Detection of Microbial Translocation in HIV and SIV Infection Using the Limulus Amebocyte Lysate Assay is Masked by Serum and Plasma

Objective: Microbial translocation (MT) is thought to be a major contributor to the pathogenesis of HIV-related immune activation, and circulating lipopolysaccharide (LPS) from Gram-negative bacteria is the principle measurement of this process. However, related research has been impeded by inconsistent LPS test results. Methods: Specimens were obtained from HIV-infected adults enrolled in the PEARLS study (ACTG A5175) and HIV-HCV co-infected participants enrolled in a study of liver disease staging using MRI elastography. Pig-tailed macaque specimens were obtained from SIV-infected and –uninfected animals. Samples were tested for LPS using the LAL assay with diazo-coupling modifications to improve sensitive detection. Results: When exogenous LPS was added to macaque plasma,.25 % inhibition of LPS detection was found in 10/10 (100%) samples at 20 % plasma concentration compared to control; in contrast 5/10 (50%) samples at 2 % plasma concentration (p = 0.07) and 0/10 (0%) at 0.1 % plasma concentration (p = 0.004) showed.25 % inhibition of LPS detection. Similarly, when LPS was added to human serum,.25 % inhibition of LPS detection was found in 5/12 (42%) of samples at 2 % serum concentration compared to control, while 0/12 (0%) of samples in 0.1 % serum showed.25 % inhibition of LPS detection (p = 0.07). Likewise, LPS detection in human sera without exogenous LPS was improved by dilution: LPS was detected in 2/ 12 (17%) human samples in 2 % serum, ranging from 3,436–4,736 pg/mL, compared to 9/12 (75%) samples in 0.1 % serum