Adjuvant screening of the Senecavirus A inactivated vaccine in mice and evaluation of its immunogenicity in pigs.

BACKGROUND: Senecavirus A (SVA) causes an emerging vesicular disease (VD) with clinical symptoms indistinguishable from other vesicular diseases, including vesicular stomatitis (VS), foot-and-mouth disease (FMD), and swine vesicular disease (SVD). Currently, SVA outbreaks have been reported in Canada, the U.S.A, Brazil, Thailand, Vietnam, Colombia, and China. Based on the experience of prevention and control of FMDV, vaccines are the best means to prevent SVA transmission. RESULTS: After preparing an SVA inactivated vaccine (CH-GX-01-2019), we evaluated the immunogenicity of the SVA inactivated vaccine mixed with Imject® Alum (SVA + AL) or Montanide ISA 201 (SVA + 201) adjuvant in mice, as well as the immunogenicity of the SVA inactivated vaccine combined with Montanide ISA 201 adjuvant in post-weaned pigs. The results of the mouse experiment showed that the immune effects in the SVA + 201 group were superior to that in the SVA + AL group. Results from pigs immunized with SVA inactivated vaccine combined with Montanide ISA 201 showed that the immune effects were largely consistent between the SVA-H group (200 µg) and SVA-L group (50 µg); the viral load in tissues and blood was significantly reduced and no clinical symptoms occurred in the vaccinated pigs. CONCLUSIONS: Montanide ISA 201 is a better adjuvant choice than the Imject® Alum adjuvant in the SVA inactivated vaccine preparation, and the CH-GX-01-2019 SVA inactivated vaccine can provide effective protection for pigs.

Subunit vaccine raised against the SARS-CoV-2 spike of Delta and Omicron variants.

Vaccination has proven effective against SARS-CoV-2 infection but vaccines were originally based on the wild type and emerging variants have led to a decrease in protective efficacy. There is an urgent need for broad-spectrum vaccine protection against emerging variants. A vaccine based on the Delta strain spike protein was created by optimization of vector, codon, and protein structure to produce a subunit immunogen (Delta-6P-S) containing six proline mutations, stable pre-fusion conformation, and with high expression in CHO-S cells. Immunogenicity and protective efficacy were evaluated in mice and golden hamsters using alum adjuvant. The Delta-6P-S recombinant protein induced strong immune responses in C57BL/6J mice and golden hamsters and sera had cross-neutralization activity and neutralized wild type and Beta, Delta, Omicron BA.1, BA.2, and BA.5 variant strains. Golden hamsters were immunized against Delta, Omicron BA.1, and BA.2 variants. Viral RNA detected from throat swabs, lungs and tracheas decreased significantly in vaccine-inoculated animals relative to alum-treated controls and no infectious viruses were detected in lungs and tracheas. Almost no pathological damage to lung tissue was found in vaccinated animals by contrast with those treated only with alum. The Delta-6P-S recombinant protein rapidly eliminated replicating virus in the upper and lower airways of golden hamsters and merits further investigation as a candidate anti-SARS-CoV-2 vaccine.

Synergistic pathogenicity by coinfection and sequential infection with JXA1-like HP-PRRSV and PCV2d in PCV2 antibody-positive post-weaned pigs.

Porcine reproductive and respiratory syndrome virus (PRRSV) and Porcine circovirus (PCV) are two important pathogens, which caused respiratory disease in pigs. PRRSV and PCV2 had caused great economic losses to the pig industry. Pigs coinfection with PCV2 and PRRSV were common in the clinic, PCV2 antibodies can be detected in most of the pigs. PCV2d and HP-PRRSV(JXA1-like) were two major viruses circulating in the pigs in China. In this study, HP-PRRSV (JXA1-like) and PCV2d were used to coinfect and (or) sequential infect 5-week-old weaned PCV2-antibody positive pigs and the clinical indications, pathological, virus load, and specific antibodies of the challenged post-weaned piglets were evaluated. Thirty 5-week-old post-weaned pigs were divided into six groups infected with PBS, PCV2, PRRSV, PCV2-PRRSV, PRRSV-PCV2, and Co-PRRSV-PCV2 according to the PCV2 specific antibodies. Pigs infected with PRRSV can experience diarrhea, increased body temperature, weight loss, and even death. The pigs in the PRRSV infected group and PRRSV-PCV2 infected group showed severe clinical symptoms, high mortality, and low average daily gain. The main pathological changes were widening of the lung interstitium, lung adhesion, and so on. The PRRSV-PCV2 infected group showed high levels of TNF-α and IL-2. In conclusion, PRRSV and PRRSV-PCV2 sequential infected pigs showed most pathogenic signs, and PCV2-PRRSV sequential infected pigs showed less pathogenicity than pigs of PCV2 and PRRSV coinfection and PRRSV monoinfection from day 10-14, partially suppressing the cytokine storm produced by PRRSV.

Immunogenicity and protective efficacy of a DNA vaccine inducing optimal expression of the SARS-CoV-2 S gene in hACE2 mice.

The wide spread of coronavirus disease 2019 (COVID-19) has significantly threatened public health. Human herd immunity induced by vaccination is essential to fight the epidemic. Therefore, highly immunogenic and safe vaccines are necessary to control SARS-CoV-2, whose S protein is the antigenic determinant responsible for eliciting antibodies that prevent viral entry and fusion. In this study, we developed a SARS-CoV-2 DNA vaccine expressing the S protein, named pVAX-S-OP, which was optimized according to the human-origin codon preference and using polyinosinic-polycytidylic acid as an adjuvant. pVAX-S-OP induced specific antibodies and neutralizing antibodies in BALB/c and hACE2 transgenic mice. Furthermore, we observed 1.43-fold higher antibody titers in mice receiving pVAX-S-OP plus adjuvant than in those receiving pVAX-S-OP alone. Interferon gamma production in the pVAX-S-OP-immunized group was 1.58 times (CD3+CD4+IFN-gamma+) and 2.29 times (CD3+CD8+IFN-gamma+) lower than that in the pVAX-S-OP plus adjuvant group but higher than that in the control group. The pVAX-S-OP vaccine was also observed to stimulate a Th1-type immune response. When, hACE2 transgenic mice were challenged with SARS-CoV-2, qPCR detection of N and E genes showed that the viral RNA loads in pVAX-S-OP-immunized mice lung tissues were 104 times and 106 times lower than those of the PBS control group, which shows that the vaccine could reduce the amount of live virus in the lungs of hACE2 mice. In addition, pathological sections showed less lung damage in the pVAX-S-OP-immunized group. Taken together, our results demonstrated that pVAX-S-OP has significant immunogenicity, which provides support for developing SARS-CoV-2 DNA candidate vaccines.

Genetic characterization of a new NSP2-deletion porcine reproductive and Respiratory Syndrome Virus in China.

The Porcine Reproductive and Respiratory Syndrome Virus (PRRSV) cause a huge economic loss around the pig industry worldwide; the NADC30-like PRRSV have attracted much attention outbreaks in China in recent years. Recombination between PRRSV subtypes, point mutations, insertions and deletions that contribute to the emergence of new variants in the genome. In this study, the PRRSV-HB-16-China-2019 strain's full-length genomic sequence shares 93.0% nucleotide similarity to NADC30 PRRSV without any gene insertion. Compared with VR-2332, it has an NSP2 coding region that is different from NADC30, which has a discontinuous 206-aa (111-aa from position 323 to 433 and 95-aa from position 476 to 570) deletion. Compared with other NADC30-Like strains, it has a discontinuous 75-amino acid (75-aa from position 476 to 552) deletion, which was first reported. Notably, the strain, PRRSV-HB-16-China-2019, contained an addition a 1-aa deletion in ORF5 and a unique 3-nt deletion in 3'-UTR similar to NADC30, the strain is recombined between a NADC30-like strain and a vaccine strain named RespPRRS MLV(parental strain VR-2332). Our findings indicate that PRRSV-HB-16-China-2019 is a new NSP2-deletion NADC30-like strain with certain deletions and mutations. Our results show that the emergence of the new NADC30-like strain has increased the difficulty of PRRSV prevention in China.

Pathogenicity of Seneca Valley virus in pigs and detection in Culicoides from an infected pig farm.

BACKGROUND: Porcine vesicular disease is caused by the Seneca Valley virus (SVV), it is a novel Picornaviridae, which is prevalent in several countries. However, the pathogenicity of SVV on 5-6 week old pigs and the transmission routes of SVV remain unknown. METHODS: This research mainly focuses on the pathogenicity of the CH-GX-01-2019 strain and the possible vector of SVV. In this study, 5-6 week old pigs infected with SVV (CH-GX-01-2019) and its clinical symptoms (including rectal temperatures and other clinical symptoms) were monitored, qRT-PCR were used to detect the viremia and virus distribution. Neutralization antibody assay was set up during this research. Mosquitoes and Culicoides were collected from pigsties after pigs challenge with SVV, and SVV detection within mosquitoes and Culicoides was done via RT-PCR. RESULTS: The challenged pigs presented with low fevers and mild lethargy on 5-8 days post infection. The viremia lasted more than 14 days. SVV was detected in almost all tissues on the 14th day following the challenge, and it was significantly higher in the hoofs (vesicles) and lymph nodes in comparison with other tissues. Neutralizing antibodies were also detected and could persist for more than 28 days, in addition neutralizing antibody titers ranged from 1:128 to 1:512. Mosquitoes and Culicoides were collected from the pigsty environments following SVV infection. Although SVV was not detected in the mosquitoes, it was present in the Culicoides, however SVV could not be isolated from the positive Culicoides. CONCLUSIONS: Our work has enriched the knowledge relating to SVV pathogenicity and possible transmission routes, which may lay the foundation for further research into the prevention and control of this virus.

Retrospective surveillance of porcine circovirus 4 in pigs in Inner Mongolia, China, from 2016 to 2018.

A novel circovirus designated "porcine circovirus type 4" (PCV4) was recently reported in pigs with severe clinical disease in Hunan Province, China. Relatively little is known about the molecular epidemiology of this recently discovered virus. In order to assess the prevalence of PCV4 infection in pigs and to analyze its genomic characteristics, 1683 clinical samples were collected in Inner Mongolia, China, from 2016 to 2018. The overall infection rate of PCV4 was 1.6% (27/1683) at the sample level and 21.6% (11/51) at the farm level, with rates ranging from 3.2% (1/31) to 20.0% (6/30) on different PCV4-positive pig farms. In addition, the PCV4 infection rates at both the sample and farm level increased from 2016 to 2018. This also showed that PCV4 was present in pigs in 2016 in China and therefore did not arrive later than this date. Additionally, our findings showed that PCV4 infections had no association with PCV2 or PCV3 infections. We sequenced the complete genomes of three PCV4 strains and found that the PCV4 strains had a high degree of genetic stability but shared less than 80% sequence identity with other circoviruses. We identified six amino acid mutations in the Rep protein and seven in the Cap protein. Phylogenetic analysis based on Cap and Rep sequences confirmed that the PCV4 strains grouped in an independent branch. Our findings provide important information about the prevalence and genetic characteristics of PCV4 strains.

Construction and immunogenicity of Senecavirus A virus-like particle vaccine with adjuvant.

Senecavirus A (SVA) is constantly associated with vesicular disease in pigs, and the clinical symptoms of pig infection with SVA are indistinguishable from other porcine vesicular diseases. Vaccine is one of the best methods to eliminate and control the spread of SVA. Virus-like particles (VLPs) can play important roles in prevention for infectious diseases. Here, the SVA VLPs was assembled by the baculovirus expression vector system, and the immunogenicity of the SVA VLPs mixed with different adjuvants were evaluated in mice and pigs. Two recombinant baculoviruses (rPFBD-VP1-VP3 and rPFBD-VP2-VP4) were constructed, which co-infected with Sf9 suspension cells to assemble SVA VLPs successfully. SVA VLPs mixed with ISA201 adjuvant and ISA201 +Poly(I:C) adjuvant produced higher levels of neutralizing antibody, specific antibody (total IgG, IgG1, IgG2a and IgG2b) and cytokines in the T cells. And there was no significant difference between SVA VLPs+ 201 group and SVA VLPs+Poly(I:C)+ 201 group. Pigs immunized with high dose of SVA VLPs mixed with ISA201 adjuvant could produce higher titers of neutralizing antibody and SVA-specific antibody. Furthermore, the protection rates of SVA VLPs-H and SVA VLPs-L were 100% and 80%, and the viral load of SVA VLPs-H group is the lowest in all SVA VLPs groups. It is the first time to develop the SVA VLPs using the baculovirus expression vector system, which may lay the foundation for the research and development of SVA vaccine.

Silk Fibroin-Coated Nano-MOFs Enhance the Thermal Stability and Immunogenicity of HBsAg.

Vaccines are widely regarded as one of the most effective weapons in the fight against infectious diseases. Currently, vaccines must be stored and transported at low temperatures as high temperatures can lead to a loss of vaccine conformation and reduced therapeutic efficacy. Metal-organic frameworks (MOFs), such as zeolitic imidazole framework-8 (ZIF-8), are a new class of hybrid materials with large specific surface areas, high loading rates, and good biocompatibility and are successful systems for vaccine delivery and protection. Silk fibroin (SF) has a good biocompatibility and thermal stability. In this study, the hepatitis B surface antigen (HBsAg) was successfully encapsulated in ZIF-8 to form HBsAg@ZIF-8 (HZ) using a one-step shake and one-pot shake method. Subsequently, the SF coating modifies HZ through hydrophobic interactions to form HBsAg/SF@ZIF-8 (HSZ), which enhanced the thermal stability and immunogenicity of HBsAg. Compared to free HBsAg, HZ and HSZ improved the thermostability of HBsAg, promoted the antigen uptake and lysosomal escape, stimulated dendritic cell maturation and cytokine secretion, formed an antigen reservoir to promote antibody production, and activated CD4+ T and CD8+ T cells to enhance memory T-cell production. Importantly, HSZ induced a strong immune response even after 14 days of storage at 25 °C. Furthermore, the nanoparticles prepared by the one-step shake method exhibited superior properties compared to those prepared by the one-pot shake method. This study highlights the importance of SF-coated ZIF-8, which holds promise for investigating thermostable vaccines and breaking the vaccine cold chain.

Molecular characterization of an outbreak-involved Bacillus anthracis strain confirms the spillover of anthrax from West Africa.

BACKGROUND: Anthrax, a zoonotic disease caused by the spore-forming bacterium Bacillus anthracis, remains a major global public health concern, especially in countries with limited resources. Sierra Leone, a West African country historically plagued by anthrax, has almost been out of report on this disease in recent decades. In this study, we described a large-scale anthrax outbreak affecting both animals and humans and attempted to characterize the pathogen using molecular techniques. METHODS: The causative agent of the animal outbreak in Port Loko District, Sierra Leone, between March and May 2022 was identified using the nanopore sequencing technique. A nationwide active surveillance was implemented from May 2022 to June 2023 to monitor the occurrence of anthrax-specific symptoms in humans. Suspected cases were subsequently verified using quantitative polymerase chain reaction. Full-genome sequencing was accomplished by combining long-read and short-read sequencing methods. Subsequent phylogenetic analysis was performed based on the full-chromosome single nucleotide polymorphisms. RESULTS: The outbreak in Port Loko District, Sierra Leone, led to the death of 233 animals between March 26th and May 16th, 2022. We ruled out the initial suspicion of Anaplasma species and successfully identified B. anthracis as the causative agent of the outbreak. As a result of the government's prompt response, out of the 49 suspected human cases identified during the one-year active surveillance, only 6 human cases tested positive, all within the first month after the official declaration of the outbreak. The phylogenetic analysis indicated that the BaSL2022 isolate responsible for the outbreak was positioned in the A.Br.153 clade within the TransEuroAsian group of B. anthracis. CONCLUSIONS: We successfully identified a large-scale anthrax outbreak in Sierra Leone. The causative isolate of B. anthracis, BaSL2022, phylogenetically bridged other lineages in A.Br.153 clade and neighboring genetic groups, A.Br.144 and A.Br.148, eventually confirming the spillover of anthrax from West Africa. Given the wide dissemination of B. anthracis spores, it is highly advisable to effectively monitor the potential reoccurrence of anthrax outbreaks and to launch campaigns to improve public awareness regarding anthrax in Sierra Leone.

Molecular Detection and Genetic Characterization of Japanese Encephalitis Virus in Animals from 11 Provinces in China.

Japanese encephalitis virus (JEV), which uses a mosquito primary vector and swine as a reservoir host, poses a significant risk to human and animal health. JEV can be detected in cattle, goats and dogs. A molecular epidemiological survey of JEV was conducted in 3105 mammals from five species, swine, fox, racoon dog, yak and goat, and 17,300 mosquitoes from 11 Chinese provinces. JEV was detected in pigs from Heilongjiang (12/328, 3.66%), Jilin (17/642, 2.65%), Shandong (14/832, 1.68%), Guangxi (8/278, 2.88%) and Inner Mongolia (9/952, 0.94%); in goats (1/51, 1.96%) from Tibet; and mosquitoes (6/131, 4.58%) from Yunnan. A total of 13 JEV envelope (E) gene sequences were amplified in pigs from Heilongjiang (5/13), Jilin (2/13) and Guangxi (6/13). Swine had the highest JEV infection rate of any animal species, and the highest infection rates were found in Heilongjiang. Phylogenetic analysis indicated that the predominant strain in Northern China was genotype I. Mutations were found at residues 76, 95, 123, 138, 244, 474 and 475 of E protein but all sequences had predicted glycosylation sites at 'N154. Three strains lacked the threonine 76 phosphorylation site from non-specific (unsp) and protein kinase G (PKG) site predictions; one lacked the threonine 186 phosphorylation site from protein kinase II (CKII) prediction; and one lacked the tyrosine 90 phosphorylation site from epidermal growth factor receptor (EGFR) prediction. The aim of the current study was to contribute to JEV prevention and control through the characterization of its molecular epidemiology and prediction of functional changes due to E-protein mutations.

Construction and Evaluation of Recombinant Adenovirus Candidate Vaccines for Chikungunya Virus.

Chikungunya virus (CHIKV) is a mosquito-borne virus. The emergence of CHIKV infection has raised global concern, and there is a growing need to develop safe and effective vaccines. Here, adenovirus 5 was used as the vaccine vector to construct recombinant adenoviruses expressing CHIKV E2, E1, and E2-6K-E1, respectively. And then the immunogenicity and protective efficiency against CHIKV were evaluated in BALB/c mice. Compared to the ad-wt control group, all three vaccines elicited significant humoral and cellar immune responses. The levels of neutralizing antibodies in the rAd-CHIKV-E2-6K-E1 and rAd-CHIKV-E2 groups both reached 1:256, which were 3.2 times higher than those in the rAd-CHIKV-E1 group. Furthermore, the levels of lymphocyte proliferation in rAd-CHIKV-E2-6K-E1 group were the highest. Besides, the concentrations of IFN-γ and IL-4 in mice immunized with rAd-CHIKV-E2-6K-E1 were 1.37 and 1.20 times higher than those in ad-wt immunized mice, respectively. After the challenge, mice in the rAd-CHIKV-E2-6K-E1 and rAd-CHIKV-E2 groups lost 2% of their body weight compared with 5% in the ad-wt control group. And low viral loads were detected in the heart, kidney, and blood of mice immunized with rAd-CHIKV-E2-6K-E1 and rAd-CHIKV-E2 at 3-5 dpc, which decreased by 0.4-0.7 orders of magnitude compared with the ad-wt control. Overall, these data suggest that the recombinant adenovirus is a potential candidate vaccine against CHIKV.

Codon Usage for Genetic Diversity, and Evolutionary Dynamics of Novel Porcine Parvoviruses 2 through 7 (PPV2-PPV7).

Porcine parvovirus (PPV) is the main pathogen of reproductive disorders. In recent years, a new type of porcine parvovirus has been discovered and named porcine parvovirus 2 to 7 (PPV2-PPV7), and it is associated with porcine circovirus type 2 in pigs. Codon usage patterns and their effects on the evolution and host adaptation of different PPV sub-types are still largely unknown. Here, we define six main sub-types based on the Bayesian method of structural proteins of each sub-type of PPV, including PPV2, PPV3, PPV4, PPV5, PPV6, and PPV7, which show different degrees of codon usage preferences. The effective number of codons (ENC) indicates that all PPV sub-types have low codon bias. According to the codon adaptation index (CAI), PPV3 and PPV7 have the highest similarity with the host, which is related to the main popular tendency of the host in the field; according to the frequency of optimal codons (FOP), PPV7 has the highest frequency of optimal codons, indicating the most frequently used codons in its genes; and according to the relative codon deoptimization index (RCDI), PPV3 has a higher degree. Therefore, it is determined that mutational stress has a certain impact on the codon usage preference of PPV genes, and natural selection plays a very decisive and dominant role in the codon usage pattern. Our research provides a new perspective on the evolution of porcine parvovirus (PPV) and may help provide a new method for future research on the origin, evolutionary model, and host adaptation of PPV.

Establishment of a reverse transcription real-time quantitative PCR method for Getah virus detection and its application for epidemiological investigation in Shandong, China.

Getah virus (GETV) is a mosquito-borne, single-stranded, positive-sense RNA virus belonging to the genus Alphavirus of the family Togaviridae. Natural infections of GETV have been identified in a variety of vertebrate species, with pathogenicity mainly in swine, horses, bovines, and foxes. The increasing spectrum of infection and the characteristic causing abortions in pregnant animals pose a serious threat to public health and the livestock economy. Therefore, there is an urgent need to establish a method that can be used for epidemiological investigation in multiple animals. In this study, a real-time reverse transcription fluorescent quantitative PCR (RT-qPCR) method combined with plaque assay was established for GETV with specific primers designed for the highly conserved region of GETV Nsp1 gene. The results showed that after optimizing the condition of RT-qPCR reaction, the minimum detection limit of the assay established in this study was 7.73 PFU/mL, and there was a good linear relationship between viral load and Cq value with a correlation coefficient (R 2) of 0.998. Moreover, the method has good specificity, sensitivity, and repeatability. The established RT-qPCR is 100-fold more sensitive than the conventional RT-PCR. The best cutoff value for the method was determined to be 37.59 by receiver operating characteristic (ROC) curve analysis. The area under the curve (AUC) was 0.956. Meanwhile, we collected 2,847 serum specimens from swine, horses, bovines, sheep, and 17,080 mosquito specimens in Shandong Province in 2022. The positive detection rates by RT-qPCR were 1%, 1%, 0.2%, 0%, and 3%, respectively. In conclusion, the method was used for epidemiological investigation, which has extensive application prospects.

Synergistic Pathogenicity by Coinfection and Sequential Infection with NADC30-like PRRSV and PCV2 in Post-Weaned Pigs.

Porcine reproductive and respiratory syndrome virus (PRRSV) and porcine circovirus (PCVs) are two major viruses that affect pigs. Coinfections between PRRSV and PCV2 are frequently reported in most outbreaks, with clinical presentations involving dyspnea, fever, reduced feed intake, weight loss, and death in fattening pigs. The NADC30-like PRRSV and PCV2d are the main circulating virus strains found in China. This study determines the impact of NADC30-like PRRSV and PCV2d mono-infection and coinfection on the immune system, organ pathology, and viral shedding in five-week-old post-weaned pigs. Pigs were randomly divided into six groups: PBS, PRRSV, PCV2, PRRSV-PCV2 coinfection (co), and PRRSV-PCV2 or PCV2-PRRSV sequential infections. Fever, dyspnea, decreased feed intake, weight loss, and pig deaths occurred in groups infected with PRRSV, Co-PRRSV-PCV2, and PRRSV-PCV2. The viral load was higher in Co-PRRSV-PCV2, PRRSV-PCV2, and PCV2-PRRSV than those mono-infected with PRRSV or PCV2. Additionally, cytokines (IFN-γ, TNF-α, IL-4, and IL-10) produced by pigs under Co-PRRSV-PCV2 and PRRSV-PCV2 groups were more intense than the other groups. Necropsy findings showed hemorrhage, emphysema, and pulmonary adhesions in the lungs of pigs infected with PRRSV. Smaller alveoli and widened lung interstitium were found in the Co-PRRSV-PCV2 and PRRSV-PCV2 groups. In conclusion, PRRSV and PCV2 coinfection and sequential infection significantly increased viral pathogenicity and cytokine responses, resulting in severe clinical signs, lung pathology, and death.

Genetic evolution and epidemiological analysis of Seneca Valley virus (SVV) in China.

Seneca Valley virus (SVV) is a novel Picornaviridae that is closely associated with porcine idiopathic vesicular disease (PIVD). Here, a novel SVV strain (CH-GX-01-2019) was detected and isolated from swine in Guangxi Province, China. The complete genomic sequence of CH-GX-01-2019 exhibited 93.3-98.9 % identify with other SVV isolates at the nucleotide level. CH-GX-01-2019 showed the highest level of similarity (98.9 %) with Vietnamese strains. And CH-GX-01-2019 exhibited two consecutive amino acid mutations in VP1 gene. Phylogenetic analysis based on the complete genome and the VP1 gene showed that Chinese SVV isolates can be divided into three clusters. We analyzed the geographical distributions of SVV strains in China and found that the epidemiology of SVV in China is complicated; most strains are distributed predominantly in south and central China. Between 2015 and 2019, the dominant epidemic SVV isolates in China have changed from clusters 1 and 3 to cluster 2. CH-GX-01-2019 (cluster 3) is a recombinant strain from Colombia-2016 (cluster 2) and HB-CH-2016 (cluster 1). Our findings will enhance our understanding of the prevalence and genetic variation of SVV in the swine herds of China and provide important insights into the molecular epidemiology of SVV.

Prevalence, pathogenesis, and evolution of porcine circovirus type 3 in China from 2016 to 2019.

Porcine circovirus type 3 (PCV3) infection causes substantial economic losses in pig populations since its discovery in 2016. However, PCV3 molecular epidemiology remains need further study. In order to assess the prevalence of PCV3 infection in China, 4094 clinical samples from 271 pig farms in 10 provinces of China were evaluated by PCR. It was shown that the overall prevalence of PCV3 infection was 29.3 % (1200/4094) and 74.2 % (201/271) at sample and farm levels respectively, suggesting that PCV3 infection is prevalent in China. Furthermore, a statistical analysis showed PCV3 might exacerbate PCV2 and PRRSV infection rate and have a potential association with pig clinical disease. In addition, we sequenced the entire genome of 57 PCV3 strains; homology analysis showed that PCV3 strains have more than 96 % similarities at the nucleotide level, and PCV3 shares less than 60 % similarities with other circoviruses. By comparing the total 673 PCV3 strains from the NCBI GenBank, we found the major of amino acid mutations are located in predicted epitope regions and the mutations ratio changed during PCV3 evolution. Phylogenetic analysis revealed that all isolates belonged to PCV3a and PCV3b, and increasing PCV3a and decreasing PCV3b trends were observed during PCV3 evolution. Overall, this study provides important insights for understanding PCV3 prevalence, pathogenesis, and evolution and will guide future efforts to develop effective preventive and control measures.

Pathogenicity of porcine reproductive and respiratory syndrome virus (ORF5 RFLP 1-7-4 viruses) in China.

Porcine reproductive and respiratory syndrome virus (PRRSV) is an RNA virus that causes reproductive failure in sows and respiratory problems in piglets. PRRSV infection leads to substantial pig mortality and causing huge economic losses so that disease outbreaks caused by the new PRRSV strain from other regions have caused great concern in China. In this study, we analysed the pathogenicity of the novel ORF5 RFLP 1-7-4-like PRRSV strain, named PRRSV-ZDXYL-China-2018-1 in pigs. The viral challenge test showed that PRRSV-ZDXYL-China-2018-1 infection can cause persistent fever, moderate dyspnoea, serum viraemia and interstitial pneumonia in piglets. The levels of viral loads in serum and PRRSV-specific antigen were also detected in lung tissues were used one-step Taq-Man RT-qPCR and Immunohistochemistry, respectively. At 28dpi, the level of specific antibodies was increased among infected piglets. Importantly, the new virus appeared be a moderately virulent isolate with pathogenicity compared to HP-PRRSV strain LQ (JXA1-like strain). Histological examination revealed severe monocyte haemorrhage and interstitial pneumonia associated with monocyte infiltration in the lung tissue of pigs infected with PRRSV-ZDXYL-China-2018-1 and LQ-JXA1 strains. Immunohistochemistry (IHC) results showed positive brown-red epithelial cells and macrophages in pig lungs. Therefore, it is critical to establish an effective strategy to control the spread of PRRSV in China.