Host immunity involvement in the outcome of phage therapy against hypervirulent Klebsiella pneumoniae infections.

Highly encapsulated hypervirulent Klebsiella pneumoniae (hvKp) causes severe infections. Bacteriophage therapy, an antibiotic alternative, effectively treats bacterial infections. Phage φFK1979 encoding polysaccharide depolymerases can target and disarm the capsule of hvKp FK1979, showing promise against FK1979 infection. Resistant strains induced by φFK1979 are possibly eliminated by host immunity and new phage phiR3 targeting them. We constructed varied immunocompromised FK1979 infection mouse models to assess the therapy efficacy of φFK1979 alone or in combination with phiR3. Survival rates, bacterial loads, histopathology, inflammation, and immune cell distribution of mice were studied. Prompt and adequate administration of φFK1979, rather than phiR3, significantly improved survival rates in mice with different immune statuses. However, immunocompromised mice showed lower efficacy due to reduced tolerance to low-virulence φFK1979-resistant bacteria compared to immunocompetent mice. Adding phiR3 sequentially greatly enhanced therapy efficacy for them, leading to increased survival rates and notable improvements in pathology and inflammation. Immunocompetent mice exhibited the most favorable response to φFK1979 monotherapy, as their immune system cleared φFK1979-resistant bacteria while avoiding a robust response to phiR3 combating φFK1979-resistant bacteria. This study revealed host immunity involvement in the outcome of phage therapy against infections and introduced, for the first time, personalized phage therapy strategies for hvKp-infected mice with varying immune statuses.IMPORTANCEHypervirulent Klebsiella pneumoniae (hvKp), with high capsular polysaccharide production, can cause severe invasive infections. Capsule-targeting phage poses the potential to fight against hvKp. We previously elucidated that the capsule-targeting phage induces resistance in hvKp, while phage-resistant strains exhibit sensitivity to host innate immunity and new phages targeting them. This indicated that phage-resistant strains can be eliminated by the immune system in immunocompetent patients, whereas they may require treatment with phages targeting resistant bacteria in immunocompromised patients. HvKp can infect individuals with varying immune statuses, including both immunocompetent and immunocompromised/deficient patients. This study, for the first time, developed personalized phage therapy strategies for hvKp-infected mice with different immune statuses, optimizing phage therapy against hvKp infections. This research is expected to provide a theoretical foundation and novel insights for clinical phage therapy against hvKp infections, offering significant societal benefits and clinical value.

Azomycin Orchestrate Colistin-Resistant Enterobacter cloacae Complex's Colistin Resistance Reversal In Vitro and In Vivo.

The Enterobacter cloacae complex (ECC) is a group of nosocomial pathogens that pose a challenge in clinical treatment due to its intrinsic resistance and the ability to rapidly acquire resistance. Colistin was reconsidered as a last-resort antibiotic for combating multidrug-resistant ECC. However, the persistent emergence of colistin-resistant (COL-R) pathogens impedes its clinical efficacy, and novel treatment options are urgently needed. We propose that azomycin, in combination with colistin, restores the susceptibility of COL-R ECC to colistin in vivo and in vitro. Results from the checkerboard susceptibility, time-killing, and live/dead bacterial cell viability tests showed strong synergistic antibacterial activity in vitro. Animal infection models suggested that azomycin-colistin enhanced the survival rate of infected Galleria mellonella and reduced the bacterial load in the thighs of infected mice, highlighting its superior in vivo synergistic antibacterial activity. Crystal violet staining and scanning electron microscopy unveiled the in vitro synergistic antibiofilm effects of azomycin-colistin. The safety of azomycin and azomycin-colistin at experimental concentrations was confirmed through cytotoxicity tests and an erythrocyte hemolysis test. Azomycin-colistin stimulated the production of reactive oxygen species in COL-R ECC and inhibited the PhoPQ two-component system to combat bacterial growth. Thus, azomycin is feasible as a colistin adjuvant against COL-R ECC infection.

Emergence of tet(X2) in Acinetobacter pittii confers clinical resistance to tigecycline.

OBJECTIVES: To characterize a novel transposon Tn7533 carrying the tet(X2) gene in a tigecycline-resistant Acinetobacter pittii BM4623 of clinical origin. METHODS: Gene knockout and in vitro cloning were used to verify the function of tet(X2). WGS and comparative genomic analysis were used to explore the genetic characteristics and molecular evolution of tet(X2). Inverse PCR and electroporation experiments were used to evaluate the excision and integration capabilities of Tn7533. RESULTS: A. pittii BM4623 belonged to a novel ST, ST2232 (Pasteur scheme). Knockout of tet(X2) in BM4623 restored its susceptibility to tigecycline. Cloning of the tet(X2) gene into Escherichia coli DH5α and Acinetobacter baumannii ATCC 17978 resulted in 16-fold or more increases in MICs of tigecycline. Sequence analysis showed that the region upstream of tet(X2) exhibited a high degree of diversity, while there was a 145 bp conserved region downstream of tet(X2). tet(X2) in BM4623 was located on a novel composite transposon Tn7533, which also contains multiple resistance genes including blaOXA-58. Tn7533 could be excised from the chromosome to form a circular intermediate and transferred into A. baumannii ATCC 17978 by electroporation. CONCLUSIONS: Our study demonstrates that tet(X2) is a determinant conferring clinical resistance to tigecycline in Acinetobacter species. The emergence of Tn7533 may lead to the potential dissemination of tigecycline and carbapenem resistance in Acinetobacter, which requires continuous monitoring.