Serotype K1 Research Articles

Detection of carbapenemase-producing, hypervirulent Klebsiella spp. in wastewater and their potential transmission to river water and WWTP employees

Wastewater treatment plants (WWTPs) release drug-resistant microorganisms to water bodies (with effluents), and WWTP employees are exposed to bioaerosol emissions from the processed wastewater. Bacteria of the genus Klebsiella, in particular carbapenemase-producing (CP), hyper-virulent (Hvr) strains of Klebsiella pneumoniae, play a special role in this process. Klebsiella spp. strains isolated from wastewater, river water and the upper respiratory tract of WWTP employees were analyzed in this study. The isolated strains were identified as K. pneumoniae (K. pn) or K. non-pneumoniae (K. npn). The prevalence of nine types of genes encoding resistance to beta-lactams, nine genes encoding virulence factors and K1/K2 capsular serotypes, three genes encoding multi drug effluent pump systems, and the class 1 integron-integrase gene was determined by PCR. A total of 284 Klebsiella spp. isolates were obtained in the study: 270 environmental strains and 14 strains from the upper respiratory tract. Among environmental isolates 90.7% (245/270) harbored beta-lactam resistance genes, 17.4% (47/270) were classified as CP strains, 11.1% (30/270) were classified as Hvr strains, and 1.9% (5/270) were classified as CP-Hvr strains. CP-Hvr strains were also isolated from WWTP employees. Genes encoding β-lactamases (including carbapenemases), complete efflux pump systems and the K1 serotype were identified more frequently in K. pn strains. In turn, K. npn strains were characterized by a higher prevalence of blaSHV and intI1 genes and K2 serotype gene. The strains isolated from wastewater and river water also differed in the abundance of drug resistance and virulence genes. The results of the study indicate that CP-Hvr K. pn strains are possibly transmitted from wastewater via bioareosol to the upper respiratory tract of WWTP employees. blaGES-type carbapenemases significantly contributed to the spread of drug resistance in the environment.

High-risk clones of extended-spectrum β-lactamase-producing Klebsiella pneumoniae isolated from the University Hospital Establishment of Oran, Algeria (2011-2012).

The purpose of the study was to characterize the resistome, virulome, mobilome and Clustered Regularly Interspaced Short Palindromic Repeats-associated (CRISPR-Cas) system of extended-spectrum β-lactamase producing Klebsiella pneumoniae (ESBL-KP) clinical isolates and to determine their phylogenetic relatedness. The isolates were from Algeria, isolated at the University Hospital Establishment of Oran, between 2011 and 2012. ESBL-KP isolates (n = 193) were screened for several antibiotic resistance genes (ARGs) using qPCR followed by Pulsed-Field Gel Electrophoresis (PFGE). Representative isolates were selected from PFGE clusters and subjected to whole-genome sequencing (WGS). Genomic characterization of the WGS data by studying prophages, CRISPR-Cas systems, Multi-Locus Sequence Typing (MLST), serotype, ARGs, virulence genes, plasmid replicons, and their pMLST. Phylogenetic and comparative genomic were done using core genome MLST and SNP-Based analysis. Generally, the ESBL-KP isolates were polyclonal. The whole genome sequences of nineteen isolates were taken of main PFGE clusters. Sixteen sequence types (ST) were found including high-risk clones ST14, ST23, ST37, and ST147. Serotypes K1 (n = 1), K2 (n = 2), K3 (n = 1), K31 (n = 1), K62 (n = 1), and K151 (n = 1) are associated with hyper-virulence. CRISPR-Cas system was found in 47.4%, typed I-E and I-E*. About ARGs, from 193 ESBL-KP, the majority of strains were multidrug-resistant, the CTX-M-1 enzyme was predominant (99%) and the prevalence of plasmid-mediated quinolone resistance (PMQR) genes was high with aac(6′)-lb-cr (72.5%) and qnr’s (65.8%). From 19 sequenced isolates we identified ESBL, AmpC, and carbapenemase genes: blaCTX-M-15 (n = 19), blaOXA-48 (n = 1), blaCMY-2 (n = 2), and blaCMY-16 (n = 2), as well as non-ESBL genes: qnrB1 (n = 12), qnrS1 (n = 1) and armA (n = 2). We found IncF, IncN, IncL/M, IncA/C2, and Col replicon types, at least once per isolate. This study is the first to report qnrS in ESBL-KP in Algeria. Our analysis shows the concerning co-existence of virulence and resistance genes and would support that genomic surveillance should be a high priority in the hospital environment.

Clinical and Molecular Characteristics of Carbapenem-Resistant Hypervirulent Klebsiella pneumoniae Isolates in a Tertiary Hospital in Shanghai, China.

BackgroundThe convergence of carbapenem-resistance and hypervirulence in Klebsiella pneumoniae has led to a significant public health challenge. In recent years, there have been more and more reports on carbapenem-resistant hypervirulent Klebsiella pneumoniae (CR-hvKP) isolates.Materials and MethodsClinical data of patients infected with CR-hvKP from January 2019 to December 2020 in a tertiary hospital were retrospectively evaluated. The number of isolates of Klebsiella pneumoniae, hypermucoviscous Klebsiella pneumoniae (hmKP), carbapenem-resistant hypermucoviscous Klebsiella pneumoniae (CR-hmKP) and carbapenem-resistant hypervirulent Klebsiella pneumoniae (CR-hvKP) collected during the period of 2 years was calculated. The antimicrobial resistance gene, virulence-associated gene, capsular serotype gene and multilocus sequence typing (MLST) of CR-hvKP isolates were detected by PCR.ResultsDuring the study period, a total of 1081 isolates of non-repeat Klebsiella pneumoniae were isolated, including 392 isolates of hypermucoviscous Klebsiella pneumoniae (36.3%), 39 isolates of CR-hmKP (3.6%), and 16 isolates of CR-hvKP (1.5%). About 31.2% (5/16) of CR-hvKP were isolated from 2019, and 68.8% (11/16) of CR-hvKP were isolated from 2020. Among the 16 isolates of CR-hvKP, 13 isolates were ST11 and serotype K64, 1 isolate was ST11 and serotype K47, 1 isolate was ST23 and serotype K1, and 1 isolate was ST86 and serotype K2. The virulence-associated genes entB, fimH, rmpA2, iutA, iucA were present in all of 16 CR-hvKP isolates, followed by mrkD (n=14), rmpA (n=13), aerobactin (n=2), allS (n=1). Sixteen CR-hvKP isolates all carry carbapenemase gene blaKPC-2 and extended-spectrum β-lactamase gene blaSHV. ERIC-PCR DNA fingerprinting results showed that 16 CR-hvKP isolates were highly polymorphic, and there were significant differences in bands among the isolates, presenting a sporadic state.ConclusionAlthough CR-hvKP was sporadically distributed, it showed an increasing trend year by year. Therefore, clinical attention should be paid, and necessary measures should be taken to avoid the cloning and transmission of superbacterium CR-hvKP.

Correction for Kuzina et al., "Carriage of Capsular Serotype K1 Klebsiella pneumoniae Sequence Type 23 Strains in Healthy Microbiology Laboratory Staff in Russia".

Correction for Kuzina et al., "Carriage of Capsular Serotype K1 Klebsiella pneumoniae Sequence Type 23 Strains in Healthy Microbiology Laboratory Staff in Russia".

Virulence among different types of hypervirulent Klebsiella pneumoniae with multi-locus sequence type (MLST)-11, Serotype K1 or K2 strains

BackgroundTwo different types of hypervirulent K. pneumoniae (HvKp), the MLST-11 and serotype K1/K2 strains, have been frequently described in recent studies. Although these two types of strains were described to be HvKp, their virulence was not compared. In this study, in vitro and in vivo approaches were used to assess differences in virulence.Materials and methodsA total of twenty-nine isolates, including 6 strains of each of serotype K1 and K2 isolates and 17 strains of ST11 isolates, were selected for this study. Phenotypic tests of virulence were performed by the string test and analysis of the virulent associated genes was detected by PCR. In vitro models of serum resistance and phagocytosis were used as the parameters to assess the virulence. In-frame deletion of virulence-associated genes was performed to study their contributions to virulence. The median lethal dose, i.e., the LD50, in mice was determined following IP injection.ResultsAlthough serotype K1 and K2 strains and ST11 isolates had similar virulence gene profiles, the ST11 isolates showed less serum and phagocytic resistance than the serotype K1/K2 isolates. The mouse lethality test revealed that all ST11 isolates were unable to cause lethality, even at > 107 CFU, while serotypes K1 and K2 showed an LD50 at ≤ 103 CFU. Aerobactin or capsule knockout mutants exhibited a lower LD50 than the parental strain, while capsule mutants showed a more significant decrease in LD50.ConclusionSince there was a significant difference in virulence levels between the two types of HvKp when assessed in in vitro and in vivo models, it may be better to use the designation "HvKp" for some strains based on animal studies to avoid confusion. Virulence and non-virulence could be analysed in a relative manner, especially in comparison studies.

Carriage of Capsular Serotype K1 Klebsiella pneumoniae Sequence Type 23 Strains in Healthy Microbiology Laboratory Staff in Russia.

Klebsiella pneumoniae causes both nosocomial and community-associated infections. Among the hypervirulent K. pneumoniae (hvKP) isolates, K1 is the most common capsular serotype. Here, we report the draft genome sequences of 3 K1-type (sequence type 23) K. pneumoniae strains isolated from healthy microbiology laboratory staff in Russia.

Detection of Capsular Polysaccharide Virulence Genes rmpA and magA of Klebsiella Pneumonia Isolate from Diabetic Foot Ulcer Patient in Najaf Governorate in Iraq

Klebsiella Pneumoniae is an opportunistic pathogen, which mostly cause nosocomial infections. K.Pneumoniae utilizes a variety of virulence factors, especially capsule polysaccharide, lipopolysaccharide,fimbriae, outer membrane proteins and determinants for iron acquisition and nitrogen source utilization,for survival and immune evasion during infection. A total of (36) clinical swab specimens isolated frompatient with diabetic foot ulcer in Al-Sdder Medical City (Central Diabetic Foot) in Najaf Governorate,Iraq from November 2019 to February 2020 , for detection some virulence capsular polysaccharide genessuch as rmpA and magA that related with K1 serotype of K . pneumonia and associated with antibioticresistance pattern in patient of diabetic foot ulcer. Antibiotic susceptibility test of K. Pneumoniae isolatesagainst (20) of commonly used antibiotic showed there 25 (69%) of K. pneumonia isolate from diabeticfoot patient were Multi-drug resistance (MDR),while11(31%) were extensive-drug resistance (XDR). Theresults of amplified product demonstrated that K1 serotype of K . Pneumonia was positive in 23 (64%)of totally clinical specimen than other types of serotype ( Non-K1 serotype). (magA) gene that associatedwith hypermucoid viscosity was positive in 21 (58%) of total specimens while (rmpA) gene that regulate ofcapsular polysaccharide of K .pneumonia was positive in 22 (62%) of totally samples

Isolation and Characterization of a Novel Lytic Bacteriophage against the K2 Capsule-Expressing Hypervirulent Klebsiella pneumoniae Strain 52145, and Identification of Its Functional Depolymerase.

Klebsiella pneumoniae is among the leading bacteria that cause nosocomial infections. The capsule of this Gram-negative bacterium is a dominant virulence factor, with a prominent role in defense and biofilm formation. Bacteriophages, which are specific for one bacterial strain and its capsule type, can evoke the lysis of bacterial cells, aided by polysaccharide depolymerase enzymes. In this study, we isolated and characterized a bacteriophage against the nosocomial K. pneumoniae 52145 strain with K2 capsular serotype. The phage showed a narrow host range and stable lytic activity, even when exposed to different temperatures or detergents. Preventive effect of the phage in a nasal colonization model was investigated in vivo. Phlyogenetic analysis showed that the newly isolated Klebsiella phage B1 belongs to the Webervirus genus in Drexlerviridae family. We identified the location of the capsule depolymerase gene of the new phage, which was amplified, cloned, expressed, and purified. The efficacy of the recombinant B1dep depolymerase was tested by spotting on K. pneumoniae strains and it was confirmed that the extract lowers the thickness of the bacterium lawn as it degrades the protective capsule on bacterial cells. As K. pneumoniae strains possessing the K2 serotype have epidemiological importance, the B1 phage and its depolymerase are promising candidates for use as possible antimicrobial agents.

Prevalence of pathogenic Klebsiella pneumoniae based on PCR capsular typing harbouring carbapenemases encoding genes in Uganda tertiary hospitals

BackgroundKlebsiella pneumoniae is an opportunistic pathogen that has been implicated as one of commonest cause of hospital and community acquired infections. The K. pneumoniae infections have considerably contributed to morbidity and mortality in patients with protracted ailments. The capacity of K. pneumoniae to cause diseases depends on the presence of an array virulence factors. Coexistence and expression of virulence factors and genetic determinants of antibiotic resistance complicates treatment outcomes. Thus, emergence of pathogenic MDR K. pneumoniae poses a great threat to the healthcare system. However, the carriage of antibiotic resistance among pathogenic K. pneumoniae is yet to be investigated in Uganda. We sought to investigate the carbapenem resistance profiles and pathogenic potential based on capsular serotypes of K. pneumoniae clinical isolates.MethodsThis was a cross sectional study involving use of archived Klebsiella pneumoniae isolates collected between January and December, 2019 at four tertiary hospitals in Uganda. All isolates were subject to antimicrobial susceptibility assays to determine phenotypic antibiotic resistance, pentaplex PCR to detect carbapenemases encoding genes and heptaplex PCR to identify capsular serotypes K1, K2, K3, K5, K20, K54 and K57.ResultsThe study found an overall phenotypic carbapenem resistance of 23.3% (53/227) and significantly higher genotypic resistance prevalence of 43.1% (98/227). Over all, the most prevalent gene was blaOXA-48-like (36.4%), followed by blaIMP-type (19.4%), blaVIM-type (17.1%), blaKPC-type (14.0%) and blaNDM-type (13.2%). blaVIM-type and blaOXA-48-like conferred phenotypic resistance in all isolates and 38.3% of isolates that harbored them respectively. Capsular multiplex PCR revealed that 46.7% (106/227) isolates were pathogenic and the predominantly prevalent pathotype was K5 (18.5%) followed by K20 (15.1%), K3 (7.1%), K2 (3.1%) and K1 (2.2%). Of the 106 capsular serotypes, 37 expressed phenotypic resistance; thus, 37 of the 53 carbapenem resistant K. pneumoniae were pathogenic.ConclusionThe high prevalence of virulent and antibiotic resistant K. pneumoniae among clinical isolates obtained from the four tertiary hospital as revealed by this study pose a great threat to healthcare. Our findings underline the epidemiological and public health risks and implications of this pathogen.

Prevalence of Carbapenem-Resistant Klebsiella pneumoniae Co-Harboring blaKPC-Carrying Plasmid and pLVPK-Like Virulence Plasmid in Bloodstream Infections.

This study aimed to characterize carbapenem-resistant Klebsiella pneumoniae (CR-KP) co-harboring bla KPC-2-carrying plasmid and pLVPK-like virulence plasmid. Between December 2017 and April 2018, 24 CR-KP isolates were recovered from 24 patients with bacteremia. The mortality was 66.7%. Pulsed-field gel electrophoresis and multilocus sequence typing results indicated four clusters, of which cluster A (n = 21, 87.5%) belonged to ST11 and the three remaining isolates (ST412, ST65, ST23) had different pulsotypes (cluster B, C, D). The bla KPC-2-carrying plasmids all belonged to IncFIIK type, and the size ranged from 100 to 390 kb. Nineteen strains (79.2%) had a 219-kb virulence plasmid possessed high similarity to pLVPK from CG43 with serotype K2. Two strains had a 224-kb virulence plasmid resembled plasmid pK2044 from K. pneumoniae NTUH-K2044(ST23). Moreover, three strains carried three different hybrid resistance- and virulence-encoding plasmids. Conjugation assays showed that both bla KPC-2 and rmpA2 genes could be successfully transferred to E. coli J53 in 62.5% of the strains at frequencies of 4.5 × 10−6 to 2.4 × 10−4, of which three co-transferred bla KPC-2 along with rmpA2 in large plasmids. Infection assays in the Galleria mellonella model demonstrated the virulence level of these isolates was found to be consistently higher than that of classic Klebsiella pneumoniae. In conclusion, CR-KP co-harboring bla KPC-2-carrying plasmid and pLVPK-like virulence plasmid were characterized by multi-drug resistance, enhanced virulence, and transferability, and should, therefore, be regarded as a real superbug that could pose a serious threat to public health. Hence, heightened efforts are urgently needed to avoid its co-transmission of the virulent plasmid (gene) and resistant plasmid (gene) in clinical isolates.

First Report of an Extensively Drug-Resistant ST23 Klebsiella pneumoniae of Capsular Serotype K1 Co-Producing CTX-M-15, OXA-48 and ArmA in Spain.

An extensively drug-resistant (XDR) Klebsiella pneumoniae isolate MS3802 from a tracheostomy exudate was whole-genome sequenced using MiSeq and Oxford Nanopore MinION platforms in order to identify the antimicrobial resistance and virulence determinates and their genomic context. Isolate MS3802 belonged to the clone ST23 and presented a capsular serotype K1, associated with hypervirulent K. pneumoniae (hvKp) isolates. The isolate harboured a chromosomally encoded blaCTX-M-15 gene and contained a large IncHI1B hybrid virulence/resistance plasmid carrying another copy of the blaCTX-M-15 and the virulence factors iucABCD-iutA, iroBCDN, rmpA and rmpA2. The carbapenemase gene blaOXA-48 was found in a Tn1999-like transposon and the 16S rRNA methylase armA gen located in the vicinity of other antibiotic-resistant genes on an IncM2 plasmid. This study represents, to the best of our knowledge, the first description of a blaCTX-M-15-, blaOXA-48- and armA-harbouring K. pneumoniae of ST23 and capsular serotype K1 in Spain. Our report emphasizes the importance of implementing new surveillance strategies to monitor the risk of emergence and spread of such XDR and hypervirulent K. pneumoniae isolates.

A fatal case of liver abscess caused by hypervirulent Klebsiella pneumoniae in a diabetic adolescent: A clinical and laboratory study.

ABSTRACTImportanceHypervirulent variants of Klebsiella pneumoniae (hvKp) are capable of causing life‐threatening pyogenic liver abscesses (PLAs), but hvKp caused PLAs was seldom reported in pediatric populations. Hence, there is an urgent need to raise our awareness of this phenomenon in pediatric populations.ObjectiveThis study aimed to report the clinical characteristics of hvKp that caused fatal PLA complicated by bacteremia in an adolescent and further identify the microbiological and genomic features of the causative strain.MethodsA 14‐year‐old boy with diabetes mellitus was admitted to our hospital with a diagnosis of PLA complicated by bacteremia. A hypermucoviscous hvKp strain, KPN_19‐106, was isolated from the drainage fluid present within the liver abscess cavity and blood. The hypermucoviscosity phenotype of the causative strain was determined by string test. Its virulence was measured using serum resistance assay and Galleria mellonella larvae‐killing assay. Antimicrobial susceptibility was determined by broth microdilution method. Genetic information was obtained by whole‐genome sequencing and bioinformatics analysis.ResultsKPN_19‐106 belonged to sequence type 380 and serotype K2 and exhibited stronger serum resistance and higher in vivo lethality than the well‐characterized hvKp NTUH‐K2044 strain. Although KPN_19‐106 is susceptible to most antibiotics, no sign of improvement was observed during treatment with such drugs. Whole‐genome sequencing revealed that the isolate had integrated multiple mobile genetic elements related to virulence.InterpretationAntibiotic‐susceptible hvKp can cause fatal PLA complicated by bacteremia in adolescents, with no improvement during antimicrobial therapy. The causative strain in this case had integrated multiple virulence genes and thus exhibited higher virulence both in vitro and in vivo when compared with NTUH‐K2044.

Genome-Based Analysis of a Sequence Type 1049 Hypervirulent Klebsiella pneumoniae Causing Bacteremic Neck Abscess.

Hypervirulent Klebsiella pneumoniae (hvKP) has raised grave concerns in recent years and can cause severe infections with diverse anatomic locations including liver abscess, meningitis, and endophthalmitis. However, there is limited data about neck abscess caused by hvKP. A K. pneumoniae strain Kp_whw was isolated from neck abscess. We characterized the genetic background, virulence determinates of the strain by genomic analysis and dertermined the virulence level by serum resistance assay. Kp_whw belonged to sequence type (ST) 1049 K locus (KL) 5. Kp_whw showed hypermucoviscosity phenotype and was resistant to ampicillin but susceptible to the majority of the other antimicrobial agents. A pLVPK-like virulence plasmid and a chromosomal ICEKp5-like mobile genetic element were carried by Kp_whw, resulting in the risk of dissemination of hypervirulence. The strain exhibited relative higher level of core genome allelic diversity than accessory genome profile, in comparison to hvKP of K1/K2 serotype. Kp_whw was finally demonstrated as virulent as the ST23 K1 serotype hvKP strain NTUH-K2044 in vitro. In conclusion, this work elaborates the genetic background of a clinical hvKP strain with an uncommon ST, reinforcing our understanding of virulence mechanisms of hvKP.

Development of monospecific polyclonal antibodies against hypervirulent Klebsiella pneumoniae

Abstract. Darniati, Setiyaningsih S, Agungpriyono DR, Handharyani E. 2021. Development of monospecific polyclonal antibodies against hypervirulent Klebsiella pneumoniae. Biodiversitas 22: 99-105. The aim of the research was to produce monospecific antibodies against Klebsiella pneumoniae serotype K1 and K2. Klebsiella pneumonia isolates were recovered from pneumonic lungs of Aceh cattle. The bacteria were identified by rpoB (RNA polymerase β subunit) PCR amplification specific for K. pneumoniae. The presence of capsule cluster gene magA and k2A was used to characterize capsular serotype K1 and K2, respectively. Antisera were produced using serial immunization of New Zealand White rabbits with K. pneumoniae serotypes K1 and K2. The presence of antibodies was determined by using immunodiffusion test and purification was performed by immunoaffinity purification method. K. pneumoniae antibodies began to appear on the 14th day after the first injection and progressively intensified after the 1st and 2nd booster. The cross-reactivity between the antigen was eliminated by absorbing the antisera with the opposite antigen and several serotypes of non-K1/K2. After purification, serum protein concentrations were substantially decreased from 58.48 µg/µL and 53.99 µg/µL to 2.38 µg / µL and 3.72 µg / µL for K1 and K2, respectively). The SDS-PAGE analysis showed two bands with molecular weight at 54 kDa and 25 kDa representing the heavy and light chains of immunoglobulin G, respectively. Purified polyclonal antiseras against K. pneumoniae serotypes K1 and K2 showed high affinity and specificity for homologous antigens in immunodiffusion and direct agglutination tests, and immunohistochemical staining.

The Surface Protein Fructose-1, 6 Bisphosphate Aldolase of Klebsiella pneumoniae Serotype K1: Role of Interaction with Neutrophils.

Hypermucoviscosity phenotypic Klebsiella pneumoniae (HV-Kp) serotype K1 is the predominant pathogen of a pyogenic liver abscess, an emerging infectious disease that often complicates septic metastatic syndrome in diabetic patients with poor sugar control. HV-Kpisolates were more resistant to neutrophil phagocytosis than non-HV-Kpisolates because of different pathogen-associated molecular patterns. The protein expression of HV-Kp after interaction with neutrophils is unclear. We studied KP-M1 (HV phenotype; serotype K1), DT-X (an acapsularmutant strain of KP-M1), and E. coli (ATCC 25922) with the model of Kp-infected neutrophils, using a comparative proteomic approach. One the identified protein, namely fructose-1, 6-bisphosphate aldolase (FBA), was found to be distributed in the KP-M1 after infecting neutrophils. Cell fractionation experiments showed that FBA is localized both to the cytoplasm and the outer membrane. Flow cytometry demonstrated that outer membrane-localized FBA was surface-accessible to FBA-specific antibody. The fba gene expression was enhanced in high glucose concentrations, which leads to increasing bacterial resistance to neutrophils phagocytosis and killing. The KP-M1 after FBA inhibitors and FBA-specific antibody treatment showed a significant reduction in bacterial resistance to neutrophils phagocytosis and killing, respectively, compared to KP-M1 without treatment. FBA is a highly conserved surface-exposed protein that is required for optimal interaction of HV-Kp to neutrophils.

Characterization of blaKPC-2-Harboring Klebsiella pneumoniae Isolates and Mobile Genetic Elements from Outbreaks in a Hospital in Ecuador.

Aim: To investigate the mobile genetic elements harboring blaKPC gene in carbapenem-resistant Klebsiella pneumoniae recovered during a 6-month outbreak in a high-complexity hospital from Ecuador. Results: A total of 62 isolates belonging to ST258 pilv-I-positive (n = 45), ST25 serotype K2 (n = 8), ST348 (n = 6), ST42 (n = 1), ST196 (n = 1), and ST1758 (n = 1) were collected from intensive care unit (ICU), neurosurgery, burn unit, internal medicine, pneumology, and neurology. Pulsed-field gel electrophoresis analysis showed two major clusters of ST258 and ST25 related to bloodstream infections and pneumonia circulating in ICU. The PCR assay showed that in non-ST258 isolates, the blaKPC-2 gene were located on the Tn4401a transposon inserted in the transferable pKpQIL-like IncFIIK2 plasmid; the whole-genome sequencing of ST258 clone showed two plasmids, the blaKPC-2 gene was located on nonconjugative IncR plasmid, whereas the IncFIB/IncFII plasmid lacked ß-lactamase genes. We found an IncM plasmid in blaKPC-2-harboring Klebsiella pneumoniae ST1758 clone. Conclusions: These findings highlight the presence of pKpQIL-like plasmids in non-ST258 and nonconjugative plasmids in ST258 isolates causing hospital outbreaks.

Klebsiella Distinctive Syndrome Presenting with Muscular Abscess and Osteomyelitis: Case Report

The present article describes a case of 46-year-old man who presented with progressive pain and swelling at the right lower back and buttock for three weeks. The investigation showed an abscess at the right lower back and buttock, osteomyelitis of the iliac bone, and a hepatic abscess. The causative pathogens were confirmed to be Klebsiella pneumoniae, both K1 and K2 serotype. This patient was treated with surgical management, percutaneous drainage, intravenous antibiotic, and control of his underlying disease risk factor, which was diabetic mellitus. Keywords: Klebsiella distinctive syndrome, Klebsiella liver abscess syndrome, Hypervirulent strain of Klebsiella pneumoniae, Klebsiella pneumoniae, Liver abscess

Clinical Features and Risk Factors Associated With 30-Day Mortality in Patients With Pneumonia Caused by Hypervirulent Klebsiella pneumoniae (hvKP).

BackgroundReports on metastatic or invasive infections by hypervirulent Klebsiella pneumoniae (hvKP) have increased recently. However, the effects of its virulence on clinical course and outcomes in pneumonia patients have rarely been addressed. We assessed and compared the clinical features of hvKp and classic K. pneumoniae (cKP) strains isolated from patients with pneumonia caused by K. pneumoniae. We also investigated the effects of virulence factors and the K. pneumoniae capsular serotypes K1 and K2 on mortality.MethodsIn this retrospective study, we enrolled 91 patients diagnosed as having pneumonia caused by K. pneumoniae and obtained their demographic and clinical data from medical records. We evaluated genes for K1 and K2, antimicrobial susceptibility, and the virulence genes rmpA, iutA, entB, ybtS, kfu, mrkD, and allS. Strains that possessed rmpA and iutA were defined as hvKP (N=39), while the remaining were classified as cKP (N=52). Odds ratio (OR) for the risk factors associated with 30-day mortality was calculated using the binary logistic regression model.ResultsThe 30-day mortality in all patients was 23.1%; it was 17.9% (7/39) in the hvKP group and 26.9% (14/52) in the cKP group (P=0.315). Bacteremia (OR=38.1; 95% confidence interval [CI], 2.5–570.2), altered mental status (OR=8.8; 95% CI, 1.7–45.0), and respiratory rate >30 breaths/min (OR=4.8; 95% CI, 1.2–20.0) were independent risk factors for 30-day mortality in all patients.ConclusionsOur results suggest that hypervirulence determinants do not have a significant effect on 30-day mortality in patients with pneumonia caused by K. pneumoniae.

A Synthetic Carbohydrate–Protein Conjugate Vaccine Candidate against Klebsiella pneumoniae Serotype K2

Klebsiella pneumoniae causes pneumonia and liver abscesses in humans worldwide and contains virulence factor capsular polysaccharides and lipopolysaccharides linked to the cell wall. Although capsular polysaccharides are good antigens for vaccine production and capsular oligosaccharides conjugate vaccines are proven effective against infections caused by encapsulated pathogens, there is still no Klebsiella pneumoniae vaccine available. One obstacle is that the capsular polysaccharide of a dominated Klebsiella pneumoniae serotype K2 is difficult to synthesize chemically due to the three 1,2-cis linkages in its structure. In this study, we successfully synthesized K2 capsular polysaccharides from tetra- to octasaccharides in highly a stereoselective manner. Subsequently, three synthesized glycans were conjugated to DT protein to provide glycoconjugate vaccine candidates (DT-Hexa, DT-Hepta, and DT-Octa) that were used in in vivo immunization experiments in mice. The results of immunized studies showed all three glycoconjugates elicited antibodies that recognized all of the synthetic glycans at 1:200-fold dilution. Particularly, the DT-Hepta conjugate elicited a higher level of antibodies that can recognize longer glycan (octasaccharide) even at 1:12800-fold dilution and exhibited good bactericidal activity. Our results concluded that heptasaccharide is the minimal epitope and a potential candidate for the vaccine against the K2 sero group of Klebsiella pneumoniae.

Change of Hypermucoviscosity in the Development of Tigecycline Resistance in Hypervirulent Klebsiella pneumoniae Sequence Type 23 Strains.

In this study, we developed tigecycline resistance in Klebsiella pneumoniae ST23 strains in vitro and investigated the change in virulence associated with hypermucoviscosity. In vitro-induced tigecycline-resistant (TGC-IR) K. pneumoniae mutants were obtained from three tigecycline-susceptible (TGC-S) strains, belonging to ST23 and serotype K1, by culturing in media with tigecycline in a stepwise manner. An antimicrobial susceptibility test, string test, mucoviscosity assay, and capsular polysaccharide (CPS) quantification were performed. Biofilm formation and serum resistance were evaluated, and survival rates of bacterial strains in fruit flies and macrophages were measured. Alterations of rpsJ, ramR, soxR, acrR, and marR genes were investigated and the expression levels of ramA and efflux pump genes were evaluated. The hypermucoviscosity phenotype was dramatically decreased in the TGC-IR mutants. Reduced CPS production in TGC-IR mutants was also identified. Increased resistance to most other antimicrobial agents was found in TGC-IR mutants. In addition, the TGC-IR mutants exhibited reduced biofilm formation, low serum resistance, and decreased survival rates within fruit flies and macrophages. Our study shows that development of tigecycline resistance in hypervirulent K. pneumoniae strains result in defects in virulence associated with hypermucoviscosity.