[Usefulness of anti-SS-A/Ro antibody measurement based on fluorescence enzyme immunoassay with Ro60 and Ro52 antigen].

OBJECTIVE: Anti-SS-A/Ro antibody is commonly found in the sera of patients with rheumatic disease. The antigenic components of SS-A/Ro are 60kD. and 52kD-Ro protein. Only anti-60kD SS-A/Ro antibody is detected by commonly used tests in clinical laboratories. "UniCAP EliA SS-A/Ro" is a new reagent using fluorescence enzyme immunoassay (FEIA) based on a mixture of both 60kD- and 52kD-Ro antigens. We evaluated the efficacy of detecting both anti-60kD and 52kD SS-A/Ro antibodies. PATIENTS AND METHODS: We collected sera from 264 rheumatic disease patients and 106 healthy subjects. Anti-SS-A/Ro antibodies were measured by the new reagent along with conventional method. Anti-52kD and 60kD SS-A/Ro antibodies were measured by ELISA kit in rheumatic disease patients. RESULTS: Anti-SS-A/Ro antibody levels were higher in patients with Sjögren's syndrome than those with SLE or RA. The prevalence of anti-SS-A/Ro antibody in rheumatic disease patients and healthy subjects were comparable with the conventional method, and patients with Sjögren's syndrome had highest prevalence of anti-SS-A/Ro antibody. Concordance rate between EliA and conventional method, and EliA and DID, were 96.6% and 94.3%, respectively. ELISA analysis revealed that patients with Sjögren's syndrome had anti-52kD SS-A/Ro antibody at high rates, while anti-60kD SS-A/Ro antibody was widely found in rheumatic disease patients. Three patients were positive only for anti-52kD SS-A/Ro antibody. CONCLUSION: Taken together, "UniCAP EliA SS-A/Ro" is useful as a screening test for anti-SS-A/Ro antibodies.

[A case of Salmonella-infected thoracoabdominal aortic aneurysm making final diagnosis difficult].

We report a case of thoracoabdominal aortic aneurysm (TAAA) due to Salmonella Enteritidis making final diagnosis difficult. A 63-year-old man with a history of diabetes mellitus, hypertension, and cerebral infarction was seen elsewhere for a 40 degrees C fever, vomiting, and shaking on day 1 after onset. He was diagnosed with Salmonella bacteremia and hospitalized by us for intensive care. Computed tomography (CT), magnetic resonance imaging (MRI), and ultrasound imaging did not, however, show critical findings of aneurysm, endocarditis, or osteomyelitis, and laboratory testing suggest significant inflammatory symptoms. He did not respond to antibiotics, but had an intermittent low fever during the first hospitalization. On day 48 after onset during the second hospitalization, abdominal CT showed an aneurysm -3 cm in diameter in the thoracoabdominal aorta above the renal artery- small enough to have been missed in earlier diagnosis. Surgery and TAAA graft replacement were done on day 64. Bacterial culture of the graft showed no Salmonella growth due to long-term in vivo antibiotic exposure. He recovered without significant complications, with oral ciprofloxacin antibiotic therapy continued to the present. This case indicates the importance of an early diagnosis through continuous blood culture and imaging for Salmonella sp blood stream infection.

Phenotypic and genotypic characterization of Vibrio cholerae clinically isolated in Surabaya, Indonesia.

The phenotypic and genotypic characteristics of 6 clinical strains of Vibrio cholerae isolated in Surabaya, Indonesia in 2009 were examined. The DNA fingerprints obtained suggested that these isolates were not from a single clone. Furthermore, all isolates produced cholera toxin and possessed the classical type of toxin B subunit gene, thus meaning that this is the first report of the occurrence of El Tor variants of V. cholerae in Indonesia. Although all isolates were sensitive to almost all antibiotics tested, including ampicillin, chloramphenicol, ciprofloxacin, gentamicin, levofloxacin, kanamycin, nalidixic acid, norfloxacin, streptomycin, trimethoprim-sulfamethoxazole, and tetracycline, and had no mutation in the gyrA and parC genes, they nevertheless possessed the class 1 integron that is a molecular vehicle for the acquisition of antibiotic resistance genes, suggesting that they have the potential to acquire the genetic element for drug resistance.

Risk factors and mechanisms of fluoroquinolone resistance in 156 Escherichia coli strains clinically isolated from urinary tract infections.

As fluoroquinolone-resistant strains of Escherichia coli emerge, several risk factors for fluoroquinolone resistance have become evident, such as amino acid mutations in the quinolone resistance determining regions (QRDR) of gyrA and parC and previous use of fluoroquinolone. This study investigated risk factors for fluoroquinolone resistance and amino acid mutation in the QRDR in E. coli. We investigated the statistical correlation between each amino acid mutation and resistance to levofloxacin. We examined the minimum inhibitory concentration (MIC) of levofloxacin and the amino acid mutations of gyrA and parC by direct DNA sequence in E. coli clinically isolated from urinary tract infection (UTI) patients. We investigated risk factors for levofloxacin resistance, such as age, sex, and previous use of fluoroquinolone. We found a significant correlation between the number of mutations and resistance to levofloxacin (p < 0.001) and between the presence of underlying urinary tract disease and the presence of mutations (p = 0.004) by multivariate analyses. Three mutations in QRDR were demonstrated to be significantly correlated with levofloxacin resistance. In conclusion, these findings contribute to our understanding of the molecular mechanisms and risk factors for fluoroquinolone resistance.

Correlation of overexpression of efflux pump genes with antibiotic resistance in Escherichia coli Strains clinically isolated from urinary tract infection patients.

Escherichia coli is one of the most common pathogens in urinary tract infections (UTIs), and antibiotic resistance in E. coli is becoming a serious problem in treating UTI. Efflux system overexpression is reported to contribute to E. coli resistance to several antibiotics. This study investigated the correlation of antibiotic susceptibilities with the overexpression of the efflux pump genes such as marA, yhiU, yhiV, and mdfA and with risk factors for antibiotic resistance in E. coli isolated from UTI patients. We examined the expression level of efflux pump genes using quantitative real-time reverse transcription-PCR (qRT-PCR). We also tested the in vitro susceptibilities to 12 kinds of antibiotics in 64 clinical strains of E. coli isolated from UTI patients. By multivariate analyses we revealed significant relationships between the overexpression of (i) marA and MICs of cefepime (FEP) and nalidixic acid (NAL), (ii) yhiV and MICs of minocycline (MIN), and (iii) mdfA and MICs of sitafloxacin (STX). In our investigation of the efflux pump genes, risk factors such as gender and the previous use of fluoroquinolones correlated with the overexpression of marA, and indwelling catheter use correlated with the overexpression of mdfA. In conclusion, we demonstrated that the increased expression of efflux pump genes such as marA and mdfA can lead to fluoroquinolone resistance in E. coli. These results contribute to our knowledge of the efflux system and raise the possibility of developing new agents, such as efflux pump inhibitors (EPIs), to antibiotic-resistant E. coli.

Isolated true parachute mitral valve in an asymptomatic elderly patient.

We report the extremely rare case of a 73-year-old asymptomatic patient who has an isolated true parachute mitral valve (PMV). In the echocardiographic examination, the parasternal long-axis view showed a single papillary muscle. The short-axis view revealed the presence of a symmetric mitral valve orifice with all chordae attaching to a large anterolateral papillary muscle. Because detailed examination did not reveal the presence of other complications, this patient was diagnosed as an isolated true PMV.

[Evaluation of simultaneous detection of specific antinuclear antibodies using multiplexed technology].

BioPlex2200 is a fully automated analyzer using multiplexed technology and BioPlex2200 ANA Screen can analyze 11 antinuclear antibodies (ANA). We evaluated simultaneous detection of 11 ANA and clinical utility as ANA screening test. We collected sera from 317 connective tissue disease (CTD) patients, 166 healthy subjects, and 105 sera in which anti dsDNA antibody(RIA) is requested. Detection of 11 ANA were measured by BioPlex2200 ANA Screen using BioPlex2200 along with conventional methods. We evaluated positive ratio for healthy subjects and CTD patients and concordance rate between BioPlex2200 and IF. The prevalence of disease specific ANA in CTD patients were comparable with general occurrence rate except anti dsDNA antibody (39.3%) in SLE. Concordance rate between BioPlex2200 and conventional methods were high(89.0-99.7%) except anti dsDNA antibody(ELISA: 68.8%, RIA: 58.1%). Discordant sera of Bioplex2200+/DID- for anti SS-A antibodies were observed in 30 sera, and 20 sera were positive only for anti 52kD SS-A/Ro antibody. As ANA screening test, positive ratio was low (7.2%) in healthy subjects, and comparable with that of IF(1:160). Concordance rate between BioPlex2200 and IF was low (75.1%). However, 44 sera of BioPlex2200+/IF- contained 6 samples positive for anti Jo-1 antibodies and 29 samples positive for anti-52kD SS-A/Ro antibodies. Diagnostic accuracy of the Medical Decision Support Software (MDSS) by BioPlex2200 as compared to clinical diagnosis is good for specificity. Taken together, BioPlex2200 can appropriately perform simultaneous detection of 11 ANA and detect independently anti-52kD SS-A/Ro antibody. However, detection for anti dsDNA antibody of low titers is needed to be improved.

[Comparison of detection sensitivity in rapid-diagnosis influenza virus kits].

Rapid-diagnosis kits able to detect influenza A and B virus by immunochromatography developed by different manufacturers, while useful in early diagnosis, may vary widely in detection sensitivity. We compared sensitivity results for eight virus-detection kits in current use--Quick Chaser FluA, B (Mizuho Medy), Espline Influenza A & B-N (Fujirebio), Capilia Flu A + B (Nippon Beckton Dickinson & Alfesa Pharma), Poctem Influenza A/B (Otsuka Pharma & Sysmex), BD Flu Examan (Nippon Beckton Dickinson), Quick Ex-Flu "Seiken" (Denka Seiken), Quick Vue Rapid SP Influ (DP Pharma Biomedical), and Rapid Testa FLU stick (Daiichi Pure Chemicals)--against influenza virus stocks, contained five vaccination strains (one A/H1N1, two A/H3N2, and two B) and six clinical strains (two A/H1N1, two A/H3N2, and two B). Minimum detection concentrations giving immunologically positive signals in serial dilution and RNA copies in positive dilution in real-time reverse transcriptase-polymerase chain reaction (RT-PCR) were assayed for all kits and virus stock combinations. RNA log10 copy numbers/mL in dilutions within detection limits yielded 5.68-7.02, 6.37-7,17, and 6.5-8.13 for A/H1N1, A/H3N2, and B. Statistically significant differences in sensitivity were observed between some kit combinations. Detection sensitivity tended to be relatively higher for influenza A than B virus. This is assumed due to different principles in kit methods, such as monoclonal antibodies, specimen-extraction conditions, and other unknown factors.

Emergence of fluoroquinolone-resistant strains of Salmonella enterica in Surabaya, Indonesia.

Typhoid fever remains a major health problem in developing countries. Fluoroquinolones such as ciprofloxacin emerged as the 1st-choice treatment of enteric fever, including typhoid, in the 1990s. Recently, Salmonella typhi strains with resistance to ciprofloxacin have been increasingly reported in several countries, although the fluoroquinolone-resistant clinical strain has not been reported in Indonesia. In the present study, we examined the drug susceptibility and the presence of gyrA mutations in 17 clinical strains of S. typhi isolated from Surabaya, Indonesia, in 2006 (9 strains) and 2008 (8 strains). Although all 9 isolates from 2006 were sensitive to all tested antibiotics and had no mutation in the gyrA gene, all 8 isolates from 2008 were resistant to nalidixic acid and ampicillin and had a gyrA mutation at codon 87. In addition, 3 of 8 strains from 2008 showed multiple drug resistance, including resistance to chloramphenicol, trimethoprim-sulfamethoxazole, and ciprofloxacin. Therefore, newer drugs, such as ceftriaxone, cefixime, and azithromycin, might be effective in this situation. This is the 1st report of the emergence of fluoroquinolone-resistant clinical strains of S. typhi with a gyrA mutation, and it reveals a health risk due to multidrug-resistant strains in Indonesia.

[Usefulness of the formula for predicting creatinine clearance from serum cystatin C].

Estimation of Glomerular filtration rate (GFR) is crucial for the detection of renal insufficiency. In clinical practice, GFR is generally estimated from Ccr by some prediction formulas including the serum creatinine concentration and some variables: age, sex, body size. It has been suggested that serum cystatin C is less influenced by sex, body size, muscular mass or inflammation. In several studies, serum cystatin C performed better than serum creatinine did as a marker to detect GFR reduction. We developed the formula for predicting creatinine clearance (Ccr) from serum cystatin C, serum creatinine and serum beta 2-microgroburin by multiple regression analysis. In this study, we analysed 24-hour Ccr and variables (sex, age and BMI) in 82 subjects with renal diseases to develop suitable formulas. Our serum cystatin C formula is as follows: Ccr (ml/min/1.73m2) = (12.14-0.03 x age-1.72 x serum cystatin C) 2. Correlation between measured 24-hour Ccr and predicted Ccr by our serum cystatin C formula was higher (r=0.852) than our serum creatinine formula (r=0.755), serum beta 2-microgroburin formula (r=0.793), Cockcroft-Gault formula (r=0.836) and Horio formula (r=0.825). Accuracy within 15% was higher (39.0%) than other formulas (25.6-30.5%). Our formula using serum cystatin C for predicting Ccr is a useful and precise marker for Ccr than other commonly used formulas using serum creatinine.