Evaluation of three serological tests for the diagnosis of Brucella suis in dogs using Bayesian latent class analysis.

Brucella suis infection of dogs is an emerging issue worldwide requiring specific management to address zoonotic risks and animal welfare concerns. Diagnosis in dogs is routinely based on serological testing, but to date these tests have only been validated for use in production animal species and humans. This study aimed to assess the diagnostic performance of three commonly used serological tests in dogs. Canine sera (n = 989) were tested with the Rose Bengal rapid plate agglutination test (RBRPT), the complement fixation test (CFT) and a competitive ELISA (C-ELISA). Diagnostic test performance was evaluated using a three test, two population Bayesian latent class analysis accounting for conditional dependence between the three tests. Positive and negative predictive values (PPV, NPV) were calculated for a range of expected prevalence estimates for the individual tests and test combinations interpreted in series and parallel. The RBRPT showed the highest individual Se of 0.902 (95 % posterior credible interval [PCI] 0.759-0.978) and the CFT the highest individual diagnostic specificity (Sp) of 0.914 (95 % PCI 0.886-0.946). The C-ELISA had marginally the best overall diagnostic performance (Youden's index = 0.807). The CFT and the C-ELISA interpreted in parallel returned the highest combined Se and Sp (0.988 and 0.885, respectively). All tests returned NPVs of > 0.982 in 2-8 % prevalence settings. Series interpretation of the three-test combination as well as the two-test combinations of the RBRPT and the C-ELISA and the CFT and the C-ELISA produced a PPV of 0.502 when the estimated prevalence was 8 %. While all tests are suitable for the detection of B. suis antibodies in dogs, they should not be interpreted in isolation as their diagnostic value is dependent on the pre-test probability of the disease. As such they are useful tools for the diagnosis of B. suis in dogs when exposure, history and clinical presentation indicate a risk of brucellosis.

Isolation, serological and molecular methods in screening of Burkholderia mallei in East Azerbaijan province, Iran.

Glanders caused by Burkholderia mallei is one of the most dangerous zoonotic diseases in solipeds. Clinical diagnosis of this disease in its early stages in horses, is difficult. This study investigated serological and molecular identification of B. mallei in East Azerbaijan province. In the third and fourth quarters of 2020, throughout 2021, and in the first and second quarters of 2022, the complement fixation test (CFT) was performed on 350 horses. The malleination was used to confirm the positive CFT cases. Blood samples were taken for culture and for preparing serums to perform the enzyme-linked immunosorbent assay (ELISA). Deep eye discharge, nostril, cutaneous ulcers and lymph fluid swabs were cultured, and polymerase chain reaction (PCR) was carried out. Eleven horses were CFT-positive. Based on the malleination on the 11 horses, six were affected by glanders, five were not affected (false positive), and one horse was CFT-negative despite exhibiting clinical signs. It was positive by malleination, ELISA and PCR. A total number of seven positive cases of glanders were diagnosed. The B. mallei could not be isolated, but the Burkholderia cepacia complex was isolated in one case. Except for three cases, the results of the CFT, mallein and ELISA tests were consistent. The amount of confidence interval was 95.00%. It is suggested that ELISA could be used as a complement to CFT in screening and, if positive results are observed in one of the tests, the entire herd must be examined more accurately using the mallein and western blot confirmatory tests.

European Inter-Laboratory Proficiency Test for Dourine Antibody Detection Using the Complement Fixation Test.

Dourine is a sexually transmitted parasitic disease affecting equids. Its causative agent is referred to as Trypanosoma equiperdum and the prescribed serodiagnosis method is the complement fixation test (CFT). In the context of our European Reference Laboratory mandate for equine diseases (excluding African horse sickness), we organised dourine CFT inter-laboratory proficiency tests (ILPTs) in 2015, 2018 and 2022 to evaluate the performance of the European Union network of National Reference Laboratories (NRLs) for dourine. ILPT panels were composed of horse sera with or without antibodies against Trypanosoma spp. originating from non-infected, immunised or experimentally infected horses. Twenty-two NRLs participated in at least one of the three sessions. In 2015, 2018 and 2022, the percentage of laboratories obtaining 100% of the expected results was 57, 90 and 80, respectively. These dourine CFT ILPTs showed the benefits of standardising the method's detection limit and underlined the constant need to evaluate NRLs to improve the network's performance. These results also argue in favour of the need for a representative bio-bank to improve the representativeness of ILPT samples and to allow the adoption of alternative serological methods for international surveillance of dourine.

Brucella abortus Strain RB51 Administered to Prepubescent Water Buffaloes, from Vaccination to Lactation: Kinetics of Antibody Response and Vaccine Safety.

Brucella RB51 is a live modified vaccine. Its use in water buffalo has been proposed using a vaccination protocol different to that used for cattle, but knowledge of the long-term effects of RB51 vaccination in this species remains incomplete. The aim of the study was to evaluate the safety and kinetics of antibody responses in water buffaloes vaccinated according to the protocol described for the bovine species in the WOAH Manual, modified with the use of a triple dose. Water buffaloes were vaccinated with the vaccine RB51. A booster vaccination was administered at 12 months of age. When turning 23-25 months old, female animals were induced to pregnancy. RB51-specific antibodies were detected and quantified using a CFT based on the RB51 antigen. Vaccinated animals showed a positive serological reaction following each vaccine injection, but titers and the duration of the antibody differed among animals. For 36 weeks after booster vaccination, the comparison of CFT values between vaccinated and control groups remained constantly significant. Afterwards, antibody titers decreased. No relevant changes in antibody response were recorded during pregnancy or lactation. In conclusion, results indicated that the vaccination schedule applied is safe and allows for vaccinated and unvaccinated controls to be discriminated between for up to 8 months after booster vaccination.

Serological and Bacteriological Surveillance of Glanders Among Horses in Central Region of Iran.

Glanders is the oldest and very contagious disease among horses caused by Burkholderia mallei. The disease occurs as a chronic form in horses. Hence, because of the prolonged shedding, numerous horses can potentially get infected by one horse with glanders. Glanders is endemic in Iran and this causes occasional occurrence in horse population of the country. The aim of this study was to determine the incidence of B.mallei infection in horses in two central provinces of Iran. A total of 517 serum samples were collected from stable horses in Tehran and Alborz provinces. Among the studied horses, seven presented fever, anorexia, dyspnea, subcutaneous abscesses, nasal and cutaneous discharges, emaciation, and lymphadenopathy. Nasal and ocular discharges and subcutaneous abscesses were sampled for bacterial culture and PCR. The sera were examined by means of complement fixation test (CFT) and indirect enzyme-linked immunosorbent assay (iELISA). Seropositive cases were further examined by Mallein test. The results derived from the present study indicated that only 1.35% of the studied horses were positive in CFT, iELISA and Mallein test, of which only in 42.85% B.mallei was successfully cultured on blood agar and glycerinated nutrient media and confirmed by PCR. Periodic serological tests along with quarantine can benefit reduction of the occurrence of the disease in horses in Iran.

Clinical investigation and management of Brucella suis seropositive dogs: A longitudinal case series.

BACKGROUND: Brucellosis in dogs caused by Brucella suis is an emerging zoonotic disease. OBJECTIVES: To document clinical characteristics, serology, microbiology, and clinical response to treatment in B. suis-seropositive dogs. ANIMALS: Longitudinal study of 27 privately-owned dogs. Dogs that tested positive by serology, culture, or real-time polymerase chain reaction (qPCR) were included in the study. METHODS: Clinical (physical examination and imaging) and laboratory (serology, hematology, serum biochemistry, and qPCR or culture) assessments were made at baseline and after approximately 3, 6, 12, and 18 months. RESULTS: Dogs were followed for 10 895 dog days, with 17/27 dogs completing the 18-month follow-up. Ten dogs had signs consistent with brucellosis before enrollment (n = 4), at baseline (n = 2) or during follow-up (n = 6), with 2 dogs experiencing relapse of historical signs. Antibody titers persisted for the duration of follow-up in 15/17 dogs (88%). Radiographic (n = 5) and ultrasound (n = 11) findings, of variable clinical relevance, were observed. Brucella DNA and organisms were detected in 3 dogs, all of which had clinical signs, including in the milk of a bitch around the time of whelping. Brucella DNA was not detected in blood (n = 92 samples), urine (n = 80), saliva (n = 95) or preputial swabs (n = 78) at any time during follow-up. Six dogs underwent treatment, all of which achieved clinical remission although remission was not reflected by decreasing antibody titers. CONCLUSIONS AND CLINICAL IMPORTANCE: Most dogs with B. suis infections have subclinical infections. Serology is poorly associated with clinical disease. Excretion of organisms appears rare except in whelping bitches. Clinical management using antibiotics with or without surgery is recommended.

Study on small ruminant brucellosis and owners awareness in two selected districts of southern region, Ethiopia.

INTRODUCTION: Brucellosis is one of the infectious diseases that has the greatest impact on the productivity of sheep and goats. A cross-sectional study followed by a simple random sampling technique was used to investigate the seroprevalence of brucellosis (Rose Bengal plate test; RBPT and complement fixation test; CFT) in small ruminants and its related risk variables from November 2019 to June 2020 in Kolme and Abala Abaya districts. A questionnaire was also given to owners to assess their existing knowledge of the disease. RESULT: Using the RBPT and CFT, 28 (4.1%) and 23 (3.33%) of the 690 animals were found to be seropositive for brucellosis, respectively. In this study, the seroprevalence of brucellosis detected in the Kolme district (5.3%) was greater when compared to Abala Abaya (1.0%). The odds of Brucella infection were greater for goats (odds ratio [OR] 6.0, 95% confidence interval [CI] 16 0.8-44.9) than for sheep. The odds of adult animals (OR 0.05, 95% CI 0.03-0.07) being positive for brucellosis was higher than young animals. A statistically significant difference in the seropositivity of brucellosis was detected in univariate logistic regression among districts, different age groups, herd size, parity number, and reproductive health problems except for species and sex, but in multivariate logistic regression, only reproductive health problems were revealed a statistically significant difference. Out of 138 families, 100% of respondents were unaware of brucellosis, 94.5% drink raw milk, and 74% handle animals with retained fetal membranes with their bare hands. CONCLUSION: This study showed that brucellosis was a widely spread disease in the study areas and poses a substantial public health danger. To reduce the spread of the disease in small ruminants, public health risks, and economic losses, stringent vaccination application and awareness of personal hygiene are critical.

Seroprevalence of brucellosis in communal and smallholder cattle farming in North West Province, South Africa.

Brucellosis is an important bacterial zoonosis responsible for considerable economic losses in livestock and health-related burden worldwide. The objective of this study was to determine the seroprevalence of brucellosis in communal and smallholder cattle farming in four districts of the North West province of South Africa (Dr Ruth Segomotsi Mompati, Ngaka Modiri Molema, Bojanala platinum and Dr Kenneth Kaunda districts). Seven hundred and seventy blood samples from farmed animals (n = 378) and abattoir-slaughtered animals (n = 392) were collected. In addition, milk samples (n = 22) were collected from lactating farmed cows. Rose Bengal test (RBT), complement fixation test (CFT) and milk ring test (MRT) were used to detect antibodies against Brucella species. The RBT showed a seroprevalence of 2% at 95% confidence interval (CI: 1.35-3.35), CFT confirmed an overall seroprevalence of 1.95% (95% CI: 1.14-3.12) for all four districts sampled. Although the seroprevalence of brucellosis was found to be low, the possibility of undetected cases of the disease could not be ruled out. Overall, the findings of this study confirmed that brucellosis is endemic in the surveyed areas of the North West province of South Africa.Contribution: The outcome of this study will contribute to the National Brucellosis Project organised by the Department of Agriculture, Land Reform and Rural Development (2016-2026) to assist in the effective implementation of the disease control measures with a view to prevent further outbreaks in the country's cattle population.

Production of a Bacillus anthracis Secretome with Suitable Characteristics as Antigen in a Complement Fixation Test.

In this study, we cultured the Bacillus anthracis vaccine strain Sterne 34F2 in a medium containing EDTA, and we assessed the best conditions to inhibit the activity of zinc-dependent metalloproteases to obtain a secretome containing a high concentration of non-degraded PA (PA83), as evaluated by the SDS-PAGE analysis. Then, we used this secretome as the antigen in a Complement Fixation Test (CFT) to monitor the production of antibodies against PA83 in the sera of rabbits vaccinated with Sterne 34F2 and then infected with a B. anthracis virulent strain to evaluate the potency of the vaccine. The PAS-based CFT results were compared with those obtained by using a commercial ELISA kit. The two serological tests gave similar results in terms of specificity and sensitivity, as the kinetics of the antibodies production was very similar. The Sterne 34F2 vaccine induced an antibody response to PA83, whose titer was not inferior to 1:8 in PAS-based CFT and 42 kU/mL in PA83-based ELISA, respectively, in all vaccinated rabbits. Our opinion is that the PAS-based CFT can be successfully employed in humans and in animals for epidemiological retrospective studies or post-vaccination monitoring. We also suggest the use of our method to test the efficacy of veterinary anthrax vaccines.

Designing of Western Blot Technique for Glanders Diagnosing in Iran.

Burkholderia mallei is the etiologic agent of glanders. It is difficult to diagnose this zoonotic disease in its early stages. Some methods such as the complement fixation test (CFT) cause some problems for veterinary authorities and financial losses to animal owners due to false-positive results. The mallein test requires appropriate laboratory equipment and skilled personnel. To quickly and accurately diagnose the disease, especially in areas where animals cannot be kept, new methods (such as the Western blot test [WBT]) should be used to identify the disease. This study designed and optimized the Western blot (immunoblot) test using sera from 84 glanderous equids, and the sensitivity and specificity of ELISA and CFT were compared with the WBT. ELISA tests are based on B. mallei antigens whereas a purified lipopolysaccharide-containing B. mallei antigen is used in the WBT. The sensitivity and specificity of the tests were estimated using the cut-off values recommended by the test developers. The WBT and ELISA were significantly more specific than the CFT. The ELISA based on B. mallei antigens was significantly less sensitive than the CFT. Given their comparable sensitivities and specificities, the CFT (95.7%, 98.5%), the WBT (95%, 100%) and the ELISA (85%, 100%) should be further developed. The CFT is still the prescribed technique for serological investigation of equids for trade purposes to certify individual animals without glanders. Therefore, more efforts should be made to further improve and optimize the WBT and ELISA tests.

Advances in the Diagnosis of Foot-and-Mouth Disease.

Foot-and-mouth disease (FMD) is a devastating livestock disease caused by foot-and-mouth disease virus (FMDV). Outbreaks of this disease in a country always result in conspicuous economic losses to livestock industry and subsequently lead to serious socioeconomic damages due to the immediate imposition of trade embargo. Rapid and accurate diagnoses are imperative to control this infectious virus. In the current review, enzyme-linked immunosorbent assay (ELISA)-based methods used in FMD diagnosis are extensively reviewed, particularly the sandwich, liquid-phase blocking, and solid-phase competition ELISA. The differentiation of infected animals from vaccinated animals using ELISA-based methods is also highlighted, in which the role of 3ABC polyprotein as a marker is reviewed intensively. Recently, more studies are focusing on the molecular diagnostic methods, which detect the viral nucleic acids based on reverse transcription-polymerase chain reaction (RT-PCR) and RT-loop-mediated isothermal amplification (RT-LAMP). These methods are generally more sensitive because of their ability to amplify a minute amount of the viral nucleic acids. In this digital era, the RT-PCR and RT-LAMP are progressing toward the mobile versions, aiming for on-site FMDV diagnosis. Apart from RT-PCR and RT-LAMP, another diagnostic assay specifically designed for on-site diagnosis is the lateral flow immunochromatographic test strips. These test strips have some distinct advantages over other diagnostic methods, whereby the assay often does not require the aid of an external device, which greatly lowers the cost per test. In addition, the on-site diagnostic test can be easily performed by untrained personnel including farmers, and the results can be obtained in a few minutes. Lastly, the use of FMDV diagnostic assays for progressive control of the disease is also discussed critically.

Evaluation of dot-blot test for serological diagnosis of bovine brucellosis

Abstract The objective of this study was to standardize and validate the dot-blot test for the serological diagnosis of bovine brucellosis, compare the results with those found in the 2-mercaptoethanol (2-ME) and complement fixation test (CF), and estimate the relative sensitivity and specificity of the dot-blot compared to these tests. Fifty bovine blood serum samples were used for the test standardization, and 1315 samples were used for evaluation and comparison between the tests; the results were compared using the Kappa indicator. At the end of standardization, it was established as optimal for the antigen obtained from Brucella abortus B19 after passing through a microorganism rupture process, the blood serum samples diluted at 1:100, and the conjugate at 1:30,000. The comparison of the dot-blot results with 2-ME showed Kappa index of 0.9939, sensitivity of 99.48%, and specificity 99.91%, with CF, Kappa index of 0.8226, sensitivity 100% and specificity 95.32%. Using the combination of the test results 2-ME and CF to establish the true condition of the animal, the dot-blot showed relative sensitivity of 100%, and relative specificity of 99.91%. The evaluated test proved to be effective and reliable, besides being easy to handle and interpret the results.

Extracting the barbs from complement assays: Identification and optimisation of a safe substitute for traditional buffers.

Complement assays have for many years utilised buffers based on barbitone (veronal) despite the well-recognised toxicity of this agent and the tight regulations on its use in most countries. The use of barbitone in complement assay buffers is steeped in history, from a time when no other suitable buffers were available. This is no longer the case, encouraging us to explore alternatives to barbitone for complement assays. We compared a simple, non-toxic HEPES buffer with commercially sourced complement fixation test diluent (CFD), the "gold standard" barbitone buffer, in several clinically relevant complement activity assays and across species. In classical pathway haemolysis assays in human and non-human serum, there was no difference in haemolytic curves or calculated haemolytic activity (CH50) between CFD and an optimised HEPES buffer (HBS) supplemented with cations. Alternative pathway haemolysis assays in human serum were also identical in the two buffers. In a complement fixation test for anti-erythrocyte antibodies, complement consumption was identical for the two buffer systems. The data demonstrate that barbitone-based buffers are unnecessary for assays of complement activity and can readily be replaced with safe and simple alternatives.

Evaluation of dot-blot test for serological diagnosis of bovine brucellosis.

The objective of this study was to standardize and validate the dot-blot test for the serological diagnosis of bovine brucellosis, compare the results with those found in the 2-mercaptoethanol (2-ME) and complement fixation test (CF), and estimate the relative sensitivity and specificity of the dot-blot compared to these tests. Fifty bovine blood serum samples were used for the test standardization, and 1315 samples were used for evaluation and comparison between the tests; the results were compared using the Kappa indicator. At the end of standardization, it was established as optimal for the antigen obtained from Brucella abortus B19 after passing through a microorganism rupture process, the blood serum samples diluted at 1:100, and the conjugate at 1:30,000. The comparison of the dot-blot results with 2-ME showed Kappa index of 0.9939, sensitivity of 99.48%, and specificity 99.91%, with CF, Kappa index of 0.8226, sensitivity 100% and specificity 95.32%. Using the combination of the test results 2-ME and CF to establish the true condition of the animal, the dot-blot showed relative sensitivity of 100%, and relative specificity of 99.91%. The evaluated test proved to be effective and reliable, besides being easy to handle and interpret the results.

The experience of contagious bovine pleuropneumonia ring trials in Botswana.

The Botswana National Veterinary Laboratory (BNVL) has conducted ring trials (proficiency testing) on an annual basis since 2010. Proficiency testing is carried out to evaluate the ability of veterinary laboratories to perform serological complement fixation tests (CFTs) and molecular polymerase chain reaction (PCR) tests for the diagnosis of contagious bovine pleuropneumonia (CBPP). In this paper, the authors discuss the experience gained and the lessons learned in coordinating these ring trials over a period of six years, from 2010 to 2015.The number of participating laboratories increased from five in 2010 to 11 in2015. Their performance also improved over this period. The proportion of unsatisfactory results decreased from 40% to 10% for serological testing, while questionable results decreased from 60% to 10%. The proportion of unsatisfactory results for the molecular test decreased from 33% to 0%. Systematic errors (i.e. technical errors or imperfect experimental design) were the principal causes of questionable and unsatisfactory results. An analysis of responses from customer satisfaction surveys conducted annually since 2013 provided valuable information that enabled BNVL to redesign the programme in 2014 and 2015 to improve the overall quality of the proficiency testing programme. Among the changes made were sending freeze-dried sera for CFTs and DNA for PCR instead of sera and liquid cultures.

Le Laboratoire national vétérinaire du Botswana (BNVL) effectue depuis 2010 des essais comparatifs inter-laboratoires à intervalles d'un an. Ces essais d'aptitude inter-laboratoires visent à évaluer la capacité des laboratoires vétérinaires du pays à réaliser efficacement le test de fixation du complément (FC) ainsi qu'un test moléculaire (l'amplification en chaîne par polymérase ou PCR) en vue du diagnostic de la péripneumonie contagieuse bovine (PPCB). Les auteurs décrivent l'expérience acquise et les enseignements tirés lors de la coordination d'essais comparatifs inter-laboratoires sur une période de six ans (de 2010 à2015). Le nombre de laboratoires participants est passé de cinq en 2010 à onze en 2015. Les performances des laboratoires se sont également améliorées au cours de cette période. Le pourcentage de résultats sérologiques non concluants est passé de 40 % à 10 % tandis que la part des résultats douteux est passée de60 % à 10 %. Pour ce qui concerne la technique moléculaire, la proportion de résultats non concluants est passée de 33 % à 0 %. Les résultats douteux et non concluants étaient essentiellement dus à des erreurs systématiques (c'est-à-dire à des erreurs techniques ou imputables à une conception imparfaite du protocole de test). Une analyse des réponses fournies aux enquêtes de satisfaction réalisées chaque année depuis 2013 a fourni des informations précieuses qui ont permis au BNVL de corriger le protocole du programme en 2014 et en 2015 afin d'améliorer la qualité globale du programme d'essais d'aptitude. Les principales modifications introduites ont été le recours à des sérums lyophilisés pour le test de FC et l'utilisation d'ADN lyophilisé pour les PCR au lieu des sérums et des cultures en milieu liquide.

El Laboratorio Veterinario Nacional de Botsuana (BNVL) lleva realizando pruebas interlaboratorios anuales (pruebas de competencia) desde 2010. Las pruebas de competencia sirven para evaluar la aptitud de un laboratorio veterinario para efectuar pruebas serológicas (la de fijación del complemento, FdC) y moleculares (la de reacción en cadena de la polimerasa, PCR) de diagnóstico de la perineumonía contagiosa bovina. Los autores examinan la experiencia adquirida y las enseñanzas extraídas con la coordinación de estas pruebas interlaboratorios durante el sexenio que va de 2010 a 2015. El número de laboratorios participantes pasó de 5 en 2010 a 11 en 2015. Su eficacia también mejoró durante esos años: en el caso de la técnica serológica, el porcentaje de resultados insatisfactorios pasó de un 40% a un 10 y el de resultados cuestionables de un 60% a un 10%. En el caso de la técnica molecular, el porcentaje de resultados insatisfactorios pasó de un 33% a un 0%. Los resultados insatisfactorios o cuestionables se deben principalmente a errores sistemáticos (esto es, errores técnicos o concepción imperfecta de la prueba). Al analizar las respuestas a los cuestionarios de satisfacción del cliente realizados anualmente desde 2013 se obtuvo información de gran interés, que sirvió al BNVL para remodelar el programa de pruebas de competencia en 2014 y 2015 a fin de mejorar su calidad general. Entre los cambios introducidos está el envío de sueros liofilizados para la prueba de FdC y de ADN liofilizado para la PCR, en lugar de sueros y cultivos líquidos.

Incidence of serum antibody titers against herpes simplex virus in Japanese patients.

Herpes simplex virus (HSV) establishes latency in the sensory neuronal ganglia after primary infection, and occasionally causes recurrent infection, mainly on the lips or genitalia. Previous reports revealed an age-related increase in HSV-immunoglobulin G seropositive subjects in a hospital-based study and the general population in Japan. In this report, we retrospectively analyzed the results of serological tests against HSV, in which subjects were diagnosed with or suspected as having HSV infection. A total of 1216 subjects with at least one complement fixation (CF) result were included. Of these, 771 subjects (63.4%) were positive at first visit. When stratified by age, incidence of positive patients linearly increased with age from teenagers (44.9%) to those in their 80s (88.9%). Positivity in women was higher than in men overall; significantly higher incidence was observed in women aged in their 30s, 40s and 60s. When observing changing HSV-CF titers over time in 81 initially negative patients, 18 (22%) seroconverted during the 2121-day observation period. In this study, we clearly show that distribution of HSV-CF titers is similar to previous HSV-immunoglobulin G results. This correlation is probably caused by the continual subclinical proliferation of HSV, thus maintaining CF titers. Our observations provide current data on the incidence of HSV, reconfirming that serological examination is unreliable in diagnosing recurrent herpes, and the majority of infected subjects are asymptomatic.

Incidence of serum antibody titers against varicella zoster virus in Japanese patients with herpes zoster.

Herpes zoster is an internal reactivation of varicella zoster virus following establishment of latent infection in the dorsal root ganglia during primary infection, which presents as chickenpox. Therefore, serologically, herpes zoster patients already have anti-varicella zoster virus immunoglobulin G at the onset of disease. Hence, positive serum antibody does not confirm the diagnosis of herpes zoster. We retrospectively investigated the incidence of varicella zoster virus-specific complement fixation in 865 zoster patients at initial presentation to a dermatology clinic. As a result, 66% of patients showed negative complement fixation, with patient numbers decreasing as titer increased. Paired complement fixation tests conducted within a short period showed a marked elevation in titer, and complement fixation titer gradually decreased after a year. Furthermore, incidence showed no correlation with patient age. These observations indicate that the complement fixation titer at first visit is mainly influenced by the duration from onset to presentation at clinic. Our findings indicate that a positive complement fixation result by single-point testing confirms at least recent onset of herpes zoster, while paired tests can confirm disease when primary tests are negative.

Interlaboratory ring trial to evaluate CFT proficiency of European laboratories for diagnosis of glanders in equids.

To evaluate the routine complement fixation test (CFT) used to detect Burkholderia mallei antibodies in equine sera, an interlaboratory proficiency test was held with 24 European laboratories, including 22 National Reference Laboratories for glanders. The panels sent to participants were composed of sera with or without B mallei antibodies. This study confirmed the reliability of CFT and highlighted its intralaboratory reproducibility. However, the sensitivity of glanders serodiagnosis and laboratory proficiency may be improved by standardising critical reagents, including antigens, and by developing a standard B mallei serum.

Versatile microfluidic complement fixation test for disease biomarker detection.

The complement fixation test (CFT) is a serological test that can be used to detect the presence of specific antibodies or antigens to diagnose infections, particularly diseases caused by microbes that are not easily detected by standard culture methods. We report here, for the first time, a poly(dimethylsiloxane) (PDMS)/glass slide hybrid microfluidic device that was used to manipulate the solution compartment and communication within the microchannel to establish sampler and indicator systems of CFT. Two types of on-chip CFT, solution-based and solid phase agar-based assays, were successfully demonstrated for biomarker carcinoembryonic antigen (CEA) and recombinant avian influenza A (rH7N9) virus protein detection. In addition, the feasibility of the on-chip CFT in assaying real biopsy was successfully demonstrated by specifically detecting rH7N9 and CEA in human serum. The results demonstrated that the miniaturized assay format significantly reduced the assay time and sample consumption. Exemption from protein immobilization, blocking, complicated washing steps and expensive enzyme/fluorescein conjugates highlights the merits of on-chip CFT over ELISA. Most attractively, the on-chip agar-based CFT results can be imaged and analysed by smartphone, strengthening its point-of-care application potential. We anticipate that the on-chip CFT reported herein will be a useful supplemental or back-up tool for on-chip immunoassays such as ELISA for disease diagnosis and food inspection.

Development of a Sterne-Based Complement Fixation Test to Monitor the Humoral Response Induced by Anthrax Vaccines.

Anthrax is a zoonotic disease caused by Bacillus anthracis spore-forming bacterium. Since it is primarily a disease of animals, the control in animals, and humans depend on the prevention in livestock, principally cattle, sheep, and goats. Most veterinary vaccines utilize the toxigenic, uncapsulated (pXO1+/pXO2-) B. anthracis strain 34F2 which affords protection through the production of neutralizing antibodies directed to the toxin components Protective Antigen (PA), Lethal Factor (LF), and Edema Factor (EF). The titration of specific antibodies in sera of vaccinated animals is crucial to evaluate the efficacy of the vaccination and to obtain epidemiological information for an effective anthrax surveillance. In this study, we developed a Sterne-based Complement Fixation Test (CFT) to detect specific antibodies induced in animals vaccinated with Sterne 34F2. We assessed its efficacy in laboratory animals and under field conditions by monitoring the humoral response induced by vaccination in cattle. The results indicated that the Sterne-based CFT is able to correctly identify vaccinated animals. It proved to be a very sensitive and specific test. Moreover, the Sterne-based CFT offers many benefits with regard to costs, standardization and reproducibility of the assay procedure.