However, mock gilts experienced comparable or higher mean numbers of IgA and IgG ASC in the spleen, mesenteric lymph node (MLN), and ileum compared with first, second, and third trimester gilts (Figures S8A,B), corresponding to higher PEDV RNA shedding titers in feces and PEDV-induced diarrhea of mock gilts (Figures S6A,B). Aceglutamide in the third trimester. To determine if lactogenic immunity correlated with protection, all piglets were PEDV-challenged at 3C5 days postpartum. PEDV contamination of gilts at different stages of gestation significantly affected multiple maternal systemic immune parameters prepartum, including cytokines, B cells, PEDV antibodies (Abs), and PEDV antibody secreting cells (ASCs). Pregnant second trimester gilts experienced significantly higher levels of circulating PEDV IgA and IgG Abdominal muscles and ASCs and PEDV computer virus neutralizing (VN) Abdominal muscles post PEDV contamination. Coinciding FGF11 with the significantly higher PEDV Ab responses in second Aceglutamide trimester gilts, the survival rate of their PEDV-challenged piglets was 100%, compared with 87.2, 55.9, and 5.7% for first, Aceglutamide third, and mock litters, respectively. Additionally, piglet survival positively correlated with PEDV IgA Abs and ASCs and VN Abs in milk and PEDV IgA and IgG Abs in piglet serum. Our findings have implications for gestational timing of oral attenuated PEDV maternal vaccines, whereby PEDV intestinal contamination in the second trimester optimally stimulated the gut-MG-sIgA axis resulting in 100% lactogenic immune protection in suckling piglets. Keywords: swine, PEDV, pregnancy, lactogenic immunity, gut-mammary-secretory IgA axis Introduction Diarrheal diseases in young animals account for an estimated multi-million dollar loss to the livestock industry annually due to the livestock industry annually due to mortality, reduced weight gain, treatment costs, and trade sanctions on exporting animal products from infected countries (1, 2). For example, porcine epidemic diarrhea computer virus (PEDV) is a highly virulent re-emerging enteric coronavirus that causes acute diarrhea, dehydration, and death in neonatal piglets (3). It has killed over 8.5 million piglets since its emergence in the US in 2013. In adult pigs, PEDV causes watery diarrhea, depressive disorder, and anorexia as well as agalactia and reduced reproductive overall performance (1C3). Lactogenic immunity remains the most encouraging and effective way to protect neonatal suckling piglets from enteric diseases like PEDV (4, 5). This is dependent on trafficking of pathogen-specific IgA+ plasmablasts to the mammary gland (MG) and accumulation of secretory IgA (sIgA) antibodies (Abs) in milk, defined as the gut-MG-sIgA axis (6C8). Understanding the regulation of mucosal homing receptor and chemokine expression is critical to generate sufficient lactogenic immunity for piglet protection. For example, chemokine receptor (CCR)10, a lymphocyte gut homing marker, is required for IgA+ plasmablast recruitment to the MG in mice and humans (9C11). Additionally, an increase in lymphocyte migration to the MG in swine at the end of gestation and during lactation coincides with an increase in 47 integrin, another lymphocyte gut homing marker, on B cells (12). Identifying factors that influence lymphocyte migration and the gut-MG-sIgA axis may lead to improved PEDV vaccine regimens in gestating swine, improving overall herd immunity and health and industry productivity. Maternal vaccination that increases the amount of passively transferred protective Abs in milk, induced via the gut-MG-sIgA axis, is the strategy used to protect suckling piglets from PEDV immediately after birth (4, 5). For example, in swine, high rates of protection against another porcine enteric alphacoronavirus, transmissible gastroenteritis computer virus (TGEV) in piglets is usually achieved when pregnant sows are orally infected with live virulent computer virus (5C7, 13C15). The increased rate of protection was associated with high titers of IgA Abs in colostrum and milk. This demonstrates that enteric viral contamination stimulates the intestinal mucosa influencing lactogenic immunity via the gut-MG-sIgA axis (4, 5). This model system can be used in the context of PEDV, as comparable maternal vaccination strategies are needed for initiation of the gut-MG-sIgA axis and piglet protection (4, 12, 16, 17). We showed previously in third trimester pregnant gilts that administering a higher dose of virulent PEDV increased computer virus neutralizing (VN) Ab titers in colostrum/milk and piglet protection compared with a lower dose (4). Despite this, field reports demonstrate incomplete and variable protection in orally PEDV-infected gestating swine (4). Furthermore, the optimal stage of gestation to initiate the gut-MG-sIgA axis by means of natural contamination or oral vaccination in na?ve pregnant swine to generate protective lactogenic immunity is unknown. Pregnancy modulates immunological processes that change over the course of gestation (18). For example, during the first trimester of gestation, levels of innate and proinflammatory factors increase, facilitating embryo implantation (19, 20). As pregnancy progresses, inflammatory cytokines decrease and regulatory cells and cytokines increase to support fetal growth and development to prevent rejection of the fetus (21C24). In.