Evaluation of Brucellacapt for the diagnosis of human brucellosis

OBJECTIVE: To evaluate the role of Brucellacapt in the diagnosis of human brucellosis, and the correlation with the evolution of the disease. METHODS: Twenty-six patients who were admitted to the General Hospital of Albacete (Spain) over a 2-year period and diagnosed with brucellosis were included in the study. One hundred and twenty-three serum samples collected at the time of diagnosis and at intervals during and after treatment were tested by the Coombs test, the standard seroagglutination test (SAT), and Brucellacapt (a new test based on an immunocapture-agglutination technique). To study the specificity of Brucellacapt, sera from 20 patients with other infectious diseases and 20 sera from healthy donors were included in this study. RESULTS: The sensitivity of the Brucellacapt at the moment of diagnosis was similar to the Coombs test (96 and 100%, respectively), somewhat higher than that of SAT (73%). And the specificity of the Brucellacapt (97.5%) was less than SAT and the Coombs test, that was 100%. The correlation between the classical serological tests and Brucellacapt, showed that titers in Brucellacapt and Coombs test of patients were both similar in a range of 1-2 dilutions. The correlation between Brucellacapt and Coombs (r=0.14) and between Brucellacapt and SAT (r=0.0) did not reach statistical significance. However, the correlation coefficient between Coombs and SAT was r=0.8. CONCLUSIONS: Brucellacapt and Coombs tests showed a similar sensitivity and specificity in the diagnosis of human brucellosis. In addition, as Coombs test, Brucellacapt could help to diagnose patients with long evolution of brucellosis that are not detected with SAT.     

Evaluation of serological tests for diagnosis of brucellosis

The aim of the present study was to compare serological tests (Rose Bengal [RB]; standard agglutination test [SAT]; enzyme immunoassay [EIA] for detection of IgM, IgA, and IgG; and 2-mercaptoethanol [2-ME] test) that are routinely used in patients prediagnosed with different clinical types of brucellosis (acute, subacute, or chronic), and to evaluate the results of the IgG avidity test. Ninety-two patients having titers≥1/160 as measured by SAT were included in the study. The IgG avidity test was performed in 78 patients who had positive EIA-IgG results. RB test results were positive in 88 (95.7%) patients. A statistically significant correlation was found between a positive EIA-IgM result and the diagnosis of acute brucellosis. When compared to the results of the SAT, the 2-ME test showed a lower titer in 55 (59.8%) patients, and the agreement between the 2-ME test and EIA-IgG was calculated as 84.8%. No statistical difference was found between the 40% avidity index used in the IgG avidity test and avidity maturation time (6 months). From our study, we concluded that (i) the RB and SAT tests are appropriate and reliable tests for the serological diagnosis of brucellosis; (ii) IgM can be used as a marker of acute brucellosis; (iii) the 2-ME test, similar to EIA, can be used to determine IgM levels; and (iv) the IgG avidity test should be standardized.     

Changes in IgM and IgG antibody concentrations in brucellosis over time: importance for diagnosis and follow-up

Changes in concentrations of IgM and IgG antibodies to Brucella were monitored for at least 13 mo by an enzyme-linked immunosorbent assay (ELISA) in 52 patients with culture-positive brucellosis. Two main patterns were observed. After an initial peak, 29 patients (56%) had a steady drop in their IgG levels, whereas 23 (44%) had more than one peak over time. All patients with a chronic form of brucellosis or a relapse were in the second group. In most cases, Brucella antibodies, although falling to low levels, remained measurable. Cutoff levels for IgM and IgG were calculated after considering serum antibody concentrations in people who had recovered from an infection. A separate normal range was established for occupationally exposed workers. On admission, sera from all patients contained Brucella antibody levels greater than established cutoff levels. Our results show that ELISA is an excellent method for diagnosis and follow-up of brucellosis.     

Use of the Brucella IgM and IgG flow assays in the serodiagnosis of human brucellosis in an area endemic for brucellosis

The clinical utility of two complementary tests for brucellosis, the Brucella IgM and IgG flow assays, was evaluated in a hospital in eastern Turkey. The results show that the flow assays are convenient diagnostic tests for use in endemic areas. A positive result in the flow assays was obtained in 91% and 97% of the admission sera from adult and pediatric patients with brucellosis, respectively, and the sensitivity at admission was 100% for culture-confirmed brucellosis. The assay system performed equally well in diagnosing patients at different stages of illness including patients with acute, subacute, or chronic disease and with relapse. The results of the flow assays correlated well with those of a serum agglutination test at a cut-off > or =1:160. The agreement was 92%. Application of the flow assays on serum samples collected during a village survey for brucellosis after an outbreak demonstrated their diagnostic potential as field tests.     

Specific antibodies detected during relapse of human brucellosis

We studied 10 patients who had a relapse of brucellosis for significant serological changes during relapse. By the Coombs test, the pre-relapse and post-relapse median (range) titers of antibody to Brucella were 1:1120 (1:40-1:10,240) and 1:10,240 (1:40-1:81,920), respectively (P = .0069); by enzyme-linked immunosorbent assay (ELISA) for IgG, these values were 1:2240 (1:60-1:10,240) and 1:10,240 (1:160-1:81,920; P = .0069); by ELISA for IgA, the median titer increased 3.36 times but did not reach statistical significance (P = .0929); by tube agglutination, dithiothreitol (DTT) agglutination, and ELISA for IgM, median titers did not change. Nine patients had a significant titer increase by Coombs test and IgG ELISA, three had a significant increase by tube and DTT agglutination, none had significant increases by IgM ELISA, and one had no significant increase after relapse. Our findings show that for most patients with a relapse of brucellosis, there is an increase in IgG titers, as detected by ELISA and Coombs test, but no change in IgM titers.     

Seroepidemiological study on human brucellosis in Assiut Governorate

Brucellosis is the most important zoonotic disease constituting a public health problem in Assiut Governorate, hence this study was carried out to determine the prevalence of brucellosis among humans in Assiut Governorate. A total of 7154 peripheral blood samples were collected from patients with fever at Assiut Fever Hospital during the period from 2002-2003. A full detailed anamnestic and clinical assessment in the form of questionnaire was designed for each individual to determine the risk factors with specific emphasis to age, sex, residence and occupation. All serum samples were screened for Brucella antibodies by slide agglutination test. Positive sera were further analyzed by standared tube agglutination test. Enzyme linked immunosorbent assay (ELISA) was carried out to detect IgM and IgG Brucella antibodies. Statistical analysis was performed and correlation coefficient was done between all risk factors. Results declared that the prevalence of brucellosis was (1.29 +/- 0.004 %) and (1.22 +/- 0.002 %) as detected by agglutination and ELISA, respectively. IgM antibodies were estimated in 9.8 % of the examined patients, while IgG antibodies were found in 30.4 % of the examined patients, moreover both IgM and IgG antibodies were detected in 54.3 % of the examined patients. The prevalence of brucellosis was significantly (P < 0.05) affected by sex, where the rate of detection was higher among females (1.76 +/- 0.009 %) than males (1.05 +/- 0.004 %) as detected by agglutination test. On the other hand, the prevalence rate based on ELISA was (1.64 +/- 0.39 % and 1.01 +/- 0.89 %) for females and males, respectively. Prevalence of brucellosis was higher in rural areas (1.3 +/- 0.005 % & 1.25 +/- 0.009 %) than in urban areas (1.23 +/- 0.001% & 1.12 +/- 0.01 %) as detected by agglutination test and ELISA, respectively. The prevalence of brucellosis varied significanctly between different occupational and age groups. Public health impact of brucellosis is discussed and suggestive measures for control are explained.     

Parasite-specific antibody response in Trichinella sp. 3 human infection: a one year follow-up

Sera from patients with confirmed or suspected trichinellosis were examined for 1 year to detect the presence of parasite-specific antibodies (IgG, IgM, and IgE) using the enzyme-linked immunosorbent assay (ELISA). The indirect ELISA was used to detect specific IgG (ELISA-IgG) and specific IgM (ELISA-IgM); an amplified technique proved the most reliable for detection of specific IgE (a-ELISA-IgE). The immunofluorescence (IF) test was used to detect specific IgG (IF-IgG). The patients were from an outbreak of trichinellosis in Salsomaggiore (northern Italy) in 1986. The parasite was isolated and isoenzymatically typed as Trichinella sp. 3. The specificity of our tests was greater than 95%. During the 1st period of infection, all tests used gave practically the same positivity rate (78.2-86.9%). One year after infection, ELISA-IgG gave the highest positivity rate (55%). With the other tests, the positivity rate was 20-38.5%. At the 2nd month of infection, the IF-IgG test was the most discriminating in patients with confirmed and suspected trichinellosis, but ELISA-IgG proved the most reliable test for detecting specific immunoglobulins in late human trichinellosis infection.     

2001-2015年锦州市布鲁氏菌病疫情流行病学分析

目的描述2001-2015年锦州市布病疫情流行病学特征和变化趋势,探讨下一步的防控重点。方法用描述流行病学方法对锦州市布病疫情进行分析,对试管凝集试验阳性率与发病率进行Pearson相关性分析。结果 2001-2015年锦州市布病发病率为0.45/10万-16.53/10万。发病时间集中在3-7月,占总数的66.89%。2003-2015年共采集重点人群血清5 904份,阳性率为5.01%。2007-2015年从136份血培养标本中分离出布鲁氏菌63株,其中羊种3型48株、羊种1型12株,羊种变异2株、犬种1株。结论锦州市近年布病疫情呈明显上升趋势,其优势菌株主要是羊种3型。对犬的布鲁氏菌携带情况应予以一定的关注,及时淘汰病犬。     

Limited diagnostic usefulness of antibodies to cytoplasmic proteins of Brucella in early-treated human brucellosis

Antibodies to cytoplasmic proteins (CP) of Brucella have been shown to be useful for the diagnosis of human brucellosis; however, some early-diagnosed patients lack such an antibody response while having high titers of antibodies to lipopolysaccharide (LPS). To address which factors determine this serological discrepancy in the early stages of brucellosis we examined the antibody response to CP and LPS of 21 patients involved in an outbreak of B. melitensis infection who had a short duration of clinical illness at diagnosis (3-40 d). At diagnosis, antibodies to LPS (IgM and/or IgG) were found in all patients, while anti-CP antibodies were detected in 16 subjects (76%). At 6 weeks post-diagnosis IgG to CP (with or without IgM) had been detected in 13 patients and IgM alone had been found in 4; however, 4 other patients (19%) had no response to CP. No significant differences were found between these 3 groups in terms of age, gender, antimicrobial agents or factors that could hamper the immune response. Notably, however, the 4 non-responders and 3 of the 4 patients having only IgM to CP had started antibiotic therapy within 14 d post-symptoms, while treatment was started later in 9 of 13 patients who developed anti-CP IgG. In addition, maximum titers of IgG to CP tended to be lower in early-treated patients. These results suggest that very early antibiotic therapy hampers the antibody response to Brucella CP but has little impact on the anti-LPS response. Given the higher specificity of the former and the higher sensitivity of the latter, both reactivities should be measured in order to diagnose human brucellosis.     

Specific antibody profile in human brucellosis.

The results of classic serological tests were compared with those of enzyme-linked immunosorbent assay in studies of immunoglobulins to Brucella in 761 serum samples from 75 patients with brucellosis. Except for five instances involving the IgM ELISA, all serological tests gave positive results at admission. Among the 63 patients without relapse, rates of persistent ELISA positivity (determined by the Kaplan-Meier method) 12 months after therapy were 25% for IgM, 69% for IgA, and 89% for IgG. Among the 12 patients with relapse, a second peak of ELISA IgG and IgA was often detected. The persistence of high serum antibody titers in patients without relapse was due mainly to IgG and was often associated with high titers at admission or with the presence of focal disease. Overall, serological changes were better detected by ELISA than by classic serological tests. While a second peak of ELISA IgG and IgA is a good marker of relapse, the persistence of high titers of IgG by itself is not a good predictor of chronic infection.     

布鲁氏菌病快速检测试纸条的探索

目的 建立一种操作简单、结果肉眼可视,并具有良好敏感性和特异性的适合现场快速检测布鲁氏菌病的胶体金层析方法。方法 将重组的布鲁氏菌外膜蛋白OMP22和OMP28作为胶体金标记物,抗OMP22单克隆抗体作为质控线,OMP22和OMP28蛋白作为检测线组装试纸条。结果 该试纸条检测牛布鲁氏菌标准阳性血清最大稀释度可达1:128;并且与牛结核、蓝舌病、牛病毒性腹泻、口蹄疫、牛白血病及小反刍兽疫血清无交叉反应;检测牛、羊源血清样品结果与商品化的ELISA检测试剂盒(IDEXX)符合率为95%,与虎红平板凝集试验(RBPT)检测的符合率为97.4%。结论 胶体金层析试纸条在检测布鲁氏菌病方面具有快速、敏感性高及特异性强的特点,适用于布鲁氏菌病动态流行病学调查和临床快速筛查。     

The Antibody Response to Brucella: Immunoglobulin Response Measured by Enzyme-linked lmmunosorbent Assay and Conventional Tests

Abstract: The antibody response to Brucella: immunoglobulin response measured by enzymelinked immunosorbent assay and conventional tests. G. L. Gilbert and L. A. Hawes, Aust. N.Z. J. Med ., 1981, 11, pp 40–45.
An enzyme-linked immunosorbent assay was adapted to measure total and individual classes of brucella antibody. The results were compared with those of conventional tests for brucella antibody on the sera of a number of healthy seropositive abattoir workers and several patients with either acute or suspected chronic brucellosis. IgG was the class of brucella specific immunoglobulin most commonly detected in all groups. IgM was present in the sera of 40% of seropositive abattoir workers, all but one of the patients with recent acute brucellosis or seroconversion and none of those with suspected chronic brucellosis. Many of the abattoir workers' sera which contained brucella specific IgM gave negative results in the direct agglutination test. The presence of brucella specific lgM in the sera of these men was, in most cases, associated with no past history of acute brucellosis and a relatively short period of employment in the abattoir. It is suggested that the presence of brucella specific IgM in the serum of a person occupationally exposed to 6. abortus, probably indicates a relatively recent primary infection, either symptomatic or sub clinical and has no prognostic significance. Repeated or prolonged exposure is associated with IgG brucella antibodies, often in high titre, irrespective of symptoms. It was not possible, on the basis of any serological tests performed in this study to distinguish healthy people exposed to brucella from those with symptoms consistent with chronic brucellosis     

Diagnosis and treatment of 106 cases of human brucellosis

During the year 1987, 106 cases of human brucellosis were studied prospectively at the Jordan University Hospital. The disease was more often diagnosed among adults (73.6%) than children (26.4%). Serious clinical complications were observed in 5.7% patients. An initial Brucella antibody titre greater than or equal to 160 proved to be reliable in confirming suspected cases of acute and subacute brucellosis. Culture of blood was found to be more sensitive (44.4%) and significant (P less than 0.02) than bone marrow culture (27.7%) for detecting Brucella melitensis. All patients treated with rifampicin plus tetracycline or co-trimoxazole were considered to be clinically cured by disappearance of all major clinical features of brucellosis. By contrast, 2/10 patients treated with rifampicin alone, as well as 1/56 patients treated with tetracycline and streptomycin, clinically relapsed. It is evident from this study that the treatment with rifampicin alone is not as effective in brucellosis as it is when given with another appropriate drug.     

Evaluation of A newly developed DOT-ELISA kit for the diagnosis of human brucellosis

A new dot-ELISA kit for detection of Brucella antibodies in human sera was developed and compared with that of serum agglutination test, Rose Bengal plate test, rapid slide agglutination and Coomb's antiglobulin test. Following testing of 120 human sera from suspected patients of occupational risk, 25 gave positive reaction in Rose Bengal plate test, 25 in rapid slide agglutination test, 26 in serum agglutination test, 27 in Coomb's antiglobulin test and 28 in dot-ELISA kit. Dot-ELISA kit picked up more positive than any other Serological test, indicating its superiority over the other laboratory tests for the diagnosis of brucellosis.     

Dot immunoassay with colloidal silver particles as a marker of specific antigen for testing human sera for brucellosis

Whether the dot immunoassay is suitable for the detection of Brucella antibodies in human sera by using a colloidal silver-labeled Brucella specific antigen as a diagnostic tool is assessed. The antigen was the B. abortus 19BA protein polysaccharide complex isolated by Brucella acetic acid hydrolysis. The dot immunoassay is easy-to-use, cost-effective, highly sensitive, and therefore of more informative value in detecting Brucella antigens than routine serological tests (Huddleson test, Wright agglutination test, passive hemagglutination test, long-term complement fixation test, and Coombs test). It requires no use of expensive equipment and reagents.     

An Evaluation Study of Enzyme-Linked Immunosorbent Assay (ELISA) Using Recombinant Protein Pap31 for Detection of Antibody against Bartonella bacilliformis Infection among the Peruvian Population

Reliable laboratory testing is of great importance to detect Bartonella bacilliformis infection. We evaluated the sensitivity and specificity of the enzyme-linked immunosorbent assay (ELISA) using recombinant protein Pap31 (rPap31) for the detection of antibodies against B. bacilliformis as compared with immunofluorescent assay (IFA). Of the 302 sera collected between 1997 and 2000 among an at-risk Peruvian population, 103 and 34 samples tested positive for IFA-immunoglobulin G (IgG) and IFA-IgM, respectively. By using Youden''s index, the cutoff values of ELISA-IgG at 0.915 gave a sensitivity of 84.5% and specificity of 94%. The cutoff values of ELISA-IgM at 0.634 gave a sensitivity of 88.2% and specificity of 85.1%. Using latent class analysis, estimates of sensitivity and specificity of almost all the assays were slightly higher than those of a conventional method of calculation. The test is proved beneficial for discriminating between infected and non-infected individuals with the advantage of low-cost and high-throughput capability.     

Severe thrombocytopenic purpura due to brucellosis

A case of severe thrombocytopenic purpura as the sole manifestation of brucellosis in an 11-y-old boy is presented. Clinical examination was negative and laboratory tests revealed anemia, leukopenia and severe thrombocytopenia. The initial diagnosis was idiopathic thrombocytopenic purpura and intravenous gamma globulin was given. A prompt hematological response was observed. However, on the fifth day after admission, blood culture yielded Brucella which could not be serotyped. The boy was subsequently treated with intravenous gentamicin, oral doxycycline and rifampicin and was discharged in good health. Brucellosis has occasionally been associated with mild hematological abnormalities such as anemia and leukopenia. Thrombocytopenia is rare and only in very rare cases it is severe enough to cause bleeding. Prompt recognition of this complication of brucellosis and aggressive therapy are essential, especially if there is a family history of brucellosis or if there is suspicion of exposure to infected food products.     

Seroprevalence of brucellosis in cattle and humans in the Akwapim-South district of Ghana: public health implications

A total of 183 cattle comprising 54 bulls, 53 milking cows, 76 heifers and 44 calves in the Akwapim-South district of Ghana were tested for antibodies against Brucella abortus using the Rose Bengal plate test. The results indicated that cattle in the Akwapim-South district were infected with Brucella with a mean seroprevalence of 6. 6%. There was no difference in the seroprevalence either between females 11/129 (8.5%) and males 1/54 (1.9%), or among the three different breeds of cattle (Sanga, West African short horn (WASH) and white Fulani) in the study area. However, there was a significant increase in seropositivity with respect to age. A significant association between antibodies against Brucella and a history of abortions and retained placenta in cows indicated that brucellosis might be responsible for significant economic losses to farmers in the area. However, no evidence of human brucellosis was detected by antibody screening in selected risk groups.     

Evaluation of in-house and commercial immunoassays for the sero-diagnosis of brucellosis in a non-endemic low prevalence population

The Brucella Reference Unit (BRU) at the University Hospital Aintree offers a national Brucella sero-diagnosis service for England, Wales, Eire and Northern Ireland. The United Kingdom is a non-endemic area with a very low prevalence of infection. The objective of this study was to evaluate new CE marked assays, Brucellacapt (Vircell) and Brucella IgG and IgM ELISAs (Vircell), against the standard set of in-house serological assays used at BRU. These include a micro-agglutination (MAG) assay, in-house IgG and IgM assays and a complement fixation test (CFT). One hundred and forty-three archived serum samples were re-tested by both the commercial and in-house assays. Samples were divided into four distinct groups based on the most common clinical patterns of serological profiles seen (negative, clinically significant and two forms of clinical indeterminate results). The kappa test was calculated to determine the level of agreement between the commercial and in-house results. The kappa coefficient for Brucellacapt and MAG assays was 0.90 (95% CI 0.85, 0.95) giving a very good level of agreement. Discrepancies between positive MAG and Brucellacapt assay results (5.7%) occurred only in sera with weakly reactive MAG titres of <1:160. Similarly the kappa coefficient calculated for the IgG assays was 0.81 (95% CI 0.75, 0.87) also indicating good agreement. However, the kappa coefficient for the commercial and in-house IgM assays was poor at 0.38 (95% CI, 0.30, 0.46). The weak IgM correlation was associated in some instances, with a lack of use of IgG sorbent in the in-house assay resulting in false-negative results. In a low prevalence population, the combination of in-house and commercial immunoassays offers improvements in the sero-diagnosis of brucellosis.     

布氏菌病的流行趋势及诊断

目的加强对布氏菌病的流行趋势、临床表现及实验室诊断的认识,更好地监测该病。方法回顾性分析近5年我院的70例布氏菌病患者的发病趋势、临床特征及诊断方法,包括血培养及血清学凝集方法。结果实验室血液培养阳性的19例均为临床诊断病例,而76例血清学阳性的有70例临床诊断为布氏菌病,特异度为92.11%。患者以成年男性为主,且呈逐年递增之势;临床表现主要以发热、多汗、乏力、关节痛为主。结论布氏菌病有增加趋势,须加强对其监测及临床特征的认识。血培养是诊断布氏菌病的金标准,血清凝集试验是快速有效的辅助诊断方法。