Anti-hemagglutinin monomeric nanobody provides prophylactic immunity against H1 subtype influenza A viruses.

Influenza viruses constitute a major threat to human health globally. The viral surface glycoprotein hemagglutinin (HA) is the immunodominant antigen, contains the site for binding to the cellular receptor (RBS), and it is the major target of neutralizing antibody responses post-infection. We developed llama-derived single chain antibody fragments (VHHs) specific for type A influenza virus. Four VHHs were identified and further characterized. VHH D81 bound residues in the proximity of the C-terminal region of HA1 of H1 and H5 subtypes, and showed weak neutralizing activity, whereas VHH B33 bound residues in the proximity of the N-terminal region of the HA's stem domain (HA2) of H1, H5, and H9 subtypes, and showed no neutralizing activity. Of most relevance, VHHs E13 and G41 recognized highly conserved conformational epitopes on the H1 HA's globular domain (HA1) and showed high virus neutralizing activity (ranging between 0.94 to 0.01µM), when tested against several human H1N1 isolates. Additionally, E13 displayed abrogated virus replication of a panel of H1N1 strains spanning over 80 years of antigenic drift and isolated from human, avian, and swine origin. Interestingly, E13 conferred protection in vivo at a dose as low as 0.05 mg/kg. Mice treated with E13 intranasally resulted in undetectable virus challenge loads in the lungs at day 4 post-challenge. The transfer of sterilizing pan-H1 immunity, by a dose in the range of micrograms given intranasally, is of major significance for a monomeric VHH and supports the further development of E13 as an immunotherapeutic agent for the mitigation of influenza infections.

Selection of tawny owl (Strix aluco) flight feather shaft for biomonitoring As, Cd and Pb pollution.

In this study, we determined the concentrations of As, Cd and Pb in the shaft of all primary flight feathers from ten tawny owl (Strix aluco) specimens, with the aim of selecting which shaft of the corresponding primary feather should be used in biomonitoring surveys to enable inter-individual comparisons of the levels of these metals. The birds had died between 2006 and 2013 and their bodies were stored in the various Wildlife Recovery Centres in Galicia (NW Spain). The analyses revealed a high degree of inter-shaft variability, mainly in the concentrations of As and Cd. However, it was possible to identify the most representative samples in each case: for As, the shaft of primary flight feather number 5 (S5) (which represented 11% of the total As excreted in all of the primary flight feathers); for Cd, the shaft of primary flight feather number 2 (S2) (11% of the total excreted); and for Pb, the shaft of primary flight feather number 8 (S8) (14% of the total excreted). However, the difficulties associated with the analytical determination of these pollutants in the shaft should be taken into account when this technique is applied in biomonitoring studies.

Peptide epitopes of the Taenia solium antigen Ts8B2 are immunodominant in human and porcine cysticercosis.

Ts8B2 is a gene which encodes for a member of the Taenia solium metacestode 8kDa antigen family. Since the Ts8B2-GST recombinant protein compares very favourably with other diagnostic antigens, and in order to study the antigenic nature and structure of this molecule, the Ts8B2 was expressed in prokaryotic and eukaryotic systems. The diagnostic potential of the recombinant Ts8B2 proteins was evaluated by enzyme-linked immunosorbent assays (ELISA) using a collection of serum and cerebrospinal fluid (CSF) samples from patients with clinically defined neurocysticercosis (NCC), and also sera from T. solium infected pigs. Despite the predicted glycosylation of the Ts8B2-Bac recombinant protein, there was very little difference in assay sensitivity/specificity when the Ts8B2 reagent was expressed in either prokaryotic or eukaryotic systems, suggesting that peptidic Ts8B2 epitopes are immunodominant in porcine cysticercosis and human neurocysticercosis. Conveniently, production of recombinant Ts8B2 in Escherichia coli is economical and facile, making it a feasible and practical choice as a diagnostic reagent for use in endemic areas. The Ts8B2 ELISA is particularly useful for the diagnosis of active as opposed to inactive cases of NCC and conduct of the assay is also facilitated by the fact that assay sensitivity is significantly greater when serum as opposed to CSF samples are employed.

Evaluation of recombinant HP6-Tsag, an 18 kDa Taenia saginata oncospheral adhesion protein, for the diagnosis of cysticercosis.

With the objective of providing inexpensive and reproducible assays for the detection of antibodies indicating exposure to Taenia saginata and Taenia solium, we have evaluated the diagnostic utility of the T. saginata oncosphere adhesion protein (HP6-Tsag), expressed in baculovirus (HP6-Bac) and bacteria (HP6-GST [glutathione S-transferase]), employing enzyme-linked immunosorbent assays (ELISAs) and sera from T. saginata infected cattle, T. solium infected pigs and serum and cerebrospinal fluid (CSF) samples from clinically defined T. solium neurocysticercosis (NCC) patients. The two recombinant proteins were antigenic in all three systems, with the signal to background ratio of the HP6-Bac ELISA slightly higher than that for the HP6-GST ELISA. Assay performance in cattle was similar to previously described peptide-based ELISA assays, although NCC sample sensitivity/specificity was marginally better. The sensitivity of the HP6-Bac and HP6-GST ELISAs was close for active human NCC (77.4 and 80.6% for serum and 76.9 and 73.1% for CSF samples, respectively). In inactive human NCC, however, the sensitivity of the HP6-Bac ELISA was almost twice that of the HP6-GST ELISA. Because peptides are relatively expensive and recombinant proteins are simple and economical to produce, the latter may provide useful reagents for antibody detection in countries with endemic cysticercosis/NCC.