Cytotoxic Lymphocyte-Monocyte Complex Reflects the Dynamics of Coronavirus Disease 2019 Systemic Immune Response.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection causes a variety of clinical manifestations, many of which originate from altered immune responses, either locally or systemically. Immune cell cross-talk occurs mainly in lymphoid organs. However, systemic cell interaction specific to coronavirus disease 2019 has not been well characterized. Here, by employing single-cell RNA sequencing and imaging flow cytometry analysis, we unraveled, in peripheral blood, a heterogeneous group of cell complexes formed by the adherence of CD14+ monocytes to different cytotoxic lymphocytes, including SARS-CoV-2-specific CD8+ T cells, γδ T cells, and natural killer T cells. These lymphocytes attached to CD14+ monocytes that showed enhanced inflammasome activation and pyroptosis-induced cell death in progression stage; in contrast, in the convalescent phase, CD14+ monocytes with elevated antigen presentation potential were targeted by cytotoxic lymphocytes, thereby restricting the excessive immune activation. Collectively, our study reports previously unrecognized cell-cell interplay in the SARS-CoV-2-specific immune response, providing new insight into the intricacy of dynamic immune cell interaction representing antiviral defense.

Polymorphism in the EREG gene confers susceptibility to tuberculosis.

BACKGROUND: Host genetic factors affect the immune response to Mycobacterium tuberculosis (Mtb) infection as well as the progression of the disease. Epiregulin (EREG) belongs to the epidermal growth factor (EGF) family, which binds to the epidermal growth factor receptor (EGFR) to regulate the immune response of the host during infections. Our study aimed to compare EREG levels in tuberculosis (TB) patients and healthy controls and assess whether polymorphisms in EREG increase the risk of TB. METHODS: We used ELISA to determine the plasma EREG level from 30 healthy controls and 50 tuberculosis patients. By evaluating the EREG gene from 624 TB patients and 600 healthy controls, we determined the allelic and genotypic frequencies for association with susceptibility to TB infections in this group. RESULTS: This paper shows that the pulmonary tuberculosis (PTB) and extrapulmonary tuberculosis (EPTB) groups showed a significantly higher plasma EREG level (1014 ± 733.9 pg/ml, 700.2 ± 676.6 pg/ml, respectively) than the healthy controls (277 ± 105.4 pg/ml). The rs2367707 polymorphism was associated with a higher risk of PTB and EPTB (P = 0.00051, P = 0.0012). Analyses of haplotype frequencies found that people with the haplotype CACAT had a higher risk of PTB and EPTB (P = 0.00031, OR = 1.43; P = 0.000053, OR = 1.65). Moreover, the rs6446993 polymorphism of the EREG gene was found to be associated with EPTB (P = 0.00087, OR = 1.54; 95% CI = 1.23-1.94). CONCLUSIONS: Compared to that of healthy controls, the level of EREG in the plasma of TB patients increased significantly. Based on these data, we demonstrated that EREG polymorphisms are genetic factors for susceptibility to TB and various forms of TB.

Evaluation of Matrix-Assisted Laser Desorption Ionization-Time of Flight Mass Spectrometry for Identification of Mycobacterium abscessus Subspecies According to Whole-Genome Sequencing.

This study was undertaken to evaluate the utility of matrix-assisted laser desorption ionization-time of flight mass spectrometry with the Vitek MS Plus system for identifying Mycobacterium abscessus subspecies in order to facilitate more rapid and appropriate therapy. A total of 175 clinical M. abscessus strains were identified by whole-genome sequencing analysis: 139 Mycobacterium abscessus subsp. abscessus and 36 Mycobacterium abscessus subsp. massiliense The research-use-only (RUO) Saramis Knowledge Base database v.4.12 was modified accordingly by adding 40 M. abscessus subsp. abscessus and 19 M. abscessus subsp. massiliense reference spectra to construct subspecies SuperSpectra. A blind test, used to validate the remaining 116 isolates, yielded 99.1% (n = 115) reliability and only 0.9% (n = 1) error for subspecies identification. Among the two subspecies SuperSpectra, two specific peaks were found for M. abscessus subsp. abscessus and four specific peaks were found for M. abscessus subsp. massiliense Our study is the first to report differential peaks 3,354.4 m/z and 6,711.1 m/z, which were specific for M. abscessus subsp. massiliense Our research demonstrates the capacity of the Vitek MS RUO Saramis Knowledge Base database to identify M. abscessus at the subspecies level. Moreover, it validates the potential ease and accuracy with which it can be incorporated into the IVD system for the identification of M. abscessus subspecies.

CHAMP: A Centrifugal Microfluidics-Based CRISPR/Cas12b-Combined Real-Time LAMP One-Pot Method for Mycoplasma pneumoniae Infection Diagnosis.

The Mycoplasma pneumoniae outbreak poses health risks to community residents. However, it still has limitations for current clinical diagnostic methods (qPCR nucleic acid assay or IgM immunoassay), including specialized handling, expensive equipment, prolonged turnaround time, and false positives and negatives, highlighting the need to improve clinical diagnostic methods. Herein, we present a novel centrifugal microfluidics-based method for rapidly diagnosing M. pneumoniae infections (CHAMP system). This user-friendly method combines CRISPR/Cas12b and real-time loop-mediated isothermal amplification (LAMP) in a one-pot reaction, offering high sensitivity, specificity, and simplicity for methodology. By adding fully automated nucleic acid magnetic bead-extracted samples to a prepackaged centrifugal microfluidics chip, 48 samples can be automated tested simultaneously within 15 to 60 min at 60 °C. 427 clinical nasopharyngeal swab specimens were used for validation, demonstrating good positive and negative predictive values and good diagnostic sensitivity, specificity, and significant time savings. This method is particularly suitable for detecting low nucleic acid copies of M. pneumoniae samples.

Mycobacterium tuberculosis hijacks host macrophages-derived interleukin 16 to block phagolysosome maturation for enhancing intracellular growth.

The discovery of promising cytokines and clarification of their immunological mechanisms in controlling the intracellular fate of Mycobacterium tuberculosis (Mtb) are necessary to identify effective diagnostic biomarkers and therapeutic targets. To escape immune clearance, Mtb can manipulate and inhibit the normal host process of phagosome maturation. Phagosome maturation arrest by Mtb involves multiple effectors and much remains unknown about this important aspect of Mtb pathogenesis. In this study, we found that interleukin 16 (IL-16) is elevated in the serum samples of Tuberculosis (TB) patients and can serve as a specific target for treatment TB. There was a significant difference in IL-16 levels among active TB, latent TB infection (LTBI), and non-TB patients. This study first revealed that macrophages are the major source of IL-16 production in response to Mtb infection, and elucidated that IL-16 can promote Mtb intracellular survival by inhibiting phagosome maturation and suppressing the expression of Rev-erbα which can inhibit IL-10 secretion. The experiments using zebrafish larvae infected with M. marinum and mice challenged with H37Rv demonstrated that reducing IL-16 levels resulted in less severe pathology and improved survival, respectively. In conclusion, this study provided direct evidence that Mtb hijacks the host macrophages-derived interleukin 16 to enhance intracellular growth. It is suggesting the immunosuppressive role of IL-16 during Mtb infection, supporting IL-16 as a promising therapeutic target.

Using Vitek MS v3.0 To Identify Nontuberculous Mycobacteria in Liquid Media in a Clinical Microbiology Laboratory.

Recently, the incidence of diseases caused by nontuberculous mycobacteria (NTM) has been increasing worldwide, especially in immunocompromised patients and those with potential chronic lung disease. Vitek MS v3.0 matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) has emerged as a rapid and reliable method for identifying mycobacteria in clinical laboratories. This study aimed to evaluate the performance of Vitek MS v3.0 by isolating NTM directly from automated liquid medium systems using patient samples. A total of 855 Mycobacterium growth indicator tube (MGIT)-positive liquid cultures were investigated. Among them, 658 (77.0%) liquid cultures were correctly identified to the species, group, or complex level, 192 (23.0%) resulted in no identification, and 5 (0.6%) were misidentified at the species level. DNA sequencing identified 855 NTM isolates from liquid cultures, comprising 316 isolates of rapidly growing mycobacteria (RGM) and 539 isolates of slow-growing mycobacteria (SGM). Using the Vitek MS system, the RGM integral identification rate (276/316 [87.34%]) was higher than the SGM rate (381/539 [70.69%]) (P < 0.01). It was also higher than the SGM rate for all MGIT report-positive periods. These results indicate that the Vitek MS v3.0 system can rapidly identify NTM species from liquid cultures. Further validation using molecular techniques is required. IMPORTANCE Rapid and accurate identification of nontuberculous mycobacteria (NTM) is essential for diagnosis, appropriate therapy, and infection control. Vitek MS v3.0 matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) has emerged as a rapid and reliable method for identifying mycobacteria in clinical laboratories. This study reported a clinical validation of the Vitek MS V3.0 system for identification of NTM isolates from 855 MGIT-positive liquid cultures which contained relatively large NTM types. Vitek MS v3.0 showed a promising rate for identification NTM isolates in positive liquid cultures. Vitek MS v3.0 had a better performance with RGM than with SGM. Vitek MS v3.0 results included "unidentified" or "misidentified" NTM isolates, which would also serve as an important reference for future optimization of this system. Vitek MS v3.0 represented a valuable technique for NTM identification from positive liquid cultures.

Efflux pump overexpression in conjunction with alternation of outer membrane protein may induce Acinetobacter baumannii resistant to imipenem.

BACKGROUND: To investigate the role of outer membrane proteins in Acinetobacter baumannii resistant to imipenem, 2 strains were procured from the same patient. METHODS: The imipenem-resistant strain was obtained following a period of imipenem treatment in vivo. The multilocus sequence typing and repetitive extragenic palindromic PCR results indicated that the imipenem-resistant strain originated from the sensitive one. RESULTS: Isoelectric focusing detected no carbapenemases, with neither OXA carbapenemases nor metallo-ß-lactamases found. Mass spectrophotometric analysis revealed that 3 outer membrane proteins were expressed differentially in the 2 strains: 2 downregulated proteins (OprD and CarO) and 1 upregulated the 34-kDa efflux pump protein in the resistant strain. A 32-fold decrease in the MIC for imipenem in the presence of Phe-Arg-ß-naphthylamide in the same strain indicated a possible involvement of the efflux pump mechanism in its resistance, which was consistent with the findings that the mRNA expression of the 34-kDa efflux pump gene was almost fivefold upregulated in the imipenem-resistant strain compared with that in the imipenem-sensitive strain. Such a significant difference, however, was not found in the expression of AdeB and AdeJ between the 2 strains, and AdeE was not detected. CONCLUSIONS: Our results suggested that downregulation of outer membrane proteins in conjunction with efflux pump overexpression might contribute to imipenem resistance induced in vivo in A. baumannii.

Hybrid Cu2O/TiO2 Nanocomposites with Enhanced Photocatalytic Antibacterial Activity toward Acinetobacter Baumannii.

The hybrid Cu2O-TiO2 photocatalytic composite with a p-n heterojunction was synthesized by the supercritical solvothermal route. The uniformly distributed Cu2O was stably combined with TiO2 to benefit the increase of the specific surface area, the harvesting of visible light, and the separation of photogenerated holes and electrons. As a result, photocatalytic inactivation of Acinetobacter baumannii under visible-light irradiation was facilitated. Based on the photogenerated holes acting as the main active species, the Cu2O-TiO2 photocatalytic heterojunction could induce the leakage of K+ ions and serious damage of the cell structure, leading to the final cell death. During the photoantibacterial process on Cu2O-TiO2, the RecA gene played a significant role in enhancing the survival rate of Acinetobacter baumannii. Furthermore, the great bactericidal effect of Cu2O-TiO2 nanocomposites to different bacteria indicated the potential application for the environment-friendly disinfection.

Evaluation of the VITEK MS knowledge base version 3.0 for the identification of clinically relevant Mycobacterium species.

Different Mycobacterium spp. infections may indicate varied treatment regimens in the clinic. Thus, the species-level identification of Mycobacterium spp. is one of the most important tasks for a clinical microbiology laboratory. Although matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) has emerged as a rapid and accurate method for the identification of mycobacteria, this method lacks a comprehensive evaluation of the identification accuracy for clinically collected mycobacteria using VITEK MS Knowledge Base Version 3.0 (Ver 3.0). The objectives of the present study were to evaluate the identification performance of Mycobacterium spp. using Ver 3.0 and a sample processing kit for strain inactivation and protein extraction. Among the 507 Mycobacterium isolates, 46 isolates were M. tuberculosis, and 461 isolates were nontuberculous mycobacteria (NTM) (including 27 species: 17 species were slowly growing mycobacteria (SGM), and 10 species were rapidly growing mycobacteria (RGM)). The VITEK MS V3.0 library was used to correctly identify 476/507 (93.9%) isolates (425 isolates were correctly identified initially, and 51 more isolates were correctly identified on repeat), 23/507 (4.5%) isolates were unidentified, and 8/507 (1.6%) isolates were misidentified. In summary, we showed that Mycobacterium spp. can be adequately identified by Ver 3.0 in combination with the use of a standard sample processing kit.

Synergistic Photocatalytic-Photothermal Contribution to Antibacterial Activity in BiOI-Graphene Oxide Nanocomposites.

The threat of environmental microbial contamination to the health of human beings has drawn particular attention. In order to explore the synergistic effect of photocatalytic and photothermal process to the antibacterial property, a stably combined BiOI-graphene oxide (GO) nanocomposite was constructed and prepared through a facile solvothermal method. BiOI crystals were uniformly distributed on the GO nanosheets by the formation of the Bi-C bond. On the basis of various characterizations, the great surface area, the high light harvesting with extension into NIR region, and the efficient transfer of photoinduced electrons by the conductivity of GO were demonstrated, all of which are beneficial for the photocatalytic antibacterial activity. More importantly, the photothermal effect of GO increased the temperature of the BiOI-GO composite with high photothermal conversion efficiency and induced the photogenerated electrons from BiOI crystals to obtain more energy and higher carrier mobility. Conversely, the temperature elevation of BiOI-GO composite improved its capability for light absorption and separation of photoinduced charges. As a result, the BiOI-GO composite enabled the synergistic photocatalytic-photothermal effect for the improvement of the antibacterial property for Acinetobacter baumannii with higher efficiency of TOC removal and leakage of K+ ions, in comparison with the individual photocatalytic process. Thus, the synergistic photocatalytic-photothermal contribution of BiOI-GO composite will provide significance for the potential application of environmental disinfection in the future.

Relationship between Antibiotic Susceptibility and Genotype in Mycobacterium abscessus Clinical Isolates.

This study aimed to determine the antibiotic susceptibility and resistance related genotypes of Mycobacterium abscessus. One hundred sixty-two clinical isolates were collected. Genomic data were obtained by whole genome sequencing. Single nucleotide polymorphism (SNP) analysis was conducted using the NCBI GenBank database and BLAST algorithm. The following genes were of interest: erm(41), rrl and rrs. Erm(41) was further divided into 3 sequevars: erm(41)C28, erm(41)T28, and M type [erm(41) with deletions in nucleotides 64 and 65, or 159 through 432]. Antibiotic susceptibility was assessed at 3 days (early reading time, ERT) and 14 days (late reading time, LRT) after clarithromycin (CLA) treatment. Three patterns of CLA resistance were observed. (1) Fifty-five (acquired resistance) isolates [45 erm(41)T28, 1 erm(41)C28 and 9 M type] exhibited MIC ≥8 mg/L at ERT; among these isolates, 10 had an rrl 2058/2059 mutation. (2) Sixty-two subsp. abscessus and 2 subsp. massiliense (induced resistance) isolates exhibited MIC ≤4 mg/L at ERT, but ≥8 mg/L at LRT. (3) Forty-three (sensitive and intermediate) isolates [14 erm(41)C28, 1 erm(41)T28, and 28 M type] exhibited MIC ≤4 mg/L at both ERT and LRT. No rrs 1408 mutation or other meaningful SNP was found in 3 amikacin-resistant isolates. No correlation was found between rrl, erm(41) or rrs and susceptibility to the 8 other antibiotics tested. The rrl and erm(41) genotypes could predict the CLA resistance of M. abscessus clinical isolates. China has a large number of CLA-resistant M. abscessus isolates with erm(41)T28 sequevar. Treatment of M. abscessus infections should be based upon a comprehensive consideration of factors that include genotype and geographic location.

Rapid detection of mutations in erm(41) and rrl associated with clarithromycin resistance in Mycobacterium abscessus complex by denaturing gradient gel electrophoresis.

Clarithromycin resistance is increasing dramatically among Mycobacterium abscessus complex. The main resistance mechanisms are mutations in the erm(41) and rrl genes. Here we report PCR-based denaturing gradient gel electrophoresis (DGGE) as an alternative method for rapidly detection of mutations in erm(41) and rrl among M. abscessus isolates. Four primer sets targeting the full-length erm(41) gene and a 354bp fragment of the rrl gene were designed. A combination of 16 different DGGE patterns were observed for erm(41) gene, including 16 in M. abscessus subsp. abscessus and 1 in M. abscessus subsp. massiliense. Six DGGE patterns were obtained for rrl gene. Mutations in the erm(41) and rrl detected by DGGE were 100% identical to mutations detected by DNA sequencing. This is the first report to identify PCR-based DGGE as a practical, relatively inexpensive technique for rapidly detecting mutations in the erm(41) and rrl genes associated with clarithromycin resistance in M. abscessus complex.

Antigens Rv0310c and Rv1255c are promising novel biomarkers for the diagnosis of Mycobacterium tuberculosis infection.

This study aimed to identify novel immunogenic epitopes from Mycobacterium tuberculosis (MTB) that could be used in tuberculosis (TB) diagnostics. To determine the diagnostic potential of mycobacterial antigens in serodiagnosis of TB, 256 patients were enrolled in a study and divided into two groups: 126 smear-positive pulmonary TB patients (SPPT) and 130 smear-negative pulmonary TB patients (SNPT); 152 bacillus Calmette-Guerin (BCG)-vaccinated healthy people were used as a control. Murine results showed that antigens Rv0310c-E from RD 8 and Rv1255c-E from RD 10 were strongly immunogenic to Th1 cells and induced a great humoral response. Receiver operating characteristic analysis indicated that Rv0310c-E (area under the curve (AUC): 0.800) and Rv1255c-E (AUC: 0.808) performed better than ESAT-6 (AUC: 0.665) and CFP-10 (AUC: 0.623) proteins but were comparable with Rv3425 (AUC: 0.788) protein in a human serum IgG analysis. Rv0310c-E demonstrated the highest diagnostic ability for the SPPT group (Youden index: 0.5602, sensitivity: 69.84%, specificity: 86.18%), while Rv1255c-E demonstrated the highest diagnostic ability for the SNPT group (Youden index: 0.5674, sensitivity: 73.84%, specificity: 82.89%). In addition, combination analysis found that antigen Rv0310c-E, coupled with the Rv3425 protein (Youden index: 0.6098, sensitivity: 87.30%, specificity: 73.68%) had the strongest performance for TB diagnostics of the SPPT group, and the single antigen Rv1255c-E was strongest for the SNPT group. These results suggest that antigens Rv0310c-E and Rv1255c-E are potential antigens for TB serodiagnostic tests, which may facilitate detection of MTB in smear-negative and smear-positive patients.

Association of Single-Nucleotide Polymorphism in the Hepcidin Promoter Gene with Susceptibility to Extrapulmonary Tuberculosis.

BACKGROUND: Hepcidin is a 25-amino acid peptide produced by the liver in response to inflammation and iron overload. It is encoded by the hepcidin antimicrobial peptide (HAMP) gene and plays a key role in innate immunity. Previous studies have reported that a -582 A>G polymorphism in the HAMP promoter (HAMP-P) affects hepcidin expression, causing susceptibility to various bacterial and viral pathogens. However, it is not known whether the HAMP-P -582 A>G polymorphism is associated with tuberculosis (TB) susceptibility. AIMS: The objective of the current study was to examine the relationship between the HAMP-P -582 A>G polymorphism and TB susceptibility in a Chinese Han population. METHODS: Han Chinese subjects examined included 500 pulmonary TB, 386 extrapulmonary TB, and 600 healthy control subjects. We analyzed correlations between the hepcidin promoter -582 A>G polymorphism and disease susceptibility and then examined the regulatory effects of the -582 A>G variant on hepcidin production in CD14+ monocyte cultures stimulated with lipoarabinomannan derived from Mycobacterium tuberculosis. RESULTS: Our findings indicate that the HAMP-P -582 A>G polymorphism (rs10421768) is associated with susceptibility to extrapulmonary TB, but not pulmonary TB. CD14+ monocytes from individuals with the rs10421768 GG genotype secreted significantly less hepcidin in response to M. tuberculosis lipoarabinomannan compared with cells from individuals with either the AA or AG genotypes. CONCLUSIONS: The G allele of the HAMP-P -582 A>G gene may play a critical role in TB susceptibility.

Polymorphisms in the prostaglandin receptor EP2 gene confers susceptibility to tuberculosis.

OBJECTIVES: Prostaglandin E2 (PGE2) is an important lipid mediator of the inflammatory immune response during acute and chronic infections. PGE2 modulates a variety of immune functions via four receptors (EP1-EP4), which mediate distinct PGE2 effects. Mice lacking EP2 are more susceptible to infection by Mycobacterium tuberculosis (M.tb), have a higher bacterial load, and increase size and number of granulomatous lesions. Our aim was to assess whether single nucleotide polymorphisms (SNPs) in EP2 increase the risk of tuberculosis. METHODS: DNA re-sequencing revealed five common EP2 variants in the Chinese Han population. We sequenced the EP2 gene from 600 patients and 572 healthy controls to measure SNP frequencies in association with tuberculosis infections (TB) within the population. RESULTS: The rs937337 polymorphism is associated with increased risk to tuberculosis (p=0.0044, odds ratio [OR], 1.67; 95% confidential interval,1.22-2.27). The rs937337 AA genotype and the rs1042618 CC genotype were significantly associated with TB. An estimation of the frequencies of haplotypes revealed a single protective haplotype GACGC for tuberculosis (p=0.00096, odds ratio [OR], 0.56; 95% confidential interval, 0.41-0.77). Furthermore, we determined that the remaining SNPs of EP2 were nominally associated with clinical patterns of disease. CONCLUSIONS: We identified genetic polymorphisms in EP2 associated with susceptibility to tuberculosis within a Chinese population. Our data support that EP2 SNPs are genetic predispositions of increased susceptibility to TB and to different clinical patterns of disease.

Characterization of Mycobacterium Abscessus Subtypes in Shanghai of China: Drug Sensitivity and Bacterial Epidemicity as well as Clinical Manifestations.

The aim of the study was to investigate the epidemic characteristics of Mycobacterium abscessus in Shanghai.Fifty-five strains from 55 M. abscessus pulmonary disease patients were isolated. Drug sensitivity was measured by a broth microdilution method. Subtypes of M. abscessus were identified by DNA sequencing. Multilocus sequence typing (MLST), mining spanning tree (MST), and pulsed-field gel electrophoresis (PFGE) were used to analyze sequence types (ST) and clonal complexes (CC). Clinical manifestations were assessed by CT imaging.We identified 42 A isolates, 11 M, and 2 B-subtypes. A and M were highly sensitive to tigecycline and amikacin (97.6-100%). The A-type easily developed drug resistance against clarithromycin. Both types were highly resistance to sulfonamides, moxifloxacin, doxycycline, imipenem, and tobramycin. MLST analysis identified 41 STs including 32 new STs. The MST algorithm distributed 55 isolates into 12 separate CC. The PFGE analysis exhibited 53 distinct restriction patterns and the M-type was closely clustered according to their ST and CC numbers. CT imaging showed that tree-in-bud and patch shadow were commonly observed in M-type, whereas pulmonary cavities were often found in A-type infection patients (P < 0.001).ST1 in A and ST23 in M-type were the main epidemic strains in Shanghai. The M-type appeared to be prone to epidemic nosocomial transmission.

PPE26 induces TLR2-dependent activation of macrophages and drives Th1-type T-cell immunity by triggering the cross-talk of multiple pathways involved in the host response.

The pathophysiological functions and the underlying molecular basis of PE /PPE proteins of M. tuberculosis remain largely unknown. In this study, we focused on the link between PPE26 and host response. We demonstrated that PPE26 can induce extensive inflammatory responses in macrophages through triggering the cross-talk of multiple pathways involved in the host response, as revealed by iTRAQ-based subcellular quantitative proteomics. We observed that PPE26 is able to specifically bind to TLR2 leading to the subsequent activation of MAPKs and NF-κB signaling. PPE26 functionally stimulates macrophage activation by augmenting pro-inflammatory cytokine production (TNF-α, IL-6 and IL-12 p40) and the expression of cell surface markers (CD80, CD86, MHC class I and II). We observed that PPE26-treated macrophages effectively polarizes naïve CD4(+) T cells to up-regulate CXCR3 expression, and to secrete IFN-γ and IL-2, indicating PPE26 contributes to the Th1 polarization during the immune response. Importantly, rBCG::PPE26 induces stronger antigen-specific TNF-α and IFN-γ activity, and higher levels of the Th1 cytokines TNF-α and IFN-γ comparable to BCG. Moreover, PPE26 effectively induces the reciprocal expansion of effector/memory CD4(+)/CD8(+) CD44(high)CD62L(low) T cells in the spleens of mice immunized with this strain. These results suggest that PPE26 may be a TLR2 agonist that stimulates innate immunity and adaptive immunity, indicating that PPE26 is a potential antigen for the rational design of an efficient vaccine against M. tuberculosis.

A novel recombinant BCG-expressing pro-apoptotic protein BAX enhances Th1 protective immune responses in mice.

One-third of the world's population is infected with Mycobacterium tuberculosis (MTB). The protective efficacy of bacille Calmette Guérin (BCG) vaccine against tuberculosis (TB) in adults is highly controversial even though the BCG vaccine has been available for more than 90 years. Because BCG is effective against infantile tuberculosis meningitis and miliary tuberculosis in young children and provides cost-effective prevention from tuberculosis for developing countries, it would be desirable to modify the existing BCG vaccine to provide more comprehensive protection. In our study, we constructed a novel recombinant BCG strain expressing pro-apoptotic BAX (rBCG::BAX) and demonstrated that it significantly induced the apoptosis of macrophages infected with rBCG::BAX both in vitro and in vivo. In addition, it significantly enhanced Ag85B-specific IFN-γ enzyme-linked immunospot responses, IFN-γ secretion, IL-2 secretion and the ratio of Ag85B-specific IgG2b/IgG1, and it significantly decreased Ag85B-specific IL-4. Furthermore, it presumably facilitated antigen presentation by inducing a significant up-regulation in the expression of MHC-II and B7.1 (CD80) co-stimulatory molecules on macrophages. In conclusion, these results suggest that the rBCG::BAX strain elicited predominantly a Th1 protective immune responses and might be a potential tuberculosis vaccine candidate for further study.

Tree-in-bud pattern of chest CT images for diagnosis of Mycobacterium abscesses.

OBJECTIVES: Changes of chest CT images in Mycobacterium and non-Mycobacterium abscesses in patients with lung disease were with a view to making an early diagnosis. METHODS: 124 primary patients diagnosed with non-tuberculosis Mycobacterium infections with a positive sputum acid-fast smear were enrolled in this retrospective study. CT images and clinical data of these patients were analyzed. RESULTS: The 52 Mycobacterium abscess lung disease cases included bronchiectasis 82.7% (43/52), which was more easily detected bilaterally than unilaterally (29/52 vs. 14/52), lung consolidation 44.2% (23/52), nodules 44.2% (22/52), cavities 32.7% (17/52), tree-in-bud pattern 42.3% (22/52) and patchy shadow 63.5% (33/52) in CT images. Tree-in-bud pattern was more common in Mycobacterium abscess compared with non-Mycobacterium abscess lung disease (42.3% vs. 18.1%, P = 0.004). A significant difference of the lung area involved by tree-in-bud in CT was found between non-Mycobacteria abscess and Mycobacterium abscess lung disease (17.0% vs. 7.2%, P < 0.001), and tree-in-bud occurred more readily unilaterally (21.2% vs. 6.9%, P = 0.029), and in the inferior lobe of the right lung (3.2% vs. 0.2%, P = 0.029) in Mycobacterium abscess lung disease. Patchy shadow was more common in non-Mycobacterium abscess lung disease (63.5% vs. 80.1%, P = 0.041). Further multi-factor analysis confirmed that tree-in-bud was an independent predictor of Mycobacterium abscess lung disease. CONCLUSIONS: Different CT results existed between non-Mycobacterium abscess and Mycobacterium abscess lung diseases. The tree-in-bud pattern might be helpful to choose a suitable therapy in patients, with an acid-fast bacilli smear-positive diagnosis of lung disease.