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Abstract Orthobunyavirus oropouche ense virus (OROV), the causative agent of Oropouche fever, is widely dispersed in Brazil and South America, causing sporadic outbreaks. Due to the similarity of initial clinical symptoms caused by OROV with other arboviruses found in overlapping geographical areas, differential diagnosis is challenging. As for most neglected tropical diseases, there is a shortage of reagents for diagnosing and studying OROV pathogenesis. We therefore developed and characterized mouse monoclonal antibodies and, one of them recognizes the OROV nucleocapsid in indirect immunofluorescent (IFA) and immunohistochemistry (IHC) assays. Considering that it is the first monoclonal antibody produced for detecting OROV infections, we believe that it will be useful not only for diagnostic purposes but also for performing serological surveys and epidemiological surveillance on the dispersion and prevalence of OROV in Brazil and South America.

Orthobunyavirus oropouche ense virus (OROV), the causative agent of Oropouche fever, is widely dispersed in Brazil and South America, causing sporadic outbreaks. Due to the similarity of initial clinical symptoms caused by OROV with other arboviruses found in overlapping geographical areas, differential diagnosis is challenging. As for most neglected tropical diseases, there is a shortage of reagents for diagnosing and studying OROV pathogenesis. We therefore developed and characterized mouse monoclonal antibodies and, one of them recognizes the OROV nucleocapsid in indirect immunofluorescent (IFA) and immunohistochemistry (IHC) assays. Considering that it is the first monoclonal antibody produced for detecting OROV infections, we believe that it will be useful not only for diagnostic purposes but also for performing serological surveys and epidemiological surveillance on the dispersion and prevalence of OROV in Brazil and South America. [ABSTRACT FROM AUTHOR]

Eshetu Nigussie GeletaDepartment of Medical Laboratory Science, Madda Walabu University, Bale Goba, EthiopiaCorrespondence: Eshetu Nigussie GeletaDepartment of Medical Laboratory Science, Madda Walabu University, PO Box 302, Bale Goba 4540, EthiopiaTel +251 91 356 0128Email eshetunba@gmail.comBackground: Dengue fever (DF) is a re-emerging public health threat in Ethiopia. Yet, little is known about the epidemiology and risk factors of dengue infection in the region. In this study, the seroprevalence and associated risk factors of dengue virus infection were assessed in the Borena Zone health facilities.Methods: A hospital-based cross-sectional study was conducted from July to August 2016. A total of 519 consecutive acute febrile patients attending the outpatient departments of Teltelle Health Center, Yabello and Moyale Hospital were enrolled. Data on socio-demographic and environmental risk factors were collected using a structured questionnaire. Three to five milliliter blood samples were collected from all participants and screened for dengue virus exposure using indirect immunofluorescent assay.Results: The overall prevalence of anti-DENV IgG and IgM was 22.9% and 7.9%, respectively. DF serostatus was influenced by gender (adjusted odd ratio (AOR)=1.72; 95% CI 1.01–2.94), place of residence (AOR=2.69; 95%CL 1.55–4.64) that had a higher rate of exposure and recalling of a recent mosquito bite (AOR=2.98; 95% CI 1.51–5.89) probably imply recent and/or ongoing active transmission.Conclusion: This study showed that DF could potentially emerge as a public health threat in the study area. In addition to that, the observed low awareness of participants underlines the urgent need for further community-based studies to determine the environmental, and host factors that determine the extent of exposure to dengue virus infection in the area for appropriate control and prevention planning.Keywords: Borena, dengue virus, indirect immunofluorescent assay, Ethiopia

Abstract Neosporosis is caused by an obligate intracellular protozoan, Neospora caninum . It is considered one of the most widespread and frequent causes of abortion in cattle worldwide. To evaluate the prevalence of anti-N. caninum antibodies and associated risk factors, serum samples were collected from 2,452 bovines at 262 farms in the northern Pantanal, state of Mato Grosso, Brazil. Each farmer was asked to fill out a questionnaire for subsequent epidemiological data analysis. Anti-N. caninum antibodies were detected by means of the indirect immunofluorescent assay (IFA), using a cut-off dilution of 1:100. The overall anti-N. caninum antibodies prevalence was 25.44% (Confidence Interval - CI 95%; 20.10%; 30.78%), and the anti-N. caninum antibodies prevalence per herd was 76.72% (CI 95%; 71.60%; 81.84%). The presence of dogs, occurrence of abortion in cows, and sale of cattle for breeding were statistically associated with seropositivity in herds, while the risk of females being seropositive for N. caninum was higher in animals ≤ 6-years-old and in the presence of dogs. A spatial analysis indicated that the relative risk of the disease is spatially constant and that the farms with the highest prevalence of anti-N. caninum antibodies are located south of the region under study.

In Iran, leptospirosis is endemic to Caspian Sea littoral. The disease appears as a seasonal infection mostly affecting people in rural areas involved in farming. We investigated the prevalence of this infection among suspected patients in Gilan Province by an indirect immunofluorescent assay (IFA), and two PCR protocols, a nested-PCR and a real-time PCR (qPCR), targeting rrs and lipL32 genes, respectively. We also identified the common Leptospira species by sequencing a partial sequence of rrs gene. Out of the 128 sera examined by IFA, 25.78% were positive with the antibody titers ≥1/80. The antibody titer in 39.06% of sera ranged from 1/10 to 1/140, and 35. 16% showed no antibodies, all considered negative. Nested PCR and qPCR detected Leptospira DNA in 20.31% and 18.75% of the sera, respectively. The two PCR assays had 98.43% agreement ( K  = 0.93) and showed an inverse correlation with the IFA titers. Also, three pathogenic Leptospira species, L . kirschneri (n = 10),  L. introgans (n = 8), and L. borgpetersenii (n = 2) were identified from the clinical specimens in the study area. In our hands both PCR assays proved very efficient for early diagnosis of illness and could be used in combination with IFA for both diagnosis and epidemiological studies, but nested PCR was cheaper and appeared more appropriate for our laboratories in rural settings. [ABSTRACT FROM AUTHOR]

[b]Introduction[i][/i][/b][i]. Borrelia burgdorferi [/i]sensu lato (s.l.) and Anaplasma phagocytophilum are well known zoonotic pathogens, whereas[i] Ehrlichia canis[/i] is usually considered to be of veterinary concern, although on the basis of recent reports it also seems to be able to infect humans. [b]objective[/b]. The aim of the study was to determine the seroprevalence of [i]B. burgdorferi [/i]s.l., A. phagocytophilum and [i]E. canis[/i] in an Italian canine population, and to verify if there are differences between dogs living in urban areas and those from a rural environment. [b]materials and method.[/b] Blood sera of 1,965 dogs, 1,235 from cities and 730 from rural areas, were tested by indirect immunofluorescent assay (IFAT). [b]results[/b]. The overall seroprevalence was highest for E. canis (7.07%), followed by [i]A. phagocytophilum[/i] (4.68%), and [i]B. burgdorferi[/i] s.l. (1.47%). Rural dogs showed the highest seroprevalence to [i]B. burgdorferi[/i] s.l. and [i]A. phagocytophilum[/i]. No significant differences were observed between rural and urban [i]E. canis[/i]-positive dogs. A low percentage (1.32%) of dogs with dual seropositivity was detected, and no triple positive reactions were observed. No significant differences were detected in the seroprevalence of the three agents in relationship to the age and gender of the dogs. Seroprevalence in the five years considered were not statistically different, except for the lowest rate for [i]E. canis[/i] observed in 2012. [b]conclusions[/b]. The results confirm the presence of [i]B. burgdorferi[/i] s.l., [i]A. phagocytophilum[/i] and [i]E. canis[/i] in Italian dogs in both urban and rural areas. Monitoring pet dogs, which share the same environment with their owners, is useful for identifying the presence of tick-borne disease agents of both veterinary and public health significance

Introduction: Indirect immunofluorescence test is a most common test for the detection of Systemic Lupus Erythematosis (SLE), however, it is very expensive and is not easily available for all field laboratories. We aimed to evaluate the diagnostic performance of five micrometer thick sections prepared from bovine thymus for the detection of SLE patients. Methods: Totally 48 serum samples (25 negative and 23 positive) were drained from SLE suspected patients. Bovine thymus was purchased from the slaughterhouses. Five micrometer thick sections prepared from bovine thymus. Also rat liver was prepared through the laboratory animal dissection, and assayed in parallel with HEp-2 tests. Two fold serial dilutions (1:40 to 1:640) were prepared for each serum sample using phosphate buffer solution and assayed for ANA. Immunostained cells were observed using a blue filter equiped fluorescent microscope with 200x magnification and any bright green fluorescence accumulation in the cell nucleus was considered as positive for the presence of ANA. Results: Our findings showed sections prepared from thymus cells can be used for the detection of ANA in patient samples and the calculated sensitivity, specificity and accuracy of the test were 82.6%, 100% and 91.7%, respectively. Positive predictive value and negative predictive value of the test were 100% and 86.2%, respectively. Conclusion: We concluded that thymic cells can be easily prepared and used for detection of ANA in SLE patients. However, minor improvements in section preparation and assay conditions as well as determination of a new cut off point is necessary to gain 100% efficiency of the assay. [ABSTRACT FROM AUTHOR]

Se muestra la utilidad e idoneidad de la técnica de inmunofluorescencia indirecta (IFI) como prueba suplementaria confirmatoria de la infección por VIH-1 con base en siete años de experiencia de uso permanente en el Instituto Nacional de Salud (INS), siguiendo un método de confirmación (flujograma), recomendado por la Organización Panamericana de la Salud y el Federal Center for AIDS de Canadá. La especificidad de la prueba IFI del INS (IFI-VIH1-INS), utilizando como prueba de referencia la inmunoelectrotransferencia o Western Blot (WB), fue del 100% con 925 sueros estudiados. La concordancia entre la IFI y dos pruebas diferentes de WB fue similar. Sin embargo, aunque la especificidad de la prueba es excelente, su sensibilidad de 61% con 6.137 sueros probados es discretamente inferior a la de otras pruebas IFI informadas en la literatura, por cuenta de un alto número de resultados indeterminados, de los cuales 41% (975), también fue indeterminado por WB. La utilización de la IFI-VIH1-INS como prueba suplementaria confirmatoria en más de 6.000 sueros, entre 1993 y 2000, permitió ahorrar al programa de VIH de la Red Nacional de Laboratorios de Colombia un valor cercano a los $340'000.000 o US $170.000.

Introduction. Borrelia burgdorferi sensu lato (s.l.) and Anaplasma phagocytophilum are well known zoonotic pathogens, whereas Ehrlichia canis is usually considered to be of veterinary concern, although on the basis of recent reports it also seems to be able to infect humans. Objective. The aim of the study was to determine the sero-prevalence of 6. burgdorferi s.l., A. phagocytophilum and E. canis in an Italian canine population, and to verify if there are differences between dogs living in urban areas and those from a rural environment. Materials and method. Blood sera of 1,965 dogs, 1,235 from cities and 730 from rural areas, were tested by indirect immunofluorescent assay (IFAT). Results. The overall seroprevalence was highest for E. canis (7.07%), followed by A. phagocytophilum (4.68%), and B. burgdorferi s.l. (1.47%). Rural dogs showed the highest sero-prevalence to B. burgdorferi s.l. and A. phagocytophilum. No significant differences were observed between rural and urban E canis-positive dogs. A low percentage (1.32%) of dogs with dual seropositivity was detected, and no triple positive reactions were observed. No significant differences were detected in the sero-prevalence of the three agents in relationship to the age and gender of the dogs. Sero-prevalence in the five years considered were not statistically different, except for the lowest rate for E. canis observed in 2012. Conclusions. The results confirm the presence of B. burgdorferi s.l., A. phagocytophilum and E canis in Italian dogs in both urban and rural areas. Monitoring pet dogs, which share the same environment with their owners, is useful for identifying the presence of tick-borne disease agents of both veterinary and public health significance. [ABSTRACT FROM AUTHOR]

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The article examines the seroprevalence of Q fever among the dairy cattle in the Konya province, Central Anatolia, Turkey. It cites that 40 of the 322 cattle sera were seropositive in the Immunoflurescent Antibody (IFA) test. It reports that no pathological history, abortion, dead or weak offspring or infertility in the 40 seropositive cattle when samples were drawn. It adds that no tick was found either on cattle or in housing areas upon examination. It notes that the IFA test is considered to be the reference method in serologically diagnosing Q fever.

Abstract: Indirect immunofluorescent assay for antinuclear antibodies (IIFA-ANA) on HEp-2 cell substrate is a widely used test for diagnosis of connective tissue diseases. Recently, serum-free fibroblast cell line McCoy-Plovdiv has been developed to provide an alternative substrate. The aim of the present study was to evaluate the diagnostic performance of IIFA-ANA on McCoy-Plovdiv cell substrate for systemic lupus erythematosus (SLE) and rheumatoid arthritis (RA) and compare it with that of the standard HEp-2 cell substrate. Sera from 72 patients with RA and 23 patients with SLE were tested with IIFA-ANA on both substrates. The control group consisted of 100 sera from healthy individuals. The agreement (Cohen''s κ) and the diagnostic performance (ROC analysis) of both methods were evaluated. IIFA-ANA on McCoy-Plovdiv cells proved to be comparable with the standard IIFA-ANA on HEp-2 cell substrate. [Copyright &y& Elsevier]

Abstract: We describe a seroepidemiological survey of malaria prevalence in two areas of low endemicity: Intervales State Park and Alto Ribeira State Tourist Park (PETAR). Both are located in the Vale do Ribeira in the state of São Paulo, Brazil. In this study, 318 subjects from both areas had their blood analyzed for the presence of malaria parasites by thin and thick blood smears. One hundred and sixty-three (51.2%) of the subjects were from Intervales State Park and 155 (48.7%) were from PETAR. We analyzed all the samples by indirect immunofluorescent assay (IFA) to detect antibodies against asexual forms of Plasmodium vivax and Plasmodium malariae and enzyme immunosorbent assay (ELISA) to detect the presence of antibodies against circumsporozoite proteins (CSP) from P. vivax VK210, human P. vivax-like/Plasmodium simiovale, P. vivax VK247 and Plasmodium brasilianum/P. malariae. The presence of Plasmodium species was detected by polymerase chain reaction (PCR). Eighteen of the subjects analyzed had positive IFA results for IgM against P. malariae antigens, and three others were positive for P. vivax antigens. Positivity of IgG antibodies against P. vivax detected by IFA was high in samples from both Intervales State Park and PETAR (32.0% and 49.0%, respectively), while positivity for P. malariae was lower (16.0% and 19.3% in Intervales State Park and PETAR, respectively). ELISA tests showed a higher prevalence of antibodies against P. vivax VK210 (35.0%) in samples from Intervales State Park and against human P. vivax-like (29.7%) in samples from PETAR. PCR reactions revealed the presence of parasites in several of the samples analyzed. In Intervales State Park, one subject was infected by P. malariae and two by Plasmodium falciparum, while in PETAR, one subject was positive for P. falciparum and three for both P. falciparum and P. vivax parasites. The areas where these parks are located belong to the Atlantic Forest habitat, and inhabitants frequently, see monkeys. Our data suggest that monkeys may constitute a natural reservoir for malaria in both areas. [Copyright &y& Elsevier]

Abstract: The antibody response against Leishmania (Leishmania) amazonensis crude antigen was measured through the indirect immunofluorescent assay (IFA) and the immunoenzymatic assay (ELISA) in 114 patients with cutaneous leishmaniasis (CL) in Brazil. Fifty-four patients were infected by Leishmania (Viannia) braziliensis, and 60 patients had L. (V.) guyanensis infection. Patients were comparable by age, sex, disease duration and the Montenegro skin test diameter. L. (V.) braziliensis-infected patients showed significant lower number of ulcerated lesions, greater ulcerated area and higher proportion of lymph node enlargement. Sensitivity of IFA was 79.6% (95% CI 66.1–88.9) and 71.7% (95% CI 58.4–82.2) for L. (V.) braziliensis and L. (V.) guyanensis-infected patients, respectively (P=0.324). Sensitivity of ELISA was 98.2% (95% CI 88.8–99.9) and 85.0% (95% CI 72.9–92.5) for L. (V.) braziliensis and L. (V.) guyanensis-infected patients, respectively (P=0.018). Significant differences were observed in the magnitude of the antibody response before treatment with higher levels detected in L. (V.) braziliensis-infected patients by both serologic techniques. Eighty-four patients had serologic evaluations before and 12 weeks after treatment with meglumine antimoniate, 20mg/kg/day for 20 days. Significant lower optic density values were observed after treatment with both species independent of cure or failure. Our data showed that L. (V.) braziliensis induces a higher antibody response against L. (L.) amazonensis antigens than L. (V.) guyanensis and that down-modulation of the antibody response occurs shortly during disease evolution after treatment. Moreover the data support the use of ELISA as a better tool for detection of antibodies in CL. [Copyright &y& Elsevier]

Jovičić V, Grego E, Lako B, Ristović B, Lepšanović Z, Stajković N. Improved serodiagnosis of early Lyme borreliosis: Immunoblot with local Borrelia afzelii strain. APMIS 2003;111:1053–9. To improve the serodiagnosis of Lyme borreliosis (LB) the performances of four tests were evaluated. An indirect immunofluorescent assay based on Borrelia burgdorferi s.s. , enzyme-linked immunosorbent assay (ELISA) based on local isolates of Borrelia afzelii and B. burgdorferi s.s., and immunoblot (IB) of B. afzelii were prepared. The serum panels contained 214 serum samples: control group (n=120) and patients at different stages of LB (n=94). The specificity of IB was 96%, of in-house ELISA 93%, and of IFA 89%. In early LB the sensitivity of IFA was 36%, ELISA 67%, and IB 93%. In late-stage LB the sensitivity was: 72% for IFA, 80% for ELISA, and 94% for IB. Comparison of in-house and Behring ELISA showed that the sensitivity of the serological assay could be increased when the test was based on local Borrelia strains. IgM and IgG antibodies from sera of patients with early and late LB most frequently demonstrated reactivity to OspC. The other significant proteins in early LB were: p39, p41 in IgM IB, and p83/100, p39, Osp17 in IgG IB; in late LB: p39, p41 in IgM IB, and p83/100, Osp17, p21 and p43 in IgG IB. Using IB based on local B. afzelii isolates improves the serodiagnosis of early LB in our geographical region. [ABSTRACT FROM AUTHOR]

Abstract: In a study, carried out in 2000, of the clinical and parasitological status of a Wichi Aboriginal community living in the suburbs of Tartagal, northern Salta, Argentina, 154 individuals were screened for parasitic infections. Ninety-five faecal samples were also obtained from the same population. Ninety-three percent of the subjects were positive for 1 or more of the parasites investigated by direct test and 70.5% of them had parasitic superinfection. The most frequent helminths were Strongyloides stercoralis (50.5%) and hookworm (47.4%). We found low reinfection rates and a long reinfection period after treatment and provision of safe water and sanitation. Serum reactivity of these patients was analysed by enzyme-linked immunosorbent assay and indirect immunofluorescent assay and 22.1% of them had anti-Toxocara antibodies, 16.2% were positive for a complex antigen of Leishmania braziliensis, 29.9% were positive for a complex Trypanosoma cruzi antigen, and 17.5% were positive for a specific Trypanosoma cruzi antigen, Ag1 63136/cruzipain. [Copyright &y& Elsevier]

We used A chain extracted from urine of patient with myeloma (IgD λ) as antigen for immunizing BALB/C mice, and 86 hybridoma cell clones secreting monoclonal antibodies (McAb) were obtained after fusing twice and cloning 3-4 times. 15 of these clones secreted monoclonal anti-idiotypic antibodies (anti-Id McAb). The results showed that 12 of 15 anti-Id antibodies reacted only with homologous λ chain and IgD, not with A chain, κ chain, IgG, IgA, IgM, IgD, IgE, albumin and paraglobulin from normal subjects. Indirect immunofluorescent assay demonstrated that positive reaction rate between anti-Id McAbs and peripheral blood lymphocytes or bone marrow cells of the patient with myeloma was up to 23 %. No reaction was observed between anti-Id McAb and peripheral blood lymphocytes or bone marrow cells from normal subjects. Some of these McAbs presented positive reaction with plasmacytoma cell lines cultured in our laboratory. [ABSTRACT FROM AUTHOR]

Two kinds of immunofluorescent assay, the indirect immunofluorescent assay (IIF) and the improved indirect immunofluorescent assay (improved-IIF), were used to detect the intracellular cytomegalovirus (CMV) antigens. It was found that the improved-IIF could demonstrate CMV antigens not only earlier but also in a smaller amount than the IIF. It is obvious that the improved-IIF is a more sensitive method than the IIF. The possible principles for the enhanced sensitivity of improved-IIF were discussed briefly. [ABSTRACT FROM AUTHOR]