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PubMed1993

Serological evidence of pneumonia virus of mouse (PVM) infection in laboratory rats.

PVM specific antibody was determined using 3 serological tests (Parker's original hemagglutination-inhibition [HI] test, modified HI test, and indirect fluorescent antibody [IFA] test). IFA test was found to be the most sensitive method, so that IFA test was considered to be the method of choice for a small scale microbiological monitoring in animal facilities. A total of 1,280 sera including rats, mice, hamsters and rabbits were tested for PVM antibody. The prevalence of PVM antibody was 17.2% (116/674) in conventional rats and 30.0% (160/200) in SPF rats obtained from commercial breeders and research facilities. Furthermore, two samples of 144 conventional rabbits (1.4%) were showed specific antibody positive to the virus. However, hamsters and mice showed no evidence of PVM infection. These results suggest that PVM infection is prevalent, especially in laboratory rats in Japan.

Animals↗
PubMed2004

Comparison between immunomagnetic separation, coupled with immunofluorescence, and the techniques of Faust et al. and of Lutz for the diagnosis of Giardia lamblia cysts in human feces.

In the present study, the performance of Immunomagnetic Separation technique, coupled with Immunofluorescence (IMS-IFA), was compared with the FAUST et al. and Lutz parasitological techniques for the detection of Giardia lamblia cysts in human feces. One hundred and twenty-seven samples were evaluated by the three techniques at the same time showing a rate of cyst detection of 27.5% by IMS-IFA and 15.7% by both Faust et al. and Lutz techniques. Data analysis showed a higher sensitivity of IMS-IFA for the detection of G. lamblia cysts in comparison with the techniques of FAUST et al. and Lutz. The use of this methodology as a routine procedure enables the processing of many samples simultaneously, in order to increase recovery rate of G. lamblia cysts and reduce the time of sample storage.

Animals↗
PubMedDate not supplied

Seroepidemiological studies of cutaneous leishmaniasis in the Campeche state of México.

Seroepidemiological studies of cutaneous leishmaniasis were carried out in 169 individuals in a rural area of the Campeche state of México. Fifty showed cutaneous lesions suggestive of leishmaniasis, 70% were parasite positive and 96% skin test positive. An overall 40% positivity to skin test with Montenegro's antigen was found. Most of the affected individuals were males from 11 to 30 years-old. Antibodies were determined by immunofluorescent antibody test (IFA) and by Western blot. Two antigen preparations were used, one from a Leishmania mexicana strain which produced localized cutaneous leishmaniasis (LCL) and the other from a diffuse cutaneous leishmaniasis (DCL). In the general population from the area of study 19% gave positive IFA tests with DCL antigen and 20% with LCL antigen while for the patients 67% gave positive IFA tests with DCL and 71% with LCL. By Western blot analysis most of the patients recognized more antigens in the DCL than in the LCL strain. In the DCL strain 78% of patients recognized a 105 kDa, 34% a 139 kDa, 28% a 117 kDa and 26% a 205 kDa MW antigen. In the LCL strain 40% of patients recognized a 205 kDa and 22% a 175 kDa antigens.

Adult↗
PubMed2000

Production and characterization of monoclonal antibodies to Brazilian isolates of bovine viral diarrhea virus.

Three Brazilian isolates of bovine viral diarrhea virus (BVDV), antigenically distinct from the standard North American isolates, were selected to immunize BALB/c mice in order to obtain hybridoma cells secreting anti-BVDV monoclonal antibodies (mAbs). Two hybridoma clones secreting mAbs, reacting specifically with BVDV-infected cells (mAbs 3.1C4 and 6.F11), were selected after five fusions and screening of 1001 hypoxanthine-aminopterin-thymidine-resistant clones. These mAbs reacted in an indirect fluorescent antibody (IFA) assay with all 39 South and North American BVDV field isolates and reference strains available in our laboratory, yet failed to recognize other pestiviruses, namely the hog cholera virus. The mAbs reacted at dilutions up to 1:25,600 (ascitic fluid) and 1:100 (hybridoma culture supernatant) in IFA and immunoperoxidase (IPX) staining of BVDV-infected cells but only mAb 3.1C4 neutralized virus infectivity. Furthermore, both mAbs failed to recognize BVDV proteins by IPX in formalin-fixed paraffin-embedded tissues and following SDS-PAGE and immunoblot analysis of virus-infected cells, suggesting they are probably directed to conformational-type epitopes. The protein specificity of these mAbs was then determined by IFA staining of CV-1 cells transiently expressing each of the BVDV proteins: mAb 3. 1C4 reacted with the structural protein E2/gp53 and mAb 6.F11 reacted with the structural protein E1/gp25. Both mAbs were shown to be of the IgG2a isotype. To our knowledge, these are the first mAbs produced against South American BVDV isolates and will certainly be useful for research and diagnostic purposes.

Animals↗
PubMedDate not supplied

Serologic and molecular evidence of coinfection with multiple vector-borne pathogens in dogs from Thailand.

Forty-nine dogs from Thailand were evaluated for serologic evidence of exposure or polymerase chain reaction (PCR) evidence of infection with vectorborne pathogens, including Ehrlichia sp. (Ehrlichia canis, Ehrlichia chaffeensis, Ehrlichia equi, and Ehrlichia risticii), Bartonella vinsonii subsp. berkhoffi (Bvb), spotted fever group (SFG) rickettsiae (Rickettsia rickettsii), Typhus group (TG) rickettsiae (Rickettsia canada, Rickettsia prowazekii, and Rickettsia typhi), and Babesia sp. (Babesia canis and Babesia gibsonii). All study dogs had at least 1 of 3 entry criteria: fever, anemia, or thrombocytopenia. By immunofluorescence antibody (IFA) testing, seroreactivity was most prevalent to E chaffeensis (74%) and E canis (71%) antigens, followed by E equi (58%), Bvb (38%), E risticii (38%), R prowazekii (24%), B canis (20%), R rickettsii (12%), R canada (4%), and B gibsonii (4%) antigens. There was 100% concordance between E canis IFA and Western blot immunoassay (WI) for 35 of 35 samples; 2 samples were IFA and WI reactive only to E equi antigens. By PCR amplification, 10 dogs were found to be infected with E canis, 5 with Ehrlichia platys, and 3 with B canis. Sequencing of PCR products was undertaken to compare Ehrlichia strains from Thailand to strains originating from the United States. Partial DNA sequence analysis confirmed infection with E canis and E platys, with identical 16S rRNA sequence alignment to E canis (U26740) and to E platys (M83801), as reported in GenBank. Partial E canis P28.1 and P28.2 amino acid sequences from Thai dogs were divergent from analogous sequences derived from North American E canis (AF082744) strains, suggesting that the Thai dogs were infected with a geographically distinct strain of E canis compared to North American strains. The results of this study indicate that dogs in Thailand have substantial exposure to vectorborne diseases and that coinfection with these pathogens may be common.

Amino Acid Sequence↗
PubMed2003

Relationships of resting energy expenditure with body fat distribution and abdominal fatness in Japanese population.

Body fat distribution and abdominal fatness are indicators of risks for coronary heart disease. However, the relationships between resting energy expenditure (REE) and the body fat distribution or the abdominal fatness are unclear. We examined the relationships of REE with whole-body fat distribution (waist, hip and waist-to-hip ratio: WHR) and abdominal fatness (intra-abdominal fat: IF and subcutaneous fat: SF) after adjustment for body composition. 451 men and 471 women were subdivided into two groups, 40-59 years: middle-aged group and 60-79 years: elderly group. REE was measured by an indirect calorimetry system. Percentage of fat mass (%FM), fat mass (FM) and fat-free mass (FFM) were assessed by a dual-energy x-ray absorptiometry method. The IF area (IFA) and SF area (SFA) at the level of the umbilicus were measured using computed tomography. Circumference of waist and hip were measured in a standing position. The WHR, waist circumference and SFA did not significantly (p>0.05) associate with the REE after adjusting for FM, FFM and age in any of the groups. The adjusted REE was significantly and inversely correlated with hip (r=-0.159, p<0.05) and IFA (r=-0.131, p<0.05) in the elderly men. These results suggest that lower REE may contribute to greater hip and IFA rather than WHR and waist in elderly men.

Abdomen↗
PubMedDate not supplied

Malaria in Brazilian military personnel deployed to Angola.

BACKGROUND: Malaria represents one of the most important infectious disease threats to deployed military forces; most personnel from developed countries are nonimmune personnel and are at high risk of infection and clinical malaria. This is especially true for forces deployed to highly-endemic areas in Africa and Southeast Asia where drug-resistant malaria is common. METHODS: We conducted an outbreak investigation of malaria cases in Angola where a total of 439 nonimmune Brazilian troops were deployed for a 6-month period in 1995-1996. A post-travel medical evaluation was also performed on 338 (77%) of the 439 soldiers upon return to Brazil. Questionnaire, medical record, thick/thin smear, and serum anti-Plasmodium falciparum antibody titer (by IFA) data were obtained. Peak serum mefloquine (M) and methylmefloquine (MM) metabolite levels were measured in a subsample of 66 soldiers (42 cases, 24 nonmalaria controls) who were taking weekly mefloquine prophylaxis (250 mg). RESULTS: Seventy-eight cases of malaria occurred among the 439 personnel initially interviewed in Angola (attack rate = 18%). Four soldiers were hospitalized, and 3 subsequently died of cerebral malaria. Upon return to Brazil, 63 (19%) of 338 soldiers evaluated were documented to have had clinical symptoms and a diagnosis of malaria while in Angola. In addition, 37 (11%) asymptomatically infected individuals were detected upon return (< 1% parasitemia). Elevated, post-travel anti-P. falciparum IFA titers (> or = 1:64) were seen in 101 (35%) of 292 soldiers tested, and was associated with a prior history of malaria in-country (OR = 3.67, 95% CI 1.98-6.82, p <.001). Noncompliance with weekly mefloquine prophylaxis (250 mg) was associated with a malaria diagnosis in Angola (OR = 3.75, 95% CI 0.97-17.41, p =.03) but not with recent P. falciparum infection (by IFA titer). Mean peak levels (and ratios) of serum M and MM were also found to be lower in those who gave a history of malaria while in Angola. CONCLUSIONS: Malaria was a significant cause of morbidity among Brazilian Army military personnel deployed to Angola. Mefloquine prophylaxis appeared to protect soldiers from clinical, but not subclinical, P. falciparum infections. Mefloquine noncompliance and an erratic chemoprophylaxis prevention policy contributed to this large outbreak in nonimmune personnel. This report highlights the pressing need for development of newer, more efficacious and practical, prophylactic drug regimens that will reduce the malaria threat to military forces and travelers.

Angola↗
PubMed2001

Evaluation of a polyvalent enzyme-linked immunosorbent assay incorporating a recombinant p44 antigen for diagnosis of granulocytic ehrlichiosis in dogs and horses.

OBJECTIVE: To develop and evaluate a polyvalent ELISA incorporating a highly specific recombinant antigen (p44) for diagnosis of granulocytic ehrlichiosis in dogs and horses. ANIMALS: 32 dogs and 43 horses. PROCEDURE: Results of the ELISA were compared with results of indirect fluorescent antibody (IFA) staining and western immunoblotting incorporating whole-cell antigen. RESULTS: For the canine and equine samples, percentages of samples with positive IFA staining, western immunoblotting, and ELISA results were similar. For 29 (91 %) canine samples and 30 (70%) equine samples, results of IFA staining, western immunoblotting, and the ELISA were in complete agreement. Results of the ELISA for 3 canine serum samples known to contain antibodies to Ehrlichia canis and 12 equine serum samples known to contain antibodies to E risticii were negative. CONCLUSIONS AND CLINICAL RELEVANCE: Results of the present study suggest that a polyvalent ELISA incorporating a recombinant p44 antigen is suitable for detecting antibodies to E equi in dogs and horses.

Animals↗
PubMed2003

Seroprevalence of antibodies against Leishmania spp among dogs in the United States.

OBJECTIVE: To determine seroprevalence of antibodies against Leishmania spp among dogs other than Foxhounds in the United States. DESIGN: Cross-sectional study. SAMPLE POPULATION: 957 serum samples from dogs throughout the United States submitted between January 2000 and August 2001 to the Diagnostic Center for Population and Animal Health at Michigan State University for serologic testing for tick-borne diseases. PROCEDURE: Samples were tested for antibodies against Leishmania spp with an immunofluorescent antibody (IFA) assay. Samples with positive results were submitted to the Centers for Disease Control and Prevention for confirmatory testing. RESULTS: Results of the IFA assay were negative for 939 of 957 samples. For 16 samples, titers were from 1:16 to 1:64, and titers in these dogs were considered likely to be a result of cross-reactivity with antibodies directed against other organisms. For the remaining 2 samples, the titers were > or = 1:128. One of these samples was from a blood donor dog that had never had any clinical signs of leishmaniasis. Follow-up samples from both dogs also had Leishmania IFA titers > or = 1:128. Both dogs had antibodies against Trypanosoma cruzi, as determined with a radioimmunoprecipitation assay. CONCLUSIONS AND CLINICAL RELEVANCE: Results suggested that the seroprevalence of antibodies against Leishmania spp in dogs in the United States was low. However, results further suggested that leishmaniasis may not be limited to Foxhounds in the United States.

Animals↗
PubMed1995

Rapid simple and nested polymerase chain reaction for the diagnosis of Pneumocystis carinii pneumonia.

We have developed a rapid and easy extraction procedure for polymerase chain reaction (PCR) protocols. Using this simplified step, we evaluated the sensitivity and the specificity of a simple PCR using the primers of Wakefield et al, and of a nested PCR, using new internal primers selected by us, in a total of 89 bronochoalveolar lavage (BAL) fluid samples from 43 immunosuppressed patients. In 13 patients, Pneumocystis carinii pneumonia (PCP) was diagnosed by immunofluorescent antibody (IFA) staining performed on BAL cells cytospun on microscope slides. In seven of these patients we attempted to estimate the post-treatment persistence of P. carinii in BAL, by PCR. After a rapid 2-h extraction procedure, simple and nested PCR were positive in all cases of PCP. SImple and nested PCR both had a 100% sensitivity and a 98 and 84% specificity respectively, compared to IFA. After completion of treatment, BAL liquids from asymptomatic patients were no longer positive by both PCR techniques, whereas the BAL fluid of a patient who was still symptomatic was positive by simple and nested PCR. In follow-up BAL fluids of patients with proven PCP, persistence of P. carinii was detected for a longer period by nested PCR than by simple PCR. Simple PCR is a very rapid and sensitive assay for the diagnosis of PCP in BAL fluid and gives clear-cut results in the case of doubtful IFA staining results. Nested PCR seems to improve the sensitivity of the detection of P. carinii in BAL fluid, but the clinical relevance of a positive result remains to be investigated..

Base Sequence↗
PubMed1981

Comparison of specific immunoglobulin G, M and agglutinating antibodies against Legionella pneumophila.

Sera from 25 patients showing a 4-fold or greater rise in polyvalent IFA titer against Legionella pneumophila were examined for the presence of agglutinating antibodies and the titers of specific IgG and IgM immunoglobulins. Agglutinating antibodies were detected in 20 patients (80%) at titers paralleling and amounts of specific IgM present. In sera from the remaining 5 patients (20%), agglutinating antibodies and specific IgM were absent. In this study the class of antibody produced following infection with L. Pneumophila varied from patient to patient. Specific IgM or IgG only was produced or both classes of immunoglobulin appeared concurrently. IFA, using a polyvalent antihuman conjugate, detected both IgG and IgM but cross reactions among the L. pneumophila serogroups occurred in sera from 40.5% of the patients we examined. Tube agglutination measured mainly specific IgM and no cross reactions were evident with this technique. The value of a rapid tube agglutination test as an adjunct to IFA in the serodiagnosis of legionellosis was illustrated in this study.

Agglutinins↗
PubMed2004

Serologic and molecular biologic methods for SARS-associated coronavirus infection, Taiwan.

Severe acute respiratory syndrome (SARS) has raised a global alert since March 2003. After its causative agent, SARS-associated coronavirus (SARS-CoV), was confirmed, laboratory methods, including virus isolation, reverse transcriptase-polymerase chain reaction (RT-PCR), and serologic methods, have been quickly developed. In this study, we evaluated four serologic tests ( neutralization test, enzyme-linked immunosorbent assay [ELISA], immunofluorescent assay [IFA], and immunochromatographic test [ICT]) for detecting antibodies to SARS-CoV in sera of 537 probable SARS case-patients with correlation to the RT-PCR. With the neutralization test as a reference method, the sensitivity, specificity, positive predictive value, and negative predictive value were 98.2%, 98.7%, 98.7%, and 98.4% for ELISA; 99.1%, 87.8%, 88.1% and 99.1% for IFA; 33.6%, 98.2%, 95.7%, and 56.1% for ICT, respectively. We also compared the recombinant-based western blot with the whole virus-based IFA and ELISA; the data showed a high correlation between these methods, with an overall agreement of >90%. Our results provide a systematic analysis of serologic and molecular methods for evaluating SARS-CoV infection.

Antibodies, Viral↗
PubMed1987

Serological cross-reactivity between Sarcocystis and Toxoplasma in pigs.

The development of antibody titers and cross reation between Sarcocystis and Toxoplasma were investigated by means of IFA test and ELISA in pigs experimentally infected with 1.5 x 10(6) S. suicanis sporocysts and 10,000 T. gondii oocysts, respectively. The intact and soluble Sarcocystis antigens were prepared from the bradyzoites harvested by peptic digestion of infected pork. The intact and soluble Toxoplasma, antigens were prepared from the tachyzoites in mouse peritoneal cavity. IgG antibodies in pigs infeded with Sarcocystis and Toxoplasma, respectively were detected first at 2 weeks post infection on both IFA test and ELISA. The antibody titer to Toxoplasma reached its maximum at 6 weeks post infection and decreased thereafter. The antibody titer to Sarcocystis reached its maximum terminally. The cross-reaction titer in pigs infected with Toxoplasma against Sarcocystis antigen was up to 1:16 in IFA test and up to 1:32 in ELISA. The titer in control group was below 1:4 in both reactions.

Journal Article↗
PubMed2006

Immunofluorescent test for simultaneous detection of Edwardsiella ictaluri and Flavobacterium columnare.

Enteric septicemia of catfish (ESC) and columnaris disease are 2 bacterial diseases significantly affecting the aquaculture industry, and thus rapid diagnosis of disease is imperative for making judicious management decisions. A rapid indirect fluorescent antibody (IFA) test with antibody conjugated fluorochromes having 2 different spectral properties (Alexa Fluor 488-emitting green fluorescence, and Alexa Fluor 594-emitting red fluorescence) was compared with bacteriological culture (accepted standard) for simultaneous detection of Edwardsiella ictaluri (EI) and Flavobacterium columnare (FC) in 3 groups of experimentally infected channel catfish (Ictalurus punctatus Rafinesque), and a fourth group that acquired an aquarium-infection with F. columnare. A total of 303 samples (derived from kidney, brain and nares) from 101 fish were concurrently examined by both tests. Fish in the 3 experimentally infected groups (I to III) were culture positive for the bacteria with which they were infected, and fish in Group IV, (the spontaneously infected fish) revealed F. columnare only. The IFA test compared favorably in sensitivity (EI= 80.7 %; FC = 87.2%) and specificity (EI = 83.9%; FC = 88.9%) with the standard bacteriological culture. The positive predictive value (EI = 96.2% Group I, 90.8% Group II, 93.7% Groups I and II combined; FC = 95.2% Group II, 95.3% Groups II, III and IV combined) was high, while the negative predictive value (EI = 66.7% Group I, 31.3% Group II, 59.5% Groups I and II combined; FC = 73.7% Group II, 72.7% Groups II, III and IV combined) was relatively low. The IFA test will serve as an efficient tool for rapid simultaneous detection of E. ictaluri and F. columnare in outbreaks of disease.

Animals↗
PubMed1983

Specificity and crossreactivity of immune serum and hybridoma antibodies to various species of avian coccidia.

The species-specificity and crossreactivity of serum antibodies (Ab) from birds immunized specifically with six different species of coccidia and of 24 hybridoma antibodies (Ab) developed against four species of chicken and two species of turkey coccidia were determined by use of the indirect immunofluorescent antibody (IFA) test on air-dried sporozoites. With few exceptions, the immune chicken sera were found to crossreact with all species of coccidia tested. Seven of the hybridoma Ab were species-specific, while the other 17 Ab demonstrated varying degrees of crossreactivity. Similar types of IFA patterns were seen with both the species-specific and crossreactive hybridoma Ab. Some of the crossreactive hybridoma Ab produced one type of IFA pattern with the sporozoites against which they were originally raised and different patterns with other species of sporozoites. The development of the hybridoma Ab has made it possible to identify the species of coccidia found in mixed infections and check the purity of laboratory strains.

Animals↗
PubMed1975

Development of antibodies to Plasmodium vivax as measured by two different serologic techniques.

Sera from individuals infected with Plasmodium vivax were tested for the presence of malarial antibodies using the indirect fluorescent antibody (IFA) and the indirect hemagglutination (IGA) tests. The primary infection resulted in the conversion of all sera to a positive response in the IFA test, whereas only 50% gave a positive IGA response. There was a direct relationship between the duration of the primary parasitemia and percentage giving positive IGA response. Relapse resulted in high level positive IFA and IGA responses.

Antibodies↗
PubMed1976

Adaptation of a microimmunofluorescence test to the study of human Rickettsia tsutsugamuskh antibody.

A microimmunofluorescence (micro-IFA) test has been adapted to the study of naturally occurring antibody to Rickettsia tsutsugamushi in humans. The micro-IFA test was comparable to the present immlnofluorescent assay in sensitivity and reproducibility, but offered distinct advantages in the quantity of reagents necessary. Also the micro-IFA could be used to simultaneously titer a serum to as many as nine strains of the organism resulting in a great saving of time.

Antibodies, Bacterial↗
PubMed1977

The seroepidemiology of malaria in Middle America. IV. Passage of malaria antibodies from mothers to infants.

In an area of El Salvador moderately endemic for malaria, use of the indirect fluorescent antibody test (IFA) showed that 44% of the infants born to mothers who had IFA responses to Plasmodium vivax of 1:20 or higher during the latter part of their pregnancy had positive IFA responses of 1:10 or higher to this antigen. No serum from an infant was positive in the absence of some level of malarial response in the mother.

Antibodies↗