A glycoengineered antigen exploiting a conserved protein O-glycosylation pathway in the Burkholderia genus for detection of glanders infections.

We recently described a protein O-glycosylation pathway conserved in all species of the Burkholderia genus that results in the synthesis and incorporation of a trisaccharide glycan to membrane-exported proteins. Here, we exploited this system to construct and evaluate a diagnostic tool for glanders. Burkholderia mallei, the causative agent of glanders, is a highly infectious and fatal zoonotic pathogen that infects horses, mules, donkeys, and occasionally humans. A highly sensitive and specific diagnostic tool is crucial for the control, elimination, and eradication of B. mallei infections. We constructed plasmids carrying synthetic genes encoding a modified, previously unannotated Burkholderia glycoprotein containing three glycosylation sequons fused to the cholera toxin B-subunit. The resulting proteins were glycosylated in the B. cenocepacia K56-2 parental strain, but not in glycosylation-deficient mutants, as determined by SDS-PAGE and fluorescent lectin blots. One of these glycoproteins was used as an antigen in ELISA and western blots to screen a panel of serum samples collected from glanders-infected and healthy horses, which were previously investigated by complement fixation test and indirect ELISA based on a semi-purified fraction of B. mallei. We show that ELISA and western blot assays based on our glycoprotein antigen provide 100% specificity, with a sensitivity greater than 88%. The glycoprotein antigen was recognized by serum samples collected from patients infected with B. pseudomallei, B. mallei, B. multivorans, and B. cenocepacia. Our results indicate that protein O-glycosylation in Burkholderia can be exploited as a biomarker for diagnosis of Burkholderia-associated infections.

Airway Epithelial Derived Cytokines and Chemokines and Their Role in the Immune Response to Respiratory Syncytial Virus Infection.

The airway epithelium is the primary target of respiratory syncytial virus infection. It is an important component of the antiviral immune response. It contributes to the recruitment and activation of innate immune cells from the periphery through the secretion of cytokines and chemokines. This paper provides a broad review of the cytokines and chemokines secreted from human airway epithelial cell models during respiratory syncytial virus (RSV) infection based on a comprehensive literature review. Epithelium-derived chemokines constitute most inflammatory mediators secreted from the epithelium during RSV infection. This suggests chemo-attraction of peripheral immune cells, such as monocytes, neutrophils, eosinophils, and natural killer cells as a key function of the epithelium. The reports of epithelium-derived cytokines are limited. Recent research has started to identify novel cytokines, the functions of which remain largely unknown in the wider context of the RSV immune response. It is argued that the correct choice of in vitro models used for investigations of epithelial immune functions during RSV infection could facilitate greater progress in this field.

Endothelin and vasopressin influence splanchnic blood flow distribution during and after cardiopulmonary bypass.

OBJECTIVE: Gastrointestinal blood flow can be compromised during and after cardiopulmonary bypass. Endothelin has been shown to be involved in the intestinal microcirculatory disturbance of sepsis. The aim of the present study was to analyze the involvement of the endothelin system on intestinal blood flow regulation during cardiopulmonary bypass and the effect of vasopressin given during cardiopulmonary bypass. METHODS: A total of 24 pigs were studied in 4 groups (n = 6): group I, sham; group II, ischemia/reperfusion with 1 hour of superior mesenteric artery occlusion; group III, cardiopulmonary bypass for 1 hour; and group IV, 1 hour of cardiopulmonary bypass plus vasopressin administration, maintaining the baseline arterial pressure. All the pigs were reperfused for 90 minutes. During the experiment, the hemodynamics and jejunal microcirculation were measured continuously. The jejunal mucosal expression of endothelin-1 and its receptor subtypes A and B were determined using polymerase chain reaction. RESULTS: During cardiopulmonary bypass, superior mesenteric artery flow was preserved but marked jejunal microvascular impairment occurred compared with baseline (mucosal capillary density, 192.2 ± 5.4 vs 150.8 ± 5.1 cm/cm(2); P = .005; tissue blood flow, 501.7 ± 39.3 vs 332.3 ± 27.9 AU; P = .025). The expression of endothelin-1 after cardiopulmonary bypass (3.2 ± 0.4 vs 12.2 ± 0.8 RQ, P = .006) and endothelin subtype A (0.7 ± 0.2 vs 2.4 ± 0.6 RQ; P = .01) was significantly increased compared to the sham group. Vasopressin administration during cardiopulmonary bypass led to normal capillary density (189.9 ± 3.9 vs 178.0 ± 6.3; P = .1) and tissue blood flow (501.7 ± 39.3 vs 494.7 ± 44.4 AU; P = .4) compared with baseline. The expression of endothelin-1 (3.2 ± 0.4 vs 1.8 ± 0.3 RQ; P = .3) and endothelin subtype A (0.7 ± 0.2 vs 0.9 ± 0.2 RQ; P = .5) was not different from the sham group. CONCLUSIONS: Cardiopulmonary bypass leads to microvascular impairment of jejunal microcirculation, which is associated with the upregulation of endothelin-1 and endothelin subtype A. The administration of vasopressin minimizes these cardiopulmonary bypass-associated alterations.