Dataset Information

Isolation of Four Lytic Phages Infecting Klebsiella pneumoniae K22 Clinical Isolates from Spain.

ABSTRACT: The emergence of multi-drug-resistant bacteria represents a major public-health threat. Phages constitute a promising alternative to chemical antibiotics due to their high host specificity, abundance in nature, and evolvability. However, phage host specificity means that highly diverse bacterial species are particularly difficult to target for phage therapy. This is the case of Klebsiella pneumoniae, which presents a hypervariable extracellular matrix capsule exhibiting dozens of variants. Here, we report four novel phages infecting K. pneumoniae capsular type K22 which were isolated from environmental samples in Valencia, Spain. Full genome sequencing showed that these phages belong to the Podoviridae family and encode putative depolymerases that allow digestion of specific K22 K. pneumoniae capsules. Our results confirm the capsular type-specificity of K. pneumoniae phages, as indicated by their narrow infectivity in a panel of K. pneumoniae clinical isolates. Nonetheless, this work represents a step forward in the characterization of phage diversity, which may culminate in the future use of large panels of phages for typing and/or for combating multi-drug-resistant K. pneumoniae.

SUBMITTER: Domingo-Calap P

PROVIDER: S-EPMC7013548 | biostudies-literature | 2020 Jan

REPOSITORIES: biostudies-literature

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Isolation of Four Lytic Phages Infecting <i>Klebsiella pneumoniae</i> K22 Clinical Isolates from Spain.

Domingo-Calap Pilar P Beamud Beatriz B Vienne Justine J González-Candelas Fernando F Sanjuán Rafael R

International journal of molecular sciences 20200109 2

The emergence of multi-drug-resistant bacteria represents a major public-health threat. Phages constitute a promising alternative to chemical antibiotics due to their high host specificity, abundance in nature, and evolvability. However, phage host specificity means that highly diverse bacterial species are particularly difficult to target for phage therapy. This is the case of <i>Klebsiella pneumoniae</i>, which presents a hypervariable extracellular matrix capsule exhibiting dozens of variants ...[more]

PMID: 31936552

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Project description:The bacterial pathogen Klebsiella pneumoniae causes urinary tract infections in immunocompromised patients. Generally, the overuse of antibiotics contributes to the potential development and the spread of antibiotic resistance. In fact, certain strains of K. pneumoniae are becoming increasingly resistant to antibiotics, making infection by these strains more difficult to treat. The use of bacteriophages to control pathogens may offer a non-antibiotic-based approach to treat multidrug-resistant (MDR) infections. However, a detailed understanding of phage-host interactions is crucial in order to explore the potential success of phage-therapy for treatment. In this study, we investigated the molecular epidemiology of nine carbapenemase-producing K. pneumoniae isolates from a local hospital in Shanghai, China. All strain isolates belong to sequence type 11 (ST11) and harbor the blaKPC-2 gene. The S1-PFGE (S1 nuclease pulsed field gel electrophoresis) pattern of the isolates did not show any relationship to the multilocus sequence typing (MLST) profiles. In addition, we characterized phage 117 and phage 31 and assessed the potential application of phage therapy in treating K. pneumoniae infections in vitro. The results of morphological and genomic analyses suggested that both phages are affiliated to the T7 virus genus of the Podoviridae family. We also explored phage-host interactions during growth in both planktonic cells and biofilms. The phages' heterogeneous lytic capacities against K. pneumoniae strains were demonstrated experimentally. Subsequent culture and urine experiments with phage 117 and host Kp36 initially demonstrated a strong lytic activity of the phages. However, rapid regrowth was observed following the initial lysis which suggests that phage resistant mutants were selected in the host populations. Additionally, a phage cocktail (117 + 31) was prepared and investigated for antimicrobial activity. In Luria Broth (LB) cultures, we observed that the cocktail showed significantly higher antimicrobial activity than phage 117 alone, but this was not observed in urine samples. Together, the results demonstrate the potential therapeutic value of phages in treating K. pneumoniae urinary tract infections.

Project description:BACKGROUND: Members of the genus Klebsiella are among the leading microbial pathogens associated with nosocomial infection. The increased incidence of antimicrobial resistance in these species has propelled the need for alternate/combination therapeutic regimens to aid clinical treatment. Bacteriophage therapy forms one of these alternate strategies. METHODS: Electron microscopy, burst size, host range, sensitivity of phage particles to temperature, chloroform, pH, and restriction digestion of phage DNA were used to characterize Klebsiella phages. RESULTS AND CONCLUSIONS: Of the 32 isolated phages eight belonged to the family Myoviridae, eight to the Siphoviridae whilst the remaining 16 belonged to the Podoviridae. The host range of these phages was characterised against 254 clinical Enterobacteriaceae strains including multidrug resistant Klebsiella isolates producing extended-spectrum beta-lactamases (ESBLs). Based on their lytic potential, six of the phages were further characterised for burst size, physicochemical properties and sensitivity to restriction endonuclease digestion. In addition, five were fully sequenced. Multiple phage-encoded host resistance mechanisms were identified. The Siphoviridae phage genomes (KP16 and KP36) contained low numbers of host restriction sites similar to the strategy found in T7-like phages (KP32). In addition, phage KP36 encoded its own DNA adenine methyltransferase. The ?KMV-like KP34 phage was sensitive to all endonucleases used in this study. Dam methylation of KP34 DNA was detected although this was in the absence of an identifiable phage encoded methyltransferase. The Myoviridae phages KP15 and KP27 both carried Dam and Dcm methyltransferase genes and other anti-restriction mechanisms elucidated in previous studies. No other anti-restriction mechanisms were found, e.g. atypical nucleotides (hmC or glucosyl hmC), although Myoviridae phage KP27 encodes an unknown anti-restriction mechanism that needs further investigation.

Project description:Klebsiella pneumoniae is among the most common causes of hospital-acquired infections and has emerged as an urgent threat to public health due to carbapenem antimicrobial resistance. K. pneumoniae commonly colonizes hospitalized patients and causes extraintestinal infections such as urinary tract infection, bloodstream infection (septicemia), and pneumonia. If colonization is an intermediate step before infection, then detection and characterization of colonizing isolates could enable strategies to prevent or empirically treat K. pneumoniae infections in hospitalized patients. However, the strength of the association between colonization and infection is unclear. To test the hypothesis that hospitalized patients become infected with their colonizing strain, 1,765 patients were screened for rectal colonization with K. pneumoniae, and extraintestinal isolates from these same patients were collected over a 3-month period in a cohort study design. The overall colonization prevalence was 23.0%. After adjustment for other patient factors, colonization was significantly associated with subsequent infection: 21 of 406 (5.2%) colonized patients later had extraintestinal infection, compared to 18 of 1,359 (1.3%) noncolonized patients (adjusted odds ratio [OR], 4.01; 95% confidence interval, 2.08 to 7.73; P < 0.001). Despite a high diversity of colonizing isolates, 7/7 respiratory, 4/4 urinary, and 2/5 bloodstream isolates from colonized patients matched the patient corresponding rectal swab isolates, based on wzi capsular typing, multilocus sequence typing (MLST), and whole-genome sequence analysis. These results suggest that K. pneumoniae colonization is directly associated with progression to extraintestinal infection. IMPORTANCE K. pneumoniae commonly infects hospitalized patients, and these infections are increasingly resistant to carbapenems, the antibiotics of last resort for life-threatening bacterial infections. To prevent and treat these infections, we must better understand how K. pneumoniae causes disease and discover new ways to predict and detect infections. This study demonstrates that colonization with K. pneumoniae in the intestinal tract is strongly linked to subsequent infection. This finding helps to identify a potential time frame and possible approach for intervention: the colonizing strain from a patient could be isolated as part of a risk assessment, and antibiotic susceptibility testing could guide empirical therapy if the patient becomes acutely ill.

Dataset Information

Isolation of Four Lytic Phages Infecting Klebsiella pneumoniae K22 Clinical Isolates from Spain.

Publications

Isolation of Four Lytic Phages Infecting <i>Klebsiella pneumoniae</i> K22 Clinical Isolates from Spain.

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