Rapid detection of Salmonella enterica serovar Choleraesuis in blood cultures by a dot blot enzyme-linked immunosorbent assay.

A dot blot enzyme-linked immunosorbent assay (ELISA) with a monoclonal antibody specific to phase1-c Salmonella was developed for the direct detection of Salmonella enterica serovar Choleraesuis in blood cultures. This system was applied to the identification of serovar Choleraesuis, and the results were compared with those obtained by a conventional biochemical method. It was revealed that all 12 samples identified to be infected with serovar Choleraesuis were positive on testing by the ELISA. In contrast, 77 samples infected with bacteria commonly isolated from the blood were not reactive by the ELISA. The calculated sensitivity and specificity of the established assay are 100%.

Immunological detection of Salmonella paratyphi A in raw prawns.

A slot blot enzyme-linked immunosorbent assay, using monoclonal antibodies specific only for Salmonella paratyphi A, to detect S. paratyphi A contamination in raw prawns has been established. When artificially contaminated prawn samples were tested. S. paratyphi A contamination could be identified correctly within 20 h. No false positives from samples artificially contaminated by other microorganisms were obtained. The sensitivity was such that as few as 1 S. paratyphi A organism per g of raw prawn could be detected. Therefore, the assay constituted a promising test for the rapid and specific detection of S. paratyphi A in prawns.

Antimicrobial resistance of 100 serial gram-negative isolates in two intensive care units.

To determine antimicrobial resistance pattern among gram-negative bacteria isolated from suspected sources of infections in patients hospitalized in two Intensive Care Units (ICUs) at Siriraj Hospital from September 1991 to December 1991, minimal inhibitory concentrations of one-hundred consecutive gram-negative isolates for various antimicrobials were performed using the microbroth dilution method. Of all gram-negative bacterial isolates, 25 per cent were Pseudomonas aeruginosa, 22 per cent Acinetobacter anitratus, 16 per cent Klebsiella pneumoniae, 12 per cent enterobacter, 8 per cent E.coli, 5 per cent non-fermenter, 4 per cent pseudomonas, 3 per cent arizona, 2 per cent A. lwoffii, 1 per cent Aeromonas hydrophila, 1 per cent Aeromonas hydrophila, 1 per cent Proteus rettgeri, and 1 per cent shigella. The in vitro MIC study revealed that 50, 48, 43, 61, 59, 34, 47, 52, 31, 15 per cent of gram-negative isolates were resistant to gentamicin, tobramycin, amikacin, cefotaxime, ceftriaxone, ceftazidime, aztreonam, piperacillin, ciprofloxacin and imipenem respectively. In addition, 64 and 71 per cent of the isolates were resistant to aminoglycosides and cephalosporins being used in the same patients 48 hours before cultures were obtained respectively. The possible spread of resistant gram-negative isolates by cross contamination was not evident by looking at MIC co-variation in sequential isolates of P. aeruginosa. It was concluded that antimicrobial resistance was highly prevalent among gram-negative bacteria isolated from patients already hospitalized in the ICUs. Potent antimicrobials such as imipenem, newer fluoro-quinolones, ceftazidime and amikacin, are often needed for therapy of serious gram-negative bacterial infections in the ICUs.

Dot-blot ELISA for the detection of IgM RF and IgA RF.

Dot-blot ELISA was developed for the detection of IgM RF and IgA RF. Normal rabbit IgG (NRIgG), concentration 100 micrograms/ml, was used as the antigen for dotting on the 0.45 microns pore size nitrocellulose membrane. Serum, conjugate and substrate incubation conditions were at room temperature for 1 hour, 1 hour and 3 minutes, respectively. The membrane with NRIgG dot could be sotred for 6 weeks before use in the assay. Positive results of IgM RF, at the serum dilution 1:800, were found in 31/51 patients with either classical or definite rheumatoid arthritis and 3/68 normal healthy individuals. Positive IgA RF, at the serum dilution 1:100, was found in 27/51 of the former and none of the latter. Significant concordance with high agreement index was found between the results of the dot-blot ELISA developed and those obtained from ELISA performed in microtitre plate (Kappa greater than or equal to 0.78 for IgM RF and 0.83 for IgA RF, p less than 0.001).

Re-evaluation of ELISA and latex agglutination test for rheumatoid factor detection in the diagnosis of rheumatoid arthritis.

An indirect ELISA for the determination of each isotype (IgM, IgG, IgA, IgD, IgE) of rheumatoid factors (RF) was performed with sera obtained from 77 patients with either classical or definite rheumatoid arthritis (RA) and 319 controls, using rabbit IgG as the antigen. The results were compared with those of a commercial latex agglutination test, using denatured human gamma globulin as the antigen for rheumatoid factor determination. At the cut-off level at which positive results were found in less than 5% of normal controls, ELISA for IgM RF determination had sensitivity, specificity, efficiency, positive predictive value and negative predictive value of 46.75%, 98.12%, 88.13%, 85.71%, 88.41%, while those for IgA RF were 46.75%, 93.42%, 84.34%, 63.16%, 87.91% and for IgG RF were 59.74%, 92.16%, 85.86%, 64.78%, 90.46%, respectively. These indices by latex agglutination test were 83.11%, 93.73%, 91.67%, 76.19% and 95.83%, respectively. IgD RF titre greater than or equal to 1:5 was detected in 19/77 RA patients and 4/200 normal controls while IgE RF titre greater than or equal to 1:5 was detected only in 7/77 RA patients. Thus, ELISA did not appear to have any advantage over latex agglutination test for diagnosis of RA.

HIV infection in male patients attending a sexually transmitted disease clinic.

Three hundred and fifty-two heterosexual males, attending a sexually transmitted disease clinic at Siriraj Hospital from December 1989 to February 1991 were studied for the prevalence of HIV infection. Of these, 334 men reported prostitutes as the main source of their sexually transmitted disease. No one had received blood transfusion in the last 5 years, and there was no history of intravenous drug use, homosexuality or bisexuality. HIV antibody was found in the sera of 24 men (6.8%). HIV seropositivity was associated with serologic makers of syphilis (P < 0.05) but was not associated with present genital ulcers on physical examination or other STDs. These data indicate the high rate of female prostitutes to male transmission of HIV infection in the presence of sexually transmitted disease and confirms the relationship between syphilis and HIV infection. HIV/AIDS educational programmes and campaigns to promote condom use among prostitutes and clients are an urgent need in Thailand.

PIP: The study subjects were recruited among heterosexual men attending the male sexually transmitted disease (STDs) clinic operated by the Dermatology Unit, Siriraj Hospital, Mahidol University, Bangkok, Thailand. The subjects had no history of intravenous drug use, homosexuality, or bisexuality, had not received blood transfusion in the preceding 5 years, and claimed that they had contracted the disease from prostitutes. Between December 1989 and February 1991, 352 men enrolled in the study who had a median age of 28 years (range 15-63 years). The participants completed a questionnaire about occupation and clinical symptoms of STDs. Sera were assayed for VDRL and TPHA. HIV antibody screening was performed by gel particle agglutination or ELISA technique, and the specimens were confirmed as positive by HIV antibody Immunoblot technique. Of the 328 HIV seronegative men, 44% had nonspecific urethritis, 13.3% had gonorrhea, 13.1% had genital ulcers (including syphilis and chancroid), and 7.6% had syphilis (including positive VDRL or TPHA 1:160). 334 of 352 men (94.9%) reported prostitutes as the source of their STDs. HIV antibody was detected in 19 (5.4%) of 352 men. Only 100 of the 333 men whose first HIV antibody was negative returned to the clinic for a second HIV antibody test within 12 weeks, and HIV antibody was detected in 5 (5.0%) of these 100 men. Thus, the HIV antibody was found in a total of 24 (6.8%) of 352 men. This rate was 15 times the rate found in blood samples from healthy donors at Siriraj Hospital during the period between 1989 and 1990. The HIV seropositivity was significantly associated with syphilis (including positive VDRL or TPHA 1:160), but was not associated with genital ulcers (including syphilitic ulcer and chancroid). None of the 24 seropositive men had clinical evidence of AIDS-related complex or full-blown AIDS. The lack of association between HIV seropositivity and genital ulcer remains to be further investigated.