Abstract
Background
Staphylococcus aureus (S. aureus) is an important human etiologic agent. An investigation of the characteristics of common genotypes of S. aureus relating to pathogenicity and antibiotic resistance may provide a foundation to prevent infection.
Methods
This study collected 275 S. aureus isolates from Zhengzhou city in China, including 148 isolates from patient samples and 127 isolates from ready‐to‐eat food samples. Antimicrobial susceptibility testing was performed using the broth dilution method. Molecular characteristics of antimicrobial resistance, virulence, and genotypes were identified by polymerase chain reaction (PCR).
Results
In total, 34.18% (94/275) of S. aureus isolates were MRSA. Compared with food isolates, clinical isolates had significantly higher antibiotic resistance rates, carrying resistance genes such as acc(6′)/aph(2′), aph(3′)‐III, ermA, and ermB and virulence genes such as tetM, sea, seb, pvl, and etb. MRSA‐t030‐agrI‐SCC mec III and MSSA‐t002‐agr II were the most common strain types among clinical strains, and MRSA‐t002‐agr II‐SCC mec III and MSSA‐t002‐agr II were the most common strain types among food strains. Additionally, some strains in the agr group were also spa type‐specific, suggesting that there may be phenotypic consistency.
Conclusion
Clinical isolates contained higher numbers of resistance genes and demonstrated higher antibiotic resistance, while 2 source strains exhibited high toxicity. These results indicate that bacteria with different origins may have undergone different evolutionary processes. As resistance and virulence factors in food bacteria can be transmitted to humans, food handlers should strictly follow hygienic measures during food production to ensure the safety of human consumers.
Keywords: antimicrobial resistance, genotypes, methicillin‐resistant S. aureus, resistance genes, virulence genes
1. INTRODUCTION
Staphylococcus aureus (S. aureus) is an important Gram‐positive human etiologic agent that can cause a wide range of diseases, such as pneumonia, toxic shock syndrome, and food poisoning.1 S. aureus contamination is the third leading cause of food poisoning worldwide and is always related to nonstandard health measures. Based on β‐lactam susceptibility, S. aureus is commonly divided into 2 categories: methicillin‐susceptible S. aureus (MSSA) and methicillin‐resistant S. aureus (MRSA).2 Since the emergence of resistant strains, especially MRSA, treatment of S. aureus infection is becoming more difficult, and this pathogen represents a serious threat to human health.3
The reasons underlying the difficulty in treating S. aureus infection in recent decades are partly attributable to the virulence factors produced by this pathogen. Various virulence determinants, including hemolysins, staphylococcal enterotoxins, toxic shock syndrome toxin‐1 (TSST‐1), and Panton‐Valentine leukocidin (PVL), are associated with S. aureus infections.4 In general, different combinations of virulence genes may determine disparate infection outcomes.
At present, many molecular typing methods are used for the genotypic characterization of S. aureus. The main methods include pulsed‐field gel electrophoresis (PFGE), multilocus sequence typing (MLST), staphylococcal protein A (spa) typing, and accessory gene regulator (agr) typing. PFGE has been acknowledged as the “gold standard” for molecular typing of S. aureus because of its excellent discriminatory power and ability to type most bacterial species.5 However, as it is difficult to compare results between laboratories and over time, PFGE is not appropriate for long‐term and wide‐ranging epidemiological studies.6 MLST and spa, both developed recently, are sequence‐based typing methods. MLST depends on 7 housekeeping loci and nucleotide variations among them for typing.7 Spa typing relies on the features, order, and number of repeat sequences in the x region of spa.8 Furthermore, spa typing is easier and less costly than MLST and PFGE.6 Agr typing relies on detecting polymorphisms in the amino acid sequence of the agr‐encoding auto‐inducing peptide and its corresponding receptor and always encompasses 4 major groups (agrI‐IV).9 It would be interesting to investigate different spa and agr types in S. aureus isolates with different origins. The aim of this study was to determine the prevalence of MRSA, antimicrobial resistance, and virulence‐associated genes carried in bacterial strains and to compare the diversity of these factors in S. aureus obtained from different sources. Moreover, the molecular genetics underlying the relationships between isolates from clinical and food origins were analyzed using the spa and agr typing methods.
2. MATERIALS AND METHODS
2.1. Bacteria
In this study, we collected 275 S. aureus isolates from Zhengzhou city, Henan Province, China, from July 2013 to December 2015. Of the 275 isolates, 148 isolates were from patients at 2 hospitals in Zhengzhou city and included 85 isolates from secretions, 10 from transudation, 7 from cerebrospinal fluid, 20 from blood, 11 from phlegm, and 15 from urine. The remaining 127 isolates were from ready‐to‐eat food samples, comprising 55 isolates from fresh meat, 18 from meat products, 17 from fruits and vegetables, and 37 strains from cereal products. All bacterial strains were stored at −80°C before use.
2.2. Antimicrobial susceptibility testing and DNA isolation
Antimicrobial susceptibility testing was performed using the broth dilution method according to the recommendations of the Clinical and Laboratory Standards Institute. Using the reference strain ATCC 29213 as a control, genomic DNA of S. aureus was extracted using a QIAamp DNA Mini Kit (Qiagen, Germany) according to the specifications for Gram‐positive bacteria.
2.3. Detection of resistance and virulence genes
A total of 9 resistance genes (mecA, ermA, ermB, ermC, msrA, aac(6′)/aph(2′’), aph(3′)‐III, ant(4′,4″), and tetM) were detected by polymerase chain reaction (PCR) as described below.10, 11 In addition, PCR was used to detect the presence of 9 virulence genes: hla, hlb, sea, seb, sec, pvl, eta, etb, and tsst.12, 13 mecA gene amplification was performed to identify MRSA isolates. SCCmec typing of MRSA strains was analyzed by multiplex PCR as previously described.14
2.4. spa and agr typing
The spa repeating region was amplified by PCR as described by Harmsen.15 spa types were determined using the spa server website (http://spa.ridom.de/spaserver/). agr allele types were determined through multiplex PCR as previously described.16
2.5. Statistical analyses
The statistical methods used for analysis were the Chi‐square test and Fisher's exact test. P < .05 was considered statistically significance. Statistical analyses of all data were performed using SPSS version 21.0 software.
3. RESULTS
3.1. Antibiotic resistance of Staphylococcus aureus isolates
Among the 275 S. aureus isolates, 34.18% (94/275) were MRSA; among these, 37.84% (56/148) were from patient samples and 29.92% (38/127) were from food samples. No significant difference (P > .05) was observed between the 2 sources (ie, clinical and food samples) of S. aureus isolates. However, there were different rates of resistance to various antibiotics among all isolates, including very high rates of resistance to penicillin, erythromycin, azithromycin, and clindamycin at 94.91%, 70.18%, 67.64%, and 64.00%, respectively. Resistance rates to tetracycline, trimethoprim/sulfamethoxazole, levofloxacin, gentamicin, moxifloxacin, and rifampin were 44.73%, 33.09%, 24.00%, 18.91%, 12.73%, and 7.27%, respectively. All strains were susceptible to vancomycin. Only 1 strain from a clinical source was resistant to linezolid. More details regarding antibiotic resistance are shown in Table 1. Moreover, resistance rates to erythromycin, azithromycin, rifampin, moxifloxacin, levofloxacin, gentamicin, and tetracycline in clinical MRSA isolates were significantly higher than those in clinical MSSA isolates (P < .05). No significant difference in antibiotic resistance rates was observed between MRSA and MSSA isolates from food strains (P > .05). Overall, the prevalence of resistance to penicillin, erythromycin, azithromycin, clindamycin, rifampin, moxifloxacin, gentamicin, and levofloxacin among clinical strains was higher than that among food strains (P < .05). Rates of resistance to trimethoprim/sulfamethoxazole, linezolid, vancomycin, and tetracycline showed no differences between the 2 sources (P > .05).
Table 1.
Antibiotic resistance of Staphylococcus aureus isolates, n (%)
| Antibiotic | Patients | No. of patients (n = 148) | Food | No. of food (n = 127) | P c | Total (n = 275) | ||||
|---|---|---|---|---|---|---|---|---|---|---|
| MRSA (n = 56) | MSSA (n = 92) | P a | MRSA (n = 38) | MSSA (n = 89) | P b | |||||
| Penicillin | 55 (98.21) | 90 (97.83) | 1.000 | 145 (97.97) | 36 (94.74) | 80 (89.89) | .586 | 116 (91.34) | .013 | 261 (94.91) |
| Erythromycin | 52 (92.86) | 66 (71.74) | .002 | 118 (79.73) | 25 (65.79) | 50 (56.18) | .313 | 75 (59.06) | .000 | 193 (70.18) |
| Azithromycin | 52 (92.86) | 68 (73.91) | .004 | 120 (81.08) | 16 (42.11) | 50 (56.18) | .146 | 66 (51.97) | .000 | 186 (67.64) |
| Clindamycin | 45 (80.36) | 61 (66.30) | .066 | 106 (71.62) | 25 (65.79) | 45 (50.56) | .114 | 70 (55.12) | .004 | 176 (64.00) |
| Trimethoprim/sulfamethoxazole | 22 (39.29) | 30 (32.61) | .409 | 52 (35.14) | 10 (26.32) | 29 (32.58) | .483 | 39 (30.71) | .437 | 91 (33.09) |
| Linezolid | 1 (1.79) | 0 | .378 | 1 (0.68) | 0 | 0 | ‐ | 0 | .538 | 1 (0.36) |
| Vancomycin | 0 | 0 | ‐ | 0 | 0 | 0 | ‐ | 0 | ‐ | 0 |
| Rifampin | 19 (33.93) | 1 (1.09) | .000 | 20 (13.51) | 0 | 0 | ‐ | 0 | .000 | 20 (7.27) |
| Moxifloxacin | 24 (42.86) | 11 (11.96) | .000 | 35 (23.65) | 0 | 0 | ‐ | 0 | .000 | 35 (12.73) |
| Levofloxacin | 33 (58.93) | 29 (31.52) | .001 | 62 (41.89) | 0 | 4 (4.49) | 1.000 | 4 (3.15) | .000 | 66 (24.00) |
| Gentamicin | 26 (46.43) | 18 (19.57) | .001 | 44 (29.73) | 5 (13.16) | 3 (3.37) | .093 | 8 (6.30) | .000 | 52 (18.91) |
| Tetracycline | 39 (69.64) | 30 (32.61) | .000 | 69 (46.62) | 17 (44.74) | 37 (41.57) | .741 | 54 (42.52) | .495 | 123 (44.73) |
The prevalence of antibiotic resistance among MRSA and MSSA isolates of clinical origin was compared.
The prevalence of antibiotic resistance among MRSA and MSSA isolates of food origin was compared.
The prevalence of antibiotic resistance among clinical isolates and food isolates was compared.
3.2. Distribution of resistance genes in Staphylococcus aureus isolates
The most frequent resistance gene among the 275 S. aureus isolates was ant(4′,4′’), with 73.45% (202/275) of isolates being positive, followed by acc(6′)/aph(2′’) (70.55%), ermC (64.36%), aph(3′)‐III (48.36%), ermB (37.45%), ermA (29.09%), msrA (21.82%), and tetM (18.91%). More information about the distribution of resistance genes is provided in Table 2. The ermA, msrA, and tetM carriage rates were significantly higher for MRSA than MSSA among clinical isolates (P < .05). There was no significant difference in resistance genes between MRSA and MSSA among food isolates (P > .05). Furthermore, no isolates of food origin were positive for ermA and tetM. In general, the acc(6′)/aph(2′), aph(3′)‐III, ermA, ermB, and tetM carriage rates among isolates of clinical origin were higher than among those of food origin (P < .05). Conversely, the frequency of ant(4′,4′’) in food isolates was higher than that among clinical isolates (P < .05).
Table 2.
Distribution of resistance genes, n (%)
| Gene | Patients | No. of patients (n = 148) | Food | No. of food (n = 127) | P c | Total (n = 275) | ||||
|---|---|---|---|---|---|---|---|---|---|---|
| MRSA (n = 56) | MSSA (n = 92) | P a | MRSA (n = 38) | MSSA (n = 89) | P a | |||||
| acc(6′)/aph(2′’) | 47 (83.93) | 82 (89.13) | .359 | 129 (87.16) | 20 (52.63) | 45 (50.56) | .831 | 65 (51.18) | .000 | 194 (70.55) |
| aph(3′)‐III | 34 (60.71) | 64 (69.57) | .270 | 98 (66.22) | 11 (28.95) | 24 (26.97) | .819 | 35 (27.56) | .000 | 133 (48.36) |
| ant(4′,4′’) | 40 (71.43) | 61 (66.30) | .516 | 101 (68.24) | 30 (78.95) | 71 (79.78) | .916 | 101 (79.53) | .035 | 202 (73.45) |
| ermA | 38 (67.86) | 42 (45.65) | .009 | 80 (54.05) | 0 | 0 | ‐ | 0 | .000 | 80 (29.09) |
| ermB | 27 (48.21) | 40 (43.48) | .575 | 67 (45.27) | 14 (36.84) | 22 (24.72) | .165 | 36 (28.35) | .004 | 103 (37.45) |
| ermC | 37 (66.07) | 52 (56.52) | .250 | 89 (60.14) | 28 (73.68) | 60 (67.42) | .483 | 88 (69.29) | .114 | 177 (64.36) |
| msrA | 18 (32.14) | 14 (15.22) | .015 | 32 (21.62) | 11 (28.95) | 17 (19.10) | .220 | 28 (22.05) | .932 | 60 (21.82) |
| tetM | 30 (53.57) | 22 (23.91) | .000 | 52 (35.14) | 0 | 0 | ‐ | 0 | .000 | 52 (18.91) |
The prevalence of resistance genes among MRSA and MSSA isolates of clinical origin was compared.
The prevalence of resistance genes among MRSA and MSSA isolates of food origin was compared.
The prevalence of resistance genes among clinical isolates and food isolates was compared.
3.3. Prevalence of virulence genes in Staphylococcus aureus isolates
Of the 275 isolates, the percentage of hla was highest (97.09%), followed by sea (69.82%), hlb (66.91%), seb (57.45%), pvl (52.36%), eta (45.82%), sec (38.91%), tsst (37.82%), and etb (34.55%). Most virulence genes demonstrated no difference in frequency between MRSA and MSSA isolates of clinical origin (P > .05), with the exception of the sec gene, which had a higher percentage in MSSA than in MRSA isolates (P < .05). Interestingly, only the etb percentage was higher in MRSA strains than in MSSA strains among food isolates (P < .05). The sea, seb, pvl, and etb carriage rates among isolates of clinical origin were significantly higher than those among isolates of food origin (P < .05). In contrast, the hla and hlb carriage rates were higher among food isolates than among clinical isolates (P < .05). Moreover, all food strains were positive for hla. The prevalence of virulence genes in S. aureus isolates is shown in Table 3.
Table 3.
The prevalence of virulence genes in Staphylococcus aureus isolates, n (%)
| Gene | Patients | No. of patients (n = 148) | Food | No. of food (n = 127) | P b | Total (n = 275) | ||||
|---|---|---|---|---|---|---|---|---|---|---|
| MRSA (n = 56) | MSSA (n = 92) | P b | MRSA (n = 38) | MSSA (n = 89) | P a | |||||
| sea | 54 (96.43) | 81 (88.04) | 0.148 | 135 (91.22) | 18 (47.37) | 39 (43.82) | 0.713 | 57 (44.88) | 0.000 | 192 (69.82) |
| seb | 47 (83.93) | 74 (80.43) | 0.594 | 121 (81.76) | 12 (31.58) | 25 (28.09) | 0.692 | 37 (29.13) | 0.000 | 158 (57.45) |
| sec | 16 (28.57) | 46 (50.00) | 0.010 | 62 (41.89) | 14 (36.84) | 31 (34.83) | 0.828 | 45 (35.43) | 0.273 | 107 (38.91) |
| pvl | 39 (69.64) | 59 (64.13) | 0.492 | 98 (66.22) | 17 (44.74) | 29 (32.58) | 0.192 | 46 (36.22) | 0.000 | 144 (52.36) |
| hla | 52 (92.86) | 88 (95.65) | 0.723 | 140 (94.59) | 38 (100) | 89 (100) | ‐ | 127 (100) | 0.008 | 267 (97.09) |
| hlb | 33 (58.93) | 54 (58.70) | 0.978 | 87 (58.78) | 32 (84.21) | 65 (73.03) | 0.175 | 97 (76.38) | 0.002 | 184 (66.91) |
| eta | 20 (35.71) | 45 (48.91) | 0.117 | 65 (43.92) | 16 (42.11) | 45 (50.56) | 0.382 | 61 (48.03) | 0.495 | 126 (45.82) |
| etb | 29 (51.79) | 47 (51.09) | 0.934 | 76 (51.35) | 10 (26.32) | 9 (10.11) | 0.019 | 19 (14.96) | 0.000 | 95 (34.55) |
| tsst | 27 (48.21) | 35 (38.04) | 0.224 | 62 (41.89) | 16 (42.11) | 26 (29.21) | 0.157 | 42 (33.07) | 0.133 | 104 (37.82) |
The prevalence of virulence genes among MRSA and MSSA isolates of clinical origin was compared.
The prevalence of virulence genes among MRSA and MSSA isolates of food origin was compared.
The prevalence of virulence genes among clinical isolates and food isolates was compared.
3.4. Molecular characteristics of Staphylococcus aureus genotypes
A total of 63 spa types were found among the 275 S. aureus isolates. The most prominent spa type was t030, which accounted for 15.64% (43/275) of isolates, followed by t002 (10.55%), t189 (8.36%), t091 (6.55%), t437 (6.18%), t701 (5.82%), and t127 (5.45%). The main spa types among clinical isolates were t030 (29.05%, 43/148), t437 (8.78%, 13/148), t189 (6.08%, 9/148), and t002 (5.41%, 8/148), while the main spa types among food isolates were t002 (16.54%, 21/127), t189 (11.02%, 14/127), t091 (11.02%, 14/127), and t127 (11.02%, 14/127). The newly identified spa types t15478, t15920, t15922, t15923, t16751, and t16752 were all present in MSSA strains; t15478 was present in clinical strains, while t15920, t15922, t15923, t16751, and t16752 were present in food strains. The molecular characteristics of spa types are shown in Table S1. Among the 56 MRSA strains of clinical origin, 52 strains had 3 SCCmec types (types II, III, and IVa). Of these, SCCmec type III was the most common, detected in 69.64% (39/56) of clinical MRSA isolates, followed by SCCmec type IVa in 19.64% (11/56) and SCCmec type II in 3.57% (2/56) of clinical MRSA isolates. No SCCmec type was established for the remaining 4 MRSA clinical strains. Of the 38 MRSA strains of food origin, 27 had 2 SCCmec types; SCCmec type III accounted for 60.53% (23/38) and SCCmec type IVa for 10.53% (4/38). The remaining 11 MRSA food strains had no SCCmec type. For agr typing, the proportions of the 4 groups among the 2 sources of S. aureus isolates were different. In clinical isolates, agr groups I, III, II, and IV were present in 64.19%, 12.84%, 11.49%, and 4.05% of isolates, respectively, while the agr proportions in food isolates for groups I, II, III, and IV were 56.69%, 22.05%, 17.32%, and 3.94%, respectively. However, there were 11 clinical isolates that were negative for any of the 4 agr groups. Among the strains of clinical origin, MRSA‐t030‐agrI‐SCCmecIII (19/56) and MRSA‐t437‐agrI‐SCCmecIVa (7/56) were the most common types among MRSA isolates, while MSSA‐t002‐agrII (8/92) and MSSA‐t189‐agrI (7/92) were the most common among MSSA isolates. Among strains of food origin, the most common were MRSA‐t002‐agrII‐SCCmecIII (6/38), MRSA‐t701‐agrI‐SCCmecIII (3/38), and MRSA‐t437‐agrI‐SCCmecIVa (3/38) for MRSA and MSSA‐t002‐agrII (15/89), MSSA‐t189‐agrI (12/89), and MSSA‐t127‐agrIII (12/89) for MSSA.
4. DISCUSSION
Staphylococcus aureus is one of the most common pathogenic bacteria in the clinic and in the community, and the presence or absence of resistance and virulence genes is closely related to the pathogenicity of S. aureus infection. Analysis of the features of antibiotic resistance and detecting the distribution of resistance and virulence genes in S. aureus isolates from different sources will guide rational drug use among clinicians and prevent the dissemination of infection.
In the present study, 34.18% of all S. aureus isolates (275) were MRSA. The MRSA rate was ~37.84% among clinical isolates, which was consistent with the prevalence of MRSA in China.17 All 148 clinical isolates demonstrated differing degrees of resistance to antibiotics with the exception of vancomycin, and resistance to penicillin reached 97.97%. This revealed that antibiotic resistance was widely distributed among clinical strains, and the drug resistance of clinical bacteria remains serious. Compared with MSSA, resistance rates to erythromycin, azithromycin, rifampin, moxifloxacin, levofloxacin, gentamicin, and tetracycline in MRSA were higher, and similarly, the ermA, msrA, and tetM carriage rates among MRSA were also higher than the carriage rates among MSSA. S. aureus possesses 2 main methods to obtain antibiotic resistance: The first is mutations to bacterial chromosomal genes, and the second is the acquisition of resistance genes from other organisms via plasmids or mobile genetic elements (MGEs).18, 19 The molecular basis of resistance among MRSA isolates is the MGE designated staphylococcal cassette chromosome mec (SSCmec),20 which contains the mecA gene encoding resistance to β‐lactam antibiotics via an altered penicillin‐binding protein (PBP2a).21 In addition, MRSA strains obtain a variety of resistance genes that undergo transfer under the pressure of antibiotic selection, leading to high resistance and multidrug resistance rates in MRSA isolates. Notably, 1 MRSA strain was resistant to linezolid. This number seems low, as there have been some outbreaks of healthcare infections associated with linezolid resistance, such as a linezolid‐resistant MRSA outbreak in a Spanish ICU caused by the extensive use of this antibiotic.22 Therefore, it is important to use new antibiotics appropriately despite the fact that they treat pathogens effectively. The presence of virulence genes in the 148 clinical strains was high, ranging from 41.89% to 94.59%; in particular, the carriage rate for pvl (66.22%) was much higher than that described in an earlier report (10.7%).23 Moreover, our study found that only the sec rate was higher in MSSA isolates than in MRSA isolates. This finding was in agreement with a previous study. However, the same previous study also detected higher frequencies of pvl and tsst genes in MSSA strains and higher frequencies of sea and eta genes in MRSA strains among clinical isolates,24 unlike our results, which showed no differences in the sea, seb, hla, hlb, pvl, eta, etb, and tsst rates between MRSA and MSSA strains of clinical origin. The reasons underlying these differences may be related to the different strain origins.
Staphylococcus aureus is a major food‐borne pathogen. Symptoms of S. aureus infection include nausea, vomiting, abdominal cramping, and sometimes diarrhea.25 In this study, 127 isolates were collected from food samples. Of these, 38 (29.92%) isolates were MRSA; this rate did not differ from that among clinical strains, indicating that MRSA contamination in food has become a serious problem. Interestingly, among the 127 food isolates, there were no differences in antibiotic resistance rates between MRSA and MSSA strains, unlike clinical isolates. Similarly, there were no differences in resistance gene rates between MRSA and MSSA strains, probably because exposure to agricultural and environmental sources of antibiotics initiated the development of resistance in these strains. MRSA strains did not show higher rates of resistance than MSSA strains among isolates of food origin, unlike clinical isolates, probably because hospitals utilize many antibiotics to maintain selective pressure, According to antimicrobial susceptibility testing, the penicillin resistance rate reached 91.34%, making it the least effective drug. Second to penicillin was erythromycin, demonstrating a resistance rate of 59.06%. This result differed from a report indicating that resistance to trimethoprim/sulfamethoxazole was most common in bacteria isolated from goat milk.26 The present study did not identify food‐derived isolates resistant to linezolid, vancomycin, rifampin, and moxifloxacin, but 94.49% of food isolates were resistant to at least 1 antibiotic. Furthermore, a higher rate (51.97%) of multidrug resistance (resistance to 3 or more antimicrobial categories) was identified in ready‐to‐eat food samples compared with a rate of 3.8% in Malaysia.27 The 127 food isolates were also screened for virulence genes to determine their pathogenic risk. All food strains were positive for hla, in accordance with a study conducted in Asturias, which found that all strains isolated from cow milk harbored the hla virulence gene.28 Moreover, it was reported that hla was related to an outbreak of staphylococcal food poisoning.29 Another important factor underlying staphylococcal food poisoning is the presence of enterotoxins, although the percentage of staphylococcal enterotoxin genes in isolates of food origin varies in different studies. Different levels of enterotoxin genes indicate different pathogenic potential. This current study showed that sea was present in 44.88% of food isolates, sec in 35.43%, and seb in 29.13%; these percentages were higher than those identified in chicken products from Egypt,30 indicating that the genes may significantly contribute to staphylococcal food poisoning. The transmission of bacterial virulence factors to humans through the food chain as well as the higher multidrug resistance rates in those strains represents large risks for human infection. When resistance gene carriage rates were compared between MRSA and MSSA strains of food origin, only the etb rate was higher in MRSA strains; other genes demonstrated no differences. Combined with the results obtained for clinical isolates, our study suggested that the difference in virulence gene carriage rates between MRSA and MSSA strains might be insignificant.
Although some reports of vancomycin resistance have emerged,31 the 275 strains examined in this study were all sensitive to vancomycin, suggesting that vancomycin remains an effective drug for the treatment of serious S. aureus infections in our region. When we tested resistance to twelve types of antibiotics, clinical strains had higher resistance rates to penicillin, erythromycin, azithromycin, clindamycin, rifampin, moxifloxacin, gentamicin, and levofloxacin than food strains. This revealed that clinical isolates demonstrated higher levels of drug resistance compared with food isolates. The acquisition of antibiotic resistance in clinical S. aureus isolates may be attributable to the inappropriate use of antibiotics in hospitals. In addition, drug resistance can spread by plasmid transfer between strains and cross‐contamination in hospitals, resulting in higher resistance among clinical strains. Accordingly, we found that the carriage rates of the resistance genes acc(6′)/aph(2′), aph(3′)‐III, ermA, ermB, and tetM were higher among clinical isolates than among isolates of food origin; only the rate of ant(4′,4′’) was higher in food isolates. This inconsistency between resistance phenotypes and genes may be associated with the differential replication and expression of resistance‐related plasmids in bacteria, as well as other mechanisms of drug resistance. Moreover, we found that clinical strains carried more resistance genes (≧ 5) compared with food strains (Table S2). Consistent with this, the genomes of clinical strains contained more virulence genes (≧ 5) (Table S3), and virulence gene rates were not low among either source of isolates; almost every strain contained at least 2 virulence genes, indicating that strains from both clinical and food sources had high virulence potential. These results also suggested that food contaminated with S. aureus has higher pathogenic potential, with implications for human health. Thus, food safety issues should be taken seriously. During processes associated with food production and sales, food handlers should endeavor to follow hygienic measures to ensure the safety of human consumers.
Among the 275 S. aureus isolates, a total of 63 spa types were identified. Clinical isolates contained 37 spa types, and food strains included 38 spa types. The most prevalent spa type was t030, although it was only found in clinical strains. This finding was in agreement with a previous survey showing that t030 is one of the most common spa types in Chinese hospitals.32 t030 demonstrates high resistance to antibiotics that are commonly used in clinical settings, which may be the reason for its widespread distribution in hospitals, and our results indicate that some spa types may only appear in strains with specific origins. The most common spa type among food strains was t002, followed by t127, t189, and t901. It was previously reported that t127 is associated with serious human infections in the United States and Germany,33 indicating food safety measures must be strengthened to minimize the infection rate. t189 is frequently identified in food isolates; for example, 1 study showed that the most prevalent spa type in raw and processed food commodities in Shanghai was t189.34 In our study, t189 was present in 6.08% of clinical strains and ranked third in prevalence. This result indicates the important correlation between food‐originating S. aureus and popular spa types in clinical isolates. A previous study showed that different spa types may contain different numbers of virulence genes,35 but the present study did not corroborate those findings. Of the 275 S. aureus strains obtained from 2 sources, MRSA SCCmec type III was the most common. It was previously reported that the most common MRSA type in Asian regions other than Japan and South Korea, which primarily carry the SCCmecII genotype, is SCCmec type III.36 In MRSA strains of clinical origin, the most common type was MRSA‐t030‐agrI‐SCCmecIII, which was consistent with the results of a previous study in central‐southern China.10 We also found that some strains belonging to the same spa type were present in the same agr group. For example, strains carrying spa type t127 were all present in agr group III. In addition, t189, t091, and t078 strains all belonged to agr group I. These results imply that some spa types may belong to 1 specific agr group, and there are relationships between different genotypes.
In conclusion, clinical isolates contained more resistance genes and demonstrated higher rates of antibiotic resistance than food strains, although all strains from both sources exhibited high virulence. These results indicate that bacteria of different origins may undergo different evolutionary processes. S. aureus can infect humans through food contamination, so it is necessary to follow strict hygienic measures during food production processes. To date, many researchers have reported on S. aureus isolates with different origins worldwide, but studies comparing the characteristics of clinical strains and food strains are scarce. Our results provide information with implications for human health and food safety.
Supporting information
ACKNOWLEDGMENTS
The National Natural Science Foundation of China (no. 81401700) and the Henan Province University Science and Technology Innovation Talent Projects (17HASTIT045) supported this work.
Luo K, Shao F, Kamara KN, et al. Molecular characteristics of antimicrobial resistance and virulence determinants of Staphylococcus aureus isolates derived from clinical infection and food. J Clin Lab Anal. 2018;32:e22456 10.1002/jcla.22456
KL and FS contributed equally to this work.
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