Amelioration of Maternal Immune Activation-Induced Autism Relevant Behaviors by Gut Commensal Parabacteroides goldsteinii.

Autism spectrum disorder (ASD) is characterized by cognitive inflexibility and social deficits. Probiotics have been demonstrated to play a promising role in managing the severity of ASD. However, there are no effective probiotics for clinical use. Identifying new probiotic strains for ameliorating ASD is therefore essential. Using the maternal immune activation (MIA)-based offspring ASD-like mouse model, a probiotic-based intervention strategy was examined in female mice. The gut commensal microbe Parabacteroides goldsteinii MTS01, which was previously demonstrated to exert multiple beneficial effects on chronic inflammation-related-diseases, was evaluated. Prenatal lipopolysaccharide (LPS) exposure induced leaky gut-related inflammatory phenotypes in the colon, increased LPS activity in sera, and induced autistic-like behaviors in offspring mice. By contrast, P. goldsteinii MTS01 treatment significantly reduced intestinal and systemic inflammation and ameliorated disease development. Transcriptomic analyses of MIA offspring indicated that in the intestine, P. goldsteinii MTS01 enhanced neuropeptide-related signaling and suppressed aberrant cell proliferation and inflammatory responses. In the hippocampus, P. goldsteinii MTS01 increased ribosomal/mitochondrial and antioxidant activities and decreased glutamate receptor signaling. Together, significant ameliorative effects of P. goldsteinii MTS01 on ASD relevant behaviors in MIA offspring were identified. Therefore, P. goldsteinii MTS01 could be developed as a next-generation probiotic for ameliorating ASD.

A comparison of the immunological potency of Burkholderia lipopolysaccharides in endotoxemic BALB/c mice.

Lipopolysaccharide is one of the virulence factors of the soil-borne pathogens Burkholderia pseudomallei, B. thailandensis, B. cenocepacia and B. multivorans, which cause septic melioidosis (often in B. pseudomallei infections but rarely in B. thailandensis infections) or cepacia syndromes (commonly in B. cenocepacia infections but rarely in B. multivorans infections). The inflammatory responses in Burkholderia LPS-induced endotoxemia were evaluated in this study. Prior to induction, the conserved structures and functions of each purified LPS were determined using electrophoretic phenotypes, the ratios of 3-hydroxytetradecanoic to 3-hydroxyhexadecanoic acid and endotoxin units. In an in vitro assay, cytokine expression of myeloid differentiation primary response gene 88 and Toll/IL-1 receptor domain containing adapter-inducing INF-ß-dependent signaling-dependent signaling differed when stimulated by different LPS. Endotoxemia was induced in mice by s.c. injection as evidenced by increasing serum concentrations of 3-hydroxytetradecanoic acid and the septic prognostic markers CD62E and ICAM-1. During endotoxemia, splenic CD11b+ I-A+ , CD11b+ CD80+ , CD11b+ CD86+ and CD11b+ CD11c+ subpopulations increased. After induction with B. pseudomallei LPS, there were significant increases in splenic CD49b NK cells and CD14 macrophages. The inflamed CD11b+ CCR2+ , CD11b+ CD31+ , CD11b+ CD14+ , resident CD11b+ CX3 CR1+ and progenitor CD11b+ CD34+ cells showed delayed increases in bone marrow. B. multivorans LPS was the most potent inducer of serum cytokines and chemokines, whereas B. cenocepacia LPS induced relatively low concentrations of the chemokines MIP-1α and MIP-1ß. Endotoxin activities did not correlate with the virulence of Burkholderia strains. Thus factors other than LPS and/or other mechanisms of low activity LPS must mediate the pathogenicity of highly virulent Burkholderia strains.

Improvement in T helper 1-related immune responses in BALB/c mice immunized with an HIV-1 gag plasmid combined with a chimeric plasmid encoding interleukin-18 and flagellin.

Both flagellin (fliC) and IL-18 (INF-γ-inducing factor) have been developed as adjuvants for improving immunogenicity in DNA-vaccinated hosts. An HIV-1 gag plasmid encodes a protein harboring broad epitopes for cytotoxic T-lymphocytes. In this study, the immunogenicity of BALB/c mice immunized with an HIV-1 gag plasmid (pVAX/gag) combined with a chimeric plasmid encoding IL-18 fused to flagellin (pcDNA3/IL-18_fliC) or a single plasmid encoding IL-18 (pcDNA3/IL-18) and/or flagellin (pcDNA3/fliC) was assessed. Through in vitro transcription and translation, it was demonstrated that both mRNA and protein were appropriately expressed by each construct. The IL-18 and flagellin fusion protein, which could be detected in supernatants from transfected cells, was effective in inducing IFN-γ by lymphocytes. Following i.m. immunization, expressions of flagellin or IL-18 were detected in muscle cells by immunohistochemistry analysis from 72 hr. At 12 weeks post-immunization, both gag-specific IgG in sera and spleen cell proliferation were high in all murine groups. However, the IgG2a/IgG1 ratio, Th1 cytokine (IL-2 and IFN-γ) production and proportion of gag-specific CD3(+) CD8(+) IFN-γ-secreting cells were significantly higher in the murine group co-immunized with pVAX/gag plasmid and pcDNA3/IL-18_fliC than in the mice immunized with pVAX/gag plasmid combined with either pcDNA3/fliC or pcDNA3/IL-18 plasmid or both. These findings suggest that a chimeric plasmid encoding IL-18 fused to flagellin can be used as an adjuvant-like plasmid to improve the Th1 immune response, particularly for induction of CD3(+) CD8(+) IFN-γ-secreting cells in gag plasmid-vaccinated mice.

Types of Septic Cardiomyopathy: Prognosis and Influencing Factors - A Clinical Study.

Objective: To explore the prognostic outcomes associated with different types of septic cardiomyopathy and analyze the factors that exert an influence on these outcomes. Methods: The data collected within 24 hours of ICU admission included cardiac troponin I (cTnI), N-terminal pro-Brain Natriuretic Peptide (NT-proBNP); SOFA (sequential organ failure assessment) scores, and the proportion of vasopressor use. Based on echocardiographic outcomes, septic cardiomyopathy was categorized into left ventricular (LV) systolic dysfunction, LV diastolic dysfunction, and right ventricular (RV) systolic dysfunction. Differences between the mortality and survival groups, as well as between each cardiomyopathy subgroup and the non-cardiomyopathy group were compared, to explore the influencing factors of cardiomyopathy. Results: A cohort of 184 patients were included in this study, with LV diastolic dysfunction having the highest incidence rate (43.5%). The mortality group had significantly higher SOFA scores, vasopressor use, and cTnI levels compared to the survival group; the survival group had better LV diastolic function than the mortality group (p < 0.05 for all). In contrast to the non-cardiomyopathy group, each subgroup within the cardiomyopathy category exhibited elevated levels of cTnI. The subgroup with left ventricular diastolic dysfunction demonstrated a higher prevalence of advanced age, hypertension, diabetes mellitus, coronary artery disease, and an increased mortality rate; the RV systolic dysfunction subgroup had higher SOFA scores and NT-proBNP levels, and a higher mortality rate (P < 0.05 for all); the LV systolic dysfunction subgroup had a similar mortality rate (P > 0.05). Conclusion: Patients with advanced age, hypertension, diabetes mellitus, or coronary artery disease are more prone to develop LV diastolic dysfunction type of cardiomyopathy; cardiomyopathy subgroups had higher levels of cTnI. The RV systolic dysfunction cardiomyopathy subgroup had higher SOFA scores and NT-proBNP levels. The occurrence of RV systolic dysfunction in patients with sepsis significantly increased the mortality rate.

Corrigendum: No safe renal warm ischemia time-The molecular network characteristics and pathological features of mild to severe ischemia reperfusion kidney injury.

[This corrects the article DOI: 10.3389/fmolb.2022.1006917.].

Early Diastolic Peak Velocity of Mitral Valve Annulus and Right Ventricular Systolic Tricuspid Annular Velocity as Predictors in Assessing Prognosis of Patients with Sepsis.

Objective: To analyze the epidemiological data of patients with septic cardiomyopathy and investigate the relationship between ultrasonic parameters and prognosis of patients with sepsis. Methods: In this study, we enrolled patients with sepsis who were treated at the Department of Critical Care Medicine in the Beijing Electric Power Hospital (No.1 Taipingqiao Xili, Fengtai District, Beijing) from January 2020 to June 2022. All patients received standardized treatment. Their general medical status and 28-day prognosis were recorded. Transthoracic echocardiography was performed within 24 hours after admission. We compared the ultrasound indexes between the mortality group and the survival group at the end of 28 days. We included parameters with significant difference in the logistic regression model to identify the independent risk factors for prognosis and evaluated their predictive value using receiver operating characteristic (ROC) curve. Results: We included 100 patients with sepsis in this study; the mortality rate was 33% and the prevalence rate of septic cardiomyopathy was 49%. The peak e' velocity and right ventricular systolic tricuspid annulus velocity (RV-Sm) of the survival group were significantly higher than those of the mortality group (P < 0.05). Results of logistic regression analysis showed that the peak e' velocity and RV-Sm were independent risk factors for prognosis. The area under curve of the peak e' velocity and the RV-Sm was 0.657 and 0.668, respectively (P < 0.05). Conclusion: The prevalence rate of septic cardiomyopathy in septic patients is high. In this study, we found that the peak e' velocity and right ventricular systolic tricuspid annulus velocity were important predictors of short-term prognosis.

Case cluster shifting and contaminant source as determinants of melioidosis in Taiwan.

OBJECTIVES: To assess the geographical distribution of melioidosis contamination sources and the association between the location of melioidosis cases and positive sampling sites for Burkholderia pseudomallei in Taiwan. METHODS: Data on the location of melioidosis cases from 2002 to 2011 were combined with the geographical distribution of B. pseudomallei as indicated by the detection of specific flagella gene products measured from 2005 to 2011. Temporal and spatial analyses were used to determine the incidence, cluster shifts and associations between the two datasets. RESULTS: Melioidosis cases clustered in two 'hot-spot' areas with incidence rates that were significantly higher than in neighbouring towns. The incidence rates in the northern area gradually decreased, while the rates in the southern area increased and were temporally associated with the appearance of B. pseudomallei-specific flagella genes in water samples. CONCLUSIONS: Melioidosis hot-spot areas were present in Taiwan. Water contaminated with B. pseudomallei serves as a potential transmission vehicle and is correlated with an increase in melioidosis cases; this correlation was stronger than that for B. pseudomallei-contaminated soil.

No safe renal warm ischemia time-The molecular network characteristics and pathological features of mild to severe ischemia reperfusion kidney injury.

Ischemic acute kidney injury (AKI) has always been a hot and difficult research topic in the field of renal diseases. This study aims to illustrate the safe warm ischemia time of kidney and the molecular network characteristics and pathological features of mild to severe ischemia reperfusion kidney injury. We established varying degrees of renal injury due to different ischemia time (0 min, 16 min, 18 min, 20 min, 22 min, 24 min, 26 min, 28 min, and 30 min) on unilateral (left kidney) ischemia-reperfusion injury and contralateral (right kidney) resection (uIRIx) mouse model. Mice were sacrificed 24 h after uIRIx, blood samples were harvested to detect serum creatinine (Scr), and kidney tissue samples were harvested to perform Periodic Acid-Schiff (PAS) staining and RNA-Seq. Differentially expressed genes (DEGs) were identificated, time-dependent gene expression patterns and functional enrichment analysis were further performed. Finally, qPCR was performed to validated RNA-Seq results. Our results indicated that there was no absolute safe renal warm ischemia time, and every minute of ischemia increases kidney damage. Warm ischemia 26min or above in mice makes severe kidney injury, renal pathology and SCr were both significantly changed. Warm ischemia between 18 and 26 min makes mild kidney injury, with changes in pathology and renal molecular expression, while SCr did not change. No obvious pathological changes but significant differences in molecular expression were found less than 16min warm ischemia. There are two key time intervals in the process of renal ischemia injury, 0 min-16 min (short-term) and 26 min-28 min (long-term). Gene expression of immune-related pathways were most significantly down-regulated in short-term ischemia, while metabolism-related pathways were the mainly enriched pathway in long-term ischemia. Taken together, this study provides novel insights into safe renal artery occlusion time in partial nephrectomy, and is of great value for elucidating molecular network characteristics and pathological features of mild to severe ischemia reperfusion kidney injury, and key genes related to metabolism and immune found in this study also provide potential diagnostic and therapeutic biomarkers for AKI.

Burkholderia multivorans acts as an antagonist against the growth of Burkholderia pseudomallei in soil.

In this study, it was demonstrated, by using agar diffusion tests and a Transwell system, that Burkholderia multivorans NKI379 has an antagonistic effect against the growth of B. pseudomallei. Bacterial representatives were isolated from agricultural crop soil and mixed to construct a partial bacterial community structure that was based on the results of reproducible patterns following PCR-denaturing gradient gel electrophoresis analysis of total soil chromosomes. The antagonistic effect of B. multivorans on B. pseudomallei was observed in this imitate community. In a field study of agricultural crop soil, the presence of B. pseudomallei was inversely related to the presence of the antagonistic strains B. multivorans or B. cenocepacia. B. multivorans NKI379 can survive in a broader range of pH, temperatures and salt concentrations than B. pseudomallei, suggesting that B. multivorans can adapt to extreme environmental changes and therefore predominates over B. pseudomallei in natural environments.

The Ferric Citrate Uptake System Encoded in a Novel bla CTX-M-3- and bla TEM-1-Harboring Conjugative Plasmid Contributes to the Virulence of Escherichia coli.

Escherichia coli is one major cause of bacterial infections and can horizontally acquire antimicrobial resistance and virulence genes through conjugation. Because conjugative plasmids can rapidly spread among bacteria of different species, the plasmids carrying both antimicrobial resistance and virulence genes may pose a significant threat to public health. Therefore, the identification and characterization of these plasmids may facilitate a better understanding of E. coli pathogenesis and the development of new strategies against E. coli infections. Because iron uptake ability is a potential virulence trait of bacteria, we screened for E. coli conjugative plasmids able to confer both iron uptake ability and ampicillin resistance. The plasmid pEC41, which was derived from the bacteremia clinical isolate EC41, was identified. EC41, which carried the fimH27 allele, belonged to sequence type (ST) 405 and phylogroup D. According to the sequencing analyses, pEC41 was 86 kb in size, and its backbone structure was almost identical to that of another highly conjugative plasmid, pCTX-M3, in which the extended-spectrum ß-lactamase gene bla CTX-M-3 was originally identified. pEC41 carried bla CTX-M-3 and bla TEM-1. The ferric citrate uptake (fec) system was identified in pEC41 and was responsible for conferring iron uptake ability. The fec system contributes to the pathogenesis of EC41 in systemic infections but not in urinary tract infections (UTIs). However, this system promoted competitive fitness of a cystitis-associated clinical isolate to colonize urinary tracts. Additionally, the distribution of the fec system was related to E. coli isolates associated with human bacteremia and UTIs. In summary, the present study identified a novel conjugative plasmid, pEC41, which conferred both antimicrobial resistance and an extra iron uptake ability to E. coli. The iron uptake ability was encoded in the fec system and contributed to E. coli pathogenesis. This study is the first to show that the fec system is a virulence factor in E. coli.

Distribution of melioidosis cases and viable Burkholderia pseudomallei in soil: evidence for emerging melioidosis in Taiwan.

A survey for the prevalence if Burkholderia pseudomallei in soil in Taiwan found that its incidence is comparable to that in other regions of the world where melioidosis is endemic. The presence of identical genetic patterns among the clinical and environmental isolates evaluated suggested a link between the pathogens present in contaminated soil and the emergence of indigenous melioidosis.

Peptidoglycan Endopeptidase Spr of Uropathogenic Escherichia coli Contributes to Kidney Infections and Competitive Fitness During Bladder Colonization.

Uropathogenic E scherichia coli (UPEC) is the most common pathogen of urinary tract infections (UTIs). Antibiotic therapy is the conventional measure to manage such infections. However, the rapid emergence of antibiotic resistance has reduced the efficacy of antibiotic treatment. Given that the bacterial factors required for the full virulence of the pathogens are potential therapeutic targets, identifying such factors may facilitate the development of novel therapeutic strategies against UPEC UTIs. The peptidoglycan (PG) endopeptidase Spr (also named MepS) is required for PG biogenesis in E. coli. In the present study, we found that Spr deficiency attenuated the ability of UPEC to infect kidneys and induced a fitness defect during bladder colonization in a mouse model of UTI. Based on the liquid chromatography (LC)/mass spectrometry (MS)/MS analysis of the bacterial envelope, spr deletion changed the levels of some envelope-associated proteins, suggesting that Spr deficiency interfere with the components of the bacterial structure. Among the proteins, FliC was significantly downregulated in the spr mutant, which is resulted in reduced motility. Lack of Spr might hinder the function of the flagellar transcriptional factor FlhDC to decrease FliC expression. The motility downregulation contributed to the reduced fitness in urinary tract colonization. Additionally, spr deletion compromised the ability of UPEC to evade complement-mediated attack and to resist intracellular killing of phagocytes, consequently decreasing UPEC bloodstream survival. Spr deficiency also interfered with the UPEC morphological switch from bacillary to filamentous shapes during UTI. It is known that bacterial filamentation protects UPEC from phagocytosis by phagocytes. In conclusion, Spr deficiency was shown to compromise multiple virulence properties of UPEC, leading to attenuation of the pathogen in urinary tract colonization and bloodstream survival. These findings indicate that Spr is a potential antimicrobial target for further studies attempting to develop novel strategies in managing UPEC UTIs.

Identification of non-fermenting Gram-negative bacteria of clinical importance by an oligonucleotide array.

Many species of non-fermenting Gram-negative bacilli (non-fermenters) are important opportunistic and nosocomial pathogens. Identification of most species of non-fermenters by phenotypic characteristics can be difficult. In this study, an oligonucleotide array was developed to identify 38 species of clinically relevant non-fermenters. The method consisted of PCR-based amplification of 16S-23S rRNA gene intergenic spacer (ITS) regions using bacterial universal primers, followed by hybridization of the digoxigenin-labelled PCR products with oligonucleotide probes immobilized on a nylon membrane. A total of 398 strains, comprising 276 target strains (i.e. strains belonging to the 38 species to be identified) and 122 non-target strains (i.e. strains not included in the array), were analysed by the array. Four target strains (three reference strains and one clinical isolate) produced discrepant identification by array hybridization. Three of the four discordant strains were found to be correctly identified by the array, as confirmed by sequencing of the ITS and 16S rRNA genes, with the remaining one being an unidentified species. The sensitivity and specificity of the array for identification of non-fermenters were 100 and 96.7%, respectively. In summary, the oligonucleotide array described here offers a very reliable method for identification of clinically relevant non-fermenters, with results being available within one working day.

Colonization of a medical center in Southern Taiwan by epidemic strains of carbapenem- and multidrug-resistant Acinetobacter baumannii and the genetic organization of their integrons.

A total of 46 carbapenem- and multidrug-resistant (CR- and MDR-)Acinetobacter baumannii bacteremic isolates from a Taiwanese medical center were investigated over the period 2000 to 2006 using randomly amplified polymorphic DNA (RAPD) profiling and by analysing the genetic organization of their integrons. The results of RAPD patterns revealed that before 2003 each CR- and MDR-A. baumannii bacteremic isolate was independent, but after 2003 the isolates appeared to belong in four epidemic strains and persisted in the hospital. All the CR- and MDR-A. baumannii strains harbored class I integron (intI1) genes. PCR amplification and nucleotide sequencing showed that the cassette genes of intI1 were found to form four different antibiotic-resistant gene alignments in those strains. The bla(IMP-1) gene in the cassette genes of intI1 was identified in a clone, which raised great concern that clonal spread of this strain or of an integron-mediated horizontal gene may have occurred.

Distinct Pathogenic Patterns of Burkholderia pseudomallei Isolates Selected from Caenorhabditis elegans and Dictyostelium discoideum Models.

Burkholderia pseudomallei is a selective agent that causes septic melioidosis and exhibits a broad range of lethal doses in animals. Host cellular virulence and phagocytic resistance are pathologic keys of B. pseudomallei. We first proposed Caenorhabditis elegans as the host cellular virulence model to mimic bacterial virulence against mammals and second established the resistance of B. pseudomallei to predation by Dictyostelium discoideum as the phagocytosis model. The saprophytic sepsis-causing Burkholderia sp. (B. pseudomallei, Burkholderia thailandensis, Burkholderia cenocepacia, and Burkholderia multivorans) exhibited different virulence patterns in both simple models, but B. pseudomallei was the most toxic. Using both models, attenuated isolates of B. pseudomallei were selected from a transposon-mutant library and a panel of environmental isolates and reconfirmed by in vitro mouse peritoneal exudate cell association and invasion assays. The distinct pathological patterns of melioidosis were inducted by different selected B. pseudomallei isolates. Fatal melioidosis was induced by the isolates with high virulence in both simple models within 4-5 day, whereas the low-virulence isolates resulted in prolonged survival greater than 30 day. Infection with the isolates having high resistance to D. discoideum predation but a low C. elegans killing effect led to 83% of mice with neurologic melioidosis. By contrast, infection with the isolates having low resistance to D. discoideum predation but high C. elegans killing effect led to 20% cases with inflammation in the salivary glands. Our results indicated that individual B. pseudomallei isolates selected from simple biological models contribute differently to disease progression and/or tissue tropism.

Extremely high prevalence and genetic diversity of hepatitis C virus infection among HIV-infected injection drug users in Taiwan.

BACKGROUND: An outbreak of human immunodeficiency virus (HIV) type 1 infection among injection drug users (IDUs) occurred in Taiwan, and thereafter, injection drug use became the most frequent risk factor for HIV infection in Taiwan. We sought to study the prevalence of and genotypes causing hepatitis C virus (HCV) infection among HIV-infected IDUs in Taiwan. METHODS: A multicenter, longitudinal cohort study of 990 HIV-infected IDUs was conducted from 1993 through 2006. Blood samples were collected and analyzed for the presence of antibody to HCV and to determine the genotype of HCV. RESULTS: The overall prevalence of HCV infection among HIV-infected IDUs was 96.6%. The annual prevalence increased from 65.5% before 2002 to 98.6% in 2006. The main circulating HCV genotypes were 1a (accounting for 29.2% of samples), 6a (23.5%), and 3a (20.2%), whereas 1b, the most predominant genotype circulating in the general population in Taiwan, accounted for only 13.2% of samples. Genotypes 2b (accounting for 6.6% of samples), 6k (2.9%), 2a (1.6%), 6g (1.6%), and 3b (1.2%) were present in only a few IDUs. Multivariate logistic regression analysis revealed that duration of injection drug use and a travel history to China or Southeast Asia were significantly associated with infection due to HCV genotypes 1a, 3, and 6. CONCLUSIONS: Our study demonstrated a high prevalence of HCV infection among HIV-infected IDUs in Taiwan, with a predominance of infection due to genotypes 1a, 6a, and 3a, as a result of the impact of IDUs' behavior and their drug trafficking route. Our study revealed that HCV infection in IDUs originated from a geographically large transmission network that was mainly distinct from that associated with other HCV-infected individuals; this transmission network has also been documented in association with HIV infection in IDUs.

Transmission Modes of Melioidosis in Taiwan.

In Taiwan, melioidosis is an emerging disease that suddenly increased in the Er-Ren River Basin, beginning in 2005 and in the Zoynan region during 2008⁻2012, following a typhoon. Additionally, the disease sporadically increased in a geography-dependent manner in 2016. Subcutaneous inoculation, ingestion, and the inhalation of soil or water contaminated with Burkholderia pseudomallei are recognized as the transmission modes of melioidosis. The appearance of environmental B. pseudomallei positivity in northern, central and southern Taiwan is associated with disease prevalence (cases/population: 0.03/100,000 in the northern region, 0.29/100,000 in the central region and 1.98/100,000 in the southern region). However, melioidosis-clustered areas are confined to 5 to 7.5 km² hot spots containing high-density populations, but B. pseudomallei-contaminated environments are located >5 km northwestern of the periphery of these hot spots. The observation that the concentration of B. pseudomallei-specific DNA in aerosols was positively correlated with the incidence of melioidosis and the appearance of a northwesterly wind in a hot spot indicated that airborne transmission had occurred in Taiwan. Moreover, the isolation rate in the superficial layers of a contaminated crop field in the northwest was correlated with PCR positivity in aerosols collected from the southeast over a two-year period. The genotype ST58 was identified by multilocus sequence typing in human and aerosol isolates. The genotype ST1001 has increased in prevalence but has been sporadically distributed elsewhere since 2016. These data indicate the transmission modes and environmental foci that support the dissemination of melioidosis are changing in Taiwan.

Burkholderia pseudomallei-loaded cells act as a Trojan horse to invade the brain during endotoxemia.

Neurologic melioidosis occurs in both human and animals; however, the mechanism by which the pathogen Burkholderia pseudomallei invades the central nervous system (CNS) remains unclear. B. pseudomallei-loaded Ly6C cells have been suggested as a putative portal; however, during melioidosis, lipopolysaccharide (LPS) can drive disruption of the blood-brain barrier (BBB). This study aims to test whether the Trojan horse-like mechanism occurs during endotoxemia. The expression levels of cerebral cytokines, chemokines and cell adhesion molecules; the activation of astrocytes, microglia and endothelial cells; and the increased vascular permeability and brain-infiltrating leukocytes were evaluated using B. pseudomallei, B. thailandensis, B. cenocepacia and B. multivorans LPS-induced brains. Accordingly, different degrees of BBB damage in those brains with endotoxemia were established. The B. multivorans LPS-induced brain exhibited the highest levels of disruptive BBB according to the above mediators/indicators. Into these distinct groups of endotoxemic mice, B. pseudomallei-loaded Ly6C cells or free B. pseudomallei were adoptively transferred at equal bacterial concentrations (103 CFU). The bacterial load and number of cases of meningeal neutrophil infiltration in the brains of animals treated with B. pseudomallei-loaded Ly6C cells were higher than those in brains induced by free B. pseudomallei in any of the endotoxemic groups. In particular, these results were reproducible in B. multivorans LPS-induced brains. We suggest that B. pseudomallei-loaded cells can act as a Trojan horse and are more effective than free B. pseudomallei in invading the CNS under septic or endotoxemic conditions even when there is a high degree of BBB disruption.

Performance characteristics of the limulus amebocyte lysate assay and gas chromatography-mass spectrum analysis of lipopolysaccharides relative to nitric oxide production by peritoneal exudates of cells.

Limulus amebocyte lysate (LAL) assay and gas chromatography-mass spectrum (GC-MS) analysis are usually used for the quantification of lipopolysaccharides (LPS) from the environment. The LAL assay measures endotoxin units to represent the LPS biological function but GC-MS analysis measures decanoic (C(10:0)) and dodecanoic (C(12:0)) 3-hydorxy fatty acids (3-OH FA) concentration to represent the LPS chemical composition. A study was carried out using these methods to evaluate their degree of correlation. Using the culture supernatants of 30 independent strains of Pseudomonas aeruginosa, the bacterial supernatants gave of 0.53+/-0.45 and 2.49+/-1.75mg/l of C(10:0) and C(12:0) 3-OH FA, respectively, compared to 17.96+/-13.28mg/l of LPS with the LAL assay (1ng/ml of LPS congruent with0.78EU/ml). The 3-OH FA concentration relative to the endotoxin unit dose in the supernatants exhibited a positive correlation (r(2)=0.5182, C(10:0); r(2)=0.4359, C(12:0)). When supernatants having a high level of endotoxin were used to treat peritoneal exudates cells, nitric oxide (NO) was generated in a dose-dependent manner (r(2)=0.6174). To determine if either C(10:0) or C(12:0) 3-OH FA can act as an indictor of LPS quantity was correlated with this immunostimulatory effect, the correlation of these 3-OH FA concentrations against the produced NO levels was evaluated. This also exhibited a positive correlation; however, the two indicators of 3-OH FA gave different dose-responsible performances (r(2)=0.3211, C(10:0); r(2)=0.4527, C(12:0)).

Protective effects of a freeze-dried extract of vegetables and fruits on the hydroxyl radical-mediated oxidative damage of DNA and decrease of erythrocytes deformability.

The protective effects of a freeze-dried extracts of vegetables and fruits (BauYuan; BY) on the hydroxyl radical-mediated DNA strand breakages and the structural integrity of human red blood cells (RBCs) were investigated. First, the supercoiled plasmid (pEGFP-C1) DNA was subjected to oxidation damage by an ascorbate-fortified Fenton reaction and the protective effects were analyzed by agarose gel electrophoresis. In the absence of BY extracts, exposure of the high-throughput .OH-generating system (Fe2+ concentration >1.0 microM) caused a complete fragmentation of DNA. Supplementation of BY extract (1 mg/mL) to the plasmid DNA prior to the exposure could prevent it significantly. In contrast, as the plasmid exposed to a low-grade .OH-generating system (Fe2+<0.1 microM), the BY extract (1 mg/mL) provided an almost complete protection. Next, the cell deformabilities were measured to assess the protection effects of various BY extracts on human erythrocytes exposed to the oxidative insults. We found that both the aqueous extract and the organic solvent-derived extracts could strongly protect human RBCs from the reactive oxygen species (ROS)-mediated decrease in the deformability indices. The results implicated that the BY extracts could effectively protect the cell membrane integrity via scavenging ROS which enabling RBCs to maintain a balance of water content and surface area to prevent the drop of cell deformability.