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The nasal microbiota is a key contributor to animal health, and characterizing the nasal microbiota composition is an important step towards elucidating the role of its different members. Efforts to characterize the nasal microbiota composition of domestic pigs and other farm animals frequently report the presence of bacteria that are typically found in the gut, including many anaerobes from the Bacteroidales and Clostridiales orders. However, the in vivo role of these gut-microbiota associated taxa is currently unclear. Here, we tackled this issue by examining the prevalence, origin, and activity of these taxa in the nasal microbiota of piglets. First, analysis of the nasal microbiota of farm piglets sampled in this study, as well as various publicly available data sets, revealed that gut-microbiota associated taxa indeed constitute a substantial fraction of the pig nasal microbiota that is highly variable across individual animals. Second, comparison of herd-matched nasal and rectal samples at amplicon sequencing variant (ASV) level showed that these taxa are largely shared in the nasal and rectal microbiota, suggesting a common origin driven presumably by the transfer of fecal matter. Third, surgical sampling of the inner nasal tract showed that gut-microbiota associated taxa are found throughout the nasal cavity, indicating that these taxa do not stem from contaminations introduced during sampling with conventional nasal swabs. Finally, analysis of cDNA from the 16S rRNA gene in these nasal samples indicated that gut-microbiota associated taxa are indeed active in the pig nasal cavity. This study shows that gut-microbiota associated taxa are not only present, but also active, in the nasal cavity of domestic pigs, and paves the way for future efforts to elucidate the function of these taxa within the nasal microbiota.

The view on antimicrobials has dramatically changed due to the increased knowledge on the importance of microbiota composition in different body parts. Antimicrobials can no longer be considered only beneficial, but also potentially deleterious for favourable bacterial populations. Still, the use of metaphylactic antimicrobial treatment at early stages of life is a practice in use in porcine production. Many reports have shown that antibiotics can critically affect the gut microbiota, however the effect of perinatal antimicrobial treatment on the nasal microbiota has not been explored yet. To gain insights on the potential changes in nasal microbial composition due to antimicrobial treatments, piglets from two different farms were sampled at weaning. The nasal microbiota was analysed when antimicrobial treatment was used early in life, and later, when no antimicrobial treatment was used during the lactation period. Removal of perinatal antimicrobials resulted in an increased bacterial diversity in nasal microbiota at weaning. Concurrently, elimination of antimicrobials produced an increase in the relative abundance of Prevotella and Lactobacillus , and a decrease in Moraxella and Bergeyella . These changes in microbiota composition were accompanied by an improvement of the piglets’ health and a higher productivity in the nursery phase.

Glaesserella (formerly Haemophilus ) parasuis causes Glässer’s disease, which results in high economic loss in the swine industry. To understand the polymicrobial interactions of G. parasuis and the nasal microbiota, the statistical association patterns of nasal colonizing bacteria with virulent and non-virulent strains of G. parasuis were studied accounting for the farm management practices as potential risk factors for the occurrence of Glässer’s disease. The nasal microbiota from 51 weaned-piglets from four farms with Glässer’s disease and three farms with no respiratory diseases was previously characterized and included in this study. The presence of virulent and/or non-virulent G. parasuis strains in the nasal cavities was determined in order to establish the potential association with other members of the nasal microbiota. Multivariate logistic and linear regression models were performed among the various members of nasal microbiota and G. parasuis . The multi-site production system and disease presence in the farm were both significantly associated with the presence of G. parasuis virulent strains in the nose of the piglets. Differential bacterial associations were observed with virulent or non-virulent G. parasuis . Chitinophagaceae , Corynebacteriaceae and Corynebacterium were positively associated with the virulent G. parasuis strains, while Enterobacteriaceae, Peptostreptococcaceae , Clostridium XI, and Escherichia/Shigella were negatively associated with virulent G. parasuis . On the other hand, Flavobacteriaceae , Planobacterium , and Phascolarctobacterium were positively associated with the non-virulent G. parasuis strains, while Rikenellaceae , Enterococcaceae , Odoribacter , and Corynebacterium were negatively associated with non-virulent G. parasuis . In conclusion, the nasal microbiota communities showed variations in the association with the G. parasuis strains type.

Glaesserella parasuis is the etiological agent of Glässer’s disease, a common pathology in the pork industry with higher prevalence in the postweaning period. Vaccination is one of the strategies to control this disease. Here, we investigated the effect that sow vaccination against virulent strains of G. parasuis had in the nasal microbiota of their offspring. Nasal swabs from fifteen days-old piglets from vaccinated (vs-P, n = 11) and unvaccinated sows (cs-P, n = 11) were obtained and DNA was extracted for 16S amplicon sequencing. Microbiota composition was different, with lower diversity in vs-P, and a strong clustering of the groups in beta diversity analysis. Among the 1509 sequences associated to either study group, all the sequences classified as G. parasuis (10 ASVs) had lower relative abundance in the vs-P group. A list of 32 inferred metabolic pathways were statistically different between groups. A distinctive structure of the two microbial networks was detected, with modules in the cs-P not conserved in the vs-P network. In conclusion, vaccination of the sows had a large effect in the microbiota composition of their offspring that went beyond the effect on the targeted pathogen. The mechanisms underneath these changes may include alteration of the microbiota network due to the elimination of the targeted pathogen and/or immunological changes.

The nasal microbiota plays an important role in animal health and the use of antibiotics is a major factor that influences its composition. Here, we studied the consequences of an intensive antibiotic treatment, applied to sows and/or their offspring, on the piglets’ nasal microbiota. Four pregnant sows were treated with crystalline ceftiofur and tulathromycin (CT sows ) while two other sows received only crystalline ceftiofur (C sows ). Sow treatments were performed at D-4 (four days pre-farrowing), D3, D10 and D17 for ceftiofur and D-3, D4 and D11 for tulathromycin. Half of the piglets born to CT sows were treated at D1 with ceftiofur. Nasal swabs were taken from piglets at 22–24 days of age and bacterial load and nasal microbiota composition were defined by 16 s rRNA gene qPCR and amplicon sequencing. Antibiotic treatment of sows reduced their nasal bacterial load, as well as in their offspring, indicating a reduced bacterial transmission from the dams. In addition, nasal microbiota composition of the piglets exhibited signs of dysbiosis, showing unusual taxa. The addition of tulathromycin to the ceftiofur treatment seemed to enhance the deleterious effect on the microbiota diversity by diminishing some bacteria commonly found in the piglets’ nasal cavity, such as Glaesserella , Streptococcus , Prevotella , Staphylococcus and several members of the Ruminococcaceae and Lachnospiraceae families. On the other hand, the additional treatment of piglets with ceftiofur resulted in no further effect beyond the treatment of the sows. Altogether, these results suggest that intensive antibiotic treatments of sows, especially the double antibiotic treatment, disrupt the nasal microbiota of their offspring and highlight the importance of sow-to-piglet microbiota transmission.

Glaesserella parasuis strains were characterized by serotyping PCR, vtaA virulence marker Leader Sequence (LS)-PCR, clinical significance, and geographic region. Overall, the serovars 4, 5/12, 7, 1, and 13 were the most commonly detected. Serovars of greatest clinical relevance were systemic isolates that had a higher probability of being serovar 5/12, 13, or 7. In comparison, pulmonary isolates had a higher likelihood of being serovars 2, 4, 7, or 14. Serovars 5/12 and 13 have previously been considered disease-associated, but this study agrees with other recent studies showing that serovar 7 is indeed associated with systemic G. parasuis disease. Serovar 4 strains illustrated how isolates can have varying degrees of virulence and be obtained from pulmonary, systemic, or nasal sites. Serovars 8, 9, 15, and 10 were predominantly obtained from nasal samples, which indicates a limited clinical significance of these serovars. Additionally, most internal G. parasuis isolates were classified as virulent by LS-PCR and were disease-associated isolates, including serovars 1, 2, 4, 5/12, 7, 13, and 14. Isolates from the nasal cavity, including serovars 6, 9, 10, 11, and 15, were classified as non-virulent by LS-PCR. In conclusion, the distribution of G. parasuis serovars remains constant, with few serovars representing most of the strains isolated from affected pigs. Moreover, it was confirmed that the LS-PCR can be used for G. parasuis virulence prediction of field strains worldwide.

The nasal microbiota composition is associated with the health status of piglets. Sow-contact in early life is one of the factors influencing the microbial composition in piglets; however, its impact has never been assessed in the nasal microbiota of piglets reared in controlled environmental conditions. Nasal microbiota of weaning piglets in high-biosecurity facilities with different time of contact with their sows (no contact after farrowing, contact limited to few hours or normal contact until weaning at three weeks) was unveiled by 16S rRNA gene sequencing. Contact with sows demonstrated to be a major factor affecting the nasal microbial composition of the piglets. The nasal microbiota of piglets that had contact with sows until weaning, but were reared in high biosecurity facilities, was richer and more similar to the previously described healthy nasal microbiota from conventional farm piglets. On the other hand, the nasal communities inhabiting piglets with no or limited contact with sows was different and dominated by bacteria not commonly abundant in this body site. Furthermore, the length of sow–piglet contact was also an important variable. In addition, the piglets raised in BSL3 conditions showed an increased richness of low-abundant species in the nasal microbiota. Artificially rearing in high biosecurity facilities without the contact of sows as a source of nasal colonizers had dramatic impacts on the nasal microbiota of weaning piglets and may introduce significant bias into animal research under these conditions.

Streptococcus suis is a zoonotic pathogen of swine involved in arthritis, polyserositis, and meningitis. Colonization of piglets by S. suis is very common and occurs early in life. The clinical outcome of infection is influenced by the virulence of the S. suis strains and the immunity of the animals. Here, the role of innate immunity was studied in cesarean-derived colostrum-deprived piglets inoculated intranasally with either virulent S. suis strain 10 (S10) or non-virulent S. suis strain T15. Colonization of the inoculated piglets was confirmed at the end of the study by PCR and immunohistochemistry. Fever (≥40.5 °C) was more prevalent in piglets inoculated with S10 compared to T15 at 4 h after inoculation. During the 3 days of monitoring, no other major clinical signs were detected. Accordingly, only small changes in transcription of genes associated with the antibacterial innate immune response were observed at systemic sites, with S10 inducing an earlier response than T15 in blood. Local inflammatory response to the inoculation, evaluated by transcriptional analysis of selected genes in nasal swabs, was more sustained in piglets inoculated with the virulent S10, as demonstrated by transcription of inflammation-related genes, such as IL1B , IL1A , and IRF7 . In contrast, most of the gene expression changes in trachea, lungs, and associated lymph nodes were observed in response to the non-virulent T15 strain. Thus, S. suis colonization in the absence of systemic infection induces an innate immune response in piglets that appears to be related to the virulence potential of the colonizing strain.

Medicated feed is a common strategy to control the occurrence of Streptococcus suis disease in swine production, but feed additives may constitute an alternative to metaphylaxis. In a farm with post-weaning S. suis disease, the following additives were tested: lysozyme (Lys), medium chain fatty acids plus lysozyme (FA + Lys), FA plus a natural anti-inflammatory (FA + antiinf) and amoxicillin (Amox). During the course of the study, FA + antiinf and Amox groups showed lower prevalence of clinical signs compatible with S. suis disease than the rest of the groups. Piglets from the FA + antiinf group showed high diversity and richness in their nasal and faecal microbiota. Diet supplements did not have major effects on the faecal microbiota, where the genus Mitsuokella was the only differentially present in the FA + Lys group. In the nasal microbiota, piglets from FA + antiinf presented higher differential abundance of a sequence variant from Ruminococcaceae and lower abundance of an unclassified genus from Weeksellaceae . In general, we detected more significant changes in the nasal than in the feacal microbiota, and found that parity of the dams affected the microbiota composition of their offspring, with piglets born to gilts exhibiting lower richness and diversity. Our results suggest that additives could be useful to control post-weaning disease when removing antimicrobials in farms.

Haemophilus parasuis is part of the microbiota of the upper respiratory tract in swine. However, virulent strains can cause a systemic disease known as Glässer’s disease. Several virulence factors have been described in H. parasuis including the virulence-associated trimeric autotransporters (VtaAs). VtaA2 is up-regulated during infection and is only found in virulent strains. In order to determine its biological function, the vtaA2 gene was cloned with its native promotor region in pACYC184, and the transformed Escherichia coli was used to perform functional in vitro assays. VtaA2 was found to have a role in attachment to plastic, mucin, BSA, fibronectin and collagen. As other VtaAs from H. parasuis , the passenger domain of VtaA2 contains collagen domains. In order to examine the contribution of the collagen repeats to VtaA2 function, a recombinant vtaA2 without the central collagen domains was obtained and named vtaA2OL . VtaA2OL showed similar capacity than VtaA2 to adhere to plastic, mucin, BSA, fibronectin and plasma but a reduced capacity to adhere to collagen, suggesting that the collagen domains of VtaA2 are involved in collagen attachment. No function in cell adhesion and invasion to epithelial alveolar cell line A549 or unspecific binding to primary alveolar macrophages was found. Likewise VtaA2 had no role in serum or phagocytosis resistance. We propose that VtaA2 mediates adherence to the host by binding to the mucin, found in the upper respiratory tract mucus, and to the extracellular matrix proteins, present in the connective tissue of systemic sites, such as the serosa.