Effects of different ratios of omega-6:omega-3 fatty acids in the diet of sows on the proteome of milk-derived extracellular vesicles.

Milk is a nutrient-rich biofluid that contains several biocomponents with distinctive functions, including extracellular vesicles (EV). Milk EV have been associated with the regulation of the newborn's immune system and to influence essential cellular development. The EV proteome comprises the protein constituents and cargo; changes in these compartments could impact their role mediating communication. The ratio of dietary ω-6 to ω-3 polyunsaturated fatty acids (PUFA) is known to affect health and inflammation, and to induce changes in milk fatty acid composition, but no reports have included the milk EV fraction so far. We isolated EV from milk samples obtained on days 0, 7, and 14 after parturition from sows receiving either a standard diet or a test diet enriched in ω-3 (ω6:ω3 = 4:1). Small milk-derived EV were isolated using ultracentrifugation coupled with size exclusion chromatography, and characterized by nanoparticle tracking analysis, transmission electron microscopy, and Western blotting. Using a TMT-based high-resolution quantitative approach, the proteomics analysis revealed variations in the milk EV proteome within the diet groups with differences in the abundance of spondin-2 and 78 kDa glucose-regulated protein. Future studies are encouraged to explore further dietary effects on milk EV composition and their relation to the offspring's development. SIGNIFICANCE: Milk EV are known as key players mediating the regulation of the infant's immune system and growth. The EV proteome comprises the protein constituents and protein cargo, and any changes in this system could impact their role in intercellular communication. This study aimed at evaluating how different ω-6:ω-3 ratios in the maternal diet could translate to the milk EV proteome. This is relevant for basic research, but also has applied aspects in animal nutrition and health and may provide new perspectives for feeding additives.

Differential acute-phase protein responses in dogs seropositive or seronegative for Neospora caninum.

Neospora caninum is one of the most prevalent Apicomplexa parasites that causes abortion in cattle, as it infects dogs as its definitive host, causing subclinical disease or active neosporosis, marked by meningoencephalitis, and myopathies with muscle and neuromuscular signs of disease. This study aimed to evaluate the acute phase protein response in dogs seropositive and seronegative for N. caninum. Serum samples of 72 dogs were tested by an immunofluorescence antibody test using N. caninum NC-1 strain, and the study population was divided into four groups: symptomatic - muscular and/or neuromuscular signs - and seropositive (n = 16); symptomatic and seronegative (n = 9); asymptomatic and seropositive (n = 34); and asymptomatic and seronegative (n = 13). C-reactive protein (CRP) was measured via immunoturbidimetric assay and serum haptoglobin (Hp) via hemoglobin-binding capacity assay. In the symptomatic groups, seropositive dogs had higher levels of Hp, but not CRP, while seronegative dogs had higher CRP levels. There was no difference in CRP concentration in asymptomatic dogs. Dogs with neuromuscular signs had higher concentrations for Hp in the group seropositive. Hp concentration did not differ between dogs seropositive and seronegative dogs for each group. Serum Hp and CRP could not sufficiently alone flag subclinical infections. Measurement of CRP and Hp concentrations could be clinically valuable to the diagnosis of neurological diseases, and their relative change may indicate the stage of the infection, although their sole use is not able to support the diagnosis of canine neosporosis. Further studies are encouraged to evaluate the specific dynamics of acute phase proteins in canine neosporosis. Serum samples of 72 dogs were tested by an immunofluorescence antibody test using N. caninum NC-1 strain, and the study population was divided into four groups: (1) dogs with muscular and/or neuromuscular signs and seropositive for N. caninum; (2) dogs with muscular and/or neuromuscular signs and seronegative for N. caninum; (3) dogs seropositive for N. caninum with no neuromuscular signs; and (4) healthy dogs and seronegative for N. caninum. The study evaluated if N. caninum infection could have pathophysiological changes activating the acute phase response and an increase in the concentration of acute phase proteins in serum.

Comparative proteome profiling in exosomes derived from porcine colostrum versus mature milk reveals distinct functional proteomes.

Exosomes are membranous vesicles of endocytic origin, recently been considered as major players in cell-cell communication. Milk is highly complex, and diverse biocomponents provide adequate nutrition, transfer immunity, and promote adequate neonate development. Milk exosomes are suggested to have a key role in these processes, yet to be further explored, and the alteration of the exosomes' cargo in different stages of lactation stages is important for understanding the factors relevant in nursing and also for improving milk replacer products both for humans and animals. We isolated exosomes from porcine milk in different lactation stages and analyzed their content using a TMT-based high-resolution quantitative proteomic approach. Exosomes were isolated using ultracentrifugation coupled with size exclusion chromatography to enrich milk-derived exosomes in samples obtained at day 0, 7, and 14 after parturition, and characterized by nanoparticle tracking analysis, transmission electron microscopy, and Western blotting. Quantitative proteomics analysis revealed different proteome profiles for colostrum exosomes and milk exosomes. The functional analysis highlighted pathways related to the regulation of homeostasis to be upregulated in colostrum exosomes, and pathways such as endothelial cell development and lipid metabolism to be upregulated in mature milk exosomes. This study endorses the importance of exosomes as active biocomponents of milk and provides knowledge for future studies exploring their role in the regulation of immunity and growth of the newborn. SIGNIFICANCE: The identified functional proteome and protein-protein interaction networks identified in our study help to elucidate the role of milk exosomes in different lactation periods. The results generated herein are of relevance for the basic understanding of their impact on the infant's development but also for bringing forward the manufacturing of milk replacers.

Survey of Toxoplasma gondii, Neospora caninum and Sarcocystis neurona antibodies in wild red-tailed Amazon parrots (Amazona brasiliensis).

Toxoplasma gondii, Neospora caninum and Sarcocystis neurona are obligate intracellular parasites within the phylum Apicomplexa. The red-tailed Amazon parrot (Amazona brasiliensis) is a near-threatened species of psittacine that is endemic to the Atlantic Forest of Brazil and has been designated as a bioindicator because of its sensitivity to environmental qualitative status and changes. The aim of this study was to evaluate the presence of antibodies against T. gondii, N. caninum and S. neurona in wild red-tailed Amazon parrot nestlings on Rasa Island, Brazil. Blood samples were collected from 51 parrots and plasma samples were stored at - 20 °C until immunofluorescence antibody tests (IFAT) were performed. Antigen slides were prepared using tachyzoites of T. gondii (RH strain) and, N. caninum (NC-1 strain) and using merozoites of S. neurona (SNR37 strain). Plasma samples were tested at initial dilutions of 1:16 for T. gondii, 1:50 for N. caninum and 1:5 for S. neurona. An anti-chicken antibody conjugated with FITC was used as a secondary antibody at 1:50 dilution. No antibodies for any of these three protozoa were found, thus suggesting that these wild red-tailed Amazon parrot nestlings had not been exposed to these parasites.

Survey of Toxoplasma gondii, Neospora caninum and Sarcocystis neurona antibodies in wild red-tailed Amazon parrots (Amazona brasiliensis) / Pesquisa de anticorpos anti-Toxoplasma gondii, anti-Neospora caninum e anti-Sarcocystis neurona em papagaios-de-cara-roxa (Amazona brasiliensis) de vida livre

Abstract Toxoplasma gondii, Neospora caninum and Sarcocystis neurona are obligate intracellular parasites within the phylum Apicomplexa. The red-tailed Amazon parrot (Amazona brasiliensis) is a near-threatened species of psittacine that is endemic to the Atlantic Forest of Brazil and has been designated as a bioindicator because of its sensitivity to environmental qualitative status and changes. The aim of this study was to evaluate the presence of antibodies against T. gondii, N. caninum and S. neurona in wild red-tailed Amazon parrot nestlings on Rasa Island, Brazil. Blood samples were collected from 51 parrots and plasma samples were stored at - 20 °C until immunofluorescence antibody tests (IFAT) were performed. Antigen slides were prepared using tachyzoites of T. gondii (RH strain) and, N. caninum (NC-1 strain) and using merozoites of S. neurona (SNR37 strain). Plasma samples were tested at initial dilutions of 1:16 for T. gondii, 1:50 for N. caninum and 1:5 for S. neurona. An anti-chicken antibody conjugated with FITC was used as a secondary antibody at 1:50 dilution. No antibodies for any of these three protozoa were found, thus suggesting that these wild red-tailed Amazon parrot nestlings had not been exposed to these parasites.

Resumo Toxoplasma gondii, Neospora caninum e Sarcocystis neurona são protozoários intracelulares do filo Apicomplexa. O papagaio-de-cara-roxa (Amazona brasiliensis) é um psitacídeo endêmico da floresta atlântica, considerado uma espécie quase ameaçada de extinção e bioindicadora por sua sensibilidade às mudanças no ambiente. O objetivo do presente estudo foi detectar a presença de anticorpos contra T. gondii, N. caninum e S. neurona em filhotes de papagaios-de-cara-roxa (Amazona brasiliensis) de vida livre na Ilha Rasa, Brasil. Amostras de sangue foram coletadas de 51 papagaios e armazenadas a - 20ºC até a realização da Reação de Imunofluorescência Indireta (RIFI). As lâminas de RIFI com os antígenos, foram preparadas com taquizoítos de T. gondii (cepa RH) e N. caninum (cepa NC-1) e com merozoítos de S. neurona (cepa SNR37). Os plasmas foram diluídos em PBS (Ph 7,2) nas diluições 1:16 para T. gondii, 1:50 para N. caninum e 1:5 para S. neurona. O conjugado anti-IgG de galinhas marcado com fluoresceína (FITC) foi utilizado na diluição de 1:50. Não foram detectados anticorpos para os três protozoários nas amostras sugerindo que os filhotes de papagaios-de-cara-roxa não foram expostos aos protozoários.

Serosurvey of Toxoplasma gondii, Sarcocystis sp. and Neospora caninum in geese (Anser sp.) from urban parks and captivity.

Geese, ducks, mallards, and swans are birds of the order Anseriformes, which are found in the wild, in zoos and parks, and raised for meat consumption. Toxoplasma gondii, Sarcocystis sp., and Neospora caninum are protozoans of several species of animals. Wild and domestic birds can serve as intermediate hosts, disseminators and potential sources of infection of these protozoa to humans through contaminated meat. The aims of this study were: (i) to perform a serological survey of T. gondii, Sarcocystis sp. and N. caninum in geese (Anser sp.) from public parks and from captivity and (ii) to compare seroprevalence between these two locations. Antibodies were detected by Immunofluorescence antibody test using the serum of 149 geese. Antibodies to Sarcocystis sp., T. gondii, and N. caninum were detected in 28.18%, 18% and 0.67% of geese, respectively; 57% of geese from urban parks and 26.53% of geese from captivity were seropositive for at least one protozoa. The results indicate environmental contamination, particularly for the occurrence of antibodies against T. gondii - a zoonosis that causes toxoplasmosis and is transmitted through oocyte ingestion. This is the first serological survey of T. gondii, Sarcocystis sp. and N. caninum in geese from urban parks in Curitiba, Brazil.

Serosurvey of Toxoplasma gondii, Sarcocystis sp. and Neospora caninum in geese (Anser sp.) from urban parks and captivity / Soroprevalência de Toxoplasma gondii, Sarcocystis sp. e Neospora caninum em gansos (Anser sp.) de parques urbanos e cativeiro

Abstract Geese, ducks, mallards, and swans are birds of the order Anseriformes, which are found in the wild, in zoos and parks, and raised for meat consumption. Toxoplasma gondii, Sarcocystis sp., and Neospora caninum are protozoans of several species of animals. Wild and domestic birds can serve as intermediate hosts, disseminators and potential sources of infection of these protozoa to humans through contaminated meat. The aims of this study were: (i) to perform a serological survey of T. gondii, Sarcocystis sp. and N. caninum in geese (Anser sp.) from public parks and from captivity and (ii) to compare seroprevalence between these two locations. Antibodies were detected by Immunofluorescence antibody test using the serum of 149 geese. Antibodies to Sarcocystis sp., T. gondii, and N. caninum were detected in 28.18%, 18% and 0.67% of geese, respectively; 57% of geese from urban parks and 26.53% of geese from captivity were seropositive for at least one protozoa. The results indicate environmental contamination, particularly for the occurrence of antibodies against T. gondii - a zoonosis that causes toxoplasmosis and is transmitted through oocyte ingestion. This is the first serological survey of T. gondii, Sarcocystis sp. and N. caninum in geese from urban parks in Curitiba, Brazil.

Resumo Gansos, patos, marrecos e cisnes são aves da ordem Anseriformes, encontrados em vida livre, zoológicos, parques e criados para consumo da carne. Toxoplasma gondii, Sarcocystis sp. e Neospora caninum são protozoários capazes de infectar diversas espécies animais. Aves domésticas e silvestres podem ser hospedeiras intermediárias e servir como disseminadoras e potenciais fontes de infecção para seres humanos por meio da carne. O objetivo do estudo foi 1) realizar a soroprevalência de T. gondii, Sarcocystis sp. e N. caninum em gansos (Anser sp.) provenientes de parques públicos e de um cativeiro e 2) comparar a soroprevalência entre os locais. Foi realizada sorologia de 149 Anser sp. pelo método da reação de imunofluorescência indireta. Anticorpos para Sarcocystis sp., T. gondii e N. caninum foram encontrados em 28,18%, 18%, e 0,67% dos animais, respectivamente; 57% dos gansos dos parques públicos e 26,53% dos animais cativos foram soropositivos para algum dos protozoários. A ocorrência de anticorpos para tais protozoários indica contaminação ambiental, ressaltando a alta prevalência de anticorpos para T. gondii, zoonose transmitida por ingestão dos oocistos. Sugere-se investigação da água e medidas ambientais para reduzir a contaminação dos animais e do ambiente. Este é o primeiro trabalho que avaliou sorologicamente gansos provenientes de parques urbanos de Curitiba, Paraná para T. gondii, Sarcocystis sp. e N. caninum.