scholarly journals Molecular characterization and serodiagnostic potential of two serpin proteins in Psoroptes ovis var. cuniculi

2020 ◽  
Vol 13 (1) ◽  
Author(s):  
Xiaobin Gu ◽  
Yuhang Chen ◽  
Chongyang Zhang ◽  
Yue Xie ◽  
Nengxing Shen ◽  
...  

Abstract Background Psoroptes ovis var. cuniculi is a common ectoparasite of wild and domestic rabbits worldwide that causes economically devastating losses in commercial rabbit husbandry and significantly affects the overall health of rabbits. Serine proteinase inhibitor (serpin) is present in almost all organisms that are involved in host–pathogen interactions, inflammatory responses, and reproductive development, among others. However, very little research has been carried out on P. ovis var. cuniculi serpins. Methods Two serpin genes of P. ovis var. cuniculi (Pso c 27 and PsoSP2 cDNAs) were cloned and molecularly characterized. The transcriptional profiles and tissue localization of these two serpins in P. ovis var. cuniculi were investigated by quantitative real-time PCR and immunohistochemistry, respectively. The potential function of recombinant Pso c 27 and PsoSP2 (rPso c 27 and rPsoSP2) in the serodiagnosis of P. ovis var. cuniculi infestation in rabbits was evaluated using a newly devleoped indirect enzyme-linked immunosorbent assay. Results Both the 523-residue Pso c 27 and the 240-residue PsoSP2 proteins contained typical serpin domains and signatures. Both Pso c 27and PsoSP2 cDNAs were expressed throughout the life-cycle; specifically, the cDNAs showed significantly higher expression in female mites than in larva, nymph, and male mites (Pso c 27: F(3, 8) = 1935.953, P < 0.0001; PsoSP2: F(3, 8) = 660.669, P < 0.0001). The native Pso c 27 and PsoSP2 proteins localized in the ovary and mouthparts of adult female mites, respectively. Compared to rPsoSP2, rPso c 27 showed better diagnostic efficiency, with higher values of sensitivity, specificity, and area under the receiver operating characteristic curve (rPso c 27 vs rPsoSP2: 96.0 vs 90.0%; 90.91 vs 78.18%; 0.988 vs 0.964, respectively). Moreover, rPso c 27 showed seropositivity in 80% of the rabbits as early as the 2 weeks post-infestation, prior to visible clinical signs and microscopy-positive of skin scrapings. Conclusions These results suggest that these two serpins may play essential roles in reproductive development, serum-feeding, and pathogenicity of P. ovis var. cuniculi. Compared to PsoSP2, Pso c 27 appears to be a potential antigen for serodiagnosis of P. ovis var. cuniculi infestation in rabbits, especially at the early stage of infestation.

2020 ◽  
Author(s):  
Xiaobin Gu ◽  
Yuhang Chen ◽  
Chongyang Zhang ◽  
Yue Xie ◽  
Nengxing Shen ◽  
...  

Abstract Background: Psoroptes ovis var. cuniculi is a global common ectoparasite of wild and domestic rabbits and causes an economically devastating loss and serious welfare issues of commercial rabbit husbandry. Serine proteinase inhibitor (serpin) is present in almost all organisms that are involved in host-pathogen interactions, inflammatory responses, and reproductive development, etc. However, the research on P. ovis var. cuniculi serpins is still limited.Methods: In this study, two serpin genes of P. ovis var. cuniculi (Pso c 27 and PsoSP2 cDNAs) were cloned, and the molecular characterization was analyzed. The transcriptional profiles and tissue localization of these two serpins in P. ovis var. cuniculi were investigated by quantitative real-time PCR and immunohistochemistry, respectively. The potential function of recombinant Pso c 27 and PsoSP2 (rPso c 27and rPsoSP2) in the serodiagnosis of P. ovis var. cuniculi infestation in rabbits were evaluated by indirect enzyme-linked immunosorbent assay (iELISA). Results: Both of the 523 residue Pso c 27 and the 240 residue PsoSP2 proteins contained typical serpin domains and signatures. Both Pso c 27and PsoSP2 cDNAs expressed throughout the life-cycle, more specifically, significantly higher expression in female mites than the larva, nymph, and male mites (Pso c 27, F(3, 8) = 1935.953, P < 0.0001; PsoSP2, F(3, 8) = 660.669, P < 0.0001). The native Pso c 27 and PsoSP2 localized in ovary and mouthpart of adult female mites, respectively. Compared to rPsoSP2, the rPso c 27 displayed better diagnostic efficiency with higher values of sensitivity, specificity and the area under the receiver operating characteristic curve (AUC) (rPso c 27 vs rPsoSP2: 96.0 vs 90.0%; 90.91 vs 78.18%; 0.988 vs 0.964, respectively). Moreover, the rPso c 27 showed seropositive in 80% rabbits as early as the 2 weeks post-infestation (p.i.), prior to visible clinical signs and microscopy-positive of skin scrapings. Conclusions: These results suggested that these two serpins may play essential roles in reproductive development, serum-feeding, and pathogenicity of P. ovis var. cuniculi. Compared to PsoSP2, Pso c 27 appeared as a potential antigen for serodiagnosis of P. ovis var. cuniculi infestation in rabbits, especially at the early stage of infestation.


2020 ◽  
Author(s):  
Xiaobin Gu ◽  
Yuhang Chen ◽  
Chongyang Zhang ◽  
Yue Xie ◽  
Nengxing Shen ◽  
...  

Abstract Background: Psoroptes cuniculi is a global common ectoparasite of wild and domestic rabbits and causes an economically devastating loss and serious welfare issues of commercial rabbit husbandry. Serine proteinase inhibitor (Serpin) is present in almost all organisms that are involved in host-pathogen interactions, inflammatory responses, and reproductive development, etc. However, the research on P. cuniculi serpins is still limited.Methods: In this study, two serpin genes of P. cuniculi (Pso c 27 and PsoSP2 cDNAs) were cloned, and the molecular characterization was analyzed. The transcriptional profiles and tissue localization of these two serpins in P. cuniculi were investigated by quantitative real-time PCR and immunohistochemistry, respectively. The potential function of recombinant Pso c 27 and PsoSP2 (rPso c 27and rPsoSP2) in the serodiagnosis of P. cuniculi infestation in rabbits were evaluated by indirect enzyme-linked immunosorbent assay (iELISA). Results: Both of the 523 residue Pso c 27 and the 240 residue PsoSP2 proteins contained typical serpin domains and signatures. Both Pso c 27and PsoSP2 cDNAs expressed throughout the life cycle, more specifically, significantly higher expression in female mites than the larva, nymph, and male mites (Pso c 27, F(3, 8) = 1935.953, p < 0.0001; PsoSP2, F(3, 8) = 660.669, p < 0.0001). The native Pso c 27 and PsoSP2 localized in ovary and mouthpart of adult female mites, respectively. Compared to rPsoSP2, the rPso c 27 displayed better diagnostic efficiency with higher values of sensitivity, specificity and the area under the receiver operating characteristic curve (AUC) (rPso c 27 - rPsoSP2: 96.0% - 90.0%; 90.91% - 78.18%; 0.988 - 0.964, respectively). Moreover, the rPso c 27 showed seropositive in 80% rabbits as early as the 2nd week post-infestation (p.i.), prior to visible clinical signs and microscopy-positive of skin scrapings. Conclusions: These results suggested that these two serpins may play essential roles in reproductive development, serum-feeding, and pathogenicity of P. cuniculi. Compared to PsoSP2, Pso c 27 appeared as a potential antigen for serodiagnosis of P. cuniculi infestation in rabbits, especially at the early stage of infestation.


2020 ◽  
Author(s):  
Xiaobin Gu ◽  
Yuhang Chen ◽  
Chongyang Zhang ◽  
Yue Xie ◽  
Nengxing Shen ◽  
...  

Abstract Background: Psoroptes cuniculi is a global common ectoparasite of wild and domestic rabbits and causes an economically devastating loss and serious welfare issues of commercial rabbit husbandry. Serine proteinase inhibitor (serpin) is present in almost all organisms that are involved in host-pathogen interactions, inflammatory responses, and reproductive development, etc. However, the research on P. cuniculi serpins is still limited.Methods: In this study, two serpin genes of P. cuniculi (Pso c 27 and PsoSP2 cDNAs) were cloned, and the molecular characterization was analyzed. The transcriptional profiles and tissue localization of these two serpins in P. cuniculi were investigated by quantitative real-time PCR and immunohistochemistry, respectively. The potential function of recombinant Pso c 27 and PsoSP2 (rPso c 27and rPsoSP2) in the serodiagnosis of P. cuniculi infestation in rabbits were evaluated by indirect enzyme-linked immunosorbent assay (iELISA). Results: Both of the 523 residue Pso c 27 and the 240 residue PsoSP2 proteins contained typical serpin domains and signatures. Both Pso c 27and PsoSP2 cDNAs expressed throughout the life-cycle, more specifically, significantly higher expression in female mites than the larva, nymph, and male mites (Pso c 27, F(3, 8) = 1935.953, P < 0.0001; PsoSP2, F(3, 8) = 660.669, P < 0.0001). The native Pso c 27 and PsoSP2 localized in ovary and mouthpart of adult female mites, respectively. Compared to rPsoSP2, the rPso c 27 displayed better diagnostic efficiency with higher values of sensitivity, specificity and the area under the receiver operating characteristic curve (AUC) (rPso c 27 vs rPsoSP2: 96.0 vs 90.0%; 90.91 vs 78.18%; 0.988 vs 0.964, respectively). Moreover, the rPso c 27 showed seropositive in 80% rabbits as early as the 2 weeks post-infestation (p.i.), prior to visible clinical signs and microscopy-positive of skin scrapings. Conclusions: These results suggested that these two serpins may play essential roles in reproductive development, serum-feeding, and pathogenicity of P. cuniculi. Compared to PsoSP2, Pso c 27 appeared as a potential antigen for serodiagnosis of P. cuniculi infestation in rabbits, especially at the early stage of infestation.


2020 ◽  
Author(s):  
Xiaobin Gu ◽  
Yuhang Chen ◽  
Chongyang Zhang ◽  
Yue Xie ◽  
Nengxing Shen ◽  
...  

Abstract Background: Psoroptes ovis a common ectoparasite of wild and domestic animals, leads to cutaneous inflammation, extreme pruritus, scaly lesions, and causes an economically devastating loss of animal husbandry and animal welfare issues. Serine proteinase inhibitor (Serpin) is present in almost all organisms that are involved in host-pathogen interactions, inflammatory responses, and reproductive development, etc. However, the research on P. ovis serpins is still limited.Methods: In this study, two serpins of P. ovis (PsoSP1 and PsoSP2) were cloned, and the molecular characterization was analyzed by bioinformatics. The transcriptional profiles and tissue localization of PsoSP1 and PsoSP2 in P. ovis were investigated by quantitative real-time PCR and immunohistochemistry, respectively. The potential function of recombinant PsoSP1 and PsoSP2 (rPsoSP1 and rPsoSP2) in the serodiagnosis of P. ovis infestation in rabbits were evaluated by indirect enzyme-linked immunosorbent assay (iELISA). Results: Both of the 523 residue PsoSP1 and the 240 residue PsoSP2 proteins contained typical serpin domains and signatures. Both PsoSP1 and PsoSP2 expressed throughout the life cycle, more specifically, significantly higher expression in female mites than the larva, nymph, and male mites (p < 0.001). The native PsoSP1 and PsoSP2 proteins localized in ovary and mouthpart of adult female mites, respectively. Compared to rPsoSP2, the rPsoSP1 displayed better diagnostic efficiency with higher values of sensitivity, specificity and the area under the receiver operating characteristic curve (AUC) than rPsoSP1 by iELISA (rPsoSP1 - rPsoSP2: 96.0% - 90.0%; 90.91% - 78.18%; 0.988 - 0.964, respectively). Moreover, the rPsoSP1 showed seropositive in 80% rabbits as early as the 2nd week post-infestation (p.i.), prior to visible clinical signs and microscopy-positive of skin scrapings. Conclusions: These results suggested that these two serpins may play essential roles in reproductive development, blood-feeding, and pathogenicity of P. ovis. Compared to PsoSP2, PsoSP1 appeared as a potential antigen for serodiagnosis of P. ovis infestation in rabbits, especially at the early stage of infestation.


2020 ◽  
Author(s):  
Xiaobin Gu ◽  
Yuhang Chen ◽  
Chongyang Zhang ◽  
Yue Xie ◽  
Nengxing Shen ◽  
...  

Abstract Background: Psoroptes cuniculi is a global common ectoparasite of wild and domestic rabbits and causes an economically devastating loss and serious welfare issues of commercial rabbit husbandry. Serine proteinase inhibitor (Serpin) is present in almost all organisms that are involved in host-pathogen interactions, inflammatory responses, and reproductive development, etc. However, the research on P. cuniculi serpins is still limited.Methods: In this study, two serpin genes of P. cuniculi (Pso c 27 and PsoSP2 cDNAs) were cloned, and the molecular characterization was analyzed. The transcriptional profiles and tissue localization of these two serpins in P. cuniculi were investigated by quantitative real-time PCR and immunohistochemistry, respectively. The potential function of recombinant Pso c 27 and PsoSP2 (rPso c 27and rPsoSP2) in the serodiagnosis of P. cuniculi infestation in rabbits were evaluated by indirect enzyme-linked immunosorbent assay (iELISA). Results: Both of the 523 residue Pso c 27 and the 240 residue PsoSP2 proteins contained typical serpin domains and signatures. Both Pso c 27and PsoSP2 cDNAs expressed throughout the life cycle, more specifically, significantly higher expression in female mites than the larva, nymph, and male mites (p < 0.001). The native Pso c 27 and PsoSP2 localized in ovary and mouthpart of adult female mites, respectively. Compared to rPsoSP2, the rPso c 27 displayed better diagnostic efficiency with higher values of sensitivity, specificity and the area under the receiver operating characteristic curve (AUC) (rPso c 27 - rPsoSP2: 96.0% - 90.0%; 90.91% - 78.18%; 0.988 - 0.964, respectively). Moreover, the rPso c 27 showed seropositive in 80% rabbits as early as the 2nd week post-infestation (p.i.), prior to visible clinical signs and microscopy-positive of skin scrapings. Conclusions: These results suggested that these two serpins may play essential roles in reproductive development, serum-feeding, and pathogenicity of P. cuniculi. Compared to PsoSP2, Pso c 27 appeared as a potential antigen for serodiagnosis of P. cuniculi infestation in rabbits, especially at the early stage of infestation.


Author(s):  
I. K. Avdosieva ◽  
O. I. Chajkovska ◽  
O. B. Basarab ◽  
V. V. Regenchuk

One of the main problems of poultry diseases is respiratory diseases. Among them a special place is occupied by ornithobacteriosis (ORT). Losses from ORT consist of: direct losses as a result of the disease - death of chickens, increased culling due to lameness, low live weight gain (up to 40%), reduction of carcass categories, reduction of egg production by 6-20%; indirect losses associated with the immune-suppressive effect of ornithobacteria, which increase the risk of other infections and prevent the formation of post-vaccination immunity. The diagnosis is established on the basis of epizootological data, clinical signs, pathological and anatomical changes, bacteriological and serological tests, positive bioassay. In most cases, infections caused by ornithobacteria are not diagnosed in time, the pathogen is difficult to isolate due to complications of other pathogens, or because experts are currently insufficiently aware of the ability of ORT to cause disease. ORT can be isolated by bacteriological method only at an early stage of the disease. The most relevant method of diagnosis is PCR. The advantage of the method is not only the isolation of DNA of individual cells of the pathogen in the sample, but also the ability to detect all serotypes. In addition, PCR is a successful diagnosis in the detection of ORT nucleic acid not only in tissue samples, but also in feces, eggs, dust, which is important for timely diagnosis. Enzyme-linked immunosorbent assay (ELISA) is used to control the presence of ornithobacteriosis in bird. The presence of antibodies to this pathogen in poultry of many species indicates its wide circulation. Thus, when conducting serological monitoring of blood serum from different age groups of broilers aged 1-44 days, the percentage of positive samples ranged from 40 to 100, which indicates the circulation of the field strain of the pathogen ornithobacteriosis. The percentage of positive serum from broilers to ORT was: from 1 to 5 days - from 88 to 50, from 6 to 10 days serum were negative, while at 17, 21 days and from 32 days to the end of cultivation (44 days) – 100 %. The percentage of positive batches at the end of fattening ranged from 42-53 days in the range from 75 to 100%, indicating the circulation of the field strain of the ornithobacteriosis pathogen antibiotic therapy against this disease.


Author(s):  
Tao Li ◽  
Li Wang ◽  
Huihui Wang ◽  
Xuemei Li ◽  
Shubing Zhang ◽  
...  

AbstractObjectiveThis study aimed to explore the diagnostic value of serum severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) nucleocapsid(N) protein assay in the early stage of SARS-COV-2 infection.MethodSerum N protein in SARS-COV-2 infected patients and non-SARS-COV-2 infected population was measured by enzyme-linked immunosorbent assay (ELISA) double antibody sandwich assay. Colloidal gold immunochromatography assay is used to detect serum N protein antibodies in the above population.Results50 cases of SARS-CoV-2 nucleic acid positive and SARS-CoV-2 antibody negative patients had a serum N protein positive rate of 76%, including 2% with a concentration of 10.00-49.99 pg / mL, 8% with a concentration of 50.00-99.99 pg / mL, 22% with a concentration of 100.00 - 299.99 pg/mL, and 44% with a concentration≥ 300.00 pg / mL. 37 samples of patients with serum SARS-CoV-2 antibody positive after infection had a serum SARS-CoV-2 N protein positive rate of 2.7%, of which 2.7% had the concentration of 10.00-49.99 pg / mL and 0% had the concentration of 50.00-99.99 pg / mL, 100.00 −299.99 pg / mL, and >300.00 pg / mL. Serum N protein test results of 633 non-SARS-COV-2 infected patients including pregnant women, other respiratory infections, and increased rheumatoid factor were all negative, having a serum N protein concentration less than 10.00 pg/mL, with a specificity of 100%. Using SPSS 19.0 to calculate the receiver operating characteristic curve, the area under the curve was 0.9756 (95% confidence interval 0.9485-1.000, p <0.0001), sensitivity and specificity were 92% (95% confidence interval 81.16% to 96.85%) and 96.84% (95% confidence interval 95.17% to 97.15%). The best CUTOFF value is 1.850 pg / mL.ConclusionThe measurement of SARS-COV-2 serum N protein has a high diagnostic value for the infected patients before the antibody appears, and shortens the window period of serological diagnosis. The laboratory needs to establish an individual CUTOFF value according to purpose of the application.


Cartilage ◽  
2020 ◽  
pp. 194760352095814
Author(s):  
Tachatra Ungsudechachai ◽  
Sittisak Honsawek ◽  
Jiraphun Jittikoon ◽  
Wanvisa Udomsinprasert

Objectives This study aimed to determine possible associations between transcriptional and translational levels of clusterin (CLU) in the systemic and local joint environments with the severity of knee osteoarthritis (OA) and to investigate CLU mRNA expression in knee OA fibroblast-like synoviocytes (FLSs) stimulated with tumor necrosis factor-α. Design Circulating and synovial fluid CLU levels in 259 knee OA patients were quantified using an enzyme-linked immunosorbent assay. Relative CLU mRNA expression in 50 knee OA synovial tissues and 4 knee OA FLSs was determined using real-time polymerase chain reaction. Results Plasma CLU levels of knee OA patients were significantly higher than paired synovial fluid samples. Compared with early-stage knee OA patients, those with advanced-stage OA had considerably increased plasma and synovial fluid CLU levels. There were significant positive associations of plasma and synovial fluid CLU levels with radiographic severity of knee OA. Plasma CLU levels were directly correlated with its synovial fluid levels and high-sensitivity C-reactive protein levels in the patients. Receiver-operating characteristic curve analysis unveiled the potential utility of plasma CLU as a novel biomarker for knee OA severity (AUC = 0.80), with a sensitivity of 71.4% and a specificity of 73.3%. Marked upregulation of CLU mRNA expression was observed in both the inflamed synovial tissues and FLSs of knee OA. Conclusion Increased CLU mRNA and protein levels in the systemic and local joint environments of knee OA might reflect knee OA severity, especially systemic and synovial inflammation.


2018 ◽  
Vol 2018 ◽  
pp. 1-10 ◽  
Author(s):  
Jie Hou ◽  
Manli Zhang ◽  
Yuhong Ding ◽  
Xinrui Wang ◽  
Tao Li ◽  
...  

Aim. To analyze changes in peripheral blood monocytes and their clinical significance in patients with early stage of idiopathic membranous nephropathy (IMN). Methods. A total of 27 patients with early stage of IMN and 16 age- and sex-matched healthy controls (HCs) were recruited for the study. The monocyte subset counts in circulation were measured by flow cytometry, and serum interleukin- (IL-) 10 and IL-12 concentrations were tested by enzyme-linked immunosorbent assay. The potential association between clinical signs and monocyte subset counts was analyzed statistically. Results. Compared with the HCs, the patients with early stage of IMN had higher counts of CD14+CD163+, CD14+CD163+CD206+, and CD14+CD163+CD206+CD115+ M2-like monocytes. The CD14+CD163+CD206+ M2-like cell counts and intracellular IL-10 concentrations in the monocytes were positively correlated with progression in proteinuria. The levels of serum IL-10 were significantly higher in early IMN patients than in the HCs. Furthermore, CD14+CD163+CD206+ M2-like cell counts in the patients with incipient IMN were also positively related with 24 h urinary albumin levels and the values of serum M-type phospholipase A2 receptor (PLA2R). Conclusion. CD14+CD163+CD206+ M2-like monocytes may contribute to the pathologic process in early-stage IMN and could serve as potential markers for evaluating the disease severity.


2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Wanvisa Udomsinprasert ◽  
Napat Angkathunyakul ◽  
Jiraphun Jittikoon ◽  
Usa Chaikledkaew ◽  
Paisarn Vejchapipat ◽  
...  

AbstractThis study aimed to determine whether mRNA and protein levels of cartilage oligomeric matrix protein (COMP), a glycoprotein responsible for modulating homeostasis of extracellular matrix, in the systemic and local liver environments were associated with clinical parameters of biliary atresia (BA) patients and might serve as a biomarker for BA severity. COMP protein levels in the circulation of 96 BA patients and 56 healthy controls and its mRNA and protein expressions in the liver of 20 BA patients and 5 non-BA patients were evaluated using enzyme-linked immunosorbent assay, real-time polymerase chain reaction, and immunohistochemistry, respectively. In the circulation of BA patients, COMP levels were significantly higher than those in healthy controls. Compared with early-stage BA patients, those with advanced-stage including jaundice, fibrosis, and hepatic dysfunction had significantly increased circulating COMP levels. Raised circulating COMP levels were found to be independently correlated with degree of liver fibrosis. Survival analysis showed that elevated circulating COMP levels were significantly associated with decreased survival of BA patients. Receiver-operating characteristic curve analysis unveiled a diagnostic value of circulating COMP as a non-invasive biomarker of BA (AUC = 0.99), with a sensitivity of 100.0% and a specificity of 98.2%. In the liver, both COMP mRNA and protein expressions of BA patients with fibrosis were significantly greater than those of BA patients without fibrosis and non-BA patients. Collectively, increased circulating COMP might reflect unfavorable outcome of BA patients and have potential as a novel biomarker for the disease severity following Kasai-operation.


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