Protocol for Nearly Full-Length Sequencing of HIV-1 RNA from Plasma

Nearly full-length genome sequencing of HIV-1 using peripheral blood mononuclear cells (PBMC) DNA as a template for PCR is now a relatively routine laboratory procedure. However, this has not been the case when using virion RNA as the template and this has made full genome analysis of circulating viruses difficult. Therefore, a well-developed procedure for sequencing of full-length HIV-1 RNA directly from plasma was needed. Plasma from U.S. donors representing a range of viral loads (VL) was used to develop the assay. RNA was extracted from plasma and reverse-transcribed. Two or three overlapping regions were PCR amplified to cover the entire viral genome and sequenced for verification. The success of the procedure was sensitive to VL but was routinely successful for VL greater than 105 and the rate declined in proportion to the VL. While the two-amplicon strategy had an advantage of increasing the possibility of amplifying a single species of HIV-1, the three-amplicon strategy was more successful in amplifying samples with low viral loads. This protocol provides a useful tool for molecular analysis to understand the HIV epidemic and pathogenesis, as well as diagnosis, therapy and future vaccine strategies

Use of renal allografts from donors positive for hepatitis B core antibody confers minimal risk for subsequent development of clinical hepatitis B virus disease

Background. The risk associated with transplantation of renal allografts from hepatitis B virus core antibody-positive (HBcAb(+)), hepatitis B virus surface antigen-negative (HBsAg(-)) donors is not well defined. Methods. Over 4 years, we performed 45 kidney transplants from IgG HBcAb(+), IgM HBcAb(-), HBsAg(-) donors into recipients with a history of prior hepatitis B virus (HBV) infection or reported vaccination. We examined HBV-related outcomes in these 45 patients, in comparison with 45 recipients of allografts from HBcAb(-) donors (matched for transplant type, date, and pretransplant HBV antibodies). We sought evidence for HBV transmission by testing posttransplant sera for the presence of HBcAb, hepatitis B virus surface antibody, and HBsAg. Additionally, we analyzed alanine aminotransferase profiles and allograft survival rates for all patients. Results. No patient receiving an allograft from an HBcAb(+) donor developed clinical HBV infection. No patient receiving an allograft from an HBcAb(+) donor had HBsAg detected through retrospective testing of stored sera or through prospective routine clinical evaluation and care. However, among the HBcAb(+) kidney recipients, 27% developed new HBcAb and/or hepatitis B virus surface antibody after transplant; in contrast, only 4% of control patients developed new antibody responses (relative risk=4.94; confidence interval 1.07-22.83). Among the recipients of HBcAb(+) organs, 18% developed elevated transaminases after transplant, in comparison with 36% of the controls. No association was found between \u27seroconverter\u27 status and elevated alanine aminotransferase profiles in either group. Conclusions. Transplantation of renal allografts from HBcAb(+), HBsAg(-) donors was not associated with clinically detectable HBV disease or antigenemia. However, recipients had a significantly increased risk of HBV seroconversion, consistent with exposure to HBV antigen. These results suggest that HBcAb(+) kidneys can be safely used if transplanted into appropriate recipients, but highlight the need for effective HBV vaccination and vaccine-response monitoring in potential recipients

Toxoplasma gondii Immunoglobulin G Antibodies and Nonfatal Suicidal Self-Directed Violence

Objective: The primary aim was to relate Toxoplasma gondii seropositivity and serointensity to scores on the self-rated Suicide Assessment Scale (SUAS-S). Another aim was to reevaluate the previously reported positive association between T gondii serointensity and a history of nonfatal suicidal self-directed violence. Method: This cross-sectional, observational study compared T gondii serointensity and seropositivity in plasma from 54 adult suicide attempters (inpatients at Lund University Hospital, Lund, Sweden) and 30 adult control subjects (randomly selected from the municipal population register in Lund, Sweden) recruited between 2006 and 2010.The potential of patients and controls for self-directed violence was evaluated with the SUAS-S. Psychiatric diagnoses were made according to DSM-IV criteria. Plasma samples were tested for immunoglobulin G antibodies to T gondii, cytomegalovirus, and herpes simplex virus type 1. Data were analyzed using multivariable logistic regression to investigate the association between T gondii serointensity or seropositivity and a history of nonfatal suicidal self-directed violence; multivariable linear regression was used to explore the relationship between T gondii serointensify or seropositivity and the SUAS-S. Both regression models included sex, age, and body mass index as covariates. Results: Seropositivity of T gondii (adjusted odds ratio [OR]=7.12; 95% CI, 1.66-30.6; P=.008) and serointensity of T gondii (adjusted OR=2.01; 95% CI, 1.09-3.71; P=.03) were positively associated with a history of nonfatal suicidal self-directed violence. Seropositivity of T gondii was associated with higher SUAS-S scores, a relationship significant for the whole sample (P=.026), but not for suicide attempters only. No significant associations with other pathogens were identified. Conclusions: These results are consistent with previous reports on the association between T gondii infection and nonfatal suicidal self-directed violence. Confirming these results in future large longitudinal studies and including suicide as an outcome may lead to novel individualized approaches in suicide prevention. J Clin Psychiatry 2012;73 (8):1069-1076 (c) Copyright 2012 Physicians Postgraduate Press, Inc