into account the risk of isoniazid-related hepatotoxicity and post-exposure TB (2 years post contact) using the TST, QFT-GIT and T-SPOT.TB IGRAs. Results: Examining costs alone, the TST/IGRA dual screening strategies (TST/T-SPOT.TB and TST/QFT-GIT; £162,387 and £157,048 per 1000 contacts, respectively) cost less than their single strategy counterparts (T-SPOT.TB and QFT-GIT; £203,983 and £202,921 per 1000 contacts) which have higher IGRA test costs and greater numbers of persons undergoing LTBI treatment. However, IGRA alone strategies direct healthcare interventions and costs more accurately to those that are truly infected. Subsequently, less contacts need to be treated to prevent an active case of TB (T-SPOT.TB and QFT-GIT; 61.7 and 69.7 contacts) in IGRA alone strategies. IGRA single strategies also prevent more cases of post-exposure TB. However, this greater effectiveness does not outweigh the lower incremental costs associated with the dual strategies. Consequently, when these costs are combined with effectiveness, the IGRA dual strategies are more cost-effective than their single strategy counterparts. Comparing between the IGRAs, T-SPOT.TB-based strategies (single and dual; £39,712 and £37,206 per active TB case prevented, respectively) were more cost-effective than the QFT-GIT-based strategies (single and dual; £42,051 and £37,699 per active TB case prevented, respectively). Using the TST alone was the least cost-effective (£47,840 per active TB case prevented). Cost effectiveness values were sensitive to changes in LTBI prevalence, IGRA test sensitivities/specificities and IGRA test costs. Conclusion: A dual strategy is more cost effective than a single strategy but this conclusion is sensitive to screening test assumptions and LTBI prevalence.