An investigation of fingerstick blood collection for point-of-care HIV-1 viral load monitoring in South Africa

Background. Viral load (VL) quantification is an important tool in determining newly developed drug resistance or problems with adherence to antiretroviral therapy (ART) in HIV-positive patients. VL monitoring is becoming the standard of care in many resource-limited settings. Testing in resource-limited settings may require sampling by fingerstick because of general shortages of skilled phlebotomists and the expense of venepuncture supplies and problems with their distribution.Objective. To assess the feasibility and ease of collecting 150 µL capillary blood needed for the use of a novel collection device following a classic fingerstick puncture.Methods. Patients were recruited by the study nurse upon arrival for routine ART monitoring at the Themba Lethu Clinic in Johannesburg, South Africa. Each step of the fingerstick and blood collection protocol was observed, and their completion or omission was recorded.Results. One hundred and three patients consented to the study, of whom three were excluded owing to the presence of callouses. From a total of 100 patients who consented and were enrolled, 98% of collection attempts were successful and 86% of participants required only one fingerstick to successfully collect 150 µL capillary blood. Study nurse adherence to the fingerstick protocol revealed omissions in several steps that may lower the success rate of capillary blood collection and reduce the performance of a subsequent VL assay.Conclusion. The findings of this study support the feasibility of collecting 150 µL of capillary blood via fingerstick for point-of-care HIV-1 VL testing in a resource-limited setting

Rapid, Point-of-Care Extraction of Human Immunodeficiency Virus Type 1 Proviral DNA from Whole Blood for Detection by Real-Time PCR ▿

PCR detection of human immunodeficiency virus type 1 (HIV-1) proviral DNA is the method recommended for use for the diagnosis of HIV-1 infection in infants in limited-resource settings. Currently, testing must be performed in central laboratories, which are usually located some distance from health care facilities. While the collection and transportation of samples, such as dried blood spots, has improved test accessibility, the results are often not returned for several weeks. To enable PCR to be performed at the point of care while the mothers wait, we have developed a vertical filtration method that uses a separation membrane and an absorbent pad to extract cellular DNA from whole blood in less than 2 min. Cells are trapped in the separation membrane as the specimen is collected, and then a lysis buffer is added. The membrane retains the DNA, while the buffer washes away PCR inhibitors, which get wicked into the absorbent blotter pad. The membrane containing the entrapped DNA is then added to the PCR mixture without further purification. The method demonstrates a high degree of reproducibility and analytical sensitivity and allows the quantification of as few as 20 copies of HIV-1 proviral DNA from 100 μl of blood. In a blinded study with 182 longitudinal samples from infants (ages, 0 to 72 weeks) obtained from the Women and Infants Transmission Study, our assay demonstrated a sensitivity of 99% and a specificity of 100%

The nematode Caenorhabditis elegans displays a chemotaxis behavior to tuberculosis-specific odorants

A simple, affordable diagnostic test for pulmonary tuberculosis (TB) is urgently needed to improve detection of active Mycobacterium tuberculosis. Recently, it has been suggested that animal behavior can be used as a biosensor to signal the presence of human disease. For example, the giant African pouched rats can detect tuberculosis by sniffing sputum specimens while trained honeybees respond to three of the volatile organic compounds (VOCs) detected in the breath of TB positive patients by proboscis extension. However, both rats and honeybees require animal housing facilities and professional trainers, which are outside the scope of most disease testing facilities. Here, we report that the innate olfactory behavioral response of the roundworm nematode Caenorhabditis elegans can be used to detect the TB-specific VOCs methyl p-anisate, methyl nicotinate, methyl phenylacetate and o-phenylanisole, in chemotaxis assays. Dauer larvae, a long-lived stress resistant alternative development state of C. elegans in which the animals can survive for extended periods of time in dry conditions with no food, were also demonstrated to detect the VOCs. We propose that exposing naive dauer larvae to TB-related VOCs and recording their response in this behavioral assay could lead to the development of a new method for TB diagnostics using breath as the sample type. Keywords: Tuberculosis, Caenorhabditis elegans, Chemotaxis, Volatile organic compounds, Diagnostics, Odorant

Immiscible Phase Nucleic Acid Purification Eliminates PCR Inhibitors with a Single Pass of Paramagnetic Particles through a Hydrophobic Liquid

Extraction and purification of nucleic acids from complex biological samples for PCR are critical steps because inhibitors must be removed that can affect reaction efficiency and the accuracy of results. This preanalytical processing generally involves capturing nucleic acids on microparticles that are then washed with a series of buffers to desorb and dilute out interfering substances. We have developed a novel purification method that replaces multiple wash steps with a single pass of paramagnetic particles (PMPs) though an immiscible hydrophobic liquid. Only two aqueous solutions are required: a lysis buffer, in which nucleic acids are captured on PMPs, and an elution buffer, in which they are released for amplification. The PMPs containing the nucleic acids are magnetically transported through a channel containing liquid wax that connects the lysis chamber to the elution chamber in a specially designed cartridge. Transporting PMPs through the immiscible phase yielded DNA and RNA as pure as that obtained after extensive wash steps required by comparable purification methods. Our immiscible-phase process has been applied to targets in whole blood, plasma, and urine and will enable the development of faster and simpler purification systems

Latino Men and Familial Risk Communication About Prostate Cancer

PURPOSE: This study investigated how familial communication about prostate cancer risk and screening affects sons of men with prostate cancer. It is important to engage Latino families shared decision making toward early detection because first degree relatives of men with PCa are at heightened risk and Latino men are diagnosed at more advanced stages of disease than Non-Hispanic White men. METHODS: The team conducted semi-structured interviews with seventeen sons of PCa survivors. Eight participants completed a follow up interview an average of seven months later. Therefore, our sample includes twenty-five transcripts. The sons are Latinos living in Southern California. Data were analyzed with a mix of a priori topical codes and grounded theory techniques. RESULTS: Sons were under informed about both familial risk and screening options. They became sensitized to PCa, desired information, and held protective intentions. Hopeful intentions came up against cultural taboos around sex, reproductive health, and intimacy that limited discussions between fathers and sons. Fathers were a valued source of information, but play various roles, which affect sons’ screening intentions. Open communication between father and son promoted awareness of screening and familial risk. DISCUSSION: Uncertainty about familial risk and screening options, especially early detection strategies, was exacerbated by cultural taboos around PCa. Fathers could have been primary and credible advocates for shared decision making, but sons found it difficult to learn from their fathers’ experience. NURSING IMPLICATIONS: Findings from our study can inform community based interventions with Latino families, help to culturally tailor health messaging, and sensitize clinicians to a group which needs concerted counseling about PCa risk and screening