Capsular serotype K1 or K2, rather than magA and rmpA, is a major virulence determinant for Klebsiella pneumoniae liver abscess in Singapore and Taiwan

Capsular serotypes, magA, and rmpA have been documented in high prevalence for Klebsiella pneumoniae liver abscess. To investigate the regional difference and the correlation of capsular serotype, magA, and rmpA with virulence, 73 isolates were collected in Singapore and Taiwan. Capsular serotypes were determined by countercurrent immunoelectrophoresis, the presence of magA and rmpA was determined by PCR, and virulence was determined by phagocytosis and mouse inoculation. Isolates from Singapore were similar to those from Taiwan in genomic heterogeneity, prevalence of serotype, and the presence of magA and rmpA. The most common serotype was K1 (34/73; 46.6%), followed by K2 (15/73; 20.5%). magA was restricted to serotype K1. All K1 or K2 isolates and 66.7% (16/24) of isolates that were neither serotype K1 nor serotype K2 (non-K1/K2) carried rmpA. Serotype K1 or K2 isolates demonstrated significantly more phagocytic resistance and virulence than did rmpA-positive and -negative groups of non-K1/K2 isolates. In the non-K1/K2 group, the virulence profiles of rmpA-positive strains from Taiwan and Singapore were different by phagocytosis assay and in the mouse model, indicating that factors other than rmpA contributed to virulence. The characteristics of K. pneumoniae liver abscess in Singapore and Taiwan are similar. Capsular serotype K1 or K2 plays a more important role than magA and rmpA in determining virulence in K. pneumoniae liver abscess.     

Emerging severe and fatal infections due to Klebsiella pneumoniae in two university hospitals in France

Severe infections caused by hypermucoviscous Klebsiella pneumoniae have been reported in Southeast Asian countries over the past several decades. This report shows their emergence in France, with 12 cases observed during a 2-year period in two university hospitals. Two clones (sequence type 86 [ST86] and ST380) of serotype K2 caused five rapidly fatal bacteremia cases, three of which were associated with pneumonia, whereas seven liver abscess cases were caused by K1 strains of ST23.     

Evidence for clonal dissemination of the serotype K1 Klebsiella pneumoniae strain causing invasive liver abscesses in Korea

Seventy-three liver abscess isolates of serotype K1 Klebsiella pneumoniae from a nationwide collection in Korea were genotypically characterized using pulsed-field gel electrophoresis and multilocus sequence typing. We found that serotype K1 K. pneumoniae strains that caused liver abscesses in Korea were genotypically related and that most were sequence type 23.     

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 CC23^K1 and CC65^K2, 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.     

Clinical spectrum and molecular characteristics of Klebsiella pneumoniae causing community-acquired extrahepatic abscess

BACKGROUND AND PURPOSE: Klebsiella pneumoniae causes a wide spectrum of infections, including abscess and non-abscess formation. This study investigated the clinical spectrum and molecular characteristics of community-acquired Klebsiella infection with primary extrahepatic abscess. METHODS: From April 2004 through March 2007, a total of 18 strains of K. pneumoniae, 11 from blood and 7 from focal purulent specimens, were recovered from a medical center in southern Taiwan. The clinical data were collected from medical records. Hypermucoviscosity phenotype was defined as positive string test. The virulence genes, including rmpA (regulator of mucoid phenotype), magA (specific to K1 capsule serotype), k(2)A (specific to K2 capsule serotype), and kfu (an iron uptake system) were amplified by polymerase chain reaction using specific primers. RESULTS: Twelve men and 6 women with ages ranging from 37 to 74 years were enrolled. Fifteen patients had underlying diabetes mellitus. The duration of hospitalization ranged from 1 to 96 days. Three patients died by the end of treatment. All of the K. pneumoniae strains carried rmpA and 16 strains showed the hypermucoviscosity phenotype. Of the 18 strains, 7 strains were positive for k(2)A and 4 strains carried magA. kfu was identified in 4 magA-positive strains and 2 magA-negative/k(2)A-negative strains. CONCLUSION: Diabetes mellitus was the most frequent underlying disease among our patients. The rmpA and/or hypermucoviscosity phenotype were the most common virulence factors in K. pneumoniae isolates causing extrahepatic abscesses, among which K2 capsule serotype (k(2)A(+)) was more prevalent than K1 capsule serotype (magA(+)).     

Virulence and plasmid transferability of KPC Klebsiella pneumoniae at the Veterans Affairs Healthcare System of New Jersey

Klebsiella pneumoniae carbapenemase (KPC)-producing Klebsiella pneumoniae infections are associated with high mortality; however, little is known about the virulence determinants of KPC-producing K. pneumoniae. At the Veterans Affairs New Jersey Healthcare System (VA NJHCS), we investigated the virulence and plasmid transferability of 60 clinically unique KPC-containing K. pneumoniae isolates. All 60 isolates were negative for known virulence factors K1, K2, and K5 capsular antigens; rmpA; and the aerobactin gene by polymerase chain reaction. Isolates varied in their susceptibility to neutrophil phagocytosis, but were less resistant than the virulent serotype K1 isolate. Additionally, no deaths were seen on murine lethality studies. Conjugation results of this study showed that the bla(KPC) gene can be transferred into an Escherichia coli J-53 strain but not to E. coli JP-995. However, the stability is very limited as E. coli J-53 does not retain bla(KPC)-containing plasmids for any period of time. The lack of virulence factors in the set of KPC-producing K. pneumoniae studied suggests that morbidity and mortality may be due to detection issues or lack of effective antibiotics.     

Interaction of Klebsiella capsule type 7 with human polymorphonuclear leucocytes.

Klebsiella serotype K7 is found among the capsule types that are most prevalent in respiratory tract isolates. To evaluate the significance of the K7 antigen in bacteria-leucocyte interactions, K7-encapsulated Klebsiella pneumoniae strains and their non-capsulate mutants were investigated. The K7 isolates were compared to K2-capsulate strains and their respective K- derivatives. K7-capsulate bacteria were less hydrophilic, and more readily phagocytosed and killed by human polymorphonuclear leucocytes (PMNL) than K2 strains. Loss of the K7 antigen resulted in increased surface hydrophobicity but did not affect phagocytosis and killing, whereas loss of the K2 capsule caused greater susceptibility to the phagocytic and killing action of PMNL. Both the K7 and K2 antigen stimulated the extracellular release of lysozyme from neutrophils but not of myeloperoxidase, indicating degranulation of only secondary granules. All K- mutants induced the release of both lysozyme and myeloperoxidase. Our results suggest that, in contrast to the K2 antigen, the K7 capsular polysaccharide does not confer antiphagocytic properties on bacteria. However, the K7 antigen is able to impede the extracellular release of primary granule enzymes.     

Familial spread of a virulent clone of Klebsiella pneumoniae causing primary liver abscess

Capsule-forming Klebsiella pneumoniae K1 caused primary liver abscess in two household members of a family. The causative isolates had identical pulsed-field gel electrophoresis patterns and were determined to be sequence type 23. An additional member of the family was found to carry the same strain without clinical manifestation.     

Identification of a major cluster of Klebsiella pneumoniae isolates from patients with liver abscess in Taiwan

Klebsiella pneumoniae has emerged as the leading liver abscess pathogen in Taiwan, with the percentage rising from 30% in the 1980s to over 80% in the 1990s. Most of the patients with K. pneumoniae liver abscess are diabetic and without biliary tract disease. Some patients develop serious extrahepatic complications such as endophthalmitis, meningitis, lung abscess, and necrotizing fasciitis. Pulsed-field gel electrophoresis (PFGE) was used for cluster analysis of 96 isolates from patients with liver abscess and 60 isolates from patients with other diseases. A total of 136 PFGE types were identified. Among the 96 liver abscess-associated isolates, 60 (62.5%) were classified in major cluster A. Cluster A included 41 PFGE types (types 1 to 41) which had a genetic similarity of at least 72.4% +/- 9.4%. The PFGE patterns of cluster A strains are so similar that they could have originated from the same ancestor. This study demonstrates that cluster A plays an important role in the high incidence of K. pneumoniae liver abscess in Taiwan.     

Increase in prevalence of Streptococcus pneumoniae serotype 6C at Eight Children's Hospitals in the United States from 1993 to 2009

Streptococcus pneumoniae serotype 6C, which was described in 2007, causes invasive disease in adults and children. We investigated the prevalence of 6C among pediatric isolates obtained from eight children's hospitals in the United States. S. pneumoniae isolates were identified from a prospective multicenter study (1993 to 2009). Fifty-seven serotype 6C isolates were identified by multiplex PCR and/or Quellung reaction. Five were isolated before 2000, and the prevalence increased over time (P < 0.000001). The median patient age was 2.1 years (range, 0.2 to 22.5 years). Clinical presentations included bacteremia (n = 24), meningitis (n = 7), pneumonia (n = 4), abscess/wound (n = 3), mastoiditis (n = 2), cellulitis (n = 2), peritonitis (n = 1), septic arthritis (n = 1), otitis media (n = 10), and sinusitis (n = 3). By broth microdilution, 43/44 invasive serotype 6C isolates were susceptible to penicillin (median MIC, 0.015 μg/ml; range, 0.008 to 2 μg/ml); all were susceptible to ceftriaxone (median MIC, 0.015 μg/ml; range, 0.008 to 1 μg/ml). By disk diffusion, 16/44 invasive isolates (36%) were nonsusceptible to erythromycin, 19 isolates (43%) were nonsusceptible to trimethoprim-sulfamethoxazole (TMP-SMX), and all isolates were clindamycin susceptible. Multilocus sequence typing (MLST) revealed 24 sequence types (STs); 9 were new to the MLST database. The two main clonal clusters (CCs) were ST473 and single-locus variants (SLVs) (n = 13) and ST1292 and SLVs (n = 23). ST1292 and SLVs had decreased antibiotic susceptibility. Serotype 6C causes disease in children in the United States. Emerging CC1292 expressed TMP-SMX resistance and decreased susceptibility to penicillin and ceftriaxone. Continued surveillance is needed to monitor changes in serotype prevalence and possible emergence of antibiotic resistance in pediatric pneumococcal disease.     

A k2A-positive Klebsiella pneumoniae causes liver and brain abscess in a Saint Kitt's man

Klebsiella pneumoniae isolated in community-acquired pneumonia is increasingly found in primary pyogenic liver abscesses. The presence of magA in K. pneumoniae has been implicated in hypermucoviscosity and virulence of liver abscess isolates. The K2 serotype has also been strongly associated with hypervirulence. We report the isolation of non-magA, K2 K. pneumoniae strain from a liver abscess of a Saint Kitt''s man who survived the invasive syndrome.     

Development of an enzyme-linked immunosorbent assay method for typing and quantitation of Klebsiella pneumoniae lipopolysaccharide: application to serotype O1.

We describe a method for the typing and quantitation of Klebsiella pneumoniae serotype O1 lipopolysaccharide (LPS) based on inhibition in an enzyme-linked immunosorbent assay of a reaction of known O1 LPS antigen and anti-O1 antibody by unknown LPS extracts. Serotype O1 was found in 32% of the 124 K. pneumoniae clinical isolates tested, showing that this serotype is frequent among the eight O serotypes which have been described previously.     

Correlation of the virulence of Klebsiella pneumoniae K1 and K2 with the presence of a plasmid encoding aerobactin.

Nine isolates of Klebsiella pneumoniae belonging to capsular serotypes K1 and K2 were assayed for virulence in mice. Virulent isolates (50% lethal dose of less than 10(3) microorganisms) and avirulent isolates (50% lethal dose of over 10(6) microorganisms) were selected. Supplementation of a defined minimal medium with transferrin markedly reduced the growth of avirulent strains but had no significant effect on the growth of virulent strains. All isolates produced enterochelin, but only production of aerobactin could be correlated with virulence. The genes encoding aerobactin and its receptor protein were located on a 180-kilobase plasmid. They were cloned into the mobilizable vector pSUP202. Homology was demonstrated with the aerobactin operon of the Escherichia coli plasmid pColV-K30. Transfer of the recombinant plasmid pKP4 into an avirulent recipient enhanced virulence by 100-fold. These experiments demonstrated that aerobactin is an essential factor of pathogenicity in K. pneumoniae.     

Emergence of Serotype IV Group B Streptococcus Adult Invasive Disease in Manitoba and Saskatchewan,Canada, Is Driven by Clonal Sequence Type 459 Strains

Serotype IV group B Streptococcus (GBS) is emerging in Canada and the United States with rates as high as 5% of the total burden of adult invasive GBS disease. To understand this emergence, we studied the population structure and assessed the antimicrobial susceptibility of serotype IV isolates causing adult invasive infection in Manitoba and Saskatchewan, Canada, between 2010 and 2014. Whole-genome sequencing was used to determine multilocus sequence typing information and identify genes encoding antimicrobial resistance in 85 invasive serotype IV GBS strains. Antimicrobial susceptibility testing was performed by standard methods. Strain divergence was assessed using genome-wide single-nucleotide polymorphism analysis. Serotype IV strains were responsible for 16.9% of adult invasive GBS infections in Manitoba and Saskatchewan during the period. The majority of serotype IV isolates (89%) were clonally related, tetracycline-, erythromycin-, and clindamycin-resistant sequence type 459 (ST459) strains that possessed genes tetM and ermTR. Genome comparisons between ST459 and serotype V ST1 GBS identified several areas of recombination in an overall similar genomic background. Serotype IV ST459 GBS strains are expanding and causing a substantial percentage of adult invasive GBS disease. This emergence may be linked to the acquisition of resistance to tetracycline, macrolides, and lincosamides.     

Hypermucoviscosity,rmpA, and aerobactin are associated with community-acquired Klebsiella pneumoniae bacteremic isolates causing liver abscess in Singapore

Introduction

This retrospective study investigated the clinical etiology of community-acquired bacteremic Klebsiella pneumoniae infections, and characterized laboratory and genetic markers which may be associated with primary liver abscess (PLA).

Aerobactin production of serotyped Escherichia coli from urinary tract infections

Aerobactin production was examined by a bioassay in 467 Escherichia coli urinary strains from girls. All strains were of known O:K:H serotype. 139, 119 and 112 strains were isolates from pyelonephritis (Py), cystitis (Cy) and asymptomatic bacteriuria (ABU), respectively, and 97 were from fecal samples of healthy girls (FN). The incidence of aerobactin production was significantly higher among Py strains than among ABU and FN strains (P less than 0.001) and also significantly higher than among Cy strains (P less than 0.01). Aerobactin production was associated with serotype, e.g. the majority of O6:K2:H1 strains and of O16:K1:H6 were positive while e.g. the O6:K13:H1 strains were negative. There was no consistent pattern of coappearance of aerobactin and hemolysin.     

Somatic Serogroups, Capsular Types, and Species of Fecal Klebsiella in Patients with Ankylosing Spondylitis

The purpose of the present study was to find out whether patients with ankylosing spondylitis (AS) carry fecal Klebsiella strains that belong to serotypes or species specific for AS. Somatic serotypes (O groups), capsular (K) serotypes, and biochemically identified species were determined for fecal klebsiellae isolated from 187 AS patients and 195 control patients. The controls were patients with fibromyalgia or rheumatoid arthritis. The 638 isolates of Klebsiella that were obtained represented 161 strains; 81 from AS patients and 80 from the controls. The average number of Klebsiella strains per patient was 1.7 for the AS group and 1.5 for the control group. The most common O group was O1, which was observed for isolates from 23 of 187 AS patients and 24 of 195 control patients. Next in frequency was group O2, which was observed for isolates from 17 AS patients and 15 control patients. Regarding the K serotypes, 59 different types were identified, revealing a heterogeneous representation of Klebsiella strains, without a predominance of any serotype. By biochemical identification, Klebsiella pneumoniae was the most frequently occurring species, being found in 45 AS patients and 45 control patients. Next in the frequency was K. oxytoca, which was observed in 26 AS patients and in 29 control patients. K. planticola and K. terrigena occurred in only a minority of patients. Altogether, when analyzed either separately or simultaneously according to O groups, K serotypes, and biochemically identified species, no evidence of the existence of AS-specific Klebsiella strains was obtained. These findings do not indicate participation of Klebsiella in the etiopathogenesis of AS.     

Extended-spectrum beta-lactamase genes of Klebsiella pneumoniae strains in Taiwan: recharacterization of shv-27, shv-41, and tem-116

Klebsiella pneumoniae causing primary liver abscess (PLA) is emerging. This study identified the beta-lactamases genes of K. pneumoniae isolates in Taiwan. The susceptibilities of beta-lactam antibiotics of 30 K. pneumoniae strains associated with primary liver abscess and 30 noninvasive strains were analyzed. The beta-lactamase genes of randomly selected 24 strains from community-acquired infection and 7 extended-spectrum beta-lactamases (ESBL) strains were identified by PCR and DNA sequencing. Protein expression and the ESBL phenotype of beta-lactamase were determined. All 60 strains were ampicillin resistant and cefotaxime susceptible, whereas no strain was ESBL producing. In the 24 selected strains, shv-1a was found in 14, shv-1 in 7; shv-26, shv-27, and shv-41 were detected in one. However, all of these 24 strains had the tem-116 gene. In 7 ESBL-producing K. pneumoniae strains, shv-5a was found in 5, whereas shv-5 and ctx-m-9 group were detected in 1 strain. Two previously reported ESBL genes, shv-27 and tem-116, as well as a suspected ESBL gene, shv-41, were found in non-ESBL-producing strains. Transformation of these genes conferred ampicillin resistance but not the ESBL-producing phenotype in Escherichia coli. Beta-lactamase protein expression of these strains was further confirmed by western blotting. In conclusion, ESBL is rare in community-acquired K. pneumoniae infection and is not associated with PLA in Taiwan. The shv-5a, shv-5, and ctx-m-9 groups are present in ESBL-producing strains in Taiwan, but shv-27, shv-41, and tem-116 are not ESBL genes.     

Capsular polysaccharide synthesis regions in Klebsiella pneumoniae serotype K57 and a new capsular serotype

Community-acquired pyogenic liver abscess caused by Klebsiella pneumoniae is an emerging infectious disease. We explored the capsular polysaccharide synthesis (cps) regions of three non-K1, non-K2 K. pneumoniae strains, A1142, A7754, and A1517, from Taiwanese patients experiencing pyogenic liver abscess. Two of the strains, A1142 and A7754, belonged to capsular serotype K57, while the third belonged to a new capsular serotype, different from the previously reported 77 serotypes. Deletion and complementation experiments suggested that a unique K57 gene, a homologue of wzy, was essential for K57 capsular synthesis and confirmed that this gene cluster was a genetic coding region for K57. Compared to K1 and K2 strains, the three strains were all serum sensitive, suggesting that host factors might also be involved in the three patients. PCR using primers from specific genes for K57 was more sensitive and specific than traditional serotyping. The remaining strain, A1517, did not react to the antisera from any of the 77 serotypes, and none of the 77 reference strains reacted to the serum against this strain. Moreover, PCR analyses using various primer pairs from the serotype-specific open reading frames did not reveal cross-reactivity to any of the 77 reference strains, suggesting that this strain likely represents a new capsular type. We conclude that sequences from these two cps regions are very useful in detecting K57 and the new cps genotype.     

Analysis of group B streptococcal isolates from infants and pregnant women in Portugal revealing two lineages with enhanced invasiveness

The populations of group B streptococcus (GBS) associated with vaginal carriage in pregnant women and invasive neonatal infections in Portugal were compared. GBS isolates were characterized by serotyping, pulsed-field gel electrophoresis (PFGE) profiling, and multilocus sequence typing (MLST). Serotypes III and V accounted for 44% of all colonization isolates (n = 269), whereas serotypes III and Ia amounted to 69% of all invasive isolates (n = 64). Whereas serotype Ia was associated with early-onset disease (EOD), serotype III was associated with late-onset disease (LOD). Characterization by PFGE and MLST identified very diverse populations in carriage and invasive disease. Serotype Ia was represented mainly by a single PFGE cluster defined by sequence type 23 (ST23) and the infrequent ST24. In contrast, serotype III was found in a large number of PFGE clusters and STs, but a single PFGE cluster defined by ST17 was found to be associated with invasive disease. Although serotype III was associated only with LOD, ST17 showed an enhanced capacity to cause both EOD and LOD. Our data reinforce the evidence for enhanced invasiveness of ST17 and identify a lineage expressing serotype Ia capsule and represented by ST23 and ST24 as having enhanced potential to cause EOD.