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Abstract Passive acoustic monitoring (PAM) has gained increasing popularity to study behaviour, habitat preferences, distribution and community assembly of birds and other animals. Automated species classification algorithms like ‘BirdNET’ are capable of detecting and classifying avian vocalizations within extensive audio data, covering entire species assemblages. PAM reveals substantial potential for biodiversity monitoring that informs evidence‐based conservation. Nevertheless, fully realizing this potential remains challenging, especially due to the issue of false‐positive species detections. Here, we introduce an optimized thresholding framework, which incorporates contextual information extracted from the time‐series of automated species detections (i.e. covariates on quality and quantity of species' detections measured at varying time intervals) to improve the differentiation of true and false positives. We verified a sample of BirdNET detections per species and modelled species‐specific thresholds using conditional inference trees. These thresholds were designed to minimize false‐positive detections while maximizing the preservation of true positives in the dataset. We tested this framework for a large dataset of BirdNET detections (5760 h of audio data, 60 sites) recorded over an entire breeding season. Our results revealed considerable interspecific variability of precision (percentage of true positives) within raw BirdNET data. Our optimized thresholding approach achieved high precision (≥0.9) for 70% of the 61 detected species, while species‐specific thresholds solely relying on the BirdNET confidence scores achieved high precision for only 31% of the species. Conservative universal thresholds (not species‐specific) reached high precision for 48% of the species. Our thresholding approach outperformed previous thresholding approaches and enhanced interspecific comparability for bird community analyses. By incorporating contextual information from the time‐series of species detections, the differentiation of true and false positives was substantially improved. Our approach may enhance a straightforward application of PAM in biodiversity research, landscape planning and evidence‐based conservation.

Abstract Biocrusts represent associations of lichens, green algae, cyanobacteria, fungi and other microorganisms, colonizing soils in varying proportions of principally arid biomes. The so-called grit crust represents a recently discovered type of biocrust situated in the Coastal Range of the Atacama Desert (Chile) made of microorganisms growing on and in granitoid pebbles, resulting in a checkerboard pattern visible to the naked eye on the landscape scale. This specific microbiome fulfills a broad range of ecosystem services, all probably driven by fog and dew-induced photosynthetic activity of mainly micro-lichens. To understand its biodiversity and impact, we applied a polyphasic approach on the phototrophic microbiome of this biocrust, combining isolation and characterization of the lichen photobionts, multi-gene phylogeny of the photobionts and mycobionts based on a direct sequencing and microphotography approach, metabarcoding and determination of chlorophylla+b contents. Metabarcoding showed that yet undescribed lichens within the Caliciaceae dominated the biocrust together with Trebouxia as the most abundant eukaryote in all plots. Together with high mean chlorophylla+b contents exceeding 410 mg m−2, this distinguished the symbiotic algae Trebouxia as the main driver of the grit crust ecosystem. The trebouxioid photobionts could be assigned to the I (T. impressa/gelatinosa) and A (T. arboricola) clades and represented several lineages containing five potential species candidates, which were identified based on the unique phylogenetic position, morphological features, and developmental cycles of the corresponding isolates. These results designate the grit crust as the only known coherent soil layer with significant landscape covering impact of at least 440 km2, predominantly ruled by a single symbiotic algal genus.

Background Herpes zoster (HZ) is a painful condition caused by reactivation of the varicella-zoster virus. The objectives of this study were to compare HZ incidence in adults with asthma versus adults without asthma and to compare healthcare resource use as well as direct costs in adults with HZ and asthma versus adults with asthma alone in the USA.Methods This retrospective longitudinal cohort study included adults aged ≥18 years across the USA. Patients were identified from Optum’s deidentified Clinformatics Data Mart Database, an administrative claims database, between 1 October 2015 and 28 February 2020, including commercially insured and Medicare Advantage with part D beneficiaries. Cohorts of patients with and without asthma, and separate cohorts of patients with asthma and HZ and with asthma but not HZ, were identified using International Classification of Diseases 10th Revision, Clinical Modification codes. HZ incidence, healthcare resource use and costs were compared, adjusting for baseline characteristics, between the relevant cohorts using generalised linear models.Results HZ incidence was higher in patients with asthma (11.59 per 1000 person-years) than patients without asthma (7.16 per 1000 person-years). The adjusted incidence rate ratio (aIRR) for HZ in patients with asthma, compared with patients without asthma, was 1.34 (95% CI 1.32 to 1.37). Over 12 months of follow-up, patients with asthma and HZ had more inpatient stays (aIRR 1.11; 95% CI 1.02 to 1.21), emergency department visits (aIRR 1.26; 95% CI 1.18 to 1.34) and outpatient visits (aIRR 1.19; 95% CI 1.16 to 1.22), and direct healthcare costs that were US dollars ($) 3058 (95% CI $1671 to $4492) higher than patients with asthma without HZ.Conclusion Patients with asthma had a higher incidence of HZ than those without asthma, and among patients with asthma HZ added to their healthcare resource use and costs.

Abstract Introduction Patients with psoriasis (PsO) are at increased risk of herpes zoster (HZ), but recent data on the incidence of HZ among patients with PsO and the impact of HZ on healthcare resource use (HRU) and costs for patients with PsO have not been described. Methods This retrospective, longitudinal, cohort study estimated HZ incidence in cohorts of adults with vs without PsO (PsO + vs PsO–) and HRU and costs among those with PsO, with vs without HZ (PsO + /HZ + vs PsO + /HZ–) using Optum’s de-identified Clinformatics Data Mart Database during 2015–2020. Patients with psoriatic arthritis were excluded from all four cohorts. Comparisons between cohorts used generalized linear models to adjust outcomes based on various baseline characteristics. Results The incidence rate of HZ was significantly higher in the PsO + (n = 144,115) vs PsO– (n = 23,837,237) cohorts at 11.35 vs 7.67 per 1000 patient-years; adjusted incidence rate ratio (aIRR): 1.21, 95% confidence interval (CI): 1.16–1.25. HRU (outpatient, emergency department, and inpatient) was significantly higher in the PsO + /HZ + (n = 1859) vs PsO + /HZ– (n = 78,664) cohorts during 1 month and 3 months after HZ diagnosis (e.g., outpatient visits during month: 2.83 vs 1.30 per patient; aIRR: 1.96; 95% CI 1.86–2.06). Mean all-cause costs were also significantly higher in the PsO + /HZ + vs PsO + /HZ– cohort during both month ($5020 vs $2715 per patient; adjusted cost difference: $1390; 95% CI $842–$1964) and 3 months ($12,305 vs $8256; adjusted cost difference: $1422; 95% CI $280–$2889) after HZ diagnosis. Conclusion These findings show the increased incidence of HZ among patients with PsO and the clinical and economic burdens of HZ in this population. Considering the high prevalence of PsO, insights into the impact of HZ in these patients provide valuable evidence to inform clinical decision-making. Graphical abstract

Abstract Background Many types of interstitial lung diseases (ILDs) may transition to progressive chronic-fibrosing ILDs with rapid lung function decline and a negative survival prognosis. In real-world clinical settings, forced vital capacity (FVC) measures demonstrating progressive decline may be linked to negative outcomes, including increased risks of costly healthcare resource utilization (HRU). Thus, we assessed the relationship between rate of decline in lung function and an increase in HRU, specifically inpatient hospitalization, among patients with chronic fibrosing ILD. Methods This study utilized electronic health records from 01-Oct-2015 to 31-Oct-2019. Eligible patients (≥ 18 years old) had ≥ 2 fibrosing ILD diagnosis codes, clinical activity for ≥ 15 months, and ≥ 2 FVC tests occurring 6 months apart. Patients with missing demographic data, IPF, or use of nintedanib or pirfenidone were excluded. Two groups were defined by relative change in percent of predicted FVC (FVC% pred) from baseline to 6 months: significant decline (≥ 10%) vs. marginal decline/stable FVC (decrease

Abstract Objective To estimate the incremental healthcare resource utilization (HRU) and cost burden posed by herpes zoster (HZ) in adult patients with rheumatoid arthritis (RA) in the United States. Methods A retrospective cohort study was conducted using an administrative claims database containing commercial and Medicare Advantage with Part D data, between October 2015 and February 2020. Patients with RA and HZ (RA+/HZ+) or RA without HZ (RA+/HZ−) were identified based on diagnosis codes and relevant medications. Outcomes measured included HRU and medical, pharmacy, and total costs at month 1, quarter 1, and year 1 after the index date (HZ diagnosis for RA+/HZ+ cohort, randomly assigned for RA+/HZ− cohort). Generalized linear models incorporating propensity scores and other covariates were used to estimate differences in outcomes between cohorts. Results A total of 1866 patients from the RA+/HZ+ cohort and 38,846 patients from the RA+/HZ− cohort were included. Hospitalizations and emergency department visits occurred more frequently in the RA+/HZ+ than the RA+/HZ− cohort, especially in the month after HZ diagnosis (adjusted incidence rate ratio [95% confidence interval (CI)] for hospitalizations: 3.4 [2.8; 4.2]; emergency department visits: 3.7 [3.0; 4.4]). Total costs were also higher in the month after HZ diagnosis (mean adjusted cost difference [95% CI]: $3404 [$2089; $4779]), with cost differences driven by increased medical costs ($2677 [$1692; $3670]). Conclusions These findings highlight the high economic burden of HZ among individuals with RA in the United States. Strategies to reduce the risk of HZ in patients with RA (such as vaccination) may serve to reduce this burden. Video abstract

ABSTRACTThe recombinant zoster vaccine (RZV) was licensed in the US for prevention of herpes zoster (HZ) in 2017. We conducted a literature search (January 1, 2017–August 1, 2023) using PubMed, Embase, and Scopus to consolidate the real-world evidence related to RZV. Overall, RZV effectiveness against HZ was high across the studied populations in real-world settings, including adults aged ≥ 50 years and patients aged ≥ 18 years with immunodeficiency or immunosuppression. Effectiveness was higher with two doses versus one dose, especially in elderly people and immunocompromised individuals. The safety profile of RZV was broadly consistent with that established in clinical trials. RZV does not appear to increase the risk of disease flares in patients with immune-mediated diseases. Approximately two-thirds of individuals received a second RZV dose within 2–6 months after the first dose. Collectively, RZV effectiveness against HZ was high, and these real-world studies reaffirm its favorable benefit–risk profile.

Abstract Background Interstitial lung diseases (ILDs) include a variety of parenchymal lung diseases. The most common types of ILDs are idiopathic pulmonary fibrosis (IPF), autoimmune ILDs and hypersensitivity pneumonitis (HP). There is limited real world data on care patterns of patients with chronic fibrosing ILDs with a progressive phenotype other than IPF. Therefore, the aim of this study is to describe care patterns in these patients. Methods This retrospective cohort study used claims data from 2015 to 2019 from the Optum Research Database. The study population included adults (≥ 18 years old) with at least two diagnosis codes for fibrosing ILD during the identification period (1OCT2016 to 31DEC2018). A claim-based algorithm for disease progression was used to identify patients likely to have a progressive fibrotic phenotype using progression proxies during the identification period. Index date was the first day of progression proxy identification after fibrosing ILD diagnosis. Patients were required to have continuous enrollment for 12 months before (baseline) and after (follow-up) index date. Patients with an IPF diagnosis were excluded. Descriptive statistics were used to describe the patient population and care patterns. Results 11,204 patients were included in the study. Mean age of the patient population was 72.7 years, and 54.5% were female. Unclassified ILDs (48.0%), HP (25.2%) and autoimmune ILDs (16.0%) were the most common ILD types. Other respiratory conditions were prevalent among patients including chronic obstructive pulmonary disease (COPD) (58.9%), obstructive sleep apnea (OSA) (25.0%) and pulmonary hypertension (9.8%). During baseline, 65.3% of all patients had at least one pulmonology visit, this proportion was higher during follow-up, at 70.6%. Baseline and follow-up use for HRCT were 39.9% and 48.8%, and for pulmonary function tests were 43.7% and 48.5% respectively. Use of adrenal corticosteroids was higher during follow-up than during baseline (62.5% vs. 58.0%). Anti-inflammatory and immunosuppressive medication classes were filled by a higher percentage of patients during follow-up than during baseline. Conclusions Comprehensive testing is essential for diagnosis of a progressive phenotype condition, but diagnostic tests were underutilized. Patients with this condition frequently were prescribed anti-inflammatory and immunosuppressive medications.

We apply the theory of quadratic differentials, to present a classification of orthogonal pairs of foliations of the hyperbolic plane by hyperbolic conics. Light rays are represented by trajectories of meromorphic differentials, and mirrors are represented by trajectories of the quadratic differential that represents the geometric mean of two such differentials. Using the notion of a hyperbolic conic as a mirror, we classify the types of orthogonal pairs of foliations of the hyperbolic plane by confocal conics. Up to diffeomorphism, there are nine types: three of these types admit one parameter up to isometry; the remaining six types are unique up to isometry. The families include all possible hyperbolic conics.

Rain-fed mountain granite rock basins are temporary habitats conditioned by a fluctuating environment and the unpredictability of precipitation or flooding rates. These small highland freshwater habitats remain largely unexplored at the microbial level. The aim of this work is to report the presence in these habitats of genetic sequences of microbial eukaryotes that are pathogens and potential pathogens of humans, wildlife, cattle, crops as well as of other microorganisms. We sequenced the hypervariable region v4 of the 18S rDNA gene from environmental DNA of sediments taken from 21 rock basins in a National Park in Spain. More than a fifth (21%) of the eukaryotic Operational Taxonomic Units (OTUs) found are ascribed to pathogenic (within 11 Phyla) and potential pathogenic (within 1 phylum, the Chytridiomycota) microorganisms. Some OTUs retrieved are of agro-economic and public health importance (e.g., Pythium spp., Lagenidium spp., Candida spp. and Vermamoeba vermiformis). In 86% of the basins, the most abundant OTUs were affiliated to Chytridiomycota, a broad fungal group including saprozoic and parasitic taxa. Two OTUs affiliated to chytrids were significantly correlated with high concentrations of heavy metals. The high proportion of chytrid-like microbial sequences found emphasises the role of these freshwater habitats for adding knowledge regarding the ecological trade-offs of the still rather unknown Chytridiomycota. Our results show that rain-fed rock basins may be model habitats for the study and surveillance of microbial community dynamics and genetics of (mainly opportunistic) microbial pathogens.

Due to COVID-19, vaccinations dropped in 2020 and 2021. We estimated the impact of reduced recombinant zoster vaccine (RZV) use on herpes zoster (HZ) cases, complications, and quality-adjusted life-year (QALY) losses among older adults. Various scenarios were compared with Markov models using data from national sources, clinical trials, and literature. Missed series initiations were calculated based on RZV distributed doses. In 2020, 3.9 million RZV series initiations were missed, resulting in 31,945 HZ cases, 2,714 postherpetic neuralgia cases, and 610 lost QALYs. Scenarios further projected disease burden increases if individuals remain unvaccinated in 2021 or the same number of initiations are missed in 2021. Health professionals should emphasize the importance of vaccination against all preventable diseases during the COVID-19 era.

Protists dominate eukaryotic diversity and play key functional roles in all ecosystems, particularly by catalyzing carbon and nutrient cycling. To date, however, a comparative analysis of their taxonomic and functional diversity that compares the major ecosystems on Earth (soil, freshwater and marine systems) is missing. Here, we present a comparison of protist diversity based on standardized high throughput 18S rRNA gene sequencing of soil, freshwater and marine environmental DNA. Soil and freshwater protist communities were more similar to each other than to marine protist communities, with virtually no overlap of Operational Taxonomic Units (OTUs) between terrestrial and marine habitats. Soil protists showed higher γ diversity than aquatic samples. Differences in taxonomic composition of the communities led to changes in a functional diversity among ecosystems, as expressed in relative abundance of consumers, phototrophs and parasites. Phototrophs (eukaryotic algae) dominated freshwater systems (49% of the sequences) and consumers soil and marine ecosystems (59% and 48%, respectively). The individual functional groups were composed of ecosystem- specific taxonomic groups. Parasites were equally common in all ecosystems, yet, terrestrial systems hosted more OTUs assigned to parasites of macro-organisms while aquatic systems contained mostly microbial parasitoids. Together, we show biogeographic patterns of protist diversity across major ecosystems on Earth, preparing the way for more focused studies that will help understanding the multiple roles of protists in the biosphere.

In the original publication [...]

This study focuses on the foraminiferal distribution on intertidal mudflats of two contrasted estuaries (Auray and Vie) along the French Atlantic coast. In both estuaries, the foraminiferal communities are dominated by Haynesina germanica and the Ammonia tepida group. Stations located near the outlets show a high diversity and abundance of species of the genus Elphidium. Stations in the inner estuary show a higher proportion of agglutinated species (Ammotium salsum, Ammobaculites agglutinans). Multivariate statistical analysis suggests that the distance to the sea and the percentage of fine sediment (Elphidium spp. and Quinqueloculina spp.). These results suggest that the MII explains a substantial part of the faunal variability on estuarine intertidal mudflats, and can be used to detect deviations from the natural distribution patterns in response to anthropogenic pollution.

In this paper, we propose a marine influence index (MII), which is thought to give an integrated quantitative description of the complex of the environmental parameters controlling the foraminiferal fauna in estuarine intertidal mudflats. The MII contains three components, as follows: (1) the relative distance along the salinity gradient, (2) the emergence time relative to a reference tidal cycle, and (3) the relative importance of river outflow in the 30 days before sampling the foraminiferal fauna. Although these three parameters all have a strong relation with salinity, they also implicitly include other environmental parameters, such as the introduction of marine and continental organic matter and biota, hydrodynamic energy, or temperature. In order to show the functioning of this new index, MII is calculated for 28 stations in the Auray and Vie estuaries, for two different periods. The next step will be to compare the MII with faunal data sets. Ideally, this comparison should allow us to find strong correlations between some characteristics of the foraminiferal assemblages and the MII. If such strong correlations were indeed found, any major deviation of this relationship could then be interpreted as being due to strong anthropogenic disturbance.

This review provides a synthesis of current knowledge on the morphological and functional traits of testate amoebae, a polyphyletic group of protists commonly used as proxies of past hydrological changes in paleoecological investigations from peatland, lake sediment and soil archives. A trait-based approach to understanding testate amoebae ecology and paleoecology has gained in popularity in recent years, with research showing that morphological characteristics provide complementary information to the commonly used environmental inferences based on testate amoeba (morpho-)species data. We provide a broad overview of testate amoeba morphological and functional traits and trait-environment relationships in the context of ecology, evolution, genetics, biogeography, and paleoecology. As examples we report upon previous ecological and paleoecological studies that used trait-based approaches, and describe key testate amoebae traits that can be used to improve the interpretation of environmental studies. We also highlight knowledge gaps and speculate on potential future directions for the application of trait-based approaches in testate amoeba research.

Coal mine spoil is widespread in US coal mining regions, and the potential long-term leaching of toxic metal(loid)s is a significant and underappreciated issue. This study aimed to determine the flux of contaminants from historic mine coal spoil at a field site located in Appalachian Ohio (USA) and link pore water composition and solid-phase composition to the weathering reaction stages within the soils. The overall mineralogical and microbial community composition indicates that despite very different soil formation pathways, soils developing on historic coal mine spoil and an undisturbed soil are currently dominated by similar mineral weathering reactions. Both soils contained pyrite coated with clays and secondary oxide minerals. However, mine spoil soil contained abundant residual coal, with abundant Fe- and Mn- (oxy)hydroxides. These secondary phases likely control and mitigate trace metal (Cu, Ni, and Zn) transport from the soils. While Mn was highly mobile in Mn-enriched soils, Fe and Al mobility may be more controlled by dissolved organic carbon dynamics than mineral abundance. There is also likely an underappreciated risk of Mn transport from coal mine spoil, and that mine spoil soils could become a major source of metals if local biogeochemical conditions change.

Intracellular neurofibrillary tangles and extracellular senile plaques are potential targets for active and passive immunotherapies. In this study we used the transgenic mouse model P301S for active immunizations with peptide vaccines composed of a double phosphorylated tau neoepitope (pSer202/pThr205, pThr212/pSer214, pThr231/pSer235) and an immunomodulatory T cell epitope from the tetanus toxin or tuberculosis antigen Ag85B. Importantly, the designed vaccine combining Alzheimer’s disease (AD) specific B cell epitopes with foreign (bacterial) T cell epitopes induced fast immune responses with high IgG1 titers after prophylactic immunization that subsequently decreased over the observation period. The effectiveness of the immunization was surveyed by evaluating the animal behavior, as well as the pathology in the brain by biochemical and histochemical techniques. Immunized mice clearly lived longer with reduced paralysis than placebo-treated mice. Additionally, they performed significantly better in rotarod and beam walk tests at the age of 20 weeks, indicating that the disease development was slowed down. Forty-eight weeks old vaccinated mice passed the beam walk test significantly better than control animals, which together with the increased survival rates undoubtedly prove the treatment effect. In conclusion, the data provide strong evidence that active immune therapies can reduce toxic effects of deposits formed in AD.

Abandoned mine lands continue to serve as non-point sources of acid and metal contamination to water bodies long after mining operations have ended. Although soils formed from abandoned mine spoil can support forest vegetation, as observed throughout the Appalachian coal basin, the effects of vegetation on metal cycling in these regions remain poorly characterized. Iron (Fe) and manganese (Mn) biogeochemistry were examined at a former coal mine where deciduous trees grow on mine spoil deposited nearly a century ago. Forest vegetation growing on mine spoil effectively removed dissolved Mn from pore water; however, mineral weathering at a reaction front below the rooting zone resulted in high quantities of leached Mn. Iron was taken up in relatively low quantities by vegetation but was more readily mobilized by dissolved organic carbon produced in the surface soil. Dissolved Fe was low below the reaction front, suggesting that iron oxyhydroxide precipitation retains Fe within the system. These results indicate that mine spoil continues to produce Mn contamination, but vegetation can accumulate Mn and mitigate its leaching from shallow soils, potentially also decreasing Mn leaching from deeper soils by reducing infiltration. Vegetation had less impact on Fe mobility, which was retained as Fe oxides following oxidative weathering.

Rational design of peptide vaccines becomes important for the treatment of some diseases such as Alzheimer’s disease (AD) and related disorders. In this study, as part of a larger effort to explore correlations of structure and activity, we attempt to characterize the doubly phosphorylated chimeric peptide vaccine targeting a hyperphosphorylated epitope of the Tau protein. The 28-mer linear chimeric peptide consists of the double phosphorylated B cell epitope Tau229-237[pThr231/pSer235] and the immunomodulatory T cell epitope Ag85B241-255 originating from the well-known antigen Ag85B of the Mycobacterium tuberculosis, linked by a four amino acid sequence -GPSL-. NMR chemical shift analysis of our construct demonstrated that the synthesized peptide is essentially unfolded with a tendency to form a β-turn due to the linker. In conclusion, the -GPSL- unit presumably connects the two parts of the vaccine without transferring any structural information from one part to the other. Therefore, the double phosphorylated epitope of the Tau peptide is flexible and accessible.

BACKGROUND: Organotypic brain slice cultures represent an excellent compromise between single cell cultures and complete animal studies, in this way replacing and reducing the number of animal experiments. Organotypic brain slices are widely applied to model neuronal development and regeneration as well as neuronal pathology concerning stroke, epilepsy and Alzheimer's disease (AD). AD is characterized by two protein alterations, namely tau hyperphosphorylation and excessive amyloid β deposition, both causing microglia and astrocyte activation. Deposits of hyperphosphorylated tau, called neurofibrillary tangles (NFTs), surrounded by activated glia are modeled in transgenic mice, e.g. the tauopathy model P301S. METHODOLOGY/PRINCIPAL FINDINGS: In this study we explore the benefits and limitations of organotypic brain slice cultures made of mature adult transgenic mice as a potential model system for the multifactorial phenotype of AD. First, neonatal (P1) and adult organotypic brain slice cultures from 7- to 10-month-old transgenic P301S mice have been compared with regard to vitality, which was monitored with the lactate dehydrogenase (LDH)- and the MTT (3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) assays over 15 days. Neonatal slices displayed a constant high vitality level, while the vitality of adult slice cultures decreased significantly upon cultivation. Various preparation and cultivation conditions were tested to augment the vitality of adult slices and improvements were achieved with a reduced slice thickness, a mild hypothermic cultivation temperature and a cultivation CO(2) concentration of 5%. Furthermore, we present a substantial immunohistochemical characterization analyzing the morphology of neurons, astrocytes and microglia in comparison to neonatal tissue. CONCLUSION/SIGNIFICANCE: Until now only adolescent animals with a maximum age of two months have been used to prepare organotypic brain slices. The current study provides evidence that adult organotypic brain slice cultures from 7- to 10-month-old mice independently of the transgenic modification undergo slow programmed cell death, caused by a dysfunction of the neuronal repair systems.

The extracellular domain of the influenza A virus protein matrix protein 2 (M2e) is remarkably conserved between various human isolates and thus is a viable target antigen for a universal influenza vaccine. With the goal of inducing protection in multiple mouse haplotypes, M2e-based multiple antigenic peptides (M2e-MAP) were synthesized to contain promiscuous T helper determinants from the Plasmodium falciparum circumsporozoite protein, the hepatitis B virus antigen and the influenza virus hemagglutinin. Here, we investigated the nature of the M2e-MAP-induced B cell response in terms of the distribution of antibody (Ab) secreting cells (ASCs) and Ab isotypes, and tested the protective efficacy in various mouse strains.Immunization of BALB/c mice with M2e-MAPs together with potent adjuvants, CpG 1826 oligonucleotides (ODN) and cholera toxin (CT) elicited high M2e-specific serum Ab titers that protected mice against viral challenge. Subcutaneous (s.c.) and intranasal (i.n.) delivery of M2e-MAPs resulted in the induction of IgG in serum and airway secretions, however only i.n. immunization induced anti-M2e IgA ASCs locally in the lungs, correlating with M2-specific IgA in the bronchio-alveolar lavage (BAL). Interestingly, both routes of vaccination resulted in equal protection against viral challenge. Moreover, M2e-MAPs induced cross-reactive and protective responses to diverse M2e peptides and variant influenza viruses. However, in contrast to BALB/c mice, immunization of other inbred and outbred mouse strains did not induce protective Abs. This correlated with a defect in T cell but not B cell responsiveness to the M2e-MAPs.Anti-M2e Abs induced by M2e-MAPs are highly cross-reactive and can mediate protection to variant viruses. Although synthetic MAPs are promising designs for vaccines, future constructs will need to be optimized for use in the genetically heterogeneous human population.