Predicting Antibiotic Tolerance in hvKP and cKP Respiratory Infections Through Biofilm Formation Analysis and Its Resistance Implications.

Introduction: Respiratory infections are a major global health concern, with Klebsiella pneumoniae standing out due to its evolving antibiotic resistance. This study compares the resistance profiles of hypervirulent Klebsiella pneumoniae (hvKP) and classical Klebsiella pneumoniae (cKP), aiming to shed light on their clinical implications. Methods: We analyzed 86 cases, comprising 42 hvKP and 44 cKP strains, using comprehensive antimicrobial susceptibility testing and clinical data evaluation to assess antibiotic tolerance and resistance mechanisms. Results: Our findings reveal distinct resistance patterns between hvKP and cKP, highlighting the role of chromosomal mutations and plasmid-mediated gene transfer in conferring antibiotic resistance. Notably, hvKP strains exhibited unique resistance trends, including the production of extended-spectrum ß-lactamases (ESBLs) and carbapenemases, differing from those of cKP. Discussion: This research underscores the importance of continuous surveillance and the development of targeted therapies against antibiotic-resistant Klebsiella pneumoniae. It emphasizes the critical need for judicious antibiotic use and novel therapeutic approaches to combat respiratory infections caused by these increasingly resistant pathogens.

Combination effects of baicalin with linezolid against Staphylococcus aureus biofilm-related infections: in vivo animal model.

Staphylococcus aureus is a gram-positive bacterium that can produce biofilm, and biofilm-associated infections are difficult to control. Biofilm prevents antibiotics from penetrating and killing the bacteria. Combined use of antimicrobials is a common strategy to treat S. aureus biofilm-related infections. In this in vivo study, the clinically isolated strain of S. aureus 17546 (t037) was selected to establish a biofilm-associated infection rat model, and baicalin and linezolid were used to treat the infection. CFU counting was used to determine the bacteria within the biofilm, the biofilm structure was viewed using scanning electron microscopy (SEM), histopathology was performed, and inflammatory factors were analyzed by ELISA. Baicalin was efficient in destroying the biofilm and exerted a synergistic bactericidal effect when combined with linezolid. Based on these findings, baicalin combined with linezolid may be efficacious in controlling S. aureus biofilm-related infections.

Synergistic Effects of Baicalin and Levofloxacin Against Hypervirulent Klebsiella pneumoniae Biofilm In Vitro.

Hypervirulent Klebsiella pneumoniae (hvKp) strains that form biofilms have recently emerged worldwide; however, the mechanisms underlying biofilm formation and disruption remain elusive. In this study, we established a hvKp biofilm model, investigated its in vitro formation pattern, and determined the mechanism of biofilm destruction by baicalin (BA) and levofloxacin (LEV). Our results revealed that hvKp exhibited a strong biofilm-forming ability, forming early and mature biofilms after 3 and 5 d, respectively. Early biofilm and bacterial burden were significantly reduced by BA + LEV and EM + LEV treatments, which destroyed the 3D structure of early biofilms. Conversely, these treatments were less effective against mature biofilm. The expression of both AcrA and wbbM was significantly downregulated in the BA + LEV group. These findings indicated that BA + LEV might inhibit the formation of hvKp biofilm by altering the expression of genes regulating efflux pumps and lipopolysaccharide biosynthesis.

Antibacterial synergy between linezolid and baicalein against methicillin-resistant Staphylococcus aureus biofilm in vivo.

Methicillin-resistant Staphylococcus aureus (MRSA) can form biofilms, which prevents the penetration of antibiotics, decreasing their efficacy. This study investigated whether baicalein has synergistic antibacterial effects with linezolid in vivo. We cultivated MRSA 17546 biofilms on silicone implants and inserted them into the air pouches of rat models. The rats were treated with linezolid, baicalein, or a combination therapy for three consecutive days. All treatments reduced the number of colony-forming units (CFU) in the biofilms compared to the control (p < 0.05). However, by day two, the CFU counts were significantly lower in the combination group than in the individual treatment groups (p < 0.05). Histological analysis of the air pouches showed that the severity of the inflammatory cell infiltration was severe in the combination therapy group. In the combination group, the biofilm structure on the implant's surface was sparse and more free colonies could be seen by scanning electron microscopy (SEM); by day three, no obvious biofilm was observed. The serum levels of Staphylococcus enterotoxin A (SEA), C-reactive protein (CRP), and procalcitonin (PCT) were the lowest in the group where rats were treated with the combination of baicalein and linezolid (p < 0.05) compared to other groups. The results suggest that baicalein may inhibit the accessory gene regulator system, reducing the expression of SEA, thus lowering CRP and PCT levels. Furthermore, the inhibitory effect was more pronounced when baicalein was combined with linezolid. These results provide an important basis for the development of a new combination regimen to treat patients with biofilm-associated MRSA infections.

High mobility group protein B1 (HMGB1) interacts with receptor for advanced glycation end products (RAGE) to promote airway smooth muscle cell proliferation through ERK and NF-κB pathways.

BACKGROUND: High-mobility graoup box protein 1 (HMGB1) has been shown to mediate a wide range of pathologic responses by interacting with RAGE (receptor for advanced glycation endproducts) and TLRs (Toll-like receptors). Our previous study showed that HMGB1 has been involved in pathogenesis of airway remodeling in an allergen-induced chronic mice asthma model. Increased airway smooth muscle (ASM) mass is a vital feature of airway remodeling. OBJECTIVE: To evaluate the effect of HMGB1 on proliferation of ASMs and the underlying mechanisms. METHODS: Rat airway smooth muscle cells (RASMs) were obtained by primary explant techniques. We investigated the effect of HMGB1 on the proliferation of RASMs. To identify which receptors and signaling pathways be involved in proliferation of RASMs, we performed western blot and CCK-8 assay by specific receptor blockade and inhibition of MAPK (p38, JNK and ERK) and NF-κB signaling pathways. RESULTS: HMGB1 stimulated RASMs proliferation in a dose- and time-dependent manner and also increased proliferating cell nuclear antigen (PCNA) and RAGE expression of RASMs. The inhibitor of RAGE, but not TLR2 and TLR4, reversed HMGB1-induced RASM proliferation and PCNA expression. Incubation of RASMs with HMGB1 caused a rapid increase in P65 and ERK phosphorylation. RASM proliferation and PCNA expression toward HMGB1 were significantly inhibited by the inhibitors of ERK and NF-κB. CONCLUSION: HMGB1 induces proliferation of RASMs through a RAGE-dependent activation of ERK and NF-κB signaling pathways.

Generation of Methicillin-Resistant Staphylococcus Aureus Biofilm Infection in an Immunosuppressed Rat Model.

BACKGROUND Methicillin-resistant Staphylococcus aureus (MRSA) is a common pathogen responsible for many related infections, and immunosuppressed individuals are more susceptible. Its pathogenicity is associated with its virulence factors, resistance to antibiotics, and ability to form biofilm (BF). MRSA-BF infections in immunosuppressed patients pose great difficulties to clinical treatment. MATERIAL AND METHODS The study aimed to establish a model of MRSA-BF infection in rats with cyclophosphamide (CTX)-induced immunosuppression. For this, rats were administered CTX on days 1 and 4. White blood cells (WBC) were counted, then rats were inoculated with a clinical MRSA 17546 (t037) on day 5. Rats were sacrificed on days 6-10 and tissue samples were examined by scanning electron microscopy. RESULTS Using the dose of CTX: 150 (mg/kg) + 100 (mg/kg) is better than the other 2 programs as the survival rates of the immunocompromised rats were higher than in the other 2 immunosuppressive groups. The survival rate was not different between rats in the clean environment and in the SPF environment. However, the survival rate was affected by the sample acquisitions. Importantly, WBC counts started to decline on day 4, and then started to rise on day 9. Moreover, MRSA-BFs were formed earlier in immunosuppressed rats compared to the normal rats, as shown by scanning electron microscopy. CONCLUSIONS The study successfully established an immunosuppressed rat model of MRSA-BF infection, which provides methodological and data support for establishment of such animal models and is useful reference for related research. Our results may help further investigation of MRSA-BF infection.

Baicalin inhibits biofilm formation, attenuates the quorum sensing-controlled virulence and enhances Pseudomonas aeruginosa clearance in a mouse peritoneal implant infection model.

The quorum sensing (QS) circuit plays a role in the precise regulation of genes controlling virulence factors and biofilm formation in Pseudomonas aeruginosa. QS-controlled biofilm formation by Pseudomonas aeruginosa in clinical settings has remained controversial due to emerging drug resistance; therefore, screening diverse compounds for anti-biofilm or anti-QS activities is important. This study demonstrates the ability of sub-minimum inhibitory concentrations (sub-MICs) of baicalin, an active natural compound extracted from the traditional Chinese medicinal Scutellaria baicalensis, to inhibit the formation of Pseudomonas aeruginosa biofilms and enhance the bactericidal effects of various conventional antibiotics in vitro. In addition, baicalin exerted dose-dependent inhibitory effects on virulence phenotypes (LasA protease, LasB elastase, pyocyanin, rhamnolipid, motilities and exotoxin A) regulated by QS in Pseudomonas aeruginosa. Moreover, the expression levels of QS-regulatory genes, including lasI, lasR, rhlI, rhlR, pqsR and pqsA, were repressed after sub-MIC baicalin treatment, resulting in significant decreases in the QS signaling molecules 3-oxo-C12-HSL and C4-HSL, confirming the ability of baicalin-mediated QS inhibition to alter gene and protein expression. In vivo experiments indicated that baicalin treatment reduces Pseudomonas aeruginosa pathogenicity in Caenorhabditis elegans. Greater worm survival in the baicalin-treated group manifested as an increase in the LT50 from 24 to 96 h. In a mouse peritoneal implant infection model, baicalin treatment enhanced the clearance of Pseudomonas aeruginosa from the implants of mice infected with Pseudomonas aeruginosa compared with the control group. Moreover, the combination of baicalin and antibiotics significantly reduced the numbers of colony-forming units in the implants to a significantly greater degree than antibiotic treatment alone. Pathological and histological analyses revealed mitigation of the inflammatory response and reduced cell infiltration in the peritoneal tissue surrounding the implants after baicalin treatment. Measurement of the cytokine levels in the peritoneal lavage fluid of mice in the baicalin treatment group revealed a decrease in IL-4, an increase in interferon γ (IFN-γ), and a reversed IFN-γ/IL-4 ratio compared with the control group, indicating that baicalin treatment activated the Th1-induced immune response to expedite bacterial load clearance. Based on these results, baicalin might be a potent QS inhibitor and anti-biofilm agent for combating Pseudomonas aeruginosa biofilm-related infections.

Baicalein Inhibits Staphylococcus aureus Biofilm Formation and the Quorum Sensing System In Vitro.

Biofilm formed by Staphylococcus aureus significantly enhances antibiotic resistance by inhibiting the penetration of antibiotics, resulting in an increasingly serious situation. This study aimed to assess whether baicalein can prevent Staphylococcus aureus biofilm formation and whether it may have synergistic bactericidal effects with antibiotics in vitro. To do this, we used a clinically isolated strain of Staphylococcus aureus 17546 (t037) for biofilm formation. Virulence factors were detected following treatment with baicalein, and the molecular mechanism of its antibiofilm activity was studied. Plate counting, crystal violet staining, and fluorescence microscopy revealed that 32 µg/mL and 64 µg/mL baicalein clearly inhibited 3- and 7-day biofilm formation in vitro. Moreover, colony forming unit count, confocal laser scanning microscopy, and scanning electron microscopy showed that vancomycin (VCM) and baicalein generally enhanced destruction of biofilms, while VCM alone did not. Western blotting and real-time quantitative polymerase chain reaction analyses (RTQ-PCR) confirmed that baicalein treatment reduced staphylococcal enterotoxin A (SEA) and α-hemolysin (hla) levels. Most strikingly, real-time qualitative polymerase chain reaction data demonstrated that 32 µg/mL and 64 µg/mL baicalein downregulated the quorum-sensing system regulators agrA, RNAIII, and sarA, and gene expression of ica, but 16 µg/mL baicalein had no effect. In summary, baicalein inhibited Staphylococcus aureus biofilm formation, destroyed biofilms, increased the permeability of vancomycin, reduced the production of staphylococcal enterotoxin A and α-hemolysin, and inhibited the quorum sensing system. These results support baicalein as a novel drug candidate and an effective treatment strategy for Staphylococcus aureus biofilm-associated infections.

Diagnosis of pneumocystis pneumonia using serum (1-3)-ß-D-Glucan: a bivariate meta-analysis and systematic review.

BACKGROUND: The (1-3)-ß-D-Glucan (BG) assay has been approved for making a diagnosis of invasive fungal disease. However, the role of serum-BG assay for the diagnosis of pneumocystis pneumonia (PCP) is controversial, especially between patients with human immunodeficiency virus (HIV) and non-HIV. We conducted a meta-analysis to determine the difference of the overall accuracy of serum-BG assay for the diagnosis of PCP in immunocompromised patients with and without HIV. METHODS: After a systematic review of English-language studies and manual researching, sensitivity (Se), specificity (Sp), and other measures of accuracy of serum-BG for the diagnosis of PCP were pooled using random-effects models for bivariate meta-analysis. Summary receiver operating characteristic (SROC) curve was used to summarize overall test performance. Subgroup analyses were performed to explore the heterogeneity in Se and Sp. RESULTS: Thirteen studies met our inclusion criteria. The summary estimates for serum-BG assay for definite PCP were as follows: Se, 0.91 [95% confidence interval (CI), 0.88-0.93]; Sp, 0.75 (95% CI, 0.68-0.81). As for the patients with and without HIV, the Se and Sp were 0.92 and 0.78, 0.85 and 0.73, respectively. Significant heterogeneity between Se was presented (P=0.04). CONCLUSIONS: Contrary to the results of the previous meta-analysis, a negative result of serum-BG determination is sufficient for ruling out PCP only in HIV cases. For non-HIV patients, the results should be interpreted in parallel with clinical and radiological findings. Besides, further prospective studies with larger sample size are needed to confirm the diagnosis strategy of BG detection.