scholarly journals A glycoengineered antigen exploiting a conserved protein O-glycosylation pathway in the Burkholderia genus for diagnosis of glanders infections

2020 ◽  
Author(s):  
Guanbo Wang ◽  
Lena Glaser ◽  
Nichollas E. Scott ◽  
Yasmine Fathy Mohamed ◽  
Rebecca Ingram ◽  
...  
Keyword(s):  
Diagnostic Tool ◽  
Complement Fixation ◽  
Serological Test ◽  
Animal Health ◽  
Cholera Toxin B Subunit ◽  
Burkholderia Mallei ◽  
Serum Samples ◽  
Western Blots ◽  
Glycoprotein Antigen ◽  
Glycosylation Pathway

ABSTRACTWe recently described a protein O-glycosylation pathway conserved in all species of the Burkholderia genus that results in synthesis and incorporation of a trisaccharide glycan to membrane-exported proteins. Here, we exploited this system to construct and evaluate a diagnostic tool for glanders. Burkholderia mallei, the causative agent of glanders, is a highly infectious and fatal zoonotic pathogen that mainly infects horses, mules, donkeys and occasionally humans. A highly sensitive and specific diagnostic tool is crucial for the control, elimination and eradication of B. mallei infections. We constructed plasmids carrying synthetic genes encoding a modified, previously unannotated Burkholderia glycoprotein containing three glycosylation sequons fused to the cholera toxin B-subunit. The resulting proteins were glycosylated in the B. cenocepacia K56-2 parental strain, but not in glycosylation-deficient mutants, as determined by SDS-PAGE and fluorescent lectin blots. One of these glycoproteins was used as an antigen in ELISA and western blots to screen a panel of serum samples collected from glanders-infected and healthy horses previously investigated by complement fixation test and indirect ELISA based on a semi-purified fraction of B. mallei. We show that ELISA and western blot assays based on our glycoprotein antigen provide 100 % specificity, with a sensitivity greater than 88%. The glycoprotein antigen was recognized by serum samples collected from patients infected with B. pseudomallei, B. mallei, B. multivorans and B. cenocepacia. Our results indicate that protein O-glycosylation in Burkholderia can be exploited as a biomarker for diagnosis of Burkholderia-associated infections.IMPORTANCEGlanders is a severe zoonotic disease caused by the Gram-negative bacterium Burkholderia mallei, which affects horses, mules and donkeys, as well as humans. B. mallei is also considered a category B biothreat agent. Due to insufficient pathognomonic symptoms in the early stages of glanders, diagnosis can be difficult. Complement fixation is the most accurate and reliable serological test prescribed by the World Organization for Animal Health; however, this test has a considerable number of false-positive results. We have recently described a conserved protein O-glycosylation pathway present in all species of the Burkholderia genus; we also demonstrated that Burkholderia-infected humans develop anti-glycan antibodies. Here, we exploited this system to construct and evaluate a synthetic glycoengineered protein antigen as a diagnostic tool for glanders. Our results show 100 % specificity in the detection of antibodies from infected horses, indicating that protein O-glycosylation in Burkholderia can be exploited as a biomarker for diagnosis of Burkholderia-associated infections.

scholarly journals A Century of syphilis serology

2007 ◽  
Vol 28 (1) ◽  
pp. 25
Author(s):  
Peter W Robertson
Keyword(s):  
Complement Fixation Test ◽  
Complement Fixation ◽  
Congenital Syphilis ◽  
Serological Test ◽  
Serological Diagnosis ◽  
Serological Tests ◽  
Western Blots ◽  
Comprehensive Review ◽  
The Subject ◽  
Syphilis Serology

The year 2006 represented the centenary of the first diagnostic serological test, the Wasserman complement fixation test for the diagnosis of syphilis. In this article I will relate some developments I have observed in the day-to-day running of a diagnostic serology laboratory over the thirty years preceding this centenary. I have assumed some background knowledge and elected to discuss only selected aspects of serological tests for syphilis (STS), rather than attempt a comprehensive review of the subject. A number of areas such as the serological diagnosis of congenital syphilis, the use of and interpretation of western blots in ?problem? sera and diagnosis of other treponemal diseases have not been included.

Rapid detection of Chlamydia/Chlamydophila group in samples collected from swine herds with and without reproductive disorders

2014 ◽  
Vol 17 (2) ◽  
pp. 367-369 ◽  
Author(s):  
K. Rypula ◽  
A. Kumala ◽  
P. Lis ◽  
K. Niemczuk ◽  
K. Płoneczka-Janeczko ◽  
...  
Keyword(s):  
Real Time ◽  
Real Time Pcr ◽  
Rapid Detection ◽  
Complement Fixation Test ◽  
Complement Fixation ◽  
Fixation Test ◽  
Reproductive Disorders ◽  
Serum Samples ◽  
Swine Herds

Abstract The study was carried out in seven reproductive herds of pigs. In three of them reproductive disorders were observed. Three herds consisted of 10-50 and four consisted of 120-500 adult sows and they were called small and medium, respectively. Fifty-seven adult sows were randomly selected from herds. Serum samples were tested using the complement fixation test and swabs from both eyes and from the vaginal vestibule were examined using real-time PCR. All serum samples were negative. Infected sows were present in each of the study herds. In total, there were 28 positive samples (53%, 28/48) in real-time PCR in sows with reproductive disorders and 35 (53%, 35/66) in sows selected from herds without problems in reproduction. One isolate proved to be Chlamydophila pecorum, whereas all the remaining were Chamydia suis

scholarly journals Seropositivity to canine tick-borne pathogens in a population of sick dogs in Italy

2021 ◽  
Vol 14 (1) ◽  
Author(s):  
Jairo Alfonso Mendoza-Roldan ◽  
Giovanni Benelli ◽  
Marcos Antonio Bezerra-Santos ◽  
Viet-Linh Nguyen ◽  
Giuseppe Conte ◽  
...  
Keyword(s):  
Borrelia Burgdorferi ◽  
Serological Test ◽  
Control Strategies ◽  
Central Italy ◽  
Rickettsia Conorii ◽  
High Seroprevalence ◽  
The South ◽  
Serum Samples ◽  
Serological Tests ◽  
Distribution Maps

Abstract Background Canine vector-borne diseases (CVBDs) associated to ticks are among the most important health issues affecting dogs. In Italy, Ehrlichia canis, Anaplasma spp., Rickettsia conorii and Borrelia burgdorferi (s.l.) have been studied in both healthy canine populations and those clinically ill with suspected CVBDs. However, little information is currently available on the overall prevalence and distribution of these pathogens in the country. The aim of this study was to assess the prevalence and distribution of tick-borne pathogens (TBPs) in clinically suspect dogs from three Italian macro areas during a 15-year period (2006–2020). Methods A large dataset (n = 21,992) of serological test results for selected TBPs in three macro areas in Italy was analysed using a Chi-square test to evaluate the associations between the categorical factors (i.e. macro area, region, year, sex and age) and a standard logistic regression model (significance set at P = 0.05). Serological data were presented as annual and cumulative prevalence, and distribution maps of cumulative positive cases for TBPs were generated. Results Of the tested serum samples, 86.9% originated from northern (43.9%) and central (43%) Italy. The majority of the tests was requested for the diagnosis of E. canis (47%; n = 10,334), followed by Rickettsia spp. (35.1%; n = 7725), B. burgdorferi (s.l.) (11.6%; n = 2560) and Anaplasma spp. (6.2%; n = 1373). The highest serological exposure was recorded for B. burgdorferi (s.l.) (83.5%), followed by Rickettsia spp. (64.9%), Anaplasma spp. (39.8%) and E. canis (28.7%). The highest number of cumulative cases of Borrelia burgdorferi (s.l.) was recorded in samples from Tuscany, central Italy. Rickettsia spp. was more prevalent in the south and on the islands, particularly in dogs on Sicily older than 6 years, whereas Anaplasma spp. was more prevalent in the north and E. canis more prevalent in the south and on the islands. Conclusions The results of this study highlight the high seroprevalence and wide distribution of the four TBPs in dogs with clinically suspected CVBDs from the studied regions of Italy. The very high seroprevalence of B. burgdorferi (s.l.) exemplifies a limitation of this study, given the use of clinically suspect dogs and the possibility of cross-reactions when using serological tests. The present research provides updated and illustrative information on the seroprevalence and distribution of four key TBPs, and advocates for integrative control strategies for their prevention. Grapic abstract

scholarly journals Sero-epidemiological survey of bluetongue disease in one-humped camel (Camelus dromedarius) in Kassala State, Eastern Sudan

2021 ◽  
Vol 74 (1) ◽  
Author(s):  
Molhima M. Elmahi ◽  
Mohammed O. Hussien ◽  
Abdel Rahim E. Karrar ◽  
Amira M. Elhassan ◽  
Abdel Rahim M. El Hussein
Keyword(s):  
Risk Factors ◽  
Potential Risk ◽  
Animal Health ◽  
Water Source ◽  
Economic Losses ◽  
Enzyme Linked Immunosorbent Assay ◽  
Serum Samples ◽  
Cross Sectional ◽  
Potential Risk Factors ◽  
Eastern Sudan

Abstract Background Bluetongue (BT) is a vector-borne viral disease of ruminant and camelid species which is transmitted by Culicoides spp. The causative agent of BT is bluetongue virus (BTV) that belongs to genus Orbivirus of the family Reoviridae. The clinical disease is seen mainly in sheep but mostly sub-clinical infections of BT are seen in cattle, goats and camelids. The clinical reaction of camels to infection is usually not apparent. The disease is notifiable to the World Organization for Animal Health (OIE), causing great economic losses due to decreased trade and high mortality and morbidity rates associated with bluetongue outbreaks. The objective of this study was to investigate the seroprevalence of BTV in camels in Kassala State, Eastern Sudan and to identify the potential risk factors associated with the infection. A cross sectional study using a structured questionnaire survey was conducted during 2015–2016. A total of 210 serum samples were collected randomly from camels from 8 localities of Kassala State. The serum samples were screened for the presence of BTV specific immunoglobulin (IgG) antibodies using a competitive enzyme-linked immunosorbent assay (cELISA). Results Seropositivity to BTV IgG was detected in 165 of 210 camels’ sera accounting for a prevalence of 78.6%. Potential risk factors to BTV infection were associated with sex (OR = 0.061, p-value = 0.001) and seasonal river as water source for drinking (OR = 32.257, p-value = 0.0108). Conclusions Sex and seasonal river as water source for drinking were considered as potential risk factors for seropositivity to BTV in camels. The high prevalence of BTV in camels in Kassala State, Eastern Sudan, necessitates further epidemiological studies of BTV infection in camels and other ruminant species to better be able to control BT disease in this region.

scholarly journals Brucella abortus RB51 and Hot Saline Extract from Brucella ovis as Antigens in a Complement Fixation Test Used To Detect Sheep Vaccinated with Brucella abortus RB51

2001 ◽  
Vol 8 (1) ◽  
pp. 119-122 ◽  
Author(s):  
Rosanna Adone ◽  
Franco Ciuchini
Keyword(s):  
Immune Responses ◽  
Brucella Abortus ◽  
Complement Fixation Test ◽  
Complement Fixation ◽  
Infected Sheep ◽  
Fixation Test ◽  
Anticomplementary Activity ◽  
Serum Samples ◽  
Saline Extract ◽  
Brucella Ovis

ABSTRACT The efficacy of Brucella abortus RB51 and hot saline extract (HSE) from Brucella ovis as antigens in complement fixation (CF) tests was comparatively evaluated in detecting immune responses of sheep vaccinated with B. abortus strain RB51. For this study, four 5-month-old sheep were vaccinated subcutaneously with 5 × 109 CFU of RB51, and two sheep received saline. Serum samples collected at different times after vaccination were tested for the presence of antibodies to RB51 by a CF test with RB51 as antigen, previously deprived of anticomplementary activity, and with HSE antigen, which already used as the official antigen to detectB. ovis-infected sheep. The results showed that vaccinated sheep developed antibodies which reacted weakly against HSE antigen and these antibodies were detectable for 30 days after vaccination. However, antibodies to RB51 could be detected for a longer period after vaccination by using homologous RB51 antigen in CF tests. In fact, high titers were still present at 110 days postvaccination with RB51 antigen. Sera from sheep naturally infected with B. ovisalso reacted to RB51 but gave lower titers than those detected by HSE antigen. As expected, all sera from RB51-vaccinated sheep remained negative when tested with standard S-type Brucella standard antigens.

scholarly journals Seroprevalence of infectious bursal disease and its potential risk factors in backyard chicken production of Waliso district, South Western Shoa Zone, Ethiopia

2021 ◽  
Vol 20 (1) ◽  
pp. 95-105
Author(s):  
Chala Bedasa ◽  
◽  
Ararsa Duguma ◽  
Asamenew Tesfaye ◽  
Tadele Tolosa ◽  
...  
Keyword(s):  
Risk Factors ◽  
Potential Risk ◽  
Animal Health ◽  
Infectious Bursal Disease ◽  
Flock Size ◽  
Serum Samples ◽  
Significant Difference ◽  
Bursal Disease ◽  
Potential Risk Factors ◽  
Backyard Chicken

A cross sectional study on infectious bursal disease was conducted in apparently healthy backyard chicken at Waliso district of Southwestern Shoa, central oromia, Ethiopia from from November, 2018 to October, 2019. A total of 282 chickens were randomly selected to estimate seroprevalence of IBD infection and to identify the likely potential risk factors for the disease. Serum samples collected and serological test conducted in laboratory at National Animal Health Diagnosis and Investigation Center Sebeta, Ethopia. Out of 282 serum samples tested 224 were positive for indirect ELISA technique and the overall seroprevalence of IBDV in the study area was found to be 79.43% at individual level. Educational level of owners, kebeles and flock size significantly affect seroprevalence of IBD in the study area. The effect of difference in managements like source of replacement, frequency of house cleaning, use of disinfectant and isolation practice has a significant effect on IBDV sero-prevalence. A lower seroprevalence of IBDV was reported in good hygienic level of house (26.7%) than poor level of chicken house hygiene (96.4%) with statistically significant difference (P < 0.05). The seroprevalence of IBDV in the present study associated with chicken management, flock size, owner education level and other animal related risk factors for occurrence of the disease. Therefore, awareness on chicken health management, and importance of immunization would help to minimize the prevalence of the disease and play crucial role in the control of the disease. Furthermore, characterizing virus strains circulating in the area in future study is recommended.

scholarly journals Experimental determination of the pharmacokinetic properties of trimethoprim and sulfamethoxazole combination in the blood serum of broiler chickens

2021 ◽  
Author(s):  
K Putecova ◽  
K Nedbalcova ◽  
I Bartejsova ◽  
M Zouharova ◽  
K Matiaskova ◽  
...  
Keyword(s):  
Oral Administration ◽  
Broiler Chickens ◽  
Animal Health ◽  
Pharmacokinetic Parameters ◽  
Serum Samples ◽  
Commercial Preparation ◽  
Single Oral Administration ◽  
Thermo Fisher Scientific ◽  
Medicinal Substances

A rapid, simple and highly efficient analytical method for the targeted determination of trimethoprim and sulfamethoxazole in serum samples has been developed and used to measure the pharmacokinetic curve of these medicinal substances after administration to chicken broilers. The pharmacokinetics properties of trimethoprim and sulfamethoxazole were investigated in clinically healthy broiler chickens after the single oral administration of the commercial preparation Methoxasol (Eurovet Animal Health, B.V., The Netherlands) at a dose of 0.275 ml/kg b.w. After a single dose drug administration, the chickens were sacrificed by decapitation under general anaesthesia by Isoflurin 1 000 mg/g (Vetpharma AH, Spain) and the blood was collected at precisely defined intervals: 15, 30, 45, 60, 90, 120, 180, 360 and 720 min after the administration. The serum concentrations of amoxicillin were determined using Q Exactive tandem mass spectrometer (Thermo Fisher Scientific, USA) in conjunction with liquid chromatography. The detected pharmacokinetic parameters of trimethoprim after the oral administration were C<sub>max</sub> = 2.1 ± 1.0 µg/ml; T<sub>max</sub> = 1.5 h; t<sub>½</sub> = 0.88 h; k<sub>el</sub> = 0.009 3 ± 0.001 1 1/h; AUC<sub>t</sub> = 2.901 ± 1.4 µg.h/ml; AUC<sub>∞</sub> = 2.907 ± 1.5 µg.h/ml; V<sub>d</sub> = 2.632 l/kg; Cl = 2.7 l/h. The pharmacokinetic parameters of sulfamethoxazole after the oral administration were C<sub>max</sub> = 47.1 ± 15.3 µg/ml; T<sub>max</sub> = 1 h; t<sub>½</sub> = 1.92 h; k<sub>el</sub> = 0.004 6 ± 0.000 3 1/h; AUC<sub>t</sub> = 89.676 ± 26.9 µg.h/ml; AUC<sub>∞</sub> = 94.612 ± 28.4 µg.h/ml; V<sub>d</sub> = 0.584 l/kg; Cl = 0.21 l/h. To the best of our knowledge, this is the first pharmacokinetic study of the combination of sulfamethoxazole and trimethoprim in broiler chickens.

scholarly journals Quantitative serological evaluation as a valuable tool in the COVID-19 vaccination campaign

2021 ◽  
Vol 0 (0) ◽  
Author(s):  
Davide Ferrari ◽  
Alessandra Mangia ◽  
Maria Sestina Spanò ◽  
Lucia Zaffarano ◽  
Marco Viganò ◽  
...  
Keyword(s):  
Immune Response ◽  
General Population ◽  
Serological Test ◽  
Vaccination Campaign ◽  
Age Groups ◽  
Vaccine Dose ◽  
Serum Samples ◽  
Serological Tests ◽  
Antibody Persistence ◽  
Definition Of

Abstract Objectives After exceptional research efforts, several vaccines were developed against SARS-CoV-2 which sustains the pandemic COVID-19. The Comirnaty vaccine showed high efficacy in clinical trials and was the first to be approved for its distribution to the general population. We evaluated the immune response induced by the first vaccine dose in different sex/age groups and subjects with or without naturally present anti-SARS-CoV-2 antibodies. Methods As part of an Italian multicenter project (Covidiagnostix), serum samples from 4,290 health-professionals were serologically tested the day of the first vaccination dose, and 21 days later, using two different instrumentations (Siemens-Healthineers and Roche). Results In total, 97% of samples showed the presence of specific antibodies 21 days after the vaccination dose; the percentage of non-responders increased with age in both genders. Remarkably, naturally seropositive individuals showed antibody persistence up to 11 months and an exceptionally higher vaccination response compared to subjects never infected by SARS-CoV-2. Conclusions This study highlighted the importance of the serological test i) to identify naturally SARS-CoV-2 seropositive individuals and ii) to evaluate the antibody level elicited by the first vaccination dose. Both tests, highlighted differences in the immune response, when subjects were stratified by sex and age, and between naturally seropositive and seronegative subjects. The data obtained show how serological tests could play a crucial role in the triage of the population subjected to the vaccination campaign for COVID-19. The definition of suitable instrumentation-specific thresholds is needed to correctly follow eventually acquired post-vaccination immunity in the general population.

scholarly journals Development of Immunochromatographic Assay for the Rapid Detection of Mycoplasma capricolum subsp. capripneumoniae Antibodies

2022 ◽  
Vol 12 ◽  
Author(s):  
Zhen Zhu ◽  
Guanggang Qu ◽  
Changjiang Wang ◽  
Lei Wang ◽  
Jige Du ◽  
...  
Keyword(s):  
Immunoglobulin G ◽  
Colloidal Gold ◽  
Complement Fixation Test ◽  
Complement Fixation ◽  
Fixation Test ◽  
Immunochromatographic Assay ◽  
Economic Losses ◽  
Serum Samples ◽  
Mycoplasma Capricolum ◽  
Prokaryotic Expression System

Mycoplasma capricolum subsp. capripneumoniae (Mccp) is the cause of contagious caprine pleuropneumonia (CCPP), which is a highly significant respiratory disease in goats leading to significant economic losses in Africa and Asia. Currently available procedures for the diagnosis of CCPP have some limitations in sensitivity, specificity, operation time, requirement of sophisticated equipment or skilled personnel, and cost. In this study, we developed a rapid, sensitive, and specific colloidal gold-based immunochromatographic assay (GICA) strip for the efficient on-site detection of antibodies against Mccp in the serum within 10 min. For the preparation of this colloidal GICA strip, recombinant P20 protein, the membrane protein of Mccp, was expressed by Escherichia coli prokaryotic expression system after purification was used as the binding antigen in the test. The rabbit anti-goat immunoglobulin G labeled with the colloidal gold was used as the detection probe, whereas the goat anti-rabbit immunoglobulin G was coated on the nitrocellulose membrane as the control line. The concentration of the coating antibody was optimized, and the effectiveness of this colloidal GICA strip was evaluated. Our results proved that the detection limit of the test strip was up to 1:64 dilutions for the Mccp antibody-positive serum samples with no cross-reactivity with other pathogens commonly infecting small ruminants,including goat pox virus, peste des petits ruminants virus, foot-and-mouth disease virus type A, or other mycoplasmas. Moreover, the colloidal GICA strip was more sensitive and specific than the indirect hemagglutination assay for the detection of Mccp antibodies. The 106 clinical serum samples were detected by the colloidal GICA strip compared with the complement fixation test, demonstrating an 87.74% concordance with the complement fixation test. This novel colloidal GICA strip would be an effective tool for the cost-effective and rapid diagnosis of CCPP in the field.

scholarly journals Serodiagnosis of Louse-Borne Relapsing Fever with Glycerophosphodiester Phosphodiesterase (GlpQ) fromBorrelia recurrentis

2000 ◽  
Vol 38 (10) ◽  
pp. 3561-3571 ◽  
Author(s):  
Stephen F. Porcella ◽  
Sandra J. Raffel ◽  
Merry E. Schrumpf ◽  
Martin E. Schriefer ◽  
David T. Dennis ◽  
...  
Keyword(s):  
Serological Test ◽  
Geometric Mean ◽  
Eastern Africa ◽  
Enzyme Linked Immunosorbent Assay ◽  
Relapsing Fever ◽  
Serum Samples ◽  
Serological Tests ◽  
Whole Cell ◽  
Convalescent Phase ◽  
Glycerophosphodiester Phosphodiesterase

Human louse-borne relapsing fever occurs in sporadic outbreaks in central and eastern Africa that are characterized by significant morbidity and mortality. Isolates of the causative agent,Borrelia recurrentis, were obtained from the blood of four patients during a recent epidemic of the disease in southern Sudan. TheglpQ gene, encoding glycerophosphodiester phosphodiesterase, from these isolates was sequenced and compared with the glpQ sequences obtained from other relapsing-fever spirochetes. Previously we showed that GlpQ of Borrelia hermsii is an immunogenic protein with utility as a serological test antigen for discriminating tick-borne relapsing fever from Lyme disease. In the present work, we cloned and expressed theglpQ gene from B. recurrentis and used recombinant GlpQ in serological tests. Acute- and convalescent-phase serum samples obtained from 42 patients with louse-borne relapsing fever were tested with an indirect immunofluorescence assay (IFA) and an enzyme-linked immunosorbent assay (ELISA) that used whole cells ofB. recurrentis and with immunoblotting to whole-cell lysates of the spirochete and Escherichia coli producing recombinant GlpQ. The geometric mean titers of the acute- and convalescent-phase serum samples measured by IFA were 1:83 and 1:575, respectively. The immunoblot analysis identified a high level of reactivity and seroconversion to GlpQ, and the assay was more sensitive than the whole-cell IFA and ELISA using purified, recombinant histidine-tagged GlpQ. Serum antibodies to GlpQ and other antigens persisted for 27 years in one patient. We conclude that assessment of anti-GlpQ antibodies will allow serological confirmation of louse-borne relapsing fever and determination of disease prevalence.

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