Clinical utility of PCR assay of a target Sequence on the the 31 - kilodalton brucella antigen DNA in human brucellosis

Human brucellosis poses a significant public health problem in many developing countries and requires fast and accurate diagnosis. The diagnosis of brucellosis is frequently difficult to establish, not only because clinically, the disease may mimic other infectious and non infectious disease, but also because the established diagnostic methods are not always successful. In order to overcome some of the limitations of the conventional microbiological techniques in the diagnosis of human brucellosis, a PCR-Enzyme Linked Immunosorbent Assay [PCR-ELISA] with genus-specific primers was developed for the amplification of a 223-bp sequence of a gene encoding for the synthesis of a 31-KDa immunogenic membrane protein specific for the Brucella genus [BCSP31], with a detection limit of 10 f g bacterial DNA [2 bacterial cells] for the rapid diagnosis of human brucellosis. In this study, we collected peripheral blood samples from 80 patients with active brucellosis diagnosed on the basis compatible clinical picture with positive blood culture and/or positive serology; and also from control subjects composed of 25 volunteer blood donors, 14 patients with febrile illness due to factors other than Brucella etiology, and 11 asymptomatic occupationally exposed persons. The diagnostic yield of PCR-ELISA assay in human Brucellosis was evaluated by comparing its results in the patient and control groups with those of the conventional microbiological techniques such as serology, using the Standard Tube Brucella Agglutination Test [SAT], and blood cultures for Brucella using the semiautomatic Bactec 9240 system. Out of the 80 patients with active brucellosis, 37 [46.25%] were blood culture positive for Brucella, and 78 [97.5%] were serology positive. All of these 80 cases were PCR-ELISA was 100% followed by 97.5% of serology and 46.25% of blood culture. Five cases among the patient group showed focal [localized] manifestations, three of them presented with chest manifestations or disease and two with osteoarthyitis, diagnosis of these cases was established on the basis of clinical and serological critia and confirmed by PCR-ELISA assay. All of the 50 control blood samples were blood culture negative. While, 2 samples from the occupationally exposed group showed high titer specific brucella antibodies >/-160. PCR-ELISA was able to detect 3 positive cases in the control group, all of them were also from the occupationally exposed asymptomatic group. Thus, the specificity of blood culture was 100% followed by 96% of serology and 94% of PCR-ELISA. The corrected specificity of the PCR-ELISA and serology after exclusion of the occupationally exposed asymptomatic group was 100%. We concluded that PCR assay is far more sensitive than the conventiomal techniques and this, coupled with its speed and reduction in risk to laboratory workers, makes this technique a very useful practical tool for the laboratory diagnosis of human brucellosis

Post antibiotic effect and its efficacy on enhanced microbial susceptibility to phagocytosis

In the present work the in vitro post antibiotic effects of Amoxycillin/ Clavulanic acid [Augmentin] on Staphylococcus aureus and of Amikacin and Cefotaxime on Pseudomonas aeruginosa were determined by the broth technique. These organisms were hospital strains isolated from two patients in Ain Shams University Hospitals. The exposure of Staph. aureus to 5X minimum inhibitory concentration [MIC] of Augmentin for 2 hours produced a significant post-antibiotic effect [PAE] of 1.7 hour. As well as, exposure of Pseudomonas aeruginosa to 5X MIC of Amikacin and Cefotaxime for 1 hour produced a significant PAE of 2.4 hours and 1.8 hour respectively. Amikacin had a longer PAE than Cefotaxime under the same conditions. Moreover, post-antibiotic leucocyte enhancement [PALE] or the effect of previous exposure to antibiotics on the susceptibility of bacteria to the phagocytic and bactericidal activities of polymorphonuclear leucocytes [PMNL] had been tested. The PAE of Augmentin on S. aureus and of Amikacin or Cefotaxime on Ps. aeruginosa caused a significant increase in their susceptibility to the antibacterial activities of human PMNL, compared with the untreated control cells under the same test conditions. We concluded that, the PAE has many biological significances, this effect may justify the use of PAE inducing antimicrobial agents with longer dosing interval than those currently employed without loss of efficacy. Also, PAE induces the post-antibiotic leucocyte enhancement, organisms in the PAE phase are more susceptible to the antibacterial activity of human PMNL. Further studies are necessary to determine the full PAE clinical significances

Correlation of a simple biochemical marker and intraamniotic infection in patients with preterm premature rupture of membranes

Subclinical intramniotic infection has been associated with preterm labour and preterm premature rupture of membranes. Prompt diagnosis of subclinical intraamniotic infection is important in that it may prevent the maternal and neonatal morbidity associated with clinical chorioamnionitis. While amniotic fluid cultures are widely used to assess the microbiologic state of the amniotic cavity, results are not immediately available and are not capable of covering a wide range of organisms with different culture requirements. Rapid methods for detection of infection such as Gram stain and acridine orange stain of AF are insensitive. This study evaluated AF glucose concentration as an early parameter of intraamniotic infection. Amniocentesis for 40 patients with preterm PROM