Immunogenicity and protective efficacy of DNA vaccines encoding MAP0586c and MAP4308c of Mycobacterium avium subsp. paratuberculosis secretome.

&lt;p&gt;Mycobacterium avium subsp. paratuberculosis (MAP), the etiological agent of chronic enteritis of the small intestine in domestic and wild ruminants, causes substantial losses to livestock industry. Control of this disease is seriously hampered by the lack of adequate diagnostic tools, vaccines and therapies. In this study, we have evaluated the vaccine potential of two MAP proteins, i.e. MAP0586c and MAP4308c, previously identified by postgenomic and immunoproteomic analysis of MAP secretome as novel serodiagnostic antigens. Immunizations of BALB/c and C57BL/6 mice with plasmid DNA encoding MAP0586c and MAP4308c induced strong Th1 type immune responses to both antigens, whereas antibody responses were only induced upon immunization with DNA encoding MAP4308c. Homologous boosting of DNA vaccinated mice with recombinant protein resulted in strong antibody responses against both proteins. Using synthetic overlapping peptides, immunodominant H-2(d) and H-2(b) restricted Th1 T cell epitopes were identified. Finally, MAP infected mice generated strong MAP0586c-specific T cell responses and MAP0586c DNA vaccination could protect BALB/c but not C57BL/6 mice against MAP challenge mice to the same extent as the Mycobacterium bovis BCG vaccine, indicating that this putative transglycosylase is an interesting vaccine candidate that warrants further investigation.&lt;/p&gt;</p

Bronchoalveolar lavage fluid peptidomics suggests a possible matrix metalloproteinase-3 role in bronchopulmonary dysplasia

Bronchoalveolar lavage fluid (BALF) is an important diagnostic source to investigate molecular changes occurring in lung disorders. The objective of this study was to assess and compare the peptidomic profiles of BALF from premature neonates with and without bronchopulmonary dysplasia (BPD). Samples were obtained on the 3rd day of life from 34 neonates with gestational age a parts per thousand currency sign32 weeks. Two pools of samples from patients with and without BPD were analyzed by high performance liquid chromatography. Several differentially expressed peptides were collected and sequenced. Moreover, samples from single donors were analyzed by liquid chromatography-electrospray ionization mass spectrometry to define the molecular mass values of various peptides and to quantify their expression. Levels of some matrix metalloproteinases and their tissue inhibitors were also determined in single samples. Neonates of the BPD group (N = 16) showed significantly lower mean gestational age and birth weight with respect to the no-BPD group (N = 18; P < 0.0001). Levels of six peptides were significantly higher in BPD patients (P < 0.05). Two of them were identified as the albumin fragments 1-21 (2,428 Da) and 399-406 (956 Da). Levels of matrix metalloproteinase-3 (MMP-3) enzyme probably involved in albumin fragment generation were also significantly higher in the BPD group compared to the no-BPD group (P < 0.05), whereas the levels of tissue inhibitor of metalloproteinases-1 were significantly lower (P < 0.05). Levels of albumin fragments and MMP-3 showed a significant correlation (P < 0.05). This study shows that proteomic techniques can be applied to investigate the involvement of proteolytic enzymes on the airways of mechanically ventilated premature infants

Association of high levels of \u3b1-defensins and S100A proteins with Candida mannan detection in bronchoalveolar lavage fluid of preterm neonates

BACKGROUND: Candida mannan (Mn) detection in bronchoalveolar lavage fluid (BALF) was shown to be useful for earlier identification and preemptive therapy targeting in preterm infants at high risk of invasive Candida infection. We investigated whether early detection of Candida Mn in BALF is associated with the presence of some neutrophilic products, as markers of prenatal infection/inflammation. METHODS: BALF specimens were collected during the first 48 h of life from mechanically ventilated preterm newborns. Samples were analyzed by high-performance liquid chromatography-electrospray ionization-mass spectrometry. The relative amounts of \u3b1-defensins 1-4 and S100A proteins were measured by extracted ion current peak area. Absolute and differential white cell counts in BALF were obtained. Mn antigen concentrations were determined by the Platelia Candida antigen kit. RESULTS: Twenty-five studied neonates were divided into two groups: Mn-positive group and Mn-negative group. Levels of \u3b1-defensins 1-4 and S100A12 were significantly higher in the Mn-positive group than in the Mn-negative group. Moreover, positive significant correlations between the absolute number of neutrophils and the levels of \u3b1-defensins 1-4 and S100A8 were observed. CONCLUSION: The detection of Mn antigen in BALF of preterm infants is consistent with evidence of an innate immune response in their lungs as demonstrated by higher levels of \u3b1-defensins and S100A proteins

Association of high levels of alpha-defensins and S100A proteins with Candida mannan detection in bronchoalveolar lavage fluid of preterm neonates

Background.Candida mannan (Mn) detection in bronchoalveolar lavage fluid (BALF) was shown to be useful for earlier identification and preemptive therapy targeting in preterm infants at high risk of invasive candida infection (ICI). We investigated whether early detection of Candida Mn in BALF is associated with the presence of some neutrophilic products, as markers of pre-natal infection/inflammation.Methods.BALF specimens were collected during the first 48 hours of life from preterm newborns mechanically ventilated. Samples were analyzed by high-performance liquid chromatography (HPLC)-electrospray ionization-mass spectrometer. The \u3b1-defensins 1-4 and S100A proteins relative amounts were measured by extracted ion current peak area. Absolute and differential white cells counts in BALF were obtained. Mn antigen concentrations were determined by the Platelia Candida antigen kit.Results.Twenty-five studied neonates were divided into two groups: Mn-positive group and Mn-negative group. \u3b1-defensins 1-4 and S100A12 levels were significantly higher in the Mn-positive group respect to the Mn-negative group. Moreover, positive significant correlations between the absolute number of neutrophils and the \u3b1-defensins 1-4 and S100A8 levels were observed.Conclusion.The detection of Mn-antigen in BALF of preterm infants is consistent with evidence of an innate immune response in their lungs as demonstrated by higher levels of \u3b1-defensins and S100A proteins