High-level prokaryotic expression of envelope exterior of membrane protein of porcine epidemic diarrhea virus

Gao, Shandian; Zha, Enhui; Baoxian, Li; Qiao, Xin; Tang, Lijie; Li, Yijing

doi:10.1016/j.vetmic.2007.03.027

Cited by 8 publications

(6 citation statements)

References 19 publications

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“…However, these researchers did not assess the cross-reactivity of the ELISA with other coronaviruses. The M protein of PEDV protrudes from the viral envelope, and is considered a superior diagnostic antigen compared with other PEDV proteins (Shenyang et al, 2007). In the current study, we chose the PEDV M protein for use as an ELISA antigen because it is highly conserved among PEDV strains, and because it can elicit the formation of protective antibodies (Zhang et al, 2012).…”

Section: Discussionmentioning

confidence: 99%

Development of an enzyme-linked immunosorbent assay for the monitoring and surveillance of antibodies to porcine epidemic diarrhea virus based on a recombinant membrane protein

Fan

Zuo

Shen

et al. 2015

Journal of Virological Methods

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The recent dramatic increase in reported cases of porcine epidemic diarrhea (PED) in pig farms is a potential threat to the global swine industry. Therefore, the accurate diagnosis, serological monitoring, and surveillance of specific antibodies in pigs resulting from porcine epidemic diarrhea virus (PEDV) infection or vaccination would be essential in helping to control the spread of PED. We developed and validated an indirect enzyme-linked immunosorbent assay (ELISA) based on the recombinant membrane (M) protein of PEDV. To detect PEDV antibodies in eight herds, 382 serum samples were collected from sows that had been immunized with a PED vaccine, and screened using the developed ELISA in parallel with a serum neutralization (SN) assay. Of the tested samples, 276 were positive for the presence of PEDV antibodies according to both assays, while 98 were negative. An excellent agreement between the ELISA and the SN assay was observed (kappa=0.947; 95% confidence interval=0.910-0.984; McNemar's test, P=0.727). No cross-reaction was detected for the developed ELISA with other coronaviruses or other common pig pathogens. The developed ELISA could be used for serological evaluation and indirect diagnosis of PED infection.

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Section: Discussionmentioning

confidence: 99%

Development of an enzyme-linked immunosorbent assay for the monitoring and surveillance of antibodies to porcine epidemic diarrhea virus based on a recombinant membrane protein

Fan

Zuo

Shen

et al. 2015

Journal of Virological Methods

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“…In addition, the nucleotide sequence of the M gene is highly conserved among different PEDV strains but shows little homology with other coronaviruses (Kim and Chae 2000). Thus, the M protein has been used as a target antigen for PEDV detection (Ren et al 2011;Shenyang et al 2007).…”

Section: Introductionmentioning

confidence: 99%

Single-domain antibodies as promising experimental tools in imaging and isolation of porcine epidemic diarrhea virus

Yang

Yin

et al. 2018

Appl Microbiol Biotechnol

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Single-domain antibody (sdAb) or nanobody possesses specific features non-accessible for conventional antibodies that make them suitable for research and biotechnological applications. Porcine epidemic diarrhea virus (PEDV) causes lethal diarrhea in piglets, resulting in great economic losses all over the world. To detect and isolate PEDV rapidly and accurately is important for the control and further research of the clinical PEDV strains. In this study, four sdAb fragments (sdAb-Mc19/29/30/37) targeting the membrane (M) protein of PEDV were selected from sdAb library that was constructed through M protein-immunized Camelus bactrianus. The selected sdAb-Mcs were solubly expressed in Escherichia coli. The functional characteristics analysis revealed that the recombinant sdAb-Mcs have excellent binding activity and specificity to M protein but have no neutralizing activity to PEDV. For further application, sdAb-Mc37 was conjugated with quantum dots to synthesize a nanoprobe for imaging PEDV in vero cells. The observed fluorescence in vero cells clearly reflects that PEDV virions can be reliably recognized and labeled by the nanoprobe. Furthermore, the sdAb-Mc29 was conjugated with superparamagnetic nanobeads to construct immunomagnetic nanobeads (IMNBs) used to isolate PEDV. One PEDV strain was successfully isolated from clinical fecal sample, suggesting IMNBs as a novel and efficient tool suitable for PEDV isolation from clinical samples. This study provided a novel application and substantiated the suitability of sdAb as a specific binder for the isolation of viruses.

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“…Specifically, there has been an interest to develop a recombinant protein (Balamurugan et al, 2010). For example, several immune assays based on recombinant proteins (S, M, N) from PEDV have been developed using E. coli as an expression system (Cao et al, 2013;Pan et al, 2015;Ren et al, 2011;Shenyang et al, 2007). Also, a recombinant TGEV N protein was developed with high sensitivity and specificity (Liu et al, 2001).…”

Section: Discussionmentioning

confidence: 99%

Cloning, expression, and characterization of potential immunogenic recombinant porcine deltacoronavirus (PDCoV) membrane protein from a synthetic gene.

Cuevas-Romero

Montes

Cerriteño-Sánchez

et al. 2022

Preprint

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Porcine Deltacoronavirus (PDCoV) is an emergent swine coronavirus that infects epithelia cells from the small intestine, and inducing watery diarrhea, vomiting and dehydration. Clinical signs are more aggressive in piglets causing high mortality rates (>40%) representing serious economic losses. Despite the importance of PDCoV as an emerging coronavirus, little is known about the currently prevalence in México. We select from GenBank a group of 138 sequences and obtained a consensus PDCoV membrane protein (M-PDCoV) sequence of 216 a.a. A Maximum Likelihood phylogenetic tree was constructed and evaluate the relationship between the 138 sequences. Also, a protein tertiary structure analysis was performed to analyze and compare the topological differences. The phylogeny and the tertiary structure analysis showed that M-PDCoV is highly conserved and therefore suitable to use as an antigen in a diagnostic system. Hence, an expression system performed in E. coli BL21 (DE3) using the vector pET-SUMO with a His-tag was prepared, resulting in a synthetic M gene of 654 pb to produce a recombinant M-PDCoV protein ( rM-PDCoV). Western blot test confirmed the rM-PDCoV immune detection in 8 of 17 sera samples. The rM-PDCoV was able to successfully stimulate immune response in immunized mice to produce antibodies after day 7 of immunization (P < 0.001). Our results show that rM-PDCoV is suitable to use in diagnostic systems like an ELISA.

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High-level prokaryotic expression of envelope exterior of membrane protein of porcine epidemic diarrhea virus

Cited by 8 publications

References 19 publications

Development of an enzyme-linked immunosorbent assay for the monitoring and surveillance of antibodies to porcine epidemic diarrhea virus based on a recombinant membrane protein

Development of an enzyme-linked immunosorbent assay for the monitoring and surveillance of antibodies to porcine epidemic diarrhea virus based on a recombinant membrane protein

Single-domain antibodies as promising experimental tools in imaging and isolation of porcine epidemic diarrhea virus

Cloning, expression, and characterization of potential immunogenic recombinant porcine deltacoronavirus (PDCoV) membrane protein from a synthetic gene.

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