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1.  Inflammatory Responses in Blood Samples of Human Immunodeficiency Virus-Infected Patients with Pulmonary Infections 
We analyzed the characteristics of the inflammatory response occurring in blood during pulmonary infections in human immunodeficiency virus (HIV)-infected patients. A prospective study of consecutive hospital admissions of HIV-infected patients with new-onset radiologic pulmonary infiltrates was carried out in a tertiary university hospital from April 1998 to May 2001. Plasma cyclic AMP receptor protein (CRP), interleukin 1β (IL-1β), IL-6, IL-8, IL-10, and tumor necrosis factor alpha (TNF-α) levels were determined at the time of admission and 4, 5, and 6 days later. Patients were included in a protocol addressed to study etiology and outcome of disease. A total of 249 episodes of infection were included, with the main diagnoses being bacterial pneumonia (BP) (118 episodes), Pneumocystis carinii pneumonia (PCP) (41 episodes), and mycobacteriosis (36 episodes). For these three patient groups, at the time of admission the median CRP and cytokine levels were as follows: CRP, 10.2, 3.8 and 5 mg/dl, respectively (P = 0.0001); IL-8, 19, 3, and 2.9 pg/ml (P = 0.045); and TNF-α, 46.4, 44, and 75 pg/ml, respectively (P = 0.029). There were no significant differences in levels of IL-1β, IL-6, or IL-10 among the patient groups. A total of 23 patients died. At the time of admission, HIV-infected patients with BP had higher plasma CRP and IL-8 levels than did PCP and mycobacteriosis patients. TNF-α levels were higher in patients with mycobacteriosis. An elevated IL-8 level (>61 pg/ml) at the time of admission was an independent factor associated with higher mortality (odds ratio, 12; 95% confidence interval, 1.2 to 235.5).
doi:10.1128/CDLI.11.3.608-614.2004
PMCID: PMC404570  PMID: 15138189
2.  Single-Tube Balanced Heminested PCR for Detecting Mycobacterium tuberculosis in Smear-Negative Samples 
Journal of Clinical Microbiology  2000;38(3):1166-1169.
In order to achieve more sensitive and specific results for the rapid diagnosis of tuberculosis, we have developed a new method, named balanced heminested PCR, which avoids the inconvenience of asymmetric amplification and has the advantages of single-tube heminested PCR. This was achieved by replacing the outer primer that participates in both rounds of amplification in the standard heminested technique by another primer containing the sequence of the inner primer attached at its 5′ end. When both techniques were tested for the IS6110 target of Mycobacterium tuberculosis complex in 80 smear-negative culture-positive sputum samples and 60 control samples, the results showed 100% specificity for both techniques and sensitivities of 60 and 75% for heminested PCR and balanced heminested PCR, respectively (P = 0.02). In conclusion, the balanced heminested technique shows a higher sensitivity than that of the standard heminested, and it could be applied to any PCR by attaching the inner primer at the 5′ end of the opposite outer primer. Thus, the balanced heminested technique provides a target for the inner primer in both strands, avoiding asymmetric amplification and thereby resulting in a more efficient amplification, and, in practice, a higher sensitivity without loss of specificity and with a minimum risk of cross-contamination.
PMCID: PMC86365  PMID: 10699014

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