Bo-Moon Shin

Multicentre study of the prevalence of toxigenic Clostridium difficile in Korea: results of a retrospective study 2000-2005.

The prevalence of toxigenic Clostridium difficile in Korea has been reported to be approximately 60-80%. Although the prevalence of the tcdA(-)tcdB(+) C. difficile strain was less then 5% prior to the year 2000, it has become an emerging nosocomial pathogen in Korea. Therefore, we have attempted to determine the multicentre nationwide prevalence of tcdA(+)tcdB(+) and tcdA(-)tcdB(+) C. difficile for epidemiological purposes. C. difficile strains (n=724, 30 from 2000, 80 from 2001, 74 from 2002, 76 from 2003, 179 from 2004, 285 from 2005) were obtained retrospectively from January 2000 to December 2005 from in-patients at 6 hospitals, all of whom were suspected of having C. difficile-associated disease (CDAD), colitis or pseudomembranous colitis. The numbers of participating hospitals varied yearly (1 in 2000, 2 in 2001-2003, 3 in 2004, 5 in 2005). The hospitals were located in Seoul (n=4), Kyunggi Province (n=1) and Busan (n=1), Korea. PCR assays for tcdA and tcdB genes were conducted using 724 unduplicated C. difficile isolates. The mean prevalence of tcdA(+)tcdB(+) and tcdA(-)tcdB(+) C. difficile strains over the 6 years was 51.8 % (38.4-59.3%) and 25.8%(10-56.0%), respectively. The mean prevalence of tcdA(-)tcdB(+) C. difficile strains was less than 7% until 2002, but began to increase in 2003 (13.2%) and achieved a peak in 2004 (50.3%). In 2005, the mean prevalence of tcdA(+)tcdB(+) and tcdA(-)tcdB(+) C. difficile strains was 47.7% (30.9-60.3%) and 27.0% (17.6-54.8%), respectively. This nationwide epidemiological study showed that tcdA(-)tcdB(+) C. difficile strains have already spread extensively throughout Korea, and our results provide basic data regarding the controversies currently surrounding the toxigenicity of tcdA(-)tcdB(+) C. difficile. The use of enzyme immunoassays capable of detecting both TcdA and TcdB is strongly recommended for the diagnosis of CDAD in microbiology laboratories, in order to control the spread of the tcdA(-)tcdB(+) strains of C. difficile.

Seroprevalence of Hepatitis B Virus among Health Care Workers in Korea

We studied the seroprevalence of HBsAg, anti-HBs and anti-HBc and the vaccination histories among health care workers (HCWs) at a large suburban referral hospital in Korea. The purpose of this study was to determine the immune status of HCWs against hepatitis B virus and we also wanted to prepare a practical guideline to protect HCWs from occupational exposure. During December, 2003, 571 HCWs (56 physicians, 289 nurses, 113 technicians and 113 aid-nurses) aged between 21 and 74 yr were included in the surveillance. The positive rates of HBsAg and anti-HBs were 2.4% (14/571) and 76.9% (439/571), respectively. The positive rate of anti-HBs was lower in the physician group, and this was associated with the male gender and older age. Of the 439 anti-HBs positive cases, 320 cases (73.1%) were anti-HBc negative and this was significantly associated with a past history of HBV vaccination. The distribution of the anti-HBs levels was not associated with age (except for HCWs in their sixties), gender or occupation. Our study revealed that the seroprevalence rates of HBsAg and anti-HBs in HCWs in Korea were not different from those of the general population. Based on this surveillance, we can make reasonable decisions in case of occupational exposure to hepatitis B virus.

Algorithm Combining Toxin Immunoassay and Stool Culture for Diagnosis of Clostridium difficile Infection

ABSTRACT Enzyme immunoassays (EIA) to detect glutamate dehydrogenase or toxins A (TcdA) and B (TcdB), a cytotoxicity assay, and bacteriologic culture have disadvantages when applied individually to diagnosis of Clostridium difficile infections. Stool specimens (n = 1,596) were subjected to toxin detection via an enzyme-linked fluorescent immunoassay (ELFA; Vidas CDAB assay) and bacteriologic culture for toxigenic C. difficile in a three-step algorithm with additional toxigenic culture. Isolates (n = 163) from ELFA-negative stool specimens were examined via ELFA for toxin production. We amplified tcdA and tcdB from C. difficile isolates and tcdB from stool specimens that were ELFA positive or equivocal and culture negative, and we compared the results to those obtained with the three-step algorithm. More than 26% of stool specimens (419/1,596) were culture positive, yielding 248 isolates (59.2%) with both toxin genes (tcdA- and tcdB-positive isolates), 88 isolates (21.0%) with either tcdA or tcdB, and 83 (19.8%) that had no toxin genes (tcdA- and tcdB-negative isolates). Among 49 (culture-negative/ELFA-positive or -equivocal) stool specimens, 53.1% (26/49) represented tcdB-positive isolates. Therefore, the total number of PCR-positive cases was 362, and 27.1% (98/362) of these were detected through toxigenic culture. The sensitivity, specificity, positive predictive value (PPV), and negative predictive value (NPV) were 63.3%, 96.7%, 90.5%, and 92.4% (ELFA alone); 92.8%, 93.3%, 80.2%, and 97.8% (culture); and 70.7%, 91.4%, 95.5%, and 100% (three-step algorithm ELFA and bacterial culture with toxigenic culture), respectively, with culture and PCR for tcdA and tcdB as the standards. Thus, sensitivity and specificity were highest using culture and ELFA, respectively, but we recommend the three-step algorithm comprising EIA to detect both toxins and toxigenic culture for C. difficile as a practical method for achieving better PPV and NPV.

Frequent Detection of Pandemic (H1N1) 2009 Virus in Stools of Hospitalized Patients

A novel subtype of influenza A virus called pandemic (H1N1) 2009 virus showed global outbreaks in 2009. Previous studies reported that gastrointestinal symptoms such as diarrhea, vomiting, and abdominal pain were frequent in patients with pandemic (H1N1) 2009 virus infection (3, 4, 5). However, the frequency of viral shedding in this pandemic (H1N1) 2009 virus infection was not well known. We recovered pandemic (H1N1) 2009 virus from stools of hospitalized H1N1-positive patients. From 10 November to 31 December 2009, 1,774 patients were diagnosed as having pandemic (H1N1) 2009 virus infections by real-time reverse transcription-PCR (RT-PCR) from nasopharyngeal swab samples in a tertiary-care hospital located in Seoul, South Korea. Among them, 96 patients required hospitalization due to progression to pneumonia or febrile convulsion, acute gastroenteritis, or underlying illnesses. Their stool samples were requested to be collected at admission, irrespective of gastrointestinal symptoms. A total of 65 patients submitted their stool samples at 0 to 8 days after the onset of flu symptoms and were included in this study (Table ​(Table1).1). Twelve patients (18.5%) complained of diarrhea during the hospitalization period. All of them were self-limited, and only one patient showed a pathogen, Salmonella group B, in routine stool cultures. This study was approved by the Sanggye Paik Hospital institutional review board. TABLE 1. Demographic characteristics categorized by the presence of viral RNA in stools among the study population hospitalized with pandemic (H1N1) 2009 virus Viral RNA in stool was isolated using the QIAamp virus RNA minikit (Qiagen, Crawley, United Kingdom). Pandemic (H1N1) 2009 virus was detected by two kinds of RT-PCR kits, the Anyplex FluA new H1N1 kit (SeeGene, Seoul, South Korea) and TaqMan Influenza A MGB assays (Applied Biosystems). Pandemic (H1N1) 2009 virus was recovered from stools of 16 patients (24.6%). Nine of them (56.3%) did not complain of any gastrointestinal symptoms during hospitalization. Only 4 patients simultaneously presented diarrhea and fecal excretion of pandemic (H1N1) 2009 virus. Comparing stool pandemic (H1N1) 2009-positive and -negative groups, no difference of frequency was noticed for diarrhea (P = 0.683), nausea (P = 0.746), vomiting (P = 0.898), and abdominal pain (P = 0.977) (Table ​(Table11). The clinical significance and pathophysiology of fecal influenza A virus excretion are not yet clear (1, 9). However, avian influenza A (H5N1) virus has been known to cause severe gastrointestinal manifestations and to replicate in human intestinal tissues (2, 7, 8). To et al. detected pandemic (H1N1) 2009 virus in stool from 4 of 9 patients with positive viral culture from the specimen with the highest viral load (6). This finding suggested the fecal shedding of viable pandemic (H1N1) 2009 virions. In this study, fecal excretion of pandemic (H1N1) 2009 virus was not correlated with the presence of gastrointestinal symptoms as described for avian influenza A (H5N1) virus (7). This study has some limitations: outpatients were not included and virus cultures and quantifications were not performed. However, our data show that considerable numbers of patients shed pandemic (H1N1) 2009 virus in their stool, even in the absence of diarrhea. The frequent detection of fecal pandemic (H1N1) 2009 virus excretion deserves attention in the study of infection control to prevent viral dissemination.

Characterization of cases of Clostridium difficile infection (CDI) presenting at an emergency room: molecular and clinical features differentiate community-onset hospital-associated and community-associated CDI in a tertiary care hospital.

ABSTRACT Definition of community-onset, hospital-acquired Clostridium difficile infection (CO-HA-CDI) is difficult in patients presenting with diarrhea at hospitals or outpatient clinics, especially 4 to 12 weeks after the last discharge. We performed C. difficile stool culture for 272 diarrheic patients visiting the emergency room (ER) between January 2006 and June 2010. C. difficile was isolated from 36 cases (13.2%), and isolation rates increased year by year, from 10.1% in 2008 to 12.4% in 2009 and 16.7% in 2010. Among 32 toxin-positive isolates, 13 (40.6%) and 19 (59.4%) were associated with CO-HA-CDI and community-acquired CDI (CA-CDI), respectively, if cases with CDI diagnosed within 12 weeks after discharge were considered hospital associated. The majority (70%) of CO-HA-CDI cases occurred within 2 weeks after hospital discharge, although the interval from discharge to onset of symptoms was as long as 10 weeks. We found via tcdA and tcdB and repetitive sequence PCR analysis, that toxin A-positive/toxin B-positive isolates were the most prevalent in both CO-HA-CDI (53.8%) and CA-CDI (94.7%) cases. Toxin A-negative/toxin B-positive isolates were also still highly associated with HA-CDI cases but were also observed in CA-CDI cases. Younger age, fewer underlying diseases, lack of prior antibiotic use, and genetic diversity of isolates in repetitive sequence PCR were the main characteristics in CA-CDI cases visiting the ER.

Evaluation of four commercial IgG- and IgM-specific enzyme immunoassays for detecting Mycoplasma pneumoniae antibody: comparison with particle agglutination assay.

Diagnosis of Mycoplasma pneumoniae infection is important due to its variable clinical manifestations and absence of response to beta-lactams. Introduction of enzyme immunoassays (EIAs) for serologic diagnosis of M. pneumoniae has made it possible to separate the analyses of specific IgG and IgM antibodies. We compared four different commercial EIAs, ImmunoWELL IgG, IgM (GenBio), Medac IgG, IgA, IgM (Medac), Platelia IgG, IgM (Sanofi Pasteur), and Ridascreen IgG, IgA, IgM (r-Biopharm) with indirect particle agglutination assay (PA), Serodia-MycoII (Fujirebio). We tested 91 specimens from 73 pediatric patients (2-17 yr) hospitalized at a tertiary-care hospital between December 2005 and January 2006. The measurements of IgM EIAs were correlated with PA titers (Spearmans correlation coefficient, from 0.89 to 0.92) with high concordance rates, ranging from 82.4% to 92.3%. However, some negative IgM-EIA results in PA-positive specimens indicated that serial samplings with convalescent sera would be necessary to confirm M. pneumoniae infection.

Comparison of BD GeneOhm Cdiff and Seegene Seeplex ACE PCR Assays Using Toxigenic Clostridium difficile Culture for Direct Detection of tcdB from Stool Specimens

ABSTRACT We evaluated the performances of 2 PCR assays (BD GeneOhm and Seegene ACE) for direct detection of tcdB from stool specimens. The concordance rate between BD and Seegene was 96.3%. The sensitivities, specificities, positive predictive values (PPVs), and negative predictive values (NPVs) of BD and Seegene were 95.7%, 96.5%, 91.8%, and 98.2% and 90.0%, 97.1%, 92.6%, and 96.0%, respectively.

Comparison of VIDAS CDAB and CDA immunoassay for the detection of Clostridium difficile in a tcdA- tcdB+ C. difficile prevalent area.

Enzyme immunoassays for TcdA and/or TcdB are widely used for diagnosis of C. difficile infection. This study compared the performance of the new VIDAS C. difficile Toxin A & B assay (CDAB) with that of the existing VIDAS C. difficile Toxin A II assay (CDA) in a tcdA(-)tcdB(+) prevalent area. A total of 555 fecal samples were cultured and tested using CDAB and CDA. C. difficile was isolated in 150 samples and the concordance rate was 81.8% (454/555) between CDAB and CDA. PCR assays for tcdA and/or tcdB were used as a confirmatory test on C. difficile strains recovered from culture positive cases (n=150) and on fecal specimens in culture negative/CDAB positive or equivocal cases (n=27). The number of tcdA(+)tcdB(+), tcdA(-)tcdB(+), and tcdA(-)tcdB(-) strains on culture positive isolates (n=150) were 75 (50.0%), 41 (27.3%), and 34 (22.7%), respectively. PCR assays for tcdB gene alone in stool specimens (n=27) showed positivity in five cases. The sensitivity of VIDAS CDAB was higher than that of VIDAS CDA (65.3% vs. 29.8%), by more than 2-fold. The specificity of CDAB was almost the same as CDA (93.8% vs. 94.5%). Toxigenic culture of C. difficile isolates in culture positive/VIDAS CDAB negative cases (n=62) additionally detected 22 VIDAS CDAB positive and 9 VIDAS CDAB equivocal cases. The VIDAS CDAB assay detects more tcdA(+)tcdB(+) strains (60% vs. 45.3%) and tcdA(-)tcdB(+) strains (70.7% vs. 0%) compared with VIDAS CDA.

Comparison of ChromID Agar and Clostridium difficile Selective Agar for Effective Isolation of C. difficile from Stool Specimens

Background ChromID Clostridium difficile agar (IDCd; bioMérieux SA, France) is a recently developed chromogenic medium for rapid and specific isolation of C. difficile. We compared the performance of IDCd with that of Clostridium difficile Selective Agar (CDSA). Methods A total of 530 fresh stool specimens were collected from patients with clinical signs compatible with C. difficile infection, and cultures for C. difficile were performed on IDCd and CDSA. C. difficile colonies were identified by spore staining, odor, use of an ANI identification test kit (bioMérieux SA), and multiplex PCR for tcdA, tcdB, and tpi. Results The concordance rate between IDCd and CDSA was 90.6% (480/530). The positivity rates on IDCd on days 1 and 2 (55.6% and 85.0%, respectively) were significantly higher than those on CDSA (19.4% and 75.6%, respectively) (P<0.001 for day 1 and P=0.02 for day 2), but the detection rates on IDCd and CDSA on day 3 were not different (89.4% vs. 82.8%, P=0.0914). On day 3, the recovery rates for non-C. difficile isolates on IDCd and CDSA were 30.2% (160/530) and 22.1% (117/530), respectively (P=0.0075). Clostridium spp. other than C. difficile were the most prevalent non-C. difficile isolates on both media. Conclusions The culture positivity rates on IDCd and CDSA were not different on day 3 but IDCd may allow for rapid and sensitive detection of C. difficile within 2 days of cultivation.

Evaluation of Xpert C. difficile, BD MAX Cdiff, IMDx C. difficile for Abbott m2000, and Illumigene C. difficile Assays for Direct Detection of Toxigenic Clostridium difficile in Stool Specimens

Background We evaluated the performance of four commercial nucleic acid amplification tests (NAATs: Xpert C. difficile, BD MAX Cdiff, IMDx C. difficile for Abbott m2000, and Illumigene C. difficile) for direct and rapid detection of Clostridium difficile toxin genes. Methods We compared four NAATs on the same set of 339 stool specimens (303 prospective and 36 retrospective specimens) with toxigenic culture (TC). Results Concordance rate among four NAATs was 90.3% (306/339). Based on TC results, the sensitivity and specificity were 90.0% and 92.9% for Xpert; 86.3% and 89.3% for Max; 84.3% and 94.4% for IMDx; and 82.4% and 93.7% for Illumigene, respectively. For 306 concordant cases, there were 11 TC-negative/NAATs co-positive cases and 6 TC-positive/NAATs co-negative cases. Among 33 discordant cases, 18 were only single positive in each NAAT (Xpert, 1; Max, 12; IMDx, 1; Illumigene, 4). Positivity rates of the four NAATs were associated with those of semi-quantitative cultures, which were maximized in grade 3 (>100 colony-forming unit [CFU]) compared with grade 1 (<10 CFU). Conclusions Commercial NAATs may be rapid and reliable methods for direct detection of tcdA and/or tcdB in stool specimens compared with TC. Some differences in the sensitivity of the NAATs may partly depend on the number of toxigenic C. difficile in stool specimens.