[Show abstract][Hide abstract] ABSTRACT: Procalcitonin (PCT) can discriminate bacterial from viral systemic infections and true bacteremia from contaminated blood cultures. The aim of this study was to evaluate PCT diagnostic accuracy in discriminating Gram-positive, Gram-negative, and fungal bloodstream infections. A total of 1,949 samples from patients with suspected bloodstream infections were included in the study. Median PCT value in Gram-negative (13.8 ng/mL, interquartile range (IQR) 3.4-44.1) bacteremias was significantly higher than in Gram-positive (2.1 ng/mL, IQR 0.6-7.6) or fungal (0.5 ng/mL, IQR 0.4-1) infections (P < 0.0001). Receiver operating characteristic analysis showed an area under the curve (AUC) for PCT of 0.765 (95% CI 0.725-0.805, P < 0.0001) in discriminating Gram-negatives from Gram-positives at the best cut-off value of 10.8 ng/mL and an AUC of 0.944 (95% CI 0.919-0.969, P < 0.0001) in discriminating Gram-negatives from fungi at the best cut-off of 1.6 ng/mL. Additional results showed a significant difference in median PCT values between Enterobacteriaceae and nonfermentative Gram-negative bacteria (17.1 ng/mL, IQR 5.9-48.5 versus 3.5 ng/mL, IQR 0.8-21.5; P < 0.0001). This study suggests that PCT may be of value to distinguish Gram-negative from Gram-positive and fungal bloodstream infections. Nevertheless, its utility to predict different microorganisms needs to be assessed in further studies.
[Show abstract][Hide abstract] ABSTRACT: Objective
Regulatory T cells (Treg) play a critical role in the prevention of autoimmunity, and the suppressive activity of these cells is impaired in rheumatoid arthritis (RA). The aim of the present study was to investigate function and properties of Treg of RA patients in response to purified polysaccharide glucuronoxylomannogalactan (GXMGal).
Flow cytometry and western blot analysis were used to investigate the frequency, function and properties of Treg cells.
GXMGal was able to: i) induce strong increase of FOXP3 on CD4+ T cells without affecting the number of CD4+CD25+FOXP3+ Treg cells with parallel increase in the percentage of non-conventional CD4+CD25−FOXP3+ Treg cells; ii) increase intracellular levels of TGF-β1 in CD4+CD25−FOXP3+ Treg cells and of IL-10 in both CD4+CD25+FOXP3+ and CD4+CD25−FOXP3+ Treg cells; iii) enhance the suppressive activity of CD4+CD25+FOXP3+ and CD4+CD25−FOXP3+ Treg cells in terms of inhibition of effector T cell activity and increased secretion of IL-10; iv) decrease Th1 response as demonstrated by inhibition of T-bet activation and down-regulation of IFN-γ and IL-12p70 production; v) decrease Th17 differentiation by down-regulating pSTAT3 activation and IL-17A, IL-23, IL-21, IL-22 and IL-6 production.
These data show that GXMGal improves Treg functions and increases the number and function of CD4+CD25−FOXP3+ Treg cells of RA patients. It is suggested that GXMGal may be potentially useful for restoring impaired Treg functions in autoimmune disorders and for developing Treg cell-based strategies for the treatment of these diseases.
[Show abstract][Hide abstract] ABSTRACT: Background:
Procalcitonin (PCT) levels can be used to predict bacteremia and DNAemia in patients with sepsis. In this study, the diagnostic accuracy of PCT in predicting blood culture (BC) results and DNAemia, as detected by real-time PCR (RT-PCR), was compared with that of other markers of inflammation commonly evaluated in patients with suspected sepsis, such as C-reactive protein (CRP), erythrocyte sedimentation rate (ESR), and white blood cell (WBC) count.
A total of 571 patients for whom BC, blood RT-PCR, PCT, CRP, ESR, and WBC count were requested for laboratory diagnosis of sepsis were included in the study. Receiver operating characteristic curve analysis was performed to compare the ability of the above biomarkers to predict BC and blood RT-PCR results.
A total of 108 pathogens were identified by BC (79 pathogens, 14.5% positive rate) and/or RT-PCR (90 pathogens, 16.5% positive rate), after exclusion of 26 contaminated samples. The PCT areas under the curve (AUCs) in predicting BC (0.843; 95% CI 0.796-0.890; p < 0.0001) and RT-PCR (0.916; 95% CI 0.888-0.945; p < 0.0001) results were significantly greater than AUCs found for CRP, ESR, and WBC count.
PCT showed a better diagnostic accuracy than CRP, ESR, and WBC count in predicting DNAemia and bacteremia in patients with suspected sepsis.
No preview · Article · Sep 2014 · Infectious Diseases
[Show abstract][Hide abstract] ABSTRACT: Infective endocarditis (IE) is a life-threatening condition, burdened by high mortality. Current guidelines recommend that, in case of negative culture result, tissues from excised heart valves or vegetations from patients with suspected IE should be referred for broad-range bacterial PCR and sequencing. In this proof-of-concept study, the diagnostic utility of the commercially available multiplex real-time PCR system SeptiFast (SF), performed on cardiac valves, was evaluated in a selected population of 20 patients with definite IE of known origin, in comparison with culture. A significant difference was found between SF and culture in the rate of pathogen detection (19 versus 3 respectively; chi-square 14.06; P=0.0002). SF sensitivity was 95%; specificity, 100%; positive predictive value (PPV), 100%; and negative predictive value (NPV), 83.3%. Culture sensitivity was 15%; specificity, 100%; PPV, 100%; and NPV, 22.7%. SF assay, performed on culture-negative excised heart valves, can be useful for the etiological diagnosis of IE.
Full-text · Article · May 2014 · Diagnostic microbiology and infectious disease
[Show abstract][Hide abstract] ABSTRACT: In healthy subjects, Cytomegalovirus infection can be asymptomatic or manifest as mononucleosis syndrome, but organ disease has also been reported. However, in immunocompromised patients this infection can lead to its most significant and severe disease and even mortality. When Cytomegalovirus causes a gastrointestinal tract infection, it more commonly manifests with luminal tract disease and is usually characterized by ulcerative lesions. Appendicitis is a rare manifestation, and has been reported mainly in human immunodeficiency virus-infected patients or patients with other causes of immunocompromise.
The authors report on a case of acute primary Cytomegalovirus infection complicated with acute appendicitis due to Cytomegalovirus in an apparently immunocompetent 24-year-old Caucasian man also suffering from primary sclerosing cholangitis and ulcerative colitis. Diagnosis was based on clinical manifestations, serology results, as well as microbiological and histological findings. Treatment consisted of surgery and anti-Cytomegalovirus therapy.
Cytomegalovirus should be included among the etiologic agents of acute appendicitis in patients with primary sclerosing cholangitis and ulcerative colitis. Currently, there are no definitive data regarding the frequency of Cytomegalovirus appendicitis and the role of anti-Cytomegalovirus treatment in human immunodeficiency virus-negative and apparently immunocompetent subjects.
Full-text · Article · Mar 2014 · Journal of Medical Case Reports
[Show abstract][Hide abstract] ABSTRACT: The encapsulated fungal pathogen Cryptococcus neoformans represents a significant agent of life-threatening infections in immunocompromised subjects. A unique characteristic of Cryptococcus species is the presence of a polysaccharide capsule, which is essential for virulence and endows Cryptococcus with potent immunoregulatory properties. This review provides an overview of the immunological properties of the principal components of C. neoformans capsule.
[Show abstract][Hide abstract] ABSTRACT: Sepsis is a syndrome characterized by a systemic inflammatory response due to severe infection. Early detection of causal agents and appropriate antimicrobial treatment reduce mortality. Conventional microbiological methods often do not provide time critical results for an optimal early management. We used an in-house protocol based on Tween 80 to process 109 positive blood cultures for bacteria and yeast identification by matrix-assisted laser desorption/ionization-time-of-flight mass spectrometry (MALDI-TOF MS), and results were compared to standard reference or automated methods. MALDI-TOF MS correctly identified 91.7% of the isolates. Correct identification was obtained for 57/62 (91.9%) aerobic/facultative anaerobic Gram-positive isolates, 53 (85.5%) at species level, and 4 (6.4%) at the genus level; 32/32 (100%) aerobic/facultative anaerobic Gram-negative isolates, 31 (96.9%) at species level, and 1 (3.1%) at the genus level; 7/7 (100%) obligate anaerobes, all at the genus level; 3/7 (42.8%) fungi, all at genus level. Overall, the median identification time of MALDI-TOF MS vs reference standard methods was significantly shorter: median (interquartile range) 7.1h (4.7-10.2) vs 48.1h (32.5-50.0), p<0.0001. MALDI-TOF MS is a valuable tool for rapid identification of pathogens in septic patients. An in-house protocol based on Tween 80 can be used to process positive blood cultures.
No preview · Article · Mar 2013 · International journal of medical microbiology: IJMM
[Show abstract][Hide abstract] ABSTRACT: The persistence of activated T cells in rheumatoid arthritis (RA) synovium may be attributable to increased homing, increased retention or a possible imbalance between cell proliferation and programmed cell death. Induction of apoptosis may represent a potential therapeutic approach. Galactoxylomannan (GalXM) from the opportunistic fungus Cryptococcus neoformans can interact with T cells and induce T-cell apoptosis through the inhibition of CD45 phosphatase activity. The aim of this study was to determine the effect of GalXM on circulating T cells from patients with RA and the underlying mechanisms. GalXM immunomodulating effect on apoptosis and signal transduction pathway involved in IL-17A production was evaluated on T cells. RA T-cell apoptosis, higher than that of control T cells, was further increased by GalXM through induction of caspase-3 activation. Activated T cells expressing the CD45RO molecule and producing IL-17A were the main target of GalXM-induced apoptosis. GalXM induced consistent impairment of IL-17A production and inhibition of STAT3, which was hyperactivated in RA. In conclusion, GalXM triggered apoptosis of activated memory T cells and interfered with IL-17A production in RA. These data suggest therapeutic targeting of deleterious Th17 cells in RA and other autoimmune diseases.
[Show abstract][Hide abstract] ABSTRACT: We previously reported that a synthetic peptide with sequence identical to a CDR of a mouse monoclonal antibody specific for difucosyl human blood group A exerted an immunomodulatory activity on murine macrophages. It was therapeutic against systemic candidiasis without possessing direct candidacidal properties. Here we demonstrate that a selected peptide, N10K, putatively deriving from the enzymatic cleavage of the constant region (Fc) of human IgG(1), is able to induce IL-6 secretion and pIkB-α activation. More importantly, it causes an up-regulation of Dectin-1 expression. This leads to an increased activation of β-glucan-induced pSyk, CARD9 and pIkB-α, and an increase in the production of pro-inflammatory cytokines such as IL-6, IL-12, IL-1β and TNF-α. The increased activation of this pathway coincides with an augmented phagocytosis of non opsonized Candida albicans cells by monocytes. The findings suggest that some Fc-peptides, potentially deriving from the proteolysis of immunoglobulins, may cause an unexpected immunoregulation in a way reminiscent of innate immunity molecules.
[Show abstract][Hide abstract] ABSTRACT: Infective endocarditis (IE) is a life-threatening condition in which identification of the causative pathogen is pivotal for subsequent effective therapy. Blood culture (BC) is considered the gold standard for microbial diagnosis of IE, but in up to one-third of patients, it can be negative due to a number of possible factors, e.g., prior antimicrobial therapy, fastidious microorganisms, or low bacterial load. The aim of this study was to assess the diagnostic utility of the commercially available PCR based system SeptiFast (SF) for microbial diagnosis of IE from clinical blood specimens. The results indicate that, this test, in combination with BC, could represent a valuable tool for microbial diagnosis of IE. Reduction of SF detection limit set for coagulase-negative staphylococci and streptococci could improve the sensitivity of the test.
Full-text · Article · Mar 2012 · Journal of Medical Microbiology
[Show abstract][Hide abstract] ABSTRACT: Early identification of causative pathogen in sepsis patients is pivotal to improve clinical outcome. SeptiFast (SF), a commercially
available system for molecular diagnosis of sepsis based on PCR, has been mostly used in patients hospitalized in hematology
and intensive care units. We evaluated the diagnostic accuracy and clinical usefulness of SF, compared to blood culture (BC),
in 391 patients with suspected sepsis, hospitalized in a department of internal medicine. A causative pathogen was identified
in 85 patients (22%). Sixty pathogens were detected by SF and 57 by BC. No significant differences were found between the
two methods in the rates of pathogen detection (P = 0.74), even after excluding 9 pathogens which were isolated by BC and were not included in the SF master list (P = 0.096). The combination of SF and BC significantly improved the diagnostic yield in comparison to BC alone (P < 0.001). Compared to BC, SF showed a significantly lower contamination rate (0 versus 19 cases; P < 0.001) with a higher specificity for pathogen identification (1.00, 95% confidence interval [CI] of 0.99 to 1.00, versus
0.94, 95% CI of 0.90 to 0.96; P = 0.005) and a higher positive predictive value (1.00, 95% CI of 1.00 to 0.92%, versus 0.75, 95% CI of 0.63 to 0.83; P = 0.005). In the subgroup of patients (n = 191) who had been receiving antibiotic treatment for ≥24 h, SF identified more pathogens (16 versus 6; P = 0.049) compared to BC. These results suggest that, in patients with suspected sepsis, hospitalized in an internal medicine
ward, SF could be a highly valuable adjunct to conventional BC, particularly in patients under antibiotic treatment.
Full-text · Article · Feb 2012 · Journal of clinical microbiology
[Show abstract][Hide abstract] ABSTRACT: Early detection of aetiological agents is pivotal for adequate therapy for bacterial infections. Although culture is still considered the mainstay for laboratory diagnosis, it often lacks sensitivity, especially in patients already treated with antibiotics. The present study investigated the potential clinical utility of the commercial real-time-PCR-based system SeptiFast (SF), originally intended for diagnosis of sepsis from blood specimens, in the aetiological diagnosis of other bacterial infections, in patients undergoing antibiotic therapy. A total of 53 non-blood specimens were analysed for microbial pathogen detection by conventional culture and with SF real-time PCR: 19 (35.8%) synovial fluids, 9 (17.0%) cardiac valve tissues and 25 (47.2%) purulent exudates from various body sites. Overall, the number of specimens positive for a pathogen by SF (26/53; 49.1%) was significantly greater (P=0.001) than that of specimens positive by culture (10/53; 18.9%). In particular, SF was superior to culture for pathogen detection in cardiac valve tissues and synovial fluids. The analysis of concordance showed a fair agreement between the two methods (kappa value=0.314; 95% confidence interval=0.531-0.097). Even with the limitation of the low number of specimens, this study confirmed the great potential of diagnosing bacterial infections by a molecular approach, and indicates that the real-time PCR SF system can be used for specimens other than blood, from patients undergoing antibiotic treatment.
Preview · Article · Aug 2011 · Journal of Medical Microbiology
[Show abstract][Hide abstract] ABSTRACT: Cryptococcus neoformans is an opportunistic fungal pathogen responsible for life-threatening infections in immunocompromised individuals and occasionally in those with no known immune impairment. The fungus is endowed with several virulence factors, including capsular polysaccharides that play a key role in virulence. The capsule is composed of 90-95% glucuronoxylomannan (GXM), 5-8% galactoxylomannan (GalXM) and <1% mannoproteins. Capsular polysaccharides are shed into tissue where they produce many deleterious effects. Since GalXM has a smaller molecular mass, the molar concentration of GalXM in polysaccharide that is shed could exceed that of GXM in C. neoformans exopolysaccharides. Moreover, GalXM exhibits a number of unusual biologic properties both in vitro and in vivo. Here, we summarize the principal immunomodulatory effects of GalXM described during the last 20 years, particularly the mechanisms leading to induction of apoptosis in T lymphocytes, B lymphocytes and macrophages. Since the capacity of GalXM to induce widespread immune suppression is believed to contribute to the virulence of C. neoformans, this property might be exploited therapeutically to dampen the aberrant activation of immune cells during autoimmune disorders.
[Show abstract][Hide abstract] ABSTRACT: Breastfeeding has a major impact on CMV epidemiology. Postnatal CMV reactivation's incidence during lactation is nearby the maternal seroprevalence. Although perinatal CMV infection has practically no consequences in term newborn, it may cause, in some cases, a severe symptomatic disease in preterm newborns. The aims of the present study are to evaluate the rate and clinical expression of CMV infection breast milk transmitted in preterm infants and to check the safety of the freezing treated breast milk.
The study included fifty-seven preterm infants and their CMV seropositive mothers. Fresh breast milk samples have been collected from 1(st) to 9(th) postpartum week. Both fresh breast milk and 72, 96, 120 hours frozen samples have been examined, checking the presence of CMV; urine samples have been tested too.
70.2% of tested mothers showed reactivation of the infection, and CMV-positive breast milk during the six weeks postpartum has been found. However, only one infant was infected by CMV, developing hepatic affection concomitantly with a multi-system involvement, as shown CMV DNA detection in urine, saliva, blood, gastric aspirate, and stools.
Freezing breast milk at -20°C and pasteurization may respectively reduce or eliminate the viral load.
Full-text · Article · Jan 2011 · Italian Journal of Pediatrics
[Show abstract][Hide abstract] ABSTRACT: Previously, we reported that Galactoxylomannan (GalXM) activates the extrinsic and intrinsic apoptotic pathways through an interaction with the glycoreceptors on T cells. In this study we establish the role of the glycoreceptor CD45 in GalXM-induced T cell apoptosis, using CD45(+/+) and CD45(-/-) cell lines, derived from BW5147 murine T cell lymphoma. Our results show that whereas CD45 expression is not required for GalXM association by the cells, it is essential for apoptosis induction. In CD45(+/+) cells, CD45 triggering by GalXM reduces the activation of Lck, ZAP70 and Erk1/2. Conversely, in CD45(-/-) cells, Lck was hyperphosphorylated and did not show any modulation after GalXM stimulation. On the whole, our findings provide evidence that the negative regulation of Lck activation occurs via CD45 engagement. This appears to be related to the capacity of GalXM to antagonize T cell activation and induce T cell death. Overall this mechanism may be responsible for the immune paralysis that follows GalXM administration and could explain the powerful immunosuppression that accompanies cryptococcosis.
[Show abstract][Hide abstract] ABSTRACT: It has been documented that, independently from the specificity of the native antibody (Ab) for a given antigen (Ag), complementarity determining regions (CDR)-related peptides may display differential antimicrobial, antiviral and antitumor activities.
In this study we demonstrate that a synthetic peptide with sequence identical to V(H)CDR3 of a mouse monoclonal Ab (mAb) specific for difucosyl human blood group A is easily taken up by macrophages with subsequent stimulation of: i) proinflammatory cytokine production; ii) PI3K-Akt pathway and iii) TLR-4 expression. Significantly, V(H)CDR3 exerts therapeutic effect against systemic candidiasis without possessing direct candidacidal properties.
These results open a new scenario about the possibility that, beyond the half life of immunoglobulins, Ab fragments may effectively influence the antiinfective cellular immune response in a way reminiscent of regulatory peptides of innate immunity.
[Show abstract][Hide abstract] ABSTRACT: For many years the development of new azole antifungals has been quite empirically based. More recently, the publication of the crystal structure of CYP51 of Mycobacterium tuberculosis (MT-CYP51) provided new opportunities to rationalize the knowledge about antifungal action of this class of compounds. Recent studies reported that a 'channel 2 opened' conformation of the enzyme could better explain the interaction with ketoconazole (KTZ)-like drugs. Conformational changes were made on our model of Candida albicans CYP51 (CA-CYP51) previously reported and docking experiments were performed. The results allowed new KTZ analogues to be designed, by predicting that the 1,4-benzoxazine moiety could replace the KTZ aryl-piperazinyl chain. The synthesis of derivatives 12 and 13 was planned. The in vitro antifungal activity was evaluated against different Candida species and low and high capsulated strains of Cryptococcus neoformans. Since the in vitro activity do not necessarily correlate with the in vivo antifungal activity the newly synthesized compounds were also tested in a murine model of systemic C. albicans infection. The therapeutic effect was evaluated in terms of animal survival and of fungal growth in the kidneys, the target organ in systemic candidiasis.
[Show abstract][Hide abstract] ABSTRACT: The major virulence factor of Cryptococcus neoformans is its capsular polysaccharide, which is also released into tissues. The shed polysaccharide is composed of glucuronoxylomannan, galactoxylomannan (GalXM), and mannoproteins. In a previous study, we demonstrated a direct interaction of purified soluble GalXM with T cells that induced their apoptosis. In this study, we focus on the mechanisms involved in the apoptotic effect of GalXM. In our experimental system, we analyzed the effect of GalXM on purified human T cells and Jurkat cells, a T cell line routinely used for apoptotic studies. Our results reveal that GalXM activates the extrinsic and intrinsic apoptotic pathways through the cleavage and recruitment of caspase-8. Caspase-8 elicits the downstream executioner caspase-3, caspase-6, and caspase-7 both directly and indirectly, via Bid cleavage and caspase-9 activation. These effects appeared to be primarily mediated by the interaction of GalXM with the glycoreceptors, which differed in human T and Jurkat cells. CD45 was primarily involved in Jurkat cells apoptosis while CD7 and CD43 mediated human T cell apoptosis. Our results highlight a new mechanism by which a microbial product can contribute to virulence through direct interaction with T cell glycoreceptors, thereby triggering lymphocyte apoptosis.
Preview · Article · Jun 2009 · The Journal of Immunology
[Show abstract][Hide abstract] ABSTRACT: Dectin-1 is a pattern recognition receptor found on monocytes and dendritic cells (DC) able to recognize beta-1,3 and beta-1,6 glucans. It is thought to act via the spleen tyrosine kinase (Syk) to initiate immune response against infectious agents such as Candida albicans, one of the leading causes of invasive fungal disease in immunocompromised individuals. This study addresses the importance of this receptor in the context of human DC response to C. albicans. Upon blockage of Dectin-1, immature DC are less able than untreated cells to bind, phagocytose, and kill C. albicans via oxidative burst. In fact, a consistent decrease in superoxide anion, but not nitric oxide production, was manifested when the Syk pathway was inhibited. C. albicans-induced cytokine production via Dectin-1 recognition is mediated by the Syk activation pathway. Indeed, specific Syk inhibition significantly suppressed the production of IL-12, IL-6, and TNF-alpha. Finally, we observed that Dectin-1 engagement was also involved in DC maturation and subsequent lymphocyte activation. Collectively, these findings identify Dectin-1 as a key receptor influencing critical biological functions of DC in response to C. albicans leading to T cells response alteration. These effects are largely, though not completely, mediated by Syk activation.
No preview · Article · May 2009 · Microbes and Infection
[Show abstract][Hide abstract] ABSTRACT: The glucocorticoid-induced TNFR-related (GITR) protein is a member of the tumor necrosis factor receptor superfamily influencing natural and acquired immune response. GITR is activated by its ligand, GITRL, mainly expressed on antigen presenting cells. Previously, we demonstrated that GITR plays a role in regulating immune response to Candida albicans. Here we analyzed whether GITRL-GITR interaction influences the recognition of C. albicans by regulating the expression of pattern recognition receptors on splenic dendritic cells. Our report demonstrates that under physiological conditions and during candidiasis the GITRL-GITR system affects TLR-2 and TLR-4 expression on DC. These changes correlate with decrease in: MyD88 activation; CD80 and CD40 expression on DC; T cell activation response, including CD28 expression, IL-2 and IFN-gamma production. Our results point out that, during fungal infection, GITRL-GITR interaction modulates TLR-4 and TLR-2 expression, thereby altering the antigen presentation process, and suggesting a role of GITRL-GITR interaction in resistance against infectious diseases.