scholarly journals GP5 Protein-based ELISA for the Detection of PRRSV Antibodies

2016 ◽  
Vol 19 (3) ◽  
pp. 495-501 ◽  
Author(s):  
Y. Wang ◽  
J. Guo ◽  
S. Qiao ◽  
Q. Li ◽  
J. Yang ◽  
...  
Keyword(s):  
Cell Culture ◽  
Western Blot ◽  
Neutralizing Antibodies ◽  
Economic Losses ◽  
Respiratory Syndrome Virus ◽  
Flask Culture ◽  
E Coli ◽  
Swine Pathogen ◽  
And Control ◽  
Glycoprotein 5

AbstractPorcine reproductive and respiratory syndrome virus (PRRSV) is an important swine pathogen, causing huge economic losses each year worldwide. Immunization with vaccines containing the glycoprotein 5 (GP5) of PRRSV is the main measure to induce neutralizing antibodies and control the disease. Here, we developed a GP5 protein-based ELISA for detecting antibodies against PRRSV. The overall yield of purified GP5 inE. coliflask culture was more than 45 mg/L cell culture. Western blot and IFA indicated that the GP5 protein was highly immunogenic. After optimization and validation with IDEXX PRRS using 566 clinical sera, the DSN, DSP, and accuracy of GP5-ELISA were 81.39%, 75.96%, and 80.39%, respectively. Besides, GP5-ELISA is highly specific, showing no cross-reactions with sera against other important swine pathogens. Hence, GP5 is a good diagnostic antigen and the GP5 protein-based ELISA has the potential to be used in the field.

A Porcine Reproductive and Respiratory Syndrome Virus Vaccine Candidate Based on PRRSV Glycoprotein 5 and the Toll-Like Receptor 5 Agonist Salmonella typhimurium Flagellin

2015 ◽  
Vol 25 (1) ◽  
pp. 56-59 ◽  
Author(s):  
Hongqin Song ◽  
Dan Xiong ◽  
Jing Wang ◽  
Xianyue Zhai ◽  
Guangcheng Liang
Keyword(s):  
Salmonella Typhimurium ◽  
Neutralizing Antibodies ◽  
Expression Vectors ◽  
Respiratory Syndrome Virus ◽  
Toll Like Receptor ◽  
Pcr Method ◽  
Overlap Pcr ◽  
Toll Like Receptor 5 ◽  
Cellular Immune ◽  
Glycoprotein 5

Glycoprotein 5 (GP5) from porcine reproductive and respiratory syndrome virus (PRRSV) is a key inducer of neutralizing antibodies. A truncated GP5 gene lacking the signal peptide and transmembrane sequences was amplified via an overlap PCR method and inserted into prokaryotic expression vectors, pET32a or pGEX-6p-1, to add an His or GST tag, respectively. His-tagged GP5 was induced with IPTG, verified by SDS-PAGE and Western blotting, and purified to serve as an immunogen accompanied with the <i>Salmonella typhimurium</i> flagellin (FliC), a Toll-like receptor 5 (TLR5) agonist. Levels of TLR5 and cytokine mRNAs in spleens of mice following injection with FliC were detected by qRT-PCR to verify the activation of innate immunity. FliC was used as an adjuvant and administered with the GP5 to C57BL/6 mice via intraperitoneal injection. Coadministration of GP5 with FliC induced a significantly enhanced GP5-specific IgG and IFN-&#947; response compared with administration of GP5 alone, and the GP5-specific titer in the GP5 + FliC coadministration group was elevated almost twofold after the third immunization. These results indicate that FliC is an effective adjuvant, increasing the induction of antibodies against GP5 with the induction of both humoral and cellular immune responses.

scholarly journals Indirect ELISA for diagnosis of Brucella ovis infection in rams

2014 ◽  
Vol 66 (6) ◽  
pp. 1695-1702 ◽  
Author(s):  
S.A. França ◽  
J.P.S. Mol ◽  
E.A. Costa ◽  
A.P.C. Silva ◽  
M.N. Xavier ◽  
...  
Keyword(s):  
Western Blot ◽  
Recombinant Proteins ◽  
Indirect Elisa ◽  
High Sensitivity ◽  
Economic Losses ◽  
Well Testing ◽  
E Coli ◽  
Sheep Industry ◽  
Brucella Ovis ◽  
Sexually Mature

Brucella ovis is a major cause of epididymitis in sexually mature rams, resulting in subfertility, infertility, and economic losses for the sheep industry worldwide. The aim of this study was to develop an indirect ELISA (iELISA) using recombinant proteins, namely rBoP59 and rBP26, as antigens for serological diagnosis of B. ovisinfection. The BoP59 and BP26 recombinant proteins were expressed in E. coli and purified by affinity chromatography. Antigenicity was tested by Western blot and iELISA. Standardization of iELISA was performed with 500ng and 1µg BoP59 and rBP26 per well, testing serum from uninfected and experimentally infected rams. rBP26 was effective in distinguishing positive from negative rams. The rBP26 iELISA developed in this study is the first to use a completely purified rBP26 as antigen resulting in high sensitivity (100%) and specificity (90.2%), and an overall accuracy equal to 1.0.

scholarly journals Xanthohumol inhibits PRRSV proliferation and alleviates oxidative stress induced by PRRSV via the Nrf2–HMOX1 axis

2019 ◽  
Vol 50 (1) ◽  
Author(s):  
Xuewei Liu ◽  
Zhongbao Song ◽  
Juan Bai ◽  
Hans Nauwynck ◽  
Yongxiang Zhao ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Therapeutic Strategy ◽  
Antioxidant Response ◽  
Economic Losses ◽  
Respiratory Syndrome Virus ◽  
Related Factor ◽  
Nuclear Factor ◽  
Swine Industry ◽  
Therapeutic Drugs ◽  
Swine Pathogen

Abstract Porcine reproductive and respiratory syndrome virus (PRRSV) is a prevalent and endemic swine pathogen that causes significant economic losses in the global swine industry. Commercial vaccines provide limited protection against this virus, and no highly effective therapeutic drugs are yet available. In this study, we first screened a library of 386 natural products and found that xanthohumol (Xn), a prenylated flavonoid found in hops, displayed high anti-PRRSV activity by inhibiting PRRSV adsorption onto and internalization into cells. Transcriptome sequencing revealed that Xn treatment stimulates genes associated with the antioxidant response in the nuclear factor-erythroid 2-related factor 2 (Nrf2) signalling pathway. Xn causes increased expression of Nrf2, HMOX1, GCLC, GCLM, and NQO1 in Marc-145 cells. The action of Xn against PRRSV proliferation depends on Nrf2 in Marc-145 cells and porcine alveolar macrophages (PAMs). This finding suggests that Xn significantly inhibits PRRSV proliferation and decreases viral-induced oxidative stress by activating the Nrf2–HMOX1 pathway. This information should be helpful for developing a novel prophylactic and therapeutic strategy against PRRSV infection.

scholarly journals Efficacy of a live attenuated highly pathogenic PRRSV vaccine against a NADC30-like strain challenge: implications for ADE of PRRSV

2021 ◽  
Vol 17 (1) ◽  
Author(s):  
Xin-xin Chen ◽  
Xinyu Zhou ◽  
Tengda Guo ◽  
Songlin Qiao ◽  
Zhenhua Guo ◽  
...  
Keyword(s):  
Neutralizing Antibodies ◽  
Molecular Mechanisms ◽  
Clinical Signs ◽  
Bodyweight Gain ◽  
Economic Losses ◽  
Respiratory Syndrome Virus ◽  
Experimental Conditions ◽  
Highly Pathogenic ◽  
Highly Pathogenic Prrsv

Abstract Background Porcine reproductive and respiratory syndrome virus (PRRSV) infection can cause severe reproductive failure in sows and respiratory distress in pigs of all ages, leading to major economic losses. To date, there are still no effective strategies to prevent and control PRRSV. Antibody-dependent enhancement (ADE), a phenomenon in which preexisting non-neutralizing antibodies or sub-neutralizing antibodies facilitate virus entry and replication, may be a significant obstacle in the development of effective vaccines for many viruses, including PRRSV. However, the contribution of ADE to PRRSV infection remains controversial, especially in vivo. Whether attenuated PRRSV vaccines prevent or worsen subsequent disease in pigs infected by novel PRRSV strains requires more research. In the present study, in vivo experiments were conducted to evaluate ADE under different immune statuses, which were produced by waiting different lengths of time after vaccination with a commercially available attenuated highly pathogenic PRRSV (HP-PRRSV) vaccine (JXA1-R) before challenging the pigs with a novel heterologous NADC30-like strain. Results Piglets that were vaccinated before being challenged with PRRSV exhibited lower mortality rates, lower body temperatures, higher bodyweight gain, and lower viremia. These results demonstrate that vaccination with JXA1-R alleviated the clinical signs of PRRSV infection in all vaccinated groups. Conclusions The obtained data indicate that the attenuated vaccine test here provided partial protection against the NADC30-like strain HNhx. No signs of enhanced PRRSV infection were observed under the applied experimental conditions. Our results provide some insight into the molecular mechanisms underlying vaccine-induced protection or enhancement in PRRSV.

scholarly journals Virulence-Associated Genes and Genetic Diversity of Avian Pathogenic (APEC) and Fecal (AFEC) E. Coli Isolates from Chickens

2021 ◽  
Author(s):  
Zhen Zhu ◽  
Mingze Cao ◽  
Weiwei Wang ◽  
Qiqi Zhu ◽  
Chengye Wang ◽  
...  
Keyword(s):  
Genetic Background ◽  
Prevention And Control ◽  
Virulence Genes ◽  
Etiologic Agent ◽  
The Other ◽  
Economic Losses ◽  
Molecular Characteristics ◽  
Phylogenetic Groups ◽  
E Coli ◽  
And Control

Abstract Background: Avian pathogenic E. coli (APEC) is the etiologic agent of serious colibacillosis and causes extensive economic losses. To examine the genetic background of APEC, we characterized the serotypes, virulence genes, phylogenetic classification and MLST of 392 APEC and 586 AFEC strains isolated from infected chickens. Results: The results showed that the most predominant serotypes were O78 (13.47%), O2 (9.16%), O18 (5.39%), O20 (4.42%) and O25 (4.09%). The major serotypes O78 (13.47%) and O2 (9.16%) were significantly higher in the APEC isolates than in the AFEC isolates. Among the 16 analyzed virulence-associated genes (VAGs), iroN (100%), ompT (100%), fimC (92.46%), iss (77.91%) and irp2 (71.98%) were the most frequently identified. Over half (54.85%) of the strains possessed > 8–13 VAGs, and 85.23% of the strains carried iroN-ompT-fimC-iss/irp2 VAG patterns. According to the phylogenetic analysis, phylogroups A (32.11%) and B2 (31.36%) proved to be the most prevalent phylogenetic groups in the AFEC and APEC isolates, respectively. The strains that belonged to phylogroup B2 were associated with more VAGs. Based on MLST, 46 STs belonging to 15 different clonal complexes were identified, and 4 were novel. ST88 (10.67%) was found to be the most dominant ST, and it possessed at least 9 VAGs and belonged to phylogroups B2 or D. Furthermore, the isolates belonging to B2-O78/O2-ST88 were the most likely APEC isolates to be associated with epidemics, and they carried more VAGs than the other strains. Conclusions: Our findings have enriched our knowledge of the molecular characteristics of APEC isolates from chickens, which will be important for the prevention and control of avian colibacillosis.

scholarly journals A DNA Prime Immuno-Potentiates a Modified Live Vaccine against the Porcine Reproductive and Respiratory Syndrome Virus but Does Not Improve Heterologous Protection

Viruses ◽  
2019 ◽  
Vol 11 (6) ◽  
pp. 576 ◽  
Author(s):  
Cindy Bernelin-Cottet ◽  
Céline Urien ◽  
Maxence Fretaud ◽  
Christelle Langevin ◽  
Ivan Trus ◽  
...  
Keyword(s):  
T Cell ◽  
Antibody Response ◽  
Cell Response ◽  
Neutralizing Antibodies ◽  
T Cell Response ◽  
Economic Losses ◽  
Respiratory Syndrome Virus ◽  
T Cell Epitopes ◽  
Immune Correlates ◽  
Heterologous Protection

The porcine reproductive and respiratory syndrome virus (PRRSV), an RNA virus inducing abortion in sows and respiratory disease in young pigs, is a leading infectious cause of economic losses in the swine industry. Modified live vaccines (MLVs) help in controlling the disease, but their efficacy is often compromised by the high genetic diversity of circulating viruses, leading to vaccine escape variants in the field. In this study, we hypothesized that a DNA prime with naked plasmids encoding PRRSV antigens containing conserved T-cell epitopes may improve the protection of MLV against a heterologous challenge. Plasmids were delivered with surface electroporation or needle-free jet injection and European strain-derived PRRSV antigens were targeted or not to the dendritic cell receptor XCR1. Compared to MLV-alone, the DNA-MLV prime- boost regimen slightly improved the IFNγ T-cell response, and substantially increased the antibody response against envelope motives and the nucleoprotein N. The XCR1-targeting of N significantly improved the anti-N specific antibody response. Despite this immuno-potentiation, the DNA-MLV regimen did not further decrease the serum viral load or the nasal viral shedding of the challenge strain over MLV-alone. Finally, the heterologous protection, achieved in absence of detectable effective neutralizing antibodies, was not correlated to the measured antibody or to the IFNγ T-cell response. Therefore, immune correlates of protection remain to be identified and represent an important gap of knowledge in PRRSV vaccinology. This study importantly shows that a naked DNA prime immuno-potentiates an MLV, more on the B than on the IFNγ T-cell response side, and has to be further improved to reach cross-protection.

scholarly journals Antagonizing Interferon-Mediated Immune Response by Porcine Reproductive and Respiratory Syndrome Virus

2014 ◽  
Vol 2014 ◽  
pp. 1-9 ◽  
Author(s):  
Rong Wang ◽  
Yan-Jin Zhang
Keyword(s):  
Immune Response ◽  
Neutralizing Antibodies ◽  
Innate Immune ◽  
Economic Losses ◽  
Respiratory Syndrome Virus ◽  
First Line ◽  
Swine Industry ◽  
Antiviral Signaling ◽  
Cell Mediated Immune Response ◽  
Host Innate Immune Response

Interferons (IFNs) are important components in innate immunity involved in the first line of defense to protect host against viral infection. Porcine reproductive and respiratory syndrome virus (PRRSV) leads to severe economic losses for swine industry since being first identified in early 1990s. PRRSV interplays with host IFN production and IFN-activated signaling, which may contribute to the delayed onset and low level of neutralizing antibodies, as well as weak cell-mediated immune response in infected pigs. PRRSV encodes several proteins that act as antagonists for the IFN signaling. In this review, we summarized the various strategies used by PRRSV to antagonize IFN production and thwart IFN-activated antiviral signaling, as well as the variable interference with IFN-mediated immune response by different PRRSV strains. Thorough understanding of the interaction between PRRSV and host innate immune response will facilitate elucidation of PRRSV pathogenesis and development of a better strategy to control PRRS.

The effect of type of water supply on water quality in a developing community in South Africa

1997 ◽  
Vol 35 (11-12) ◽  
pp. 35-40 ◽  
Author(s):  
B. Genthe ◽  
N. Strauss ◽  
J. Seager ◽  
C. Vundule ◽  
F. Maforah ◽  
...  
Keyword(s):  
Water Quality ◽  
Water Supply ◽  
Cross Sectional Study ◽  
Cross Sectional ◽  
E Coli ◽  
Guideline Values ◽  
Developing Communities ◽  
Quality Guideline ◽  
And Control ◽  
The Relationship

Efforts to provide water to developing communities in South Africa have resulted in various types of water supplies being used. This study examined the relationship between the type of water supply and the quality of water used. Source (communal taps, private outdoor and indoor taps) and point-of-use water samples were examined for heterotrophic plate counts (HPC), total and faecal coliforms, E. coli, and coliphages. Ten percent of samples were also analysed for enteric viruses, Giardia and Cryptosporidium. Approximately 320 households were included in a case-control study. In addition, a cross-sectional study was conducted. Both studies examined the relationship between different types of water facilities and diarrhoea among pre-school children. The source water was of good microbial quality, but water quality was found to have deteriorated significantly after handling and storage in both case and control households, exceeding drinking water quality guideline values by 1-6 orders of magnitude. Coliphage counts were low for all water samples tested. Enteric viruses and Cryptosporidium oocysts were not detected. Giardia cysts were detected on one occasion in case and control in-house samples. Comparisons of whether in-house water, after handling and storage, complied with water quality guideline values demonstrated households using communal taps to have significantly poorer quality than households using private outdoor or indoor taps for HPC and E. coli (χ2 = 14.9, P = 0.001; χ2 = 6.6, P = 0.04 respectively). A similar trend (although not statistically significant) was observed for the other microbial indicators. The cross-sectional study demonstrated an apparent decrease in health risk associated with private outdoor taps in comparison to communal taps. This study suggests that a private outdoor tap is the minimum level of water supply in order to ensure the supply of safe water to developing communities.

Miniature Bioreactors for Rapid Bioprocess Development of Mammalian Cell Culture

2012 ◽  
Vol 59 (1) ◽  
Author(s):  
Mohd Helmi Sani ◽  
Frank Baganz
Keyword(s):  
Cell Culture ◽  
Mammalian Cell ◽  
Process Development ◽  
Mammalian Cell Culture ◽  
Small Scale ◽  
Cell Culture Process ◽  
Deep Wells ◽  
Working Volume ◽  
Culture Process ◽  
And Control

At present, there are a number of commercial small scale shaken systems available on the market with instrumented controllable microbioreactors such as Micro–24 Microreactor System (Pall Corporation, Port Washington, NY) and M2P Biolector, (M2P Labs GmbH, Aachen, Germany). The Micro–24 system is basically an orbital shaken 24–well plate that operates at working volume 3 – 7 mL with 24 independent reactors (deep wells, shaken and sparged) running simultaneously. Each reactor is designed as single use reactor that has the ability to continuously monitor and control the pH, DO and temperature. The reactor aeration is supplied by sparging air from gas feeds that can be controlled individually. Furthermore, pH can be controlled by gas sparging using either dilute ammonia or carbon dioxide directly into the culture medium through a membrane at the bottom of each reactor. Chen et al., (2009) evaluated the Micro–24 system for the mammalian cell culture process development and found the Micro–24 system is suitable as scaledown tool for cell culture application. The result showed that intra-well reproducibility, cell growth, metabolites profiles and protein titres were scalable with 2 L bioreactors.

scholarly journals The Combined Effect of Weaning Stress and Immune Activation during Pig Gestation on Serum Cytokine and Analyte Concentrations

Animals ◽  
2021 ◽  
Vol 11 (8) ◽  
pp. 2274
Author(s):  
Haley E. Rymut ◽  
Laurie A. Rund ◽  
Courtni R. Bolt ◽  
Maria B. Villamil ◽  
Bruce R. Southey ◽  
...  
Keyword(s):  
Immune Activation ◽  
Management Practices ◽  
Interleukin 1 ◽  
Interleukin 2 ◽  
Blood Chemistry ◽  
Respiratory Syndrome Virus ◽  
Weaning Stress ◽  
And Control ◽  
Metabolic Hormone

Weaning stress can elicit changes in the metabolic, hormone and immune systems of pigs and interact with prolonged disruptions stemming from maternal immune activation (MIA) during gestation. The present study advances the characterization of the combined effects of weaning stress and MIA on blood chemistry, immune and hormone indicators that inform on the health of pigs. Three-week-old female and male offspring of control gilts or gilts infected with the porcine reproductive and respiratory syndrome virus were allocated to weaned or nursed groups. The anion gap and bilirubin profiles suggest that MIA enhances tolerance to the effects of weaning stress. Interleukin 1 beta and interleukin 2 were highest among weaned MIA females, and cortisol was higher among weaned relative to nursed pigs across sexes. Canonical discriminant analysis demonstrated that weaned and nursed pigs have distinct chemistry profiles, whereas MIA and control pigs have distinct cytokine profiles. The results from this study can guide management practices that recognize the effects of the interaction between MIA and weaning stress on the performance and health of pigs.

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