Angélica Hollunder Klippel

Leishmania presents a complex life cycle that involves both invertebrate and vertebrate hosts. By regulating gene expression, protein synthesis, and metabolism, the parasite can adapt to various environmental conditions. This regulation occurs mainly at the post‐transcriptional level and may involve epitranscriptomic modifications of RNAs. Recent studies have shown that mRNAs in humans undergo a modification known as N4‐acetylcytidine (ac4C) catalyzed by the enzyme N‐acetyltransferase (NAT10), impacting mRNAs stability and translation. Here, we characterized the NAT10 homologue of L. mexicana, finding that the enzyme exhibits all the conserved acetyltransferase domains although failed to functionally complement the Kre33 mutant in Saccharomyces cerevisiae. We also discovered that LmexNAT10 is nuclear, and seems essential, as evidenced by unsuccessful attempts to obtain null mutant parasites. Phenotypic characterization of single‐knockout parasites revealed that LmexNAT10 affects the multiplication of procyclic forms and the promastigote‐amastigote differentiation. Additionally, in vivo infection studies using the invertebrate vector Lutzomyia longipalpis showed a delay in the parasite differentiation into metacyclics. Finally, we observed changes in the cell cycle progression and protein synthesis in the mutant parasites. Together, these results suggest that LmexNAT10 might be important for parasite differentiation, potentially by regulating ac4C levels.

Leishmania presents a complex life cycle that involves both invertebrate and vertebrate hosts. By regulating gene expression, protein synthesis, and metabolism, the parasite can adapt to various environmental conditions. This regulation occurs mainly at the post-transcriptional level and may involve epitranscriptomic modifications of RNAs. Recent studies have shown that mRNAs in humans undergo a modification known as N4-acetylcytidine (ac4C) catalyzed by the enzyme N-acetyltransferase (NAT10), impacting mRNAs stability and translation. Here, we characterized the NAT10 homologue of L. mexicana , finding that the enzyme exhibits all the conserved acetyltransferase domains and although failed to functionally complement the Kre33 mutant in Saccharomyces cerevisia e, has in vitro acetyltransferase activity. We also discovered that LmexNAT10 is nuclear, and seems essential, as evidenced by unsuccessful attempts to obtain null mutant parasites. Phenotypic characterization of single-knockout parasites revealed that LmexNAT10 affects the multiplication of procyclic forms and the promastigote-amastigote differentiation. Additionally, in vivo infection studies using the invertebrate vector Lutzomyia longipalpis showed a delay in the parasite differentiation into metacyclics. Finally, we observed changes in the cell cycle progression and protein synthesis in the mutant parasites. Together, these results suggest that LmexNAT10 might be important for parasite differentiation, potentially by regulating ac4C levels.

Accurate insect specimen identification is usually a crucial first step in a forensic entomological analysis. It is traditionally done by morphological classification using identification keys. However, due to sensibility limitations in the identification of animal species based only on their morphology, new methods have been developed, including species identification by DNA barcodes. The objective of this study was to identify forensically important species of Diptera in Espirito Santo state using DNA barcodes. For this, adult flies were collected in Espirito Santo, Southeast Region of Brazil. After DNA extraction, COI gene was amplified and sequenced. All sequences were matched to BOLD platform and alternatively to GenBank MegaBLAST. As result, 281 adult flies were collected and identified morphologically. From these, 36% of samples were classified as Calliphoridae, 34% of Muscidae and 30% of Sarcophagidae. Approximately 10% of all collected samples were analyzes by DNA. It was possible to identify only 35.7% of tested samples, probably due to lack of samples deposited in databases. Therefore, more efforts should be made to deposit a greater variety of dipterous in databases to allow the use of this technique in forensic routine, especially in BOLD

Road mortality is the leading source of biodiversity loss in the world, especially due to fragmentation of natural habitats and loss of wildlife. The survey of the main species victims of roadkill is of fundamental importance for the better understanding of the problem, being necessary, for this, the correct species identification. The aim of this study was to verify if DNA barcodes can be applied to identify road-killed samples that often cannot be determined morphologically. For this purpose, 222 vertebrate samples were collected in a stretch of the BR-101 highway that crosses two Discovery Coast Atlantic Forest Natural Reserves, the Sooretama Biological Reserve and the Vale Natural Reserve, in Espírito Santo, Brazil. The mitochondrial COI gene was amplified, sequenced and confronted with the BOLD database. It was possible to identify 62.16% of samples, totaling 62 different species, including Pyrrhura cruentata, Chaetomys subspinosus, Puma yagouaroundi and Leopardus wiedii considered Vulnerable in the National Official List of Species of Endangered Wildlife. The most commonly identified animals were a bat (Molossus molossus), an opossum (Didelphis aurita) and a frog (Trachycephalus mesophaeus) species. Only one reptile was identified using the technique, probably due to lack of reference sequences in BOLD. These data may contribute to a better understanding of the impact of roads on species biodiversity loss and to introduce the DNA barcode technique to road ecology scenarios.