Original Paper Received: May 13, 2013 Accepted after revision: October 10, 2013 Published online: December 18, 2013
Neonatology 2014;105:142–148 DOI: 10.1159/000356733
Prevalence and Molecular Characterization of Staphylococcus aureus Colonization among Neonatal Intensive Care Units in Taiwan Chen-Yen Kuo a–d Yi-Chuan Huang a, e Daniel Tsung-Ning Huang a, f Hsin Chi a, f Chun-Yi Lu a, g Luan-Yin Chang a, g Chia-Yu Chi a, h Yu-Huai Ho a, i Yhu-Chering Huang a, b, d Ching-Chuan Liu a, j Li-Min Huang a, g
a Taiwan Pediatric Infectious Diseases Alliance, b Department of Pediatrics, Chang Gung Memorial Hospital at Linkou, and c Graduate Institute of Clinical Medical Sciences, d College of Medicine, Chang Gung University, Taoyuan, e Division of Infectious Diseases, Department of Pediatrics, Kaohsiung Chang Gung Memorial Hospital, Kaohsiung, f Department of Pediatrics, Mackay Memorial Hospital, and g Department of Pediatrics, National Taiwan University Hospital, College of Medicine, National Taiwan University, Taipei, h National Institute of Infectious Diseases and Vaccinology, National Health Research Institutes, Zhunan, i Department of Pediatrics, Buddhist Tzu Chi General Hospital, Hualien, and j Department of Pediatrics, National Cheng-Kung University Hospital, Tainan, Taiwan
Abstract Background: Staphylococcus aureus, particularly methicillinresistant (MRSA), is an important pathogen in neonatal intensive care units (NICUs). Carriage of S. aureus is a significant risk factor for subsequent infection. Objectives: To determine the current status of MRSA prevalence among NICUhospitalized infants in Taiwan, we conducted this pilot island-wide survey. Methods: On two designated dates in 2011, each patient who stayed in the NICUs of 7 participating hospitals was included. Nasal and umbilical swabs were obtained and sent for detection of S. aureus. The prevalence and risk factors for MRSA carriage were analyzed. MRSA strains were tested for antimicrobial susceptibility and un-
© 2013 S. Karger AG, Basel 1661–7800/13/1052–0142$38.00/0 E-Mail [email protected] www.karger.com/neo
derwent molecular characterization. Results: A total of 251 subjects were included. The overall prevalence of S. aureus and MRSA carriage was 13 and 4.4%, respectively. Previous skin and soft tissue infection was the only predictor in multivariate analysis (OR 40.36; 95% CI 2.32–702.64; p = 0.011). Among 11 MRSA isolates, 3 pulsotypes were identified, with one major type (73%). Nine isolates carried a type IV staphylococcal chromosomal cassette, and 2 carried the type VT. All but one MRSA isolate belonged to linage sequence type 59, the community clone in Taiwan. Conclusions: On a designated date, 4.4% of the infants staying in NICUs in Taiwan carried almost genetically identical community strains of MRSA. MRSA colonization in these infants was significantly associated with previous skin and soft tissue infection. © 2013 S. Karger AG, Basel
C.-Y.K. and Y.-C.H. contributed equally to this work.
Dr. Yhu-Chering Huang Department of Pediatrics, Chang Gung Memorial Hospital No. 5, Fu-Hsin St Kweishan, Taoyuan 333 (Taiwan) E-Mail ychuang @ cgmh.org.tw
Downloaded by: Thammasat University 203.131.222.2 - 10/9/2014 10:33:18 PM
Key Words Staphylococcus aureus · Colonization · Neonatal intensive care unit · Taiwan
Methicillin-resistant Staphylococcus aureus (MRSA) is an important pathogen in neonatal intensive care units (NICUs) [1, 2]. Outbreaks of MRSA in NICUs have been not infrequently reported in the past decade [3, 4]. Colonizing S. aureus may serve as an endogenous reservoir for subsequent overt clinical infections or may spread to other patients [5–7]. MRSA poses a significantly greater risk for the development of subsequent infections than does methicillin-sensitive S. aureus (MSSA) [7]. Identification of MRSA colonization is important for the prevention of MRSA transmission in NICUs [4, 8, 9]. In Taiwan, S. aureus, particularly MRSA, is among the most common nosocomial pathogens and accounts for more than 10% of late-onset bloodstream infections in NICUs in most hospitals [3, 10, 11]. A previous study conducted in Northern Taiwan revealed that, via weekly surveillance cultures, MRSA colonization was detected in more than 40% of infants during their NICU stay in 2003–2004 [4]. However, the incidence rate of health care-associated MRSA infection in NICUs has declined gradually since the mid-2000s [11, 12]. To determine the current status of MRSA prevalence among NICU-hospitalized infants in Taiwan, we conducted this pilot islandwide survey among 7 medical centers. In addition, the risk factors of MRSA colonization among these infants and the molecular characteristics of these MRSA isolates were analyzed. Materials and Methods
samples were inoculated via the streak plate method onto Trypticase soy agar with 5% sheep blood plates. The plates were incubated at 37 ° C overnight. Identification of S. aureus was performed via morphologic evaluation, Gram staining, and coagulase tests of strains grown on agar plates. To identify MRSA, cefoxitin disks were used with the disk diffusion method according to the recommendation of the Clinical and Laboratory Standard Institutes.
Molecular Characterization of MRSA Isolates The molecular methods in this study included pulsed-field gel electrophoresis (PFGE) with SmaI digestion, staphylococcal chromosomal cassette mec (SCCmec) typing, and multilocus sequence typing (MLST). The presence of Panton-Valentine leukocidin (PVL) genes was also determined. All of the procedures have been described previously [13–18]. The genotypes of PFGE criteria were designated, as in our previous studies [13–17], in alphabetical order; any new genotype, if identified, was designated consecutively. PFGE patterns with fewer than 4 band differences from an existing genotype were defined as subtypes of that genotype and labeled with Arabic number suffixes. Two isolates were considered to be indistinguishable, highly related, or distinct if they had the same subtype, the same genotype, or a different type, respectively. Antimicrobial Susceptibility Testing Susceptibility testing of MRSA isolates to antibiotics was performed via the disk diffusion method. The following antibiotics were tested: penicillin, vancomycin, daptomycin, ciprofloxacin, linezolid, fusidic acid, clindamycin, teicoplanin, erythromycin, and sulfamethoxazole-trimethoprim. Demographics and Anthropometric Variables of the Subjects To identify potential risk factors for MRSA colonization, the following data were collected for each infant: demographics, date of hospitalization, date of ICU hospitalization, underlying diseases, current diseases, antibiotic use during hospitalization, previous MRSA infection, previous skin and soft tissue infection, and catheterization. We compared the characteristics of the infants with and without MRSA colonization.
Study Design This study was approved by the institutional review board of each participating hospital. There are about 250 beds in level-III NICUs in Taiwan. Infants who stayed at the NICUs of 7 medical centers in Taiwan on two designated dates, i.e. October 11 (first survey), 2011, and December 12 (second survey), 2011, were eligible for this study. All of the NICUs participating in this study are level III and provide about 150 beds, with a bed occupancy rate of around 80%. Thus, the number of neonates enrolled into this study was representative of 60% of infants in all NICUs in Taiwan on a single day. The 7 medical centers included: Hualien Tzu-Chil General Hospital (hospital A) in Eastern Taiwan; Mackay Memorial Hospital Taipei (hospital B) and the Tamsui Branch (hospital C), National Taiwan University Hospital (hospital D), and Chang Gung Memorial Hospital at Linkou (hospital E) in Northern Taiwan, and Chang Gung Memorial Hospital at Kaohsiung (hospital F) and National Cheng-Kung University Hospital (hospital G) in Southern Taiwan. Two specimens from the nares and umbilicus, respectively, were obtained from each infant and sent to the central laboratory at Chang Gung Memorial Hospital at Linkou for detection of S. aureus and further molecular characterization [4]. Swab
Statistics Statistical analyses were performed using SPSS software (SPSS, Inc. Chicago, Ill., USA). The χ2 test and Fisher’s exact test were used to compare dichotomous variables. The Mann-Whitney test was used to compare two nonparametric continuous variables. Potential factors found to be significantly associated with MRSA colonization in univariate analyses were further analyzed with a multivariate logistic regression model. Two-tailed p values
Neonatology 2014;105:142–148 DOI: 10.1159/000356733
Prevalence and Molecular Characterization of Staphylococcus aureus Colonization among Neonatal Intensive Care Units in Taiwan Chen-Yen Kuo a–d Yi-Chuan Huang a, e Daniel Tsung-Ning Huang a, f Hsin Chi a, f Chun-Yi Lu a, g Luan-Yin Chang a, g Chia-Yu Chi a, h Yu-Huai Ho a, i Yhu-Chering Huang a, b, d Ching-Chuan Liu a, j Li-Min Huang a, g
a Taiwan Pediatric Infectious Diseases Alliance, b Department of Pediatrics, Chang Gung Memorial Hospital at Linkou, and c Graduate Institute of Clinical Medical Sciences, d College of Medicine, Chang Gung University, Taoyuan, e Division of Infectious Diseases, Department of Pediatrics, Kaohsiung Chang Gung Memorial Hospital, Kaohsiung, f Department of Pediatrics, Mackay Memorial Hospital, and g Department of Pediatrics, National Taiwan University Hospital, College of Medicine, National Taiwan University, Taipei, h National Institute of Infectious Diseases and Vaccinology, National Health Research Institutes, Zhunan, i Department of Pediatrics, Buddhist Tzu Chi General Hospital, Hualien, and j Department of Pediatrics, National Cheng-Kung University Hospital, Tainan, Taiwan
Abstract Background: Staphylococcus aureus, particularly methicillinresistant (MRSA), is an important pathogen in neonatal intensive care units (NICUs). Carriage of S. aureus is a significant risk factor for subsequent infection. Objectives: To determine the current status of MRSA prevalence among NICUhospitalized infants in Taiwan, we conducted this pilot island-wide survey. Methods: On two designated dates in 2011, each patient who stayed in the NICUs of 7 participating hospitals was included. Nasal and umbilical swabs were obtained and sent for detection of S. aureus. The prevalence and risk factors for MRSA carriage were analyzed. MRSA strains were tested for antimicrobial susceptibility and un-
© 2013 S. Karger AG, Basel 1661–7800/13/1052–0142$38.00/0 E-Mail [email protected] www.karger.com/neo
derwent molecular characterization. Results: A total of 251 subjects were included. The overall prevalence of S. aureus and MRSA carriage was 13 and 4.4%, respectively. Previous skin and soft tissue infection was the only predictor in multivariate analysis (OR 40.36; 95% CI 2.32–702.64; p = 0.011). Among 11 MRSA isolates, 3 pulsotypes were identified, with one major type (73%). Nine isolates carried a type IV staphylococcal chromosomal cassette, and 2 carried the type VT. All but one MRSA isolate belonged to linage sequence type 59, the community clone in Taiwan. Conclusions: On a designated date, 4.4% of the infants staying in NICUs in Taiwan carried almost genetically identical community strains of MRSA. MRSA colonization in these infants was significantly associated with previous skin and soft tissue infection. © 2013 S. Karger AG, Basel
C.-Y.K. and Y.-C.H. contributed equally to this work.
Dr. Yhu-Chering Huang Department of Pediatrics, Chang Gung Memorial Hospital No. 5, Fu-Hsin St Kweishan, Taoyuan 333 (Taiwan) E-Mail ychuang @ cgmh.org.tw
Downloaded by: Thammasat University 203.131.222.2 - 10/9/2014 10:33:18 PM
Key Words Staphylococcus aureus · Colonization · Neonatal intensive care unit · Taiwan
Methicillin-resistant Staphylococcus aureus (MRSA) is an important pathogen in neonatal intensive care units (NICUs) [1, 2]. Outbreaks of MRSA in NICUs have been not infrequently reported in the past decade [3, 4]. Colonizing S. aureus may serve as an endogenous reservoir for subsequent overt clinical infections or may spread to other patients [5–7]. MRSA poses a significantly greater risk for the development of subsequent infections than does methicillin-sensitive S. aureus (MSSA) [7]. Identification of MRSA colonization is important for the prevention of MRSA transmission in NICUs [4, 8, 9]. In Taiwan, S. aureus, particularly MRSA, is among the most common nosocomial pathogens and accounts for more than 10% of late-onset bloodstream infections in NICUs in most hospitals [3, 10, 11]. A previous study conducted in Northern Taiwan revealed that, via weekly surveillance cultures, MRSA colonization was detected in more than 40% of infants during their NICU stay in 2003–2004 [4]. However, the incidence rate of health care-associated MRSA infection in NICUs has declined gradually since the mid-2000s [11, 12]. To determine the current status of MRSA prevalence among NICU-hospitalized infants in Taiwan, we conducted this pilot islandwide survey among 7 medical centers. In addition, the risk factors of MRSA colonization among these infants and the molecular characteristics of these MRSA isolates were analyzed. Materials and Methods
samples were inoculated via the streak plate method onto Trypticase soy agar with 5% sheep blood plates. The plates were incubated at 37 ° C overnight. Identification of S. aureus was performed via morphologic evaluation, Gram staining, and coagulase tests of strains grown on agar plates. To identify MRSA, cefoxitin disks were used with the disk diffusion method according to the recommendation of the Clinical and Laboratory Standard Institutes.
Molecular Characterization of MRSA Isolates The molecular methods in this study included pulsed-field gel electrophoresis (PFGE) with SmaI digestion, staphylococcal chromosomal cassette mec (SCCmec) typing, and multilocus sequence typing (MLST). The presence of Panton-Valentine leukocidin (PVL) genes was also determined. All of the procedures have been described previously [13–18]. The genotypes of PFGE criteria were designated, as in our previous studies [13–17], in alphabetical order; any new genotype, if identified, was designated consecutively. PFGE patterns with fewer than 4 band differences from an existing genotype were defined as subtypes of that genotype and labeled with Arabic number suffixes. Two isolates were considered to be indistinguishable, highly related, or distinct if they had the same subtype, the same genotype, or a different type, respectively. Antimicrobial Susceptibility Testing Susceptibility testing of MRSA isolates to antibiotics was performed via the disk diffusion method. The following antibiotics were tested: penicillin, vancomycin, daptomycin, ciprofloxacin, linezolid, fusidic acid, clindamycin, teicoplanin, erythromycin, and sulfamethoxazole-trimethoprim. Demographics and Anthropometric Variables of the Subjects To identify potential risk factors for MRSA colonization, the following data were collected for each infant: demographics, date of hospitalization, date of ICU hospitalization, underlying diseases, current diseases, antibiotic use during hospitalization, previous MRSA infection, previous skin and soft tissue infection, and catheterization. We compared the characteristics of the infants with and without MRSA colonization.
Study Design This study was approved by the institutional review board of each participating hospital. There are about 250 beds in level-III NICUs in Taiwan. Infants who stayed at the NICUs of 7 medical centers in Taiwan on two designated dates, i.e. October 11 (first survey), 2011, and December 12 (second survey), 2011, were eligible for this study. All of the NICUs participating in this study are level III and provide about 150 beds, with a bed occupancy rate of around 80%. Thus, the number of neonates enrolled into this study was representative of 60% of infants in all NICUs in Taiwan on a single day. The 7 medical centers included: Hualien Tzu-Chil General Hospital (hospital A) in Eastern Taiwan; Mackay Memorial Hospital Taipei (hospital B) and the Tamsui Branch (hospital C), National Taiwan University Hospital (hospital D), and Chang Gung Memorial Hospital at Linkou (hospital E) in Northern Taiwan, and Chang Gung Memorial Hospital at Kaohsiung (hospital F) and National Cheng-Kung University Hospital (hospital G) in Southern Taiwan. Two specimens from the nares and umbilicus, respectively, were obtained from each infant and sent to the central laboratory at Chang Gung Memorial Hospital at Linkou for detection of S. aureus and further molecular characterization [4]. Swab
Statistics Statistical analyses were performed using SPSS software (SPSS, Inc. Chicago, Ill., USA). The χ2 test and Fisher’s exact test were used to compare dichotomous variables. The Mann-Whitney test was used to compare two nonparametric continuous variables. Potential factors found to be significantly associated with MRSA colonization in univariate analyses were further analyzed with a multivariate logistic regression model. Two-tailed p values
