Porcine Circovirus Type Research Articles

Detection of Lawsonia intracellularis by oral fluids and fecal samples in Canadian swine

Objectives: The study objectives were to 1) describe the proportion of Lawsonia intracellularis-positive samples in unvaccinated and vaccinated Canadian swine herds during the mid- and late-finishing phases; 2) compare the probability of detecting L intracellularis by quantitative polymerase chain reaction using fecal samples (FS) and oral fluids (OF); and 3) investigate risk factors of L intracellularis detection using FS and OF. Material and methods: Site demographics and vaccination protocols were obtained from 40 Canadian swine sites via questionnaire. Three OF and 3 FS were collected per site once during the mid-finisher (15-17 wk of age) and once during the late-finisher (20-22 wk of age) production stages. Results: Half of all investigated production sites were positive for L intracellularis. A 2-fold increase in L intracellularis detection rate was observed for OF compared to FS (odds ratio = 2.36; 95% CI, 1.24-4.49; P = .009). The presence of porcine circovirus type 2 (PCV2) had a 5-fold increased risk of L intracellularis positivity compared to sites without PCV2 (incidence rate ratio [IRR] = 4.99; 95% CI, 1.29-20.23; P = .02). A higher positive rate was found for sites with L intracellularis outbreaks within the last 2 years (IRR = 3.08; 95% CI, 1.51-6.37; P = .002). Implications: This study presents evidence that OF may have a higher detection rate compared to FS for L intracellularis. Herds with PCV2 or exposure to recent L intracellularis outbreaks may be at increased risk of harboring L intracellularis and warrant additional investigation.

MALDI-TOF nucleic acid mass spectrometry for simultaneously detection of fourteen porcine viruses and its application

BackgroundMixed infections of multiple viruses significantly contribute to the prevalence of swine diseases, adversely affecting global livestock production and the economy. However, effectively monitoring multiple viruses and detecting mixed infection samples remains challenging. This study describes a method that combines single-base extension PCR with matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) to detect important porcine viruses. ResultsOur approach accurately and simultaneously identified 14 porcine viruses, including porcine circovirus types 1–3, porcine bocaviruses groups 1–3, African swine fever virus, pseudorabies virus, porcine parvovirus, torque teno sus virus, swine influenza virus, porcine reproductive and respiratory syndrome virus, classical swine fever virus, and foot-and-mouth disease virus. The low limit of detection for multiplex identification ranges from 13.54 to 1.59 copies/μL. Inter- and intra-assay stability was found to be ≥98.3 %. In a comprehensive analysis of 114 samples, the assay exhibited overall agreement with qPCR results of 97.9 %. ConclusionsThe developed MALDI-TOF NAMS assay exhibits high sensitivity, specificity, and reliability in detecting and distinguishing a wide spectrum of porcine viruses in complex matrix samples. This underscores its potential as an efficient diagnostic tool for porcine-derived virus surveillance and swine disease control.

Importance of sequential infection order of porcine circovirus type 2 and porcine reproductive and respiratory syndrome virus to divergent clinical outcomes

This study was designed to investigate the different sequential order of infection for porcine circovirus type 2 (PCV2) and porcine reproductive and respiratory syndrome virus (PRRSV). Thirty-six pigs were randomly assigned to six different treatment groups. The first (hereafter referred to as PRRSV-PCV2) group was inoculated with PRRSV first followed by PCV2d. The second (hereafter referred to as PCV2+PRRSV) group was co-infected with both viruses at the same timepoint (42 days of age). The third (hereafter referred to as PCV2-PRRSV) group was inoculated with PCV2d first followed by PRRSV. A fourth group was only inoculated with PCV2d at 42 days of age, while a fifth group was only inoculated with PRRSV at the same timepoint. The sixth group served as a negative control group. The most important observation discovered that PRRSV only had a potentiation effect on PCV2 in both PRRS-PCV2 and PCV2+PRRSV groups. Both PRRSV-PCV2 and PCV2+PRRSV groups experienced a significant reduction in growth performance compared with control pigs. In addition, PRRSV-PCV2 and PCV2+PRRSV groups exhibited a greater severity in their clinical signs, and/or had higher PCV2 blood and lymphoid viral loads that resulted in a stronger severity of lymphoid lesions compared with PCV2-PRRSV group. Serum TNF-α levels were significantly higher in both PRRS-PCV2 and PCV2+PRRSV groups compared with those in PCV2-PRRS, PCV2, and PRRSV groups. The results of this study demonstrated that divergent clinical outcomes are dependent on the sequential infection order of PCV2 and PRRSV.

Comparative immunogenic and immunoprotective activities of PCV2d Cap and Rep antigens delivered by an efficient eukaryotic expression system engineered into a Salmonella vaccine vector

Porcine circovirus type 2 (PCV2) stands as a predominant etiological agent in porcine circovirus-associated diseases. To manage the spread of the disease, it is necessary to develop a next-generation vaccine expressing PCV2 antigens that target the prevailing genotype such as PCV2d. A bacterial-mediated vaccine delivery by live-attenuated Salmonella has attracted interest for its low-cost production and highly effective vaccine delivery. Thus, in this study, we utilized the advantages of the Salmonella-mediated vaccine delivery by cloning PCV2d cap and rep into a eukaryotic expression plasmid pJHL204 and electroporation into an engineered live-attenuated Salmonella Typhimurium JOL2500 (Δlon, ΔcpxR, ΔsifA, Δasd). The eukaryotic antigen expression by JOL2995 (p204:cap) and JOL2996 (p204:rep) was confirmed in vitro and in vivo which showed efficient antigen delivery. Furthermore, vaccination of mice model with the vaccine candidates elicited humoral and cell-mediated immune responses as depicted by high levels of PCV2-specific antibodies, CD4+ and CD8+ T cells, and neutralizing antibodies, especially by JOL2995 (p204:cap) which correlated with the significant decrease in the viral load in PCV2d-challenged mice. Interestingly, JOL2996 (p204:rep) may not have elicited high levels of neutralizing antibodies and protective efficacy, but it elicited considerably higher cell-mediated immune responses. This study demonstrated Salmonella-mediated vaccine delivery system coupled with the eukaryotic expression vector can efficiently deliver and express the target PCV2d antigens for strong induction of immune response and protective efficacy in mice model, further supporting the potential application of the Salmonella-mediated vaccine delivery system as an effective novel approach in vaccine strategies for PCV2d.

The PCV3 Cap Virus-like Particle Vaccine with the Chimeric PCV2-Neutralizing Epitope Gene Is Effective in Mice.

Porcine circovirus type 3 (PCV3) infection can cause symptoms similar to those of porcine circovirus type 2 (PCV2) infection, and coinfections with both PCV2 and PCV3 are observed in the swine industry. Consequently, developing chimeric vaccines is essential to prevent and control porcine circovirus infections. In this study, we used both E. coli and mammalian expression systems to express PCV3 Cap (Cap3) and a chimeric gene containing the PCV2-neutralizing epitope within the PCV3 Cap (Cap3-Cap2E), which were assembled into virus-like particle (VLP) vaccines. We found that Cap3 lacking nuclear localization signal (NLS) could not form VLPs, while Cap3 with a His-tag successfully assembled into VLPs. Additionally, the chimeric of PCV2-neutralizing epitopes did not interfere with the assembly process of VLPs. Various immunization approaches revealed that pCap3-Cap2E VLP vaccines were capable of activating high PCV3 Cap-specific antibody levels and effectively neutralizing both PCV3 and PCV2. Furthermore, pCap3-Cap2E VLPs demonstrated a potent ability to activate cellular immunity, protecting against PCV3 infection and preventing lung damage in mice. In conclusion, this study successfully developed a PCV3 Cap VLP vaccine incorporating chimeric PCV2-neutralizing epitope genes, providing new perspectives for PCV3 vaccine development.

Cytotoxicity effect and transcriptome analysis of PCV3-infected cells revealed potential viral pathogenic mechanisms

Porcine circovirus type 3 (PCV3) has become an important pathogen in the global swine industry and poses a threat to pig health, but its pathogenic mechanism remains unknown. In this study, we constructed an innovative, linear infectious clone of PCV3 for rescuing the virus, and explored the transcriptome of infected cells to gain insights into its pathogenic mechanisms. Subsequently, an in vivo experiment was conducted to evaluate the pathogenicity of the rescued virus in pig. PCV3 nucleic acid was distributed across various organs, indicating systemic circulation via the bloodstream and viremia. Immunohistochemical staining also revealed a significant presence of PCV3 antigens in the spleen, lungs, and lymph nodes, indicating that PCV3 had tropism for these organs. Transcriptome analysis of infected ST cells revealed differential expression of genes associated with apoptosis, immune responses, and cellular metabolism. Notably, upregulation of genes related to the hypoxia-inducible factor-1 pathway, glycolysis, and the AGE/RAGE pathway suggests activation of inflammatory responses, ultimately leading to onset of disease. These findings have expanded our understanding of PCV3 pathogenesis, and the interplay between PCV3 and host factors.

Establishment and preliminary application of duplex fluorescence quantitative PCR for porcine circoviruses type 2 and type 3

Porcine circovirus types 2 (PCV2) and 3 (PCV3) are the two most prevalent porcine circoviruses in China, all of which can infect swine herds and cause serious diseases. To detect coinfection with PCV2 and PCV3, primers and probes for duplex PCV2 and PCV3 real-time PCR were designed to target their cap genes based on the constructed plasmids pUC57-PCV2 and pUC57-PCV3. The established duplex PCV2 and PCV3 real-time PCRs were specific to PCV2 and PCV3 and showed no cross-reactions with other porcine viral pathogens. The limit of detection was 5 and 50 copies for the PCV2 and PCV3 plasmids, respectively. The intra- and interassay repeatability had coefficients of variation below 3 %. The established methods were used to analyze clinical samples from Liaoning and Jilin provinces of China. The coinfection rates of PCV2 and PCV3 in pigs extensively fed in Liaoning and Jilin, large-scale farmed pigs in Liaoning and large-scale farmed pigs in Jilin were 15.0 % (6/40), 36.7 % (11/30) and 35.4 % (62/175), respectively. This study established a useful duplex PCV2 and PCV3 real-time PCR method that can be used for the detection of PCV2 and PCV3 in local clinical samples.

Porcine circovirus type 2 ORF5 induces an inflammatory response by up-regulating miR-21 levels through targeting nuclear ssc-miR-30d

Porcine circovirus type 2 (PCV2) infection leads to multi-system inflammation in pigs, and this effect can be achieved by upregulating host miR-21. The underlying mechanism of miR-21 regulates PCV2-induced inflammation is already known, however, how PCV2 regulates miR-21 levels and function using both autonomic and host factors remains to be further revealed. Here we present the first evidence that PCV2 ORF5 induces an inflammatory response by up-regulating miR-21 level through targeting nuclear miR-30d. In this study, we found that overexpression of ORF5 significantly increased miR-21 level and promoted the expression of inflammatory cytokines and activation of the NF-κB pathway, while ORF5 mutation had the opposite effect. Moreover, the differential expression of miR-21 could significantly change the pro-inflammatory effect of ORF5, indicating that ORF5 promotes inflammatory response by up-regulating miR-21. Bioinformatics analysis and clinical detection found that nuclear miR-30d was significantly down-regulated after ORF5 overexpression and PCV2 infection, and targeted pri-miR-21 and PCV2 ORF5. Functionally, we found that miR-30d inhibited the levels of miR-21 and inflammatory cytokines in cells. Mechanistically, we demonstrated that ORF5 inhibits miR-30d expression levels through direct binding but not via the circRNA pathway, and miR-30d inhibits miR-21 levels by targeting pri-miR-21. In summary, the present study revealed the molecular mechanism of ORF5 upregulation of miR-21, further refined the molecular chain of PCV2-induced inflammatory response and elucidated the role of miRNAs in it.

Development of a novel PCV2 and PCV3 vaccine using virus-like vesicles incorporating Venezuelan equine encephalomyelitis virus-containing vesicular stomatitis virus glycoprotein.

Porcine circovirus disease (PCV) causes substantial economic losses in the pig industry, primarily from porcine circovirus type 2 (PCV2) and porcine circovirus type 3 (PCV3). Novel vaccines are necessary to prevent and control PCV infections. PCV coat proteins are crucial for eliciting immunogenic proteins that induce the production of antibodies and immune responses. A vaccine platform utilizing Semliki Forest virus RNA replicons expressing vesicular stomatitis virus glycoprotein (VSV-G), was recently developed. This platform generates virus-like vesicles (VLVs) containing VSV-G exclusively, excluding other viral structural proteins. In our study, we developed a novel virus-like vesicle vaccine by constructing recombinant virus-like vesicles (rVLVs) that also express EGFP. These rVLVs were created using the RNA replicon of Venezuelan equine encephalomyelitis (VEEV) and New Jersey serotype VSV-G. The rVLVs underwent characterization and safety evaluation in vitro. Subsequently, rVLVs expressing PCV2d-Cap and PCV3-Cap proteins were constructed. Immunization of C57 mice with these rVLVs led to a significant increase in anti-porcine circovirus type 2 and type 3 capsid protein antibodies in mouse serum. Additionally, a cellular immune response was induced, as evidenced by high production of IFN-γ and IL-4 cytokines. Overall, this study demonstrates the feasibility of developing a novel porcine circovirus disease vaccine based on rVLVs.

Development a high-sensitivity sandwich ELISA for determining antigen content of porcine circovirus type 2 vaccines

Porcine circovirus type 2 (PCV2) is intensely prevalent in global pig farms. The PCV2 vaccine is an important means of preventing and controlling PCV2. The quality control of PCV2 vaccines is predominantly based on detection techniques such as animal testing and neutralizing antibody titration. Measuring the content of effective proteins in vaccines to measure vaccine efficacy is an excellent alternative to traditional methods, which can greatly accelerate the development speed and testing time of vaccines. In this study, we screened a monoclonal antibody (mAb) that can effectively recognize not only the exogenous expression of PCV2 Cap protein but also PCV2 virus. The double antibody sandwich ELISA (DAS-ELISA) was developed using this mAb that specifically recognize PCV2 Cap. The minimum protein content detected by this method is 3.5 ng/mL. This method can be used for the quality control of PCV2 inactivated vaccine and subunit vaccine, and the detection results are consistent with the results of mice animal experiments. This method has the advantages of simple operation, good sensitivity, high specificity and wide application. It can detect the effective antigen Cap protein content of various types of PCV2 vaccines, which not only shorten the vaccine inspection time but also save costs.

Design of a New Vaccine Prototype against Porcine Circovirus Type 2 (PCV2), M. hyopneumoniae and M. hyorhinis Based on Multiple Antigens Microencapsulation with Sulfated Chitosan.

This work evaluated in vivo an experimental-multivalent-vaccine (EMV) based on three Porcine Respiratory Complex (PRC)-associated antigens: Porcine Circovirus Type 2 (PCV2), M. hyopneumoniae (Mhyop) and M. hyorhinis (Mhyor), microencapsulated with sulfated chitosan (M- ChS + PRC-antigens), postulating chitosan sulphate (ChS) as a mimetic of the heparan sulfate receptor used by these pathogens for cell invasion. The EMV was evaluated physicochemically by SEM (Scanning-Electron-Microscopy), EDS (Energy-Dispersive-Spectroscopy), Pdi (Polydispersity-Index) and zeta potential. Twenty weaned pigs, distributed in four groups, were evaluated for 12 weeks. The groups 1 through 4 were as follows: 1-EMV intramuscular-route (IM), 2-EMV oral-nasal-route (O/N), 3-Placebo O/N (M-ChS without antigens), 4-Commercial-vaccine PCV2-Mhyop. qPCR was used to evaluate viral/bacterial load from serum, nasal and bronchial swab and from inguinal lymphoid samples. Specific humoral immunity was evaluated by ELISA. M-ChS + PRC-antigens measured between 1.3-10 μm and presented low Pdi and negative zeta potential, probably due to S (4.26%). Importantly, the 1-EMV protected 90% of challenged animals against PCV2 and Mhyop and 100% against Mhyor. A significant increase in antibody was observed for Mhyor (1-EMV and 2-EMV) and Mhyop (2-EMV), compared with 4-Commercial-vaccine. No difference in antibody levels between 1-EMV and 4-Commercial-vaccine for PCV2-Mhyop was observed. Conclusion: The results demonstrated the effectiveness of the first EMV with M-ChS + PRC-antigens in pigs, which were challenged with Mhyor, PCV2 and Mhyop, evidencing high protection for Mhyor, which has no commercial vaccine available.

A multiplex digital PCR assay for detection and quantitation of porcine circovirus type 2 and type 3.

Porcine circovirus (PCV) has become a major pathogen, causing major economic losses in the global pig industry, and PCV type 2 (PCV2) and 3 (PCV3) are distributed worldwide.We designed specific primer and probe sequences targeting PCV2 Cap and PCV3 Rap anddeveloped a multiplex crystal digital PCR (cdPCR) method after optimizing the primerconcentration, probe concentration, and annealing temperature.The multiplex cdPCR assay permits precise and differential detection of PCV2 and PCV3, with a limit of detection of 1.39 × 101 and 1.27 × 101 copies/reaction, respectively, and no cross-reaction with other porcine viruses was observed. The intra-assay and interassay coefficients of variation (CVs) were less than 8.75%, indicating good repeatability and reproducibility. To evaluate the practical value of this assay, 40 tissue samples and 70 feed samples were tested for both PCV2 and PCV3 by cdPCR and quantitative PCR (qPCR). Using multiplex cdPCR, the rates of PCV2 infection, PCV3 infection, and coinfection were 28.45%, 1.72%, and 12.93%, respectively, and using multiplex qPCR, they were 25.00%, 0.86%, and 4.31%, respectively This highly specific and sensitive multiplex cdPCR thus allows accurate simultaneous detection of PCV2 and PCV3, and it is particularly well suited for applications that require the detection of small amounts of input nucleic acid or samples with intensive processing and complex matrices.

Metagenomics to Identify Viral Communities Associated with Porcine Respiratory Disease Complex in Tibetan Pigs in the Tibetan Plateau, China.

Tibetan pig is a unique pig breed native to the Qinghai-Tibet Plateau. To investigate viral communities associated with porcine respiratory disease complex (PRDC), 167 respiratory samples were collected from Tibetan pigs in the Ganzi Tibetan autonomous prefecture of Sichuan province. Following library construction and Illunima Novaseq sequencing, 18 distinct viruses belonging to 15 viral taxonomic families were identified in Tibetan pigs with PRDC. Among the 18 detected viruses, 3 viruses were associated with PRDC, including porcine circovirus type 2 (PCV-2), Torque teno sus virus (TTSuV), and porcine cytomegalovirus (PCMV). The genomic sequences of two PCV-2 strains, three TTSuV strains, and one novel Porprismacovirus strain were assembled by SOAPdenovo software (v2). Sequence alignment and phylogenetic analysis showed that both PCV-2 strains belonged to PCV-2d, three TTSuVs were classified to TTSuV2a and TTSuV2b genotypes, and the Porprismacovirus strain PPMV-SCgz-2022 showed a close genetic relationship with a virus of human origin. Recombination analysis indicated that PPMV-SCgz-2022 may have originated from recombination events between Human 16,806 × 66-213 strain and Porcine 17,668 × 82-593 strain. Furthermore, the high proportion of single infection or co-infection of PCV2/TTSuV2 provides insight into PRDC infection in Tibetan pigs. This is the first report of the viral communities in PRDC-affected Tibetan pigs in this region, and the results provides reference for the prevention and control of respiratory diseases in these animals.

Porcine Circovirus Type 3 (PCV3) in Poland: Prevalence in Wild Boar Population in Connection with African Swine Fever (ASF).

Human health is dependent on food safety and, therefore, on the health of farm animals. One of the most significant threats in regard to swine diseases is African swine fever (ASF). Infections caused by porcine circoviruses (PCVs) represent another important swine disease. Due to the ubiquitous nature of PCV2, it is not surprising that this virus has been detected in ASFV-affected pigs. However, recent data indicate that coinfection of PCV3 and ASFV also occurs. It is still unclear whether PCV infection plays a role in ASFV infection, and that subject requires further analysis. The aim of this study was to assess whether PCV3 and PCV4 are present in the wild boar population in Poland (real-time PCR). The analysis was performed on wild boar samples collected for routine ASF surveillance in Poland, between 2018 and 2021. By extension, the obtained data were compared in regard to ASFV presence in these samples, thus investigating the odds of ASFV infection on the grounds of the PCV carrier state in free-ranging Suidae in Poland. In addition, sequencing of PCV3 and phylogenetic analysis were performed, based on a full genome and a capsid gene. In the current study, we demonstrated the high prevalence of PCV3 in the wild boar population in Poland; meanwhile, PCV4 was not detected. The odds of ASFV infection on the grounds of the PCV3 carrier state in free-ranging Suidae in Poland was more than twice as high. Ten full genome sequences of PCV3 were obtained, all of them belonging to clade 3a. The similarity between them was in the range of 98.78-99.80%.

Establishment of an ELISA Based on a Recombinant Antigenic Protein Containing Multiple Prominent Epitopes for Detection of African Swine Fever Virus Antibodies.

African swine fever virus (ASFV) poses a significant threat to the global pig industry, necessitating accurate and efficient diagnostic methods for its infection. Previous studies have often focused on a limited number of epitopes from a few proteins for detecting antibodies against ASFV. Therefore, the current study aimed to use multiple B-cell epitopes in developing an indirect Enzyme-Linked Immunosorbent Assay (ELISA) for enhanced detection of ASFV antibodies. For the expression of recombinant protein, k3 derived from 27 multiple peptides of 11 ASFV proteins, such as p72, pA104R, pB602L, p12, p14.5, p49, pE248R, p30, p54, pp62, and pp220, was used. To confirm the expression of the recombinant protein, we used the Western blotting analysis. The purified recombinant K3 protein served as the antigen in our study, and we employed the indirect ELISA technique to detect anti-ASFV antibodies. The present finding showed that there was no cross-reactivity with antibodies targeting Foot-and-mouth disease virus (FMDV), Porcine circovirus type 2 (PCV2), Pseudorabies virus (PRV), Porcine reproductive and respiratory syndrome virus (PRRSV), and Classical swine fever virus (CSFV). Moreover, the current finding was sensitive enough to find anti-ASFV in serum samples that had been diluted up to 32 times. The test (k3-iELISA) showed diagnostic specificity and sensitivity of 98.41% and 97.40%, respectively. Moreover, during the present investigation, we compared the Ingenasa kit and the k3-iELISA to test clinical pig serum, and the results revealed that there was 99.00% agreement between the two tests, showing good detection capability of the k3-iELISA method. Hence, the current finding showed that the ELISA kit we developed can be used for the rapid detection of ASFV antibodies and used as an alternative during serological investigation of ASF in endemic areas.

277 Effect of yeast probiotics in lactation and nursery diets on progeny lifetime growth performance, immune response, and carcass characteristics

Abstract Mixed parity sows (n = 28; Line 241; DNA) and their offspring were used in a farrow-to-finish study to evaluate the effect of live yeast supplementation during lactation with or without yeast extract supplementation during the nursery period on sow and litter performance and lifetime growth performance, antibody titers, and carcass characteristics. Sows were blocked by parity and body weight (BW) on d 110 of gestation and allotted to 1 of 2 dietary treatments with 14 replications per treatment which consisted of a standard corn-soybean meal lactation diet with or without a yeast-based probiotic (0.10% Actisaf Sc 47 HR+; Phileo by Lesaffre, Milwaukee, WI). Following weaning, pigs (n = 350; 241×600, DNA; initial BW = 6.1 ± 0.02 kg) were randomly assigned in a 2 × 2 factorial design within sow treatment to 1 of 2 nursery diets which consisted of a control diet or a diet with pre-probiotic combination (0.10% MS309; Phileo by Lesaffre, Milwaukee, WI) fed for 42 d after weaning followed by common diets until marketing. Two nursery pens were combined in the finisher, such that there were 5 and 10 pigs/pen with 18 and 9 replications/treatment during the nursery and finishing periods, respectively. There were no significant effects of live yeast supplementation on lactation performance (P > 0.079). There was a sow × nursery diet interaction (P = 0.024) during the nursery period where pigs from sows fed Actisaf had increased average daily gain (ADG) when fed control nursery diets compared with pigs from control sows fed control nursery diets (Table). Pigs fed MS309 in the nursery from either sow treatment were intermediate. Pigs from sows fed Actisaf tended to be heavier at marketing (P = 0.067) with heavier hot carcass weight (HCW; P = 0.101), but there were no differences in overall finishing growth performance with the inclusion of yeast in lactation diets or nursery diets (P > 0.10). A subset of pigs were sampled at 7 time points to determine serum porcine circovirus type 2 (PCV2) and Mycoplasma hyopneumoniae antibody sample-to-positive (S/P) ratios and at 5 time points to ascertain percent inhibition of Lawsonia intracellularis. For all 3 immunological criteria, there were no sow diet × nursery diet × day interactions. There was a sow diet × day interaction for PCV2 S/P ratio (P = 0.097) where offspring from Actisaf sows tended to have greater PCV2 S/P ratios at 101 d of age compared with offspring from control sows (P = 0.046). There was a PCV2 S/P ratio nursery diet × day interaction (P = 0.036) where pigs fed MS309 had reduced PCV2 S/P ratios at 66, 78, and 162 d of age (P ≤ 0.022). In conclusion, feeding a pre/probiotic combination in the nursery did not affect performance or immune criteria. Conversely, feeding a live yeast probiotic during lactation resulted in tendencies for increased nursery ADG, final BW, and HCW.

269 Influence of herbal active D on nursery pig growth performance, bone characteristics and blood measures

Abstract Pigs [n = 2.268; L337×1050 PIC; initial body weight (BW) = 5.5 ± 0.17 kg] were used in a 42-d growth study to evaluate the effects of herbal active D (a source of calcitriol) on growth performance, bone characteristics, and serum criteria of nursery pigs. Pigs were weaned at approximately 21-d of age and randomly allotted to 1 of 3 dietary treatments in a randomized complete block design. There were 27 pigs/pen and 28 replications/treatment across 2 rooms with pens blocked by BW and weaning date. Treatment diets were corn-soybean meal-based and consisted of a control diet (1,650 IU/kg of vitamin D3), or the control diet with 120 or 200 mg/kg of herbal active D (Phytobiotics, St. Louis, MO). Blood samples were collected from 25 gilts/treatment on d 21 and 42 to assess 25(OH)D3 and cytokine concentrations. At the end of the study, 10 pigs/treatment were euthanized and the right fibula, metacarpal, 2nd rib, and 10th rib were collected to determine bone density, bone breaking strength, and percentage bone ash by utilizing the de-fatted processing method. Overall (d 0 to 42), there was a marginally significant (P = 0.067) reduction in gain to feed ratio (G:F) as herbal active D increased, but no effect on final BW, average daily gain (ADG) and average daily feed intake (ADFI), or mortality (P > 0.10; Table). For bone breaking strength and bone ash, there was no evidence of a treatment × bone interaction (P > 0.10). Percentage of bone ash increased linearly across all bones as herbal active D increased (P = 0.026). Treatment did not affect bone ash weight and breaking strength (P > 0.10). However, metacarpals and 10th ribs had the greatest bone ash weight followed by the fibula with 2nd rib having the least (P < 0.05). Metacarpals had greater breaking strength compared with all other bones, followed by the fibula and 10th rib, with the 2nd rib having the least (P < 0.001). There was a bone × treatment interaction for bone density, where increasing herbal active D increased bone density for the 2nd rib (P = 0.012), but there were no treatment differences for other bones (P > 0.10). Additionally, there was no difference (P > 0.10) between treatments for porcine circovirus type 2 S/P ratio, porcine reproductive and respiratory syndrome, Mycoplasma hyopneumoniae, 25(OH)D3 status or circulating cytokine concentrations except for IL-8 concentrations which increased linearly (P = 0.027) as herbal active D increased. Antibodies and cytokine concentrations were greater on d 21 compared with d 42 (P < 0.001), except for IL-1ra and IL-8 having no significant (P > 0.10) day effect. In summary, herbal active D had minimal impact on growth or serum parameters; however, increasing herbal active D increased percentage bone ash.

Establishment of real-time viral pathogen detection method-surface-enhanced Raman scattering system for porcine circovirus type II

The transition towards large-scale intensive pig farming methods has resulted in an increase in cases of complex, mixed infections, and secondary infections. Porcine circovirus (PCV), the causative agent of PCV disease (PCVD), is a small nonenveloped DNA virus containing a single-stranded circular genome. Previously, PCV was even considered nonpathogenic to pigs. However, a novel strain of PCV, designated PCV-2, has been associated with various disease syndromes in pigs over the last 5 years. PCV-2 primarily induces post-weaning multisystemic wasting syndrome, characterized by clinical signs such as debility, dyspnea, palpable lymphadenopathy, diarrhea, and pallor or icterus. PCV-2 is now recognized as an important emerging pathogen. Early diagnosis and disease prevention have become crucial for effective pig farm management. However, clinical diagnosis may be complicated by antibiotic treatment and atypical disease symptoms. To ensure accurate pathology diagnosis, it is essential to integrate robust laboratory diagnostics with traditional methods. Surface-enhanced Raman Scattering (SERS) spectroscopy has emerged as a potentially powerful technique for whole-organism fingerprinting, enabling rapid identification of bacteria. Biosensors utilizing SERS offer promising capabilities for sensitive and quick detection of bacterial pathogens, thereby reducing diagnosis time. In this study, our aim was to characterize and evaluate a SERS-based diagnostic system for detecting and identifying PCV-2 present in pooled swine sera and feces. We compared the spectra of PCV-2 recovered from the specimens to those of pure PCV-2 to determine the PCV-2 molecular fingerprint. Our results demonstrated successful detection, identification, and classification of PCV-2 in swine sera and feces using SERS. SERS provided reproducible molecular spectroscopic signatures suitable for analytical applications. This approach presents a new and potent tool for real-time surveillance of animal viral pathogens in clinical settings.

Frequency of seropositivity against porcine circovirus type 2 (PCV2) in the metropolitan area of Monterrey, Nuevo León, and its peripheral area

El circovirus porcino tipo 2 (PCV2) es un virus de tipo DNA que tiene afinidad por células del sistema inmune y que genera depleción de linfocitos por lo que favorece el desarrollo de enfermedades causadas por otros agentes oportunistas, así mismo está relacionado con la generación de diferentes síndromes. Por lo anterior, este virus produce importantes afecciones a la industria porcícola, sin embargo, fácilmente se pueden prevenir los síndromes asociados a PCV2 mediante la adecuada aplicación de medidas de bioseguridad y vacunación. Por otro lado, las unidades de producción a pequeña escala (UPPs) suelen carecer de este tipo de manejo preventivo, así como de vigilancia rutinaria por parte de un médico veterinario. Si bien, el PCV2 se considera un virus ampliamente distribuido, no existen reportes de su presencia en las UPPs en Nuevo León. Se determinó la presencia de anticuerpos contra PCV2 mediante el uso de un kit comercial y se realizó la biometría hemática de los animales. Se localizaron 48 UPPs en las que se encontró un 91.67% de positividad, así como un 89.7% de seropositividad en los animales. En la biometría se encontró que la HGB y el HCT se presentaron disminuidos en los individuos que resultaron positivos a anticuerpos comparados contra los negativos (P=0.03 y P=0.01, respectivamente), por el contrario, el valor de células blancas totales se encontró disminuido en los individuos que resultaron negativos a la presencia de anticuerpo contra PCV2 (P=0.01).

PCV2 Induced Endothelial Derived IL-8 Affects MoDCs Maturation Mainly via NF-κB Signaling Pathway.

Porcine circovirus type 2 (PCV2) infection can cause immunosuppressive diseases in pigs. Vascular endothelial cells (VECs), as the target cells for PCV2, play an important role in the immune response and inflammatory regulation. Endothelial IL-8, which is produced by porcine hip artery endothelial cells (PIECs) infected with PCV2, can inhibit the maturation of monocyte-derived dendritic cells (MoDCs). Here, we established a co-culture system of MoDCs and different groups of PIECs to further investigate the PCV2-induced endothelial IL-8 signaling pathway that drives the inhibition of MoDC maturation. The differentially expressed genes related to MoDC maturation were mainly enriched in the NF-κB and JAK2-STAT3 signaling pathways. Both the NF-κB related factor RELA and JAK2-STAT3 signaling pathway related factors (IL2RA, JAK, STAT2, STAT5, IL23A, IL7, etc.) decreased significantly in the IL-8 up-regulated group, and increased significantly in the down-regulated group. The expression of NF-κB p65 in the IL-8 up-regulated group was reduced significantly, and the expression of IκBα was increased significantly. Nuclear translocation of NF-κB p65 was inhibited, while the nuclear translocation of p-STAT3 was increased in MoDCs in the PCV2-induced endothelial IL-8 group. The results of treatment with NF-κB signaling pathway inhibitors showed that the maturation of MoDCs was inhibited and the expression of IL-12 and GM-CSF at mRNA level were lower. Inhibition of the JAK2-STAT3 signaling pathway had no significant effect on maturation, and the expression of IL-12 and GM-CSF at mRNA level produced no significant change. In summary, the NF-κB signaling pathway is the main signaling pathway of MoDC maturation, and is inhibited by the PCV2-induced up-regulation of endothelial-derived IL-8.