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Ascaris lumbricoides and Ascaris suum are nematode parasites that infect millions of people and pigs worldwide, respectively. Reports of cross-infection and hybridization between the two species has stimulated molecular epidemiological studies of the Ascaris genus. In this study, we evaluated the dynamics of Ascaris transmission between Guarani indigenous schoolchildren, pigs, and the environment of a village in the state of Paraná, southern Brazil. Parasitological and molecular analyses of fecal samples from humans and pigs, and soil samples from the village were carried out. Eggs of Ascaris spp. were observed in 8.4% (7/83) of human samples, 44.4% (8/18) of pig samples, and 8.9% (6/68) of soil samples. PCR amplification of the ITS-1 locus of the rDNA gene in samples that were positive in the parasitological examination revealed mixed infection and/or hybrids of the two species, A. lumbricoides and A. suum, in human and swine hosts. The soil, which was contaminated by both human and swine feces, also contained eggs of the two Ascaris species and hybrids, thus constituting a source of Ascaris infection for both hosts. DNA from A. lumbricoides and A. suum, individually, was detected in samples from humans and pigs, respectively, and the soil, while DNA from hybrid and/or Ascaris spp. was detected in samples from both hosts and the soil. The results of this study confirm the necessity of a One Health approach with the correct disposal of both human and animal feces to control the spread of human and swine ascariasis.

SARS-CoV-2 is the virus responsible for a respiratory disease called COVID-19 that devastated global public health. Since 2020, there has been an intense effort by the scientific community to develop safe and effective prophylactic and therapeutic agents against this disease. In this context, peptides have emerged as an alternative for inhibiting the causative agent. However, designing peptides that bind efficiently is still an open challenge. Here, we show an algorithm for peptide engineering. Our strategy consists of starting with a peptide whose structure is similar to the interaction region of the human ACE2 protein with the SPIKE protein, which is important for SARS-COV-2 infection. Our methodology is based on a genetic algorithm performing systematic steps of random mutation, protein–peptide docking (using the PyRosetta library) and selecting the best-optimized peptides based on the contacts made at the peptide–protein interface. We performed three case studies to evaluate the tool parameters and compared our results with proposals presented in the literature. Additionally, we performed molecular dynamics (MD) simulations (three systems, 200 ns each) to probe whether our suggested peptides could interact with the spike protein. Our results suggest that our methodology could be a good strategy for designing peptides.

Background Rhodnius robustus and Rhodnius pictipes are vectors of Trypanosoma cruzi, the etiologic agent of Chagas disease (CD), that are found in the Brazilian Amazon region. Susceptibility to infection and vector competence depend on the parasite-vector relationship. Our objective was to evaluate the interaction between T. cruzi and these two triatomine vectors in pure and mixed experimental infections of T. cruzi strains from the same or different geographic regions. Methods Fifth-instar nymphs of R. robustus and R. pictipes were fed on mice infected with four T. cruzi strains, namely genotypes TcIAM, TcIMG, TcIIPR, and TcIVAM, respectively, from the Brazilian states of Amazonas, Minas Gerais and Paraná. Over a period of 120 days, excreta were examined every 20 days to assess vector competence, and intestinal contents (IC) were examined every 30 days to determine susceptibility to infection. Results The highest positive rate in the fresh examination (%+FE, 30.0%), the highest number of parasitic forms (PF, n = 1969) and the highest metacyclogenesis rate (%MC, 53.8%) in the excreta were recorded for R. robustus /TcIVAM. Examination of the IC of R. pictipes revealed a higher number of PF in infections with TcIAM (22,680 PF) and TcIIPR (19,845 PF) alone or in association (17,145 PF), as well as a %+FE of 75.0% with TcII, in comparison with the other genotypes. The highest %MC (100%) was recorded for the mixed infections of TcIAM with TcIIPR or TcIVAM in the IC of R. pictipes . Conclusions Overall, both species were found to be susceptible to the T. cruzi strains studied. Rhodnius robustus showed vector competence for genotypes TcIVAM and TcIAM+TcIVAM and R. pictipes for TcIAM+TcIVAM and TcIAM+TcIIPR; there was elimination of infective forms as early as at 20 days. Our results suggest that both the genetics of the parasite and its geographic origin influence the susceptibility to infection and vector competence, alone or in association. Graphical Abstract

Equine piroplasmosis stands out among the diseases that affect Equidae in Brazil and the world. It is caused by the protozoa Theileria equi and Babesia caballi. The objective of the present study was to carry out the molecular characterization of T. equi using equine blood samples collected in the 5 geographic regions of Brazil. Samples from all over the country were tested for the presence of T. equi by real-time PCR. The 18S rRNA sequences (∼1,600 bp) obtained from 23 samples taken from naturally infected horses were characterized by sequencing and analyzed to identify the genotypes and the possible sites of genetic variability. Thirteen different T. equi 18S rRNA sequences were identified, and 2 different genotypes were demonstrated to be in circulation in Brazil. Alignment entropy analysis demonstrated the existence of three hypervariable regions (V2, V4, and V8) within the 18S rRNA sequence of T. equi. The V2 region is located between nucleotides 63 and 75, V4 is located between nucleotides 524 and 586, and V8 is located between nucleotides 1,208 and 1,226. The hypervariable region V4 demonstrated the greatest variation within the 18S rRNA sequence of T. equi. Phylogenetic analysis based on the 18S rRNA sequences revealed the formation of 3 distinct clades (A, B, and C). The Brazilian samples belonged to 2 clades (A and C). The present study describes the characterization and heterogeneity of the circulating T. equi 18S rRNA sequences in Brazil. The results confirm that the country is an endemic area for the disease, and they indicate that at least 2 distinct T. equi genotypes are naturally infecting equines in Brazil.

Context:Central precocious puberty (CPP) results from premature activation of the hypothalamic–pituitary–gonadal axis. Few genetic causes of CPP have been identified, with the most common being mutations in the paternally expressed imprinted gene MKRN3.Objective:To identify the genetic etiology of CPP in a large multigenerational family.Design:Linkage analysis followed by whole-genome sequencing was performed in a family with five female members with nonsyndromic CPP. Detailed phenotyping was performed at the time of initial diagnosis and long-term follow-up, and circulating levels of Delta-like 1 homolog (DLK1) were measured in affected individuals. Expression of DLK1 was measured in mouse hypothalamus and in kisspeptin-secreting neuronal cell lines in vitro.Setting:Endocrine clinic of an academic medical center.Patients:Patients with familial CPP were studied.Results:A complex defect of DLK1 (∼14-kb deletion and 269-bp duplication) was identified in this family. This deletion included the 5′ untranslated region and the first exon of DLK1, including the translational start site. Only family members who inherited the defect from their father have precocious puberty, consistent with the known imprinting of DLK1. The patients did not demonstrate additional features of the imprinted disorder Temple syndrome except for increased fat mass. Serum DLK1 levels were undetectable in all affected individuals. Dlk1 was expressed in mouse hypothalamus and in kisspeptin neuron-derived cell lines.Conclusion:We identified a genomic defect in DLK1 associated with isolated familial CPP. MKRN3 and DLK1 are both paternally expressed imprinted genes. These findings suggest a role of genomic imprinting in regulating the timing of human puberty.Through a combination of linkage analysis and whole genome-sequencing, a mutation in the paternally expressed imprinted gene DLK1 in a family with central precocious puberty is identified.

The identification of loss-of-function mutations in MKRN3 in patients with central precocious puberty in association with the decrease in MKRN3 expression in the medial basal hypothalamus of mice before the initiation of reproductive maturation suggests that MKRN3 is acting as a brake on gonadotropin-releasing hormone (GnRH) secretion during childhood. In the current study, we investigated the mechanism by which MKRN3 prevents premature manifestation of the pubertal process. We showed that, as in mice, MKRN3 expression is high in the hypothalamus of rats and nonhuman primates early in life, decreases as puberty approaches, and is independent of sex steroid hormones. We demonstrated that Mkrn3 is expressed in Kiss1 neurons of the mouse hypothalamic arcuate nucleus and that MKRN3 repressed promoter activity of human KISS1 and TAC3, 2 key stimulators of GnRH secretion. We further showed that MKRN3 has ubiquitinase activity, that this activity is reduced by MKRN3 mutations affecting the RING finger domain, and that these mutations compromised the ability of MKRN3 to repress KISS1 and TAC3 promoter activity. These results indicate that MKRN3 acts to prevent puberty initiation, at least in part, by repressing KISS1 and TAC3 transcription and that this action may involve an MKRN3-directed ubiquitination-mediated mechanism.

After obtaining certification of the absence of transmission of the Trypanosoma cruzi by Triatoma infestans in 2006, other native species of protozoan vectors have been found in human dwellings within municipalities of the State of Paraná, Southern Brazil. However, the spatial distribution of T. cruzi vectors and how climatic and landscape combined variables explain the distribution are still poorly understood. The goal of this study was to predict the potential distribution of T. cruzi vectors as a proxy for Chagas disease transmission risk using Ecological Niche Models (ENMs) based on climatic and landscape variables. We hypothesize that ENM based on both climate and landscape variables are more powerful than climate-only or landscape-only models, and that this will be true independent of vector species. A total of 2,662 records of triatomines of five species were obtained by community-based entomological surveillance from 2007 to 2013. The species with the highest number of specimens was Panstrongylus megistus (73%; n = 1,943), followed by Panstrongylus geniculatus (15.4%; 411), Rhodnius neglectus (6.0%; 159), Triatoma sordida (4.5%; 119) and Rhodnius prolixus (1.1%; 30). Of the total, 71.9% were captured at the intradomicile. T. cruzi infection was observed in 19.7% of the 2,472 examined insects. ENMs were generated based on selected climate and landscape variables with 1 km2 spatial resolution. Zonal statistics were used for classifying the municipalities as to the risk of occurrence of synanthropic triatomines. The integrated analysis of the climate and landscape suitability on triatomines geographical distribution was powerful on generating good predictive models. Moreover, this showed that some municipalities in the northwest, north and northeast of the Paraná state have a higher risk of T. cruzi vector transmission. This occurs because those regions present high climatic and landscape suitability values for occurrence of their vectors. The frequent invasion of houses by infected triatomines clearly indicates a greater risk of transmission of T. cruzi to the inhabitants. More public health attention should be given in the northern areas of the State of Paraná, which presents high climate and landscape suitabilities for the disease vectors. In conclusion, our results-through spatial analysis and predictive maps-showed to be effective in identifying areas of potential distribution and, consequently, in the definition of strategic areas and actions to prevent new cases of Chagas' disease, reinforcing the need for continuous and robust surveillance in these areas.

Abstract Human puberty is a dynamic biological process determined by the increase in the pulsatile secretion of GnRH triggered by distinct factors not fully understood. Current knowledge reveals fine tuning between an increase in stimulatory factors and a decrease in inhibitory factors, where genetic and epigenetic factors have been indicated as key players in the regulation of puberty onset by distinct lines of evidence. Central precocious puberty (CPP) results from the premature reactivation of pulsatile secretion of GnRH. In the past decade, the identification of genetic causes of CPP has largely expanded, revealing hypothalamic regulatory factors of pubertal timing. Among them, 3 genes associated with CPP are linked to mechanisms involving DNA methylation, reinforcing the strong role of epigenetics underlying this disorder. Loss-of-function mutations in Makorin Ring-Finger Protein 3 (MKRN3) and Delta-Like Non-Canonical Notch Ligand 1 (DLK1), 2 autosomal maternally imprinted genes, have been described as relevant monogenic causes of CPP with the phenotype exclusively associated with paternal transmission. MKRN3 has proven to be a key component of the hypothalamic inhibitory input on GnRH neurons through different mechanisms. Additionally, rare heterozygous variants in the Methyl-CpG-Binding Protein 2 (MECP2), an X-linked gene that is a key factor of DNA methylation machinery, were identified in girls with sporadic CPP with or without neurodevelopmental disorders. In this mini-review, we focus on how the identification of genetic causes of CPP has revealed epigenetic regulators of human pubertal timing, summarizing the latest knowledge on the associations of puberty with MKRN3, DLK1, and MECP2.

Abstract Context Loss-of-function mutations of makorin RING finger protein 3 (MKRN3) are the most common monogenic cause of familial central precocious puberty (CPP). Objective To describe the clinical and hormonal features of a large cohort of patients with CPP due to MKRN3 mutations and compare the characteristics of different types of genetic defects. Methods Multiethnic cohort of 716 patients with familial or idiopathic CPP screened for MKRN3 mutations using Sanger sequencing. A group of 156 Brazilian girls with idiopathic CPP (ICPP) was used as control group. Results Seventy-one patients (45 girls and 26 boys from 36 families) had 18 different loss-of-function MKRN3 mutations. Eight mutations were classified as severe (70% of patients). Among the 71 patients, first pubertal signs occurred at 6.2 ± 1.2 years in girls and 7.1 ± 1.5 years in boys. Girls with MKRN3 mutations had a shorter delay between puberty onset and first evaluation and higher follicle-stimulating hormone levels than ICPP. Patients with severe MKRN3 mutations had a greater bone age advancement than patients with missense mutations (2.3 ± 1.6 vs 1.6 ± 1.4 years, P = .048), and had higher basal luteinizing hormone levels (2.2 ± 1.8 vs 1.1 ± 1.1 UI/L, P = .018) at the time of presentation. Computational protein modeling revealed that 60% of the missense mutations were predicted to cause protein destabilization. Conclusion Inherited premature activation of the reproductive axis caused by loss-of-function mutations of MKRN3 is clinically indistinct from ICPP. However, the type of genetic defect may affect bone age maturation and gonadotropin levels.