K2 Strains Research Articles

Identification of three capsule depolymerases in a bacteriophage infecting Klebsiella pneumoniae capsular types K7, K20, and K27 and therapeutic application

BackgroundKlebsiella pneumoniae capsular types K1, K2, K5, K20, K54, and K57 are prevalent hypervirulent types associated with community infections, and worrisomely, hypervirulent strains that acquired drug resistance have been found. In the search for alternative therapeutics, studies have been conducted on phages that infect K. pneumoniae K1, K2, K5, and K57-type strains and their phage-encoded depolymerases. However, phages targeting K. pneumoniae K20-type strains and capsule depolymerases capable of digesting K20-type capsules have rarely been reported. In this study, we characterized a phage that can infect K. pneumoniae K20-type strains, phage vB_KpnM‐20.MethodsA phage was isolated from sewage water in Taipei, Taiwan, its genome was analyzed, and its predicted capsule depolymerases were expressed and purified. The host specificity and capsule-digesting activity of the capsule depolymerases were determined. The therapeutic effect of the depolymerase targeting K. pneumoniae K20-type strains was analyzed in a mouse infection model.ResultsThe isolated Klebsiella phage, vB_KpnM‐20, infects K. pneumoniae K7, K20, and K27-type strains. Three capsule depolymerases, K7dep, K20dep, and K27dep, encoded by the phage were specific to K7, K20, and K27-type capsules, respectively. K20dep also recognized Escherichia coli K30-type capsule, which is highly similar to K. pneumoniae K20-type. The survival of K. pneumoniae K20-type-infected mice was increased following administration of K20dep.ConclusionsThe potential of capsule depolymerase K20dep for the treatment of K. pneumoniae infections was revealed using an in vivo infection model. In addition, K7dep, K20dep, and K27dep capsule depolymerases could be used for K. pneumoniae capsular typing.

Functional Characterization of Plasmid-Borne rmpADC Homologues in Klebsiella pneumoniae.

Expression of the hypermucoviscosity (HMV) phenotype and capsular polysaccharide (CPS) biosynthesis in Klebsiella pneumoniae were reported to be encoded by genes located in the chromosomal rmp locus. However, the functions of the rmp locus in the virulence plasmid remained unclear, and most of the rmp loci in clinical K. pneumoniae are plasmid carried. In this study, we investigated the functional characteristics of plasmid-borne rmp homologues in clinical hypervirulent K. pneumoniae (hvKP) strains by cloning and introducing such gene homologues into K. pneumoniae strains of different capsule types, followed by the evaluation of phenotypic changes in these strains. Acquisition of the plasmid-borne prmpADC and prmpA2D2 loci were found to result in an increase in mucoviscosity and CPS production in K1 and K2 K. pneumoniae, while only the prmpA2D2 locus contributed to phenotypic changes in the ST11/KL64 strain. Consistently, both rmpD and rmpD2 increased HMV in K1 and K2 K. pneumoniae, while only rmpD2 contributed to HMV in the ST11/KL64 strain; rmpC contributed to CPS overproduction in K1 and K2 strains but not in the ST11/KL64 strain. Furthermore, we proposed a logistic molecular basis of the HMV phenotype of K. pneumoniae on which prmpD2-mediated HMV is attributed to the increase of cell-free CPS production. Our data confirm that the rmp homologues carried by the virulence plasmid play a key role in virulence expression in K. pneumoniae, but the phenotype is highly dependent on the genetic background of the host strain and explained why most of the clinical ST11 strains carry only the prmpA2D2 locus. IMPORTANCE Klebsiella pneumoniae has become the most frequently isolated bacterial pathogen in hospital settings, with a very high mortality rate worldwide. Factors contributing to the virulence of K. pneumoniae are the overproduction of capsular polysaccharide (CPS) as well as the hypermucoviscosity (HMV) phenotype. These two phenotypes were reported to be regulated by rmpA/A2 homologues, which are often carried by virulence plasmids. Here, we determined the functional role of two plasmid-borne rmpA in mediating expression of the HMV phenotype and CPS production in K. pneumoniae. Different capsule types exhibited differences in the expression of HMV and CPS production although they harbored an identical plasmid-borne rmpA or rmpA2 locus, indicating that these virulence-related phenotypes are strongly related to the genetic background of the host strains. Our study provides a novel understanding of the regulation of virulence-related phenotypes and clinical management of K. pneumoniae infections.

Keeping alert to the hypervirulent K1, K2, K3, K5, K54 and K57 strains of Klebsiella pneumoniae within dairy production process

Klebsiella pneumoniae (Kp) is now recognized as an urgent threat to public health since the emergence of multidrug-resistant and hypervirulent isolates. We identified a hypervirulent K2 isolate from the milk samples possibly associated with an infection incident in children, which raised the alarm to the zoonotic potential of bovine mastitis Kp as a foodborne pathogen. Subsequently, numerous K1, K2, K3, K5, K54 and K57 strains were identified from mastitis milk samples, and showed high pathogenicity in infected mouse. Further analysis based on complete genomes found that these isolates showed closely evolutionary relationships with the human hypervirulent strains in diverse phylogenetic lineages, suggesting their potential risk to public health.

Hypervirulence and Multiresistance to Antibiotics in Klebsiella pneumoniae Strains Isolated from Patients with Hospital- and Community-Acquired Infections in a Mexican Medical Center.

Klebsiella pneumoniae is a pathogenic bacterium associated with different infectious diseases. This study aimed to establish the different association profiles of virulence genes related to the hypermucoviscous phenotype (HM), capsular serotypes, biofilm formation, and multidrug resistance in K. pneumoniae strains from patients with hospital- and community-acquired infections. K. pneumoniae virulence genes and capsular serotypes were identified by PCR, antibiotic susceptibility by the Kirby–Bauer method, HM by the string test, and biofilm formation by measurement in polystyrene microtiter plates. Of a total of 150 strains from patients with hospital- (n = 25) and community-acquired infections (n = 125), 53.3% (80/150) were HM-positive and 46.7% (70/150) were HM-negative. HM-positive (68/80) and HM-negative (67/70) strains were biofilm-forming. Moreover, 58.7% (47/80) HM-positive and 57.1% (40/70) HM-negative strains were multidrug-resistant. Among HM-positive, HM-negative, and serotypes K1 (25/150), K2 (48/150), and non-K1/K2 strains, (77/150) the frequently detected adhesion genes were fimH, mrkD, ycfM, and kpn; entB, irp2, irp1, and ybtS, for iron acquisition; and rmpA for protectins. The gene association pattern fimH/kpn/mrkD/ycfM/entB/irp1/irp2/ybtS/fyuA (18/150) was frequent among the strains. K. pneumoniae strains from patients with hospital- and community-acquired infections demonstrated a wide diversity of virulence gene profiles related to phenotype (hypermucoviscosity, multidrug resistance, and biofilm formation) and serotypes.

Emergence of K1 ST23 and K2 ST65 hypervirulent klebsiella pneumoniae as true pathogens with specific virulence genes in cryptogenic pyogenic liver abscesses Shiraz Iran.

Hypervirulent Klebsiella pneumoniae (hvKp) pathotype is emerging worldwide in pyogenic liver abscesses (PLAs). However, the role of virulence factors in pathogenicity remains unclear. On the other hand, the epidemiology of PLAs in Iran is unknown. From July 2020 to April 2022, bacterial species were isolated and identified from the drainage samples of 54 patients with PLAs. K. pneumoniae as the most common pathogen of pyogenic liver abscesses was identified in 20 (37%) of the 54 patients. We analyzed the clinical and microbiological characteristics of K. pneumoniae-related pyogenic liver abscesses. Antibiotic susceptibility testes and string test were performed. 16S rRNA, antibiotic resistance, and virulence genes were determined by polymerase chain reaction amplification. Clonal relatedness of isolates was identified by multilocus sequence typing. Virulence levels were assessed in the Galleria mellonella larval infection model. Four hvKp isolates (K1/K2) were found to be responsible for cryptogenic PLAs, and 16 classical K. pneumoniae isolates (non-K1/K2) were associated with non-cryptogenic PLAs. Three capsular serotype K1 strains belonged to sequence type 23 (ST23) and one K2 strain to ST65. Meanwhile, the non-K1/K2 strains belonged to other STs. ST231 was the most common strain among the classical K. pneumoniae strains. Compared with the non-K1/K2 strains, capsular serotypes K1/K2 strains were less resistant to antibiotics, had positive string test results, and had more virulence genes. In Galleria mellonella, a concentration of 106 colony-forming units of the K1 hvKp strain resulted in 100% death at 24 hours, confirming the higher virulence of the hvKp strain compared with cKp. K. pneumoniae isolates represented that the acquisition of any plasmid or chromosomal virulence genes contributes to pathogenicity and high prevalence in PLAs. Meanwhile, hvKp isolates with a specific genetic background were detected in cryptogenic PLAs.

Genetic and Functional Characterization of a Conjugative KpVP-2-Type Virulence Plasmid From a Clinical Klebsiella pneumoniae Strain.

The main mechanism of virulence in Klebsiella pneumoniae is the acquisition of K. pneumoniae virulence plasmids (KpVPs), which include two dominant types, namely, KpVP-1 (carrying iuc1, iro1, rmpA, and rmpA2) and KpVP-2 (carrying iuc2, iro2, and rmpA). Both are non-conjugative and associated with different hypervirulent clones. In contrast to KpVP-1 reported in K1, K2, and other serotypes of K. pneumoniae, KpVP-2 was only reported in K2 strains and rarely characterized. In this study, we identified a conjugative KpVP-2-type virulence plasmid from a clinical hypervirulent K. pneumoniae strain. This plasmid was generated by the integration of conjugative transfer genes into the KpVP-2-type plasmid Kp52.145 II and could be readily conjugated to Escherichia coli strain EC600 and K. pneumoniae strains of various types which are clinically existing, mediating hypervirulence. Furthermore, this kind of conjugative KpVP-2-type virulence plasmid has been disseminated in clinical settings in Hong Kong and other regions of the world. The generation of conjugative virulence plasmid may promote its transmission and explain the evolution of this type of virulence plasmid.

Epidemiological Features of Klebsiella pneumoniae Infection in the Hepatobiliary System of Patients in Yantai, China, Based on Clinical and Genetic Analyses.

PurposeTo investigate the epidemiological features of Klebsiella pneumoniae infection of the hepatobiliary system of patients in Yantai, China.MethodsThis retrospective study was conducted from January to December 2019 in Yantai Yuhuangding Hospital. Patients for whom K. pneumoniae was isolated from the hepatobiliary system were considered for inclusion. The clinical features and genetic analyses were conducted to explore the epidemiological characteristics.ResultsA total of 88 cases were enrolled, including 69 cases of hypervirulent K. pneumoniae (hvKP) and 19 cases of classical K. pneumoniae (cKP). Community-acquired infections, fever, liver abscess, and C-reactive protein (CRP) and procalcitonin (PCT) levels were significantly higher, while biliary tract disease was lower in the hvKP group compared with the cKP group. Among the 69 hvKP infections, 61 developed a liver abscess. Community-acquired infections, fever, and CRP and PCT levels were higher, whereas biliary tract disease and malignancy were lower in the liver abscess group compared with the non-liver abscess group. All strains were susceptible to the majority of antibiotics tested. All hvKP strains possessed the blaSHV, oqxA, oqxB and fosA resistance genes. K1 and K2 accounted for 78% of hvKP strains. K1 strains belonged to sequence types ST23 and ST700, whereas K2 strains belonged to ST65, ST86 and ST5212. K1 isolates possessed the most virulence determinants, followed by K2 and non-K1/K2 isolates. K2 isolates lacked the allS gene, which was rare in non K1/K2 isolates, but present in most K1 isolates. The mceG gene was only detected in K1 isolates. AllS and virulence determinants were significantly more prevalent in the liver abscess group than in the non-liver abscess group.ConclusionThe prevalence of hvKP among K. pneumoniae infections of the hepatobiliary system is high in Yantai, China. Greater vigilance of hvKP infection is required in clinical and microbiological laboratories.

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.

Variation and Distribution of L-A Helper Totiviruses in Saccharomyces sensu stricto Yeasts Producing Different Killer Toxins

Yeasts within the Saccharomyces sensu stricto cluster can produce different killer toxins. Each toxin is encoded by a medium size (1.5–2.4 Kb) M dsRNA virus, maintained by a larger helper virus generally called L-A (4.6 Kb). Different types of L-A are found associated to specific Ms: L-A in K1 strains and L-A-2 in K2 strains. Here, we extend the analysis of L-A helper viruses to yeasts other than S. cerevisiae, namely S. paradoxus, S. uvarum and S. kudriavzevii. Our sequencing data from nine new L-A variants confirm the specific association of each toxin-producing M and its helper virus, suggesting co-evolution. Their nucleotide sequences vary from 10% to 30% and the variation seems to depend on the geographical location of the hosts, suggesting cross-species transmission between species in the same habitat. Finally, we transferred by genetic methods different killer viruses from S. paradoxus into S. cerevisiae or viruses from S. cerevisiae into S. uvarum or S. kudriavzevii. In the foster hosts, we observed no impairment for their stable transmission and maintenance, indicating that the requirements for virus amplification in these species are essentially the same. We also characterized new killer toxins from S. paradoxus and constructed “superkiller” strains expressing them.

Comparison of Diabetic and Non-diabetic Human Leukocytic Responses to Different Capsule Types of Klebsiella pneumoniae Responsible for Causing Pyogenic Liver Abscess.

The major risk factor for Klebsiella liver abscess (KLA) is type 2 diabetes mellitus (DM), but the immunological mechanisms involved in the increased susceptibility are poorly defined. We investigated the responses of neutrophils and peripheral blood mononuclear cells (PBMCs) to hypervirulent Klebsiella pneumoniae (hvKP), the causative agent of KLA. DNA and myeloperoxidase levels were elevated in the plasma of KLA patients compared to uninfected individuals indicating neutrophil activation, but diabetic status had no effect on these neutrophil extracellular trap (NET) biomarkers in both subject groups. Clinical hvKP isolates universally stimulated KLA patient neutrophils to produce NETs ex vivo, regardless of host diabetic status. Ability of representative capsule types (K1, K2, and non-K1/K2 strains) to survive intra- and extra-cellular killing by type 2 DM and healthy neutrophils was subsequently examined. Key findings were: (1) type 2 DM and healthy neutrophils exhibited comparable total, phagocytic, and NETs killing against hvKP, (2) phagocytic and NETs killing were equally effective against hvKP, and (3) hypermucoviscous K1 and K2 strains were more resistant to total, phagocytic, and NETs killing compared to the non-mucoviscous, non-K1/K2 strain. The cytokine response and intracellular killing ability of type 2 DM as well as healthy PBMCs upon encounter with the different capsule types was also examined. Notably, the IL-12–IFNγ axis and its downstream chemokines MIG, IP-10, and RANTES were produced at slightly lower levels by type 2 DM PBMCs than healthy PBMCs in response to representative K1 and non-K1/K2 strains. Furthermore, type 2 DM PBMCs have a mild defect in its ability to control hvKP replication relative to healthy PBMCs. In summary, our work demonstrates that type 2 DM does not overtly impact neutrophil intra- and extra-cellular killing of hvKP, but may influence cytokine/chemokine production and intracellular killing by PBMCs.

Relationships and Evolution of Double-Stranded RNA Totiviruses of Yeasts Inferred from Analysis of L-A-2 and L-BC Variants in Wine Yeast Strain Populations.

Killer strains of S. cerevisiae secrete protein toxins that kill nonkiller yeasts. The "killer phenomenon" depends on two dsRNA viruses: L-A and M. M encodes the toxin, and L-A, the helper virus, provides the capsids for both viruses. Different killer toxins exist: K1, K2, K28, and Klus, encoded on different M viruses. Our data indicate that each M dsRNA depends on a specific helper virus; these helper viruses have nucleotide sequences that may be as much as 26% different, suggesting coevolution. In wine environments, K2 and Klus strains frequently coexist. We have previously characterized the association of Mlus and L-A-lus. Here we sequence and characterize L-A-2, the helper virus of M2, establishing the helper virus requirements of M2, which had not been completely elucidated. We also report the existence of two specific L-BC totiviruses in Klus and K2 strains with about 10% of their nucleotides different, suggesting different evolutionary histories from those of L-A viruses.

Reinfection and Relapse of Recurrent Bacteremia caused by Klebsiella Pneumoniae in a Medical Center in Taiwan

To delineate the genetic relatedness of Klebsiella pneumoniae causing recurrent bacteremia. Clinical characteristics of patients, distribution of capsular genoserotypes by capsular cps-PCR genotyping and the genetic relatedness of the paired isolates using arbitrarily primed PCR and pulsed-field gel electrophoresis were investigated. Of 3067 patients with K. pneumoniae bacteremia, 129 (4.2%) had recurrent bacteremia (301 episodes). Of 129 patients with recurrent bacteremia, 51 (39.5%) were reinfected by the same strains (paired isolates with identical randomly amplified polymorphic DNA patterns and closely related pulsotypes). Multivariate analysis demonstrated that diabetes mellitus (odds ratio: 5.82; p < 0.001) and infection due to serotype K2 (odds ratio: 13.93; p < 0.001) were associated with relapse in patients with recurrent bacteremia. Recurrent K. pneumoniae bacteremia could occur in patients with underlying hematological malignancies or suffering from primary bacteremia. Relapse infection is associated with serotype K2 strains and underlying diabetes mellitus.

Impacts of Hypervirulence Determinants on Clinical Features and Outcomes of Bacteremia Caused by Extended-Spectrum β-Lactamase-Producing Klebsiella pneumoniae.

We investigated the implications of hypervirulence determinants on clinical features of 48 adult patients with bacteremia caused by extended-spectrum β-lactamase-producing Klebsiella pneumoniae. Isolates in the hypervirulence group included any of the following virulence determinants: K1/K2 capsule serotypes, hypermucoviscosity phenotype, rmpA gene, or rmpA2 gene. Nonhypervirulence group isolates were negative for all of the above virulence factors. In this study, all isolates used were non-K1/K2 strains. Statistically significant differences were observed in clinical features of patients between the two groups. The hypervirulent isolates (n = 19), including 11 isolates with the hypermucoviscosity phenotype, 15 with the rmpA gene, and 16 with the rmpA2 gene, were more commonly recovered from diabetic patients and mainly manifested as secondary bacteremia (such as pneumonia, urinary tract infections, or other localized infections). The nonhypervirulent isolates (n = 29) were more commonly recovered from patients after prolonged hospital stays (>30 days) and mostly manifested as primary bacteremia. The overall in-hospital mortality was 56.3%. Hazard ratio (HR) analysis revealed the following positive predictors for mortality: nosocomial infection, stay in an intensive care unit, no removal of the central venous catheter, Charlson comorbidity score, and APACHE II score (≧15). The negative predictors were initial appropriate antibiotic therapy (HR 0.42) and urinary tract infection (HR 0.19). Charlson score was an independent confounder based on multivariate analysis (HR 1.43, 95% confidence interval 1.04-1.99). In conclusion, hypervirulence determinants played a role in causing secondary infections in diabetic patients; however, the presence of morbidity cofactors could themselves influence mortality, despite the absence of hypervirulence determinants.

Antibacterial activities of the methanol extracts, fractions and compounds from Harungana madagascariensis Lam. ex Poir. (Hypericaceae)

Ethnopharmacological relevanceHarungana madagascariensis Lam. ex Poir. (Hypericaceae) is used in folk medicine to treat a variety of human ailments, mainly antibacterial, antifungal, antiviral and viral infections. In the present study, the methanol extract from the leaves (HML) and bark (HMB) of this plant as well as fractions (HMBa-c), sub-fractions (HMBa1-5) and compounds isolated from HMBa and HMBb namely betulinic acid (1), madagascin (2), ferruginin A (3) and Kaempferol-3-O-β-d-glucopyranoside (4) were tested for their antimicrobial activities against a panel of 28 g-negative bacteria including multidrug resistant (MDR) phenotypes. Materials and methodsThe broth microdilution method was used to determine the minimal inhibitory concentration (MIC) and minimal bactericidal concentration (MBC) of the above samples; column chromatography was used for the fractionation and purification of the bark extract whilst the chemical structures of compounds were determined using spectroscopic techniques. ResultsCrude extract HMB together with fraction HMBa and sub-fraction HMBa3 were active on the 28 tested bacterial strains. HML as well as fractions HMBb, HMBc and sub-fractions HMBa1, HMBa2, HMBa4 and HMBa5 were selectively active. MIC values below or equal to 1024µg/mL were recorded with these samples on 92.9% (for HML and HMBa 4), 82.1% (for HMBb), 78.6% (for HMBa2), 50.0% (for HMBa5) and 42.9% (for HMBc) tested bacteria. For crude material, the lowest MIC value below 8µg/mL was obtained with HMB against Escherichia coli ATCC10536 and W3110 strains, and with sub-fraction HMBa3 against Klebsiella pneumoniae K2 strains. MIC values below 10µg/mL were recorded with compound 3 against E. coli ATCC10536, Enterobacter aerogenes ATCC13048 and EA294, Pseudomonas aeruginosa PA01, K. pneumoniae K2 and Kp55 and Enterobacter cloacae BM67. ConclusionsHarungana madagascariensis is a potential source of antimicrobial drugs to fight against MDR bacteria. The anthranol 3 is the main antibacterial constituents of the bark of the plant. HMB and compound 3 deserve further investigations to develop natural drug to combat Gram-negative bacteria and otherwise MDR phenotypes.

A Novel Saccharomyces cerevisiae Killer Strain Secreting the X Factor Related to Killer Activity and Inhibition of S. cerevisiae K1, K2 and K28 Killer Toxins.

It was determined that Kx strains secrete an X factor which can inhibit all known Saccharomyces cerevisiae killer toxins (K1, K2, K28) and some toxins of other yeast species-the phenomenon not yet described in the scientific literature. It was shown that Kx type yeast strains posess a killer phenotype producing small but clear lysis zones not only on the sensitive strain α'1 but also on the lawn of S. cerevisiae K1, K2 and K28 type killer strains at temperatures between 20 and 30°C. The pH at which killer/antikiller effect of Kx strain reaches its maximum is about 5.0-5.2. The Kx yeast were identified as to belong to S. cerevisiae species. Another newly identified S. cerevisiae killer strain N1 has killer activity but shows no antikilller properties against standard K1, K2 and K28 killer toxins. The genetic basis for Kx killer/antikiller phenotype was associated with the presence of M-dsRNA which is bigger than M-dsRNA of standard S. cerevisiae K1, K2, K28 type killer strains. Killer and antikiller features should be encoded by dsRNA. The phenomenon of antikiller (inhibition) properties was observed against some killer toxins of other yeast species. The molecular weight of newly identified killer toxins which produces Kx type strains might be about 45kDa.

Surface antigens contribute differently to the pathophysiological features in serotype K1 and K2 Klebsiella pneumoniae strains isolated from liver abscesses.

BackgroundThe virulence role of surface antigens in a single serotype of Klebsiella pneumoniae strain have been studied, but little is known about whether their contribution will vary with serotype.MethodTo investigate the role of K and O antigen in hyper-virulent strains, we constructed O and K antigen deficient mutants from serotype K1 STL43 and K2 TSGH strains from patients with liver abscess, and characterized their virulence in according to the abscess formation and resistance to neutrophil phagocytosis, serum, and bacterial clearance in liver.ResultsBoth of K1 and K2-antigen mutants lost their wildtype resistance to neutrophil phagocytosis and hepatic clearance, and failed to cause abscess formation. K2-antigen mutant became serum susceptible while K1-antigen mutant maintained its resistance to serum killing. The amount of glucuronic acid, indicating the amount of capsular polysaccharide (CPS, K antigen), was inversed proportional to the rate of phagocytosis. O-antigen mutant of serotype K1 strains had significantly more amount of CPS, and more resistant to neutrophil phagocytosis than its wildtype counterpart. O-antigen mutants of serotype K1 and K2 strains lost their wildtype serum resistance, and kept resistant to neutrophil phagocytosis. While both mutants lacked the same O1 antigen, O-antigen mutant of serotype K1 became susceptible to liver clearance and cause mild abscess formation, but its serotype K2 counterpart maintained these wildtype virulence.ConclusionWe conclude that the contribution of surface antigens to virulence of K. pneumoniae strains varies with serotypes.

K2 killer toxin-induced physiological changes in the yeast Saccharomyces cerevisiae.

Saccharomyces cerevisiae cells produce killer toxins, such as K1, K2 and K28, that can modulate the growth of other yeasts giving advantage for the killer strains. Here we focused on the physiological changes induced by K2 toxin on a non-toxin-producing yeast strain as well as K1, K2 and K28 killer strains. Potentiometric measurements were adjusted to observe that K2 toxin immediately acts on the sensitive cells leading to membrane permeability. This correlated with reduced respiration activity, lowered intracellular ATP content and decrease in cell viability. However, we did not detect any significant ATP leakage from the cells treated by killer toxin K2. Strains producing heterologous toxins K1 and K28 were less sensitive to K2 than the non-toxin producing one suggesting partial cross-protection between the different killer systems. This phenomenon may be connected to the observed differences in respiratory activities of the killer strains and the non-toxin-producing strain at low pH. This might also have practical consequences in wine industry; both as beneficial ones in controlling contaminating yeasts and non-beneficial ones causing sluggish fermentation.

Klebsiella pneumoniae subsp. pneumoniae-bacteriophage combination from the caecal effluent of a healthy woman.

A sample of caecal effluent was obtained from a female patient who had undergone a routine colonoscopic examination. Bacteria were isolated anaerobically from the sample, and screened against the remaining filtered caecal effluent in an attempt to isolate bacteriophages (phages). A lytic phage, named KLPN1, was isolated on a strain identified as Klebsiella pneumoniae subsp. pneumoniae (capsular type K2, rmpA+). This Siphoviridae phage presents a rosette-like tail tip and exhibits depolymerase activity, as demonstrated by the formation of plaque-surrounding haloes that increased in size over the course of incubation. When screened against a panel of clinical isolates of K. pneumoniae subsp. pneumoniae, phage KLPN1 was shown to infect and lyse capsular type K2 strains, though it did not exhibit depolymerase activity on such hosts. The genome of KLPN1 was determined to be 49,037 bp (50.53 %GC) in length, encompassing 73 predicted ORFs, of which 23 represented genes associated with structure, host recognition, packaging, DNA replication and cell lysis. On the basis of sequence analyses, phages KLPN1 (GenBank: KR262148) and 1513 (a member of the family Siphoviridae, GenBank: KP658157) were found to be two new members of the genus “Kp36likevirus.”

Mapping the Evolution of Hypervirulent Klebsiella pneumoniae.

ABSTRACTHighly invasive, community-acquired Klebsiella pneumoniae infections have recently emerged, resulting in pyogenic liver abscesses. These infections are caused by hypervirulent K. pneumoniae (hvKP) isolates primarily of capsule serotype K1 or K2. Hypervirulent K1 isolates belong to clonal complex 23 (CC23), indicating that this clonal lineage has a specific genetic background conferring hypervirulence. Here, we apply whole-genome sequencing to a collection of K. pneumoniae isolates to characterize the phylogenetic background of hvKP isolates with an emphasis on CC23. Most of the hvKP isolates belonged to CC23 and grouped into a distinct monophyletic clade, revealing that CC23 is a unique clonal lineage, clearly distinct from nonhypervirulent strains. Separate phylogenetic analyses of the CC23 isolates indicated that the CC23 lineage evolved recently by clonal expansion from a single common ancestor. Limited grouping according to geographical origin was observed, suggesting that CC23 has spread globally through multiple international transmissions. Conversely, hypervirulent K2 strains clustered in genetically unrelated groups. Strikingly, homologues of a large virulence plasmid were detected in all hvKP clonal lineages, indicating a key role in K. pneumoniae hypervirulence. The plasmid encodes two siderophores, aerobactin and salmochelin, and RmpA (regulator of the mucoid phenotype); all these factors were found to be restricted to hvKP isolates. Genomic comparisons revealed additional factors specifically associated with CC23. These included a distinct variant of a genomic island encoding yersiniabactin, colibactin, and microcin E492. Furthermore, additional novel genomic regions unique to CC23 were revealed which may also be involved in the increased virulence of this important clonal lineage.

Molecular epidemiology and virulence factors of pyogenic liver abscess causing Klebsiella pneumoniae in China

The molecular epidemiology and prevalence of virulence factors of isolates from patients with Klebsiella pneumoniae liver abscess (KLA) in mainland China are unknown. Klebsiella pneumoniae isolates were obtained from drainage samples aseptically collected from patients with pyogenic liver abscess (PLA). The genetic similarity of KLA isolates was analyzed by pulsed-field gel electrophoresis. The hypermucoviscosity (HV) phenotype was identified by a positive string test. The K1 and K2 genotypes, the pLVPK-derived genetic loci, aerobactin gene, kfu and alls were detected by PCR amplification. The sequence types (STs) were identified by multilocus sequence typing. Among the 51 non-repetitive KLA isolates, 49 PFGE types have been identified. In total, 19 (37.2%) and 14 (27.4%) of the 51 KLA isolates belonged to clonal complex (CC) 23 and CC65, respectively, while the other 18 isolates (35.3%) were defined as other STs. CC23 consisted of only K1 strains, while CC65 included only K2 strains. All non-K1/K2 strains were classified as STs other than CC23 and CC65. Approximately 70.6% (36/51) of KLA isolates exhibited an HV phenotype. Both K1 and K2 isolates presented significantly higher prevalence of the pLVPK-derived loci than non-K1/K2 isolates. The K1 isolates had a significantly higher prevalence of the kfu and allS genes than K2 and non-K1/K2 isolates, while the K2 isolates exhibited higher repA prevalence than K1 and non-K1/K2 isolates. The majority of KLA isolates belonged to CC23K1 and CC65K2, while other STs with non-K1/K2 capsular types have also been identified. The virulent factors exhibited diverse distribution among the different clones of KLA isolates.