Mustapha Berri

Early-life establishment of the swine gut microbiome and impact on host phenotypes

Early bacterial colonization and succession within the gastrointestinal tract has been suggested to be crucial in the establishment of specific microbiota composition and the shaping of host phenotype. Here, the composition and dynamics of faecal microbiomes were studied for 31 healthy piglets across five age strata (days 14, 36, 48, 60 and 70 after birth) together with their mothers. Faecal microbiome composition was assessed by 16S rRNA gene 454-pyrosequencing. Bacteroidetes and Firmicutes were the predominant phyla present at each age. For all piglets, luminal secretory IgA concentration was measured at day 70, and body weight was recorded until day 70. The microbiota of suckling piglets was mainly represented by Bacteroides, Oscillibacter, Escherichia/Shigella, Lactobacillus and unclassified Ruminococcaceae genera. This pattern contrasted with that of Acetivibrio, Dialister, Oribacterium, Succinivibrio and Prevotella genera, which appeared increased after weaning. Lactobacillus fermentum might be vertically transferred via breast milk or faeces. The microbiota composition coevolved with their hosts towards two different clusters after weaning, primarily distinguished by unclassified Ruminococcaceae and Prevotella abundances. Prevotella was positively correlated with luminal secretory IgA concentrations, and body weight. Our study opens up new possibilities for health and feed efficiency manipulation via genetic selection and nutrition in the agricultural domain.

Phylogenetic network analysis applied to pig gut microbiota identifies an ecosystem structure linked with growth traits

The ecological interactions within the gut microbial communities are complex and far from being fully understood. Here we report the first study that aims at defining the interaction network of the gut microbiota in pigs and comparing it with the enterotype-like clustering analysis. Fecal microbiota of 518 healthy piglets was characterized by 16S ribosomal RNA gene sequencing. Two networks were constructed at the genus and operational taxonomic unit levels. Within-network interactions mirrored the human gut microbiota relationships, with a strong co-exclusion between Prevotella and Ruminococcus genera, and were consistent with the two enterotype-like clusters identified in the pig microbiota. Remarkably, the cluster classification of the individuals was significantly associated with the body weight at 60 days of age (P=0.005) and average daily gain (P=0.027). To the best of our knowledge, this is the first study to provide an integrated overview of the porcine gut microbiota that suggests a conservation of the ecological community interactions and functional architecture between humans and pig. Moreover, we show that the microbial ecosystems and porcine growth traits are linked, which allows us to foresee that the enterotype concept may have an important role in the animal production industry.

SOCS proteins in infectious diseases of mammals.

Abstract As for most biological processes, the immune response to microbial infections has to be tightly controlled to remain beneficial for the host. Inflammation is one of the major consequences of the hosts immune response. For its orchestration, this process requires a fine-tuned interplay between interleukins, endothelial cells and various types of recruited immune cells. Suppressors of cytokine signalling (SOCS) proteins are crucially involved in the complex control of the inflammatory response through their actions on various signalling pathways including the JAK/STAT and NF-κB pathways. Due to their cytokine regulatory functions, they are frequent targets for exploitation by infectious agents trying to escape the hosts immune response. This review article aims to summarize our current knowledge regarding SOCS family members in the different mammalian species studied so far, and to display their complex molecular interactions with microbial pathogens.

Pattern recognition receptors in the gut: analysis of their expression along the intestinal tract and the crypt/villus axis

Pattern recognition receptors (PRRs) play a critical role in the detection of microorganisms and the induction of inflammatory and immune responses. Using PCR and Western‐blot analysis, this study investigated the differential expression in the intestine of 14 PRRs and nine associated cytokines. Thirty‐two pigs were used to determine the expression of these markers (1) along the proximal/distal axis of the small intestine (duodenum, jejunum, and ileum) and (2) between the intestinal segments and their respective lymphoid organs (Peyers patches [PP] and mesenteric lymph nodes [MLN]). Six additional animals were used to quantify the expression of these genes along the crypt/villus axis of jejunum, using microdissected samples. Most genes showed increased expression (1) in the distal than in the proximal parts of the small intestine (TLR3, 5, RIG‐I, IL‐1β, IL‐8, and IFN‐γ); (2) in lymphoid organs (TLR1, 2, 6, 9, 10, IL‐10, TNF‐α), especially the MLN (TLR4, 7, 8, NOD1, NOD2, NALP3, IFN‐α, IL‐6, IL‐12, and TGF‐β), than in intestinal segments. The analysis along the crypt/villus identified: (1) genes with higher expression in lamina propria (TLR1, 2, 4, 9, NOD1, NOD2, IL‐1β, IL‐10, TGF‐β, TNF‐α) and (2) genes with higher expression in the villus (TLR3, 5, 6, RIG‐I, IL‐6). These results highlight the differential expression of PRRs and cytokines along the proximal/distal and the crypt/villus axis of the intestine, contributing to a fine analysis of the complex functional architecture of the small intestine and should be related to the gut microbiota.

Innate immune response to a H3N2 subtype swine influenza virus in newborn porcine trachea cells, alveolar macrophages, and precision-cut lung slices

Viral respiratory diseases remain of major importance in swine breeding units. Swine influenza virus (SIV) is one of the main known contributors to infectious respiratory diseases. The innate immune response to swine influenza viruses has been assessed in many previous studies. However most of these studies were carried out in a single-cell population or directly in the live animal, in all its complexity. In the current study we report the use of a trachea epithelial cell line (newborn pig trachea cells – NPTr) in comparison with alveolar macrophages and lung slices for the characterization of innate immune response to an infection by a European SIV of the H3N2 subtype. The expression pattern of transcripts involved in the recognition of the virus, interferon type I and III responses, and the host-response regulation were assessed by quantitative PCR in response to infection. Some significant differences were observed between the three systems, notably in the expression of type III interferon mRNA. Then, results show a clear induction of JAK/STAT and MAPK signaling pathways in infected NPTr cells. Conversely, PI3K/Akt signaling pathways was not activated. The inhibition of the JAK/STAT pathway clearly reduced interferon type I and III responses and the induction of SOCS1 at the transcript level in infected NPTr cells. Similarly, the inhibition of MAPK pathway reduced viral replication and interferon response. All together, these results contribute to an increased understanding of the innate immune response to H3N2 SIV and may help identify strategies to effectively control SIV infection.

Development of gut immunoglobulin A production in piglet in response to innate and environmental factors

The current review focuses on pre- and post-natal development of intestinal immunoglobulin A (IgA) production in pig. IgA production is influenced by intrinsic genetic factors in the foetus as well as extrinsic environmental factors during the post-natal period. At birth, piglets are exposed to new antigens through maternal colostrums/milk as well as exogenous microbiota. This exposure to new antigens is critical for the proper development of the gut mucosal immune system and is characterized mainly by the establishment of IgA response. A second critical period for neonatal intestinal immune system development occurs at weaning time when the gut environment is exposed to new dietary antigens. Neonate needs to establish oral tolerance and in the absence of protective milk need to fight potential new pathogens. To improve knowledge about the immune response in the neonates, it is important to identify intrinsic and extrinsic factors which influence the intestinal immune system development and to elucidate their mechanism of action.

Extensive Expression Differences along Porcine Small Intestine Evidenced by Transcriptome Sequencing

The aim of this study was to analyse gene expression along the small intestine (duodenum, jejunum, ileum) and in the ileal Peyers patches in four young pigs with no clinical signs of disease by transcriptome sequencing. Multidimensional scaling evidenced that samples clustered by tissue type rather than by individual, thus prefiguring a relevant scenario to draw tissue-specific gene expression profiles. Accordingly, 1,349 genes were found differentially expressed between duodenum and jejunum, and up to 3,455 genes between duodenum and ileum. Additionally, a considerable number of differentially expressed genes were found by comparing duodenum (7,027 genes), jejunum (6,122 genes), and ileum (6,991 genes) with ileal Peyers patches tissue. Functional analyses revealed that most of the significant differentially expressed genes along small intestinal tissues were involved in the regulation of general biological processes such as cell development, signalling, growth and proliferation, death and survival or cell function and maintenance. These results suggest that the intrinsic large turnover of intestinal tissues would have local specificities at duodenum, ileum and jejunum. In addition, in concordance with their biological function, enteric innate immune pathways were overrepresented in ileal Peyers patches. The reported data provide an expression map of the cell pathway variation in the different small intestinal tissues. Furthermore, expression levels measured in healthy individuals could help to understand changes in gene expression that occur in dysbiosis or pathological states.

Mycoplasma hyopneumoniae does not affect the interferon-related anti-viral response but predisposes the pig to a higher level of inflammation following swine influenza virus infection.

In pigs, influenza A viruses and Mycoplasma hyopneumoniae (Mhp) are major contributors to the porcine respiratory disease complex. Pre-infection with Mhp was previously shown experimentally to exacerbate the clinical outcomes of H1N1 infection during the first week after virus inoculation. In order to better understand the interactions between these pathogens, we aimed to assess very early responses (at 5, 24 and 48 h) after H1N1 infection in pigs pre-infected or not with Mhp. Clinical signs and macroscopic lung lesions were similar in both infected groups at early times post-H1N1 infection; and Mhp pre-infection affected neither the influenza virus replication nor the IFN-induced antiviral responses in the lung. However, it predisposed the animals to a higher inflammatory response to H1N1 infection, as revealed by the massive infiltration of neutrophils and macrophages into the lungs and the increased production of pro-inflammatory cytokines (IL-6, IL-1β and TNF-α). Thus, it seems it is this marked inflammatory state that would play a role in exacerbating the clinical signs subsequent to H1N1 infection.

Better horizontal transmission of a US non-InDel strain compared with a French InDel strain of porcine epidemic diarrhoea virus

Abstract From the severe porcine epidemic diarrhoea (PED) epidemics that struck in 2013 in the United States of America and other countries of North and South America, two types of porcine epidemic diarrhoea virus (PEDV) were isolated, namely the InDel and the non‐InDel strains. They are differentiated by insertions/deletions in the S1 nucleotide sequence of the S gene, and differences in virulence were observed from the clinical cases. In 2014, a PED outbreak occurred in a pig farm in France, from which an InDel strain was isolated. This study aimed at comparing, under experimental conditions, the pathogenicity and the direct and indirect transmissions between a non‐InDel strain isolated from a PED‐affected piglet in 2014 in the USA and the French InDel strain. All infected pigs showed clinical signs with the non‐InDel strain although only the inoculated and direct contact pigs showed clinical signs in the InDel strain group. Although viral RNA was detected in air samples with both strains, the indirect contact pigs remained free from infection with the InDel strain in contrast to the non‐InDel group in which airborne transmission occurred in the indirect contact pigs. All infected pigs shed virus in faeces regardless of PEDV strain with 9 of 30 pigs showing intermittent faecal shedding. The transmission rate by direct contact was found to be 2.17‐fold higher than the non‐InDel strain compared with the InDel. In conclusion, the InDel strain was less pathogenic than the non‐InDel strain in our experimental conditions. The transmission route differed between the two strains. Direct contact was the main transmission route for the InDel strain, although the non‐InDel strain was transmitted through direct contact and indirectly through the air.

Immunome differences between porcine ileal and jejunal Peyer’s patches revealed by global transcriptome sequencing of gut-associated lymphoid tissues

The epithelium of the intestinal mucosa and the gut-associated lymphoid tissues (GALT) constitute an essential physical and immunological barrier against pathogens. In order to study the specificities of the GALT transcriptome in pigs, we compared the transcriptome profiles of jejunal and ileal Peyer’s patches (PPs), mesenteric lymph nodes (MLNs) and peripheral blood (PB) of four male piglets by RNA-Seq. We identified 1,103 differentially expressed (DE) genes between ileal PPs (IPPs) and jejunal PPs (JPPs), and six times more DE genes between PPs and MLNs. The master regulator genes FOXP3, GATA3, STAT4, TBX21 and RORC were less expressed in IPPs compared to JPPs, whereas the transcription factor BCL6 was found more expressed in IPPs. In comparison between IPPs and JPPs, our analyses revealed predominant differential expression related to the differentiation of T cells into Th1, Th2, Th17 and iTreg in JPPs. Our results were consistent with previous reports regarding a higher T/B cells ratio in JPPs compared to IPPs. We found antisense transcription for respectively 24%, 22% and 14% of the transcripts detected in MLNs, PPs and PB, and significant positive correlations between PB and GALT transcriptomes. Allele-specific expression analyses revealed both shared and tissue-specific cis-genetic control of gene expression.