Histo-Blood Group Antigen-Producing Bacterial Cocktail Reduces Rotavirus A, B, and C Infection and Disease in Gnotobiotic Piglets.

The suboptimal performance of rotavirus (RV) vaccines in developing countries and in animals necessitates further research on the development of novel therapeutics and control strategies. To initiate infection, RV interacts with cell-surface O-glycans, including histo-blood group antigens (HBGAs). We have previously demonstrated that certain non-pathogenic bacteria express HBGA- like substances (HBGA+) capable of binding RV particles in vitro. We hypothesized that HBGA+ bacteria can bind RV particles in the gut lumen protecting against RV species A (RVA), B (RVB), and C (RVC) infection in vivo. In this study, germ-free piglets were colonized with HBGA+ or HBGA- bacterial cocktail and infected with RVA/RVB/RVC of different genotypes. Diarrhea severity, virus shedding, immunoglobulin A (IgA) Ab titers, and cytokine levels were evaluated. Overall, colonization with HBGA+ bacteria resulted in reduced diarrhea severity and virus shedding compared to the HBGA- bacteria. Consistent with our hypothesis, the reduced severity of RV disease and infection was not associated with significant alterations in immune responses. Additionally, colonization with HBGA+ bacteria conferred beneficial effects irrespective of the piglet HBGA phenotype. These findings are the first experimental evidence that probiotic performance in vivo can be improved by including HBGA+ bacteria, providing decoy epitopes for broader/more consistent protection against diverse RVs.

Maternal immunization and vitamin A sufficiency impact sow primary adaptive immunity and passive protection to nursing piglets against porcine epidemic diarrhea virus infection.

Porcine epidemic diarrhea virus (PEDV) causes a highly contagious enteric disease with major economic losses to swine production worldwide. Due to the immaturity of the neonatal piglet immune system and given the high virulence of PEDV, improving passive lactogenic immunity is the best approach to protect suckling piglets against the lethal infection. We tested whether oral vitamin A (VA) supplementation and PEDV exposure of gestating and lactating VA-deficient (VAD) sows would enhance their primary immune responses and boost passive lactogenic protection against the PEDV challenge of their piglets. We demonstrated that PEDV inoculation of pregnant VAD sows in the third trimester provided higher levels of lactogenic protection of piglets as demonstrated by >87% survival rates of their litters compared with <10% in mock litters and that VA supplementation to VAD sows further improved the piglets' survival rates to >98%. We observed significantly elevated PEDV IgA and IgG antibody (Ab) titers and Ab-secreting cells (ASCs) in VA-sufficient (VAS)+PEDV and VAD+VA+PEDV sows, with the latter maintaining higher Ab titers in blood prior to parturition and in blood and milk throughout lactation. The litters of VAD+VA+PEDV sows also had the highest serum PEDV-neutralizing Ab titers at piglet post-challenge days (PCD) 0 and 7, coinciding with higher PEDV IgA ASCs and Ab titers in the blood and milk of their sows, suggesting an immunomodulatory role of VA in sows. Thus, sows that delivered sufficient lactogenic immunity to their piglets provided the highest passive protection against the PEDV challenge. Maternal immunization during pregnancy (± VA) and VA sufficiency enhanced the sow primary immune responses, expression of gut-mammary gland trafficking molecules, and passive protection of their offspring. Our findings are relevant to understanding the role of VA in the Ab responses to oral attenuated vaccines that are critical for successful maternal vaccination programs against enteric infections in infants and young animals.

Differential transcriptome response following infection of porcine ileal enteroids with species A and C rotaviruses.

BACKGROUND: Rotavirus C (RVC) is the major causative agent of acute gastroenteritis in suckling piglets, while most RVAs mostly affect weaned animals. Besides, while most RVA strains can be propagated in MA-104 and other continuous cell lines, attempts to isolate and culture RVC strains remain largely unsuccessful. The host factors associated with these unique RVC characteristics remain unknown. METHODS: In this study, we have comparatively evaluated transcriptome responses of porcine ileal enteroids infected with RVC G1P[1] and two RVA strains (G9P[13] and G5P[7]) with a focus on innate immunity and virus-host receptor interactions. RESULTS: The analysis of differentially expressed genes regulating antiviral immune response indicated that in contrast to RVA, RVC infection resulted in robust upregulation of expression of the genes encoding pattern recognition receptors including RIG1-like receptors and melanoma differentiation-associated gene-5. RVC infection was associated with a prominent upregulation of the most of glycosyltransferase-encoding genes except for the sialyltransferase-encoding genes which were downregulated similar to the effects observed for G9P[13]. CONCLUSIONS: Our results provide novel data highlighting the unique aspects of the RVC-associated host cellular signalling and suggest that increased upregulation of the key antiviral factors maybe one of the mechanisms responsible for RVC age-specific characteristics and its inability to replicate in most cell cultures.

Host Cell Response to Rotavirus Infection with Emphasis on Virus-Glycan Interactions, Cholesterol Metabolism, and Innate Immunity.

Although rotavirus A (RVA) is the primary cause of acute viral gastroenteritis in children and young animals, mechanisms of its replication and pathogenesis remain poorly understood. We previously demonstrated that the neuraminidase-mediated removal of terminal sialic acids (SAs) significantly enhanced RVA-G9P[13] replication, while inhibiting RVA-G5P[7] replication. In this study, we compared the transcriptome responses of porcine ileal enteroids (PIEs) to G5P[7] vs. G9P[13] infections, with emphasis on the genes associated with immune response, cholesterol metabolism, and host cell attachment. The analysis demonstrated that G9P[13] infection led to a robust modulation of gene expression (4093 significantly modulated genes vs. 488 genes modulated by G5P[7]) and a significant modulation of glycosyltransferase-encoding genes. The two strains differentially affected signaling pathways related to immune response, with G9P[13] mostly upregulating and G5P[7] inhibiting them. Both RVAs modulated the expression of genes encoding for cholesterol transporters. G9P[13], but not G5P[7], significantly affected the ceramide synthesis pathway known to affect both cholesterol and glycan metabolism. Thus, our results highlight the unique mechanisms regulating cellular response to infection caused by emerging/re-emerging and historical RVA strains relevant to RVA-receptor interactions, metabolic pathways, and immune signaling pathways that are critical in the design of effective control strategies.

Vitamin A deficiency and vitamin A supplementation affect innate and T cell immune responses to rotavirus A infection in a conventional sow model.

Rotavirus A (RVA) causes ~200,000 diarrheal deaths annually in children <5yrs, mostly in low- and middle-income countries. Risk factors include nutritional status, social factors, breastfeeding status, and immunodeficiency. We evaluated the effects of vitamin A (VA) deficiency/VA supplementation and RVA exposure (anamnestic) on innate and T cell immune responses in RVA seropositive pregnant and lactating sows and passive protection of their piglets post-RVA challenge. Sows were fed VA deficient (VAD) or sufficient (VAS) diets starting at gestation day (GD)30. A subset of VAD sows received VA supplementation from GD|76 (30,000IU/day, VAD+VA). Sows (6 groups) were inoculated with porcine RVA G5P[7] (OSU strain) or Minimal Essential Medium (mock) at GD~90: VAD+RVA; VAS+RVA; VAD+VA+RVA; VAD-mock; VAS-mock; and VAD+VA-mock. Blood, milk, and gut-associated tissues were collected from sows at several time points to examine innate [natural killer (NK), dendritic (DC) cells], T cell responses and changes in genes involved in the gut-mammary gland (MG)-immunological axis trafficking. Clinical signs of RVA were evaluated post inoculation of sows and post-challenge of piglets. We observed decreased frequencies of NK cells, total and MHCII+ plasmacytoid DCs, conventional DCs, CD103+ DCs and CD4+/CD8+ and T regulatory cells (Tregs) and NK cell activity in VAD+RVA sows. Polymeric Ig receptor and retinoic acid receptor alpha (RARα) genes were downregulated in mesenteric lymph nodes and ileum of VAD+RVA sows. Interestingly, RVA-specific IFN-γ producing CD4+/CD8+ T cells were increased in VAD-Mock sows, coinciding with increased IL-22 suggesting inflammation in these sows. VA supplementation to VAD+RVA sows restored frequencies of NK cells and pDCs, and NK activity, but not tissue cDCs and blood Tregs. In conclusion, similar to our recent observations of decreased B cell responses in VAD sows that led to decreased passive immune protection of their piglets, VAD impaired innate and T cell responses in sows, while VA supplementation to VAD sows restored some, but not all responses. Our data reiterate the importance of maintaining adequate VA levels and RVA immunization in pregnant and lactating mothers to achieve optimal immune responses, efficient function of the gut-MG-immune cell-axis and to improve passive protection of their piglets.

Immune Impairment Associated with Vitamin A Deficiency: Insights from Clinical Studies and Animal Model Research.

Vitamin A (VA) is critical for many biological processes, including embryonic development, hormone production and function, the maintenance and modulation of immunity, and the homeostasis of epithelium and mucosa. Specifically, VA affects cell integrity, cytokine production, innate immune cell activation, antigen presentation, and lymphocyte trafficking to mucosal surfaces. VA also has been reported to influence the gut microbiota composition and diversity. Consequently, VA deficiency (VAD) results in the imbalanced production of inflammatory and immunomodulatory cytokines, intestinal inflammation, weakened mucosal barrier functions, reduced reactive oxygen species (ROS) and disruption of the gut microbiome. Although VAD is primarily known to cause xerophthalmia, its role in the impairment of anti-infectious defense mechanisms is less defined. Infectious diseases lead to temporary anorexia and lower dietary intake; furthermore, they adversely affect VA status by interfering with VA absorption, utilization and excretion. Thus, there is a tri-directional relationship between VAD, immune response and infections, as VAD affects immune response and predisposes the host to infection, and infection decreases the intestinal absorption of the VA, thereby contributing to secondary VAD development. This has been demonstrated using nutritional and clinical studies, radiotracer studies and knockout animal models. An in-depth understanding of the relationship between VAD, immune response, gut microbiota and infections is critical for optimizing vaccine efficacy and the development of effective immunization programs for countries with high prevalence of VAD. Therefore, in this review, we have comprehensively summarized the existing knowledge regarding VAD impacts on immune responses to infections and post vaccination. We have detailed pathological conditions associated with clinical and subclinical VAD, gut microbiome adaptation to VAD and VAD effects on the immune responses to infection and vaccines.

Full-genome analysis and pathogenicity of a genetically distinct Russian PRRSV-1 Tyu16 strain.

Porcine reproductive and respiratory syndrome virus-1 (PRRSV-1) strains from Eastern Europe have a high diversity. All three known subtypes (1, 2, 3) of PRRSV-1 have been detected in Russia. There are two different groups of viruses belonging to the subtype 1: pan-European subtype 1 strains, and insufficiently studied Russian strains. The main objective of this study was to characterize the full genomic structure of the atypical Tyu16 strain of the Russian group subtype 1 PRRSV-1 and to assess its pathogenicity. Complete sequencing of the Tyu16 strain revealed that it did not belong to any existing subtype. Comparison of the whole genome sequence of the Tyu16 strain with that of PRRSV-1 prototype strains revealed 78.1 % (subtype 1 Lelystad), 78.1 % (subtype 2 WestSib13) and 77.7 % (subtype 3 Lena) nucleotide identity level, respectively. The coding sequence of different parts of the Tyu16 strain genome demonstrated a varying percentage identity to the different reference PRRSV-1 strains, which may indicate recombination events in its evolutionary history. We assume that among PRRSV-1 isolates, the Tyu16 is the closest relative to the common ancestor of PRRSV-1 and PRRSV-2. Low pathogenicity of the Tyu16 was demonstrated by experimental infection of 70-day-old piglets. Infected animals showed fever not exceeding 7 days, dyspnea in two out of five pigs and reduced weight gain. The virus shedding was undetectable and viremia was at low level.

First detection and full genome sequence of porcine circovirus type 3 in Russia.

Porcine circovirus type 3 (PCV3) was firstly detected in 2016 in USA. Later PCV3 was discovered in Asia, Europe, and South America. The present investigation demonstrates for the first time the circulation of PCV3 among pigs in Russia. The viruses were detected at two geographically distant unrelated commercial farms with records of reproductive failure (abortions, stillbirth), porcine dermatitis, and nephropathy syndrome (PDNS). The two farms were located in the region of Smolensk (western part of Russia) and the region of Tyumen (West Siberia, Russia). We investigated samples collected from pigs of different ages. We performed PCR for the PCV3 DNA detection. The DNA of PCV3 was detected in serum, kidney, heart, spleen, pleural effusion, and peritoneal cavity fluid samples. Two viral genomes were sequenced and the corresponding strains were named PCV3-RU/SM17 (the strain from Smolensk region) and PCV3-RU/TY17 (the strain from Tyumen region). The full genome sequences of both strains were 2000 nucleotides in length and contained at least two ORFs, encoding the Cap and Rep proteins. Full sequence alignment revealed a 99.3% identity between the PCV3-RU/SM17 and PCV3-RU/TY17 strains. Molecular analysis showed that the two strains from Russia are highly homologous to viruses identified in other countries, with a 98.5-99.6% homology for PCV3-RU/TY17 and 97.9-99.0 for PCV3-RU/SM17. The PCV3-RU/SM17 and PCV3-RU/TY17 strains were found to form a monophyletic group in a phylogenetic tree based on PCV3 complete genome sequences.

Genetic and pathogenic characterization of a Russian subtype 2 PRRSV-1 isolate.

Porcine reproductive and respiratory syndrome virus (PRRSV) causes reproductive failure and respiratory problems. Data about the virulence and pathogenicity of subtype 2 PRRSV-1 strains are limited. The main purposes of this investigation were to characterize the full genome sequence of the subtype 2 PRRSV-1 WestSib13 strain and to compare the pathogenicity with that of the subtype 1 PRRSV-1 Lelystad strain. Comparison of the whole genome sequence of the WestSib13 strain with that of PRRSV-1 prototype strains revealed a 76.2% (subtype 1 Lelystad virus) and 79.0% (subtype 3 Lena virus) identity, respectively The virulence and pathogenicity of the European subtype 2 PRRSV strain WestSib13 and the European subtype 1 PRRSV strain Lelystad were compared in 3-week-old piglets upon inoculation of 105.4 TCID50 of virus. Non-infected animals (control group) as well as animals infected with the Lelystad strain were clinically healthy until 14days post challenge. In contrast, animals infected with the WestSib13 strain demonstrated dyspnea starting at 3days post-inoculation (dpi). All piglets in this group died between 5 and 8 dpi. During that period, fever was not observed in WestSib13-infected animals. Viremia was detected in animals from both infected groups starting from 2 dpi. Viral loads in serum and lungs upon euthanasia were significantly higher (3 log10) in the WestSib13-infected than in the LV-infected animals. Taken together, this study provides the full genome sequence and the unusual virological and clinical outcome (high level viremia without fever) of the novel WestSib13 strain.