Microbiological Characteristics And Clinical Features Of Serotype K54 Klebsiella Pneumoniae

Objective:To investigate the epidemiological status and microbiological characteristics of K54 capsular serotype Klebsiella pneumoniae(K54-Kp),as well as the clinical infection characteristics and provide evidence for clinical prevention and rational drug use;at the same time,investigate the molecular epidemiological and Virulence characteristics.Method:1.Collect non-repetitive Klebsiella pneumoniae isolated from various clinical specimens in our hospital from January 2016 to December 2019.The K54-Kp isolates were screened by polymerase chain reaction(PCR),and then high viscosity was confirmed to use a string test.The quantity of capsular polysaccharide in the strain was measured via m-hydroxyphenyl colorimetry,and drug sensitivity was assessed,so as to completely comprehend the microbiological characteristics of K54-Kp.2.The clinical data of K54-Kp infected patients and non-K54-Kp patients were collected according to the ratio of 1:1 to grasp their clinical infection characteristics.3.The molecular epidemiological characteristics and distribution of virulence and drug resistance genes of K54-Kp were analyzed by pulsed field gel electrophoresis(PFGE),MLST typing and detection of virulence and drug resistance genes.4.The virulence phenotype of K54-Kp strain was reflected by crystal violet biomembrane formation test,serum resistance test and larva infection test.Results:1.From January 2016 to December 2019,328 strains of Klebsiella pneumoniae were collected in our hospital,and 30 strains of K54-Kp were screened out,with an overall detection rate of 9.15%(30/328)and an average annual detection rate of 2.29%,of which the detection rate in 2016 was 5.8%(4/68).36.67%(11/30)of the strains were positive in the drawing test,and the content of capsular polysaccharide was between NTUH-K2044 and ATCC700603 of Klebsiella pneumoniae with high virulence(P<0.05).The drug sensitivity showed a high resistance rate(> 65%)to cephalosporins,quinolones,aminoglycosides,carbapenems,and β-lactamases antibiotics.The resistance rate showed an increasing trend year by year,with the most extensive range in 2018-2019.All strains were sensitive to tigecycline.2.The average age of patients infected with K54-Kp is 60.40 13.14 years old,and the ratio of male to female is close to 3:1(76.7%:23.7%).The departments are mainly from neurosurgery(7/30,23.33%)and ICU(6/30,20%),followed by gastroenterology(5/5).The main source type of specimens was sputum(14,46.7%),followed by blood(12,40%).80%(24/30)patients had a nosocomial infection,80%had serious basic diseases and related complications and a history of invasive operation,53.33% were complicated with infection by other pathogens,all patients had a history of exposure to more than two antibiotics,and 53.33% patients had a poor prognosis.β-lactamases and carbapenem inhibitors were the most common treatments.Compared with non-K54-Kp patients,K54-Kp patients have no significant differences in sex,age,inflammatory index,invasive operation,antibiotic use,and prognosis.Still,there are statistical differences in the factors of hepatobiliary diseases in basic diseases(P=0.005).3.K54-KP has a total of 10 sequence typing(MLST),mainly ST29(11/30),and PFGE shows homology > 85%,distributed primarily in the B2 subgroup,mainly in neurosurgery.Virulence genes showed that the detection rates of Aerobactin,Sils,ALLS,WCAG,WABG,and MRKD were over 80%,among which WABG and MRKD were detected in all strains.Compared with the non-ST29-K54 strain,the tellurite resistance gene ter W was statistically different in the ST29-K54 strain(P<0.05).The detection rates of drug-resistant genes Kp C,NDM,VIM,SHV,CTX-M,TEM,and OXA-48 were 63.33%,30%,26.67%,100%,80%,60%,and 0 respectively.Eight strains carried both virulence gene rmp A2 and drug resistance gene Kp C,and eight strains jointly harbored Kp C and NDM.4.In the virulence phenotype experiment,the results of the biofilm formation experiment in vitro showed that the K54-Kp strain showed strong biofilm formation ability as a whole,and 27 strains(90%)could produce biofilm,among which three strains were strong biofilm formation strains,all of which were from blood.There was a statistical difference in biofilm formation strength between blood-infected strains and non-blood-infected strains(P<0.05),but in ST29 and non-ST29 K54-Kp strains.Most strains(17,58.6%)were sensitive to serum,and the killing ability of serum was weaker than that of the highly virulent strain NTUH-K2044.The virulence of the Galleria mellonella model was different.Six strains in the flora had the same virulence as NTUH-K2044,and four strains had lower virulence than ATCC700603.Most of the larvae of ST29 survived within 72 h,and the overall virulence was lower than that of ST11 and NTUH-K2044 in Cercospora mellifera model.Further comparative analysis showed that ST29 was superior to ST11 in carrying rmp A2 and wca G virulence genes(P<0.05).Conclusion:1.Although K54-Kp detection rate in our hospital was lower than K1/K2 detection rates,they were rising steadily.K54-Kp exhibited multidrug resistance,and the rate of drug resistance grew yearly.2.Although patients with basic hepatobiliary diseases were prone to develop a K54-Kp infection at our hospital,the distribution of wards were not consistent with them,mainly distributed in neurosurgery and ICU wards.3.Thetyping of K54-Kp MLST type was still mainly ST29 in our study,exhibiting the characteristics of clonal transmission.Ter W,a virulence gene of K54-Kp,had a close relationship to ST29,and the main drug resistance mechanisms included bla SHV,bla CTX-M,and bla Kp C.4.K54-Kp infected by blood flow in our hospital possessed a strong ability to form biofilm in vitro,but the overall strain’s serum killing ability and the lethality of the larvae infected by Galleria mellonellawere low,especially the ST29 K54-Kp had a lower virulence phenotype.