Effect of Demographics and Time to Sample Processing on the qPCR Detection of Pathogenic Leptospira spp. from Human Samples in the National Reference Laboratory for Leptospirosis, Brazil

Abstract

Leptospirosis diagnosis by MAT requires antibody levels that are typically present only after the first week of symptoms, many days after infection. To improve testing capacity and to develop a fast and reliable solution for the diagnosis of this disease in the first few days after clinical manifestations, the National Reference Laboratory for Leptospirosis/WHO Collaborating Center in Brazil implemented a duplex molecular method by qPCR for human samples for the detection of the gene lipL32, conserved in pathogenic Leptospira spp. In this paper, we describe the overall performance of this protocol in the first 3 months as a standard routine. Detection of pathogenic Leptospira spp. DNA was similar between blood, plasma, and tissue samples, with a limit of detection as low as one cell per sample, and among 391 samples from suspected cases, 174 (44.6%) were positive. The average RNASEP1 control gene detection cycle thresholds (Ct) were 28.4 and 29.8 for positive and negative samples, respectively. The median sample collection interval from the beginning of symptoms was 3 days for positive and 4 days for negative samples, respectively. Neither age, sex, nor the time intervals between sample collection and DNA extraction significantly influenced the results. Surprisingly, positivity was related to the time between DNA extraction and the qPCR reaction. These data support the use of this routine as a diagnostic approach to strengthen the molecular detection of leptospirosis and to develop new strategies.

Structural barriers in the prevention and control of leptospirosis in a municipality in southern Brazil: a qualitative study

“…What is perceived in the country is that this scenario of low visibility is also reinforced by the decrease in media coverage related to leptospirosis, as well as scientific studies that essentially prioritize quantitative themes, such as experimental and epidemiological studies of the disease 11,12 . However, studies with a qualitative approach are still incipient, especially those that incorporate the voices of healthcare workers in Brazil, particularly in complex health issues such as leptospirosis control in high-risk areas 13 .…”