Correlation between a Change of Drug Resistance of Klebsiella pneumoniae and Defined Daily Doses of Antimicrobial Agents from 2014 to 2018

Introduction: The prevalence of Klebsiella pneumoniae has rapidly increased in recent years and the distribution differed greatly by region, We aimed to study the relationship between antibiotic resistance and K. pneumoniae, especially carbapenem-resistant Klebsiella pneumoniae (CRKP) in our tertiary hospitals from 2014 to 2018. Methodology: The antibiotic consumption data of K. pneumoniae were expressed as the defined daily dose (DDD) per 100 inpatient days (DDDs). K. pneumoniae which isolated from clinical samples in hospital between January 2014 and December 2018 were retrospectively analyzed, and the correlation between antibiotic resistance rate and antibiotic frequency was analyzed. Results: From 2014 to 2018, a total of 2295 strains of K. pneumoniae were isolated, with the detection rates of 8.2%, 9.2%, 11.9%, 13.4% and 14.0%. There were 423 strains of CRKP, with the detection rates of 7.5%, 5.8%, 17.8% 24.2% and 25.2% respectively. K. pneumoniae showed different degrees of resistance to antibiotics and showed an increasing trend year by year to carbapenems. The resistance rate of imipenem was 2.5%, 2.8%, 9.9%, 12.3%, 13.4%, and the resistance rate of meropenem was 2.0%, 3.0%, 8.8%, 12.6%, 12.7%, respectively. The resistance


Introduction
Klebsiella pneumoniae is a gram-negative bacillus, which is an essential condition pathogenic bacterium and iatrogenic infectious bacterium in Enterobacteriaceae Klebsiella. In recent years, the bacterial resistance rate has been increasing with the widespread use of antibacterial drugs and the rapid change of bacteria, especially carbapenem-resistant K. pneumoniae (CRKP), CHINET data showed that the detection rate of CRKP was 13.8% in 2007 [1], increasing to 25.0% and 26.3% in 2018 [2]. Therefore, the rational use of antibacterial drugs has become an essential means of curbing bacterial resistance. This study monitored and analysed the drug resistance of K. pneumoniae in the hospital from January 2014 to December 2018, and at the same time analysed the frequency of use of antimicrobial drugs among inpatients during this period to provide a basis for the rational use of antimicrobial drugs.

Sample Source and Antimicrobial Testing
Klebsiella pneumoniae were isolated from various patient bodily fluids/sites including sputum, urine, pus, blood, sterile body fluid, surgical wound, submitted by the clinical department of the hospital from 2014 to 2018. The samples to exclude the same patient during a hospital and the same strain of the same part were separated and identified by Vitek-2 system (French bio-Merieux company) and the K-B method following guidelines recommended by the Clinical and Laboratory Standards Institute (CLSI), 2011 (M100-S21) [3]. The quality control strain was Klebsiella pneumoniae ATCC 700603, which was purchased from the Clinical Laboratory of the Ministry of Health.

Detecting of the Carbapenem-Resistant K. pneumoniae (CRKP)
The modified Hodge test (MHT) was used to dilute the suspension of K. pneumoniae ATCC 700603 0.5 M. turbidity bacteria with sterile normal saline at 1:10 and evenly spread on the MH plate. After drying at room temperature for 3 -10 min, meropenem was pasted in the middle, and the bacteria to be tested were streaked from the edge of the paper sheet to the edge of the plate. The results were observed after 18 -24 h incubation at 35 C. Result: when the MIC of meropenem ≥ 2 ug/ml, it was a carbapenem-resistant strain.

Data Collection and Antimicrobial Consumption
The antibacterial drug data was derived from the hospital HIS systems from January 2014 to December 2018. The antibacterial drug distribution information of the inpatient pharmacy and the relevant information in Excel. The total consumption is calculated and using the defined daily dose (DDD) classification recommended by the World Health Organization (WHO) (http://www.whocc.no/atc_ddd_index/). Inpatient days were calculated by multiplying the quarterly total number of hospital discharges with the mean number of days of hospitalization [4]. Antimicrobial agents tracked include aminoglycosides, β-lactam antibiotics (penicillins, cephalosporins, carbapenems), quinolones, sulfa antibacterial drugs.

Statistical Analysis
The trend over time for antimicrobial consumption and rates of KP was calculated independently by Pearson's correlation analysis. The Pearson's correlation coefficient (r) between the bacterial resistance rate and the DDDs value of antibacterial drugs was calculated: r > 0.8, which was considered to have a strong correlation between them; r < 0.3, the relationship between them is considered to be weak; P < 0.05 is considered statistically significant. All statistical analyses were performed using IBM SPSS Statistics 24.0.

Detection Rate of Klebsiella pneumoniae
From 2014 to 2018, a total of 2295 strains of K. pneumoniae were detected, and the detection rate increased from 8.3% in 2014 to 14.1% in 2018, showing an increasing trend year by year. Among them, 423 CRKP strains were detected, and the isolation rate was from 7.3% in 2014, 5.8% in 2015 to 25.2% in 2018. The specific data are shown in Table 1.

The Distribution of Species
A total of 2295 K. pneumoniae were isolated from sputum, urine and pus, and their composition ratios were 67.2%, 12.3% and 7.8%, respectively. But CRKP was mainly derived from sputum, blood and urine, with a composition ratio of 50.4%, 13.9% and 12.8%, respectively. The specific data are shown in Table 2.

Antibacterial Drugs Resistance Rate of K. pneumoniae and Its
Correlation with DDDs K. pneumoniae have different degrees of resistance to antibacterial drugs, and the resistance to carbapenems has been a significant increasing trend. From 2014 to 2018, The resistance rates of imipenem are 2.53%, 2.80%, 9.90%, 12.30% and 13.41%, the drug resistance rates of meropenem were 2.04%, 3.00%, 8.80%, 12.60%, and 12.72%, respectively. The drugs with the highest DDDs value were cefoperazone/sulbactam and showed an upward trend each year. The smaller values were ciprofloxacin and imipenem. The correlation between the resistance rate of KP to common antibacterial drugs and the changes in the corresponding antibacterial drugs DDDs value from 2014 to 2018 is shown in Table 3.

Correlation Analysis of the Changing Trend of Antibacterial Drugs DDDs Value and Resistance Rate of K. pneumoniae
The strength of association between DDDs of cefoperazone/sulbactam, piperacillin/tazobactam, gentamicin and resistance rate of K. pneumoniae were very high (r = 0.964, 0.980, 0.967), and that the correlation coefficient is very highly significantly different from zero (P < 0.01). The values of DDDs of amikacin and ciprofloxacin were negatively correlated with the resistance rate of K. pneumoniae (r = −0.965, −0.972, P < 0.05). The DDDs of levofloxacin had a correlation with the K. pneumoniae resistance rate of 0.8 > r > 0.6, P < 0.01. The DDDs of meropenem had a strong positive correlation with K. pneumoniae resistance rate r > 0.8, but P > 0.05 showed no difference. Imipenem 0.5 < r < 0.8, P > 0.05, its correlation is not large. For other antibacterial drugs, r < 0.8, P > 0.05, the correlation is not great.

Discussion
Our study highlights the rising resistance rates of K. pneumoniae which isolated Antimicrobial resistance (AMR) is currently one of the most urgent public health issues in the world [6]. The resistance of K. pneumoniae is mediated by a series of mechanisms, which are mainly related to the widespread use of broad-spectrum antibacterial drugs. Catalytic hydrolysis to produce KPC carbapenemase or NDM-1 metalloenzyme can hydrolyze carbapenems to produce CRKP. Resistance to other antibacterial drugs is related to the combined action of biofilm formation, loss or alteration of membrane pore proteins, and efflux pumps [7] [8] [9].
From the data, it was found that 15 antibacterial drugs of K. pneumoniae were tested for in vitro susceptibility. The increase in drug resistance rate was directly related to the amount of clinical use. The DDDs of cefoperazone/sulbactam rose from 27,283.0 to 41,241.4. The resistance rate of cefoperazone/sulbactam to K. pneumoniae increased from 9.8% to 20.5%, and the DDDs of piperacillin sodium/tazobactam sodium increased from 2395.6 to 6733. 5  to piperacillin was significantly positively correlated with AUD of ceftazidime.
The drug resistance rate of imipenem was positively correlated with AUD of amoxicillin clavulanate.
In the study, the monitoring data shows that the resistance of K. pneumoniae to carbapenem antibacterial drugs is on the rise because carbapenem antibacterial drugs are the most effective antibacterial drugs for the treatment of K. pneumoniae infections [8]. Therefore, Our hospital establishes an fore-warning mechanism for different bacteria every quarter to cope with the difficult drug resistance situation and improving the level of anti-infective treatment and controlling the clinical application of antibacterial drugs refers to the Guidelines for the Clinical Application of Antibacterial Drugs to increase the rate of clinical specimen submission and detection, Choose appropriate antibacterial drugs according to the susceptibility of bacteria.
After 5-year period analysis of drug resistance rates, although K. pneumoniae has aminoglycosides (amikacin), enzyme-inhibitors (piperacillin/tazobactam, cefoperazone/sulbactam), The sensitivity of cephalosporins (cefoxitin) and carbapenems (imipenem, meropenem) has remained at a high level, and the resistance rate of the third and fourth generation cephalosporins has increased, especially CRKP is gradually increasing. Therefore, the drug resistance rate of K. pneumoniae is highly correlated with the use of various antibacterial drugs. Rotation of antibacterial drugs can reduce the drug resistance rate of bacteria.

Conclusion
Our data demonstrate a significant increase in the prevalence of K. pneumoniae and the consumption of enzyme-inhibitors in our hospital. We found that the high incidence of CRKP is associated with increased enzyme-inhibitors usage in our hospital. Future work is needed to elucidate the factors which influence the increased consumption of enzyme-inhibitors usage in our hospital.

Funding
This work was funded by the Chengde Science and Technology Planning Project (No. 201701A094).