Increasing Concentrations of Perfluoroalkyl Acids in Scandinavian Otters (<i>Lutra lutra</i>) between 1972 and 2011: A New Threat to the Otter Population?

Liver samples from 140 otters <i>(Lutra lutra)</i> from Sweden and Norway were analyzed for 10 perfluoroalkyl carboxylic acids (PFCAs; C6–C15), 4 perfluoroalkane sulfonic acids (PFSAs; C4,C6,C8,C10) and perfluorooctane sulfonamide (FOSA). Perfluorooctane sulfonic acid (PFOS) was the dominant compound accounting for approximately 80% of the fluorinated contaminants and showing concentrations up to 16 μg/g wet weight. Perfluorononanoic acid (PFNA) was the dominant PFCA (up to 640 ng/g wet weight) closely followed by the C10 and C11 homologues. A spatial comparison between otters from southwestern Norway, southern and northern Sweden sampled between 2005 and 2011 revealed that the samples from southern Sweden had generally the largest contaminant load, but two PFCAs and FOSA were higher concentrated in the Norwegian samples. A temporal trend study was performed on otters from southern Sweden collected between 1972 and 2011. Seven PFCAs (C8–C14), PFOS and perfluorodecane sulfonic acid (PFDS) showed significantly increasing trends with doubling times between 5.5 and 13 years. The PFCAs also showed significantly increasing trends over the period 2002 to 2011. These findings together with the exceptionally high liver concentrations of PFOS are of great concern for the Scandinavian otter populations

Matrix-M Adjuvated Seasonal Virosomal Influenza Vaccine Induces Partial Protection in Mice and Ferrets against Avian H5 and H7 Challenge

<div><p>There is a constant threat of zoonotic influenza viruses causing a pandemic outbreak in humans. It is virtually impossible to predict which virus strain will cause the next pandemic and it takes a considerable amount of time before a safe and effective vaccine will be available once a pandemic occurs. In addition, development of pandemic vaccines is hampered by the generally poor immunogenicity of avian influenza viruses in humans. An effective pre-pandemic vaccine is therefore required as a first line of defense. Broadening of the protective efficacy of current seasonal vaccines by adding an adjuvant may be a way to provide such first line of defense. Here we evaluate whether a seasonal trivalent virosomal vaccine (TVV) adjuvated with the saponin-based adjuvant Matrix-M (MM) can confer protection against avian influenza H5 and H7 virus strains in mice and ferrets. We demonstrate that mice were protected from death against challenges with H5N1 and H7N7, but that the protection was not complete as evidenced by severe clinical signs. In ferrets, protection against H7N9 was not observed. In contrast, reduced upper and lower respiratory tract viral loads and reduced lung pathology, was achieved in H5N1 challenged ferrets. Together these results suggest that, at least to some extent, Matrix-M adjuvated seasonal virosomal influenza vaccine can serve as an interim measure to decrease morbidity and mortality associated with a pandemic outbreak.</p></div

Ferrets are not protected against highly pathogenic H7N9 after TVV+MM vaccination.

<p>Groups of 7–8 ferrets received two intramuscular injections with TVV, TVV+MM, PBS, PBS+MM or inactivated H7N9 virus as positive control (Control). 4 weeks later the animals were challenged with a sub-lethal dose of 10^5.5 TCID₅₀ of influenza A H7N9 A/Anhui/1/2013. Ferrets were monitored for 4 consecutive days and sacrificed at day 4 post challenge. (A) Infectious viral load in lung tissue (B) infectious throat viral load (day 1 to 4), (C) percentage of body weight change during the observation period (D) lung weight as determined after sacrifice. Dots indicate individual animals and horizontal lines represent group means (A and D). Lines represent group mean with 95% confidence interval (B) or the interquartile range (C). Asterisks indicate statistically significant differences compared to PBS injected animals (*<i>p</i><0.05, **<i>p</i><0.01, ***<i>p</i><0.001, according to the materials and methods section).</p

Ferrets are partially protected against highly pathogenic H5N1 after TVV+MM vaccination.

<p>Groups of 7–8 ferrets received two intramuscular injections with TVV, TVV+MM, PBS, PBS+MM or inactivated H5H1 virus as positive control (Control). 4 weeks later the animals were challenged with a sub-lethal dose of 10⁴ TCID₅₀ of influenza A H5N1 A/Indonesia/05/2005. Ferrets were monitored for 4 consecutive days and sacrificed at day 4 post challenge. (A) Infectious viral load in lung tissue (B) infectious throat viral load (day 1 to 4), (C) percentage of body weight change during the observation period and (D) lung weight as determined after sacrifice. Dots indicate individual animals and horizontal lines represent group means (A and D). Lines represent group mean with 95% confidence interval (B) or the interquartile range (C). Asterisks indicate statistically significant differences compared to PBS injected animals (*<i>p</i><0.05, **<i>p</i><0.01, ***<i>p</i><0.001, according to the materials and methods section).</p

TVV+MM induces cross-reactive H5 and H7 antibody responses.

<p>Mice (n = 9-10/group) were immunized 1x or 2x with TVV with or without MM. 27 days later (1 day before challenge) individual serum samples were obtained and tested for (A) vaccine homologous recH1 of A/California/07/07, (B) vaccine homologous recH3 of the A/Victoria/210/09-like A/Perth/16/09 (98.8% homologous), (C) recH5 of A/Hong Kong/156/97, and (D) recH7 of A/Netherlands/219/03 (99.6% homologous to the challenge strain A/chicken/Netherlands/621557/03) antibody responses. Serum pools of mice (n = 50/group) that received 1x or 2x TVV+MM or no immunization (-) were tested for (E) vaccine homologous recN1 A/California/04/09 and (F) recN1 of A/Hong Kong/156/97 reactive antibody responses. Black bars indicate medians of log-10 transformed ELISA titers (EU). Asterisks indicate statistically significant differences compared to the vehicle control group (*p<0.05, **p<0.01, ***p<0.001, according to the materials and methods section).</p

TVV+MM protects mice against avian H5N1.

<p>Mice (n = 9-10/group) were immunized 1x or 2x with TVV with or without MM. Four weeks later, mice were challenged with 25xLD₅₀ wild-type A/Hong Kong /156/97 (H5N1) and monitored for 21 days for survival, body weight loss and clinical symptoms. Graphs represent the Kaplan-Meier survival curve (A and D) or mean bodyweight change with 95% confidence interval (B and E) or mean clinical scores with interquartile range (C and F). Asterisks indicate statistically significant differences compared to the vehicle control group (*<i>p</i><0.05, **<i>p</i><0.01, ***<i>p</i><0.001, according to the materials and methods section).</p

TVV+MM protects mice against avian H7N7.

<p>Mice (n = 10/group) were immunized 1x or 2x with TVV with or without MM. Four weeks later, mice were challenged with 25xLD₅₀ H7N7 A/Chicken/Netherlands/62155710/03 and monitored for 21 days for survival, body weight loss and clinical symptoms. Graphs represent the Kaplan-Meier survival curve (A and D) or mean bodyweight change with 95% confidence interval (B and E) or mean clinical scores with interquartile range (C and F). Asterisks indicate statistically significant differences compared to the vehicle control group (*<i>p</i><0.05, **<i>p</i><0.01, ***<i>p</i><0.001, according to the materials and methods section).</p

The first (left panels) and second (right panels) global scores of the sPCA of SNP (top panels) and microsatellite (bottom panels) datasets for the NOS to IBS subset.

<p>The squares represent the score (white–positive, black–negative) of each genotype and are positioned according to their spatial coordinates on the map of the Baltic Sea and adjacent sub-regions. Map data: ESRI (2013).</p

Sampling locations and assignment to geographic sub-regions.

<p>(A) A map of European Seas with circles representing collection sites for individual samples from the Western Baltic Sea (WBS), the Atlantic and the Baltic Sea regions. WBS and Icelandic samples (part of the Atlantic region) are labeled with light blue and dark blue circles, respectively. (B) Collection sites for individual samples from the North Sea sub-region (NOS; purple), and Baltic Sea sub-regions: Skagerrak-northern Kattegat (SK1; pink), southern Kattegat-Belt Sea 1 (KB1; red), Belt Sea 2 (BES2; orange), Inner Baltic Sea (IBS; green). Borders between SK1 and KB1, and BES2 and IBS (dashed lines) were based on proposed borders between management units at 56.95°N latitude, and 13.5°E longitude, respectively (39). Borders between NOS and SK1, and KB1 and BES2 are based on [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0162792#pone.0162792.ref032" target="_blank">32</a>]. Geographic assignment to regions and sub-regions is summarized in a table (bottom right). Map data: ESRI (2013).</p

Association between SNP Structure analysis-derived clusters for WBS to Baltic regions and mitochondrial haplotypes (cf. Fig 5 left panels–all samples).

<p>The haplotype distribution between the two SNP clusters is significantly different (Χ² = 6.111, p = 0.047 for all haplotypes), due to a significant difference in the occurence of the haplotypes PHO4 and PHO7 (Χ² = 6.111, p = 0.013 for PHO4/PHO7 only).</p