Clinical characteristics and corresponding disease syndromes
A total of 191 non-repetitive S. aureus isolates were collected from Beijing Children’s Hospital, including 120 MRSA (62.8%) and 71 MSSA (37.2%). Seventy-nine (41.4%) strains were CA, 40 (20.9%) were HA, 46 (24.1%) isolated from asymptomatic carriers, and 26 (13.6%) from outpatients. Table 1 presented patient’s age, gender, infection characteristics, clinical sources, and the agr genotyping of the 191 S. aureus isolates, as well as the clinical characteristics (hospital length of stay, PICU transfer, PICU length of stay, antibiotics usage) of the 119 inpatients. The median age was 12.0 months (ranged from 2.0 to 72.0 months; 30 were ≤ 28 days old) and 102 (53.4%) were male; the hospital length of stay ranged from 7 to 18 days, with a median of 11 days; twenty-four (12.6%) were transferred to PICU, with a median of 18.5 days (6.5-28.0); all of them received intravenous antibiotics while in the hospital, with 54 (45.4%) received vancomycin (24, 20.2%), linezolid (29, 24.4%), meropenem (18, 15.1%) and ertapenem (4, 3.4%) either alone or as part of combination treatment. In addition to systemic antibiotics, mupirocin ointment (8, 6.7%), levofloxacin ear drops (1, 0.8%) and eye drops (1, 0.8%) were prescribed. To rule out selection bias among available isolates, we analyzed our data and found no significant differences by age (p = 0.915), gender (p = 0.442), hospital length of stay (p = 0.514), PICU length of stay (p = 0.502), or vancomycin (p = 0.288) and linezolid usage (p = 0.901) among different agr groups in patients whose isolates were available for laboratory testing. While the PICU transfer rate of agr group III was significantly higher than that of other groups (p = 0.009). There was no difference between strains from diverse clinical sources among four agr groups (p = 0.495) (Fig. 1a). Whereas the number of MRSA isolates belonged to agr group I was significantly higher than that of MSSA isolates (p < 0.001) (Fig. 1b).
Upon discharge, there were a total of 161 clinical diagnoses for these 119 inpatients (Table 2), including 66 (28.3%) cases of respiratory tract infections, 38 (16.3%) cases of skin and soft tissue infections (SSTIs), 52 (22.3%) cases of invasive infections, and 5 (2.1%) cases of exfoliative toxin-mediated diseases. The distribution of different diagnoses according to agr genotypes was shown in Table 2. The frequency of strains isolated from cellulitis patients harbored agr group III was significantly higher than that of other agr groups (p = 0.046). There was only one strain isolated from SSSS patient, which was identified as agr type IV (p = 0.021). Besides, neonatal diseases (including neonatal pneumonia, omphalitis, and bullous impetigo) were all community acquired and 95% of which belonged to agr group I (except the strain which isolated from a cellulitis patient was identified as agr group III). Among medical device-related infections, 50% of isolates were agr group II, and necrotizing pneumonia isolates were more likely to be agr group III. However, there was no statistical difference between agr groups according to these diagnoses (p > 0.05).
agr genotyping
By multiplex PCR, the agr types were successfully identified in all of the isolates. As shown in Table 1, the agr type I was the most prevalent (162, 84.8%), followed by agr type II, which was found in 16 (8.4%) isolates, whereas agr types III and IV possessed by only 9 (4.7%) and 4 (2.1%) isolates, respectively.
Molecular characteristics
The genotypic diversity was high, 191 S. aureus isolates were divided into 33 STs, ST59 (76, 39.8%) was the predominant type, followed by ST22 (24, 12.6%), ST398 (15, 7.9%), and ST25 (10, 5.2%). The frequencies of the remaining STs were ranging from 0.5% to 3.7%. The 33 STs belonged to 20 CCs, CC59 (81, 42.4%), CC22 (29, 15.2%), CC398 (15, 7.9%), CC5 (14, 7.3%), and CC25 (10, 5.2%) were the most prevalent CCs.
A total of 59 distinct spa strain types were seen. The most common type was t437 (72, 37.7%), followed by t309 (20, 10.5%), t571 (10, 5.2%), and t078 (7, 3.7%). The prevalence rates of the remaining spa types were ranging from 0.5% to 2.1%. Besides, three strains could not be determined by spa typing.
Of the 120 MRSA strains, all of the five different SCCmec types were detected. Most of which carried SCCmec type IV (77, 65.0%), followed by SCCmec type V (25, 20.8%), SCCmec type I (14, 11.7%), SCCmec type III (2, 1.7%), and SCCmec type II (1, 0.8%). Also, 0.8% (1) of the isolates could not be typed by multiplex PCR.
The genotypic characteristics of 191 S. aureus stratified by agr types were shown in Fig. 2 and Additional file 1. Except that 3 CCs (CC1, CC5, and CC45) which mostly belonged to one specific agr group partly superimposed on another agr group, all of the CC59, CC25, CC22, CC951, CC8 and CC398 strains had agr group I, CC15 isolates harbored agr group II, CC30 possessed agr group III, and CC121 harbored agr group IV. Furthermore, the distribution of prevalent spa types across the patients reflected the CC distribution. Among strains that belonged to agr group I, the most common types were CC59-t437, CC25-t078, CC22-t309, CC951-t437, and CC398-t571. While CC5 and CC15 which mostly belonged to agr group II had greater spa type diversity compared to other prominent CCs. In agr groups III and IV, the two dominant types were CC1-t114 and CC121-t2092, respectively.
Prevalence and Distribution of Virulence Genes
As illustrated in Table 3, all isolates harbored the hla (191, 100%), hlb (191, 100%) and hld (191, 100%) genes. The prevalence of the lukS/F-PV gene was 31.4% (60), while the frequencies of tst, eta and etb genes were 7.9% (15), 3.7% (7) and 1.0% (2), respectively.
All isolates possessed the icaA (191, 100%) and icaD (191, 100%) genes, the frequency of the icaC gene was also high at 93.7% (179). Notably, all of the ST25 (10) strains were icaC negative. In contrast, only 1 (0.5%) isolate harbored the bap gene, which isolated from the bronchial lavage of a cystic fibrosis patient.
All isolates harbored the eno (100%), clfA (100%) and clfB (100%) genes, followed by high frequency of fib gene (143, 74.9%). The prevalence of bbp, ebpS, cna and fnbB genes were 3.1% (6), 19.9% (38), 11.5% (22) and 16.8% (32), respectively.
Of these 19 virulence genes, the distribution of the following 6 genes differed among the isolates according to the agr genotyping: tst (p = 0.006), eta (p = 0.003), etb (p = 0.001), lukS/F-PV (p = 0.004), bbp (p < 0.001), and ebpS (p < 0.001) genes (Table 3). The tst gene was more prevalent among agr group III compared to other groups. Among 7 strains that harbored the eta gene, 2 of which also had etb gene and belonged to agr group IV. It is noted that nearly all of the strains that harbored the lukS/F-PV gene belonged to agr group I (except 1 strain belonged to agr group III, which isolated from a necrotizing pneumonia patient). Additionally, the frequencies of bbp and ebpS genes that belonged to agr group I were statistically lower than that of other groups.
The average number of virulence genes was examined based on agr genotyping, which were 10.5, 11.1, 11.7 and 13.3 for agr groups I, II, III, and IV, respectively (Fig. 3). The coexistence of the studied virulence genes was also investigated. Most of strains had 10 (81, 42.4%) and 11 (82, 42.9%) genes investigated. The number of strains had 12, 13 and 14 genes were 14 (7.3%), 3 (1.6%) and 3 (1.6%), respectively. Besides, there were 1 (0.5%) and 7 (3.7%) strains had only 8 and 9 genes investigated (Fig. 4).