Archaeome structure and function of the intestinal tract in healthy and H1N1 infected swine.

Background: Methanogenic archaea represent a less investigated and likely underestimated part of the intestinal tract microbiome in swine. Aims/Methods: This study aims to elucidate the archaeome structure and function in the porcine intestinal tract of healthy and H1N1 infected swine. We performed multi-omics analysis consisting of 16S rRNA gene profiling, metatranscriptomics and metaproteomics. Results and discussion: We observed a significant increase from 0.48 to 4.50% of archaea in the intestinal tract microbiome along the ileum and colon, dominated by genera Methanobrevibacter and Methanosphaera. Furthermore, in feces of naïve and H1N1 infected swine, we observed significant but minor differences in the occurrence of archaeal phylotypes over the course of an infection experiment. Metatranscriptomic analysis of archaeal mRNAs revealed the major methanogenesis pathways of Methanobrevibacter and Methanosphaera to be hydrogenotrophic and methyl-reducing, respectively. Metaproteomics of archaeal peptides indicated some effects of the H1N1 infection on central metabolism of the gut archaea. Conclusions/Take home message: Finally, this study provides the first multi-omics analysis and high-resolution insights into the structure and function of the porcine intestinal tract archaeome during a non-lethal Influenza A virus infection of the respiratory tract, demonstrating significant alterations in archaeal community composition and central metabolic functions.

Application of a maximal-clique based community detection algorithm to gut microbiome data reveals driver microbes during influenza A virus infection.

Influenza A Virus (IAV) infection followed by bacterial pneumonia often leads to hospitalization and death in individuals from high risk groups. Following infection, IAV triggers the process of viral RNA replication which in turn disrupts healthy gut microbial community, while the gut microbiota plays an instrumental role in protecting the host by evolving colonization resistance. Although the underlying mechanisms of IAV infection have been unraveled, the underlying complex mechanisms evolved by gut microbiota in order to induce host immune response following IAV infection remain evasive. In this work, we developed a novel Maximal-Clique based Community Detection algorithm for Weighted undirected Networks (MCCD-WN) and compared its performance with other existing algorithms using three sets of benchmark networks. Moreover, we applied our algorithm to gut microbiome data derived from fecal samples of both healthy and IAV-infected pigs over a sequence of time-points. The results we obtained from the real-life IAV dataset unveil the role of the microbial families Ruminococcaceae, Lachnospiraceae, Spirochaetaceae and Prevotellaceae in the gut microbiome of the IAV-infected cohort. Furthermore, the additional integration of metaproteomic data enabled not only the identification of microbial biomarkers, but also the elucidation of their functional roles in protecting the host following IAV infection. Our network analysis reveals a fast recovery of the infected cohort after the second IAV infection and provides insights into crucial roles of Desulfovibrionaceae and Lactobacillaceae families in combating Influenza A Virus infection. Source code of the community detection algorithm can be downloaded from https://github.com/AniBhar84/MCCD-WN.

A Multi-Omics Protocol for Swine Feces to Elucidate Longitudinal Dynamics in Microbiome Structure and Function.

Swine are regarded as promising biomedical models, but the dynamics of their gastrointestinal microbiome have been much less investigated than that of humans or mice. The aim of this study was to establish an integrated multi-omics protocol to investigate the fecal microbiome of healthy swine. To this end, a preparation and analysis protocol including integrated sample preparation for meta-omics analyses of deep-frozen feces was developed. Subsequent data integration linked microbiome composition with function, and metabolic activity with protein inventories, i.e., 16S rRNA data and expressed proteins, and identified proteins with corresponding metabolites. 16S rRNA gene amplicon and metaproteomics analyses revealed a fecal microbiome dominated by Prevotellaceae, Lactobacillaceae, Lachnospiraceae, Ruminococcaceae and Clostridiaceae. Similar microbiome compositions in feces and colon, but not ileum samples, were observed, showing that feces can serve as minimal-invasive proxy for porcine colon microbiomes. Longitudinal dynamics in composition, e.g., temporal decreased abundance of Lactobacillaceae and Streptococcaceae during the experiment, were not reflected in microbiome function. Instead, metaproteomics and metabolomics showed a rather stable functional state, as evident from short-chain fatty acids (SCFA) profiles and associated metaproteome functions, pointing towards functional redundancy among microbiome constituents. In conclusion, our pipeline generates congruent data from different omics approaches on the taxonomy and functionality of the intestinal microbiome of swine.

Re-emergence of porcine epidemic diarrhea virus in a piglet-producing farm in northwestern Germany in 2019.

BACKGROUND: Porcine epidemic diarrhea (PED) is a viral enteric disease of pigs. It affects all age classes of animals but lethality is mainly seen in suckling piglets. After its first appearance in England in 1971, Porcine epidemic diarrhea virus (PEDV) has spread worldwide. While sporadic outbreaks prevailed in Europe, the disease had high impact in Asia. Following particularly severe outbreaks in 2011, high impact cases were also reported in the United States and neighboring countries in 2013. Subsequently, outbreaks were also reported in several European countries including Germany. These outbreaks were less severe. This case report describes a recent case of PED re-emergence in Germany and the sequence analyses of the causative PEDV. CASE PRESENTATION: In spring 2019 5 years after re-introduction of PED into Central Europe, a piglet-producer in northwestern Germany experienced an outbreak that affected sows, their suckling piglets, and weaners. After initial confirmation of PEDV by real-time RT-PCR, fecal material and small intestine samples from affected pigs were subjected to metagenomic analyses employing next-generation sequencing. Phylogenetic analyses showed high identities among the PEDV sequences obtained from samples of different animals and a close relation to recent strains from Hungary and France. Compared to the PEDV strains analyzed in 2014, genetic drift could be confirmed. Changes were mainly observed in the spike protein encoding S gene segment. In addition, metagenomic analyses showed multiple Picobirnavirus reads in all investigated samples. CONCLUSION: This case report shows that PEDV is still circulating in Europe. The causative strains are moderately virulent and are still closely related to the so-called INDEL strains reported previously in Europe, including Germany. However, a genetic drift has taken place that can be seen in a novel cluster comprising strains from Germany, Hungary and France in 2019. Relevance and impact of the detected Picobirna sequences need further investigations.

Experimental H1N1pdm09 infection in pigs mimics human seasonal influenza infections.

Pigs are anatomically, genetically and physiologically comparable to humans and represent a natural host for influenza A virus (IAV) infections. Thus, pigs may represent a relevant biomedical model for human IAV infections. We set out to investigate the systemic as well as the local immune response in pigs upon two subsequent intranasal infections with IAV H1N1pdm09. We detected decreasing numbers of peripheral blood lymphocytes after the first infection. The simultaneous increase in the frequencies of proliferating cells correlated with an increase in infiltrating leukocytes in the lung. Enhanced perforin expression in αß and γδ T cells in the respiratory tract indicated a cytotoxic T cell response restricted to the route of virus entry such as the nose, the lung and the bronchoalveolar lavage. Simultaneously, increasing frequencies of CD8αα expressing αß T cells were observed rapidly after the first infection, which may have inhibited uncontrolled inflammation in the respiratory tract. Taking together, the results of this study demonstrate that experimental IAV infection in pigs mimics major characteristics of human seasonal IAV infections.

VOC breath profile in spontaneously breathing awake swine during Influenza A infection.

Influenza is one of the most common causes of virus diseases worldwide. Virus detection requires determination of Influenza RNA in the upper respiratory tract. Efficient screening is not possible in this way. Analysis of volatile organic compounds (VOCs) in breath holds promise for non-invasive and fast monitoring of disease progression. Breath VOC profiles of 14 (3 controls and 11 infected animals) swine were repeatedly analyzed during a complete infection cycle of Influenza A under high safety conditions. Breath VOCs were pre-concentrated by means of needle trap micro-extraction and analysed by gas chromatography mass spectrometry before infection, during virus presence in the nasal cavity, and after recovery. Six VOCs could be related to disease progression: acetaldehyde, propanal, n-propyl acetate, methyl methacrylate, styrene and 1,1-dipropoxypropane. As early as on day four after inoculation, when animals were tested positive for Influenza A, differentiation between control and infected animals was possible. VOC based information on virus infection could enable early detection of Influenza A. As VOC analysis is completely non-invasive it has potential for large scale screening purposes. In a perspective, breath analysis may offer a novel tool for Influenza monitoring in human medicine, animal health control or border protection.