Clinical and radiological features of immune checkpoint inhibitor-related pneumonitis in lung cancer and non-lung cancers.

OBJECTIVE: To investigate the clinical and radiological features of immune checkpoint inhibitor-related pneumonitis (ICI-P), a rare but serious pulmonary complication of cancer immunotherapy and to evaluate key differences between lung cancer (LC) and non-LC patients. METHODS: 247 patients (LC, n = 151) treated with ICI for malignancies were retrospectively screened in a single institute. The number of patients, history of other immune-related adverse events (irAE), the onset, serum KL-6 levels, and chest CT features (types of pneumonitis, symmetry, laterality, location) were recorded for the ICI-P population and compared for LC and non-LC groups. RESULTS: ICI-P was identified in 26 patients in total (LC, n = 19; non-LC, n = 7). The incidence of other irAE was significantly higher in ICI-P group (63%) compared with patients without ICI-P (34%) (p = 0.0056). An earlier onset of ICI-P was recorded in LC (78 days) compared to non-LC patients (186 days) (p = 0.0034). Serum KL-6 was significantly elevated only in the non-LC group when ICI-P was noticed (p = 0.029). Major CT findings of ICI-P, irrespective of primary disease, were organizing pneumonia pattern and ground glass opacities. LC patients commonly exhibited consolidation and traction bronchiectasis and were prone to asymmetrical shadows (p < 0.001). Non-LC patients were more likely to exhibit symmetrical infiltrations. A small fraction of both groups experienced relapse or moving patterns of ICI-P. CONCLUSION: ICI-P patients more often experienced other irAE prior to the development of ICI-P. The characteristics of ICI-P can differ in terms of the onset, KL-6 reliability, and chest CT findings between LC and non-LC patients. ADVANCES IN KNOWLEDGE: In ICI-P patients, a history of other irAE can be more frequently observed. Differences in disease onset and radiological patterns between LC and non-LC patients might be helpful to make a diagnosis of ICI-P; however, longitudinal observation of chest CT scans is advised to observe the pneumonitis activity irrespective of cancer types.

Gold nanoparticle triggered siloxane formation for polymerization based amplification in enzyme free visual immunoassay.

Herein, we report a new signal amplification scheme for quantitative biochemical analysis based on gold nanoparticle (GNPs) catalyzed polymerization of transparent silane solution to milky white and turbid siloxane. Using immunoassay as a proof of concept, GNP labeled immunoprobe was used to bind captured antigen and catalyse the polymerization reaction allowing sensitive biochemical investigation. The polymerization reaction was optimized for standard 96 well polystyrene microtiter plates and we discovered that sodium lactate acts as an enhancer in the polymerization reaction as it reduces detection time to merely 30 min. The sensing strategy was applied to detection and quantification of Salmonella Typhimurium in water and egg samples and the platform showed excellent visibly quantifiable analytical response up to 100 cells mL-1. Furthermore, clinical utility and potential of the method was validated by detecting Vi capsular polysaccharide (Vi antigen) responsible for typhoidal Salmonellosis in human serum in sandwich format with a detection limit of 1 ng mL-1. The method serves as the first report towards nanoparticle triggered polymerization for development of rapid and low cost quantitative biochemical assay.

Carbohydrate Specificity and Isotypes of Monoclonal and Polyclonal Antibodies to Conjugated Tetrasaccharide, a Synthetic Analogue of Repeating Unit of Capsular Polysaccharide of Streptococcus Pneumoniae Serotype 14.

We studied carbohydrate specificity and isotypes of antibodies to BSA-conjugated tetrasaccharide, a repeating unit of the capsular polysaccharide of Streptococcus pneumoniae serotype 14, in mouse polyclonal sera and hybridoma-synthesized products. Natural IgM antibodies to the tetrasaccharide containing epitopes similar to surface carbohydrate structures of mammalian and human cells in low titers were determined in native mouse serum by ELISA using biotinylated tetrasaccharide and synthetic capsular polysaccharide as the solid-phase antigens. Polyclonal sera to the conjugated tetrasaccharide contained IgM and all subclasses of IgG antibodies, which were detected in a higher titer when the biotinylated tetrasaccharide was used as a solid phase antigen compared to synthetic capsular polysaccharide. Monoclonal antibodies to S. pneumoniae serotype 14 tetrasaccharide were identified in an equivalent titer using either biotinylated tetrasaccharide or synthetic capsular polysaccharide. Monoclonal antibodies obtained in vitro belonged to IgM isotype and cross-reacted with secondary full-size IgG antibodies. In the serum of mice inoculated with hybridoma, IgM and IgG2a antibodies recognizing the tetrasaccharide epitope in the structure of synthetic capsular polysaccharide were simultaneously determined.

Effects of daily consumption of psyllium, oat bran and polyGlycopleX on obesity-related disease risk factors: A critical review.

The persistent obesity crisis, with its increased risk for the metabolic syndrome (MetS), type 2 diabetes, and cardiovascular disease (CVD), continues to damage the health of populations globally, including children. Diets rich in the fiber provided by fruit and vegetables support good metabolic health, although few adults and children achieve the recommended daily target. Daily fiber supplementation, particularly with soluble fiber products, such as psyllium, oat bran, or a newer product such as PolyGlycopleX, may provide a convenient solution. Literature searches were conducted to identify original research articles, systematic reviews, and meta-analyses with the search terms psyllium, oat bran, PolyGlycopleX, and PGX, AND adults and children AND overweight, obesity, and metabolic syndrome. Data source was Embase and PubMed from 1980 to 2017. The results show that the addition of a soluble fiber product, most notably psyllium, improves blood lipid profiles, particularly total and low-density lipoprotein cholesterol, as well as glycemic response, and increases satiety, and by thus improving MetS and CVD risk factors, may augment the processes initiated by weight reduction diets. Although less studied than psyllium, the available evidence has shown that ß-glucan present in oat bran has a beneficial effect on MetS and CVD risk factors, particularly blood lipids and glycemia. Early research has found PolyGlycopleX to provide similar benefits to other soluble fiber products, and suggest it may also assist with weight loss. This critical review demonstrates that soluble fiber supplements used as an adjunct to dietary and lifestyle modifications may assist with the treatment of CVD and MetS risk factors. More research is needed to further clarify the benefits of PolyGlycopleX in particular, as well as to develop safe and efficacious recommendations for fiber supplementation of all types for children in general.

Aptamer functionalized MoS2-rGO nanocomposite based biosensor for the detection of Vi antigen.

We report a novel aptamer functionalized MoS2-rGO based electrochemical method for Vi polysaccharide antigen mediated detection of enteric fever. Herein, highly selective anti-Vi aptamers were screened from a pool of oligonucleotides using a microtitre based SELEX approach and characterized for its specificity and stability. The MoS2-rGO nanocomposite was synthesized using a liquid assisted exfoliation by taking optimum ratio of MoS2 and rGO. The nanocomposite presented synergistic effect owing to easy biomolecular functionalization and enhanced conductivity. The screened anti-Vi aptamers were embedded on the MoS2-rGO nanocomposite via thiol linkage to give a stable biointerface. The developed aptasensor was characterized and further evaluated for its performance with different concentrations of Vi antigen using ferrocene labeled boronic acid as an electroactive probe. The aptasensor responded linearly in the range between 0.1 ng mL-1 to 1000 ng mL-1with a detection limit of 100 pg mL-1, and did not show any cross-reactivity with other bacterial polysaccharides indicating high specificity. The applicability of the developed aptasensor was further validated in urine and sera specimens spiked with Vi antigen.

In vivo Distribution and Clearance of Purified Capsular Polysaccharide from Burkholderia pseudomallei in a Murine Model.

Burkholderia pseudomallei is the causative agent of melioidosis, a severe infection prominent in northern Australia and Southeast Asia. The "gold standard" for melioidosis diagnosis is bacterial isolation, which takes several days to complete. The resulting delay in diagnosis leads to delayed treatments, which could result in death. In an attempt to develop better methods for early diagnosis of melioidosis, B. pseudomallei capsular polysaccharide (CPS) was identified as an important diagnostic biomarker. A rapid lateral flow immunoassay utilizing CPS-specific monoclonal antibody was developed and tested in endemic regions worldwide. However, the in vivo fate and clearance of CPS has never been thoroughly investigated. Here, we injected mice with purified CPS intravenously and determined CPS concentrations in serum, urine, and major organs at various intervals. The results indicate that CPS is predominantly eliminated through urine and no CPS accumulation occurs in the major organs. Immunoblot analysis demonstrated that intact CPS was excreted through urine. To understand how a large molecule like CPS was eliminated without degradation, a 3-dimenational structure of CPS was modeled. The predicted CPS structure has a rod-like shape with a small diameter that could allow it to flow through the glomerulus of the kidney. CPS clearance was determined using exponential decay models and the corrected Akaike Information Criterion. The results show that CPS has a relatively short serum half-life of 2.9 to 4.4 hours. Therefore, the presence of CPS in the serum and/or urine suggests active melioidosis infection and provides a marker to monitor treatment of melioidosis.

Simple alternative to sialic acid determination in meningococcal polysaccharides W or Y.

Physicochemical methods are the primary tests used to ensure that batches of meningococcal polysaccharide (PS) antigens are manufactured consistently to those shown to be safe and effective in clinical trials. Although modern physicochemical methods of analysis providing structural information about the antigens have been developed and used, simpler assays, which can be readily validated, are still in use for polysaccharide batch release. The simple and cheap method for Neisseria meningitidis serogroup W or Y polysaccharide (MenW or MenY PS) content quantification has been developed. This colorimetric method is based on the galactose or glucose quantification in MenW or MenY PS hydrolysate, respectively. Intra- and inter-assay precision and accuracy of the novel method have been demonstrated, in comparison to the same properties of the current regulatory approved method for the same purpose - sialic acid quantification. We provided the calculation of the possible future regulatory requirement for the galactose or glucose content in MenW or MenY PS, respectively, and revealed in detail the stoichiometric calculation behind it.

Differential Killing of Salmonella enterica Serovar Typhi by Antibodies Targeting Vi and Lipopolysaccharide O:9 Antigen.

Salmonella enterica serovar Typhi expresses a capsule of Vi polysaccharide, while most Salmonella serovars, including S. Enteritidis and S. Typhimurium, do not. Both S. Typhi and S. Enteritidis express the lipopolysaccharide O:9 antigen, yet there is little evidence of cross-protection from anti-O:9 antibodies. Vaccines based on Vi polysaccharide have efficacy against typhoid fever, indicating that antibodies against Vi confer protection. Here we investigate the role of Vi capsule and antibodies against Vi and O:9 in antibody-dependent complement- and phagocyte-mediated killing of Salmonella. Using isogenic Vi-expressing and non-Vi-expressing derivatives of S. Typhi and S. Typhimurium, we show that S. Typhi is inherently more sensitive to serum and blood than S. Typhimurium. Vi expression confers increased resistance to both complement- and phagocyte-mediated modalities of antibody-dependent killing in human blood. The Vi capsule is associated with reduced C3 and C5b-9 deposition, and decreased overall antibody binding to S. Typhi. However, purified human anti-Vi antibodies in the presence of complement are able to kill Vi-expressing Salmonella, while killing by anti-O:9 antibodies is inversely related to Vi expression. Human serum depleted of antibodies to antigens other than Vi retains the ability to kill Vi-expressing bacteria. Our findings support a protective role for Vi capsule in preventing complement and phagocyte killing of Salmonella that can be overcome by specific anti-Vi antibodies, but only to a limited extent by anti-O:9 antibodies.

Capsular Polysaccharide (CPS) Release by Serotype 3 Pneumococcal Strains Reduces the Protective Effect of Anti-Type 3 CPS Antibodies.

The efficacy of the serotype 3 (ST3) pneumococcal conjugate vaccine (PCV) remains unclear. While the synthesis of capsular polysaccharide (CPS) of most serotypes is wzy dependent, the strains of two serotypes, 3 and 37, synthesize CPS by the synthase-dependent pathway, resulting in a polysaccharide that is not covalently linked to peptidoglycan and can be released during growth. We hypothesized that the release of CPS during growth reduces anti-type 3 CPS antibody-mediated protection and may explain the lower efficacy of the type 3 component of PCV than that of other PCVs. The in vitro-released CPS concentrations per 10(7) CFU of ST3 and ST37 strains were significantly higher than those for the ST1, ST4, ST6B, and ST14 strains. Following intraperitoneal (i.p.) injection in mice, blood concentrations of CPS were significantly higher for the ST3 than for the ST4/5 strains. The opsonophagocytic killing assay (OPKA) titer of anti-type 3 CPS antibody was significantly reduced by type 3 CPS, culture supernatant, or serum from Streptococcus pneumoniae ST3 strain WU2-infected mice. Mice were injected with capsule-specific antibodies and challenged i.p. with or without the addition of sterile culture supernatant containing type-specific CPS. The addition of 0.2 µl of culture supernatant from WU2 inhibited passive protection, whereas 100-fold-more culture supernatant from S. pneumoniae ST4 strain TIGR4 was required for the inhibition of protection. We conclude that released type 3 CPS interferes with antibody-mediated killing and protection by anti-CPS antibodies. The relative failure of ST3 PCV may be due to CPS release, suggesting that alternative immunization approaches for ST3 may be necessary.

Pneumococcal polysaccharide vaccination in rheumatoid arthritis patients receiving tocilizumab therapy.

OBJECTIVES: We assessed the impact of tocilizumab (TCZ), a humanised monoclonal anti-interleukin-6 receptor antibody, on antibody response following administration of the 23-valent pneumococcal polysaccharide vaccine (PPV23). METHODS: A total of 190 patients with rheumatoid arthritis (RA) received PPV23. Patients were classified into TCZ (n=50), TCZ + methotrexate (MTX) (n=54), MTX (n=62) and RA control (n=24) groups. We measured serotype-specific IgG concentrations of pneumococcal serotypes 6B and 23F using ELISA and functional antibody activity using a multiplexed opsonophagocytic killing assay, reported as the opsonisation indices (OIs), before and 4-6 weeks after vaccination. Positive antibody response was defined as a 2-fold or more increase in the IgG concentration or as a ≥10-fold or more increase in the OI. RESULTS: IgG concentrations and OIs were significantly increased in all treatment groups in response to vaccination. The TCZ group antibody response rates were comparable with those of the RA control group for each serotype. MTX had a negative impact on vaccine efficacy. Multivariate logistic analysis confirmed that TCZ is not associated with an inadequate antibody response to either serotype. No severe adverse effect was observed in any treatment group. CONCLUSIONS: TCZ does not impair PPV23 immunogenicity in RA patients, whereas antibody responses may be reduced when TCZ is used as a combination therapy with MTX.

Purification and partial elucidation of the structure of an antioxidant carbohydrate biopolymer from the probiotic bacterium Bacillus coagulans RK-02.

An exopolysaccharide (EPS) was isolated from Bacillus coagulans RK-02 and purified by size exclusion chromatography. The purified, homogeneous EPS had an average molecular weight of ∼3 × 104 Da by comparison with FITC-labeled dextran standards. In vivo evaluations showed that, like other reported polysaccharides, this EPS displayed significant antioxidant activity. FTIR spectroscopy analysis showed the presence of hydroxy, carboxy, and α-glycosidic linkages and a mannose residue. GC analysis indicated that the EPS was a heteropolymer composed of glucose, mannose, galactose, glucosamine, and fucose as monomeric constituent units. Partial elucidation of the structure of the carbohydrate biopolymer based on GC-MS and NMR analysis showed the presence of two unique sets of tetrasaccharide repeating units that have 1→3 and 1→6 glycosidic linkages. This is also the first report of a Gram-positive bacterial polysaccharide with both fucose as a sugar monomer and 1→3 and 1→6 glycosidic linkages in the molecular backbone.

Endotoxins and other sepsis triggers.

Endotoxin, or more accurately termed bacterial lipopolysaccharide (LPS), is recognized as the most potent microbial mediator implicated in the pathogenesis of sepsis and septic shock. Yet despite its discovery well over a century ago, the fundamental role of circulating endotoxin in the blood of most patients with septic shock remains enigmatic and a subject of considerable controversy. LPS is the most prominent 'alarm molecule' sensed by the host's early warning system of innate immunity presaging the threat of invasion of the internal milieu by Gram-negative bacterial pathogens. In small doses within a localized tissue space, LPS signaling is advantageous to the host in orchestrating an appropriate antimicrobial defense and bacterial clearance mechanisms. Conversely, the sudden release of large quantities of LPS into the bloodstream is clearly deleterious to the host, initiating the release of a dysregulated and potentially lethal array of inflammatory mediators and procoagulant factors in the systemic circulation. The massive host response to this single bacterial pattern recognition molecule is sufficient to generate diffuse endothelial injury, tissue hypoperfusion, disseminated intravascular coagulation and refractory shock. Numerous attempts to block endotoxin activity in clinical trials with septic patients have met with inconsistent and largely negative results. Yet the groundbreaking discoveries within the past decade into the precise molecular basis for LPS-mediated cellular activation and tissue injury has rekindled optimism that a new generation of therapies that specifically disrupt LPS signaling might succeed. Other microbial mediators found in Gram-positive bacterial and viral and fungal pathogens are now appreciated to activate many of the same host defense networks induced by LPS. This information is providing novel interventions in the continuing effots to improve the care of septic patients.

[Assessment of the contribution of the immunochromatographic pneumococcal urinary antigen test to the etiological diagnosis of pneumonia in hospitalized adults]. / Bilan de la contribution de l'antigénurie pneumocoque par test immunochromatographique au diagnostic étiologique des pneumonies chez l'adulte hospitalisé.

AIM OF THE STUDY: To assess the usefulness and prescription practices of the Binax Now Streptococcus pneumoniae urinary antigen test in hospitalized adults. PATIENTS AND METHODS: The results of the pneumococcal urinary antigen tests (UAT) performed from January 2002 to September 2004 were related to that of microbiological cultures, and in positive patients to radiographic findings and C-reactive protein (CRP) levels. The evolution of the number of prescriptions and positivity rate in 2007 versus 2002-2004 was analyzed. RESULTS: The pneumococcal UAT was positive in 32 of the 278 patients included from 2002 to 2004 (11.5%). Results were concordant with that of microbiological cultures in 90% of the 247 documented cases. Pneumococcal etiology was considered to be definite in 19 patients (isolation of S. pneumoniae from blood, 17 patients; or pleural fluid, two patients), of whom 15 had a positive UAT (sensitivity: 79%); to be probable in 22 patients (positive UAT, 17 patients and/or isolation of S. pneumoniae from respiratory samples, six patients), and was retained in 39 of the 41 patients (positive predictive value: 93.7%). CRP was greater than 100mg/L in 34 of 39 documented patients and lobar alveolar radiographic opacities observed in 25 of 28 documented patients. In 2007, the dramatic increase in the number of UAT prescriptions and the diversification of prescribing units were associated to a decreased positivity rate (8.1%). CONCLUSION: Whereas the pneumococcal UAT clearly increases etiological diagnosis, pneumococcal pneumonia cannot be ruled out if negative. Indications for its use need to be refined to improve the cost-effectiveness of this test.

Inhalative vaccination with pneumococcal polysaccharide in patients with chronic obstructive pulmonary disease.

In order to determine the feasibility of inhalative vaccination with polysaccharide antigen in patients with chronic obstructive pulmonary disease (COPD), we used controlled inhalation of a defined dose of Pneumovax in a randomized 3-arm study. The vaccine was either deposited in the alveoli (alveolar vaccination) or in the large airways (bronchial vaccination) and these were compared to standard intramuscular vaccination. Adverse effects were minor and never exceeded WHO grade 2. There was frequent cough, headache and shivering in the bronchial vaccination group, frequent fatigue only in the alveolar vaccination group and no frequent adverse effects in the intramuscular vaccination group. Specific serum IgG antibody was measured before and at 4 and 12 weeks after vaccination. At 12 weeks there was a greater than twofold rise in 7 out of 10 individuals in every vaccination group. Mean antibody levels of responders at 12 weeks were 278 mg/l for alveolar vaccination, 238 mg/l for bronchial vaccination and 737 mg/l for standard intramuscular vaccination. The data show that polysaccharide vaccine can be safely administered by controlled inhalation in COPD patients and that it can induce a rapid serum antibody response.

Critical differences between pneumococcal polysaccharide enzyme-linked immunosorbent assays with and without 22F inhibition at low antibody concentrations in pediatric sera.

A comparative study was conducted between two laboratories in order to evaluate the differences between two enzyme-linked immunosorbent assay (ELISA) techniques for the detection of pneumococcal anti-capsular polysaccharide antibodies. One laboratory used an assay including heterologous 22F polysaccharide inhibition, and the other laboratory employed a non-22F reference assay. After conjugate immunization, 30 pediatric post-primary immunization sera with antipolysaccharide concentrations ranging from <0.05 to 15 mug/ml were analyzed. Aggregate reverse cumulative distribution curves combining concentrations of antibodies against serotypes 4, 6B, 9V, 14, 18C, 19F, and 23F revealed similar results for both methods at antibody levels of >1 microg/ml. However, at antibody levels of <1 microg/ml, the distribution curve measured with the 22F inhibition ELISA shifted toward lower levels. This observation suggests that the 22F inhibition assay is more specific at low antibody concentrations, which was confirmed by heterologous polysaccharide inhibition experiments. Translation of low antibody levels suggested that the proposed threshold concentration of 0.35 mug/ml determined with the non-22F ELISA corresponded to a concentration of 0.20 mug/ml with the 22F inhibition ELISA. Pneumococcal antipolysaccharide ELISA including 22F inhibition can be recommended as a reference method.

Rapid detection of pneumococcal antigen in serum samples for diagnosing pneumococcal pneumonia.

OBJECTIVES: The aim of the study is to assess the usefulness of C polysaccharide and polysaccharide capsular antigen detection by immunochromatography (ICT) and enzyme immunoassay (EIA), respectively, in serum samples for diagnosing pneumococcal pneumonia. METHODS: Adult patients included in the study were classified in the following groups: In group 1 we studied 101 serum samples from patients with pneumonia due to Streptococcus pneumoniae. In 53 cases the pneumonia was bacteremic. The second group contained 113 serum samples from patients with no pneumococcal pneumonia. Group 3 was made up of 40 serum samples from healthy subjects with no clinical or radiological signs of pneumonia. RESULTS: Using ICT, antigen was detected in 50% of patients with pneumococcal pneumonia, in 64.3 and 40.9% of patients with bacteremic and non-bacteremic pneumococcal pneumonia, respectively. Using EIA, antigens were detected in 35.8% of patients with pneumococcal pneumonia, in 45 and 22.2% of patients with bacteremic and non-bacteremic pneumococcal pneumonia, respectively. CONCLUSIONS: In conclusion, the sensitivity of the tests is low. However, in special situations, where obtaining large volume of urine is difficult, they could be a complementary method in the rapid diagnosis of pneumococcal pneumonia.

Doubly branched hexasaccharide epitope on the cell wall polysaccharide of group A streptococci recognized by human and rabbit antisera.

A number of epitope specificities associated with the cell wall polysaccharide antigen of group A streptococci were identified in a polyclonal rabbit antiserum induced in rabbits by whole group A streptococci and in polyclonal convalescent human antisera from children that had recovered from streptococcal A infections. The identification was achieved by using a series of synthetic oligosaccharides, glycoconjugates, and bacterial polysaccharide inhibitors to inhibit the binding of the group A helical polysaccharide to the polyclonal antisera. The exclusively dominant epitope expressed in the convalescent human antisera was the doubly branched extended helical hexasaccharide with the structure alpha-L-Rhap(1-->2)[beta-D-GlcpNAc(1-->3)]alpha-L-Rhap(1-->3)alpha-L-Rhap(1-->2)[beta-D-GlcpNAc(1-->3)]alpha-L-Rhap. The hexasaccharide epitope also bound with the highest immunoreactivity to the rabbit antiserum. In contrast, the human antisera did not show significant binding to the singly branched pentasaccharide with the structure alpha-L-Rhap(1-->2)alpha-L-Rhap(1-->3)alpha-L-Rhap(1-->2)[beta-D-GlcpNAc(1-->3)]alpha-L-Rhap or the branched trisaccharide alpha-L-Rhap(1-->2)[beta-D-GlcpNAc(1-->3)]alpha-l-Rhap, although both these haptens bound significantly to the same rabbit antiserum, albeit with less immunoreactivity than the hexasaccharide. Inhibition studies using streptococcal group A and B rabbit antisera and the inhibitors indicated above also suggested that the group A carbohydrate, unlike the group B streptococcal polysaccharide, does not contain the disaccharide alpha-L-Rhap(1-->2)alpha-L-Rhap motif at its nonreducing chain terminus, stressing the importance of mapping the determinant specificities of these two important streptococcal subcapsular group polysaccharides to fully understand the serological relationships between group A and group B streptococci.