An indirect ELISA for Neosporosis: Associating recombinant Neospora caninum proteins NcSRS2 and NcSAG1

Sinnott, Francine Alves; Leal, Karen Silva; Silva, Mara Thais de Oliveira; Pinho, Rodrigo Barros de; Pappen, Felipe Geraldo; Farias, Nara Amélia da Rosa; Llano, Horwald Alexander Bedoya; Melo, Débora Pereira Garcia; Borsuk, Sibele

doi:10.1016/j.vetpar.2020.109101

Cited by 6 publications

(5 citation statements)

References 24 publications

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“…Hamidinejat et al ( 35 ) developed the rNcGRA7-based indirect ELISA in cattle and water buffalo, sensitivities and specificities were 94.64 and 90.38% respectively using sera of cattle, but were 98.57 and 86.54% in the case of buffaloes respectively ( 35 ). More recently, an indirect ELISA based on rNcSRS2/rNcSAG1 was developed for the diagnosis of cattle and ovine neosporosis, and the sensitivity and specificity performed with the proteins NcSRS2 and NcSAG1 in combination had higher than alone ( 20 ). In this study, we obtained 86.7% sensitivity and higher specificity (96.1%) when using this chimeric protein, suggesting its applicability to the practical diagnosis of neosporosis.…”

Section: Discussionmentioning

confidence: 99%

“…The inclusion body was dissolved in 8 M urea solution and purified by affinity chromatography on a HisTrap™ Sepharose nickel column (Beyotime Biotechnology; Shanghai, China). And the concentration was determined using a BCA kit (Beyotime Biotechnology; Shanghai, China) as previously described ( 20 ).…”

Section: Methodsmentioning

confidence: 99%

“…The NcSRS2 protein is expressed in tachyzoites and bradyzoites, with a 44% sequence identity to T. gondii TgSRS2, the NcSAG1 protein is one of the major tachyzoite surface protein, and displays 53% identity to T. gondii TgSAG1, and the dense granule protein NcGRA7 is with a 42% sequence identity to T. gondii TgGRA7 (17)(18)(19). The sensitivity and specificity of indirect ELISA based on NcSRS2, NcSAG1, or NcGRA7 have been studied, and Alves et al (20) confirmed that sensitivity and specificity of indirect ELISA performed with the proteins NcSRS2 and NcSAG1 in combination had higher than individually (20). However, the chimeric expression of these three proteins has not been studied.…”

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confidence: 99%

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Development and application of an indirect ELISA to detect antibodies to Neospora caninum in cattle based on a chimeric protein rSRS2-SAG1-GRA7

Yang

Ayanniyi

et al. 2022

Front. Vet. Sci.

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Neospora caninum is an important apicomplexan parasite causing neosporosis in cattle. The disease is recognized as one of the most important cause of reproductive problems and abortion in cattle worldwide. In this context, we developed an indirect enzyme-linked immunosorbent assays (ELISA) with chimeric protein rSRS2-SAG1-GRA7 to diagnose antibodies to Neospora-infection. This indirect ELISA was compared to indirect fluorescent antibody test (IFAT) and western blotting (WB), and the sensitivity and specificity results of ELISA were calculated to be 86.7 and 96.1%, respectively. The overall coincidence rate was 92.6% using IFAT and WB. Additionally, 329 aborting dairy cattle serum samples were tested using this ELISA to evaluate the prevalence of N. caninum in Ningxia, China. The positive rate of N. caninum in these farms was from 19.05 to 57.89%, and the mean rate was 41.64% (±11.01%), indicating that infection with N. caninum may be one of the important causes of cattle abortion in this region. This established rSRS2-SAG1-GRA7 indirect ELISA is capable for detecting the antibodies against N. caninum, and it could be a useful screening tool for monitoring the epidemiology of neosporosis in cattle.

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Section: Discussionmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

mentioning

confidence: 99%

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Development and application of an indirect ELISA to detect antibodies to Neospora caninum in cattle based on a chimeric protein rSRS2-SAG1-GRA7

Yang

Ayanniyi

et al. 2022

Front. Vet. Sci.

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“…Neospora caninum is an obligate intracellular parasitic protozoan that has a complex life cycle and infects a number of warm-blooded animals ( 1 , 2 ). N. caninum can complete sexual reproduction in canids which is the definitive hosts of this parasite; while many important domestic and wild animals, such as cattle, sheep and camels, can serve as intermediate hosts ( 3 – 5 ). Because dogs often can move freely in the cattle herds, the risk of cattle infection with N. caninum is high, and the seroprevalence of N. caninum can be as high as 90% in some cattle herds ( 6 , 7 ).…”

Section: Introductionmentioning

confidence: 99%

Seroprevalence of Neospora caninum infection and associated risk factors in cattle in Shanxi Province, north China

et al. 2022

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Neospora caninum is an obligate intracellular parasitic protozoan that can cause abortions in cattle and pose considerable economic losses to the cattle industry. As a major livestock province, little is known of N. caninum infection in cattle in Shanxi Province, north China. In order to investigate the seroprevalence of N. caninum in cattle in Shanxi Province, 978 cattle serum samples were collected from 11 cities in three representative geographical locations in Shanxi Province, and the N. caninum-specific IgG antibodies were examined using an indirect enzyme linked immunosorbent assay (ELISA) kit commercially available. The results showed that 133 of the 978 examined cattle serum samples (13.60%, 95% CI = 11.45–15.75) were positive for N. caninum antibodies, and the seroprevalence in different cities ranged from 0 to 78.89%. The geographical location and management mode were the risk factors associated with N. caninum infection in cattle herds in Shanxi Province. Cattle in Northern and Central Shanxi Province as well as cattle whose management mode is that of large-scale cattle farming companies are more susceptible to N. caninum infection. This was the first large-scale survey of N. caninum seroprevalence and assessment of associated risk factors in cattle in Shanxi Province, which provided baseline information for the prevention and control of N. caninum infection in cattle in Shanxi Province, north China.

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“…In line with this, we obtained a Klebsiella pneumoniae goat polyclonal antibody and established an indirect ELISA method. Furthermore, a total of 1320 serum samples from clinics in Shandong Province were subjected to assess whether our assay is feasible to deploy in clinical veterinary settings [13]. Therefore, the developed an indirect ELISA detection assay is a rapid diagnostic method that could be applied to clinical veterinary diagnosis and epidemiological investigation of goat-sourced Klebsiella pneumonia [14].…”

Section: Introductionmentioning

confidence: 99%

Establishment and evaluation of an indirect ELISA for detection of antibodies to goat Klebsiella pneumonia

Chen¹,

Shang

Niu

et al. 2020

Preprint

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Background: Klebsiella pneumonia, a Gram-negative bacterium belonging to the genus Enterobacter, causes many human and livestock diseases. Notably, infected goats may develop pneumonia, septicemia, which can lead to occasional death, resulting in great economic losses in goat-farming industry. However, there are little systematic methods for detection of goat Klebsiella pneumoniae in livestock production.Results: In this study, we developed a Klebsiella pneumoniae goat polyclonal antibody and established an indirect ELISA method to detect the Klebsiella pneumoniae. After screening and optimizing the conditions for detection, we determined the optimal working dilutions of the coated-bacterial antigen, the polyclonal antibody, and the enzyme-labeled secondary antibody that were 1:800 (2.99x107 CFU/ml), 1:6400, and 1:5000, respectively. The optimal condition of coating and blocking were both 4 °C for 12 h. The optimal dilution buffers of bacterial antigen, the antibodies, and the blocking buffer were 0.05 mol/L carbonate buffer, 1% BSA phosphate buffer, and 1.5% BSA carbonate buffer, respectively. The cut-off value was determined to be 0.28, and the analytical sensitivity was 1:800 (dilution of a positive sample). Furthermore, there was no cross-reaction between the coated antigen and goat serum positive for antibodies against other bacteria, indicating that indirect ELISA could detect Klebsiella pneumoniae specifically in most cases. The average coefficients of variation of intra-assay and inter-assay were 4.37% and 5.17% indicating favorable reproducibility of indirect ELISA. In the detection of clinical veterinary samples, the positive rate of indirect ELISA was 6.74%, higher than that of conventional agglutination assays. Conclusions: Taken together, we successfully established an indirect ELISA method for detecting antibodies against Klebsiella pneumoniae in goats, which can be applied in production.

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An indirect ELISA for Neosporosis: Associating recombinant Neospora caninum proteins NcSRS2 and NcSAG1

Cited by 6 publications

References 24 publications

Development and application of an indirect ELISA to detect antibodies to Neospora caninum in cattle based on a chimeric protein rSRS2-SAG1-GRA7

Development and application of an indirect ELISA to detect antibodies to Neospora caninum in cattle based on a chimeric protein rSRS2-SAG1-GRA7

Seroprevalence of Neospora caninum infection and associated risk factors in cattle in Shanxi Province, north China

Establishment and evaluation of an indirect ELISA for detection of antibodies to goat Klebsiella pneumonia

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