Bacterial pathogens have a broad arsenal of genes that are tightly regulated and coordinated to facilitate adaptation to alter host inflammatory response and prolong intracellular bacterial survival. Salmonella enterica serovar Typhimurium utilizes a type III secretion system (T3SS) to deliver effector molecules into host cells and regulate signal transduction pathways such as NF-κB, thereby resulting in salmonellosis. SpvB, a pSLT-encoded cytotoxic protein secreted by Salmonella pathogenicity island-2 T3SS, is associated with enhanced Salmonella survival and intracellular replication. In this report, we characterized the effects of SpvB on NF-κB signaling pathway. We showed that SpvB has a potent and specific ability to prevent NF-κB activation by targeting IκB kinase β (IKKβ). Previous studies from our laboratory showed that SpvB decreases Nrf2 through its C-terminal domain. Here we further demonstrated that KEAP1, a cytoplasmic protein that interacts with Nrf2 and mediates its proteasomal degradation, is involved in SpvB-induced downregulation of IKKβ expression and phosphorylation. Reduction of KEAP1 by small-interfering RNA prevented the suppression of IKKβ and its phosphorylation mediated by SpvB. These findings revealed a novel mechanism by which Salmonella modulates NF-κB activity to ultimately facilitate intracellular bacterial survival and proliferation and delay host immune response to establish infection.

Escherichia coli (E. coli) bloodstream infections (BSIs) are among the most predominant causes of death in infants and children worldwide. NDM-5 (New Delhi Metallo-lactamase-5) is responsible for one of the main mechanisms of carbapenem resistance in E. coli. To analyze the phenotypic and genomic characteristics of NDM-5-producing E. coli from bloodstream infections (BSIs), a total of 114 E. coli strains was collected from a children’s hospital in Jiangsu province, China. Eight bla_NDM-5-carrying E. coli strains were identified which were all carbapenem-resistant and carried diverse antimicrobial resistance genes apart from bla_NDM-5. They belonged to six distinct sequence types (STs) and serotypes including one each for ST38/O7:H8, ST58/O?:H37, ST131/O25:H4, ST156/O11:H25 and ST361/O9:H30 and three strains are originating from a single clone belonging to ST410/O?:H9. Apart from bla_NDM-5, the E. coli strains isolated from BSIs also carried other β-lactamase genes, including bla_CMY-2 (n = 4), bla_CTX-M-14 (n = 2), bla_CTX-M-15 (n = 3), bla_CTX-M-65 (n = 1), bla_OXA-1 (n = 4) and bla_TEM-1B (n = 5). The bla_NDM-5 genes were located on three different types of plasmids, which were IncFII/I1 (n = 1), IncX3 (n = 4) and IncFIA/FIB/FII/Q1 (n = 3). The former two types were conjugatively transferable at frequencies of 10⁻³ and 10⁻⁶, respectively. The dissemination of NDM-producing strains, which exhibit resistance to the last-line antibiotics, carbapenems, may increase the muti-antimicrobial resistance burden among E. coli BSIs and further threaten public health.

Hypervirulent Klebsiella pneumoniae (hvKP) is increasingly reported worldwide as a major clinical and public health threat. The virulence of hvKP is attributed largely to the carriage of virulence plasmids (KpVPs). To date, two dominant types of KpVP have been identified, namely, KpVP-1 and KpVP-2. In this study, we reported two hvKP strains from bloodstream infections that carry highly identical virulence plasmids that exhibited <40% coverage compared with KpVP-1 and KpVP-2. This novel virulence plasmid was designated KpVP-3. The two hvKP have different genetic backgrounds, which belonged to ST29-K54 and ST111-K63, respectively. They were both positive for the string test, highly virulent on the Galleria mellonella infection model, and possess high-level macrophage-killing resistance in vitro . Apart from the intrinsic non-susceptibility to ampicillin, both strains were susceptible to commonly used antibiotics. The virulence plasmid carried virulence genes rmpADC , iroBCDN ( iro1 ), and the ybt locus ( ybt4 ) which was not present on either KpVP-1 or KpVP-2. It did not carry antimicrobial resistance genes but carried an incomplete conjugation machinery containing only the traH and traF genes. The KpVP-3 plasmid was stably maintained in both hvKP strains and could not be eliminated with SDS treatment or by serial passage on stress-free agar plates. KpVP-3 was non-self-transmissible under experimental conditions. Data mining suggested KpVP-3-type plasmids have emerged in different countries including China, Australia, and the USA. The emergence of this novel virulence plasmid might pose a potential threat to public health. Heightened efforts are required to study its dissemination mechanism.

Colistin is one of the last-resort antibiotics in treating infections caused by multidrug-resistant (MDR) pathogens. Unfortunately, the emergence of colistin-resistant gram-negative strains limit its clinical application. Here, we identify an FDA-approved drug, valnemulin (Val), exhibit a synergistic effect with colistin in eradicating both colistin-resistant and colistin-susceptible gram-negative pathogens both in vitro and in the mouse infection model. Furthermore, Val acts synergistically with colistin in eliminating intracellular bacteria in vitro. Functional studies and transcriptional analysis confirm that the combinational use of Val and colistin could cause membrane permeabilization, proton motive force dissipation, reduction in intracellular ATP level, and suppression in bacterial motility, which result in bacterial membrane disruption and finally cell death. Our findings reveal the potential of Val as a colistin adjuvant to combat MDR bacterial pathogens and treat recalcitrant infections.

Programmed cell death (PCD) is tightly orchestrated by molecularly defined executors and signaling pathways. NLRP6, a member of cytoplasmic pattern recognition receptors, has a multifaceted role in host resistance to bacterial infection. However, whether and how NLRP6 may contribute to regulate host PCD during Gram-negative bacterial infection remain to be illuminated. Here, we report that NLRP6 promotes RIP1 kinase-mediated necroptosis, a form of lytic PCD, in both an in vitro and in vivo model of Salmonella typhimurium infection. By downregulating TAK1-mediated p38^MAPK/MK2 phosphorylation, NLRP6 decreased RIP1 phosphorylation at residue S321 and subsequently increased RIP1 kinase-dependent MLKL phosphorylation. Suppression of p38^MAPK/MK2 cascade not only reduced the number of dead cells caused by NLRP6 but also decreased the systemic dissemination of S. typhimurium resulting from NLRP6. Taken together, our findings provide new insights into the role and regulatory mechanism of NLRP6-associated antimicrobial responses by revealing a function for NLRP6 in regulating necroptosis.

Objective: The aim of the present study is to explore the impact of the tet (A) type I variant (tetA-v1) tet A-v1) on its fitness effect in Klebsiella pneumoniae. . Methods: Clinical K. pneumoniae strains were utilized as parental strains to generate strains carrying only the plasmid vector (pBBR1MCS-5) or the tet A-v1 recombinant plasmid (ptetA-v1). tet A-v1). Antimicrobial susceptibility testing was conducted to estimate the contribution of tet A-v1 to drug resistance. Plasmid stability was evaluated by serial passage over 10 consecutive days in the absence of tigecycline. Biological fitness was examined through growth curve analysis, in vitro competition assays and a neutropenic mouse thigh infection model. Results: A 2-4-fold increase in tigecycline MIC was observed following the acquisition of tet A-v1. Without tigecycline treatment, the stability of ptetA-v1 tet A-v1 plasmids has been decreasing since day 1. The ptetA-v1 tet A-v1 plasmid in Kp89, Kp91, and Kp93 exhibited a decrease of about 20% compared to the pBBR1MCS-5 plasmid. The acquisition of the tet A-v1 gene could inhibit the growth ability of K. pneumoniae strains both in vitro and in vivo. tet A-v1 gene imposed a fitness cost in K. pneumoniae, , particularly in the CRKP strain Kp51, with a W value of approximately 0.56. Conclusion: The presence of tet A-v1 is associated with a significant fitness cost in K. pneumoniae in the absence of tigecycline, both in vitro and in vivo . (c) 2024 The Authors. Published by Elsevier Ltd on behalf of International Society for Antimicrobial Chemotherapy. This is an open access article under the CC BY-NC-ND license ( http://creativecommons.org/licenses/by-nc-nd/4.0/ )