Identification of a novel circovirus in blood sample of giant pandas (Ailuropoda melanoleuca).

The members of the family Circoviridae are considered to be one of the smallest autonomously replicating viruses that are classified into two genera, Circovirus and Cyclovirus. Circoviruses have been found in a variety of vertebrates, but whether they infect endangered protected animals has not been studied in much detail. Here, viral metagenomics and PCR methods were used to detect and verify viral nucleic acid in the blood sample from giant pandas. According to these methods, the complete genome sequence of a novel circovirus, the giant panda associated circovirus (GPCV) from the blood sample of three giant pandas was identified. The GPCV genome is 2090 bp in size and reveals two putative ambisense open-reading frames, encoding the major structural capsid protein and the replication associated protein, respectively, the latter having two predicted introns. Pairwise sequence comparison and phylogenetic analyses indicated GPCV was a putative new species within genus Circovirus based on the species demarcation criteria of the International Committee on the Taxonomy of Viruses. It is the first time that circovirus has been identified from blood sample of giant pandas. These efforts will contribute to future analyses to illuminate the evolutionary relationships between classified and newly identified members of the family Circoviridae.

Porcine circovirus type 2 (PCV2) infection in Hebei Province from 2016 to 2019: a retrospective study.

Porcine circovirus type 2 (PCV2) is the primary causative agent of porcine circovirus-associated diseases in swine, the most common of which are postweaning multisystemic wasting syndrome (PMWS) and porcine dermatitis and nephropathy syndrome (PDNS). To investigate the prevalence and genetic diversity of PCV2 in Hebei Province, Northern China, from 2016 to 2019, a total of 448 suspected cases of PCV2 infection were studied, and 179 samples were positive for PCV2. A pathological and histopathological examination suggested PCV2 to be cause of the observed lesions. Phylogenetic analysis showed that four genotypes were prevalent in Hebei Province: PCV2a, 2b, 2d, and 2e. Analysis of PCV2 strains using RDP4 and SimPlot showed that there were genetic recombination events among PCV2 strains in Hebei Province. A total of 3284 serum samples were screened by ELISA, and the positive rate of PCV2 antibodies was 73.9% (2428/3284). This study provides a scientific reference for the prevention and treatment of PCV2 in Hebei Province.

A comparative efficacy test of 1 versus 2 doses of CIRCOQ PCV2 subunit vaccine against naturally occurring PCV2-type d in piglets with high maternally derived antibodies (MDAs) on a Vietnamese swine farm.

The aim of this study was to evaluate the protective efficacy of the CIRCOQ porcine circovirus type 2 (PCV2) subunit vaccine in piglets with high maternally derived antibodies (MDAs) against disease caused by natural infection with PCV2d. A total of 130 weaned, 21-day-old healthy pigs was allocated into 3 trial groups. The signs of respiratory disorder were higher in unvaccinated pigs than in vaccinated pigs at 13 to 17 weeks old (P < 0.05), 18 to 22 weeks old (P < 0.001), and 23 to 27 weeks old (P < 0.01). The unvaccinated pigs had an early rate of dermatitis at 8 to 12 weeks old (10.0%), 13 to 17 weeks old (30.0%), 18 to 22 weeks old (46.7%), and 23 to 27 weeks old (33.3%), while there were no cases of dermatitis in vaccinated pigs. There was a significant difference (P < 0.05) in the mortality of pigs in the unvaccinated group and the 2-dosed vaccinated group. PCV2 viremia was detected in the blood and peaked at 105 days old in both unvaccinated pigs (Ct-adj = 8.40) and pigs vaccinated with 1 dose (Ct-adj = 6.37), while no detectable PCV2 virus was found in the blood of pigs vaccinated with 2 doses. At 77 and 105 days old, the PCV2 viremia load (Ct-adj) of unvaccinated pigs and those vaccinated with 1 dose was significantly higher (P < 0.05) than that of the 2-dosed vaccinated pigs. The body weight (BW), average weight gain (AWG), and average daily gain (ADG) in both groups of vaccinated pigs were significantly higher (P < 0.05) than those of unvaccinated pigs. The study vaccine was significantly efficacious in protecting vaccinated pigs against clinical symptoms, blood viral load, and mortality, as well as improving productivity, compared with unvaccinated pigs.

Le but de la présente étude était d'évaluer l'efficacité protectrice du vaccin sous-unitaire CIRCOQ du circovirus porcin de type 2 (PCV2) chez les porcelets ayant une grande quantité d'anticorps d'origine maternelle (MDA) contre la maladie causée par une infection naturelle par le PCV2d. Un total de 130 porcs sains sevrés âgés de 21 jours a été réparti dans trois groupes d'essai. Les signes de troubles respiratoires étaient plus élevés chez les porcs non vaccinés que chez les porcs vaccinés âgés de 13 à 17 semaines (P < 0,05), de 18 à 22 semaines (P < 0,001) et de 23 à 27 semaines (P < 0,01). Les porcs non vaccinés avaient un taux précoce de dermatite entre 8 et 12 semaines (10,0 %), 13 à 17 semaines (30,0 %), 18 à 22 semaines (46,7 %) et 23 à 27 semaines (33,3 %), alors qu'il n'y a eu aucun cas de dermatite chez les porcs vaccinés. Il y avait une différence significative (P < 0,05) dans la mortalité des porcs dans le groupe non vacciné et le groupe vacciné à deux doses. La virémie du PCV2 a été détectée dans le sang et a atteint un pic à 105 jours chez les porcs non vaccinés (Ct-adj = 8,40) et les porcs vaccinés avec une dose (Ct-adj = 6,37), tandis qu'aucun virus PCV2 détectable n'a été détecté dans le sang des porcs vacciné avec deux doses. À 77 et 105 jours, la charge de virémie PCV2 (Ct-adj) des porcs non vaccinés et de ceux vaccinés avec une dose était significativement plus élevée (P < 0,05) que celle des porcs vaccinés à deux doses. Le poids corporel (BW), le gain de poids moyen (AWG) et le gain quotidien moyen (ADG) dans les deux groupes de porcs vaccinés étaient significativement plus élevés (P < 0,05) que ceux des porcs non vaccinés. Le vaccin de l'étude s'est avéré significativement efficace pour protéger les porcs vaccinés contre les symptômes cliniques, la charge virale sanguine et la mortalité, ainsi que pour améliorer la productivité, par rapport aux porcs non vaccinés.(Traduit par Docteur Serge Messier).

A nanobody-horseradish peroxidase fusion protein-based competitive ELISA for rapid detection of antibodies against porcine circovirus type 2.

BACKGROUND: The widespread popularity of porcine circovirus type 2(PCV2) has seriously affected the healthy development of the pig industry and caused huge economic losses worldwide. A rapid and reliable method is required for epidemiological investigation and evaluating the effect of immunization. However, the current methods for PCV2 antibody detection are time-consuming or very expensive and rarely meet the requirements for clinical application. we have constructed the platform for expressing the nanobody(Nb)­horseradish peroxidase(HRP) fusion protein as an ultrasensitive probe to detect antibodies against the Newcastle disease virus(NDV), previously. In the present work, an Nb-HRP fusion protein-based competitive ELISA(cELISA) for rapid and simple detection antibodies against PCV2 was developed using this platform to detect anti-PCV2 antibodies in clinical porcine serum. RESULTS: Using phage display technology, 19 anti-PCV2-Cap protein nanobodies were screened from a PCV2-Cap protein immunized Bactrian camel. With the platform, the PCV2-Nb15­HRP fusion protein was then produced and used as a sensitive reagent for developing a cELISA to detect anti­PCV2 antibodies. The cut­off value of the cELISA is 20.72 %. Three hundreds and sixty porcine serum samples were tested by both newly developed cELISA and commercial kits. The sensitivity and specificity were 99.68 % and 95.92 %, respectively. The coincidence rate of the two methods was 99.17 %. When detecting 620 clinical porcine serum samples, a good consistent (kappa value = 0.954) was found between the results of the cELISA and those of commercial kits. CONCLUSIONS: In brief, the newly developed cELISA based PCV2-Nb15­HRP fusion protein is a rapid, low-cost, reliable and useful nanobody-based tool for the serological evaluation of current PCV2 vaccine efficacy and the indirect diagnosis of PCV2 infection.

Establishment of a Rep' protein antibody detection method to distinguish natural infection with PCV2 from subunit vaccine immunization.

Introduction. PCV2 is a DNA virus that exists widely in pigs and has caused great economic losses to the pig industry worldwide. In the existing commercial PCV2 enzyme-linked immunosorbent assay (ELISA) kits both natural infection with PCV2 and vaccine immunization produce results that are positive for PCV2 Cap antibodies and therefore they cannot diagnose PCV2 infection in immunized pig farms.Aim. To establish a PCV2 non-structural protein antibody detection method that distinguishes between antibodies resulting from natural prior exposure (infection) and those induced by subunit vaccine immunization.Methodology. Based on the non-structural Rep' protein, we established an indirect ELISA (iELISA) using sera from guinea pigs and piglets.Results. The results for iELISA for guinea pig serum showed that animals vaccinated with a whole-virus inactivated PCV2 vaccine had 100 % (10/10) Cap antibody positivity and 100 % (10/10) Rep' antibody positivity. Guinea pigs vaccinated with a recombinant subunit PCV2 vaccine had 100 % (10/10) Cap antibody positivity, while no (0/10) guinea pigs were Rep' antibody-positive. The combined detection results for the Rep' iELISA and a PCV2 Antibody Test kit (Commercial) showed that pigs vaccinated with a whole-virus inactivated PCV2 vaccine or PCV2 SD/2017 had 100 % (5/5) Cap antibody positivity and 100 % (5/5) Rep' antibody positivity. Pigs vaccinated with a recombinant subunit PCV2 vaccine had 100 % (5/5) Cap antibody positivity, while no (0/10) pigs were Rep' antibody-positive.Conclusion. This paper describes an effective iELISA method that can distinguish natural infection with PCV2 (Cap and Rep positive) or inoculation with a whole-virus inactivated vaccine (Cap and Rep positive) from subunit vaccine immunization (Cap-positive, Rep-negative). These comparative assays could be very useful in the control of PCV2 in pig herds.

A molecular survey and phylogenetic analysis of porcine circovirus type 3 using oral fluid, faeces and serum.

BACKGROUND: Porcine circovirus type 3 is the most recently discovered porcine circovirus, and an emerging pathogen. In this study the status of its presence on some Slovenian farms is reported. The effectiveness of the vaccine against porcine circovirus type 2 was assessed against porcine circovirus type 3. Group samples of oral fluid, faeces and individual serum samples were taken from six different pig categories and tested for presence of viral DNA, using both real time and conventional PCR. Positive samples were subjected to direct Sanger sequencing. Nucleotide sequences were analyzed and compared to GenBank PCV3 sequences. RESULTS: Positive samples were sent for genome sequencing, which confirmed the presence of virus in all different pig categories on five farms. A high to moderate correlation of strong statistical significance was found between individual serum samples, oral fluid and faeces. Slovenian PCV3 was found to be distributed in a way similar to that of other countries. Slovenian PCV3 nt sequences are highly related, sharing more than 99.5% nt identity. On one farm a commercially available vaccine against porcine circovirus type 2 was used on 3-week-old pigs. It did not affect the presence of porcine circovirus type 3 in oral fluid or sera of any of the seven age groups of pigs, each with two control groups. CONCLUSIONS: The results constitute the first discovery of the virus in Slovenia. Genome sequencing has revealed a high degree of similarity between Slovenian and GenBank isolates.

Fine mapping of linear B cell epitopes on capsid protein of porcine circovirus 3.

Porcine circovirus type 3 (PCV3) is an emerging swine pathogen associated with acute porcine dermatitis and nephropathy syndrome (PDNS)-like clinical signs, reproductive failure, and multisystemic inflammation. Current evidence shows that PCV3 is spread worldwide, and its high incidence may pose a threat to the global pig industry. Capsid (Cap) protein is the sole structural protein which plays an important role in inducing protective immunity against PCV3 infection. In this study, monoclonal antibodies (mAbs) against Cap protein of PCV3 were produced by the hybridoma technique. Subsequently, 12 serial overlapping peptides (P1 to P12) spanning the entire region of Cap were synthesized to determine the B cell epitope regions using the mAbs. Results from dot-blot and peptide ELISA identified that P3, P9, and P10 were the major B cell antigenic regions. Fine mapping by shorter N- and C-terminal truncated peptides confirmed that the motifs 57NKPWH61, 140KHSRYFT146, and 161QSLFFF166 were linear B cell epitopes, which were highly conserved among different PCV3 strains. Interestingly, we found that the motif 140KHSRYFT146 was highly conserved in all reported types of PCVs (i.e., PCV1, PCV2, PCV3, and PCV4), except for the substitution (Y → K → R) of the first residue. This is the first research to identify B cell epitopes of PCV3 Cap, and these findings may lead to a better understanding of the antibody-antigen interaction and provide some guidance for PCV3 vaccine design.Key points• The recombinant Cap protein of PCV3 was expressed and purified in soluble form. • PCV3 Cap-specific mAbs prepared in this study had no cross-reactivity with PCV1/PCV2 Cap. • This is the first report of three conserved linear B cell epitopes on PCV3 Cap. • The minimal residues of the epitopes were 57-61 aa, 140-146 aa, and 161-166 aa.

Haemato-biochemical alterations and oxidative stress associated with naturally occurring porcine circovirus2 infection in pigs.

Porcine circovirus2 (PCV2) infection in pigs is one of the major causes of economic loss to the farmers in terms of low production, slow growth and increase post-weaning mortality rate. The effect of PCV2 infection on haemogram, serum biochemical profile and oxidant/anti-oxidant status is not well established in pigs. In the present study, haemogram, serum biochemical profile and oxidant/anti-oxidant status were assessed in pigs confirmed positive for PCV2 infections as evidenced by commercially available enzyme-linked immunosorbent assay kit (n = 151) and polymerase chain reaction (PCR) (n = 42) among a total of 306 number of pigs included in the study. Non-infected healthy pigs (n = 6) served as healthy control. The total erythrocyte count (TEC), haemoglobin (Hb), packed cell volume (PCV), total leukocyte count (TLC), differential leukocyte count (DLC) and thrombocyte count were measured. The levels of total protein, albumin, globulin, total bilirubin, direct bilirubin, blood urea nitrogen (BUN), creatinine and glucose and enzymes viz. alanine transaminase (ALT), aspartate transaminase (AST), gamma-glutamyl transferase (GGT) and alkaline phosphatase (ALP) were measured. Oxidative stress indicators such as plasma malondialdehyde (MDA) and total anti-oxidant activity (TAOA) were measured using commercially available kits. The mean values of TLC, lymphocytes and thrombocyte count were significantly (P < 0.05) low in PCV2-infected pigs. The levels of globulin, AST, GGT, BUN and creatinine were significantly increased (P < 0.05) whereas levels of albumin and glucose significantly (P < 0.05) decreased in PCV2-infected pigs. The significant increase (P < 0.05) in MDA level and significant decrease (P < 0.05) in TAOA level were noticed in PCV2-infected animals as compared with healthy control. The present study supports immunosuppression, possible multiple organ damage and oxidative stress associated with naturally occurring PCV2 infection in pigs. Timely vaccination and managemental practices can reduce PCV2 infection in farms. In spite of many research studies, there is still paucity of detailed systemic study on haemato-biochemical alteration and oxidative stress associated with PCV2 infection.

Do porcine parvoviruses 1 through 7 (PPV1-PPV7) have an impact on porcine circovirus type 2 (PCV2) viremia in pigs?

Infections with porcine parvoviruses 1 through 7 (PPV1-PPV7) and porcine circovirus type 2 (PCV2) are widespread in pig population. PCV2 is involved in a number of disease syndromes collectively called PCV2-associated diseases (PCVD). It is well elucidated, that PPV1 may act as a triggering factor of PCVD through supporting PCV2 replication. Less is known about the PPV2-PPV7 impact on PCV2 viremia, but several authors suggested an association between these viruses. In order to provide a better understanding of PCV2 and PPVs co-infections, 519 serum samples from eight Polish swine farms were tested by real-time PCR to assess the possible impact of PPV1-PPV7 on PCV2 viremia. Among all 519 serum samples, 30.6 % were positive for PCV2 and PPVs detection rates ranged from 2.9 % (PPV1) to 26.6 % (PPV2). Within 159 serum samples categorized as PCV2-positive, the prevalence rates of PPVs ranged from 7.5 % (PPV1) to 37.1 % (PPV6). The level of PCV2 viremia was significantly higher only in serum samples positive for PPV1 and PPV7 compared to samples negative for these PPVs. Moreover, the correlation between Ct values for PPV7 and PCV2 was observed. Thus, our results suggested that apart from PPV1, also PPV7 stimulate the replication of PCV2.

Porcine circovirus 3 is highly prevalent in serum and tissues and may persistently infect wild boar (Sus scrofa scrofa).

Porcine circovirus 3 (PCV-3) prevalence has been minimally investigated in wild boar; dynamics of infection and viral tissue distribution are currently unknown. In this study, serum samples from 518 wild boar (from years 2004 to 2018) were used to study frequency of infection. Also, serum samples from 19 boar captured and recaptured at least two times for a period of time from 1 month to 1 year were collected to determine PCV-3 infection dynamics. Finally, to elucidate PCV-3 DNA organic distribution, sera, different tissues and faeces were obtained from 35 additional wild boar. PCV-3 DNA was extracted and amplified with a conventional PCR. For the PCV-3 PCR-positive sera from the longitudinally sampled and different tissue types, a quantitative PCR was performed. Genome sequence was obtained from a number of PCV-3 PCR-positive samples from different years, different time-points of infection and tissues. Obtained results confirmed the susceptibility of wild boar to the virus, showing high frequency of PCV-3 detection (221 out of 518, 42.66%) and demonstrating circulation at least since 2004. Compiled data indicate the possibility of long-term infections, since 5 out of 10 PCV-3 PCR-positive boars longitudinally sampled showed positivity in samplings separated for more than 5 months. All tested tissue types' harboured PCV-3 genome, with the highest percentage of PCR positivity in submandibular lymph node, tonsil, lung, liver, spleen and kidney. The amount of DNA in all tested PCV-3 PCR-positive samples was moderate to low. All partial and complete PCV-3 sequences obtained from wild boar displayed high nucleotide identity, higher than 98%. In conclusion, this study further confirms that wild boar is susceptible to PCV-3 infection, showing high frequency of detection in this animal species. Furthermore, PCV-3 can be found in different tissues of wild boar and is apparently able to cause persistent infection.

High prevalence of cyclovirus Vietnam (CyCV-VN) in plasma samples from Madagascan healthy blood donors.

Cycloviruses, small ssDNA viruses belonging to the Circoviridae family, have been suggested as possible causes of enteric, respiratory and neurological disorders in human patients. One of these species, cyclovirus-Vietnam (CyCV-VN), initially isolated from cerebrospinal fluid samples of patients with unexplained neurological disorders, has since been reported in serum samples from chronically patients infected with HBV, HCV or HIV, in Italy. On the other hand, CyCV-VN was not detected in serum samples from healthy individuals. Here, we report on a high prevalence of 43.4% (40/92) of CyCV-VN in plasma samples from asymptomatic blood donors from Madagascar. Interestingly, this virus was not detected by metagenomics and PCR in six other African countries, suggesting regional differences in CyCV-VN prevalence across Africa. Phylogenetic analysis based on the complete genomes showed that CyCV-VN sequences isolated from blood were most closely related to sequences previously reported from human stool in Madagascar. Further investigations using larger cohorts are required to determine the global epidemiology, the natural history and the pathological significance, if any, of CyCV-VN infection in humans.

Porcine circovirus type 3 (PCV3) shedding in sow colostrum.

The major objective of this work was to investigate the shedding of porcine circovirus type 3 (PCV3) in sow colostrum. PCV3 titers in the serum and colostrum samples of 38 sows were determined using qPCR. Interestingly, this is the first report regarding the identification of PCV3 from the colostrum samples. In the studied farm, the prevalence of PCV3 in the colostrum samples was 44.74% (17/38). When sows were grouped based on the PCV3 titers in the serum into the "High-viremic", "Low-viremic" and "Non-viremic" sows, it was shown that the High-viremic sows showed significantly higher PCV3 colostrum prevalence (100%; 9/9) with the PCV3 titers ranging from 4.01 to 7.33 genomic copies/mL. The results indicated that PCV3 in the colostrum might be partly influenced by the viremic stage of the infection. However, the results also showed that approximately 41% of sows shedding PCV3 with low titers in the colostrum (7/17) were non-viremic sows. In conclusion, this study identified the presence of PCV3 in sow colostrum. Clinical impacts and mechanisms of colostrum shedding of PCV3 should be further investigated.

Establishment and application of an indirect ELISA for porcine circovirus 3.

Porcine circovirus type 3 (PCV3) was initially reported in 2016 in the United States of America. Since then, the virus has been detected on swine farms in Poland, South Korea, and China using PCR. However, a serological survey of PCV3 in pig populations has never been conducted. In this study, for the first time, we established an indirect enzyme-linked immunosorbent (ELISA) assay and performed a national retrospective serological survey for PCV3. Our results showed that the PCV3-postive rate increased from 22.35% to 51.88% between 2015 and 2017. The above results suggest PCV3 has spread widely in China with increased positive rates since 2015.

The Hsp90 inhibitor 17-DMAG decreases infection of porcine circovirus type 2 in mice.

Although several factors affecting porcine circovirus type 2 (PCV2) infection have been reported, their precise roles are far from clear. The aim of this study was to determine whether 17-dimethylaminoethylamino-17-demethoxygeldanamycin (17-DMAG), an inhibitor of Hsp90, could significantly affect PCV2 infection and immune responses in BALB/c mice. Intraperitoneal injection of 17-DMAG significantly reduced viral loads in the blood and tissues of mice infected with PCV2, compared with control groups. The 17-DMAG treatment decreased serum interleukin (IL)-10 and tumor necrosis factor(TNF)-α levels, but it did not have a significant effect on the IL-1ß level. These data demonstrate that 17-DMAG is highly effective in suppressing PCV2 replication in BALB/c mice, indicating that it has potential value as an antiviral drug against PCV2 infection.

Establishment and application of a competitive enzyme-linked immunosorbent assay differentiating PCV2 antibodies from mixture of PCV1/PCV2 antibodies in pig sera.

BACKGROUND: Porcine cirovirus type 1 (PCV1) and type 2 (PCV2) are circulating in Chinese pig herds and the infected pigs develop antibodies to both viruses. Current commercial available ELISA kits cannot differentiate PCV2-specific antibodies from the mixtures of PCV1 and PCV2 antibodies in PCV1/2-infected or PCV2-vaccinated pigs. Therefore, the need for developing PCV2-specific ELISA methods is urgent to evaluate PCV2 antibody level in exclusion of PCV1 antibody interference after PCV2 vaccination. RESULTS: Virus-like particles (VLPs) of PCV2 based on the recombinant Cap protein were expressed in Escherichia coli. A competing ELISA was established by using the VLPs as coating antigen and a PCV2-specific monoclonal antibody as the competing antibody. The competing ELISA was compared with the results obtained by using an immunoperoxidase monolayer assay on 160 serum samples. The sensitivity and specificity of this competing ELISA were determined as 96.5 and 96.0 %, at 2 standard deviation from the mean or 91.8 and 100 % at 3 standard deviations from the mean. Next, a serological survey of 1297 vaccinated serum samples collected from commercial pig herds in Beijing, Hunan and Henan provinces in China was conducted. The results showed that 85.9 % of sera having positive PCV2 antibodies. CONCLUSIONS: The competing ELISA we developed in this study was both sensitive and specific to PCV2 and was suitable for large-scale PCV2 antibody monitoring in exclusion of PCV1 antibody interference after PCV2 vaccination.

Genomic Analysis of the Chicken Infectious Anemia Virus in a Specific Pathogen-Free Chicken Population in China.

The antibody to chicken infectious anemia virus (CIAV) was positive in a specific pathogen-free (SPF) chicken population by ELISA test in our previous inspection, indicating a possible infection with CIAV. In this study, blood samples collected from the SPF chickens were used to isolate CIAV by inoculating into MSB1 cells and PCR amplification. A CIAV strain (SD1403) was isolated and successfully identified. Three overlapping genomic fragments were obtained by PCR amplification and sequencing. The full genome sequence of the SD1403 strain was obtained by aligning the sequences. The genome of the SD1403 strain was 2293 bp with a nucleotide identity of 94.8% to 98.5% when compared with 30 referred CIAV strains. The viral proteins VP2 and VP3 were highly conserved, but VP1 was not relatively conserved. Both amino acids 139 and 144 of VP1 were glutamine, which was in accord with the low pathogenic characteristics. In this study, we first reported that CIAV exists in Chinese SPF chicken populations and may be an important reason why attenuated vaccine can be contaminated with CIAV.

Cyclovirus Vietnam DNA in immunodeficient patients.

BACKGROUND: Cyclovirus Vietnam (CyCV-VN) is a CyCV detected in 2013 from cerebrospinal fluid (CSF) samples of patients with neurological disorders. Information on prevalence, pathogenesis and disease association of CyCV-VN is still very patchy. OBJECTIVES AND STUDY DESIGN: In this study, we have used a PCR assay targeting the Rep gene to investigate the prevalence of CyCV-VN infection in blood and CSF samples of 346 Italian subjects. RESULTS: Overall, 7% of blood samples were positive for CyCV-VN while the virus was not detected in any of the CSF samples. The prevalence of CyCV-VN was relatively high in HIV positive patients (21%), modest in patients with HBV or HCV infection (6%), and low in transplant recipient patients (2%). Positive patients showed low levels of CyCV-VN viremia. The virus was not detected in serum samples from healthy individuals. Longitudinal analysis of serum samples obtained from selected patients showed a stable or transient presence of circulating CyCV-VN. CONCLUSIONS: The present study is the first to demonstrate CyCV-VN DNA circulation in Italy and to cast light on some biological aspects of this novel virus of men.

RNA-Seq Analysis Reveals Genes Underlying Different Disease Responses to Porcine Circovirus Type 2 in Pigs.

Porcine circovirus type 2 (PCV2), an economically important pathogen, causes postweaning multisystemic wasting syndrome (PMWS) and other syndrome diseases collectively known as porcine circovirus-associated disease (PCVAD). Previous studies revealed breed-dependent differences in porcine susceptibility to PCV2; however, the genetic mechanism underlying different resistance to PCV2 infection remains largely unknown. In this study, we found that Yorkshire × Landrace (YL) pigs exhibited serious clinical features typifying PCV2 disease, while the Laiwu (a Chinese indigenous pig breed, LW) pigs showed little clinical symptoms of the disease during PCV2 infection. At 35 days post infection (dpi), the PCV2 DNA copy in YL pigs was significantly higher than that in LW pigs (P < 0.05). The serum level of IL-4, IL-6, IL-8, IL-12 and TGF-ß1 in LW pigs and TNF-α in YL pigs increased significantly at the early infected stages, respectively; while that of IL-10 and IFN-γ in YL pigs was greatly increased at 35 dpi. RNA-seq analysis revealed that, at 35 dpi, 83 genes were up-regulated and 86 genes were down-regulated in the lung tissues of LW pigs, while in YL pigs, the numbers were 187 and 18, respectively. In LW pigs, the differentially expressed genes (DEGs) were mainly involved in complement and coagulation cascades, metabolism of xenobiotics by cytochrome P450, RIG-I-like receptor signaling and B cell receptor signaling pathways. Four up-regulated genes (TFPI, SERPNC1, SERPNA1, and SERPNA5) that are enriched in complement and coagulation cascades pathway were identified in the PCV2-infected LW pigs, among which the mRNA expression of SERPNA1, as well as three genes including TGF-ß1, TGF-ß2 and VEGF that are regulated by SERPNA1 was significantly increased (P < 0.05). We speculate that higher expression of SERPNA1 may effectively suppress excessive inflammation reaction and reduce the pathological degree of lung tissue in PCV2-infected pigs. Collectively, our findings indicate that the susceptibility to PCV2 infection depends on a genetic difference between LW and YL pigs, and SERPNA1 likely plays an important role in the resistance of LW pigs to PCV2 infection.

Serological Survey of Porcine circovirus-2 in Captive Wild Boars (Sus scrofa) from Registered Farms of South and South-east Regions of Brazil.

This study aimed to survey captive wild boars for antibodies against Porcine circovirus-2 (PCV-2) in registered farms. Serum samples (n = 1305) were collected from 90-day-old wild boars from 118 farms of the Brazilian South-east region, including the states of Minas Gerais and São Paulo, and South region, including the states of Paraná, Rio Grande do Sul and Santa Catarina. All herds (100%) presented reactive animals, in varying numbers and from low-to-high antibody titres, with the occurrence ranging from 82 to 89%. Considering farms, the average prevalence was of 84.9% (P < 0.05) and ranged from 54.1 to 94.95%. Regarding the geographic regions studied, the prevalence was of 100%, with PCV2 antibodies detected in wild boars of all regions. This study provides the first evidence of PCV2 antibodies in captive wild boars in Brazil.

Application of a pathogen microarray for the analysis of viruses and bacteria in clinical diagnostic samples from pigs.

Many of the disease syndromes challenging the commercial swine industry involve the analysis of complex problems caused by polymicrobial, emerging or reemerging, and transboundary pathogens. This study investigated the utility of the Lawrence Livermore Microbial Detection Array (Lawrence Livermore National Laboratory, Livermore, California), designed to detect 8,101 species of microbes, in the evaluation of known and unknown microbes in serum, oral fluid, and tonsil from pigs experimentally coinfected with Porcine reproductive and respiratory syndrome virus (PRRSV) and Porcine circovirus-2 (PCV-2). The array easily identified PRRSV and PCV-2, but at decreased sensitivities compared to standard polymerase chain reaction detection methods. The oral fluid sample was the most informative, possessing additional signatures for several swine-associated bacteria, including Streptococcus sp., Clostridium sp., and Staphylococcus sp.