Lynx pardinus is one of the world’s most endangered felines inhabiting the Iberian Peninsula. The present study was performed to identify the presence of microsporidia due to the mortality increase in lynxes. Samples of urine (n = 124), feces (n = 52), and tissues [spleen (n = 13), brain (n = 9), liver (n = 11), and kidney (n = 10)] from 140 lynxes were studied. The determination of microsporidia was evaluated using Weber’s chromotrope stain and Real Time-PCR. Of the lynxes analyzed, stains showed 10.48% and 50% positivity in urine and feces samples, respectively. PCR confirmed that 7.69% and 65.38% belonged to microsporidia species. The imprints of the tissues showed positive results in the spleen (38.46%), brain (22.22%), and liver (27.27%), but negative results in the kidneys. PCR confirmed positive microsporidia results in 61.53%, 55.55%, 45.45%, and 50%, respectively. Seroprevalence against Encephalitozoon cuniculi was also studied in 138 serum samples with a positivity of 55.8%. For the first time, the results presented different species of microsporidia in the urine, feces, and tissue samples of Lynx pardinus. The high titers of anti-E. cuniculi antibodies in lynx sera confirmed the presence of microsporidia in the lynx environment. New studies are needed to establish the impact of microsporidia infection on the survival of the Iberian lynx.

Rodents are recognized as reservoirs of a wide range of pathogens, including microsporidia. The presence of microsporidia in the environment of mainland Spain and its islands has become increasingly known, as the number of studies has multiplied over time. The present study was conducted to determine the occurrence and diversity of microsporidia in three rodent species (Mus musculus, Rattus norvegicus, and Rattus rattus) in the Canary Islands, Spain. Ninety-three fecal samples were obtained from wild rodents on La Gomera and Gran Canaria Islands. Each sample was tested using Weber’s modified trichrome staining and immunofluorescence antibody tests (IFATs) against the Encephalitozoon genus and Enterocytozoon bieneusi. The microscopy-positive samples were subsequently analyzed using a nested polymerase chain reaction (PCR) followed by Sanger sequencing. The staining technique showed 38.7% (36/93) positivity, whereas the IFATs for Encephalitozoon spp. and Ent. bieneusi revealed 3.2% (3/93) and 6.5% (6/93) positivity, respectively. Finally, the nested PCR and nucleotide sequence analysis confirmed a 9.7% (9/93) occurrence of Ent. bieneusi and 17.2% occurrence (16/93) of different undetermined microsporidia species, whereas no Encephalitozoon spp. were detected. Seven different Ent. bieneusi genotypes were detected as follows: three known (AAE1, D, and SBM1) and four novel (GRE1, GRE2, LGE1, and LGE2), all of which belonged to Group 1. The results demonstrate, for the first time, that microsporidia are present in the rodent populations of the Canary Islands. Further studies are needed to determine the impact of the presence of microsporidia in rodents on the zoonotic transmission of these parasites.

The aim of this paper is to study the presence and temporal variation/distribution of microsporidian species (Enterocytozoon bieneusi and Encephalitozoon spp.) in chickens from different farms in Bombali district (Sierra Leone). Fresh faecal samples were collected from twenty chickens in April 2019, while twelve were collected in Summer 2022. Six faecal samples gave positive results microscopically (spores of Encephalitozoon spp. were observed in five samples, and E. bieneusi in one). Five samples from 2019 were found to be positive by molecular methods: two E. intestinalis and E. hellem, and one E. bieneusi; meanwhile, ten samples from 2022 were positive for spores of the genus Encephalitozoon, specifically from eight chicken.

Aptamer development has traditionally relied on iterative, labor-intensive experimental approaches. Recent advances in bioinformatics and computational power now enable the rational in silico design of high-affinity nucleic acid ligands, drastically reducing time and cost. Here, we present an integrated computational pipeline for designing ssDNA aptamers targeting the receptor-binding domain (RBD) of the SARS-CoV-2 Spike (S) protein. A library of 4000 randomly generated 20-nucleotide sequences with defined base ratios (C:G:A:T = 3:3:2:2) was screened based on secondary structure and free energy. The 100 most stable sequences (ΔG ≤ −4.1 kcal/mol) were modeled in three dimensions, yielding 1000 conformations. The 100 structures with the lowest potential energies (≤−78,051.7 kcal/mol) were selected for molecular docking with the RBD in the closed, open, and transitional states. Two aptamers—model-2-seq_2714 (seq_2714) and model-5-seq_1397 (seq_1397)—were selected for experimental assessment based on their favorable predicted docking scores against the RBD. Western blot and dot blot assays showed detectable recognition of the recombinant RBD by both aptamers under the experimental conditions tested. Exploratory testing in a limited number of clinical and environmental samples further showed that detectable signals could be obtained in selected complex matrices. These findings provide an initial proof-of-principle assessment of a computational-first strategy for prioritizing candidate aptamers for subsequent experimental testing.