Successful control of an extended-spectrum beta-lactamase-producing Klebsiella pneumoniae ST307 outbreak in a neonatal intensive care unit.

BACKGROUND: In this study, we evaluated the genetic relatedness of extended-spectrum beta-lactamase-producing Klebsiella pneumoniae (ESBL-KPN) isolates from an outbreak in a neonatal intensive care unit (NICU) in August 2017, We implemented an active countermeasure to control this outbreak successfully. METHODS: The incidence of healthcare-associated ESBL-KPN bacteremia was evaluated before and after initiating enhanced infection control (IC) practices in January 2018. Surveillance cultures were set up and monitored for neonates, medical personnel, and NICU environments. Molecular analyses, including pulse-field gel electrophoresis (PFGE), sequence typing, and ESBL genotyping, were performed for the isolated KPN strains. RESULTS: After implementing the enhanced IC procedures, the healthcare-associated bacteremia rate decreased from 6.0 to 0.0 per 1000 patient-days. Samples from neonates (n = 11/15, 73.3%), medical personnel (n = 1/41, 2.4%), and medical devices and the environments (6/181, 3.3%) tested positive for ESBL-KPN in the surveillance cultures in December 2017. Active surveillance cultures revealed that 23 of 72 neonates who were screened (31.9%) were colonized with ESBL-KPN between January and March 2018. All the isolates demonstrated closely related PFGE patterns and were identified as ST307 strain carrying the CTX-M-15 gene. CONCLUSIONS: Contaminated NICU environments and medical devices, as well as transmission by medical personnel, appeared to be the source of the outbreak of ESBL-KPN infection. We employed an enhanced IC strategy during January-March 2018 and successfully controlled the clonal outbreak of CTX-M-15-positive KPN. ST307 has emerged as an important bacteremia-causing pathogen in the NICU and should be carefully monitored.

A large outbreak of Salmonella enterica serovar Thompson infections associated with chocolate cake in Busan, Korea.

OBJECTIVES: This study aimed to reveal the epidemiologic characteristics of the outbreak of gastroenteritis caused by Salmonella enterica serovar Thompson in Busan Metropolitan City and to identify points for improvement to prevent of food-borne disease outbreak. METHODS: This was a case-control study. The control group comprised asymptomatic students in the same classes of the cases. The presence or absence of symptoms, ingestion of each food provided by school meal service, and commonly ingested foods in addition to those foods in meal service were investigated. Moreover, specimens collected from rectal swab, preserved foods, and environmental surface were tested. RESULTS: Of the 6,092 subjects, 1,111 (1,083 students, 22 school personnel, and 6 foodservice employees) were included in the case group; this corresponded to an 18.4% attack rate. Symptoms included diarrhea (n=1,051, 94.6%), abdominal pain (n=931, 83.8%), febrile sensation (n=502, 45.2%), and vomiting (n=275, 24.8%). The epidemic curves of each 10 schools were unimodal. Investigation of food intake showed a significantly high odds ratio for chocolate cake in 5 out of the 10 schools. Laboratory test detected Salmonella enterica serovar Thompson both in rectal swab specimens of 9 schools and in collected preserved chocolate cakes of 9 schools. Pulsed-field gel electrophoresis test result showed that Salmonella enterica seorvar Thompson isolated from human and foods were the same. CONCLUSIONS: The source of infection for the Salmonella enterica serovar Thompson outbreak in the 10 schools of Busan Metropolitan City is chocolate cake. Traceback investigation for origin of contaminated food in food-borne disease outbreak and safety control during food production should be more enhanced.

DISTRIBUTION OF LEGIONELLA PNEUMOPHILA SEROGROUPS ISOLATED FROM WATER SYSTEMS OF PUBLIC FACILITIES IN BUSAN, SOUTH KOREA.

Legionella pneumophila is the major causes of legionellosis worldwide. The distribution of L. pneumophila was investigated in water systems of public facilities in Busan, South Korea during 2007 and 2013-2014. L. pneumophila was isolated from 8.3% of 3,055 samples, of which the highest isolation rate (49%) was from ships and the lowest 4% from fountains. Serogroups of L. pneumophila isolated in 2007 were distributed among serogroups (sgs) 1-7 with the exception of sg 4, while those of isolates during 2013 and 2014 included also 11 sgs ( 1, 2, 3, 4, 5, 6, 7, 8, 12, 13, 15). L. pneumophila sg 1 was predominated among isolates from fountains (75%), hotels (60%), buildings (44%), hospitals (38%), and public baths (37%), whereas sg 3 and sg 7 was the most prevalent from ships (46%) and factories (40%), respectively. The predominated serogroup of L. pneumophila isolates from hot and cooling tower water was sg 1 (35% and 46%, respectively), while from cold water was sg 3 (29%). These results should be useful for epidemiological surveys to identify sources of outbreaks of legionellosis in Busan, South Korea.

Short communication: Proteomic characterization of tuberculin purified protein derivative from Mycobacterium bovis.

Bovine tuberculin purified protein derivative (bPPD) is used as an intradermal test (IT) reagent to detect bovine tuberculosis (bTB) in most countries. Identification of bPPD proteins is critical to understanding the immunological reaction of IT at the molecular level. While bPPD from the United Kingdom (UK) and Brazil (BR) have been recently defined at the proteomic level, bPPD from the Republic of Korea (KR) has not yet been analyzed. Here, bPPD KR proteome was examined for the first time. In total, 271 proteins were identified, including Mycobacterium bovis-specific proteins Mb0854c and Mb2898, and 42 known T cell antigens. On comparing with proteomes of bPPD UK and BR, 33 proteins were found to be common among all three bPPDs, of which 15 proteins were T cell antigens. M. bovis-specific antigens with T cell activity in bPPD may be novel candidates for use as alternatives to currently available bPPD in diagnostics.

Mycobacterium avium paratuberculosis in wild boars in Korea.

Mycobacterium avium subsp. paratuberculosis (MAP) causes chronic infectious enteritis in various domestic and wild mammals and is widely distributed globally. Interspecies transmission has been frequently reported. We investigated the presence of MAP from December 2010 to March 2011 in blood and feces collected from 222 hunter-killed wild boars. We collected 197 serum and 180 fecal samples and examined them by culture, PCR, and enzyme-linked immunosorbent assay. We investigated the status of MAP infection and the MAP genotypes in the wild boar population of Korea by using IS900 PCR and IS1311-restriction endonuclease analysis typing. Of the 180 fecal samples cultured, MAP colonies were recovered from two. By PCR, 18 animals were positive for MAP and one serum sample had a strong humoral response to MAP. The PCR-positive DNA samples from the colonies and the feces samples were genotyped as "cattle type" and "bison type," which are major MAP genotypes infecting domestic species in Korea. Our study provides new information on mycobacterial infection among wild boars, and suggests that a more effective program should be developed to monitor mycobacterial infections in wild animal populations in Korea.

In vitro activities of antimicrobials against Brucella abortus isolates from cattle in Korea during 1998-2006.

In vitro activities of 13 antibiotics were assessed against 85 Brucella abortus isolates from naturally infected cattle in the Republic of Korea during 1998-2006, using broth microdilution test. Tetracyclines showed the most excellent activity against B. abortus, displaying MIC values of 0.5 µg/ml or below. In particular, minocycline showed the lowest MIC50/90 values (0.125/0.125 µg/ml) in this study. Among four fluoroquinolones tested, ciprofloxacin (MIC50/90, 0.5/1 µg/ml) and norfloxacin (MIC50/90, 8/8 µg/ml) had the most and the least activities, respectively. Gentamicin (MIC50/90, 1/1 µg/ml) was more effective than streptomycin, erythromycin, rifampin, and chloramphenicol (MIC50/90, 2/2 µg/ml).

Quantitative Rose Bengal Test for diagnosis of bovine brucellosis.

The Rose Bengal Test (RBT) is the most widely used screening test for brucellosis in both humans and animals. Owing to its apparent simplicity of reading, however, interpretations of the RBT results can be affected by personal experience. This study describes a simple way to improve the accuracy and uniformity of reading the RBT reaction by counting the number of agglutinated particles using transparent OHP film with Quantity One, which was originally designed to count the bacterial colony numbers on agar plates. Using this system, the reactivities of three Rose Bengal antigens from different sources against international standard serum (1,000 units, VLA, UK) could be numerically measured: the intensity scale ranged from zero to around 1,600. This system enabled us to compare the antigenicity of Rose Bengal antigens from three different sources by using statistical analyses such as regression and mean intensity. Collectively, mathematical measuring of the reaction intensity used in this study may help interpret subtle test results by providing more reliable data and additional statistical information on the herd. In addition, the method would also be applicable to other agglutination test for other diseases.

The development of a selective medium for the Brucella abortus strains and its comparison with the currently recommended and used medium.

The Brucella spp. are fastidious and relatively slow-growing organisms. The isolation of such strains in a variety of specimens often requires the use of a selective medium to reduce or eliminate the growth of unexpected microorganisms. The modified Brucella selective (MBS) medium, which contains improved antibiotic mixtures, erythritol as the only carbon source, and neutral red as a pH indicator, showed good selectivity for the Brucella abortus strains, including the RB51 vaccine strain. Erythritol in the MBS medium was able to promote and/or recover the delayed growth of the B. abortus strains through the antibiotic mixtures. The Brucella colonies, which assumed a pinkish color at their central part, were easily differentiated from other organisms. The MBS medium also allows the isolation of the Brucella strains even in contaminated specimens and/or in specimens containing small numbers of viable organisms. Moreover, this medium can be applied to environmental samples for the isolation of the Brucella strains, and it can thus offer epidemiologic traceback sources for the dissemination or transfer of diseases. Therefore, the MBS medium can be applied as a useful tool of important control measures in the eradication programs.

A genetic comparison of Brucella abortus isolates from animals and humans by using an MLVA assay.

The MLVA assay is known to have a high ability to identify and discriminate Brucella species, so that it can be used as an epidemiological tool to discriminate Brucella isolates originating from restricted geographic sources. In this study, the genetic profiles of 38 B. abortus isolates from humans were analyzed and compared with genotypes from animal isolates in South Korea. As a result, it was found that they did not show high genetic diversity and were compacted. They were clustered together with animal isolates, showing a significant correlation to regional distributions. With its ability to prove a significant genetic correlation among B. abortus isolates from animals and humans in South Korea, the MLVA assay could be utilized as part of a program to control and eradicate brucellosis, one of the major zoonoses. This study represents the first data of genetic correlation of B. abortus isolates from humans and animals in South Korea.

Application and evaluation of the MLVA typing assay for the Brucella abortus strains isolated in Korea.

BACKGROUND: A Brucella eradication program has been executed in Korea. To effectively prevent and control brucellosis, a molecular method for genetic identification and epidemiological trace-back must be established. As part of that, the MLVA typing assay was evaluated and applied to B. abortus isolates for analyzing the characteristics of the regional distribution and relationships of foreign isolates. RESULTS: A total of 177 isolates originating from 105 cattle farms for the period 1996 to 2008 were selected as representatives for the nine provinces of South Korea. A dendrogram of strain relatedness was constructed in accordance with the number of tandem repeat units for 17 loci so that it was possible to trace back in the restricted areas. Even in a farm contaminated by one source, however, the Brucella isolates showed an increase or decrease in one TRs copy number at some loci with high DI values. Moreover, those 17 loci was confirmed in stability via in-vitro and in-vivo passage, and found to be sufficiently stable markers that can readily identify the inoculated strain even if minor changes were detected. In the parsimony analysis with foreign Brucella isolates, domestic isolates were clustered distinctively, and located near the Central and Southern American isolates. CONCLUSION: The MLVA assay has enough discrimination power in the Brucella species level and can be utilized as a tool for the epidemiological trace-back of the B. abortus isolates. But it is important to consider that Brucella isolates may be capable of undergoing minor changes at some loci in the course of infection or in accordance with the changes of the host.

Definition of purified enzyme-linked immunosorbent assay antigens from the culture filtrate protein of Mycobacterium bovis by proteomic analysis.

Enzyme-linked immunosorbent assay (ELISA) has been developed as the ancillary diagnosis of bovine tuberculosis at ante-mortem to overcome the disadvantages of intradermal skin test. In this study, the antigenic proteins were purified, applied to bTB ELISA, and identified through proteomic analysis. Culture filtrate protein of Mycobacterium bovis was fractionated by MonoQ column chromatography, and examined the antigenicity by immunoblotting. The antigenic 20 kDa protein was in-gel digested and identified the antigenome by LTQ mass spectrometer and peptide match fingerprinting, which were MPB64, MPB70, MPB83, Fas, Smc, Nrp, RpoC, Transposase, LeuA, and MtbE. The 20 kDa protein exhibited the highest antigenicity to bTB positive cattle in ELISA and would be useful for bTB serological diagnosis.