Trichinella spiralis Infecting Wild Boars in West, Southwest, and Northwest of Romania: Evidence of an Underrated Risk.

The species of the genus Trichinella are etiological agents distributed all over the world and are able to infect mammals, birds, and reptiles. Trichinella spiralis is the species most adapted to domestic and wild pigs and is also the most important etiological agent of trichinellosis. The wild boar (Sus scrofa) is a nocturnal omnivorous mammal belonging to the Suidae family. S. scrofa has a great appetite and its diet includes a variety of small prey such as mice, rats, and other rodents, as well as carcasses of larger animals. The aim of this study was the identification and the molecular characterization of Trichinella larvae isolated from the muscle tissue of S. scrofa specimens collected in different counties of Romania. The muscle samples were examined by artificial digestion and the larvae identified at the species level by multiplex PCR. T. spiralis, a species that is able to infect a considerable number of different host species including humans, was identified. In Romania, S. scrofa is an important reservoir species for T. spiralis and plays an important role in linking the domestic and the wild cycle of Trichinella, with serious repercussions for human health.

Identification and Molecular Characterization of Giant Liver Fluke (Fascioloides magna) Infection in European Fallow Deer (Dama dama) in Romania-First Report.

Fascioloidosis is a parasitic disease of primary wild and domestic ruminants, caused by giant liver fluke, Fascioloides magna. The definitive host of the liver fluke in its area of origin (North America) is the white-tailed deer (Odocoileus virginianus). In Europe, the red deer (Cervus elaphus) and European fallow deer (Dama dama) are definitive hosts and the most sensitive hosts to F. magna infection, on which the parasite exerts serious pathogenic effects. In this study, we analyzed fecal samples and livers of 72 D. dama from 11 hunting grounds in Arad County, Romania. Of the 72 fecal samples and livers from D. dama, trematodes of the genus Fascioloides were identified in four (5.56%). Sequencing revealed that the trematodes identified in the samples were similar to the sequence of F. magna (GenBank no. EF534992.1, DQ683545.1, KU232369.1). The sequence obtained from the molecular analysis has been deposited in GenBank® under accession number OQ689976.1. This study describes the first report of giant liver fluke (F. magna) infection in D. dama in Romania.

First Molecular Data of Gongylonema pulchrum (Rhabditida: Gongylonematidae) in European Fallow Deer Dama dama from Romania.

Due to its adaptive versatility to numerous types of habitats, extremely diverse both in terms of composition and specificity, developed in various areas of the Western Plains of Romania, the European fallow deer (Dama dama) is a species with high ecological plasticity. In this area, the D. dama interacts with other species of wild fauna but also with numerous domestic animals, an important aspect in terms of the sanitary-veterinary status of animal populations, as well as the existence of a potential risk of infection with various species of parasites that can cause the D. dama specimens to obtain certain diseases and even zoonoses. A total of 133 esophagi from D. dama have been examined for helminths. Of the 133 esophagus samples from D. dama, nematodes of the genus Gongylonema were identified in 25 (18.80%). Sequencing revealed that the nematode identified in the samples was 99% similar to the sequence of Gongylonema pulchrum (GenBank no. LC026018.1, LC388754.1, AB646061). The present research is the first report of the nematode G. pulchrum from D. dama in Romania.

First Identification of Trichinella pseudospiralis in a Golden Jackal (Canis aureus) in Romania.

Trichinella spp. are etiological zoonotic agents that spread throughout the world and affect mammals, birds, and reptiles. Within this genus, Trichinella pseudospiralis is the only recognized non-encapsulated species known to infect mammals and birds. This species has been reported in the majority of European countries, and the real epidemiological scenario of this species remains to be defined because its detection in mammals is much lower than that of the capsulated species. The aim of this study was to examine the presence of Trichinella larvae isolated from the muscles of a jackal from the hunting fund of 36 Murfatlar, Constanta County, Romania. The muscle samples were examined by artificial digestion, and the larvae were identified at the species level by multiplex PCR. The presence of larvae belonging to T. pseudospiralis, a species more frequently reported in carnivorous birds, was observed. This study describes the first identification of T. pseudospiralis in a jackal. The results suggest that there is an urgent need to investigate which species of mammals and/or birds act as reservoirs for this zoonotic nematode in Romania.

First Identification and Molecular Characterization of Trichinella britovi (Nematoda: Trichinellidae) from the Pine Marten (Martes martes Linnaeus, 1758) in Romania.

Trichinella spp. are etiological zoonotic agents spread throughout the world and affect mammals, birds, and reptiles; they evolve via two cycles: domestic and sylvatic. Martes martes is a carnivorous nocturnal mammal from the family Mustelidae. In Romania, this host is widespread in all forests of the country. Martes martes has an extremely voracious appetite, feeding on fruit and also on a variety of small animals, including rodents such as mice and rats. The aim of this study was the identification and molecular characterization of Trichinella larvae isolated from the muscle tissue of Martes martes collected in different counties of Romania. The muscle samples were examined via artificial digestion, and the larvae were identified at the species level via multiplex PCR. The presence of larvae belonging to Trichinella britovi, a species frequently identified in wild carnivores in temperate zones, was observed. Although T. britovi has been already reported in several host species in Romania, this is the first time this species has been observed in a Martes martes specimen. This finding contributes to our knowledge about the host species involved in the maintenance of the Trichinella sylvatic cycle in Romania, and it confirms that this parasite is consistently present in the wild fauna of this country.

The First Identification of Trichinella britovi in the Raccoon Dog (Nyctereutes procyonoides) in Romania.

Trichinella spp. are nematodes distributed throughout the world that affect an impressive number of host animals (mammals, birds, and reptiles) involved in the evolution of two cycles, the domestic and the sylvatic. The raccoon dog (Nyctereutes procyonoides) is an omnivorous mammal with great ecological plasticity. The expansion of the raccoon dog in Europe is associated with the risk of the introduction and spread of different pathogens, especially zoonotic ones (Trichinella, Echinococcus). Currently, the raccoon dog's range in Romania is limited to the Danube Delta area, the Lower Danube Meadow, and the Prut Meadow. The aim of this study was to examine the presence of Trichinella larvae isolated from the muscles of raccoon dog from six hunting funds of Giurgeni, Ialomița County, Romania. The muscle samples were examined via artificial digestion, and the obtained larvae were processed via multiplex PCR. The PCR-amplified ESV and ITS1 DNA fragments were then sequenced for species confirmation. The species Trichinella britovi, which is the most common species identified in wild carnivores in temperate zones, was confirmed. Although T. britovi has been reported in several host animals in Romania, this case report confirms its presence in the raccoon dog for the first time.

First Morphological and Molecular Identification of Demodex injai in Golden Jackal (Canis aureus Linnaeus, 1758) in Romania.

Demodicosis is one of the most important external parasitic diseases found in carnivores. Three species of the Demodex mite inhabit the skin of dogs and related species, D. canis being the most prevalent. This paper describes the first case of infestation with D. injai in a golden jackal in Romania. An emaciated golden jackal female body found in Timiș County, western Romania, was examined at Parasitology Department of Faculty of Veterinary Medicine, Timișoara. The gross lesions were present on different regions of the body: feet, tail, axillary and inguinal areas, and skin folds as well, consisting of erythema, extensive severe alopecia with lichenification, seborrhea, and scaling. In order to establish diagnosis, microscopic examination of skin scrapes, trichogram (hair plucking), acetate tape test (impression), fungal culture, and PCR were performed. Both microscopic measurements and PCR analysis have confirmed the presence of D. injai.

Design of a Synthetic Long Peptide Vaccine Targeting HPV-16 and -18 Using Immunoinformatic Methods.

Human papillomavirus types 16 and 18 cause the majority of cervical cancers worldwide. Despite the availability of three prophylactic vaccines based on virus-like particles (VLP) of the major capsid protein (L1), these vaccines are unable to clear an existing infection. Such infected persons experience an increased risk of neoplastic transformation. To overcome this problem, this study proposes an alternative synthetic long peptide (SLP)-based vaccine for persons already infected, including those with precancerous lesions. This new vaccine was designed to stimulate both CD8+ and CD4+ T cells, providing a robust and long-lasting immune response. The SLP construct includes both HLA class I- and class II-restricted epitopes, identified from IEDB or predicted using NetMHCPan and NetMHCIIPan. None of the SLPs were allergenic nor toxic, based on in silico studies. Population coverage studies provided 98.18% coverage for class I epitopes and 99.81% coverage for class II peptides in the IEDB world population's allele set. Three-dimensional structure ab initio prediction using Rosetta provided good quality models, which were assessed using PROCHECK and QMEAN4. Molecular docking with toll-like receptor 2 identified potential intrinsic TLR2 agonist activity, while molecular dynamics studies of SLPs in water suggested good stability, with favorable thermodynamic properties.

Optimization and applications of echo state networks with leaky-integrator neurons.

Standard echo state networks (ESNs) are built from simple additive units with a sigmoid activation function. Here we investigate ESNs whose reservoir units are leaky integrator units. Units of this type have individual state dynamics, which can be exploited in various ways to accommodate the network to the temporal characteristics of a learning task. We present stability conditions, introduce and investigate a stochastic gradient descent method for the optimization of the global learning parameters (input and output feedback scalings, leaking rate, spectral radius) and demonstrate the usefulness of leaky-integrator ESNs for (i) learning very slow dynamic systems and replaying the learnt system at different speeds, (ii) classifying relatively slow and noisy time series (the Japanese Vowel dataset--here we obtain a zero test error rate), and (iii) recognizing strongly time-warped dynamic patterns.