Lung disease caused by non-tuberculous mycobacteria (NTM), relatives of Mycobacterium tuberculosis, is increasing. M. abscessus is the most prevalent rapid growing NTM. This environmental pathogen is intrinsically resistant to most commonly used antibiotics, including anti-tuberculosis drugs. Current therapies take years to achieve cure, if cure if achieved. Thus, there is an urgent medical need to identify new, more efficacious treatments. Here, we explore the possibility of repurposing antibiotics developed for other indications. We asked whether novel two-drug combinations of clinically used antibiotics can be identified that show synergistic activity against this mycobacterium. An in vitro checkerboard titration assay was employed to test 180 dual combinations of 41 drugs against the clinical isolate M. abscessus Bamboo. The most attractive novel combination was further profiled against reference strains representing three sub-species (M. abscessus subsp. abscessus, massiliense and bolletii) and a collection of clinical isolates. This resulted in the identification of a novel synergistic antibiotic pair active against the M. abscessus complex: the glycopeptide teicoplanin with the glycylcycline tigecycline showed inhibitory activity at 2-3 μM (teicoplanin) and 1-2 μM (tigecycline). This novel combination can now be tested in M. abscessus animal models of infection and/or patients.

The mink circovirus (MiCV), a newly discovered pathogen, is associated with diarrhea in farmed minks. The prevalence and economic importance of this virus remain poorly understood, and a quantitative method for diagnosis of MiCV infection has not been established. This research aims to develop a highly specific, sensitive, and quantitative assay for MiCV. A Real-Time quantitative polymerase chain reaction (qPCR) assay was developed to detect different isolates of the MiCV in mink samples. The qPCR system is highly sensitive with a detection limit of as low as 10 viral DNA copies. The specificity of this qPCR assay was supported by the absence of cross-reaction with other pathogens. The coefficients of variation were low for both inter-assay and intra-assay variabilities. In addition, the results also expressed the distribution of MiCV in infectious mink tissues with high levels of virus in the skeletal muscle and heart. The heart occupied a higher proportion than other tissues, which can be considered the primary source of test material. This qPCR method could be a useful tool for epidemiological studies and disease management. This method for MiCV is highly specific, sensitive, repeatable, quantitative, and can rapidly determine viral load levels in different tissues samples.

Plants recognize unrelated viruses by the antiviral defense system called RNA interference (RNAi). RNAi processes double-stranded viral RNA into small RNAs (sRNAs) of 21-24 nucleotides, the reassembly of which into longer strands in silico allows virus identification by comparison with the sequences available in databases. The aim of this study was to compare the virus detection sensitivity of sRNA-based virus diagnosis with the established virus species-specific polymerase chain reaction (PCR) approach. Viruses propagated in tobacco plants included three engineered, infectious clones of Potato virus A (PVA), each carrying a different marker gene, and an infectious clone of Potato virus Y (PVY). Total RNA (containing sRNA) was isolated and subjected to reverse-transcription real-time PCR (RT-RT-PCR) and sRNA deep-sequencing at different concentrations. RNA extracted from various crop plants was included in the reactions to normalize RNA concentrations. Targeted detection of selected viruses showed a similar threshold for the sRNA and reverse-transcription quantitative PCR (RT-qPCR) analyses. The detection limit for PVY and PVA by RT-qPCR in this study was 3 and 1.5 fg of viral RNA, respectively, in 50 ng of total RNA per PCR reaction. When knowledge was available about the viruses likely present in the samples, sRNA-based virus detection was 10 times more sensitive than RT-RT-PCR. The advantage of sRNA analysis is the detection of all tested viruses without the need for virus-specific primers or probes.

A dysbiotic microbiome can potentially contribute to the pathogenesis of many different diseases including cancer. Breast cancer is the second leading cause of cancer death in women. Thus, we investigated the diversity of the microbiome in the four major types of breast cancer: endocrine receptor (ER) positive, triple positive, Her2 positive and triple negative breast cancers. Using a whole genome and transcriptome amplification and a pan-pathogen microarray (PathoChip) strategy, we detected unique and common viral, bacterial, fungal and parasitic signatures for each of the breast cancer types. These were validated by PCR and Sanger sequencing. Hierarchical cluster analysis of the breast cancer samples, based on their detected microbial signatures, showed distinct patterns for the triple negative and triple positive samples, while the ER positive and Her2 positive samples shared similar microbial signatures. These signatures, unique or common to the different breast cancer types, provide a new line of investigation to gain further insights into prognosis, treatment strategies and clinical outcome, as well as better understanding of the role of the micro-organisms in the development and progression of breast cancer.

Goose parvovirus (GPV) remains as a worldwide problem in goose industry. For this reason, it is necessary to develop a new diagnostic approach that is easier and faster than conventional tests. A rapid immunochromatographic assay based on antibody colloidal gold nanoparticles specific to GPV was developed for the detection of GPV in goose allantoic fluid and supernatant of tissue homogenate. The monoclonal antibodies (Mab) was produced by immunizing the BALB/c mice with purified GPV suspension, and the polyclonal antibody (pAb) was produced by immunizing the rabbits with recombinant VP3 protein. The colloidal gold was prepared by the reduction of gold salt with sodium citrate coupled with Mab against GPV. The optimal concentrations of the coating antibody and capture antibody were determined to be 1.6 mg/ml and 9 μg/ml. With visual observation, the lower limit was found to be around 1.2 μg/ml. Common diseases of goose were tested to evaluate the specificity of the immune colloidal gold (ICG) strip, and no cross-reaction was observed. The clinical detection was examined by carrying out the ICG strip test with 92 samples and comparing the results of these tests with those obtained via agar diffusion test and polymerase chain reaction (PCR) test. Therefore, the ICG strip test was a sufficiently sensitive and accurate detection method for a rapid screening of GPV.

We investigated the impact of temperature on the microbial turnover of organic matter (OM) in a hydrothermal vent system in Guaymas Basin, by calculating microbial bio- and necromass turnover times based on the culture-independent D:L-amino acid model. Sediments were recovered from two stations near hydrothermal mounds (<74°C) and from one cold station (<9°C). Cell abundance at the two hydrothermal stations dropped from 108 to 106 cells cm-3 within ∼5 m of sediment depth resulting in a 100-fold lower cell number at this depth than at the cold site where numbers remained constant at 108 cells cm-3 throughout the recovered sediment. There were strong indications that the drop in cell abundance was controlled by decreasing OM quality. The quality of the sedimentary OM was determined by the diagenetic indicators %TAAC (percentage of total organic carbon present as amino acid carbon), %TAAN (percentage of total nitrogen present as amino acid nitrogen), aspartic acid:β-alanine ratios, and glutamic acid:γ-amino butyric acid ratios. All parameters indicated that the OM became progressively degraded with increasing sediment depth, and the OM in the hydrothermal sediment was more degraded than in the uniformly cold sediment. Nonetheless, the small community of microorganisms in the hydrothermal sediment demonstrated short turnover times. The modeled turnover times of microbial bio- and necromass in the hydrothermal sediments were notably faster (biomass: days to months; necromass: up to a few hundred years) than in the cold sediments (biomass: tens of years; necromass: thousands of years), suggesting that temperature has a significant influence on the microbial turnover rates. We suggest that short biomass turnover times are necessary for maintance of essential cell funtions and to overcome potential damage caused by the increased temperature.The reduced OM quality at the hyrothemal sites might thus only allow for a small population size of microorganisms.

Adjuvant 5-fluorouracil (5-FU)-based chemotherapy, including FOLFOX (5-FU, leucovorin, and oxaliplatin), is recommended for colorectal cancer. However, intestinal mucositis remains a common adverse effect for which no effective preventive strategies are available. To develop a convenient and novel way to alleviate mucositis, we investigated the effect of Lactobacillus casei variety rhamnosus (Lcr35) on FOLFOX-induced mucosal injury. BALB/c mice subcutaneously injected with syngeneic CT26 colorectal adenocarcinoma cells were orally administered Lcr35 daily before, during, and after 5-day injection of FOLFOX regimen, for 14 days. The following methods were used: diarrhea score for toxicity, ELISA for cytokine production, histopathology for intestinal injury, immunohistochemistry for apoptosis/proliferation and regulatory proteins, RT-PCR for cytokine mRNA expression, and DNA sequencing for fecal gut microbiota. FOLFOX administration to colorectal cancer-bearing mice significantly inhibited tumor growth and the accompanying marked diarrhea and intestinal injury histologically characterized by the shortening of villi and destruction of intestinal crypts. Preventive administration of Lcr35 dose-dependently reduced the severity of diarrhea and intestinal mucositis without affecting the anti-tumor effect of FOLFOX. The numbers of apoptotic, NF-κB-, and BAX-activated cells increased after FOLFOX, and these responses were mitigated by Lcr35. TNF-α and IL-6 upregulation by FOLFOX treatment was attenuated by Lcr35. The fecal gut microbiota composition of Firmicutes and Bacteroidetes disturbed by FOLFOX was significantly reversed by Lcr35 toward a preferential profile. In conclusion, the oral probiotic Lcr35 prevented FOLFOX-induced intestinal mucositis in colorectal cancer-bearing mice. The putative mechanism might involve modulation of gut microbiota and proinflammatory responses with suppression of intrinsic apoptosis in intestinal injury.

Recent metagenomic surveys of microbial community suggested that species associated with the class Acidimicrobiia are abundant in diverse aquatic environments such as acidic mine water, waste water sludge, freshwater, or marine habitats, but very few species have been cultivated and characterized. The current taxonomic framework of Acidimicrobiia is solely based on 16S rRNA sequence analysis of few cultivable representatives, and no molecular, biochemical, or physiological characteristics are known that can distinguish species of this class from the other bacteria. This study reports the phylogenomic analysis for 20 sequenced members of this class and reveals another three major lineages in addition to the two recognized families. Comparative analysis of the sequenced Acidimicrobiia species identified 15 conserved signature indels (CSIs) in widely distributed proteins and 26 conserved signature proteins (CSPs) that are either specific to this class as a whole or to its major lineages. This study represents the most comprehensive phylogenetic analysis of the class Acidimicrobiia and the identified CSIs and CSPs provide useful molecular markers for the identification and delineation of species belonging to this class or its subgroups.

The hemibiotrophic plant pathogen Colletotrichum higginsianum infects Brassicaceae and in combination with Arabidopsis thaliana, represents an important model system to investigate various ecologically important fungal pathogens and their infection strategies. After penetration of plant cells by appressoria, C. higginsianum establishes large biotrophic primary hyphae in the first infected cell. Shortly thereafter, a switch to necrotrophic growth occurs leading to the invasion of neighboring cells by secondary hyphae. In a forward genetic screen for virulence mutants by insertional mutagenesis, we identified mutants that penetrate the plant but show a defect in the passage from biotrophy to necrotrophy. Genome sequencing and pulsed-field gel electrophoresis revealed that two mutants were lacking chromosome 11, encoding potential pathogenicity genes. We established a chromosome loss assay to verify that strains lacking this small chromosome abort infection during biotrophy, while their ability to grow on artificial media was not affected. C. higginsianum harbors a second small chromosome, which can be lost without effects on virulence or growth on agar plates. Furthermore, we found that chromosome 11 is required to suppress Arabidopsis thaliana plant defense mechanisms dependent on tryptophan derived secondary metabolites.

Gut microbial dysbiosis is correlated with the development of hepatocellular carcinoma (HCC). Therefore, analyzing the changing patterns in gut microbiota during HCC development, especially before HCC occurrence, is essential for the diagnosis and prevention of HCC based on gut microbial composition. However, these changing patterns in HCC are poorly understood, especially considering the sex differences in HCC incidence and mortality. Here, with an aim to determine the relationship between gut microbiota and HCC development in both sexes, and to screen potential microbial biomarkers for HCC diagnosis, we studied the changing patterns in the gut microbiota from mice of both sexes with liver-specific knockout of Tsc1 (LTsc1KO) that spontaneously developed HCC by 9-10 months of age and compared them to the patterns observed in their wide-type Tsc1fl/fl cohorts using high-throughput sequencing. Using the LTsc1KO model, we were able to successfully exclude the continuing influence of diet on the gut microbiota. Based on gut microbial composition, the female LTsc1KO mice exhibited gut microbial disorder earlier than male LTsc1KO mice during the development of HCC. Our findings also indicated that the decrease in the relative abundance of anaerobic bacteria and the increase in the relative abundance of facultative anaerobic bacteria can be used as risk indexes of female HCC, but would be invalid for male HCC. Most of the changes in the gut bacteria were different between female and male LTsc1KO mice. In particular, the increased abundances of Allobaculum, Erysipelotrichaceae, Neisseriaceae, Sutterella, Burkholderiales, and Prevotella species have potential for use as risk indicators of female HCC, and the increased abundances of Paraprevotella, Paraprevotellaceae, and Prevotella can probably be applied as risk indicators of male HCC. These relationships between the gut microbiota and HCC discovered in the present study may serve as a platform for the identification of potential targets for the diagnosis and prevention of HCC in the future.

Human campylobacteriosis is the leading food-borne zoonosis in industrialized countries. This study characterized the clonal population structure, antimicrobial resistance profiles and occurrence of antimicrobial resistance determinants of a set of Campylobacter jejuni strains isolated from broiler carcasses in Belgium. Minimum inhibitory concentrations (MICs) against five commonly-used antibiotics (ciprofloxacin, nalidixic acid, tetracycline, gentamicin, and erythromycin) were determined for 204 C. jejuni isolates. More than half of the isolates were resistant to ciprofloxacin or nalidixic acid. In contrast, a lower percentage of screened isolates were resistant to gentamicin or erythromycin. C. jejuni isolates resistant to ciprofloxacin and/or nalidixic acid were screened for the substitution T86I in the quinolone resistance determining region (QRDR) of the gyrA gene, while C. jejuni isolates resistant to tetracycline were screened for the presence of the tet(O) gene. These resistance determinants were observed in most but not all resistant isolates. Regarding resistance to erythromycin, different mutations occurred in diverse genetic loci, including mutations in the 23S rRNA gene, the rplD and rplV ribosomal genes, and the intergenic region between cmeR and cmeABC. Interestingly, and contrary to previous reports, the A2075G transition mutation in the 23S rRNA gene was only found in one strain displaying a high level of resistance to erythromycin. Ultimately, molecular typing by multilocus sequence typing revealed that two sequence types (ST-824 and ST-2274) were associated to quinolones resistance by the presence of mutations in the gene gyrA (p = 0.01). In addition, ST-2274 was linked to the CIP-NAL-TET-AMR multidrug resistant phenotype. In contrast, clonal complex CC-45 was linked to increased susceptibility to the tested antibiotics. The results obtained in this study provide better understanding of the phenotypic and the molecular basis of antibiotic resistance in C. jejuni, unraveling some the mechanisms which confer antimicrobial resistance and particular clones associated to the carriage and spread of resistance genes.

Control of biogenic amines (BAs) is critical to guarantee the safety of fermented meat products. The aim of this study is to evaluate the BAs formation and degradation abilities of lactic acid bacteria from Chinese bacon to obtain the beneficial candidate for BAs control. Seven lactic acid bacteria were selected from the typical Chinese bacon products, identified as Lactobacillus curvatus by 16S rDNA analysis. Then, genes analysis and high-performance liquid chromatography (HPLC) analysis were performed to evaluate the BAs formation and degradation abilities of as-selected strains. All L. curvatus strains were confirmed to harbor the genes encoding the tyrosine decarboxylase and ornithine decarboxylase, and they could produce tyramine, β-phenethylamine, putrescine, and cadaverine. In comparison, the lowest concentration of total BAs was obtained in L. curvatus G-1. Meanwhile, all L. curvatus strains were positive in amines oxidase gene analysis, and they could also degrade six common BAs, especially the L. curvatus G-1 with the highest degradation percentage (above 40%) for each BA. Furthermore, fermented meat model analysis verified that the L. curvatus G-1 could significantly reduce BAs. In conclusion, L. curvatus G-1 shows a low BAs-producing ability, as well as a high BAs-degrading ability, and this study provides a promising candidate for potential BAs control in fermented meat products.

The TatD-like DNase of Plasmodium species has previously been characterized as a conserved antigen that plays an important role in immune evasion. Here, we found that TatD-like DNase is expressed, apart from the erythrocytic stage, throughout the developmental stages of the parasite in the mosquito vector. Antibodies to the molecule significantly blocked parasites development and transition in the mosquito gut. Further, mice immunized with recombinant TatD-like DNase showed significant resistance to parasite challenge. The antigenicity of the TatD-like antigen in combination with various adjuvants, including Freund's adjuvants, Montanide ISA 51 and 61, Alhydrogel (aluminum hydroxide), and levamisole was investigated. It was found that immunization of the recombinant TatD-like DNase in combination with Montanide ISA 51 induced strong humoral responses that showed significant protection against parasite challenge in a mouse model. The data further support that TatD-like DNase is a functionally important molecule in the whole development cycle of the malaria parasites and a candidate for malaria vaccine development.

Aspergillus niger is one of the most widely used fungi to study the conversion of the lignocellulosic feedstocks into fermentable sugars. Understanding the sugar uptake system of A. niger is essential to improve the efficiency of the process of fungal plant biomass degradation. In this study, we report a comprehensive characterization of the sugar transportome of A. niger by combining phylogenetic and comparative transcriptomic analyses. We identified 86 putative sugar transporter (ST) genes based on a conserved protein domain search. All these candidates were then classified into nine subfamilies and their functional motifs and possible sugar-specificity were annotated according to phylogenetic analysis and literature mining. Furthermore, we comparatively analyzed the ST gene expression on a large set of fungal growth conditions including mono-, di- and polysaccharides, and mutants of transcriptional regulators. This revealed that transporter genes from the same phylogenetic clade displayed very diverse expression patterns and were regulated by different transcriptional factors. The genome-wide study of STs of A. niger provides new insights into the mechanisms underlying an extremely flexible metabolism and high nutritional versatility of A. niger and will facilitate further biochemical characterization and industrial applications of these candidate STs.

Mycobacterium tuberculosis can proficiently enter macrophages and diminish complement activation on its cell surface. Within macrophages, the mycobacterium can suppress macrophage apoptosis and survive within the intracellular environment. Previously, we have shown that complement regulatory proteins such as factor H may interfere with pathogen-macrophage interactions during tuberculosis infection. In this study, we show that Mycobacterium bovis BCG binds properdin, an upregulator of the complement alternative pathway. TSR4+5, a recombinant form of thrombospondin repeats 4 and 5 of human properdin expressed in tandem, which is an inhibitor of the alternative pathway, was also able to bind to M. bovis BCG. Properdin and TSR4+5 were found to inhibit uptake of M. bovis BCG by THP-1 macrophage cells in a dose-dependent manner. Quantitative real-time PCR revealed elevated pro-inflammatory responses (TNF-α, IL-1β, and IL-6) in the presence of properdin or TSR4+5, which gradually decreased over 6 h. Correspondingly, anti-inflammatory responses (IL-10 and TGF-β) showed suppressed levels of expression in the presence of properdin, which gradually increased over 6 h. Multiplex cytokine array analysis also revealed that properdin and TSR4+5 significantly enhanced the pro-inflammatory response (TNF-α, IL-1β, and IL-1α) at 24 h, which declined at 48 h, whereas the anti-inflammatory response (IL-10) was suppressed. Our results suggest that properdin may interfere with mycobacterial entry into macrophages via TSR4 and TSR5, particularly during the initial stages of infection, thus affecting the extracellular survival of the pathogen. This study offers novel insights into the non-complement related functions of properdin during host-pathogen interactions in tuberculosis.

γδ T cells have broad reactivity and actively participate in protective immunity against tumors and infectious disease-causing organisms. In γδ-high species such as ruminants and other artiodactyls many γδ T cells bear the lineage-specific markers known as WC1. WC1 molecules are scavenger receptors coded for by a multigenic array and are closely related to SCART found on murine γδ T cells and CD163 found on a variety of cells. We have previously shown that WC1 molecules are hybrid pattern recognition receptors thereby binding pathogens as well as signaling co-receptors for the γδ T cell receptor. WC1+ γδ T cells can be divided into two major subpopulations differentiated by the WC1 genes they express and the pathogens to which they respond. Therefore, we hypothesize that optimal γδ T cell responses are contingent on pathogen binding to WC1 molecules, especially since we have shown that silencing WC1 results in an inability of γδ T cells from primed animals to respond to the pathogen Leptospira, a model system we have employed extensively. Despite this knowledge about the crucial role WC1 plays in γδ T cell biology, the pattern of WC1 gene expression by individual γδ T cells was not known but is critical to devise methods to engage γδ T cells for responses to specific pathogens. To address this gap, we generated 78 γδ T cell clones. qRT-PCR evaluation showed that approximately 75% of the clones had one to three WC1 genes transcribed but up to six per cell occurred. The co-transcription of WC1 genes by clones showed many combinations and some WC1 genes were transcribed by both subpopulations although there were differences in the overall pattern of WC1 genes transcription. Despite this overlap, Leptospira-responsive WC1+ memory γδ T cell clones were shown to have a significantly higher propensity to express WC1 molecules that are known to bind to the pathogen.

Targeted cancer immunotherapy with irradiated, granulocyte-macrophage colony-stimulating factor (GM-CSF)-secreting, allogeneic cancer cell lines has been an effective approach to reduce tumor burden in several patients. It is generally assumed that to be effective, these cell lines need to express immunogenic antigens coexpressed in patient tumor cells, and antigen-presenting cells need to take up such antigens then present them to patient T cells. We have previously reported that, in a phase I pilot study (ClinicalTrials.gov NCT00095862), a subject with stage IV breast cancer experienced substantial regression of breast, lung, and brain lesions following inoculation with clinical formulations of SV-BR-1-GM, a GM-CSF-secreting breast tumor cell line. To identify diagnostic features permitting the prospective identification of patients likely to benefit from SV-BR-1-GM, we conducted a molecular analysis of the SV-BR-1-GM cell line and of patient-derived blood, as well as a tumor specimen. Compared to normal human breast cells, SV-BR-1-GM cells overexpress genes encoding tumor-associated antigens (TAAs) such as PRAME, a cancer/testis antigen. Curiously, despite its presumptive breast epithelial origin, the cell line expresses major histocompatibility complex (MHC) class II genes (HLA-DRA, HLA-DRB3, HLA-DMA, HLA-DMB), in addition to several other factors known to play immunostimulatory roles. These factors include MHC class I components (B2M, HLA-A, HLA-B), ADA (encoding adenosine deaminase), ADGRE5 (CD97), CD58 (LFA3), CD74 (encoding invariant chain and CLIP), CD83, CXCL8 (IL8), CXCL16, HLA-F, IL6, IL18, and KITLG. Moreover, both SV-BR-1-GM cells and the responding study subject carried an HLA-DRB3*02:02 allele, raising the question of whether SV-BR-1-GM cells can directly present endogenous antigens to T cells, thereby inducing a tumor-directed immune response. In support of this, SV-BR-1-GM cells (which also carry the HLA-DRB3*01:01 allele) treated with yellow fever virus (YFV) envelope (Env) 43-59 peptides reactivated YFV-DRB3*01:01-specific CD4+ T cells. Thus, the partial HLA allele match between SV-BR-1-GM and the clinical responder might have enabled patient T lymphocytes to directly recognize SV-BR-1-GM TAAs as presented on SV-BR-1-GM MHCs. Taken together, our findings are consistent with a potentially unique mechanism of action by which SV-BR-1-GM cells can act as APCs for previously primed CD4+ T cells.

Untreated HIV infection is associated with progressive CD4+ T cell depletion, which is generally recovered with combination antiretroviral therapy (cART). However, a significant proportion of cART-treated individuals have poor CD4+ T cell reconstitution. We investigated associations between HIV disease progression and CD4+ T cell glucose transporter-1 (Glut1) expression. We also investigated the association between these variables and specific single nucleotide polymorphisms (SNPs) within the Glut1 regulatory gene AKT (rs1130214, rs2494732, rs1130233, and rs3730358) and in the Glut1-expressing gene SLC2A1 (rs1385129 and rs841853) and antisense RNA 1 region SLC2A1-AS1 (rs710218). High CD4+Glut1+ T cell percentage is associated with rapid CD4+ T cell decline in HIV-positive treatment-naïve individuals and poor T cell recovery in HIV-positive individuals on cART. Evidence suggests that poor CD4+ T cell recovery in treated HIV-positive individuals is linked to the homozygous genotype (GG) associated with SLC2A1 SNP rs1385129 when compared to those with a recessive allele (GA/AA) (odds ratio = 4.67; P = 0.04). Furthermore, poor response to therapy is less likely among Australian participants when compared against American participants (odds ratio: 0.12; P = 0.01) despite there being no difference in prevalence of a specific genotype for any of the SNPs analyzed between nationalities. Finally, CD4+Glut1+ T cell percentage is elevated among those with a homozygous dominant genotype for SNPs rs1385129 (GG) and rs710218 (AA) when compared to those with a recessive allele (GA/AA and AT/TT respectively) (P < 0.04). The heterozygous genotype associated with AKT SNP 1130214 (GT) had a higher CD4+Glut1+ T cell percentage when compared to the dominant homozygous genotype (GG) (P = 0.0068). The frequency of circulating CD4+Glut1+ T cells and the rs1385129 SLC2A1 SNP may predict the rate of HIV disease progression and CD4+ T cell recovery in untreated and treated infection, respectively.

Tolerance is a long-recognized property of macrophages that leads to an altered response to repeated or chronic exposure to endotoxin. The physiological role of tolerance is to limit the potential damage to host tissue that may otherwise result from prolonged production of pro-inflammatory cytokines. Tolerance is induced by all toll-like receptor (TLR) ligands tested to date, however, tolerance induced by the TLR4 ligand lipopolysaccharide (LPS) is by far the best studied. LPS tolerance involves a global transcriptional shift from a pro-inflammatory response toward one characterized by the expression of anti-inflammatory and pro-resolution factors. Although largely reversible, LPS-tolerance leads to a hybrid macrophage activation state that is pro-inflammatory in nature, but possesses distinct regulatory anti-inflammatory features. Remarkably, a comparative transcriptomic analysis of tolerance induced by different TLR ligands has not previously been reported. Here, we describe the transcriptomic profiles of mouse macrophages tolerized with ligands for TLR2, TLR3, TLR4 and TLR 9. While we identified TLR-specific transcriptional profiles in macrophages tolerized with each ligand, tolerance induced by TLR4 represented an archetype pattern, such that each gene tolerized by any of the TLRs tested was also found to be tolerized by TLR4. Pro-inflammatory cytokines are not universally suppressed in all tolerant cells, but distinct patterns of cytokine expression distinguished TLR-specific tolerance. Analysis of gene regulatory regions revealed specific DNA sequence motifs associated with distinct states of TLR tolerance, implicating previously identified as well as novel transcriptional regulators of tolerance in macrophages. These data provide a basis for the future exploitation of TLR-specific tolerant states to achieve therapeutic re-programming of the innate immune response.

Multiple sclerosis (MS) and experimental autoimmune encephalomyelitis (EAE) have been proved to be T cell-mediated autoimmune diseases. Recent researches indicate that humoral immunity is also involved in the pathogenesis of these disorders. T follicular helper (Tfh) cells are critical for B cell differentiation and antibody production. However, the role of Tfh cells in MS and EAE remains unclear. Here, we found elevated frequencies of CD4+CXCR5+PD-1+ Tfh-like cells in both MS patients and EAE. In EAE mice, Tfh-like cells, together with B cells, were found in the ectopic lymphoid structures in spinal cords. Moreover, Tfh-like cells promoted the antibody production via IL-21/IL-21R and CD40 ligand/CD40 interaction and the synergy effect of STAT3 and non-canonical NF-κB signaling pathway inside B cells. Moreover, adoptive transfer of Tfh-like cells could increase the severity and delay the remission of EAE. In conclusion, our data indicate that Tfh-like cells contribute to the pathogenesis of EAE.