or adtcosta@ ibmp.org.br.Although molecular diagnostics is well established in clinical laboratories, its full potential has not been extended to field settings. Typically, diagnostic real-time quantitative PCR (qPCR) reagents require temperature-controlled transportation and storage. Furthermore, thermocyclers are bulky and fragile, requiring good infrastructure for optimal operation. These major hurdles strongly limit use of molecularbased tests in low-resource scenarios. Herein, Trypanosoma cruzi or Plasmodium spp. DNA were detected with qPCR using commercial equipment (ABI7500 instrument) and a prototype platform comprising a portable device and a silicon chip, named Q3-Plus. In addition, a ready-to-use reaction format, where all qPCR reagents are stored on plate or on chip, was compared with the traditional freezer-stored format. No significant differences were observed in detecting T. cruzi or Plasmodium spp. DNA between thermocyclers, as well as between reagents' formats, for storage periods of up to 28 days (at 2 C to 8 C or 21 C to 23 C, respectively). When challenged with patients' samples, the Q3-Plus system performed as efficiently as the standard equipment for Plasmodium spp. DNA detection, showing it to be a valuable solution to malaria point-of-care diagnostics. Detection of T. cruzi DNA in chronic patients' samples using the Q3-Plus system yielded approximately 50% efficiency relative to the ABI7500. These results are essential to support future endeavors to bring molecular diagnostics to the point of care, where most needed.

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Trachoma, caused by the bacteria Chlamydia trachomatis, is the world leading infectious cause of blindness. The disease is associated with poor living conditions, especially in developing countries. In these countries, diagnosis is usually based on clinical evaluation, although laboratory confirmation is necessary. Serological-based tests for trachoma laboratory confirmation are cheaper than molecular-based tests, but the later are more sensitive and specific. Among the available molecular tests, qPCR has the best cost-benefit. With this in mind, the present study developed a new duplex qPCR reaction, which concomitantly detects C. trachomatis cryptic plasmid and the human 18S rRNA gene, as an internal reaction control. The new qPCR was validated using 50 previously qPCR-characterized samples for trachoma infection, and showed 95% specificity and 100% sensitivity, with an estimated LOD95 of 600 ag/µL. Next, 50 samples from an endemic area (Marajó, Pará) and 12 from a non-endemic area (Curitiba, Paraná) were investigated using direct immunofluorescence assay (DFA) or the new duplex qPCR. Among the 50 endemic samples, three were positive by clinical evaluation (6%), 18 by DFA (36%) and 48 by qPCR (96%). All samples positive by the clinic evaluation were also positive by qPCR. From the 18 DFA-positives, qPCR identified 16 as positives as well. On the other hand, 32 samples that were DFA-negative due to the low number of elementary bodies (<5 per slide) were positive by qPCR. The results show that the new duplex qPCR has sensitivity and specificity in similar levels to commercial qPCR tests available, and that qPCR indeed is more sensitive than clinical evaluation or DFA, thus allowing earlier treatment start. The ubiquitous presence of C. trachomatis DNA in samples from the endemic region confirms that constant monitoring is necessary. Additionally, effective measures for the elimination of trachoma and the detection of bacterial DNA in the active infection are of fundamental importance, and this newly developed duplex qPCR is an important tool towards this goal.