MGF360-12L of ASFV-SY18 is an immune-evasion protein that inhibits host type I IFN, NF-κB, and JAK/STAT pathways
African swine fever virus (ASFV) causes feverous and hemorrhagic disease of domestic pigs and European wild boars with high mortality, yet no commercial vaccine is currently available. Several ASFV strains with natural deletion or gene-targeted knockout of multiple MGF360 and MGF505 genes are attenuated in vitro and in vivo, and can offer full protection against homologous challenge. However, the mechanisms underlying the protection are not fully understood. This study aims to investigate the effects of MGF360-12L of ASFV-SY18 on the cGAS-STING signaling pathway and explore the potential mechanisms. We identified that ASFV-SY18 MGF360-12L could inhibit cGAS-STING, TBK1, or IRF3-5D-stimulated IFN-β expression and ISRE activation. Specifically, MGF360-12L inhibits both the activation of PRD(III-I) in a dose-dependent manner and suppresses the exogenous expression of TBK1 and IRF3-5D. MGF360-12L could block NF-κB activation induced by overexpression of cGAS-STING, TBK1, IKKβ. Downstream of the IFN-β signaling, MGF360-12L blocks the ISRE promoter activation by reducing total protein level of IRF9. Moreover, MGF360-12L protein can inhibit IFN-β-mediated antiviral effects. In conclusion, our findings suggest that MGF360-12L is a multifunctional immune-evasion protein that inhibits both the expression and effect of IFN-β, which could partially explain the attenuation of relevant gene-deleted ASFV strains and shed light on the development of efficient ASFV live attenuated vaccines in the future.
Effects of β-1,3/1,6 glucan dietary supplements on some immunological and hematological health markers in Siberian sturgeon (Acipenser baerii) infected with Aeromonas hydrophila
This experimental study aimed to investigate some hematological and immunological changes as a result of Aeromanas hydrophila infection in Siberian sturgeon (Acipenser baerii Brandt, 1869). Their feeds were supplemented with β-1,3/1,6 glucan at different ratios, 250 mg/kg (βG250); 500 mg/kg (βG500) and 750 mg/kg (βG750). To create an experimental infection, 4×106 cfu/ml Aeromonas hydrophila inoculum was intraperitoneally injected to fish. 0.1 ml intraperitoneal bacteria injection was given to the fish in 12 of 15 tanks, each consisting of 10 fish (the fish in the control group were not given bacteria). Considering the βG500 and βG750 group fish as positive (C+) and negative control (C−) groups in terms of hematological parameters, it was found that RBC (erythrocyte) and Hb (hemoglobin) values, as well as RBC indices (MCV, MCHC, MCH) significantly increased. The immunological parameters, including WBC (leukocyte), leukocyte cell percentages (lymphocyte, monocyte, neutrophil, eosinophil), as well as cytokines, including IL-1β, IL-6, IL-8, and TNF-α values showed similar increases in the βG500 and βG750 groups. It was found that the addition of 500 and 750 mg/kg doses of β-1,3/1,6 glucan to the feed stimulated non-specific immunity of fish against bacterial agents and/or septicemic diseases and beta glucan at this dosage range was determined to be ideal for fish health and that it may be a herbal immunostimulant that can be an alternative to many medicaments.
THYROID EVALUATION IN SUSPICIOUS HYPOTHYROID ADULT DOGS BEFORE AND AFTER TREATMENT
The purpose of this study was to measure circulating TSH, T4 and fT4 concentrations in dogs submitted to a clinic visit for general symptoms (weight gain, polyuria and polydipsia, changes in hair coat). Twenty-eight dogs, 14 cross-breed and 14 purebreds (Golden Retriever, Labrador, Doberman), of both sexes (14 males and 14 females), aged 8 to 14 years, were assessed. No significant differences of circulating TSH, T4, fT4 concentrations between baseline and after therapeutic treatment nor between intact and neutered females were observed. Compared to baseline values, intact males showed higher TSH concentrations (p<0.01), and castrated males lower TSH concentrations (p<0.01) after therapeutic treatment. Compared to intact males, castrated males showed baseline TSH concentrations higher (p<0.01), but lower (p<0.01) after therapeutic treatment. No significant differences of T4 andfT4 concentrations between baseline conditions and after therapeutic treatment nor between intact and castrated males were observed. The experimental sample considered in this study falls within that casuistry involving elevated TSH concentrations but low serum T4 and fT4concentrations or close to the minimum physiological cut-off, in which the common clinical signs suggestive of hypothyroidism was, essentially, overweight and neglected appearance of the hair.
The Effect of Luteolin on Spermatological Parameters, Apoptosis, Oxidative Stress Rate in Freezing Rabbit Semen.
The aim of the present study was to determine the effects of Luteolin (LUT) on semen quality, oxidative stress, apoptosis, acrosomal integrity, mitochondrial membrane potential and dead sperm ratio in rabbits. Ejaculates from six New Zealand rabbits were collected, evaluated and pooled. The pooling was divided into five groups as control (no additive) LUT 25 µM, LUT 50 µM, LUT 100 µM and LUT 200 µM and LUT added. It was then filled into a falcon tube with Tris-based extender at a final concentration of approximately 35 x 106 spermatozoa. Diluated rabbit semen samples were drawn into frozen and thawed. Frozen semen straws were thawed at 37°C in 30 seconds. According to our findings, no statistical difference was found between all doses of luteolin and the control group in the CASA (computer assisted sperm analysis) analysis performed at 4 oC. However, total motility, progressive motility and rapid sperm percentage were found to be higher in the frozen and thawed rabbit semen at a dose of LUT 50 µM compared to the other groups (p<0.05). While amplitude of lateral head displacement (ALH) and beat cross-frequency (BCF) values were found at the lowest dose of LUT 200 µM, a statistically significant difference was observed between the other groups. When the flow cytometry results were examined, no statistical difference was found between the rate of dead sperm, acrosomal integrity, mitochondrial membrane potential and apoptosis rate. Morever, the H2O2 percentage was found to be lower in all experimental groups compared to the control group (p<0.001). In conclusion, the addition of LUT in long-term storage of rabbit semen provided a protective effect for spermatozoa with its antioxidative properties against damage caused by cryopreservation.
Endogenous carbon monoxide (CO) as a modulating factor of molecular biological clock in the hypothalamic structures involved in light transmission in pig and wild boar hybrid during long and short day season
Mature males of a wild boar-pig crossbreed, during long and short day season, were used for the study which demonstrate that chemical light carrier CO, regulates the expression of biological clock genes in hypothalamus via humoral pathways. Autologous blood with experimentally elevated concentrations of endogenous CO (using lamps with white light-emitting diodes) was infused into ophthalmic venous sinus via the right dorsal nasal vein. Molecular biology methods: qPCR and Western Blot were used to determine the expression of genes and biological clock proteins. The results showed that elevated endogenous CO levels through blood irradiation induces changes in genes expression involved in functioning of the main biological clock located in suprachiasmatic nuclei. Changes in the expression of the transcription factors Bmal1, Clock and Npas2 have a similar pattern in both structures where a very large decrease in gene expression was shown after exposure to elevated endogenous CO levels. The changes in the gene expression of PER 1-2, CRY 1-2, and REV-ERB α-β and ROR β are not the same for both POA and DH hypothalamic structures indicating that both structures respond differently to the received humoral signal. Obtained results indicate that CO is a chemical light molecule whose production in organism depends on the amount of light. An adequate amount of light is an essential factor for the proper functioning of the main biological clock.
Evaluation of the anti-mycobacterial activity and composition of Carlina acaulis L. root extracts
Nontuberculous mycobacteria (NTM) have recently emerged as important bacterial pathogens of animals and humans. Of particular concern is the high level of antimicrobial resistance displayed by these organisms, which complicates treatment and potential successful outcomes. Here, we evaluated the potential of Carlina acaulis L. as a source of novel anti-mycobacterial agents. Our goal was to measure the activity of aqueous, ethanol, and chloroform C. acaulis root extracts against 99 NTM strains. GC-MS spectroscopy analyses were performed to deliver qualitative and quantitative data on the composition of C. acaulis extract. In our study, we have shown for the first time the activity of C. acaulis extracts against NTM. The highest activity was exhibited by the chloroform extract, which inhibited the growth of more than 90% of the strains at the dose of 100 μg/mL (MIC90 = 100 μg/mL). The results of the GC-MS analysis of the C. acaulis chloroform extract contributed to the identification of 37 compounds, with carlina oxide as the most representative compound (69.52%) followed by 3,4-dihydro-2H-phenanthren-1-one (6.54%) and stigmast-5-en-3-ol (4.14%). Our results indicate that C. acaulis chloroform and ethanol extracts have potential for treatment of NTM infections and that this plant contains anti-mycobacterial compounds.
Incidence and molecular characterization of extended-spectrum beta-lactamase (ESBL)-producing Salmonella enterica and Escherichia coli of avifauna origin in Pakistan
Members of Enterobacteriaceae are known to produce extended-spectrum beta-lactamases (ESBL) which hydrolyze the beta-lactam group of antibiotics. The existence of ESBL-producing Salmonella enterica (S. enterica) and Escherichia coli (E. coli) harbored by urban avifauna was investigated in this study. Dropping samples (n= 180) were collected from six different bird species in the district Jhang, Punjab province, Pakistan. Isolation and identification of ESBL isolates were made by using cefotaxime-(4 mg/L) supplemented MacConkey agar and double disc synergy test (DDST). Polymerase chain reaction (PCR) was performed for the detection of four different ESBL genes including blaCTX-M, blaTEM, blaSHV and blaOXA. A total of 42.69% isolates were confirmed as ESBL via DDST including 30.64% S. enterica and 49.54% E. coli. The incidence of ESBL S. enterica and ESBL E. coli was found highest in egret (Ardea alba) and pigeon (Columba livia) as 64.28% and 78.95%, respectively. The blaCTX-M gene was detected in 57.89% and 64.81% of isolates of S. enterica and E. coli, respectively. Among other genes in S. enterica and E. coli, blaTEM (21.05%, 20.4%); blaSHV (15.78%, 9.26%), and blaOXA (5.26%, 5.56%) were detected, respectively. All of the tested isolates were found resistant to at least one of the thirteen antimicrobial agents except meropenem. To the best of our knowledge, this is the first study reporting the incidence and genetic diversity of ESBL bacteria associated with urban avifauna in Pakistan. The urban avifauna can serve as a potential subject of bio-surveillance to monitor the emergence of antimicrobial-resistant bacteria.
A review of tuberculosis and parasitic disease co-infection in ungulates, with regard to the potential threat to European bison (Bison bonasus)
Bovine tuberculosis (BTB) is a dangerous zoonosis which presents a serious problem for endangered species such as European bison (Bison bonasus). Little is known about the influence of parasitic co-infections on the course and diagnosis of tuberculosis in animals. The best known co-infection in cattle is Fasciola hepatica and Mycobacterium bovis. The aim of this study was to review the most recent literature regarding tuberculosis and parasite co-infection in ungulates and relate the results to European bison. Our findings indicate that any comprehensive diagnosis of BTB should include parasitological monitoring, and the possible impact of such invasions on cellular response-based tuberculosis tests should be taken into account. The diagnosis of BTB is complex, as is its pathogenesis, and parasitic infestations can have a significant impact on both. This should be taken into account during further research and monitoring of tuberculosis in European bison.
Investigation of tylosin and tilmicosin residues in meat by high-performance liquid chromatography method
In this study, the presence and level of macrolide group antibiotics (tylosin and tilmicosin) were analyzed by the High-Performance Liquid Chromatography (HPLC) method in a total of 126 raw meat samples, including 42 chicken breast and 84 beef neck , available for consumption in Burdur province. The method demonstrated good linearity (R2 > 0.999) over the assayed concentration range (0.10-10 μg/mL). Intra-day and inter-day recoveries were used to express the accuracy of the method at three different levels of 0.5, 1, 2.5 μg/mL. Intraday recoveries and relative standard deviation values ranged from 97.270 (0.054)% to 98.643 (0.061)%, and inter-day recoveries and relative standard deviation values ranged from 97.057 (0.070)% to 98.197(0.042)% for tylosin. Intraday recoveries and relative standard deviation values ranged from 96.360 (0.065)% to 98.153 (0.046)%, and inter-day recoveries and relative standard deviation values ranged from 96.050 (0.058)% to 97.053 (0.096)% for tilmicosin. The limit of detection (LOD) value was calculated as 0.473 µg/kg for tylosin, 0.481 µg/kg for tilmicosin; the limit of quantification (LOQ) value was calculated as 1.561 µg/kg for tylosin, 1.587 µg/kg for tilmicosin. In general, tylosin and tilmicosin were determined in the range of 8-256 μg/kg and 30-447 μg/kg, respectively, in chicken breast meat samples; Also, they were detected in the range of 36-1209 μg/kg and 30-1102 μg/kg, respectively, in beef neck meat samples. When the obtained data were evaluated, it was found that the residues of tylosin and tilmicosin in chicken and beef meats in the market were at a much higher level than the acceptable limits specified in the regulations. This creates serious problems in terms of the ecosystem, food technology, and public health and causes significant economic losses.
Investigation of Salmonella spp. and Escherichia coli in the Snake-eyed lizard (Ophisops elegans) (Sauria, Lacertidae) in the Çankırı Province of Turkey
Zoonoses are frequently associated with wild animals. Research on reptiles either living in their natural habitat or kept as pet animals has shown that these animals frequently serve as the asymptomatic hosts of bacterial zoonotic agents, including Salmonella spp. and Escherichia coli. Studies have shown the potential of reptiles to transmit these pathogens to humans and other animals. Epidemiological research on the herpetofauna of various regions has demonstrated the high potential of reptiles as a reservoir of Salmonella spp. In the present study, Salmonella spp. were not isolated or identified from the snake-eyed lizard. Out of 150 cloacal swab samples of snake-eyed lizard 25 (16.7%) E. coli were isolated and out of these 4 (2.7%) were identified to be E. coli O157:H7 by PCR. The results suggest that Ophisops elegans could be involved in the transmission of E. coli, rather than Salmonella spp. This study demonstrates for the first time that the snake-eyed lizard acts as a cloacal carrier of E. coli O157:H7 and presents data that may aid in preventing the transmission of this strain to humans.
SNP panel for evaluation of genetic variability and relationship in roe deer
Blood samples from forty-six roe deer (Capreolus capreolus) acquired during officially approved hunting in six hunting divisions throughout Poland were used to isolate the genomic DNA. All individuals were genotyped by MD_Bovine BeadChip (Illumina) for 46.750 Single Nucleotide Polymorphism (SNP) markers. SNPs of inappropriate clusters, with a marker call rate lower than 90% and with a minor allele frequency (MAF) lower than 0.01, located on sex chromosomes and mitochondrial DNA, were removed. Altogether, 21.033 SNP markers were included for further analysis. Observed and expected heterozygosity amounted to 0.098 and 0.119, respectively. Among 21.033 markers, a panel of 148 SNPs were selected for relationship analysis. They were unlinked and had a MAF higher than 0.2. This set of SNPs showed a probability of parentage exclusion of 1.29×10-6 and 2.37×10-19 for one, and two known parents, respectively. The probability of identity was estimated at 1.8×10-40. The probabilities obtained in this study are sufficient for the monitoring and effective management of the genetic diversity of roe deer in Poland and is a cost-effective complementary tool for forensic applications.
Review and future perspectives on the integration characteristics for equine lentivirus in host genome
Despite over 40 years of research on the human immunodeficiency virus type 1 (HIV-1) vaccine, we still lack considerable progress. Equine infectious anemia virus (EIAV) is a lentivirus in the Retroviridae family, akin to HIV-1 in genome structure and antigenicity. EIA is an important infectious disease in equids, characterized by anemia, persistent infection, and repeated fevers. The EIAV attenuated vaccine in China is the only lentiviral vaccine used on a large scale. Elucidating the mechanism of waning and induction of protective immunity from this attenuated vaccine strain will provide a critical theoretical basis and reference point for vaccine research, particularly in the development of lentivirus vaccines, with far-reaching scientific value and social significance. In this paper, we summarize the information related to EIAV integration site selection, particularly for the Chinese EIAV attenuated vaccine strains on the equine genome. This may improve our mechanistic understanding of EIAV virulence reduction at the host genome level. The obtained data may help elucidate the biological characteristics of EIAV, particularly the Chinese attenuated EIAV vaccine strain, and provide valuable data regarding retroviral infections, particularly lentiviral infection and associated therapeutic vectors.
Occurrence of emerging ruminant viruses in goats in Poland
Health status of Polish goat population in regard to the viral diseases remained mostly unknown. In order to determine serological status of Polish goats for selected emerging ruminant viruses, 365 serum samples collected between 2017 and 2019 in 36 districts within 10 of Polish provinces, were tested. No antibodies specific to Peste de Petite Ruminants Virus (PPRSV) and capripoxviruses (CaPV) were found in any of the tested animals. Only single individual (0.27%) was seropositive to Blutongue Virus (BTV). Antibodies directed to Schmallenberg Virus (SBV) were detected in 46 goats which represented 12.6% of tested population. No association between seropositivity to SBV and year of sampling, province of origin, gender and age was found. In conclusion, among studied viral pathogens, currently only SBV seemed to be important for epidemiological status of Polish goats.
Microbiological studies of the round goby (Neogobius melanostomus) from the Puck Bay, southern Baltic Sea
In summer 2017 numerous dead round gobies (Neogobius melanostomus) and individuals covered with white coating were observed in the Puck Bay. The aim of our research was to determine the microbiological composition of the round goby from the Puck Bay, focusing on the presence of pathogens. Bacteria were identified by biochemical methods and, by sequencing of 16S rRNA. The dominant bacterial species were Shewanella baltica, Pseudomonas spp. and Aeromonas spp. – opportunistic pathogens, commonly present in many fish species, which may become harmful for the organism in unfavorable conditions. It was the first trial to determine the composition of the bacterial flora of N. melanostomus from that area.
Development of TaqMan-based real-time PCR assay based on the E1 gene for the quantitative detection of the Getah virus
To develop a sensitive, specific, and rapid approach for the detection Getah virus (GETV), a set of primers targeting the conserved region of the E1 gene was created. The TaqMan-based real-time PCR method for GETV detection was developed by optimizing the reaction conditions. The method demonstrated excellent specificity, and amplification did not occur with the causative agents of all prevalent swine viral infections (CSFV, PRRSV, PRV, PEDV, PTV, and JEV), except GETV. Additionally, upon assessing the sensitivity of the method, the minimum detection limit for GETV was found to be 5.94 copies/μL, which is 10 times higher than that of the traditional PCR approach. Further, the intra- and inter-assay variation coefficients were less than 1%, demonstrating good repeatability. Moreover, GETV was found in 10 of the 20 field serum samples using real-time PCR but only in three of the samples using traditional PCR. Consequently, the first GETV TaqMan-based real-time PCR approach based on the E1 gene was developed for GETV pathogenic diagnoses, and this exhibited high specificity, sensitivity, and repeatability. This assay is practical for the pathogenic diagnosis and epidemiology of GETV.
The study on bactericidal effect and ultrastructural alterations of chlorocresol nanoemulsion disinfectant against Staphylococcus aureus
Chlorocresol nanoemulsion disinfectant (CND) is an environmental disinfectant prepared with nanoemulsion as its drug carrier. This study aimed to investigate the bactericidal effect of CND on Staphylococcus aureus (S. aureus) and its effect on bacterial ultrastructure. The neutralizing effect of CND against S. aureus was first screened by suspension quantitative evaluation experiment procedure of neutralizer. Disinfection performance was evaluated by the determination of Minimal Inhibitory Concentration (MIC) and Minimum Bactericidal Concentration (MBC), quantitative bactericidal experiment, and comparative experiment of disinfection performance between 0.1% CND and 0.1% chlorocresol aqueous solution. Meanwhile, the effect of CND on the ultrastructure of S. aureus was investigated with scanning electron microscope (SEM) and transmission electron microscope (TEM) to preliminarily explore the bactericidal mechanism. The results showed that 3% Tween-80 in PBS could be screened as the neutralizer of CND against S. aureus. MIC and MBC were 100 μg/mL and 200 μg/mL, respectively. The bactericidal rates were all 100% when 0.06% and 0.08% disinfectant acted for 15 and 5 min, respectively. Furthermore, compared with 0.1% chlorocresol aqueous solution, the bactericidal effect of 0.1% CND was significantly enhanced (p<0.01). After treatment with CND for 10 min, SEM observation showed that the morphology of S. aureus cells were changed and the integrity destroyed. TEM observation showed that the cell shape changed, and the structures of the cell wall, cell membrane and cytoplasm were damaged in varying degrees. CND showed the strong bactericidal effect on S. aureus and could cause ultrastructure alterations of S. aureus.
The effect of tilmicosin and diclofenac sodium combination on cardiac biomarkers in sheep
The aim of this study was to investigate the cardiotoxic effect of the combination of tilmicosin and diclofenac sodium in sheep. Thirty-two sheep were used and were randomly divided into four equal groups as tilmicosin (T), diclofenac sodium (D), tilmicosin+diclofenac sodium (TD) and control (C) group. Group T received a single dose of tilmicosin, Group D was administered diclofenac sodium once a day for 3 days, and group TD was administered diclofenac and tilmicosin at the same doses as group T and D. Group C received NaCl in a similar way. The blood samples were taken before dosing and at 4th, 8th, 24th and 72nd hour post-dosing for measurement of cardiac markers such as H-FABP, cTn-I, CK-MB. H-FABP level of group TD was found to be significantly (p<0.05) higher than of group C at the 8th, 24th and 72nd hour and group D and T at the 72nd hour. cTn-I and CK-MB levels of group TD were found significantly (p<0.05) higher compared with other groups. In conclusion, the combined use of tilmicosin and diclofenac in sheep causes an increase in cardiac biomarkers and it can be stated that this combination of drugs may cause cardiac damage.
Identification of antimicrobial resistance genes in intestinal content from Coyote (Canis latrans).
Antibiotic resistance has become a global public health concern in the last few years. Given the widespread rate of recurrence, increasing attention is being turned toward environmental pathways that potentially contribute to antibiotic resistance genes (ARGs) dissemination outside the clinical realm. In this study, a metagenome analysis of intestinal virus-like particle fraction (VLPs) from a wild coyote (Canis latrans) revealed for the first time, multiple ARGs, such as B-lactamases and multidrug efflux pumps. Estimating ARGs presence in natural environments is critical to avoid the emergence of resistant strains.
Clinical effects of combined Lactobacillus paracasei and kestose on canine atopic dermatitis
Probiotics and prebiotics are viable bacteria with beneficial effects on the host and components that selectively act on the beneficial commensal bacteria, respectively. The combined use of probiotics and prebiotics is termed synbiotics. Probiotic intake improves dysbiosis in the intestinal microbiota and can positively affect canine atopic dermatitis (CAD). However, clinical studies on improvements in CAD using synbiotics remain limited. In this study, 15 dogs with CAD who received prednisolone, a synthetic glucocorticoid (GC) used in the treatment of CAD, for more than 90 days were continuously treated with Lactobacillus paracasei M-1 from fermented food as a probiotic, and the trisaccharide kestose as a prebiotic, for 90 days to determine their synbiotic effects on CAD. The CAD symptoms were evaluated using the canine atopic dermatitis lesion index (CADLI) and pruritus visual analog scores (PVAS). The total prednisolone use for 90 days pre- and post-administration was also evaluated. Synbiotic administration significantly reduced the CADLI (pre: median, 28.0 [22.0−40.0]; post: median, 12.0 [10.0−19.0]; p < 0.01) and PVAS (pre: median, 6.0 [5.0−7.0]; post: median, 2.0 [2.0−3.5]; p < 0.01) scores, and reduced the total prednisone use over 90 days (pre: 112.0 (25−450) mg; post: 80.0 (18.0−300.0) mg; p < 0.05) in the 15 dogs. Thus, the synbiotic activity of L. paracasei M-1 and kestose can improve CAD.