scholarly journals Serological responses to a soluble recombinant circumsporozoite protein-VK210 of Plasmodium vivax (rPvCSP-VK210) among Iranian malaria patients

2021 ◽  
Vol 26 (1) ◽  
Author(s):  
Mehdi Nateghpour ◽  
Soudabeh Etemadi ◽  
Afsaneh Motevalli Haghi ◽  
Hamid Eslami ◽  
Mehdi Mohebali ◽  
...  

Abstract Background Circumsporozoite protein (CSP) has a central immune domain that includes short regions of repeating amino acid sequences. This immunodynamic region is an epitope of B cells that can elicit an immune response in human and laboratory animals. The aim of the present study was to express the recombinant PvCSP-VK210 antigen and evaluate it for assaying antibodies obtained during human P. vivax infection by Western blotting and indirect ELISA (enzyme-linked immunosorbent assay). Method Genomic DNA of P. vivax was isolated from a blood sample of an Iranian person with vivax malaria, and by PCR, the fragment of the PvCSP-VK210 gene was amplified. The gene fragment was cut after gel purification by BamHI and HindIII enzymes and then cloned into pET28a expression vector. Finally, the recombinant pET28a was transformed into the E.coli BL21 (DE3) as the expression host. In order to produce His-tagged protein, the expression host was cultured in LB medium. The protein was purified by Ni–NTA columns and immobilized metal affinity chromatography, and after confirmation by Western blotting technique, was used as the antigen in the indirect ELISA test. Results The recombinant protein was expressed and purified as a 32-kDa protein. The sensitivity and specificity of the indirect ELISA test with the recombinant PvCSP-VK210 antigen were 61.42% and 97.14%, respectively, based on OD  =  0.313. Between the results of the microscopic test and the indirect ELISA test with the recombinant PvCSP-VK210 antigen there was a Kappa coefficient of 0.586. The positive and negative predictive value and validity of the ELISA test with the recombinant PvCSP-VK210 antigen were 95.55%, 71.57%, 79.28%, respectively. Conclusion The sensitivity of the indirect ELISA method with the recombinant PvCSP-VK210 antigen was 61.42%, which is the first report from Iran.

2022 ◽  
Vol 98 (6) ◽  
pp. 648-656
Author(s):  
G. M. Ignatyev ◽  
I. A. Leneva ◽  
A. V. Atrasheuskaya ◽  
L. I. Kozlovskaya ◽  
N. P. Kartashova ◽  
...  

Introduction. In clinical practice, the differential diagnosis of COVID-19 can be challenging during the flu season, entailing serious consequences such as delays in appropriate control measures against the SARS-CoV-2 pandemic. Another problem is posed by co-infection of SARS-CoV-2 and influenza virus (IV), which significantly contributes to the severity of the COVID-19 disease. This study was aimed to explore the cross-impact of co-administration of Russian influenza and COVID-19 vaccines on development of specific immunity in laboratory animals.Materials and methods. The study was conducted on BALB/c mice. The animals were inoculated intramuscularly with the vaccine for COVID-19 prevention (CoviVac) and the vaccine for influenza prevention (Flu-M). The sera from the immunized animals were examined separately. Three IV strains were used in the hemagglutination inhibition assay. Antibodies (Abs) against SARS-CoV-2 were detected by an enzyme-linked immunosorbent assay (ELISA). The neutralization test was performed to detect virus neutralizing antibodies against SARS-CoV-2 and IV.Results. Relatively high titers of specific Abs were found in the groups of animals inoculated with one vaccine and with two vaccines concurrently. In the groups of animals inoculated with CoviVac and with two vaccines concurrently, both in the ELISA test and in the neutralization test, the average titers of specific Abs against SARSCoV- 2 did not demonstrate any statistical difference. The group of animals inoculated concurrently with two vaccines demonstrated statistically higher titers of Abs against IV after the second immunization compared to the group of animals inoculated with Flu-M.Discussion. The study has shown that post-vaccination immunity both to IV and to SARS-CoV-2 develops after co-vaccination with two vaccines. The observed enhanced post-vaccination immune response to IV in the coimmunized laboratory animals needs further research.Conclusion. The performed studies suggest the possibility of co-administration of two vaccines to prevent influenza and COVID-19.


2020 ◽  
Vol 14 (3) ◽  
pp. 698-705
Author(s):  
Alima Maïna ◽  
Abdoulkarim Issa Ibrahim ◽  
Abdou Alassane ◽  
Hassane Adakal

La distribution et la dynamique des populations des tiques est un élément clé dans la connaissance des maladies transmises par ces vecteurs. C’est ainsi que cette étude a été conduite afin de mieux connaître l’épidémiologie de la Fièvre Hémorragique de Crimée-Congo (FHCC) dans les 8 communes du département de Boboye au Niger, où 355 sérums de bovins ont été collectés. En plus des sérums, des tiques ont été collectées sur 144 bovins, soit 18 par commune. Les sérums ont été soumis à un test ELISA (Enzyme Linked Immunosorbent Assay) indirect pour la détection d’anticorps anti-FHCC. Soixante-douze (72) éleveurs ont été interviewés sur leur connaissance de l’écologie des tiques, vecteurs du virus de la FHCC. Les résultats de l’enquête ont révélé que les éleveurs n’ont pas recours aux acaricides et que, dans leur majorité (55/72 soit 76,4 %), ils pratiquent la transhumance. L’étude a permis l’identification de 1342 tiques réparties en trois genres : Hyalomma (91,7%), Amblyomma (5,7%) et Rhipicephalus (Boophilus) (2,6%). La séroprévalence globale a été de 9,1±0,03%. Les communes de Harikanassou et Kiota ont été celles où les fortes prévalences ont été observées de 26,7 ± 12,9% et 22,5 ±12,9%. Le virus de la FHCC est en circulation chez la population animale, alors des investigations doivent être faites chez la population humaine.Mots clés : Anticorps anti-FHCC, Enzyme Linked Immunosorbent Assay Indirecte, Prévalence, Sérums, Tiques.   English Title: Crimean-Congo Hemorrhagic Fever (CCHF) ’s Epidemiology in cattle in Boboye’s department of Niger Republic To understand disease transmission by ticks, knowledge of population dynamics and distribution of these vectors are essentials. To sought that, the epidemiology of Crimean-Congo Hemorrhagic Fever (CCHF) in Niger Republic was studied by sampling 355 bovines (sera and ticks) in eight (8) local governments in Boboye’s department. Eighteen (18) bovines were sampled for ticks collection per local government making them a total of 144 bovine. Indirect ELISA test (enzyme-linked immunosorbent assay) was used to detect anti- CCHF antibodies. Seventy-two (72) farmers were surveyed on their knowledge on ticks’ ecology, main vectors of CCHF virus. The results revealed that farmers are not using acaricides, and their majority (55/72 thus 76.4%) practice Transhumance. The study allowed the identification of 1342 ticks distributed in 3 genus: Hyalomma (91.7%), Amblyomma (5.7%) and Rhipicephalus (Boophilus) (2.6%). The global seroprevalence against CCHF was (9.1 ± 0.03) %. Harikanassou and Kiota were the most affected local governments with respectively (26.7±12.9) % and (22.5±12,9) % prevalence. CCHV virus is circulating in animal population, so investigations must be made in human population. Keywords: Anti-CCHF antibodies, Indirect Enzyme Linked Immunosorbent Assay, Prevalence, Sera, Ticks.


Animals ◽  
2019 ◽  
Vol 9 (8) ◽  
pp. 548 ◽  
Author(s):  
Muhammad Ali-ul-Husnain Naqvi ◽  
Sana Zahra Naqvi ◽  
Muhammad Ali Memon ◽  
Kalibixiati Aimulajiang ◽  
Muhammad Haseeb ◽  
...  

Haemonchus contortus is recognized as one of the important health problems in small ruminants, leading to reduced production and economic loss for farmers worldwide. Prepatent diagnosis of H. contortus infection is crucial to improve control strategies as this helminth may remove up to one-fifth of total erythrocytes and may cause anemia, edema, diarrhea, and ultimately death in young animals. In this study, one of the excretory and secretory products, rHc-HCA59, was purified and used as antigen to detect specific antibodies in H. contortus infected goats during prepatent stage of infection using indirect enzyme linked immunosorbent assay (ELISA) as screening test. All goats (n = 38) were housed indoor, experimentally infected with 8000 infective larvae (L3) of H. contortus, and serum samples were collected prior to infection and at 14th day of infection. Immunoblotting was performed to confirm the results of indirect ELISA, evaluate the cross reactivity against rHc-HCA59 in sera of most common co-infecting parasites and rectify the false negative samples. Furthermore, three different batches of rHc-HCA59 were produced to evaluate the repeatability of ELISA. No eggs were detected in feces of all goats collected at 7th and 14th day of infection but, H. contortus eggs were detected at 21 days post infection in the feces. Indirect ELISA performed in this study showed 87% sensitivity and 100% specificity. The western blot analysis confirmed immunoreactivity in serum samples which scored positive in indirect ELISA and recognized the samples as negative which had OD450 lower than negative cut-off value in indirect ELISA. Furthermore, all false negative sera (n = 5) that had OD450 value between positive and negative cut-off value in rHc-HCA59 based ELISA were clearly positive in western blot. Moreover, no cross-reactivity was detected in ELISA and western blotting against rHc-HCA59 in positive sera of Toxoplasma gondii, Fasciola hepatica, and Trichinella spiralis. The results of this study concluded that combined use of indirect ELISA and western blotting with rHc-HCA59 is a potential immunodiagnostic tool for the detection of H. contortus infection during prepatent period in goats.


2014 ◽  
Vol 2014 ◽  
pp. 1-6 ◽  
Author(s):  
Harisankar Singha ◽  
Praveen Malik ◽  
Sachin K. Goyal ◽  
Sandip K. Khurana ◽  
Chiranjay Mukhopadhyay ◽  
...  

Objective. To express truncated TssB protein ofBurkholderia malleiand to evaluate its diagnostic efficacy for serological detection of glanders among equines.Materials and Methods. In an attempt to develop recombinant protein based enzyme-linked immunosorbent assay (ELISA), N-terminal 200 amino acid sequences ofB. malleiTssB protein—a type 6 secretory effector protein—were expressed in prokaryotic expression system. Diagnostic potential of recombinant TssB protein was evaluated in indirect ELISA using a panel of glanders positive (n=49), negative (n=30), and field serum samples (n=1811). Cross-reactivity of the assay was assessed with equine disease control serum and human melioidosis positive serum.Results. In comparison to CFT, diagnostic sensitivity and specificity of ELISA were 99.7% and 100%, respectively.Conclusions. The indirect ELISA method using the truncated TssB offered safer and more rapid and efficient means of serodiagnosis of glanders in equines. These data highlight the use of TssB as potential diagnostic antigen for serological diagnosis of glanders.


2010 ◽  
Vol 9 (3) ◽  
pp. 128
Author(s):  
K. M. Al-Saad ◽  
Saad Hashim Al-Husseiny

The objective of this study was to investigate Toxoplasma gondii antibodies among sheep in different regions of Basrah province (including Al-Mdayna, Shatt Al-Arab, Al-Basrah, Al-Zubayr, and Abu Al-Khasib). The study was started in Oct. 2008 and was finished in May 2009, using latex agglutination test (LAT) and indirect enzyme linked immunosorbent assay (ELISA) IgG, 309 adult sheep were randomly selected from 15 herds among different ages and both sexes and used in this study, including 62 pregnant ewes, 185 non-pregnant ewes, 14 aborted ewes, and 48 rams. Results showed, that 60.84% were seropositive by LAT, whereas 51.11% were seropositive by ELISA IgG test, among animals used in this study, results detected that 79.03% pregnant ewes (highest value), 56.75% nonpregnant ewes,71.40% aborted ewes and 50% Rams (lowest value) were seropositive by LAT, whereas 56.52% pregnant ewes, 51.11% non-pregnant ewes, 83.33% aborted ewes (highest value), and 31.25% Rams (lowest value) were seropositive by indirect ELISA IgG. Moreover, among regions of Basrah province, the details of percentage of T.gondii antibodies were 54.54% in AL-Basrah , 71.43% in Abu Al-Khasib (highest value), 57.35% in Al- Mdayna, 47.83% in Shatt Al-Arab (lowest value), and 67.16% in Al-Zubayr by LAT, whereas 63.64% in AL-Basrah (highest value), 22.73% in Abu Al-Khasib (lowest value), 57.89% in Al-Mdayna, 50% in Shatt Al-Arab and 61.90% in Al-Zubayr by indirect ELISA test. Although the difference observed in the percentage of Toxoplasma gondii antibodies among different regions of Basrah, there was no significant difference P>0.05 detected LAT, whereas in the indirect ELISA IgG test there was significant difference P<0.05. Ewes showed high percentages 62.83%, 55.40% of toxoplasmosis than rams 50 %, 31.25% by LAT and ELISA test respectively. The highest titer was 1/4 28.57% were detected in pregnant ewes and lowest titers were 1/2, 1/8, and 1/256 0.0% were detected in aborted ewes and in ramsrespectively.


2020 ◽  
Author(s):  
Charoonluk Jirapattharasate ◽  
Ruenruetai Udonsom ◽  
Apichai Prachasuphap ◽  
Kodcharad Jongpitisub ◽  
Panadda Dhepakson

Abstract Background The development of sensitive and specific methods for detecting Toxoplasma gondii infection is critical for preventing and controlling toxoplasmosis in humans and other animals. Recently, various recombinant proteins have been used in serological tests for diagnosing toxoplasmosis. The production of these antigens is associated with live tachyzoites obtained from cell cultures or laboratory animals for genomic extraction to amplify target genes. Synthetic genes have gained a key role in recombinant protein production. For the first time, we demonstrated the production of the recombinant protein of the T. gondii dense granular antigen 8 (TgGRA8) gene based on commercial gene synthesis. Recombinant TgGRA8 plasmids were successfully expressed in an Escherichia coli system. The recombinant protein was affinity-purified and characterized via sodium dodecyl sulfate-polyacrylamide gel electrophoresis and Western blotting. Furthermore, the diagnostic potential of the recombinant protein was assessed using 306 field serum samples from goats via indirect enzyme-linked immunosorbent assay (iELISA) and the latex agglutination test (LAT).Results Western blotting using known positive serum samples from goats identified a single antigen at the expected molecular weight of TgGRA8 (27 kDa). iELISA illustrated that 15.40% of goat samples were positive for T. gondii-specific IgG antibodies. In addition, TgGRA8 provided high sensitivity and specificity, with significant concordance (91.83) and kappa values (0.69) compared with the results obtained using LAT.Conclusion Our findings highlight the production of a recombinant protein from a synthetic TgGRA8 gene and the ability to detect T. gondii infection in field samples. The sensitivity and specificity of TgGRA8 demonstrated that this protein could be a good serological marker for detecting specific IgG in goat sera


2015 ◽  
Vol 7 (01) ◽  
pp. 017-020 ◽  
Author(s):  
Reza Shafiei ◽  
Bahador Sarkari ◽  
Seyed Mahmoud Sadjjadi

ABSTRACT Background: Serological diagnosis, based on antigenic fractions of the parasite can be used for the early diagnosis of human fascioliasis. The current study aimed to evaluate the efficacy of a 27 kDa immunodominant antigen of Fasciola hepatica adult worms, in an indirect enzyme-linked immunosorbent assay (ELISA) system for serological diagnosis of human fascioliasis. Materials and Methods: The immunodiagnosis of human fascioliasis, using a 27 kDa immunodominant antigen, purified from F. hepatica somatic antigens (SAs), was evaluated by Western blotting and ELISA with sera samples of human fascioliasis patients, healthy controls and patients with other parasitic infections. Results: Using western blotting, from 12 sera of fascioliasis patients, 11 sera (91.6%) detected the 27 kDa subunit. None of 30 samples from healthy controls or 32 sera from nonfascioliasis patients reacted with the 27 kDa antigen. Accordingly, sensitivity and specificity of the system was found to be 91.6% and 100%, respectively. The 27 kDa antigen was purified from the SAs and was used in an indirect ELISA system. Of 15 sera of fascioliasis patients, all (100%) were found to be positive by ELISA whereas only 4 cases (6.25%) of nonfascioliasis patients or healthy controls were false-positive by this system. Accordingly, the sensitivity and specificity of the test were 100% and 93.6%, respectively. Conclusion: Findings of this study demonstrated that both Western blotting and the indirect ELISA, based on the 27 kDa subunit of F. hepatica SA, are reliable methods for serodiagnosis of human fascioliasis.


2020 ◽  
Author(s):  
Charoonluk Jirapattharasate ◽  
Ruenruetai Udonsom ◽  
Apichai Prachasuphap ◽  
Kodcharad Jongpitisub ◽  
Panadda Dhepakson

Abstract Background The development of sensitive and specific methods for detecting Toxoplasma gondii infection is critical for preventing and controlling toxoplasmosis in humans and other animals. Recently, various recombinant proteins have been used in serological tests for diagnosing toxoplasmosis. The production of these antigens is associated with live tachyzoites obtained from cell cultures or laboratory animals for genomic extraction to amplify target genes. Synthetic genes have gained a key role in recombinant protein production. For the first time, we demonstrated the production of the recombinant protein of the T. gondii dense granular antigen 8 (TgGRA8) gene based on commercial gene synthesis. Recombinant TgGRA8 plasmids were successfully expressed in an Escherichia coli system. The recombinant protein was affinity-purified and characterized via sodium dodecyl sulfate-polyacrylamide gel electrophoresis and Western blotting. Furthermore, the diagnostic potential of the recombinant protein was assessed using 306 field serum samples from goats via indirect enzyme-linked immunosorbent assay (iELISA) and the latex agglutination test (LAT).Results Western blotting using known positive serum samples from goats identified a single antigen at the expected molecular weight of TgGRA8 (27 kDa). iELISA illustrated that 15.40% of goat samples were positive for T. gondii-specific IgG antibodies. In addition, TgGRA8 provided high sensitivity and specificity, with significant concordance (91.83) and kappa values (0.69) compared with the results obtained using LAT.Conclusion Our findings highlight the production of a recombinant protein from a synthetic TgGRA8 gene and the ability to detect T. gondii infection in field samples. The sensitivity and specificity of TgGRA8 demonstrated that this protein could be a good serological marker for detecting specific IgG in goat sera.


2010 ◽  
Vol 17 (12) ◽  
pp. 1926-1932 ◽  
Author(s):  
Emily N. Barker ◽  
Chris R. Helps ◽  
Kate J. Heesom ◽  
Christopher J. Arthur ◽  
Iain R. Peters ◽  
...  

ABSTRACT Hemoplasmas is the trivial name given to a group of erythrocyte-parasitizing bacteria of the genus Mycoplasma. Of the feline hemoplasmas, Mycoplasma haemofelis is the most pathogenic, while “Candidatus Mycoplasma haemominutum” and “Candidatus Mycoplasma turicensis” are less pathogenic. Shotgun libraries of fragmented M. haemofelis genomic DNA were constructed, and random colonies were selected for DNA sequencing. In silico-translated amino acid sequences of putative open reading frames were compared to mass spectrometry data from M. haemofelis protein spots identified as being immunogenic by two-dimensional gel electrophoresis and Western blotting. Three of the spots matched the predicted sequences of a heat shock protein 70 (DnaK) homolog, elongation factor Ts, and a fragment of phosphoglycerate kinase found during library screening. A full-length copy of the M. haemofelis dnaK gene was cloned into Escherichia coli and recombinantly expressed. Recombinant M. haemofelis DnaK was purified and then used in Western blotting and an enzyme-linked immunosorbent assay (ELISA) to investigate the humoral immune response during acute infection in cats experimentally infected with M. haemofelis, “Ca. Mycoplasma haemominutum,” or “Ca. Mycoplasma turicensis”. The recombinant M. haemofelis DnaK ELISA also was used to screen clinical samples submitted for hemoplasma PCR testing to a commercial laboratory (n = 254). Experimentally infected cats became seropositive following infection, with a greater and earlier antibody response seen in cats inoculated with M. haemofelis than those seen in cats inoculated with “Ca. Mycoplasma haemominutum” or “Ca. Mycoplasma turicensis,” by both Western blotting and ELISA. Of the clinical samples, 31.1% had antibodies detected by the ELISA but only 9.8% were positive by PCR for one or more hemoplasmas.


1992 ◽  
Vol 54 (2) ◽  
pp. 395-397
Author(s):  
Shin-ichi KAMATA ◽  
Atsushi OHKAWA ◽  
Osamu ITO ◽  
Norihide KAKIICHI ◽  
Kenichi KOMINE ◽  
...  

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