Nutrionally Induced Anovulation in the Bovine: Ovarian and Endocrine Events

Animal Breeding and Reproductio

Integration of Microfluidics, Photonic Integrated Circuits and Data Acquisition and Analysis Methods in a Single Platform for the Detection of Swine Viral Diseases

[EN] Simple Summary: The control of several swine viral diseases relies mainly on evidence-based prevention protocols due to the lack of effective treatments or vaccines. To design these protocols, laboratory investigation of viral infections is critical to confirm their occurrence and determine their epizootiology. However, laboratory confirmation of certain swine viral diseases is a time-consuming and labor-intensive process, requiring scientific personnel with relevant expertise. Point-of-Care (POC) diagnostics are tests and devices that provide clinically relevant information on-site, facilitating decision-makers to swiftly take countermeasures for disease control. In the present study, novel photonic biosensors were integrated into a single, automated POC device that can record and analyze changes in the sensors' refractive index, allowing the detection of Porcine Parvovirus (PPV) and Porcine Circovirus 2 (PCV-2) in oral fluids within 75 min. The objective of this work was to validate this device using reference and field samples (oral fluids). The system was able to detect PPV and PCV-2 in oral fluid samples satisfactorily. The device can be directly deployed in farms for the fast diagnosis of these diseases, contributing to farm biosecurity.Viral diseases challenge the health and welfare of pigs and undermine the sustainability of swine farms. Their efficient control requires early and reliable diagnosis, highlighting the importance of Point of Care (POC) diagnostics in veterinary practice. The objective of this study was to validate a novel POC system that utilizes Photonic Integrated Circuits (PICs) and microfluidics to detect swine viral pathogens using oral fluids and Porcine Parvovirus (PPV) and Porcine Circovirus 2 (PCV-2) as proofs of concept. The sensitivity and specificity of the device were calculated for both viruses, and Receiver Operating Characteristic (ROC) curves were drawn. PPV had an Area Under Curve (AUC) value of 0.820 (95% CI: 0.760 to 0.880, p < 0.0001), and its optimal efficiency threshold of detection shifts was equal to 4.5 pm (68.6% sensitivity, 77.1% specificity and Limit of Detection (LOD) value 10(6) viral copies/mL). PCV-2 had an AUC value of 0.742 (95% CI: 0.670 to 0.815, p < 0.0001) and an optimal efficiency threshold of shifts equal to 6.5 pm (69.5% sensitivity, 70.3% specificity and LOD 3.3 x 10(5) copies/mL). In this work, it was proven that PICs can be exploited for the detection of swine viral diseases. The novel device can be directly deployed on farms as a POC diagnostics tool.This research was funded by E.U.'s H2020 SWINOSTICS project under the grant agreement ID 771649.Manessis, G.; Mourouzis, C.; Griol Barres, A.; Zurita-Herranz, D.; Peransi, S.; Sánchez, C.; Giusti, A.... (2021). Integration of Microfluidics, Photonic Integrated Circuits and Data Acquisition and Analysis Methods in a Single Platform for the Detection of Swine Viral Diseases. Animals. 11(11):1-18. https://doi.org/10.3390/ani11113193118111

Point-of-Care and Label-Free Detection of Porcine Reproductive and Respiratory Syndrome and Swine Influenza Viruses Using a Microfluidic Device with Photonic Integrated Circuits

[EN] Swine viral diseases challenge the sector's sustainability by affecting productivity and the health and welfare of the animals. The lack of antiviral drugs and/or effective vaccines renders early and reliable diagnosis the basis of viral disease management, underlining the importance of point-of-care (POC) diagnostics. A novel POC diagnostic device utilizing photonic integrated circuits (PICs), microfluidics, and information and communication technologies for the detection of porcine reproductive and respiratory syndrome virus (PRRSV) and swine influenza A (SIV) was validated using spiked and clinical oral fluid samples. Metrics including sensitivity, specificity, accuracy, precision, positive likelihood ratio (PLR), negative likelihood ratio (NLR), and diagnostic odds ratio (DOR) were calculated to assess the performance of the device. For PRRSV, the device achieved a sensitivity of 83.5%, specificity of 77.8%, and DOR values of 17.66, whereas the values for SIV were 81.8%, 82.2%, and 20.81, respectively. The POC device and PICs can be used for the detection of PRRSV and SIV in the field, paving the way for the introduction of novel technologies in the field of animal POC diagnostics to further optimize livestock biosecurity.This research was funded by the EU's H2020 SWINOSTICS project under the grant agreement ID 771649.Manessis, G.; Frant, M.; Wozniakowski, G.; Nannucci, L.; Bennedetti, M.; Denes, L.; Balka, G.... (2022). Point-of-Care and Label-Free Detection of Porcine Reproductive and Respiratory Syndrome and Swine Influenza Viruses Using a Microfluidic Device with Photonic Integrated Circuits. Viruses. 14(5):1-21. https://doi.org/10.3390/v1405098812114

Viral FLICE Inhibitory Protein of Rhesus Monkey Rhadinovirus Inhibits Apoptosis by Enhancing Autophagosome Formation

Rhesus monkey rhadinovirus (RRV) is a gamma-2 herpesvirus closely related to human herpesvirus 8 (HHV8). RRV encodes viral FLICE inhibitory protein (vFLIP), which has death effector domains. Little is known about RRV vFLIP. This study intended to examine its function in apoptosis. Here we found that RRV vFLIP inhibits apoptosis induced by tumor necrosis factor-α (TNF-α) and cycloheximide. In HeLa cells with vFLIP expression, the cleavage of poly [ADP-ribose] polymerase 1 (PARP-1) and activities of caspase 3, 7, and 9 were much lower than those in controls. Cell viability of HeLa cells with vFLIP expression was significantly higher than control cells after apoptosis induction. However, RRV vFLIP appears unable to induce NF-κB signaling when tested in NF-κB reporter assay. RRV vFLIP was able to enhance cell survival under starved conditions or apoptosis induction. At early time points after apoptosis induction, autophagosome formation was enhanced and LC3-II level was elevated in cells with vFLIP and, when autophagy was blocked with chemical inhibitors, these cells underwent apoptosis. Moreover, RRV latent infection of BJAB B-lymphoblastoid cells protects the cells against apoptosis by enhancing autophagy to maintain cell survival. Knockdown of vFLIP expression in the RRV-infected BJAB cells with siRNA abolished the protection against apoptosis. These results indicate that vFLIP protects cells against apoptosis by enhancing autophagosome formation to extend cell survival. The finding of vFLIP’s inhibition of apoptosis via the autophagy pathway provides insights of vFLIP in RRV pathogenesis

Cloning of an Avian Adeno-Associated Virus (AAAV) and Generation of Recombinant AAAV Particles

Recent studies have proposed that adeno-associated viruses (AAVs) are not evolutionarily linked to other mammalian autonomous parvoviruses but are more closely linked to the autonomous parvoviruses of birds. To better understand the relationship between primate and avian AAVs (AAAVs), we cloned and sequenced the genome of an AAAV (ATCC VR-865) and generated recombinant AAAV particles. The genome of AAAV is 4,694 nucleotides in length and has organization similar to that of other AAVs. The entire genome of AAAV displays 56 to 65% identity at the nucleotide level with the other known AAVs. The AAAV genome has inverted terminal repeats of 142 nucleotides, with the first 122 forming the characteristic T-shaped palindromic structure. The putative Rep-binding element consists of a tandem (GAGY)(4) repeat, and the putative terminal resolution site (trs), CCGGT/CG, contains a single nucleotide substitution relative to the AAV(2) trs. The Rep open reading frame of AAAV displays 50 to 54% identity at the amino acid level with the other AAVs, with most of the diversity clustered at the carboxyl and amino termini. Comparison of the capsid proteins of AAAV and the primate dependoviruses indicate that divergent regions are localized to surface-exposed loops. Despite these sequence differences, we were able to produce recombinant AAAV particles carrying a lacZ reporter gene by cotransfection in 293T cells and were able to examine transduction efficiency in both chicken primary cells and several cell lines. Our findings indicate that AAAV is the most divergent AAV described to date but maintains all the characteristics unique to the genera of dependovirus

Bioengineered Bovine Papillomavirus L1 Protein Virus-like Particle (VLP) Vaccines for Enhanced Induction of CD8 T Cell Responses through Cross-Priming

Safe and effective T cell vaccines are needed for the treatment or prevention of cancers as well as infectious agents where vaccines for neutralizing antibodies have performed poorly. Recent research highlights an important role for tissue-resident memory T cells (TRM cells) in protective immunity and the role of a subset of dendritic cells that are capable of cross-priming for the induction of TRM cells. However, efficient vaccine technologies that operate through cross-priming and induce robust CD8+ T cell responses are lacking. We developed a platform technology by genetically engineering the bovine papillomavirus L1 major capsid protein to insert a polyglutamic acid/cysteine motif in place of wild-type amino acids in the HI loop. Virus-like particles (VLPs) are formed by self-assembly in insect cells infected with a recombinant baculovirus. Polyarginine/cysteine-tagged antigens are linked to the VLP by a reversible disulfide bond. The VLP possesses self-adjuvanting properties due to the immunostimulatory activity of papillomavirus VLPs. Polyionic VLP vaccines induce robust CD8+ T cell responses in peripheral blood and tumor tissues. A prostate cancer polyionic VLP vaccine was more efficacious than other vaccines and immunotherapies for the treatment of prostate cancer in a physiologically relevant murine model and successfully treated more advanced diseases than the less efficacious technologies. The immunogenicity of polyionic VLP vaccines is dependent on particle size, reversible linkage of the antigen to the VLP, and an interferon type 1 and Toll-like receptor (TLR)3/7-dependent mechanism

Point-of-Care Diagnostics for Farm Animal Diseases: From Biosensors to Integrated Lab-on-Chip Devices

Zoonoses and animal diseases threaten human health and livestock biosecurity and productivity. Currently, laboratory confirmation of animal disease outbreaks requires centralized laboratories and trained personnel; it is expensive and time-consuming, and it often does not coincide with the onset or progress of diseases. Point-of-care (POC) diagnostics are rapid, simple, and cost-effective devices and tests, that can be directly applied on field for the detection of animal pathogens. The development of POC diagnostics for use in human medicine has displayed remarkable progress. Nevertheless, animal POC testing has not yet unfolded its full potential. POC devices and tests for animal diseases face many challenges, such as insufficient validation, simplicity, and portability. Emerging technologies and advanced materials are expected to overcome some of these challenges and could popularize animal POC testing. This review aims to: (i) present the main concepts and formats of POC devices and tests, such as lateral flow assays and lab-on-chip devices; (ii) summarize the mode of operation and recent advances in biosensor and POC devices for the detection of farm animal diseases; (iii) present some of the regulatory aspects of POC commercialization in the EU, USA, and Japan; and (iv) summarize the challenges and future perspectives of animal POC testing

Aetiology, Risk Factors, Diagnosis and Control of Foot-Related Lameness in Dairy Sheep

During the last twenty years, considerable research efforts have recognized the consequences of foot-related lameness primarily in cattle, and meat and wool sheep. Despite the lack of extensive epidemiological studies, field observations and isolated research reports in dairy sheep have suggested that the problem might be more severe in semi-intensive and intensive farming systems. Footrot, contagious ovine digital dermatitis, ovine interdigital dermatitis, white line disease, and pedal joint abscess are the most common causes of foot-related lameness. Dichelobacter nodosus, Fusobacterium necrophorum, Treponema spp., and Actinomyces pyogenes are the most significant foot-related lameness-associated pathogens. Despite a documented hereditary predisposition, environmental factors are the most important in determining the occurrence of foot-related lameness. Moist and warm environment, increased parity and milk yield, inappropriate housing conditions and infrastructures, inadequate hygiene status, imbalanced nutrition, and insufficient foot care are the most critical risk factors. Furthermore, a foot-lameness control plan should include targeted implementation of claw trimming and footbathing, evidence-based planning of hygiene measures in preventive veterinary practices (i.e., antibiotic administration, vaccinations against footrot), selective breeding to footrot resistance, and, most importantly, the continuous training of farming personnel. Controlling foot-lameness in dairy sheep is critical in determining the well-being of animals, and strongly affects the farm&rsquo;s profitability and sustainability

Serological, Molecular and Culture-Based Diagnosis of Lentiviral Infections in Small Ruminants

Small ruminant lentiviruses (SRLVs) infections lead to chronic diseases and remarkable economic losses undermining health and welfare of animals and the sustainability of farms. Early and definite diagnosis of SRLVs infections is the cornerstone for any control and eradication efforts; however, a “gold standard” test and/or diagnostic protocols with extensive applicability have yet to be developed. The main challenges preventing the development of a universally accepted diagnostic tool with sufficient sensitivity, specificity, and accuracy to be integrated in SRLVs control programs are the genetic variability of SRLVs associated with mutations, recombination, and cross-species transmission and the peculiarities of small ruminants’ humoral immune response regarding late seroconversion, as well as intermittent and epitope-specific antibody production. The objectives of this review paper were to summarize the available serological and molecular assays for the diagnosis of SRLVs, to highlight their diagnostic performance emphasizing on advantages and drawbacks of their application, and to discuss current and future perspectives, challenges, limitations and impacts regarding the development of reliable and efficient tools for the diagnosis of SRLVs infections