A hypervirulent Acinetobacter baumannii strain has robust anti-phagocytosis ability.

BACKGROUND: Acinetobacter baumannii (A. baumannii) is associated with both hospital-acquired infections (HAP) and community-acquired pneumonia (CAP). In this study, we present a novel CAP-associated A. baumannii (CAP-AB) strain causing severe pneumonia in an afore healthy male patient without underlying conditions. Subsequently, we investigated the pathogenicity and immunogenicity of this CAP-AB strain using a mice pneumonia model. RESULTS: A 58-year-old male patient with no underlying conditions experienced worsening symptoms of a productive cough, sputum, and fever that developed acutely, in just 24 h. The diagnosis was severe community-acquired pneumonia (CAP) and type-1 respiratory failure. An A. baumannii strain was isolated from his sputum and blood cultures. To gain a deeper understanding of the rapid progression of its pathology, we utilized the CAP-associated A. baumannii strain YC128, a previously obtained hospital-acquired pneumonia A. baumannii (HAP-AB) strain YC156, and a highly virulent A. baumannii control strain LAC-4 to construct a mouse pneumonia model, and subsequently compared the mortality rate of the three groups. Following inoculation with 107 CFU of A. baumannii, the mortality rate for the YC128, LAC-4, and YC156 groups was 60% (6/10), 30% (3/10), and 0%, respectively. The bacterial burden within the pulmonary, liver, and spleen tissues of mice in the YC128 group was significantly higher than that of the YC156 group, and slightly higher than that of the LAC-4 group. Pathological analysis of lung tissue using HE-staining revealed that the inflammatory pathological changes in mice from the YC128 group were significantly more severe than those in the YC156 group. Additionally, CT scan images displayed more pronounced inflammation in the lungs of mice from the YC128 group compared to the YC156 group. Local levels of cytokines/chemokines such as IL-1ß, IL-6, TNF-α, and CXCL1 were assessed via RT-qPCR in lung tissues. In comparison with the YC156 strain, the highly virulent YC128 strain induced the expression of proinflammatory cytokines more rapidly and severely. Furthermore, we examined the in vitro anti-phagocytosis ability of YC128 and YC156 strains against mice peritoneal macrophages, revealing that the highly virulent YC128 isolate displayed greater resistance to macrophage uptake in contrast to YC156. Results from Whole Genome Sequencing (WGS) indicated that YC128 harbored a complete type VI secretion system (T6SS) gene cluster, while YC156 lacked the majority of genes within the T6SS gene cluster. The other virulence-related genes exhibited minimal differences between YC128 and YC156. Drawing from previous studies, we postulated that the T6SS is linked to the hypervirulence and robust anti-phagocytic ability of YC128. CONCLUSIONS: This article reports on the isolation of a novel hypervirulent CAP-AB strain, YC128, from a severe CAP patient. The results demonstrate that this CAP-AB strain, YC128, is capable of inducing fatal pneumonia and extrapulmonary dissemination in a mouse pneumonia model. Moreover, this highly virulent CAP-AB strain exhibits significantly stronger anti-phagocytic abilities compared to the HAP-AB YC156 strain. Genome sequencing comparisons reveal that the heightened hypervirulence and enhanced anti-phagocytosis abilities observed in YC128 may be attributed to the presence of the T6SS.

Phosphodiesterase-4D Knockdown in the Prefrontal Cortex Alleviates Memory Deficits and Synaptic Failure in Mouse Model of Alzheimer's Disease.

Phosphodiesterase 4 (PDE4)-dependent cAMP signaling plays a crucial role in cognitive impairment associated with Alzheimer's disease (AD). However, whether inhibition of PDE4 subtypes or their splice variants in the prefrontal cortex positively regulates synaptic plasticity and antioxidative stress, and reverses ß-amyloid 1-42 (Aß1-42, Aß42)-induced cognitive impairment still need to be clarified. The present study determined whether and how PDE4D knockdown by microinjection of lenti-PDE4D-miRNA into the prefrontal cortex reversed Aß1-42-induced cognitive impairment in behavioral, neurochemical, and molecular biology assays. The results suggested that PDE4D knockdown increased time to explore the novel object and decreased latency to leave the platform in novel object recognition and step-down passive avoidance tests. Further study suggested that PDE4D knockdown decreased the number of working memory errors in the eight-arm maze test. These effects were prevented by PKA inhibitor H89. The subsequent experiment suggested that inhibition of PDE4D in the prefrontal cortex rescued the long-term potentiation (LTP) and synaptic proteins' expression; it also increased antioxidant response by increasing superoxide dismutase (SOD) and decreasing malondialdehyde (MDA) levels. PDE4D knockdown also increased phosphorylated cAMP response element-binding protein (pCREB), brain-derived neurotrophic factor (BNDF), and anti-apoptotic proteins' expression, i.e., the ratio of Bcl-2/Bax, and decreased caspase-3 level in the prefrontal cortex. These findings extend the previous findings and support the hypothesis that RNA interference-mediated PDE4D knockdown in the prefrontal cortex ameliorated memory loss associated with synaptic failure in an AD mouse model by its antioxidant, anti-apoptotic, and neuroprotective properties.

A High-Sensitivity Method Based on Advanced Optical Waveguide Technology to Detect HBsAg.

A novel method for the detection of the hepatitis B surface antigen (HBsAg) at low concentrations, using the ultrahigh-order guided mode acting as the probe excited by a symmetrical metal-cladding waveguide, is proposed. The method using the fact of the minimum value of the absorption peaks is proportional to the concentration of the sample to be detected to realize the detection of the hepatitis B virus at extremely low concentrations. It is realized that the low concentration of the HBsAg measurement relied on the principle of the minimum value of the absorption peak and the concentration having a good linear relationship. The measurement results indicate that this new method can precisely detect HBsAg at the concentrations in the lower region of the clinical gray area (i.e., below 20 ng/mL), the lower region of the current clinical gray area of HBsAg (below 20 ng/ml) can be measured, and the resolution can be reached (2 ng/mL).

Long Noncoding RNA LINC00460 Promotes Hepatocellular Carcinoma Progression via Regulation of miR-342-3p/AGR2 Axis.

BACKGROUND: Hepatocellular carcinoma (HCC) is the leading cause of cancer-related death worldwide. LINC00460, a novel long non-coding RNA (lncRNA), was recently confirmed as an oncogene in various cancers. However, the biological function and underlying mechanism of LINC00460 in HCC is largely obscure. METHODS: Fifty pairs of tumor tissue and adjacent normal tissues from HCC patients, as well as six HCC cell lines and a normal human hepatic epithelial cell line were subjected to qRT-PCR assay to evaluate the expression levels of LINC00460. CCK-8 assays were used to detect the proliferation of HCC cells. Transwell assay was used to measure the migration and invasion abilities of HCC cells. RNA pull-down and luciferase assays were performed to verify the direct interaction between LINC00460 and miR-342-3p. A xenograft model of HCC was established to validate the in vivo function of LINC00460 in HCC progression. RESULTS: We firstly detected LINC00460 expression was significantly upregulated in both HCC tumor tissues and cell lines. The upregulation of LINC00460 was positively associated with HCC progression. Functionally, LINC00460 facilitated HCC cell proliferation, migration, and invasion capacities, which due to that LINC00460 could physically bind to and repress miR-342-3p to elevate the expression of AGR2. CONCLUSION: Our data firstly reveal the clinical relevance, biological function, and regulatory mechanism of LINC00460 in HCC development. LINC00460 promotes HCC progression by elevating AGR2 expression via sponging miR-342-3p, providing a promising therapeutic target for HCC treatment.

Biofilm formation in erythromycin-resistant Staphylococcus aureus and the relationship with antimicrobial susceptibility and molecular characteristics.

PURPOSE: In this study, we aimed to investigate biofilm formation characteristics in clinical Staphylococcus aureus (S. aureus) isolates with erythromycin (ERY) resistance from China and further analyze their correlations with antimicrobial susceptibility and molecular characteristics. METHODOLOGY: A total of 276 clinical isolates of ERY-resistant S. aureus, including 142 methicillin-resistant S. aureus (MRSA) strains and 134 methicillin-susceptible S. aureus (MSSA) strains, were retrospectively collected in China. Biofilms were determined by crystal violet staining and ERY resistance genes (ermA, ermB and ermC) were detected by polymerase chain reaction. Inducible clindamycin resistance was examined by D test and multilocus sequence typing, and clonal complexes (CCs) based on housekeeping genes were further determined. RESULTS: The frequency of biofilm formation among ERY-resistant S. aureus was 40.9% (113/276) in total and no significant difference was found for the frequency of biofilm formation between ERY-resistant MRSA and ERY-resistant MSSA (44.4% vs 37.3%, P > 0.05). In ERY-resistant MRSA isolates, the frequency of biofilm formation in ermA-positive, gentamicin-resistant and ciprofloxacin-resistant isolates was higher than that in ermA-negative, gentamicin-sensitive and ciprofloxacin-sensitive isolates, respectively (63.9% vs 23.6%, P < 0.01; 60.3% vs 27.5%, P < 0.01; 65.2% vs 26.3%, P < 0.01). In addition, tetracycline resistance facilitated biofilm formation in both ERY-resistant MRSA and MSSA and the frequency of biofilm formation in CC239- or CC7S. aureus isolates with ERY resistance was significantly higher compared with that in CC59S. aureus (both P < 0.01). CONCLUSION: The ermA gene, and gentamicin, ciprofloxacin and tetracycline resistance facilitate biofilm formation in ERY-resistant MRSA isolates and, moreover, ERY-resistant S. aureus isolates with positive biofilm formation exhibited clonality clustering regarding CC239 and CC7.

Cellular immune responses in patients with hepatitis B surface antigen seroclearance induced by antiviral therapy.

BACKGROUND: The mechanisms by which chronic hepatitis B is completely resolved through antiviral therapy are unknown, and the contribution of acquired T cell immunity to hepatitis B surface antigen (HBsAg) seroclearance has not been investigated. Therefore, we measured the T-cell responses to core and envelope antigens in patients with HBsAg seroclearance. METHODS: Fourteen subjects with HBsAg seroclearance following antiviral treatment for chronic hepatitis B, 7 HBeAg-positive immunotolerant HBV carriers and 9 HBeAg-negative inactive HBsAg carriers were recruited. HBV-specific T-cell responses to recombinant HBV core (rHBcAg) and envelope (rHBsAg) proteins and pools of core and envelope peptides were measured using an ELISPOT assay detecting interferon-gamma and intracellular cytokine staining (ICS) assays detecting interferon-gamma or interleukin 2. RESULTS: Interferon-gamma ELISPOT assays showed a low frequency of weak responses to the rHBsAg and S peptide pool in the HBsAg seroclearance group, and the response frequency to the rHBcAg and the C peptide pool was higher than to the rHBsAg (P < 0.001) and S peptide pool (P = 0.001) respectively. A higher response frequency to C than S peptide pools was confirmed in the interferon-gamma ICS assays for both CD4+ (P = 0.033) and CD8+ (P = 0.040) T cells in the HBsAg seroclearance group. The responses to C and S antigens in the inactive carriers were similar. CONCLUSIONS: There was a low frequency of CD4+ and CD8+ T cell immune responses to envelope antigens in Chinese subjects with HBsAg seroclearance following antiviral therapy. It is unlikely that these immune responses are responsible for HBsAg seroclearance in these subjects.

Presence of valine at position 27 of the hepatitis B virus core gene is associated with severe liver inflammation in Chinese patients.

Although it is widely believed that cytotoxic T lymphocytes (CTL) are responsible for severe flares of chronic hepatitis B that lead to liver failure, the published evidence to support this hypothesis is weak. The frequency of the I27V mutation in the HBV core gene, which produces a core 18-27 peptide capable of binding HLA-A*02, was compared in Chinese patients with severe liver inflammation (n = 77, including 39 with acute-on-chronic liver failure), moderate liver inflammation (n = 44) and inactive disease (n = 45). The frequency with which V27 reverted to the wild-type I27 was compared in severe liver inflammation patients who were either HLA-A*02 positive (n = 5) or negative (n = 5). The frequency of patients with a V27 positive HBV was higher in severe than in moderate liver inflammation (23.4% vs. 6.8%, P = 0.02) or inactive disease (23.4% vs. 4.7%, P = 0.006). After a minimum of 3 months follow-up, the frequency of reversion of V27 to the wild-type I27 was higher in HLA-A*02 positive than negative patients (5/5 vs. 1/5, P = 0.05). In summary, this is the first data showing an association between a specific amino acid mutation (I27V) and severe liver inflammation in patients with chronic hepatitis B. This mutation would produce a peptide that is known to bind HLA-A*02 and stimulate CTL. The high frequency of reversion to wild-type I27 in HLA-A*02 positive subjects suggests that CTL recognizing this peptide exist, and is consistent with the possibility that they contribute to the pathophysiology of severe liver inflammation in chronic hepatitis B.

[Characteristics of CDR3 of TCR beta on CD8+ T cells in chronic hepatitis B patients].

OBJECTIVE: To analyze the characteristics of CDR3 of TCRbeta on CD8+ T cells in chronic hepatitis B patients. METHODS: Eight patients with chronic hepatitis B (ALT more than 2 ULN) were enrolled in this study. CD8+ T cells were isolated from peripheral blood. RT-PCR was proformed to amplify the CDR3 of TCRbeta, and the PCR products were sequenced and analyzed. RESULTS: The chronic hepatitis B patients showed obvious clonal expansion of T cell, and three perturbation patterns of T cell expansion were showed in the CDR3 of TCRbeta, including monoclonicity, oligoclonicity and skewed peak patterns. The number of perturbation families of CD8+ subpopulation was significantly higher than that of CD8- subpopulation (10.6+/-4.7 vs. 4.1+/-3.1, t = 6.619, P less than 0.01). In 3 out of 8 patients, the number of perturbation families of CD8+ subpopulation was also higher than that of PBMCs without depleting CD8+ subpopulation. CONCLUSIONS: The characteristics of CDR3 of TCRbeta may help to understand the inflammatory response in CHB patients.