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Hypoxia is an important characteristic of Tibetan plateau environment. It can lead to apoptosis, but the mechanism of apoptosis caused by hypoxic stress needs further clarification. Here, cattle kidney cell MDBK were used as cell model. The effect of hypoxic stress on apoptosis and its molecular mechanism were explored. MDBK cells were treated with hypoxic stress, apoptosis and mitochondrial apoptotic pathway were significantly increased, and the expression of B-cell lymphoma 6 (BCL6) was significantly decreased. Overexpressing or inhibiting BCL6 demonstrated that BCL6 inhibited the apoptosis. And the increase of apoptosis controlled by hypoxic stress was blocked by BCL6 overexpressing. MDBK cells were treated with hypoxic stress, the expression and the nuclear localization of p53 were significantly increased. Overexpressing or inhibiting p53 demonstrated that hypoxic stress suppressed the expression of BCL6 through p53. Together, these results indicated that hypoxic stress induced the apoptosis of MDBK cells, and BCL6 was an important negative factor for this regulation process. In MDBK cells, hypoxic stress suppressed the expression of BCL6 through p53/BCL6-mitochondrial apoptotic pathway. This study enhanced current understanding of the molecular mechanisms underlying the regulation of apoptosis by hypoxic stress in MDBK cells. [ABSTRACT FROM AUTHOR]

This paper presents a potential strategy for the development of an antiviral agent against FMDV, specifically O Pan-Asia-2, A Iran 05 and SAT2/EGY/2012. Examining the potential use of interferon from the Bovine Ephemeral Fever Virus (BEFV) as a therapy is an intriguing approach. In order to test the cell toxicity and anti-FMDV in African green monkey kidney cell (Vero) and Baby Hamster Kidney Cell (BHK) cell lines up to serial dilutions of 10-7, BEFV was used to produce interferon in Vero and MDBK cell lines 12, 24, and 48 hours after cell infection. The in vitro findings showed that the cell safety and the ability of the prepared interferon to exhibit antiviral effects against various three FMDV serotypes, especially when administered shortly before or concurrently with the virus, are encouraging. The highest anti-FMDV effective dilution of the obtained interferon, either by Vero or MDBK cells, ranged from 10-3 to 10-4. This suggests that the interferon may be able to activate cellular antiviral mechanisms and disrupt FMDV infection. The next critical step will be conducting further in vivo studies to evaluate the efficacy, safety, and potential side effects of the interferon-based treatment. This will provide important insights into virus-host interactions and the broader therapeutic potential of this approach. Some key areas for future research could include: expanding the testing of the interferon against additional FMDV serotypes to assess its breadth of coverage; Optimizing the administration parameters, such as timing, dose, and route, to maximize the antiviral effect, Investigating the specific mechanisms by which the interferon inhibits FMDV replication and spread, Evaluating the interferon's impact on disease progression and transmission in animal models, Assessing any potential negative effects or toxicity in the treated animals. The development of effective antivirals against FMDV is critical for controlling outbreaks and protecting livestock. This research represents a promising line of investigation that warrants further study.

This paper presents a potential strategy for the development of an antiviral agent against FMDV, specifically O Pan-Asia-2, A Iran 05 and SAT2/EGY/2012. Examining the potential use of interferon from the Bovine Ephemeral Fever Virus (BEFV) as a therapy is an intriguing approach. In order to test the cell toxicity and anti-FMDV in African green monkey kidney cell (Vero) and Baby Hamster Kidney Cell (BHK) cell lines up to serial dilutions of 10-7, BEFV was used to produce interferon in Vero and MDBK cell lines 12, 24, and 48 hours after cell infection. The in vitro findings showed that the cell safety and the ability of the prepared interferon to exhibit antiviral effects against various three FMDV serotypes, especially when administered shortly before or concurrently with the virus, are encouraging. The highest anti-FMDV effective dilution of the obtained interferon, either by Vero or MDBK cells, ranged from 10-3 to 10-4. This suggests that the interferon may be able to activate cellular antiviral mechanisms and disrupt FMDV infection. The next critical step will be conducting further in vivo studies to evaluate the efficacy, safety, and potential side effects of the interferon-based treatment. This will provide important insights into virus-host interactions and the broader therapeutic potential of this approach. Some key areas for future research could include: expanding the testing of the interferon against additional FMDV serotypes to assess its breadth of coverage; Optimizing the administration parameters, such as timing, dose, and route, to maximize the antiviral effect, Investigating the specific mechanisms by which the interferon inhibits FMDV replication and spread, Evaluating the interferon's impact on disease progression and transmission in animal models, Assessing any potential negative effects or toxicity in the treated animals. The development of effective antivirals against FMDV is critical for controlling outbreaks and protecting livestock. This research represents a promising line of investigation that warrants further study. [ABSTRACT FROM AUTHOR]

Copyright of Chinese Journal of Preventive Veterinary Medicine / Zhongguo Yufang Shouyi Xuebao is the property of Chinese Journal of Preventive Veterinary Medicine Editorial Office and its content may not be copied or emailed to multiple sites or posted to a listserv without the copyright holder's express written permission. However, users may print, download, or email articles for individual use. This abstract may be abridged. No warranty is given about the accuracy of the copy. Users should refer to the original published version of the material for the full abstract. (Copyright applies to all Abstracts.)

Bovine viral diarrhea virus (BVDV) is a ubiquitous immunosuppressive etiological agent which is economically important for a wide host range in the livestock industry. Lactobacillus spp. has widely been using in the field of management and treatment of gastro-enteric disease for both humans and animals. The ability of Lacticaseibacillus casei MCJ protein-based metabolized to suppress BVDV infection in Madin–Darby Bovine Kidney cell line was demonstrated in this study. The protein-based metabolites were extracted from the cultured L. casei to obtain the safest and beneficial form of the probiotic bacteria. It is revealed that LPM have no cytotoxic effect and the cell viability remain more than 80% even after the cells are treated with 3000 µg/mL of LPM. The results of the plaque formation assay showed that LPM can reduce the viral infection rate. To know the mechanism of LPM for anti-BVDV activity, MDBK cells were exposed to LPM before, after and co-incubation of virus infection. The co-treatment of LPM with BVDV revealed the best results. The results suggest that the LPM has a potential anti-BVDV activity which could be a prospective candidate for the prevention and control of BVDV infection in an animal. [ABSTRACT FROM AUTHOR]

Butyrate is produced in the rumen from microbial fermentation and is related to several functions, including cell differentiation and proliferation. Butyrate supplementation in calves can accelerate rumen development. DNA-protein interactions, such as the CCCTC-binding factor (CTCF), play essential roles in chromatin organization and gene expression regulation. Although CTCF-binding sites have been identified recently in cattle, a deeper characterization, including differentially CTCF-binding sites (DCBS), is vital for a better understanding of butyrate's role in the chromatin landscape. This study aimed to identify CTCF-binding regions and DCBS under a butyrate-induced condition using ChIP-seq in bovine cells; 61,915 CTCF peaks were identified in the butyrate and 51,347 in the control. From these regions, 2265 DCBS were obtained for the butyrate vs. control comparison, comprising ~90% of induced sites. Most of the butyrate DCBS were in distal intergenic regions, showing a potential role as insulators. Gene ontology enrichment showed crucial terms for the induced DCBS, mainly related to cellular proliferation, cell adhesion, and growth regulation. Interestingly, the ECM-receptor interaction pathway was observed for the induced DCBS. Motif enrichment analysis further identified transcription factors, including CTCF, BORIS, TGIF2, and ZIC3. When DCBS was integrated with RNA-seq data, putative genes were identified for the repressed DCBS, including GATA4. Our study revealed promising candidate genes in bovine cells by a butyrate-induced condition that might be related to the regulation of rumen development, such as integrins, keratins, and collagens. These results provide a better understanding of the function of butyrate in cattle rumen development and chromatin landscape regulation. [ABSTRACT FROM AUTHOR]

Abstract Background Bovine viral diarrhea (BVD) which is caused by Bovine viral diarrhea virus (BVDV), is an acute, contagious disease. In spite of the use of vaccines and elimination projects, BVDV still causes severe economic losses to the cattle industry for the past few years. The current study presents a preliminary analysis of the pathogenic mechanisms from the perspective of protein expression levels in infected host cells at different points in time to elucidate the infection process associated with BVDV. Methods We used the isobaric tags for relative and absolute quantitation (iTRAQ) technology coupled with liquid chromatography-tandem mass spectrometric (LC–MS/MS) approach for a quantitative proteomics comparison of BVDV NADL-infected MDBK cells and non-infected cells. The functions of the proteins were deduced by functional annotation and their involvement in metabolic processes explored by KEGG pathway analysis to identify their interactions. Results There were 357 (47.6% downregulated, 52.4% upregulated infected vs. control), 101 (52.5% downregulated, 47.5% upregulated infected vs. control), and 66 (21.2% downregulated, 78.8% upregulated infected vs. control) proteins were differentially expressed (fold change > 1.5 or

Butyrate is produced by microbe fermentation in the rumen, and its supplementation results in rumen development. Butyrate-induced experiments are valuable in studying nutrient metabolism, cell growth, and functional genomics. This study aimed to characterize accessible chromatin regions and the dynamics of genomic accessibility prompted by butyrate-induced histone modifications in bovine cells. An average of 117,569 accessible chromatin regions were identified for all samples, and 21,347 differentially accessible regions (DARs) for butyrate. Most of the DARs were in distal intergenic regions, introns, and promoters. Gene ontology enrichment resulted in important terms related to the digestive system, regulation of epithelial cells, and cell adhesion. Ingenuity Pathway Analysis (IPA) identified critical networks (e.g., digestive system development, cell morphology and assembly, and cell cycle); canonical pathways (e.g., TGFβ, Integrin-linked kinase, and epithelial adherens junction); and upstream regulators (e.g., TGFβ1, FOS, JUNB, ATF3, and KLFs). Co-expression analysis further showcased the TGFβ and Integrin-linked kinase (ILK)-signaling pathways, which play roles in rumen development through cellular adhesions. This study is the first to provide a genome-wide characterization of differential, open chromatin regions for bovine cells by butyrate-induced treatment. These results provide valuable information for future studies of the butyrate functions in cattle gastrointestinal tract development. [ABSTRACT FROM AUTHOR]

It is reported that Notch3 and mTOR signaling pathways are involved in autophagy, and both can be activated by high glucose (HG). However, the relationship between Notch3 and mTOR and how Notch3 affects mTOR to regulate HG-induced autophagy in bovine kidney epithelial cells is still unclear. The purpose of this study is to explore how Notch3 affects mTOR to modulate HG-induced autophagy in bovine kidney cells. Our results showed that HG treatment significantly decreased the cell viability of MDBK cells in a dose-dependent manner. HG treatment significantly increased the expression of LC3-II/I ratio and Beclin1 protein and significantly decreased the expression of p62 protein. Consistently, LC3 fluorescence signal formation was detected by immunofluorescence in both dose and time-dependent manners. In addition, HG treatment significantly increased the expression of Notch3 protein and decreased the expression of the p-mTOR protein in both dose and time-dependent manners. Inhibition of Notch3 upregulated the expression of p-mTOR and p62 protein, and downregulated the expression of LC3-II/I ratio and Beclin1 protein. Besides, the function of Notch3 was investigated. In this study, inhibition of Notch3 activity significantly increased the viability of HG-stimulated MDBK cells. In summary, our results revealed that the Notch3-mediated mTOR signaling pathway was involved in HG-induced autophagy in MDBK cells. [ABSTRACT FROM AUTHOR]

In recent years, hunniviruses have been reported in a variety of animal species from many countries. Here, hunnivirus was detected in fecal samples from water buffaloes and named as BufHuV-GX-2106. The samples were inoculated into cultures of MDBK cells supplemented with TPCK trypsin and the BufHuV-GX-2106 strain was stably passaged and replicated. Electron microscopic analysis showed the BufHuV-GX-2106 virus particles were spherical and 20~30 nm in diameter. The complete genome of a plaque purified sample of BufHuV-GX-2106 was determined and analyzed. Genomic analysis revealed that the whole sequence of BufHuV-GX-2106 was ~7,601 nucleotides (nt) in length and consisted of a large open reading frame of 6,759nt, a 5′UTR, a 3'UTR and a poly(A) tail. The complete genome sequence of BufHuV-GX-2106 shares 68-85% nucleotide identities with other known hunnivirus strains, indicating high genetic heterogeneity among these viruses. Phylogenetic analysis showed that BufHuV-GX-2106 belonged to the Hunnivirus A species and was more closely related to ovine hunnivirus than other known viruses of this type. This study describes the first isolation and complete genome sequence of a hunnivirus strain from water buffaloes. In addition, this study will help to understand the mechanisms involved in the pathogenesis of Hunnivirus A among different animal species.

Copyright of Chinese Journal of Preventive Veterinary Medicine / Zhongguo Yufang Shouyi Xuebao is the property of Chinese Journal of Preventive Veterinary Medicine Editorial Office and its content may not be copied or emailed to multiple sites or posted to a listserv without the copyright holder's express written permission. However, users may print, download, or email articles for individual use. This abstract may be abridged. No warranty is given about the accuracy of the copy. Users should refer to the original published version of the material for the full abstract. (Copyright applies to all Abstracts.)

Butyrate is produced in the rumen from microbial fermentation and is related to several functions, including cell differentiation and proliferation. Butyrate supplementation in calves can accelerate rumen development. DNA-protein interactions, such as the CCCTC-binding factor (CTCF), play essential roles in chromatin organization and gene expression regulation. Although CTCF-binding sites have been identified recently in cattle, a deeper characterization, including differentially CTCF-binding sites (DCBS), is vital for a better understanding of butyrate’s role in the chromatin landscape. This study aimed to identify CTCF-binding regions and DCBS under a butyrate-induced condition using ChIP-seq in bovine cells; 61,915 CTCF peaks were identified in the butyrate and 51,347 in the control. From these regions, 2265 DCBS were obtained for the butyrate vs. control comparison, comprising ~90% of induced sites. Most of the butyrate DCBS were in distal intergenic regions, showing a potential role as insulators. Gene ontology enrichment showed crucial terms for the induced DCBS, mainly related to cellular proliferation, cell adhesion, and growth regulation. Interestingly, the ECM-receptor interaction pathway was observed for the induced DCBS. Motif enrichment analysis further identified transcription factors, including CTCF, BORIS, TGIF2, and ZIC3. When DCBS was integrated with RNA-seq data, putative genes were identified for the repressed DCBS, including GATA4. Our study revealed promising candidate genes in bovine cells by a butyrate-induced condition that might be related to the regulation of rumen development, such as integrins, keratins, and collagens. These results provide a better understanding of the function of butyrate in cattle rumen development and chromatin landscape regulation.

Poultry coccidiosis causes considerable economical losses to the livestock industry. Eimeria parasites are responsible for this disease. On a global scale, E. acervulina and E. tenella are amongst the most common Eimeria spp. infecting broilers. E. tenella is commonly used as infection model in in vivo and in vitro studies. On the other hand, E. acervulina has barely been studied under in vitro conditions. A well established and widely used in vitro model for E. tenella infection is the Madin-Darby bovine kidney cell line (MDBK); however, little is known regarding suitability of MDBK cells as host cells for E. acervulina. We infected MDBK monolayers with two different doses, 5 × 104 and 2 × 105, of E. acervulina sporozoites and evaluated cultures at 24 and 96 h post infection (hpi). For comparison, we ran an identical infection assay using E. tenella sporozoites. To assess parasite reproduction, the number of DNA copies of E. acervulina SCAR marker and E. tenella ITS-1 gene was quantified using real-time quantitative PCR. We found that the number of E. acervulina copies increased significantly at 24 hpi in comparison to E. tenella (p < 0.05). After 96 hpi, E. acervulina gene copies were considerably reduced while E. tenella continued to multiply (p < 0.05). Our results show that MDBK monolayers could be used for in vitro research aimed to study E. acervulina sporozoite cell invasion. Nevertheless, modifications of in vitro cultivation appear necessary to allow qualitative and quantitative studies over longer periods of parasite reproduction. [ABSTRACT FROM AUTHOR]

It is reported that Notch3 and mTOR signaling pathways are involved in autophagy, and both can be activated by high glucose (HG). However, the relationship between Notch3 and mTOR and how Notch3 affects mTOR to regulate HG-induced autophagy in bovine kidney epithelial cells is still unclear. The purpose of this study is to explore how Notch3 affects mTOR to modulate HG-induced autophagy in bovine kidney cells. Our results showed that HG treatment significantly decreased the cell viability of MDBK cells in a dose-dependent manner. HG treatment significantly increased the expression of LC3-II/I ratio and Beclin1 protein and significantly decreased the expression of p62 protein. Consistently, LC3 fluorescence signal formation was detected by immunofluorescence in both dose and time-dependent manners. In addition, HG treatment significantly increased the expression of Notch3 protein and decreased the expression of the p-mTOR protein in both dose and time-dependent manners. Inhibition of Notch3 upregulated the expression of p-mTOR and p62 protein, and downregulated the expression of LC3-II/I ratio and Beclin1 protein. Besides, the function of Notch3 was investigated. In this study, inhibition of Notch3 activity significantly increased the viability of HG-stimulated MDBK cells. In summary, our results revealed that the Notch3-mediated mTOR signaling pathway was involved in HG-induced autophagy in MDBK cells.

Abstract Background Crepis lacera is a plant from the Asteraceae family that is common in the Mediterranean region. Farmers believe that this plant may be deadly to small ruminants in areas of southern Italy. However, scientific evidence is lacking, and no proof exists that C. lacera is toxic to ruminants. Necropsies conducted on four sheep revealed lesions in their livers and kidneys. Results In the current study, we described sheep poisoning and isolated secondary metabolites from Crepis lacera to assess the metabolites’ biological activity both in vitro and in vivo. Phytochemical study of the aerial portions of Crepis lacera led to the isolation of five sesquiterpene lactones and two phenolic compounds. Cellular viability was evaluated in cell cultures of the bovine kidney cell line Madin Darby Bovine Kidney (MDBK) after incubation with phytochemicals. Our results showed that three sesquiterpene lactones, 8-epidesacylcynaropicrin-3-O-β-glucopyranoside (2), 8-epigrosheimin (3), and 8-β-hydroxydehydrozaluzanin C (4), were cytotoxic after 48 h of incubation. In addition, in the in vivo study, animals that received 1 mg/kg body weight (bw) of Crepis lacera extract and were then sacrificed after 48 h showed significant lesions in their liver, lungs and kidneys. These lesions were also found in rats that received 2 mg/kg bw of the same extract and sacrificed after 24 and 48 h. Conclusions These results validate the hypothesis that C. lacera is potentially dangerous when ingested in large quantities by grazing small domestic ruminants. Further studies are necessary to clarify the molecular mechanisms of Crepis spp. toxicity in animals.

Toxoplasma gondii and Neospora caninum are two major apicomplexan protozoan parasites with heteroxenous life cycles and worldwide distributions. The transplacental transmission of N. caninum causes bovine abortion, which resulting in serious economic losses to the dairy industry. Although T. gondii was also reported to cause abortions in pregnant woman and small ruminants, scarce cases about the symptom to the host cattle and the causality remains unknown. In this study, transcriptome analysis of Madin Darby bovine kidney (MDBK) cells infected with T. gondii and N. caninum was performed to uncover the differences in susceptibility of cattle to the two parasites. The results showed that 256 and 2225 differentially expressed genes (DEGs) were detected in cells infected with N. caninum and T. gondii , respectively. Moreover, significant biological differences were revealed by the functional analysis including GO and KEGG enrichment. One serpin peptidase inhibitor (SEPRINA14), which is associated with immunosuppression during pregnancy, was found to significantly decrease in cells infected with N. caninum and increase in cells infected with T. gondii -infected cells. Pattern recognition receptors TLR3 and NOD2 were also significantly upregulated in N. caninum -infected MDBK cells, but not in T. gondii. They could induce an increased inflammatory response leading to severe tissue damage. In addition, the interleukin 12 receptor subunit beta 2 (IL12β2), which plays an essential role in Th1 and Th2 cell differentiation and inflammatory bowel disease, was also markedly upregulated in the N. caninum infected cells, which led to an imbalance in the Th1 and Th2 cells by promoting the Th1 cellular response. Altogether, our findings recognized a new understanding on the differences between T. gondii and N. caninum infection of MDBK cells, where SEPRINA14, TLR3, NOD2, and IL12β2 may be the key genes that affect the difference in susceptibility of cattle to T. gondii and N. caninum , especially in pregnant animals. This study provides more clues as to why N. caninum is more likely to cause abortions in cattle. • First gene expression profiles of bovine-derived cells infected by the two parasites. • N. caninum participates in fewer immune pathways in MDBK cells than T. gondii. • Significantly differential expression of SEPRINA14, TLR3, NOD2 and IL12β2. • These key genes affect the susceptibility of cattle to T. gondii and N. caninum. [ABSTRACT FROM AUTHOR]

MicroRNAs (miRNAs) are endogenous ∼22 nt noncoding RNAs that control the translation initiation and stability of target genes in a sequence-specific manner and, thus, play important regulatory roles in animals and plants. Homologs of Dom34, called Pelota or PELO, are broadly conserved in eukaryotes and archaea. Biochemical and genetic studies indicate that eukaryotic Dom34/Pelota plays an important role in cell division, differentiation of germline stem cells, and stem cell self-renewal by controlling the expression of specific genes at the translational level. Additionally, it is reported that Pelota is specifically required for high efficiency synthesis of proteins in numerous viruses. In earlier studies, we found the Bos taurus bta-miR-2411 (shortly miR-2411 herein) was significantly upregulated by more than 2.1 times in bovine viral diarrhea virus (BVDV) strain NADL-infected Madin-Darby bovine kidney (MDBK) cells after 8 h post-infection (pi) compared to normal MDBK cells without BVDV infection. Moreover, miR-2411 overexpression significantly reduced the BVDV E1 mRNA level and viral titer. Nevertheless, the mechanisms of miR-2411 attenuating on viral replication remain unclear. Here, we report that miR-2411 as a novel microRNA regulates BVDV NADL replication via directly targeting the Pelota gene in MDBK cells. We investigated whether the potential target sequences of miR-2411, located in the Pelota 3′UTR, and miR-2411 agomir transfection attenuated Pelota mRNA and protein levels. Indeed, upon miR-2411 overexpression, BVDV NADL replication was prevented. Importantly, BVDV NADL replication levels were reversed to normal levels as a result of the Pelota rescuing experiment even though miR-2411 was existent. Overall, we profiled the unique role of miR-2411 in regulating BVDV NADL replication and provided a novel strategy for generalized inhibition of viral infection. [ABSTRACT FROM AUTHOR]

It is quite intriguing that bovines were largely unaffected by influenza A, even though most of the domesticated and wild animals/birds at the human−animal interface succumbed to infection over the past few decades. Influenza A occurs on a very infrequent basis in bovine species and hence bovines were not considered to be susceptible hosts for influenza until the emergence of influenza D. This review describes a multifaceted chronological review of literature on influenza in cattle which comprises mainly of the natural infections/outbreaks, experimental studies, and pathological and seroepidemiological aspects of influenza A that have occurred in the past. The review also sheds light on the bovine models used in vitro and in vivo for influenza-related studies over recent years. Despite a few natural cases in the mid-twentieth century and seroprevalence of human, swine, and avian influenza viruses in bovines, the evolution and host adaptation of influenza A virus (IAV) in this species suffered a serious hindrance until the novel influenza D virus (IDV) emerged recently in cattle across the world. Supposedly, certain bovine host factors, particularly some serum components and secretory proteins, were reported to have anti-influenza properties, which could be an attributing factor for the resilient nature of bovines to IAV. Further studies are needed to identify the host-specific factors contributing to the differential pathogenetic mechanisms and disease progression of IAV in bovines compared to other susceptible mammalian hosts.

Bovine parainfluenza virus type 3 (BPIV3) is one of the most important known viral respiratory pathogens of both young and adult cattle. It is also named “heat stress in transport”, causing morbidity and mass death. New variants of BPIV3 have been detected or isolated in China since 2008. Here, we isolate one BPIV3 strain (named BPIV3 BJ) in Madin-Darby bovine kidney (MDBK) cells from nasal samples collected in China. Phylogenetic analysis showed that our isolate is related to BPIV3 of the genotype A. The comparison of BPIV3-BJ and the reference Chinese isolate NM09 showed that these strains are highly divergent. We found many differences in the amino acid composition in the nucleocapsid (NP) protein among these genotype A strains. Since the NP protein has been implicated in immunization studies, our BPIV3 isolate will be useful for the development of immune assays and vaccine studies. The diversity of BPIV3 lineages that we found in China indicated ongoing evolution for immune escape. Our study highlights the importance of genetic surveillance for determining the effect of BPIV3 variability on pathogen evolution and population-scale immunity.

Caprine herpesvirus 1 (CpHV-1) infection in goats induces genital vesicular-ulcerative lesions that strictly resemble the lesions induced by herpesvirus 2 in the human host. The immunosuppressive drug Mizoribine (MIZ) was found to increase the antiviral activity of Acyclovir (ACV) against herpesvirus infections, raising interesting perspectives on new combined therapeutic strategies. In this study the anti-CpHV-1 activity in vitro of ACV alone or in combination with MIZ was characterized. When applied alone at non-toxic concentrations, ACV had a slight effect on CpHV-1 replication while in combination with MIZ a dose-dependent inhibition of the virus yield was observed with an IC50 of ACV of 28.5 μM. These findings suggest that combined therapy of ACV and MIZ is potentially exploitable in the treatment of genital infection by herpesviruses. Caprine herpesvirus 1 (CpHV-1) infection in goats induces genital vesicular-ulcerative lesions that strictly resemble the lesions induced by herpesvirus 2 in the human host. The immunosuppressive drug Mizoribine (MIZ) was found to increase the antiviral activity of Acyclovir (ACV) against herpesvirus infections, raising interesting perspectives on new combined therapeutic strategies. In this study the anti-CpHV-1 activity in vitro of ACV alone or in combination with MIZ was characterized. When applied alone at non-toxic concentrations, ACV had a slight effect on CpHV-1 replication while in combination with MIZ a dose-dependent inhibition of the virus yield was observed with an IC50 of ACV of 28.5 μM. These findings suggest that combined therapy of ACV and MIZ is potentially exploitable in the treatment of genital infection by herpesviruses. [ABSTRACT FROM AUTHOR]

A eficiência de três monocamadas celulares (linhagem contínua - células Madin and Darby Bovine Kidney/ MDBK;linhagem primária - células de útero e de oviduto bovino) foi testada para verificar a existência de especificidade celular através do desenvolvimento de embriões, desde o estádio de duas células até o de mórula compacta, em um sistema de cocultura sem fluxo externo de CO2. Depois da seleçâo, os embriões (n = 343) foram aleatoriamente distribuídos em diferentes tubos de ensaio, os quais foram colocados a 37°C durante 72 horas. Após o período de cocultura, as porcentagens de mórulas compactas obtidas foram de 87,7% em células de oviduto, 86,2% na monocamada de células uterinas e 88,3% na de células MDBK. Não foi observada diferen- ça significativa entre esses valores e, por isso mesmo, conclui-se que a especificidade celular não é importante para o desenvolvimento in vitro de embriões Mus musculus.The efficiency of three monolayers (continuous lineage - Madin and Darby Bovine Kidney/ MDBK; primary lineage - bovine celisfrom uterus and oviduct) hás been tested to verify the celular specificity through development of two cells embryos until compact morulaes stage in a coculture system without continuous CO2 flow. The selected mouse embryos (n=343) were randomiy divided into three experimental groups andplaced in closed culture tubes mantained aí 37°C during 72 hours. After the co-culture period, the porcentages of compact morulaes were 87.7% in oviduct cells, 86.2% in uterus monolayer and 88.3% in MDBK cells. It was not observed significative difference between these results and it is possible to condude that celular specificity is not importam to enhance the In vitro development of Mus musculus embryos.

The article discusses a study which examined caprine herpes virus-1 (CpHV-1) infection in cell cultures and the cytopathological changes induced by CpHV-1 in Madin-Darby bovine kidney (MDBK) cells based on double staining with two vital fluorochromes. The clinical symptoms of CpHV-1 infection depend on the age of the animal, but it affects the respiratory and genital tracts. The study revealed that a direct immunofluorescence (IF) test will be appropriate to detect CpHV-1 infection within infected MDBK cells.

Background: Bovine herpesvirus type 1 (BHV-1) is an important pathogen in cattle that is responsible for substantial economic losses. Previous studies suggest that BHV-1 may induce apoptosis in Madin-Darby bovine kidney (MDBK) cells via a mechanism only involving caspases and p53. However, the mechanism for BHV-1-induced MDBK cell apoptosis still requires more research. Methods: MDBK was used as a model to study the precise signaling pathways of apoptosis induced by BHV-1 infection. Results: BHV-1 infection activated a Fas/FasL-mediated apoptotic pathway, resulting in activation of caspase-8 and cleavage of Bid. In addition, BHV-1 infection down-regulated Bcl-2 and up-regulated Bax expression, thereby initiating the release of cytochrome c followed by caspase-9 activation. The combined activation of the extrinsic and intrinsic pathways resulted in activation of downstream effecter caspase-3 and poly ADP-ribose polymerase (PARP), leading to apoptosis. Furthermore, blocking apoptosis using caspase inhibitors improved BHV-1-infected MDBK cell viability to different extent. BHV-1 infection did not induce significant DNA fragmentation in MDBK cells pretreated with ammonium chloride (NH4Cl) or cells infected with UV-inactivated BHV-1. Blocking caspases activation increased BHV-1 replication. Conclusions: BHV-1 induces apoptosis in MDBK cells through extrinsic and intrinsic pathways and there might be cross-talk between the two pathways. In addition, BHV-1 replication may be necessary for the induction of apoptosis in BHV-1-infected cells, and prolonged cell viability benefits BHV-1 replication. [ABSTRACT FROM AUTHOR]

The aim of this paper was to examine the pathogenicity of most commonly isolated bacteria from the digestive tract of bees. Bees from 150 colonies (n=3000) were examined and 19 bacterial species were isolated, which are either permanent or temporary inhabitants of the digestive tract. Pathogenic activity of the most commonly isolated species (Enterobacter aerogenes, Klebsiella ozaenae, Klebsiella pneumonie, Citrobacter freundii, Enterobacter cloacae and Enterobacter agglomerans) was examined on seven-day-old chicken embryos and tissue of MDBK cells. Bacterial inoculation of the examined bacteria was conducted in the alantoic cavity of chicken embryos in the quantity of 0.5 mL. Control noninoculated and inoculated embryos were incubated at 38°C with about 60% relative humidity. All six bacterial species manifested pathogenic activity on chicken embryos and caused their death within 2-4 days and changes such as lagging in embryo development, bleeding and unpleasant smell. The bacteria examined in MDBK cell lines of bovine kidney tissue did not manifest cytopathogenic effect and the structure of control tissue was normal. [ABSTRACT FROM AUTHOR]

Madin Darby bovine kidney cells were stored at −80°C using trehalose. Trehalose was loaded into the cells by fluid-phase endocytosis that was facilitated by heat shock at 40°C for 1 h. Loaded cells were gradually frozen and stored at −80°C. Revival of cells was done by quick thawing and immediately seeded in the tissue culture flasks. The membrane integrity of cells was measured at different times post-storage by trypan blue dye exclusion method. It was estimated to be 96.23, 73.84, 57.33, 54.36, 25.47, 50.53 and 46.86% at 0, 7, 60, 90, 120, 160 and 180-day post-storage, respectively. Cryostorage of cells at −80°C may help to reduce the use of liquid nitrogen. [ABSTRACT FROM AUTHOR]

Abstract: Viral antigens can be obtained from infected mammalian cells cultivated on microcarriers. We have worked out parameters for the production of bovine parainfluenza 3 (PI-3) virus by Mandin–Darby Bovine Kidney (MDBK) cells cultivated on Cytodex 1 microcarriers (MCs) in spinners flasks and bioreactor using fetal bovine serum (FBS) supplemented Eagle minimal essential medium (Eagle-MEM). Medium renewal during the cell culture was shown to be crucial for optimal MCs loading (>90% MCs with confluent cell monolayers) and cell growth (2.5×106 cells/mL and a μ x (h−1) 0.05). Since cell cultures performed with lower amount of MCs (1g/L), showed good performances in terms of cell loading, we designed batch experiments with a lower concentration of MCs in view of optimizing the cell growth and virus production. Studies of cell growth with lower concentrations of MCs (0.85g/L) showed that an increase in the initial cell seeding (from 7 to 40 cells/MC) led to a different kinetic of initial cell growth but to comparable final cell concentrations ((8–10)×105 cells/mL at 120h) and cell loading (210–270 cells/MC). Upon infection with PI-3 virus, cultures showed a decrease in cell growth and MC loading directly related to the multiplicity of infection (moi) used for virus infection. Infected cultures showed also a higher consumption of glucose and production of lactate. The PI-3 virus and PI-3 antigen production among the cultures was not significantly different and attained values ranging from, respectively, 7–9log10 TCID50/mL and 1.5–2.2 OD. The kinetics of PI-3 virus production showed a sharp increase during the first 24h and those of PI-3 antigen increased after 24h. The differential kinetics of PI-3 virus and PI-3 antigen can be explained by the virus sensitivity to temperature. In view of establishing a protocol of virus production and based on the previous experiments, MDBK cell cultures performed under medium perfusion in a bioreactor of 1.2L were infected and the PI-3 virus production in 12L attained 12log10 TCID50. Other than establishing a protocol for PI-3 production in MDBK cell cultures on Cytodex 1, the experiments are proposed as a basis for approaching the development of a virus production protocol in mammalian cells cultivated on microcarriers in bioreactors. [Copyright &y& Elsevier]

Bovine viral diarrhea virus (BVDV) infection in cattle is responsible for mucosal disease; an invariably fatal syndrome characterized by the recovery of two BVDV strains: cytopathic (cp) or noncytopathic (ncp). To understand the cellular responses to cp BVDV infection, we carried out differential display-polymerase chain reaction (DD-PCR) analysis of gene expression in infected cells. Altered expression of 14 genes involved in several functions was observed in cells infected with cp BVDV: (1) immune regulation, such as CD46, FKBP-12, and osteopontin (OPN); (2) apoptosis-related cysteine proteases like calpain; (3) signaling plasma membrane proteins such as integrin β1, and prion protein; and (4) unknown function genes. Northern blot analysis of the expression of these genes in ncp BVDV infected cells revealed that while the expression of some genes was affected as in cp BVDV infected cells, others show a clearly contrary change. We postulate that a cause-effect relationship may exist between the differential gene expression alterations that characterize cp and ncp BVDV infections and the unique diseases associated with each BVDV biotype. [ABSTRACT FROM AUTHOR]

In recent years, hunniviruses have been reported in a variety of animal species from many countries. Here, hunnivirus was detected in fecal samples from water buffaloes and named as BufHuV-GX-2106. The samples were inoculated into cultures of MDBK cells supplemented with TPCK trypsin and the BufHuV-GX-2106 strain was stably passaged and replicated. Electron microscopic analysis showed the BufHuV-GX-2106 virus particles were spherical and 20~30 nm in diameter. The complete genome of a plaque purified sample of BufHuV-GX-2106 was determined and analyzed. Genomic analysis revealed that the whole sequence of BufHuV-GX-2106 was ~7,601 nucleotides (nt) in length and consisted of a large open reading frame of 6,759nt, a 5'UTR, a 3'UTR and a poly(A) tail. The complete genome sequence of BufHuV-GX-2106 shares 68-85% nucleotide identities with other known hunnivirus strains, indicating high genetic heterogeneity among these viruses. Phylogenetic analysis showed that BufHuV-GX-2106 belonged to the Hunnivirus A species and was more closely related to ovine hunnivirus than other known viruses of this type. This study describes the first isolation and complete genome sequence of a hunnivirus strain from water buffaloes. In addition, this study will help to understand the mechanisms involved in the pathogenesis of Hunnivirus A among different animal species., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Liu, Zhu, Dong, Qiao, Luo, Liu, Zou, Liu, Wu, Su, Peng, Ouyang, Chen, Li, Wei and Huang.)

Abstract Background Bovine herpesvirus type 1 (BHV-1) is an important pathogen in cattle that is responsible for substantial economic losses. Previous studies suggest that BHV-1 may induce apoptosis in Madin-Darby bovine kidney (MDBK) cells via a mechanism only involving caspases and p53. However, the mechanism for BHV-1-induced MDBK cell apoptosis still requires more research. Methods MDBK was used as a model to study the precise signaling pathways of apoptosis induced by BHV-1 infection. Results BHV-1 infection activated a Fas/FasL-mediated apoptotic pathway, resulting in activation of caspase-8 and cleavage of Bid. In addition, BHV-1 infection down-regulated Bcl-2 and up-regulated Bax expression, thereby initiating the release of cytochrome c followed by caspase-9 activation. The combined activation of the extrinsic and intrinsic pathways resulted in activation of downstream effecter caspase-3 and poly ADP-ribose polymerase (PARP), leading to apoptosis. Furthermore, blocking apoptosis using caspase inhibitors improved BHV-1-infected MDBK cell viability to different extent. BHV-1 infection did not induce significant DNA fragmentation in MDBK cells pretreated with ammonium chloride (NH4Cl) or cells infected with UV-inactivated BHV-1. Blocking caspases activation increased BHV-1 replication. Conclusions BHV-1 induces apoptosis in MDBK cells through extrinsic and intrinsic pathways and there might be cross-talk between the two pathways. In addition, BHV-1 replication may be necessary for the induction of apoptosis in BHV-1-infected cells, and prolonged cell viability benefits BHV-1 replication.

• A total of 261 DEGs were identified in CPIV3-infected MDBK cells at 24 hpi. • These DEGs mainly involved in immune system process, metabolic process, and signal transduction. • The only 7 up-regulated ISGs were integral genes during CPIV3 infection. • The 7 up-regulated ISGs inhibits CPIV3 replication in MDBK cells. Caprine parainfluenza virus type 3 (CPIV3) is the one of most common causative agents of caprine respiratory infection, resulting in significant economic losses in the goat and sheep industries. However, the molecular mechanisms and host genes involved in the pathogenesis of and immunity against CPIV3 infection remain poorly understood. In this study, we used RNA-Seq to understand the responses of madin-darby bovine kidney (MDBK) cells to CPIV3 infection. A total of 261 differentially-expressed genes (DEGs) were identified in CPIV3-infected compared with mock-infected MDBK cells at 24 h post-infection (hpi). The DEGs were mainly involved in immune system processes, metabolic processes, and signal transduction. Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis demonstrated that the most significantly enriched signaling pathways were MAPK, Wnt, PI3K-Akt, tumor necrosis factor, Toll-like receptor and ubiquitin-mediated proteolysis. STRING analysis revealed that seven interferon-stimulated genes (ISGs) were upregulated (IFI6, ISG15, OAS1Y, OAS1Z, MX1, MX2 and RSAD2) and may play a pivotal role during CPIV3 infection. Moreover, overexpression of these ISGs significantly reduced CPIV3 replication in vitro , while siRNA silencing markedly improved CPIV3 replication 24 and 48 hpi. Ours is the first study to profile the gene expression of CPIV3-infected MDBK cells. We identified seven ISGs that could be targeted in novel antiviral strategies against CPIV3. [ABSTRACT FROM AUTHOR]

• MDBK cells contaminated with bovine viral diarrhea virus (BVDV). • Novel method of removing BVDV from LSDV vaccines. • Growth of LSDV on chorioallantoic membranes of fertilized hens' eggs. Bovine viral diarrhea virus (BVDV) is a common contaminant of Madin-Darby bovine kidney (MDBK) cells as well as fetal calf serum (FCS). It is pathogenic to cattle and regulatory authorities require that veterinary vaccine stocks are free from BVDV. MDBK cells are used in the generation of recombinant lumpy skin disease virus (LSDV) and have been used for the growth of LSDV vaccines. This paper describes how vaccine stocks can be cleared of BVDV by passage through an avian host, nonpermissive to BVDV, but permissive to LSDV. LSDV vaccine stocks were shown to be cleared of BVDV after passage on the chorioallantoic membranes (CAMs) of fertilized 7-day old hens' eggs. Vaccines were passaged a second time on CAMs before being grown in primary lamb testes (LT) cells. Vaccines retained BVDV-negative status after passage on LT cells. [ABSTRACT FROM AUTHOR]

It is quite intriguing that bovines were largely unaffected by influenza A, even though most of the domesticated and wild animals/birds at the human–animal interface succumbed to infection over the past few decades. Influenza A occurs on a very infrequent basis in bovine species and hence bovines were not considered to be susceptible hosts for influenza until the emergence of influenza D. This review describes a multifaceted chronological review of literature on influenza in cattle which comprises mainly of the natural infections/outbreaks, experimental studies, and pathological and seroepidemiological aspects of influenza A that have occurred in the past. The review also sheds light on the bovine models used in vitro and in vivo for influenza-related studies over recent years. Despite a few natural cases in the mid-twentieth century and seroprevalence of human, swine, and avian influenza viruses in bovines, the evolution and host adaptation of influenza A virus (IAV) in this species suffered a serious hindrance until the novel influenza D virus (IDV) emerged recently in cattle across the world. Supposedly, certain bovine host factors, particularly some serum components and secretory proteins, were reported to have anti-influenza properties, which could be an attributing factor for the resilient nature of bovines to IAV. Further studies are needed to identify the host-specific factors contributing to the differential pathogenetic mechanisms and disease progression of IAV in bovines compared to other susceptible mammalian hosts. [ABSTRACT FROM AUTHOR]

Bovine parainfluenza virus type 3 (BPIV3) is one of the most important known viral respiratory pathogens of both young and adult cattle. It is also named "heat stress in transport", causing morbidity and mass death. New variants of BPIV3 have been detected or isolated in China since 2008. Here, we isolate one BPIV3 strain (named BPIV3 BJ) in Madin-Darby bovine kidney (MDBK) cells from nasal samples collected in China. Phylogenetic analysis showed that our isolate is related to BPIV3 of the genotype A. The comparison of BPIV3-BJ and the reference Chinese isolate NM09 showed that these strains are highly divergent. We found many differences in the amino acid composition in the nucleocapsid (NP) protein among these genotype A strains. Since the NP protein has been implicated in immunization studies, our BPIV3 isolate will be useful for the development of immune assays and vaccine studies. The diversity of BPIV3 lineages that we found in China indicated ongoing evolution for immune escape. Our study highlights the importance of genetic surveillance for determining the effect of BPIV3 variability on pathogen evolution and population-scale immunity. [ABSTRACT FROM AUTHOR]

Background: Crepis lacera is a plant from the Asteraceae family that is common in the Mediterranean region. Farmers believe that this plant may be deadly to small ruminants in areas of southern Italy. However, scientific evidence is lacking, and no proof exists that C. lacera is toxic to ruminants. Necropsies conducted on four sheep revealed lesions in their livers and kidneys. Results: In the current study, we described sheep poisoning and isolated secondary metabolites from Crepis lacera to assess the metabolites’ biological activity both in vitro and in vivo. Phytochemical study of the aerial portions of Crepis lacera led to the isolation of five sesquiterpene lactones and two phenolic compounds. Cellular viability was evaluated in cell cultures of the bovine kidney cell line Madin Darby Bovine Kidney (MDBK) after incubation with phytochemicals. Our results showed that three sesquiterpene lactones, 8-epidesacylcynaropicrin-3-O-β-glucopyranoside (2), 8-epigrosheimin (3), and 8-β-hydroxydehydrozaluzanin C (4), were cytotoxic after 48 h of incubation. In addition, in the in vivo study, animals that received 1 mg/kg body weight (bw) of Crepis lacera extract and were then sacrificed after 48 h showed significant lesions in their liver, lungs and kidneys. These lesions were also found in rats that received 2 mg/kg bw of the same extract and sacrificed after 24 and 48 h. Conclusions: These results validate the hypothesis that C. lacera is potentially dangerous when ingested in large quantities by grazing small domestic ruminants. Further studies are necessary to clarify the molecular mechanisms of Crepis spp. toxicity in animals. [ABSTRACT FROM AUTHOR]

Bovine primary cultured hepatocytes (CHs) are widely used in vitro models for liver toxicity testing. However, little is known about their whole-transcriptome profile and its resemblance to the normal liver tissue. In the present study, we profiled - by microarray - the whole-transcriptome of bovine CHs (n=4) and compared it with the transcriptomic landscape of control liver samples (n=8), as well the Madin-Darby bovine kidney (MDBK) cells (n=4). Compared with liver tissue, the bovine CHs relatively expressed (fold change >2, P<0.05) about 2155 and 2073 transcripts at a lower and higher abundance, respectively. Of those expressed at a lower abundance, many were drug biotransformation enzyme-coding genes, such as the cytochrome P450 family (CYPs), sulfotransferases, methyltransferases, and glutathione S-transferases. Also, several drug transporters and solute carriers were expressed at a lower abundance in bovine CHs. 'Drug metabolism', 'PPAR signaling', and 'metabolism of xenobiotics by CYPs' were among the most negatively-enriched pathways in bovine CHs compared with liver. A qPCR cross-validation using 8 selected genes evidenced a high correlation (r=0.95, P=0.001) with the corresponding microarray results. Although from a kidney origin, and albeit to a lower extent compared to bovine CHs, the MDBK cells showed a basal expression of many CYP-coding genes. Our study provides a whole-transcriptome-based evidence for the bovine CHs and hepatic tissue resemblance. Overall, the bovine CHs' transcriptomic profile might render it unreliable as an in vitro model to study drug metabolism., (Copyright © 2017 Elsevier Ltd. All rights reserved.)

Neospora caninum, an apicomplexan parasite, is recognized as a major bovine abortifacient. Dense granule antigens (GRAs) play important roles in the formation and modification of parasitophorous vacuoles (PVs) in Toxoplasma gondii. However, a few studies investigating GRAs have been reported in N. caninum. The aim of the present study was to characterize the dense GRA6/GRA7 of N. caninum in PVs using MDBK cells as a host cell model. Neospora caninum was inoculated into MDBK cells, and changes were observed using a transmission electron microscope (TEM). Neospora caninum GRA6/GRA7 were identified and characterized using bioinformatics, cell fractionation, and immunofluorescence. The TEM results revealed that integrated PVs were present in MDBK cells after N. caninum infection. Bioinformatics analysis showed that NcGRA6/NcGRA7 shared 28.76% and 29.66% homology with T. gondii GRA6/GRA7 (TgGRA6/TgGRA7) but had similar signal peptides, transmembrane domains, and motifs. Cell fractionation and subcellular localization analyses both showed that NcGRA6 was distributed in the lumen and intravacuolar network in soluble and transmembrane forms. The transmembrane form of NcGRA7 was observed in the PV membrane. These data lay a foundation for further study on bovine neosporosis and NcGRA6/NcGRA7 function during PV formation., (© The Author 2017. Published by Oxford University Press on behalf of the Institute of Biochemistry and Cell Biology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)