Brucellosis seroprevalence in Bali cattle with reproductive failure in South Sulawesi and Brucella abortus biovar 1 genotypes in the Eastern Indonesian archipelago.

BACKGROUND: Brucellosis is a major cause of infertility and reproductive failure in livestock. While cattle in the Eastern Indonesian archipelago suffers from reproductive problems information on bovine brucellosis in the region is fragmentary. The control of brucellosis requires a major and prolonged effort and confirmation of the infection by isolation with detailed knowledge of the spread of the infection is essential when planning a control program. RESULTS: Serological investigation of Brucella infection in beef cattle tended under extensive farming conditions revealed a high seroprevalence (19.3%; 95% CI, 17-22) in the compliment fixation tests. The results of a rapid and simple field test correlated well with the Rose Bengal test (kappa, 0.917) and indicated an acceptable sensitivity (87.5%) and specificity (98.1%) compared with the complement fixation test. Reproductive failure was reported for 39.0% of the cows with a loss of calves due to abortion or early death amounting to 19.3%. Past reproductive failure did not, however, correlate with seropositivity in the complement fixation test (RP = 1.21; P = 0.847). B. abortus biovar 1 was freshly isolated from the hygromas of two cows and together with thirty banked isolates collected since 1990 from different parts of Sulawesi and Timor eight related genotypes could be distinguished with one genotype being identical to that of an isolate (BfR91) from Switzerland. The Indonesian genotypes formed together with BfR91 and one African and one North American isolate a distinct branch on the B. abortus biovar 1 dendogram. CONCLUSIONS: Bovine brucellosis appears to be widespread in the Eastern Indonesian archipelago and calls for urgent intervention. The fresh isolation of the pathogen together with the observed high seroprevalence demonstrates the presence and frequent exposure of cattle in the area to the pathogen. The application of a rapid and simple field test for brucellosis could be very useful for the quick screening of cattle at the pen side.

Typhoid fever among hospitalized febrile children in Siem Reap, Cambodia.

Typhoid fever was confirmed by positive blood culture in 5 (3.7%) of 134 febrile children hospitalized in Cambodia. Typhoid was suspected in an additional 25 (18.7 %) blood culture-negative children based on: a positive immunoglobulin M lateral flow assay (IgMFA) (16); a positive polymerase chain reaction (PCR) for Salmonella typhi (2); or clinical assessment (7). The specificity of the IgMFA and PCR assays requires further study.

Infection of cattle in Kenya with Brucella abortus biovar 3 and Brucella melitensis biovar 1 genotypes.

Brucella melitensis biovar 1 was isolated from bovine milk samples from a herd in central Kenya, and Brucella abortus biovar 3 was isolated from aborted fetus materials and vaginal discharge fluids from cattle in central and eastern provinces of Kenya. All infections including those with B. melitensis were in cattle with reproductive problems kept in mixed herds indicating that cross infection occurs from small ruminants. Multiple-locus variable-number tandem repeat analysis genotyping revealed a close molecular homology of the B. melitensis isolates with an isolate from Israel and a close homology of the B. abortus isolates with an isolate from Uganda indicating that these genotypes have a wide geographic distribution. Infection of cattle with B. melitensis may complicate the control of brucellosis in this country.

Serological evidence for brucellosis in Bos indicus in Nigeria.

PURPOSE: Nigeria is the largest cattle-rearing nation in Africa with most animals kept under traditional husbandry practices. While bovine brucellosis does not receive much attention, a relatively high seroprevalence is found in samples submitted for laboratory testing. The aim of the study was to provide serological evidence of brucellosis in cattle from some of the main cattle-rearing states of the country and to validate a simple and rapid field test for the serodiagnosis of bovine brucellosis. METHOD: Serum samples collected in various states of Nigeria from cattle because of suspicion of brucellosis were investigated in the Rose Bengal plate test, and results were compared with a newly developed rapid field test for the detection of Brucella-specific antibodies. RESULTS: Serological evidence for the presence of brucellosis in cattle was obtained for all states included in the study and a high herd prevalence was observed. The seroprevalence was also high among trade and slaughter animals. Results of a rapid field test for the serodiagnosis of bovine brucellosis correlated well with the Rose Bengal plate test (agreement, 95.7%; kappa value, 0.80). CONCLUSIONS: The results indicate that bovine brucellosis is an important veterinarian problem in Nigeria. The easy-to-use and robust field test is most promising for field-based surveillance as it provides an immediate result allowing the prompt instigation of control measures.

Molecular epidemiology of Brucella genotypes in patients at a major hospital in central Peru.

The multiple-locus variable-number repeat analysis of 90 human Brucella melitensis isolates from a large urban area in central Peru revealed variations at 4 (Bruce07, Bruce09, Bruce18, and Bruce42) out of 16 loci investigated, of which 1 (Bruce42) also is used for species identification. Ten genotypes were identified, separated by the number of Bruce42 repeats into two groups that may have distinct phenotypic characteristics. Whereas genotypes with five or six Bruce42 repeats were cultured mainly from adult patients, genotypes with three Bruce42 repeats were isolated from children and young adolescents as well as from adults. In addition, the isolates with three Bruce42 repeats were obtained more often from patients with splenomegaly (P = 0.02) or hepatomegaly (P = 0.006). An annual variation in the diversity of genotypes was observed, possibly reflecting changes in sources of fresh dairy products, supply routes to city shops and markets, and the movement of infected dairy goat herds.

Comparative performance of lateral flow immunochromatography, iELISA and Rose Bengal tests for the diagnosis of cattle, sheep, goat and swine brucellosis.

BACKGROUND: Brucellosis is a world-wide extended zoonosis that causes a grave problem in developing economies. Animal vaccination and diagnosis are essential to control brucellosis, and the need for accurate but also simple and low-cost tests that can be implemented in low-infrastructure laboratories has been emphasized. METHODOLOGY: We evaluated bovine, sheep, goat and swine lateral flow immunochromatography assay kits (LFA), the Rose Bengal test (RBT) and a well-validated protein G indirect ELISA (iELISA) using sera of Brucella culture-positive and unvaccinated brucellosis free livestock. Sera from cattle vaccinated with S19 and RB51 brucellosis vaccines were also tested. Finally, we compared RBT and LFA using sera of white Fulani cattle of unknown bacteriological status from a brucellosis endemic area of Nigeria. RESULTS AND CONCLUSIONS: Although differences were not statistically significant, RBT showed the highest values for diagnostic sensitivity/specificity in cattle (LFA, 96.6/98.8; RBT, 98.9/100; and iELISA, 96.6/100) and the iELISA yielded highest values in sheep (LFA, 94.0/100; RBT, 92.0/100; iELISA, 100/100), goats (LFA, 95.7/96.2; RBT, 97.8/100; iELISA, 100/100) and pigs (LFA, 92.3/100; RBT, 92.3/100; iELISA, 100/100). Vaccine S19 administered subcutaneously interfered in all tests but conjunctival application minimized the problem. Although designed not to interfere in serodiagnosis, vaccine RB51 interfered in LFA and iELISA but not in the RBT. We found closely similar apparent prevalence results when testing the Nigerian Fulani cattle by RBT and LFA. Although both RBT and LFA (showing similar diagnostic performance) are suitable for small laboratories in resource-limited areas, RBT has the advantage that a single reagent is useful in all animal species. Considering these advantages, its low cost and that it is also useful for human brucellosis diagnosis, RBT might be a good choice for resource-limited laboratories.

Simple, rapid, and affordable point-of-care test for the serodiagnosis of typhoid fever.

We developed a point-of-care test for the serodiagnosis of typhoid fever in the format of an immunochromatographic lateral flow assay. The flow assay for typhoid fever is based on the detection of Salmonella enterica serotype Typhi lipopolysaccharide-specific immunoglobulin M (IgM) antibodies. The assay was evaluated on serum samples collected in a hospital in South Sulawesi, Indonesia, where typhoid fever is endemic, and the results were compared with culture and Widal test. The sensitivity of this typhoid fever IgM flow assay for samples collected at 1st diagnosis from patients with culture-confirmed typhoid fever was determined to be 59.3%. The sensitivity ranged from 41.2% to 89.5%, depending on the duration of illness. A specificity of 97.8% was calculated based on results obtained for patients with clinical suspicion of typhoid fever that was later excluded. The assay is ideal for use as a point-of-care test in health care centers that lack the expertise and facilities to perform culture or the less specific Widal test. Because of its simplicity, the assay may also be used as a field test in remote areas.

Point-of-care testing for malaria outbreak management.

A rapid antigen assay for malaria was performed on blood samples collected during a simultaneous outbreak of falciparum malaria and vivax malaria on a remote island in the Indonesian archipelago. During the outbreak, a total of 89 patients (4.3% of the population) were diagnosed with malaria within a week. Microscopic examination revealed 78 malaria slide-positive cases, of whom 49 (62.8%) were identified as P. falciparum, 7 (9.0%) as P. vivax and 22 (28.2%) as mixed P. falciparum and P. vivax infections. The rapid malaria assay showed excellent correlation with expert-confirmed routine microscopy for P. falciparum and P. vivax monoinfections and mixed infections with a parasite density >50 parasites/microl. Several slide-negative blood samples collected from febrile patients with clinical malaria tested positive in the rapid test. The estimated sensitivity calculated for the rapid test (91.0%) was slightly higher than that of microscopy (87.6%). The result indicates that rapid antigen detection for malaria could be a useful alternative to microscopy to reduce the workload during emergency outbreak situations.

Simple and rapid field tests for brucellosis in livestock.

Four simple and rapid field tests for the serodiagnosis of brucellosis in cattle, goat, sheep and swine were developed. The performance of the assays was investigated using serum samples collected in Portugal from animals originating from herds with a defined sanitary status with respect to the presence of brucellosis. The sensitivity calculated for the bovine, caprine, ovine and swine Brucella lateral flow assays based on results obtained for samples collected from animals with culture confirmed brucellosis was 90%, 100%, 90% and 73%, respectively. None of the samples from animals from herds free of brucellosis reacted in the flow assays indicating a high specificity. However, as expected, some degree of reactivity was observed when testing selected serum samples that reacted non-specific in reference tests for brucellosis.

Human brucellosis.

Human brucellosis still presents scientists and clinicians with several challenges, such as the understanding of pathogenic mechanisms of Brucella spp, the identification of markers for disease severity, progression, and treatment response, and the development of improved treatment regimens. Molecular studies have shed new light on the pathogenesis of Brucella spp, and new technologies have permitted the development of diagnostic tools that will be useful in developing countries, where brucellosis is still a very common but often neglected disease. However, further studies are needed to establish optimum treatment regimens and local and international control programmes. This Review summarises current knowledge of the pathogenic mechanisms, new diagnostic advances, therapeutic options, and the situation of developing countries in regard to human brucellosis.

Detection of Salmonella typhi by nested polymerase chain reaction in blood, urine, and stool samples.

A nested polymerase chain reaction (PCR) specific for Salmonella enterica serovar Typhi was used for the detection of the pathogen in blood, urine, and stool samples from 131 patients with clinical suspicion of typhoid fever. The sensitivity of blood culture, the PCRs with blood, urine, and feces, and the Widal test were 61.8%, 84.5%, 69.3%, 46.9%, and 39.0%, respectively. The sensitivity of the PCRs with blood (P < 0.001) and urine (P = 0.01) were significantly higher, and the sensitivity of the PCR with feces (P > 0.05) was similar to that of blood culture. Combined, the PCRs on urine and feces showed positive results for 16 (70%) of 23 typhoid patients with negative results with blood culture and PCR with blood. These results show that the PCR with blood is a sensitive method for the diagnosis of typhoid fever, and that the PCRs with urine and feces could be useful complementary tests.

Evaluation of brucellosis by PCR and persistence after treatment in patients returning to the hospital for follow-up.

Polymerase chain reaction (PCR) was applied to confirm the diagnosis of brucellosis and to study its clearance in response to the standard treatment regimen with doxycycline and rifampin at hospitals in Callao and Lima, Peru. The PCR confirmed the diagnosis in 23 (91.7%) patients with brucellosis including 12 culture-confirmed cases. For patients treated at the hospital in Callao, PCR was positive for all samples collected during and at the conclusion of treatment and for 76.9% of follow-up samples collected on average 15.9 weeks after completion of treatment. For patients treated at the hospital in Lima, PCR tests were positive for 81.8% of samples collected during treatment, for 33.3% of samples collected at the conclusion of treatment, and for > or = 50% of samples collected at first, second, and third post-treatment follow-up. Thus, Brucella DNA may persist in the serum weeks to months after completion of the standard treatment regimen.

Rapid latex agglutination test for the serodiagnosis of human brucellosis.

We developed and evaluated a user-friendly latex agglutination assay for the serodiagnosis of human brucellosis. The assay was obtained by coating colored latex beads with Brucella lipopolysaccharides and drying of the activated beads onto white agglutination cards. Individual cards were sealed in a protective foil to secure stability of the dried reagent and to obtain a test in a single assay format. The latex agglutination assay is simply performed by suspending the dried latex reagent in a drop of serum and looking for macroscopic agglutination of the latex beads by visual inspection. Results are obtained within 30 s after mixing the sample with the test reagent. The sensitivity of the assay was determined to be 89.1% (95% confidence interval [CI], 76-96) for the initial serum samples collected from patients with culture-confirmed brucellosis and the specificity is 98.2% (95% CI, 96-99). The assay is ideal for use as a field test in remote areas and as point-of-care test in hospitals and health care centers that lack the expertise and facilities to perform the more demanding classic serologic tests.

Persistence and relapse in brucellosis and need for improved treatment.

Treatment failure and relapse are major problems in the management of brucellosis. In developing countries, treatment with the oral combination doxycycline/rifampicin is preferred because of its convenience. However, its efficacy is lower than that of the doxycycline/streptomycin regimen and is likely further reduced when compliance is poor. Alternative regimens should be investigated in well designed clinical trials to determine whether treatment can be improved. Use of DNA detection as a marker for treatment success and for the prediction of relapse requires confirmation. In the absence of simple and effective treatment regimens, patient education to promote compliance is essential.

Laboratory evaluation of a simple and rapid latex agglutination assay for the serodiagnosis of typhoid fever.

A latex agglutination assay for the serodiagnosis of typhoid fever was evaluated on samples collected from patients with clinical suspicion of typhoid fever in South Sulawesi, Indonesia, where the disease is endemic. The latex assay is very easy to use, gives a rapid result and may be used as a point-of-care diagnostic test. For acute phase samples collected on average 6 days after the onset of illness, the sensitivity is 42.5% for culture-confirmed patients with typhoid fever and the specificity is 96.9%. The sensitivity improved with the duration of illness from 30.8% for samples collected during the first 4-5 days of illness to 45.5% for samples collected between days 7 and 9, and to 84.6% for the samples collected more than 9 days after the onset of illness. Testing of follow-up samples may further improve sensitivity by demonstrating seroconversion.

Review of brucellosis in Nepal.

Brucellosis is an abortifacient zoonotic disease that remains prominent in third world countries like Nepal. Brucellosis poses a public health concern, because its incidence in livestock can present substantial economic and health burdens for herders and health professionals. Several cases of bovine and human brucellosis have been reported and the prevalence is higher among livestock than among humans in Nepal. Lack of awareness, unhealthy food habit, traditional husbandry practices, and a lack of surveillance and immunization have been the major factors in maintaining a vicious cycle of propagation of the disease in human and animals. The aim of this paper is to evaluate the current status of the disease, the mechanism of infection, and pathogenesis, its zoonotic potential, diagnostic advances, treatment regimens, and the preventive measures that can be adopted in managing human brucellosis in under-developed countries such as Nepal.

Risk Factors of Typhoid Infection in the Indonesian Archipelago.

BACKGROUND: Knowledge of risk factors and their relative importance in different settings is essential to develop effective health education material for the prevention of typhoid. In this study, we examine the effect of household level and individual behavioural risk factors on the risk of typhoid in three Indonesian islands (Sulawesi, Kalimantan and Papua) in the Eastern Indonesian archipelago encompassing rural, peri-urban and urban areas. METHODS: We enrolled 933 patients above 10 years of age in a health facility-based case-control study between June 2010 and June 2011. Individuals suspected of typhoid were tested using the typhoid IgM lateral flow assay for the serodiagnosis of typhoid fever followed by blood culture testing. Cases and controls were defined post-recruitment: cases were individuals with a culture or serology positive result (n = 449); controls were individuals negative to both serology and culture, with or without a diagnosis other than typhoid (n = 484). Logistic regression was used to examine the effect of household level and individual level behavioural risk factors and we calculated the population attributable fraction (PAF) of removing each risk significant independent behavioural risk factor. RESULTS: Washing hands at critical moments of the day and washing hands with soap were strong independent protective factors for typhoid (OR = 0.38 95% CI 0.25 to 0.58 for each unit increase in hand washing frequency score with values between 0 = Never and 3 = Always; OR = 3.16 95% CI = 2.09 to 4.79 comparing washing hands with soap sometimes/never vs. often). These effects were independent of levels of access to water and sanitation. Up to two thirds of cases could be prevented by compliance to these practices (hand washing PAF = 66.8 95% CI 61.4 to 71.5; use of soap PAF = 61.9 95%CI 56.7 to 66.5). Eating food out in food stalls or restaurant was an important risk factor (OR = 6.9 95%CI 4.41 to 10.8 for every unit increase in frequency score). CONCLUSIONS: Major gains could potentially be achieved in reducing the incidence of typhoid by ensuring adherence to adequate hand-washing practices alone. This confirms that there is a pivotal role for 'software' related interventions to encourage behavior change and create demand for goods and services, alongside development of water and sanitation infrastructure.

Application of a user-friendly Brucella-specific IgM and IgG antibody assay for the rapid confirmation of Rose Bengal-positive patients in a hospital in Iran.

The Brucella IgM/IgG flow assay was used for the confirmation of brucellosis in patients from an area endemic for brucellosis and who had a Rose Bengal (RB)-positive serum sample collected at the time of first presentation for diagnosis. The flow assay confirmed the result of the RB test in 46.6% of the positive admission sera, with the majority (62.5%) of the flow assay-positive samples staining moderately strong to very strong (> or =2+). In comparison, Wright and 2-ME at the routinely used cut-off titre values of 1:320 for Wright and 1:160 for 2-ME tested positive in 37.7% of the RB-positive samples. A relatively large number of RB-positive samples agglutinated at or around the cut-off value in the Wright and 2-ME tests and 66.7% of the RB-positive samples tested positive in these confirmatory tests when using one titre step lower threshold values. The relatively high number of samples with low antibody levels supports the argument for testing follow-up samples from patients with an RB-positive sample in order to confirm the diagnosis by showing seroconversion or a rise in antibody levels.

Brucellosis in India: a deceptive infectious disease.

Brucellosis is an important but neglected disease in India. This zoonotic disease is present in all livestock systems and increased demand for dairy products accompanied with changing and intensified farming practices has raised the concern for increased spread and intensified transmission of this infection to the human population with increased risk of disease. Brucellosis can be controlled by mass vaccination of livestock. Human brucellosis can be treated with a combination of antibiotics but is very difficult to diagnose and requires laboratory testing for confirmation. Only a few recent studies have addressed the prevalence and importance of brucellosis as a human disease problem in India. The disease may be overlooked and misdiagnosed because of the difficult diagnosis and the absence and lack of experience with laboratory testing. Alertness of medical staff is needed to recognize and diagnose the disease. Awareness of risk groups is needed to take appropriate preventive measures and to accept control measures.

Brucella melitensis Biovar 1 and Brucella abortus S19 Vaccine Strain Infections in Milkers Working at Cattle Farms in the Khartoum Area, Sudan.

BACKGROUND: Human brucellosis is a preventable zoonoses that may become persistent, causing, if left untreated, severe localized disease. Occupational exposure to infected animals or animal products and consumption of fresh contaminated dairy are main risk factors. METHODS: One hundred farmworkers employed at two cattle farms one in Khartoum North and one in Omdurman were screened for the presence of specific antibodies and seropositive workers were invited to donate a blood sample for blood culture. Molecular typing was used to characterize Brucella isolates. RESULTS: Ten percent of farmworkers tested seropositive and while Brucella melitensis biovar 1 was isolated from the blood of three individuals, an isolate identical to the B. abortus S19 vaccine strain was isolated from a fourth person. All four bacteremic individuals were employed as milkers and did not have obvious disease. CONCLUSIONS: The isolation of the highly infectious pathogen B. melitensis from seropositive workers is consistent with the notion that the pathogen may persist in the blood without causing overt disease. While vaccination with strain S19 is essential for the control of bovine brucellosis the vaccine strain may be transmitted to the human population and protective measures remain important to prevent exposure also in view of the presence of B. melitensis. To create awareness for this potentially severe disease more information on the prevalence of the pathogen in different risk groups and in livestock in the Sudan is needed.