No abstract

Leishmaniases are tropical zoonotic diseases, caused by kinetoplastid parasites from the genus Leishmania. New World (NW) species are related to sylvatic cycles although urbanization processes have been reported in some South American Countries such as Colombia. Currently, few studies show the relative distribution of Leishmania species related to cutaneous Leishmaniasis (CL) in South America due to the lack of accurate surveillance and public health systems. Herein, we conducted a systematic estimation of the Leishmania species causing CL in Colombia from 1980 to 2001 via molecular typing and isoenzymes. A total of 327 Leishmania isolates from humans, sandflies and reservoirs were typed as L. panamensis 61.3% (201), L. braziliensis 27.1% (88), L. lainsoni 0.6% (2), L. guyanensis 0.9% (3), L. infantum chagasi 4% (12), L. equatoriensis 0.6% (2), L. mexicana 2.1% (8), L. amazonensis 2.8% (9) and L. colombiensis 0.6% (2). This is the first report of two new Leishmania species circulating in Colombia and suggests the need to convince the Colombian government about the need to deploy and standardize tools for the species identification to provide adequate management to individuals suffering this pathology.

Background Trypanosoma cruzi is the causative agent of Chagas disease. Due to its genetic diversity has been classified into six Discrete Typing Units (DTUs) in association with transmission cycles. In Colombia, natural T. cruzi infection has been detected in 15 triatomine species. There is scarce information regarding the infection rates, DTUs and feeding preferences of secondary vectors. Therefore, the aim of this study was to determine T. cruzi infection rates, parasite DTU, ecotopes, insect stages, geographical location and bug feeding preferences across six different triatomine species.MethodsA total of 245 insects were collected in seven departments of Colombia. We conducted molecular detection and genotyping of T. cruzi with subsequent identification of food sources. The frequency of infection, DTUs, TcI genotypes and feeding sources were plotted across the six species studied. A logistic regression model risk was estimated with insects positive for T. cruzi according to demographic and eco-epidemiological characteristics.ResultsWe collected 85 specimens of Panstrongylus geniculatus, 77 Rhodnius prolixus, 37 R. pallescens, 34 Triatoma maculata, 8 R. pictipes and 4 T. dimidiata. The overall T. cruzi infection rate was 61.2% and presented statistical associations with the departments Meta (OR: 2.65; 95% CI: 1.69–4.17) and Guajira (OR: 2.13; 95% CI: 1.16–3.94); peridomestic ecotope (OR: 2.52: 95% CI: 1.62–3.93); the vector species P. geniculatus (OR: 2.40; 95% CI: 1.51–3.82) and T. maculata (OR: 2.09; 95% CI: 1.02–4.29); females (OR: 2.05; 95% CI: 1.39–3.04) and feeding on opossum (OR: 3.15; 95% CI: 1.85–11.69) and human blood (OR: 1.55; 95% CI: 1.07–2.24). Regarding the DTUs, we observed TcI (67.3%), TcII (6.7%), TcIII (8.7%), TcIV (4.0%) and TcV (6.0%). Across the samples typed as TcI, we detected TcIDom (19%) and sylvatic TcI (75%). The frequencies of feeding sources were 59.4% (human blood); 11.2% (hen); 9.6% (bat); 5.6% (opossum); 5.1% (mouse); 4.1% (dog); 3.0% (rodent); 1.0% (armadillo); and 1.0% (cow).ConclusionsNew scenarios of T. cruzi transmission caused by secondary and sylvatic vectors are considered. The findings of sylvatic DTUs from bugs collected in domestic and peridomestic ecotopes confirms the emerging transmission scenarios in Colombia.Electronic supplementary materialThe online version of this article (doi:10.1186/s13071-016-1907-5) contains supplementary material, which is available to authorized users.

BackgroundThe diagnosis of Chagas disease is complex due to the dynamics of parasitemia in the clinical phases of the disease. The molecular tests have been considered promissory because they detect the parasite in all clinical phases. Trypanosoma cruzi presents significant genetic variability and is classified into six Discrete Typing Units TcI-TcVI (DTUs) with the emergence of foreseen genotypes within TcI as TcIDom and TcI Sylvatic. The objective of this study was to determine the operating characteristics of molecular tests (conventional and Real Time PCR) for the detection of T. cruzi DNA, parasitic loads and DTUs in a large cohort of Colombian patients from acute and chronic phases.Methodology/Principal FindingsSamples were obtained from 708 patients in all clinical phases. Standard diagnosis (direct and serological tests) and molecular tests (conventional PCR and quantitative PCR) targeting the nuclear satellite DNA region. The genotyping was performed by PCR using the intergenic region of the mini-exon gene, the 24Sa, 18S and A10 regions. The operating capabilities showed that performance of qPCR was higher compared to cPCR. Likewise, the performance of qPCR was significantly higher in acute phase compared with chronic phase. The median parasitic loads detected were 4.69 and 1.33 parasite equivalents/mL for acute and chronic phases. The main DTU identified was TcI (74.2%). TcIDom genotype was significantly more frequent in chronic phase compared to acute phase (82.1% vs 16.6%). The median parasitic load for TcIDom was significantly higher compared with TcI Sylvatic in chronic phase (2.58 vs.0.75 parasite equivalents/ml).Conclusions/SignificanceThe molecular tests are a precise tool to complement the standard diagnosis of Chagas disease, specifically in acute phase showing high discriminative power. However, it is necessary to improve the sensitivity of molecular tests in chronic phase. The frequency and parasitemia of TcIDom genotype in chronic patients highlight its possible relationship to the chronicity of the disease.

Backtracking, the reverse motion of the transcriptase enzyme on the nucleic acid template, is a universal regulatory feature of transcription in cellular organisms but its role in viruses is not established. Here we present evidence that backtracking extends into the viral realm, where backtracking by the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) RNA-dependent RNA polymerase (RdRp) may aid viral transcription and replication. Structures of SARS-CoV-2 RdRp bound to the essential nsp13 helicase and RNA suggested the helicase facilitates backtracking. We use cryo-electron microscopy, RNA–protein cross-linking, and unbiased molecular dynamics simulations to characterize SARS-CoV-2 RdRp backtracking. The results establish that the single-stranded 3′ segment of the product RNA generated by backtracking extrudes through the RdRp nucleoside triphosphate (NTP) entry tunnel, that a mismatched nucleotide at the product RNA 3′ end frays and enters the NTP entry tunnel to initiate backtracking, and that nsp13 stimulates RdRp backtracking. Backtracking may aid proofreading, a crucial process for SARS-CoV-2 resistance against antivirals.