Search

ABSTRACT Salmonella enterica is an important zoonotic intracellular bacterial pathogen that is capable of causing infections ranging from localized gastroenteritis to fatal systemic infection in humans and food-producing animals. The increasing antibiotic resistance in Salmonella isolates, especially the emergence of MDR and newer XDR strains, has compromised the efficacy of conventional antimicrobial therapy for Salmonella infections. Hence, it is desirable to develop alternative therapeutic means to tackle the antimicrobial resistance crisis. In this study, we screened plant-derived compounds to identify inhibitors of Salmonella invasion of host cells. These efforts identified fisetin as a possible protector against infection. Further mechanistic studies revealed that fisetin suppressed the function of type III secretion system 1 (T3SS-1), the virulence determinant critical for Salmonella invasion. Fisetin appears to interfere with the interaction between HilD and its relevant promoters, thereby decreasing the transcription of hilA, the central transcriptional regulator that functions to activate the expression of T3SS-1 effector proteins and structural elements. In addition, administration of fisetin in the Salmonella murine infection model resulted in reduced bacterial colonization, alleviation of histopathological destruction, and decreased proinflammatory cytokine levels. Taken together, our study establishes that the natural compound fisetin can be used as a lead compound for the development of anti-Salmonella drugs targeting T3SS-1. IMPORTANCE Salmonella spp. remains a major worldwide health concern that causes significant morbidity and mortality in both humans and animals. The spread of antimicrobial resistant strains has declined the efficacy of conventional chemotherapy. Thus, novel anti-infection drugs or strategies are needed. Anti-virulence strategy represents one of the promising means for the treatment of bacterial infections. In this study, we found that the natural compound fisetin could inhibit Salmonella invasion of host cells by targeting SPI-1 regulation. Fisetin treatment impaired the interaction of the regulatory protein HilD with the promoters of its target genes, thereby suppressing the expression of T3SS-1 effectors as well as structural proteins. Moreover, fisetin treatment could reduce pathology in the Salmonella murine infection model. Collectively, our results suggest that fisetin may serve as a promising lead compound for the development of anti-Salmonella drugs.

ABSTRACT Hypervirulent Klebsiella pneumoniae with capsular polysaccharides (CPSs) causes severe nosocomial- and community-acquired infections. Phage-derived depolymerases can degrade CPSs from K. pneumoniae to attenuate bacterial virulence, but their antimicrobial mechanisms and clinical potential are not well understood. In the present study, Klebsiella phage GH-K3-derived depolymerase Depo32 (encoded by gene gp32) was identified to exhibit high efficiency in specifically degrading the CPSs of K2 serotype K. pneumoniae. The cryo-electron microscopy structure of trimeric Depo32 at a resolution up to 2.32 Å revealed potential catalytic centers in the cleft of each of the two adjacent subunits. K. pneumoniae subjected to Depo32 became more sensitive to phagocytosis by RAW264.7 cells and activated the cells by the mitogen-activated protein kinase signaling pathway. In addition, intranasal inoculation with Depo32 (a single dose of 200 µg, 20 µg daily for 3 days, or in combination with gentamicin) rescued all C57BL/6J mice infected with a lethal dose of K. pneumoniae K7 without interference from its neutralizing antibody. In summary, this work elaborates on the mechanism by which Depo32 targets the degradation of K2 serotype CPSs and its potential as an antivirulence agent. IMPORTANCE Depolymerases specific to more than 20 serotypes of Klebsiella spp. have been identified, but most studies only evaluated the single-dose treatment of depolymerases with relatively simple clinical evaluation indices and did not reveal the anti-infection mechanism of these depolymerases in depth. On the basis of determining the biological characteristics, the structure of Depo32 was analyzed by cryo-electron microscopy, and the potential active center was further identified. In addition, the effects of Depo32 on macrophage phagocytosis, signaling pathway activation, and serum killing were revealed, and the efficacy of the depolymerase (single treatment, multiple treatments, or in combination with gentamicin) against acute pneumonia caused by Klebsiella pneumoniae was evaluated. Moreover, the roles of the active sites of Depo32 were also elucidated in the in vitro and in vivo studies. Therefore, through structural biology, cell biology, and in vivo experiments, this study demonstrated the mechanism by which Depo32 targets K2 serotype K. pneumoniae infection.

ABSTRACT Mobile colistin resistance (mcr) genes mediated by plasmids have widely disseminated throughout the world. Recently, 10 mcr genes (mcr-1 to mcr-10) and a large number of variants have been identified in more than 60 countries. However, only a few instances of Enterobacter cloacae complex (ECC) bearing mcr-10 from animal origin have been reported globally. The aim of this study was to fill a knowledge gap in mcr-10-positive ECC of animal origin and analyze the potential transmission trend and different characteristics between human and companion animal isolates. The mcr-10 gene was identified on a self-transmissible plasmid in the human isolate and non-transmissible plasmids in other three animal strains. mcr-10 was adjacent to a XerC-type tyrosine recombinase-gene, and various insertion sequences were located on the downstream of core conservative structure xerC-mcr-10, thus indicating this region might be a candidate for insertions of mobile genetic elements and mcr-10 might be mobilized by IS-mediated mechanisms. Moreover, phylogenetic analysis found that mcr-10-positive isolates were mainly distributed in the clade of Enterobacter roggenkampii, exhibiting significant species specificity. These findings indicated that mcr-10 has emerged among Enterobacter spp. within humans and companion animals, highlighting that the importance of taking effective control measures to monitor the dissemination and evolution of mcr genes. IMPORTANCE Colistin was considered as the last-resort drug against severe clinical infections caused by multidrug-resistant Gram-negative pathogens. Mobile colistin resistance (mcr) genes and its variants carried by plasmids have been reported in diverse niches in recent years, and yet few studies reported carriage of mcr-10 in ECC strains of companion animal origin. How plasmid-borne mcr-10 transmitted in opportunistic pathogens and different characteristics of mcr-10-bearing strains isolated from humans and companion animals are not well understood. In this study, we discovered mcr-10-harboring strains in multidrug-resistant ECC isolates of companion animal origin for the first time and conducted a comprehensive analysis of the genetic environment of mcr-10 from multiple countries around the world, providing the potential basis for formulating control measures to slow down the spread of colistin resistance.