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PubMed1995

Detection kinetics for positive blood culture bottles by using the VITAL automated system.

The VITAL system principle is based on homogeneous fluorescence technology. During an 11-month period, a total of 19,706 blood cultures from adult patients hospitalized in various establishments of the Montpellier Teaching Hospital were collected in VITAL bottles, of which 1,939 were declared positive. Only 204 bottles (1.04%) were false positives. The 1,735 true-positive bottles were collected from 130 patients. The final visual control permitted the detection of 10 falsely negative bottles (0.05%), of which 5 contained clinically significant microorganisms from four patients. The kinetics of detection for all microorganisms showed that 66.6% were detected within 24 h, 83.1% within 48 h, 95.5% within 120 h, and 100% within 150 h. No clinical episode would have been missed had a 5-day protocol been used instead of a 7-day protocol. Among the positive bottles, 65.7% were detected by the SLOPE algorithm, 20.1% by the DELTA algorithm, and 14.2% by the THRESHOLD algorithm. This retrospective study of our results shows that a 5-day protocol is sufficient for the detection of septic episodes using the VITAL system.

Adult↗
PubMed1977

The use of preserved milk samples in the Brucella milk ring test.

Brucella milk ring tests (BMRT's) were performed on fresh herd milk samples and pooled samples, preserved at a processing factory with potassium dichromate. Preserved milk from samples pooled over a 10-day period, gave results that were similar to an average result for individual fresh daily samples over the same period. When fresh samples gave variable results and the pooled preserved sample gave a negative result, it was considered that the latter result was more reliable, possibly due to factors causing false positive reactions being diluted. Blood testing of herds for circulating antibody indicated that the use of either fresh or preserved milk samples could occasionally produce false negative or false positive results.

Agglutination Tests↗
PubMed1979

Evaluation of three serological tests for the diagnosis of hydatid disease.

A number of techniques is used for the serological diagnosis of hydatid disease; however previous investigations suggest variable degrees or reliability for each test. This study reports an evaluation of the sensitivity and specificity of three commonly used serological tests for hydatid disease; complement fixation (CF), haemagglutination (HA) and the fluorescent antibody (FA) techniques. The results of these serological tests are compared in a series of specimens from 116 hospital patients, 21 of whom were confirmed surgically as having hydatid disease. A similar high degree of sensitivity was found for each of the three serological tests examined. The CF and HA tests gave negative results in two patients with confirmed hydatid disease, and the result of the FA test was negative in only one patient. The HA test was the most specific with no false positive reactions, but only two false positive reactions occurred with the FA test. The CF test was the least specific as the reaction was positive in nine patients without hydatid disease. When the results of both FA and HA tests were considered together, all 21 cases of hydatid disease were detected and false positive reactions occurred in only two patients.

Complement Fixation Tests↗
PubMed1989

Rapid diagnosis of acute Epstein-Barr virus infection by an indirect enzyme-linked immunosorbent assay for specific immunoglobulin M (IgM) antibody without rheumatoid factor and specific IgG interference.

An indirect enzyme-linked immunosorbent assay (ELISA) for detection of Epstein-Barr virus-specific immunoglobulin M (IgM) antibody was developed with commercial reagents. Sera containing rheumatoid factor (RF) (as little as 0.5 IU/ml) coupled with specific IgG resulted in false-positives in the ELISA. This interference was eliminated by the use of anti-human IgG antibodies to remove RF and IgG. Thus, pathogen-specific IgG complexes to which IgM-RF could be bound during the subsequent test were inhibited, and competition between specific IgG and IgM was also prevented. Of the 1,672 serum specimens tested, 353 were found to be Epstein-Barr virus IgM antibody positive by indirect immunofluorescence (IF). Compared with the IF test, the ELISA showed 96.6% sensitivity, 99.7% specificity, and 99% accuracy. Further evidence indicated that most of the 12 ELISA false-negatives were IF false-positives. There was a linear correlation between mean ELISA values and increasing IF titers (r = 0.96). However, the IF test has the disadvantages that it lacks automated reading and requires considerable technical expertise, both of which restrict the range of laboratories performing the test. The indirect ELISA has the advantages that it is simple and rapid and can be automated. All the reagents used in this assay are commercially available, have been prestandardized, and are stable.

Adolescent↗
PubMed1997

A new PCR-based seroneutralization assay in cell culture for diagnosis of hepatitis E.

A new method for the serological diagnosis of hepatitis E virus (HEV) infection based on neutralization of the virus in cell culture was developed. The test involves a short incubation of the virus in the presence of the serum sample to be tested and permissive cells. With viral replication being limited and without a cytopathic effect, viral growth in cells is evaluated by reverse transcription and PCR. The specificity of the test was established by studying sera from healthy individuals and patients with hepatitis living in France, where autochthonous hepatitis E is unknown. The kinetics and sensitivity of antibody detection were evaluated during the experimental infection of monkeys. Neutralizing antibodies were found in 79% of patients during an outbreak of hepatitis E and in 43% of patients with sporadic, acute non-A, non-B (without anti-hepatitic C virus antibodies) hepatitis. This neutralization assay is proposed as a confirmatory test for the available enzyme-linked immunosorbent assay (ELISA), which is now recognized as giving many false-positive reactions, and to improve identification of new hepatitis viruses since false-negative reactions with HEV ELISA are also encountered.

Adult↗
PubMed1992

Polymerase chain reaction: relevance for oral pathology.

The polymerase chain reaction (PCR) is a recent innovation in biotechnology for amplifying specific nucleic acid sequences by use of repeated cycles of DNA synthesis. Application of PCR has been extended to a wide variety of biological and medical disciplines; fields of application include molecular genetics, pathology, microbiology, forensic medicine, and infectious diseases. Because of its speed and powerful sensitivity, PCR has become a highly valuable method for detecting small amounts of target nucleic acid sequence with the caveat that it has certain limitations. Importantly, false-positive reactions can result from the amplification of contaminating DNA whereas false-negative reactions can result from introduction of inadequate numbers of primer sequences. The application of PCR in the context of oral pathology is presented along with a discussion of the utility of this technique to aid in the elucidation of the etiology of oral disease.

DNA↗
PubMed1996

Evaluation of dot enzyme-linked immunosorbent assay (dot-ELISA) for the serodiagnosis of canine leishmaniosis as compared with indirect immunofluorescence assay.

A dot enzyme-linked immunosorbent assay (dot-ELISA) was developed and compared with a standard indirect immunofluorescence assay (IFA) for the rapid serodiagnosis of canine leishmaniosis. The two tests were used to examine sera from Leishmania infantum-infected and control dogs. Using the dot-ELISA, 137 of 149 sera (91.9%) from infected animals gave a clearly positive reaction, whereas in the standard IFA technique 147 (98.7%) were positive at a reciprocal titer of 40 or over (titer range 40-10,240). Control sera from 75 healthy dogs, not living in endemic areas, and 11 dogs with other diseases (babesiosis, cryptococcosis, ehrlichiosis, dermatitis, and chronic hepatitis) but Leishmania-negative were used to determine the specificity of the assays. All the sera were negative with IFA (100%), whereas using the dot-ELISA only 74 sera (86%) from controls gave a negative result. In the standard IFA no cross-reactivity was noted, in the dot-ELISA a weak cross-reaction was observed with a serum sample from a dog with babesiosis. The interpretation of dot-ELISA could be easily performed by visual inspection of the nitrocellulose disks. The most remarkable feature of dot-ELISA was the high sensitivity (91.9%) and positive predictive value (92.6%), even if the test showed a specificity lower than IFA. Because of its easy performance and high sensitivity, the dot-ELISA may be a useful test to be executed in the field for the diagnosis of canine leishmaniosis.

Animals↗
PubMed1979

Antigenic components of Taenia saginata and their relevance to the diagnosis of bovine cysticercosis by immunoelectrophoresis.

22 antigenic components of Taenia saginata proglottides were detected by immunoelectrophoretic (IEP) study. Cross-absorption of the hyperimmune serum, raised in rabbits against T. saginata, with heterologous and host antigens showed that two thirds of these components were not specific for the cestode. Among the calves experimentally infected with T. saginata eggs, 3 precipitation patterns in IEP were identified depending on the evolutionary stage of the infection. The hydrosoluble extract of T. saginata was used as antigen. A short precipitating arc present near the antigen well and appearing 3 to 4 weeks post-infection of the calves and a long precipitating arc extending towards the anodic end of the slide and appearing in later stages of infection were found of high diagnostic value. No false-positive reaction or cross-reaction were found. Among the calves harbouring less than 50 cysticerci, false-negative reactions were seen.

Animals↗
PubMed2004

Validation of the Cozart microplate EIA for cocaine and metabolites in oral fluid.

The purpose of these studies was to determine the performance characteristics of the Cozart microplate EIA for the detection of cocaine and cocaine metabolites in oral fluid. A total of 217 samples were collected using the Cozart RapiScan oral fluid Collection System from donors who were receiving treatment and being monitored for their addiction. The samples were screened in the laboratory using the Cozart microplate EIA for cocaine and metabolites and confirmed using gas chromatography-mass spectrometry (GC-MS). The samples were stored frozen until analysis by GC-MS. Of the 217 samples tested, 116 were positive for cocaine and/or cocaine metabolites. The Cozart microplate EIA for cocaine, using a cutoff of 30 ng/mL benzoylecgonine equivalents in neat oral fluid, had a sensitivity of 95.7% +/- 2.0% and specificity of 100% +/- 0.7%. A series of potential adulterants of oral fluid were evaluated and shown not to alter the outcome of the test result.

Algorithms↗
PubMed2000

Evaluation of autoSCAN-W/A and the Vitek GNI+ AutoMicrobic system for identification of non-glucose-fermenting gram-negative bacilli.

The autoSCAN-W/A (W/A; Dade Behring Microscan Inc., West Sacramento, Calif.) and Vitek AutoMicrobic System (Vitek AMS; bioMérieux Vitek Systems, Inc., Hazelwood, Mo.) are both fully automated microbiology systems. We evaluated the accuracy of these two systems in identifying nonglucose-fermenting gram-negative bacilli. We used the W/A with conventional-panel Neg Combo type 12 and Vitek GNI+ identification systems. A total of 301 isolates from 25 different species were tested. Of these, 299 isolates were identified in the databases of both systems. The conventional biochemical methods were used for reference. The W/A correctly identified 215 isolates (71. 4%) to the species level at initial testing with a high probability of >/=85%. The Vitek GNI+ correctly identified 216 isolates (71.8%) to the species level at initial testing with a high probability of >/=90%. After additional testing that was recommended by the manufacturer's protocol, the correct identifications of the W/A and Vitek GNI+ improved to 96.0 and 92.3%, respectively. The major misidentified species were Sphingomonas paucimobilis and Agrobacterium radiobacter in the W/A system and Acinetobacter lwoffii, Chryseobacterium indologenes, and Comamonas acidovorans in the Vitek GNI+ system. The error rates were 4.0 and 7.6%, respectively. The overall accuracy for both systems was above 90% if the supplemental tests were applied. There was no significant difference in accuracy (P > 0.05) between the two systems.

Automation↗
PubMed2004

Near-patient testing for infection using urinalysis and immuno-chromatography strips.

Urinary tract infections require costly confirmatory tests such as a urine culture to establish the diagnosis. Elimination of the culture step would save resources; diagnosis and treatment could begin in hours rather than days. We tested a new dip-and-read strip that uses immuno-chromatography (IC) to detect infectious agents in urine. We used a goat-derived polyclonal antibody with reactivity to the cell-wall proteins of Escherichia coli (E. coli). Fluorescein linked to the anti-E. coli antibody served to trap the bacteria on a strip coated with an anti-fluorescein mouse antibody. Blue latex particles were linked to anti-E. coli antibodies by standard methods and were used for detection of E. coli. We found that the combination of leukocyte esterase and nitrite dipsticks gave negative predictive values of 93% for culture-negative urines, i.e., there were very few false-negative results. Using the same dipsticks on culture-positive specimens, the positive predictive values were unacceptably low; we obtained too many false-positive values. By contrast, the IC strips gave negative predictive values of 89%. The major advantage of the IC strips is that the positive predictive values were higher, i.e., there were fewer false-positive results. The combined use of both IC strips and urinalysis dipsticks offers the best strategy for diagnosing infection with dipsticks. The IC strip test could reduce the necessity of a urine culture in patients with suspected infections and provide rapid point-of-care testing.

Antibody-Coated Bacteria Test, Urinary↗
PubMed1987

Comparative sensitivity and specificity of ELISA and IHA for serodiagnosis of strongyloidiasis with larval antigens.

Sera from 68 patients with parasitologically proven strongyloidiasis were tested by the ELISA and IHA tests using larval antigens prepared from Strongyloides stercoralis and Strongyloides ratti. The ELISA using the S. stercoralis antigen detected the greatest number of sero-reactors (83.8%), whereas the IHA using the S. ratti antigen detected the fewest (55.9%). In addition, the S. stercoralis antigen had higher geometric mean titers than the S. ratti antigen in both the ELISA and the IHA tests. Sera from 37 presumed normal individuals also were tested by IHA and ELISA and nonspecific reactions were seen only with the IHA test. When sera from patients with parasitic infections other than strongyloidiasis were tested, the only consistent cross-reactions were with those sera from patients who had occult filariasis and acute schistosomiasis.

Antibodies↗
PubMed1994

Molecular genetic analysis of the human Lewis histo-blood group system.

The expression of Lewis histo-blood group antigen is determined by the Lewis type alpha(1,3/1,4) fucosyltransferase encoded in Fuc-TIII gene (Le gene) on chromosome 19. Weak hemagglutination reactions are often observed in the Lewis blood typing to cause false-negative reactions. The level of CA19-9 tumor marker, defined as sialylated Le(a) (sLe(a)), is influenced by the Lewis blood phenotype. The sLe(a) antigen is also considered to play a role in hematogenous metastasis of tumor cells. The accurate genotyping of Fuc-TIII gene would be, therefore, necessary for solving above questions. In addition to the two missense mutations previously reported by us, i.e. the T59G and the G508A, further sequence analyses revealed a new single base substitution, the T1067A, changing the Ile356 to Lys in the catalytic region. Polymerase chain reaction-restriction fragment length polymorphism methods were developed for the detection of these three missense mutations and applied in 15 Lewis negative (le/le) and 65 Lewis positive individuals (Le/Le or Le/le). These methods were found to be useful for genetic diagnosis for the Lewis histo-blood typing. All le alleles had the T59G mutation, whereas none of the Le alleles did. The le alleles were divided in two types, le1, having the G508A mutation, and le2, having the T1067A mutation. The T1067A mutation reduced the enzyme activity less than 10%, whereas the G508A mutation in the catalytic domain made the enzyme completely inactive. The frequency of occurrence of Le, le1, and le2 in the Japanese population by random sampling study was found to be 66, 30, and 4%, respectively.

Alleles↗
PubMed1995

Detection of Brucella abortus antigens by immunoperoxidase histochemical staining of lochia smears.

An immunoperoxidase histochemical staining technique for the detection of Brucella abortus organisms in lochia of brucellosis-infected cows was described. In comparison with the standard culture technique, the former method resulted in 36% false positive and 2.3% false negative reactions, which represents a sensitivity of 82% and a specificity of 94%. False positive reactions are presumably the result of organisms which succumbed during transit. The feasibility of the test in the confirmation of brucellosis is discussed.

Animals↗
PubMed2000

Evaluation of three methods for using the Duotip-Test device for skin testing.

The sensitivity and precision of rotation, prick and puncture methods of using the Duotip-Test for epicutaneous allergy skin testing were evaluated. Forty-one volunteers who had not taken any antihistamines within the previous two weeks were recruited. The mean age was 21.6 years (range 18 to 25 years). Histamine hydrochloride 1 mg/ml and 50% glycerol saline were used as positive and negative controls, respectively. Each method of testing was performed in triplicate on the volar surface of both forearms. Wheal and flare were measured 15 minutes later. Rotation, prick and puncture methods produced histamine mean wheal diameter +/- standard deviation of 6.61 +/- 0.87 mm, 3.86 +/- 1.03 mm, and 3.00 +/- 0.65 mm, respectively (p < 0.01). The coefficient of variation of rotation method was 13.13%. It was the only method that gave coefficient of variation lower than 20%. False negative and false positive proportions of rotation method using a 4 mm criterion for positive reaction were 1.5% and 0.75%, respectively. Rotation method was well accepted by the volunteers although it was ranked highest in pain. We concluded that the rotation method of using Duotip-Test is a highly reliable technique for skin testing.

Adolescent↗
PubMed1977

Immunodiffusion and agglutination tests for Candida in patients with neoplastic disease: inconsistent correlation of results with invasive infections.

A serological study of candidiasis was conducted with use of (1) sera sent to the laboratory by clinicians who suspected infections due to various organisms including Candida species, and (2) sera available in our serum bank from patients with candidiasis proven at autopsy and from those with documented candidemia. With this population of patients, we were able to evaluate both potential false-negative and false-positive results. Microimmunodiffusion and slide agglutination tests were used. In many cases, serial specimens were available for measurement of rises or falls in titers of agglutinating antibody. Sera from less than one-half of the patients with disseminated or invasive gastrointestinal candidiasis exhibited positive immunodiffusion reactions, titers of agglutinating antibody of greater than 1:16, or fourfold rises in titer of agglutinins. Sera from several patients with pharyngitis due to Candida and from several who were only colonized with Candida or for whom clinical or cultural evidence of candidiasis was lacking showed positive immunodiffusion reactions or agglutinin titers of greater than 1:16, and some showed fourfold rises in titer. Conversions from negative to positive immunodiffusion reactions were not consistently correlated with invasive candida infection. Using the methods described, we have not found immunodiffusion tests and titers of agglutinating antibody to be reliable indicators of invasive candida infection, since false-positive as well as false-negative reactions occur. Greater specificity for invasiveness as well as greater sensitivity in immunosuppressed patients are necessary before these tests can become important adjuncts to the evaluation of patients with suspected invasive candidiasis.

Adolescent↗