Antifungal activity of aminopyrrolnitrin against Trichophyton verrucosum in a guinea pig model of dermatophytosis.

BACKGROUND: Dermatophytosis is a common and major public health concern worldwide. Despite the increasing availability of antifungal drugs, relapses and untreated cases of dermatophyte infections are reported. Therefore, novel antifungal agents are required. Aminopyrrolnitrin (APRN) shows promise for dermatophytosis treatment because of its antifungal activity. OBJECTIVES: This study aimed to assess the antifungal properties of APRN against Trichophyton verrucosum (T. verrucosum), in both laboratory settings and a guinea pig model. METHODS: The minimum inhibitory concentrations (MICs) of APRN and enilconazole against T. verrucosum were determined according to the CLSI M38 method. The skins of 16 male guinea pigs were infected with 1.0 × 108 conidia of T. verrucosum and the animals were grouped into sets of four: negative control group (NC) received normal saline; positive control group (PC) received 2 µg/mL of enilconazole; and APRN4 and APRN8 received 4 and 8 µg/mL of APRN, respectively. Clinical, mycological and histological efficacies were measured after 10 days. RESULTS: The MIC90 of APRN and enilconazole against T. verrucosum was 4 and 2 µg/mL, respectively. The clinical scores of PC, APRN4, and APRN8 were significantly lower than those of NC. Clinical and mycological efficacies were higher for APRN8, APRN4 and PC. No fungi were observed in the skin tissues of APRN4 and APRN8, while fungi were observed in 50% of the PC. CONCLUSION: APRN showed antifungal activity against T. verrucosum in vitro and in vivo and is a potential candidate for the treatment of dermatophytosis.

Brachygnathia Inferior in Cloned Dogs Is Possibly Correlated with Variants of Wnt Signaling Pathway Initiators.

Abnormalities in animals cloned via somatic cell nuclear transfer (SCNT) have been reported. In this study, to produce bomb-sniffing dogs, we successfully cloned four healthy dogs through SCNT using the same donor genome from the skin of a male German shepherd old dog. Veterinary diagnosis (X-ray/3D-CT imaging) revealed that two cloned dogs showed normal phenotypes, whereas the others showed abnormal shortening of the mandible (brachygnathia inferior) at 1 month after birth, even though they were cloned under the same conditions except for the oocyte source. Therefore, we aimed to determine the genetic cause of brachygnathia inferior in these cloned dogs. To determine the genetic defects related to brachygnathia inferior, we performed karyotyping and whole-genome sequencing (WGS) for identifying small genetic alterations in the genome, such as single-nucleotide variations or frameshifts. There were no chromosomal numerical abnormalities in all cloned dogs. However, WGS analysis revealed variants of Wnt signaling pathway initiators (WNT5B, DVL2, DACT1, ARRB2, FZD 4/8) and cadherin (CDH11, CDH1like) in cloned dogs with brachygnathia inferior. In conclusion, this study proposes that brachygnathia inferior in cloned dogs may be associated with variants in initiators and/or regulators of the Wnt/cadherin signaling pathway.

Peptoniphilus faecalis sp. nov., isolated from swine faeces.

An obligately anaerobic, Gram-positive, non-motile, coccus-shaped bacterial strain designated AGMB00490T was isolated from swine faeces. 16S rRNA gene sequence-based phylogenetic analysis indicated that the isolate belongs to the genus Peptoniphilus and that the most closely related species is Peptoniphilus gorbachii WAL 10418T (=KCTC 5947T, 97.22 % 16S rRNA gene sequence similarity). Whole genome sequence analysis determined that the DNA G+C content of strain AGMB00490T was 31.2 mol% and moreover that the genome size and numbers of tRNA and rRNA genes were 2 129 517 bp, 34 and 10, respectively. Strain AGMB00490T was negative for oxidase and urease; positive for catalase, indole production, arginine arylamidase, leucine arylamidase, tyrosine arylamidase and histidine arylamidase; and weakly positive for phenylalanine arylamidase and glycine arylamidase. The major cellular fatty acids (>10 %) of the isolate were determined to be C16 : 0 and C18 : 1 ω9c. Strain AGMB00490T produced acetic acid as a major end product of metabolism. Accordingly, phylogenetic, physiologic and chemotaxonomic analyses revealed that strain AGMB00490T represents a novel species for which the name Peptoniphilus faecalis sp. nov. is proposed. The type strain is AGMB00490T (=KCTC 15944T=NBRC 114159T).

Phocaeicola faecicola sp. nov., isolated from porcine faeces.

An obligately anaerobic, non-motile, Gram-negative and rod-shaped strain (AGMB03916T) was isolated from faeces of a 2-week-old piglet raised at the National Institute of Animal Science in Wanju, Republic of Korea. Growth of strain AGMB03916T occurred at 30-45 °C (optimum, 37 °C), at pH 6-9 (optimum, pH 8) and in the presence of 0.5-1.0 % (w/v) NaCl. Based on the results of 16S rRNA gene sequence analyses, strain AGMB03916T was closely related to two validly published species of the genus Phocaeicola, Phocaeicola plebeius and Phocaeicola coprocola. The 16S rRNA gene sequence similarity of strain AGMB03916T compared to P. plebeius M12T (=KCTC 5793T) and P. coprocola M16T (=KCTC 5443T) were 96.3 and 95.0 %, respectively. The genomic DNA G+C content of strain AGMB03916T was 46.4 mol%. The average nucleotide identity values between strain AGMB03916T and the reference strains were 74.9-78.5 %. Cells were able to utilize d-glucose, lactose, sucrose, maltose, salicin, aesculin hydrolysis, cellobiose and raffinose. The major end product of metabolism was acetate. The major cellular fatty acids (>10 %) of the isolate were iso-C15 : 0, anteiso-C15 : 0, C16 : 0, C16 : 0 3-OH and summed feature 11 (iso-C17 : 0 3-OH and/or C18 : 2 DMA). On the basis of the genotypic, biochemical, chemotaxonomic, phenotypic and phylogenetic data, strain AGMB03916T represents a novel species of the genus Phocaeicola, for which the name Phocaeicola faecicola sp. nov. is proposed. The type strain is AGMB03916T (=KCTC 25014T=GDMCC 1.2574T).

Anaerosporobacter faecicola sp. nov. isolated from faeces of Korean cow.

A novel bacterial isolate designated as strain AGMB01083T was isolated from Korean cow faeces deposited in the National Institute of Animal Science (Wanju, Republic of Korea). The bacterium is obligate anaerobic, Gram-strain-positive, and motile. Cells are straight or curved rod-shaped, flagella and spores are observed. Growth occurs between 20-40 °C (temperature optimum of 35 °C), at pH 7-9 (pH optimum of 7), and in the presence of 0.5-1.0 % (w/v) NaCl. Based on the 16S rRNA gene sequence analysis, the strain belongs to the genus Anaerosporobacter and is most closely related to A. mobilis HY-37-4T (=KCTC5027T, similarity, 95.7 %). The DNA G+C content is 36.2 mol%, determined by the whole-genome sequence. The average nucleotide identity value between strain AGMB01083T and strain A. mobilis HY-37-4T is 75.5 %, below the interspecies identity threshold value. The major cellular fatty acids (>10 %) of strain AGMB01083T are C16 : 0, C16 : 0 dimethyl acetal (DMA), and C16 : 0 3-OH. Based on the phylogenetic, phenotypic, biochemical, chemotaxonomic, and genomic characterization, strain AGMB01083T is proposed to be a novel species, named Anaerosporobacter faecicola, in the genus Anaerosporobacter. The type strain is AGMB01083T (=KCTC 15857T=NBRC 114517T).

Description of Anaerostipes faecalis sp. nov., a new segmented filamentous bacterium isolated from swine faeces.

A novel, strictly anaerobic, gram-negative, segmented filamentous bacterium strain AGMB03513T, was isolated from the faeces of a 5-month-old pig. Phylogenetic analysis based on the 16S rRNA gene indicated that the isolate was a member of the family Lachnospiraceae, and the closest strain was Anaerostipes butyraticus. Strain AGMB03513T formed a lineage within the genus Anaerostipes and was closely related to A. butyraticus DSM 22094 T (= KCTC 15125 T, 95.8%), Anaerostipes hadrus DSM 3319 T (= KCTC 15606 T, 95.5%), Anaerostipes caccae DSM 14662 T (= KCTC 15019 T, 94.0%), and Anaerostipes rhamnosivorans DSM 26241 T (= KCTC 15316 T, 93.4%). Strain AGMB03513T grew at temperatures between 30 and 45 °C, within a pH range of 7.0-9.0, and in medium containing up to 1.5% NaCl. Cells were found to utilise D-glucose, D-mannitol, D-lactose, D-saccharose, D-maltose, D-xylose, L-arabinose, D-mannose, and D-sorbitol, and acetate was identified as the major end product of metabolism. The major components of the cellular fatty acids were C12:0, C16:0, and C18:0. In addition, the bacterium contained meso-diaminopimelic acid in the cell wall. According to the comparative analysis of the whole genome sequence, the DNA G + C content of strain AGMB03513 was 37.0 mol%. In addition, Average nucleotide identity (ANI), average amino acid identity (AAI), and digital DNA-DNA hybridisation (dDDH) values were obtained in comparisons of strain AGMB03513T with reference strains of species in the genus Anaerostipes. ANI values were found to be between 71.0 and 75.7%, AAI values between 66.6 and 73.2%, and dDDH values between 19.5 and 21.4%. All the data were below the threshold range for species determination. Based on phenotypic, phylogenetic, biochemical, chemotaxonomic, and genomic characteristics, we considered it reasonable to assign a novel species status to strain AGMB03513T, for which we propose the name Anaerostipes faecalis sp. nov. The type strain is AGMB03513T (= KCTC 25020 T = NBRC 114896 T).

Faecalicatena faecalis sp. nov., a moderately alkaliphilic bacterial strain isolated from swine faeces.

An obligately anaerobic, Gram-stain-positive, non-motile, non-spore-forming and rod-shaped strain AGMB00832T was isolated from swine faeces. Phylogenetic analysis based on the 16S rRNA gene, together with the housekeeping genes, gyrB and rpoD, revealed that strain AGMB00832T belonged to the genus Faecalicatena and was most closely related to Faecalicatena orotica KCTC 15331T. In biochemical analysis, strain AGMB00832T was shown to be negative for catalase, oxidase and urease. Furthermore, the isolate was positive for ß-glucosidase, ß-glucuronidase, glutamic acid decarboxylase, proline arylamidase, acid phosphatase and naphthol-AS-BI-phosphohydrolase. The major cellular fatty acids (> 10%) of the isolate were C14:0, C16:0 and C18:1ω11t DMA. Based on the whole genome sequence analysis, the DNA G + C content of strain AGMB00832T was 44.2 mol%, and the genome size and numbers of rRNA and tRNA genes were 5,175,159 bp, 11 and 53, respectively. The average nucleotide identity and digital DNA-DNA hybridization values between strain AGMB00832T and related strains were ≤ 77.4 and 22.5%, respectively. Furthermore, the genome analysis revealed the presence of genes for alkaline shock protein 23 and cation/proton antiporters, which may facilitate growth of strain AGMB00832T in alkaline culture condition. On the basis of polyphasic taxonomic approach, strain AGMB00832T represents a novel species within the genus Faecalicatena, for which the name Faecalicatena faecalis sp. nov. is proposed. The type strain is AGMB00832T (= KCTC 15946T = NBRC 114613T).

Anaerococcus faecalis sp. nov., Isolated from Swine Faeces.

An obligate anaerobic, Gram-stain-positive, non-spore forming, non-motile, catalase and oxidase-negative, coccoid-shaped bacterium designated AGMB00486T was isolated from swine faeces. The optimal growth of the isolate occurred at pH 8.0 and 37 ℃. Furthermore, the growth was observed in the presence of up to 4% (w/v) NaCl but not at salinity levels higher than 5%. The phylogenetic analysis based on the 16S rRNA gene sequences revealed that strain AGMB00486T was a member of the genus Anaerococcus and that the isolate was most closely related to Anaerococcus vaginalis KCTC 15028T (96.7% 16S rRNA gene sequence similarity) followed by Anaerococcus hydrogenalis KCTC 15014T (96.7%) and Anaerococcus senegalensis KCTC 15435T (96.3%). Whole-genome sequence analysis determined that the DNA G+C content of strain AGMB00486T was 30.1 mol%, and the genome size, numbers of tRNA and rRNA genes were 2,268,866 bp, 47 and 8, respectively. The average nucleotide identity values between strain AGMB00486T and the three related type strains were 77.0, 77.4 and 77.2%, respectively. The major cellular fatty acids (> 10%) of strain AGMB00486T were C14:0, C16:0 and C16:0 DMA. Accordingly, these distinct phenotypic and phylogenetic properties revealed that strain AGMB00486T represents a novel species, for which the name Anaerococcus faecalis sp. nov. is proposed. The type strain is AGMB00486T (= KCTC 15945T = CCTCC AB 202009T).

Transdifferentiation of α-1,3-galactosyltransferase knockout pig bone marrow derived mesenchymal stem cells into pancreatic ß-like cells by microenvironment modulation.

OBJECTIVE: To evaluate the pancreatic differentiation potential of α-1,3-galactosyltransferase knockout (GalTKO) pig-derived bone marrow-derived mesenchymal stem cells (BM-MSCs) using epigenetic modifiers with different pancreatic induction media. METHODS: The BM-MSCs have been differentiated into pancreatic ß-like cells by inducing the overexpression of key transcription regulatory factors or by exposure to specific soluble inducers/small molecules. In this study, we evaluated the pancreatic differentiation of GalTKO pig-derived BM-MSCs using epigenetic modifiers, 5-azacytidine (5-Aza) and valproic acid (VPA), and two types of pancreatic induction media - advanced Dulbecco's modified Eagle's medium (ADMEM)-based and N2B27-based media. GalTKO BM-MSCs were treated with pancreatic induction media and the expression of pancreas-islets-specific markers was evaluated by real-time quantitative polymerase chain reaction, Western blotting, and immunofluorescence. Morphological changes and changes in the 5'-C-phosphate-G-3' (CpG) island methylation patterns were also evaluated. RESULTS: The expression of the pluripotent marker (POU class 5 homeobox 1 [OCT4]) was upregulated upon exposure to 5-Aza and/or VPA. GalTKO BM-MSCs showed increased expression of neurogenic differentiation 1 in the ADMEM-based (5-Aza) media, while the expression of NK6 homeobox 1 was elevated in cells induced with the N2B27-based (5-Aza) media. Moreover, the morphological transition and formation of islets-like cellular clusters were also prominent in the cells induced with the N2B27-based media with 5-Aza. The higher insulin expression revealed the augmented trans-differentiation ability of GalTKO BM-MSCs into pancreatic ß-like cells in the N2B27-based media than in the ADMEM-based media. CONCLUSION: 5-Aza treated GalTKO BM-MSCs showed an enhanced demethylation pattern in the second CpG island of the OCT4 promoter region compared to that in the GalTKO BM-MSCs. The exposure of GalTKO pig-derived BM-MSCs to the N2B27-based microenvironment can significantly enhance their trans-differentiation ability into pancreatic ß-like cells.

Peribacillus faecalis sp. nov., a moderately halophilic bacterium isolated from the faeces of a cow.

A Gram-stain-positive, facultatively anaerobic, endospore-forming, rod-shaped strain, AGMB 02131T, which grew at 20-40 °C (optimum 30 °C), pH 3.0-11.0 (optimum pH 4.0) and in the presence of 0-18 % (w/v) NaCl (optimum 10 %), was isolated from a cow faecal sample and identified as a novel strain using a polyphasic taxonomic approach. The phylogenetic analysis based on 16S rRNA gene sequences along with the whole genome (92 core gene sets) revealed that AGMB 02131T formed a group within the genus Peribacillus, and showed the highest sequence similarity with Peribacillus endoradicis DSM 28131T (96.9 %), following by Peribacillus butanolivorans DSM 18926T (96.6 %). The genome of AGMB 02131T comprised 70 contigs, the chromosome length was 4 038 965 bp and it had a 38.5 % DNA G+C content. Digital DNA-DNA hybridization revealed that AGMB 02131T displayed 21.4 % genomic DNA relatedness with the most closely related strain, P. butanolivorans DSM 18926T. AGMB 02131T contains all of the conserved signature indels that are specific for members of the genus Peribacillus. The major cellular fatty acids (>10 %) of AGMB 02131T were C18 : 1ω9c, C18:0 and C16 : 0. The major polar lipids present were diphosphatidylglycerol, phosphatidylglycerol and phosphatidylethanolamine. On the basis of the phenotypic, phylogenetic, genomic and chemotaxonomic features, AGMB 02131T represents a novel species of the genus Peribacillus, for which the name Peribacillus faecalis sp. nov. is proposed. The type strain is AGMB 02131T (=KCTC 43221T=CCTCC AB 2020077T).

Preparation of Nanoemulsions of Vitamin A and C by Microfluidization: Efficacy on the Expression Pattern of Milk-Specific Proteins in MAC-T Cells.

In this study, we prepared stabilized vitamin A and C nanoemulsions, and investigated their efficacy on milk-specific proteins in bovine mammary epithelial cells (MAC-T). Emulsions of vitamin A (vit-A) and C (vit-C) were prepared using Lipoid S 75 and microfluidization. The particle size and polydispersity index (PDI) of nanoemulsified vit-A and vit-C were studied. The cytotoxic effect of nanoemulsion-free and nanoemulsified vit-A and vit-C was determined by an MTT assay. In addition, the efficacy of nanoemulsified vit-A and vit-C on the in vitro expression pattern of milk-specific proteins in MAC-T cells was investigated by quantitative RT-PCR. The results showed that the efficacies of stabilized nanoemulsions of vit-A and vit-C were 100% and 92.7%, respectively. The particle sizes were around 475.7 and 225.4 nm, and the zeta potentials were around -33.5 and -21.3 mV, respectively. The expression changes of αs2-, ß- and κ-casein were higher in the presence of a stabilized nanoemulsion of vit-A, compared with nanoemulsion-free vit-A. Furthermore, the expression changes of αs2- and ß-casein were lower and that of κ-casein was higher in the presence of a stabilized nanoemulsion of vit-C, compared with nanoemulsion-free vit-C. Thus, our findings demonstrate the efficacy of nanoemulsified vit-A and vit-C in changing the expression of milk-specific proteins in MAC-T cells.

In vitro immune responses of porcine alveolar macrophages reflect host immune responses against porcine reproductive and respiratory syndrome viruses.

BACKGROUND: Currently, an in vitro immunogenicity screening system for the immunological assessment of potential porcine reproductive and respiratory syndrome virus (PRRSV) vaccine candidates is highly desired. Thus, in the present study, two genetically divergent PRRSVs were characterized in vitro and in vivo to identify an in vitro system and immunological markers that predict the host immune response. Porcine alveolar macrophages (PAMs) and peripheral blood mononuclear cells (PBMCs) collected from PRRSV-negative pigs were used for in vitro immunological evaluation, and the response of these cells to VR2332c or JA142c were compared with those elicited in pigs challenged with the same viruses. RESULTS: Compared with VR2332c or mock infection, JA142c induced increased levels of type I interferons and pro-inflammatory cytokines (TNF-α, IL-1α/ß, IL-6, IL-8, and IL-12) in PAMs, and these elevated levels were comparable to the cytokine induction observed in PRRSV-challenged pigs. Furthermore, significantly greater numbers of activated CD4+ T cells, type I helper T cells, cytotoxic T cells and total IFN-γ+ cells were observed in JA142c-challenged pigs than in VR2332c- or mock-challenged pigs. CONCLUSIONS: Based on these results, the innate immune response patterns (particularly IFN-α, TNF-α and IL-12) to specific PRRSV strains in PAMs might reflect those elicited by the same viruses in pigs.

Suberoylanilide hydroxamic acid during in vitro culture improves development of dog-pig interspecies cloned embryos but not dog cloned embryos.

This study was conducted to investigate whether the treatment of dog to pig interspecies somatic cell nuclear transfer (iSCNT) embryos with a histone deacetylase inhibitor, to improve nuclear reprogramming, can be applied to dog SCNT embryos. The dog to pig iSCNT embryos were cultured in fresh porcine zygote medium-5 (PZM-5) with 0, 1, or 10 µM suberoylanilide hydroxamic acid (SAHA) for 6 h, then transferred to PZM-5 without SAHA. Although there were no significant differences in cleavage rates, the rates of 5-8-cell stage embryo development were significantly higher in the 10 µM group (19.5 ± 0.8%) compared to the 0 µM groups (13.4 ± 0.8%). Acetylation of H3K9 was also significantly higher in embryos beyond the 4-cell stage in the 10 µM group compared to the 0 or 1 µM groups. Treatment with 10 µM SAHA for 6 h was chosen for application to dog SCNT. Dog cloned embryos with 0 or 10 µM SAHA were transferred to recipients. However, there were no significant differences in pregnancy and delivery rates between the two groups. Therefore, it can be concluded that although porcine oocytes support nuclear reprogramming of dog fibroblasts, treatment with a histone deacetylase inhibitor that supports nuclear reprogramming in dog to pig iSCNT embryos was not sufficient for reprogramming in dog SCNT embryos.

Ascorbic acid increases demethylation in somatic cell nuclear transfer embryos of the pig (Sus scrofa).

OBJECTIVE: Investigated the effect and mechanism of ascorbic acid on the development of porcine embryos produced by somatic cell nuclear transfer (SCNT). METHODS: Porcine embryos were produced by SCNT and cultured in the presence or absence of ascorbic acid. Ten-eleven translocation 3 (TET3) in oocytes was knocked down by siRNA injection. After ascorbic acid treatment, reprogramming genes were analyzed by realtime reverse transcription-polymerase chain reaction (RT-PCR). Furthermore, relative 5-methylcytosine and 5-hydroxymethylcytosine content in pronucleus were detected by realtime PCR. RESULTS: Ascorbic acid significantly increased the development of porcine embryos produced by SCNT. After SCNT, transcript levels of reprogramming genes, Pou5f1, Sox2, and Klf were significantly increased in blastocysts. Furthermore, ascorbic acid reduced 5-methylcytosine content in pronuclear embryos compared with the control group. Knock down of TET3 in porcine oocytes significantly prevents the demethylation of somatic cell nucleus after SCNT, even if in the presence of ascorbic acid. CONCLUSION: Ascorbic acid enhanced the development of porcine SCNT embryos via the increased TET3 mediated demethylation of somatic nucleus.

Transcriptome profiles of organ tissues from pigs experimentally infected with African swine fever virus in early phase of infection.

African swine fever, caused by African swine fever virus (ASFV), is a highly contagious and fatal disease that poses a significant threat to the global pig industry. The limited information on ASFV pathogenesis and ASFV-host interactions has recently prompted numerous transcriptomic studies. However, most of these studies have focused on elucidating the transcriptome profiles of ASFV-infected porcine alveolar macrophages in vitro. Here, we analyzed dynamic transcriptional patterns in vivo in nine organ tissues (spleen, submandibular lymph node, mesenteric lymph node, inguinal lymph node, tonsils, lungs, liver, kidneys, and heart) obtained from pigs in the early stages of ASFV infection (1 and 3 d after viremia). We observed rapid spread of ASFV to the spleen after viremia, followed by broad transmission to the liver and lungs and subsequently, the submandibular and inguinal lymph nodes. Profound variations in gene expression patterns were observed across all organs and at all time-points, providing an understanding of the distinct defence strategies employed by each organ against ASFV infection. All ASFV-infected organs exhibited a collaborative response, activating immune-associated genes such as S100A8, thereby triggering a pro-inflammatory cytokine storm and interferon activation. Functional analysis suggested that ASFV exploits the PI3K-Akt signalling pathway to evade the host immune system. Overall, our findings provide leads into the mechanisms underlying pathogenesis and host immune responses in different organs during the early stages of infection, which can guide further explorations, aid the development of efficacious antiviral strategies against ASFV, and identify valuable candidate gene targets for vaccine development.

Impact of rumen cannulation surgery on rumen microbiota composition in Hanwoo steers.

Rumen cannulation is a surgical technique used to collect rumen contents from ruminants. However, rumen cannulation surgery may potentially impact the composition of the rumen microbiota. This study aimed to examine the longitudinal alterations in the rumen microbiota composition of Hanwoo steers after cannulation surgery. In this study, eight Hanwoo steers were used; four steers underwent rumen cannulation surgery (cannulation group), while the remaining four were left intact (control group). Rumen samples were collected from all eight steers using the stomach tubing method on the day before surgery (day 0) and on postoperative days 1, 4, 7, 10, 14, 17, 21, 24, and 28, resulting in 80 samples (10 timepoints × 8 animals). The microbiota of all 80 samples were analyzed using 16S rRNA gene amplicon sequencing with Quantitative Insights into Microbial Ecology version 2 (QIIME2). There were no significant differences (p > 0.05) in all major phyla and most major genera representing at least 0.5% of total sequences across all 80 samples between the control and cannulation groups on the preoperative and postoperative days. However, while the alpha diversity indices did not differ (p > 0.05) between the two groups on the preoperative day, they significantly differed (p < 0.05) between the two groups on the postoperative days. Further, the overall microbial distribution based on both unweighted and weighted principal coordinate analysis plots significantly differed (p < 0.05) between the two groups on both the preoperative and postoperative days. Orthogonal polynomial contrasts indicated that major genera and microbial diversity in the cannulation group decreased following surgery but returned to their initial states by postoperative day 28. In conclusion, this study demonstrates that rumen cannulation surgery affects some major taxa and microbial diversity, suggesting that the rumen cannulation method can alter the composition of rumen microbiota in Hanwoo steers.

Exosomes isolation from bovine serum: qualitative and quantitative comparison between ultracentrifugation, combination ultracentrifugation and size exclusion chromatography, and exoEasy methods.

Exosomes have been extensively studied as disease biomarker in humans, given their role in transporting bioactive molecules. However, despite the great potential of exosomes as noninvasive diagnostic markers and therapeutic nanocarriers for bovine diseases, few studies have been conducted on bovine exosome. Thus, this study aimed to quantitatively and qualitatively compare three isolation methods to identify a suitable method for bovine serum. Exosomes were isolated using ultracentrifugation alone (UC), a combination of ultracentrifugation and size exclusion chromatography (US), or membrane affinity-based exoEasy kit (EE). Isolated particles were evaluated using a range of complementary techniques. Transmission electron microscopy showed that all three isolation methods resulted in particles with a cup-shaped morphology. The particle concentration measured by nanoparticle trafficking analyzer of US was lower compared to those of UC and EE method. As a result of immunoblotting, exosome markers including TSG101, CD81, and HSP70 were detected in US particles, while in UC and EE, only TSG101 expression was confirmed. Particles isolated from UC and EE showed a contamination with the blood protein albumin, whereas particles from US did not show albumin contamination. In addition, to evaluate the possibility of using exosomes as biomarkers, the profiles of the small RNA in the exosomes were compared using the bioanalyzer 2100. As a result, in the EE method, the band of small RNA (25-200 nt) was most prominent, and in the US methods, a distinct band was observed in the small RNA range. Collectively, the purity of exosomes without non-exosomal contamination was highest in the US method. However, for the detection of small RNA, the EE method was found to be the most suitable. Therefore, the results suggest that the optimal isolation method varies depending on the specific purpose of exosome isolation.

Lactiplantibacillus argentoratensis AGMB00912 alleviates salmonellosis and modulates gut microbiota in weaned piglets: a pilot study.

This study aimed to evaluate the efficacy of Lactiplantibacillus argentoratensis AGMB00912 (LA) in reducing Salmonella Typhimurium infection in weaned piglets. The investigation focused on the influence of LA on the gut microbiota composition, growth performance, and Salmonella fecal shedding. The results indicated that LA supplementation significantly improved average daily gain and reduced the prevalence and severity of diarrhea. Fecal analysis revealed reduced Salmonella shedding in the LA-supplemented group. Furthermore, LA notably altered the composition of the gut microbiota, increasing the levels of beneficial Bacillus and decreasing those of harmful Proteobacteria and Spirochaetes. Histopathological examination showed less intestinal damage in LA-treated piglets than in the controls. The study also observed that LA affected metabolic functions related to carbohydrate, amino acid, and fatty acid metabolism, thereby enhancing gut health and resilience against infection. Short-chain fatty acid concentrations in the feces were higher in the LA group, suggesting improved gut microbial activity. LA supplementation enriched the population of beneficial bacteria, including Streptococcus, Clostridium, and Bifidobacterium, while reducing the number of harmful bacteria, such as Escherichia and Campylobacter. These findings indicate the potential of LA as a probiotic alternative for swine nutrition, offering protective effects to the gut microbiota against Salmonella infection.

Synchronization using PGF(2α) and estradiol with or without GnRH for timed artificial insemination in dairy cows.

This study examined the use of PGF(2α) and estradiol benzoate (EB) either with or without GnRH to synchronize estrus in dairy cows for timed artificial insemination (TAI) under field conditions. First, Holstein dairy cows with a corpus luteum (CL) received 500 µg cloprostenol and were then randomly allocated to three groups: no further treatment (control, n=236); treatment with 1 mg EB 56 h after cloprostenol (EB group, n=339); or treatment with 1 mg EB 56 h after cloprostenol followed by treatment with 100 µg gonadorelin 24 h later (EB + GnRH group, n=216). All cows received TAI 80 h after the cloprostenol injection. In a second experiment, Holstein dairy cows with a CL received 500 µg cloprostenol and were then randomly allocated to two groups: treatment with 2 mg EB 36 h later (EB group, n=284) or treatment with 2 mg EB 36 h after cloprostenol followed by 100 µg gonadorelin 24 h later (EB + GnRH group, n=229). All cows received TAI 24 h after the EB injection. Logistic analyses revealed that the odds ratio for the probability of pregnancy when 1 mg EB was administered 56 h following cloprostenol was 1.9 and 2.0 times (P<0.001) higher in the EB (39.5%) and EB + GnRH groups (40.7%), respectively, compared with the control group (25.8%). However, pregnancy rates in cows receiving 2 mg EB 24 h following cloprostenol showed no difference compared with cows treated with EB only (32.4%) or with EB + GnRH (35.8%). These results indicate that a synchronization protocol comprising PGF(2α) and EB could be used for TAI in dairy herds under field conditions.

Genome-wide association study of mammary gland tumors in Maltese dogs.

Background: A genome-wide association study (GWAS) is a valuable tool for investigating genetic and phenotypic variation in many diseases. Objective: The objective of this study was to identify variations in the genomes of Maltese dogs that are associated with the mammary gland tumor (MGT) phenotype and to assess the association between each biological condition and MGT phenotype in Maltese dogs. Methods: DNA was extracted from 22 tumor samples and 11 whole blood samples from dogs with MGTs. Genome-wide single-nucleotide polymorphism (SNP) genotyping was performed, and the top 20 SNPs associated with various conditions and genetic variations were mapped to their corresponding gene locations. Results: The genotyping process successfully identified 173,662 loci, with an overall genotype completion rate of 99.92%. Through the quality control analysis, 46,912 of these SNPs were excluded. Allelic tests were conducted to generate Manhattan plots, which showed several significant SNPs associated with MGT phenotype in intergenic region. The most prominent SNP, located within a region associated with transcription and linked to the malignancy grade of MGT, was identified on chromosome 5 (p = 0.00001) though there may be lack of statistical significance. Other SNPs were also found in several genes associated with oncogenesis, including TNFSF18, WDR3, ASIC5, STAR, and IL1RAP. Conclusion: To our knowledge, this is the first GWAS to analyze the genetic predisposition to MGT in Maltese dogs. Despite the limited number of cases, these analyzed data could provide the basis for further research on the genetic predisposition to MGTs in Maltese dogs.