Juliette Hanson

Hepatitis E virus (HEV) is a major challenge for medical and scientific communities. Among the many issues complicating HEV disease are increasingly complicated extrahepatic manifestations [1]. The...

Background/Objectives: African swine fever virus (ASFV), the causative agent of African swine fever (ASF), is a highly contagious virus affecting both domestic and feral pig populations with mortality rates approaching 100% within one week of infection. Currently, there are limited treatments or vaccines available to control the disease. Although ASF is endemic in sub-Saharan Africa, the virus has also spread widely, reaching regions of the European Union, Russia, China, Southeast Asia, and, more recently, to the Dominican Republic and Haiti, bringing the threat closer to the United States (U.S.). ASF introduction to the U.S. would have severe consequences for swine producers and the national pork industry. Consequently, there is an urgent need to develop effective vaccine strategies to manage ongoing outbreaks abroad and mitigate the risk of future ASF incursions. Recent efforts have identified several ASFV epitopes and evaluated them in experimental vaccine trials. However, these vaccine candidates have elicited limited protective immune responses and have not demonstrated full protective efficacy. Methods: In this study, we employed in silico modeling and epitope prediction tools to design a synthetic multiepitope ASF protein incorporating key immunogenic regions of ASFV. The goal was to generate a single-antigen construct capable of inducing broad and robust immune responses when formulated with an established nanoparticle-based vaccine platform. The multiepitope ASF protein was subsequently expressed and entrapped into mannose-conjugated chitosan (M-CS) nanoparticles for vaccine formulation. The candidate vaccine, formulated with M-CS nanoparticle-entrapped adjuvant (ADU S100), was administered intramuscularly to pigs, and both T- and B-cell responses were assessed following the primary (DPV 22) and booster (DPV 42) doses. Results: Our M-CS ASF protein vaccine elicited antigen-specific T- and B-cell responses, both of which are recognized as central correlates of protection against ASFV. Conclusions: These promising preliminary immunological findings suggest that this nanoparticle vaccine has the potential to confer protection against ASFV challenge, a hypothesis that will be examined in future studies.

Subclinical mastitis reduces milk yield and elicits undesirable changes in milk composition, but the mechanisms resulting in reduced milk production in affected mammary glands are incompletely understood. This study investigated the effects of sterile inflammation on mammary gland metabolism by assessing changes in milk and venous blood composition. Mid-lactation primiparous Holstein cows (n = 4) had udder halves randomly allocated to treatments; quarters of 1 udder half were infused with 2 billion cfu of formalin fixed Staphylococcus aureus (FX-STAPH) and quarters of the opposite udder half infused with saline (SAL). Blood samples were collected from the right and left subcutaneous abdominal veins in 2.6 h intervals until 40 h post challenge and analyzed for blood gas and metabolite concentrations. Milk from FX-STAPH udder halves had significantly increased SCS by first milking at 8 h post-challenge. By 16 h post-challenge, FX-STAPH udder halves had increased concentrations of protein and lactate and lower lactose concentrations than SAL udder halves. Milk fat concentrations, milk yields, energy corrected milk yields, and the ferric reducing antioxidant power of milk were not significantly different between SAL and FX-STAPH udder halves. Venous blood of FX-STAPH halves had marginally greater concentrations of saturated O₂, partial pressures of O₂, and glucose concentrations than SAL halves. Conversely, total and partial pressures of CO₂ did not differ between udder half treatments suggesting a shift in local metabolite utilization in FX-STAPH udder halves. These results indicate that changes in milk composition resulting from mastitis are accompanied by changes in some key blood metabolite concentrations. The shift in venous blood metabolite concentrations, along with the marked increase in milk lactate, suggests that local mammary tissue and/or recruited and immune cells alters metabolite usage in mammary tissues. Future studies are needed to quantify the uptake of key milk precursors during mastitis.

Hepatitis E virus (HEV) is an expanding zoonotic viral disease threat. Although HEV causes acute viral hepatitis, it is increasingly being recognized as a systemic pathogen with detection and damage in extrahepatic tissues. The presence of HEV RNA in the semen of chronically infected human patients in the absence of viremia and fecal shedding and presence of HEV in the sperm head underscores the need to understand the interaction of HEV within the male reproduction system. Male accessory glands secrete biofluids necessary for sperm nourishment and to neutralize the acidity of the vagina. The role of male accessory glands in the dissemination and persistence of HEV infection have not been studied. Using an immunosuppressed pig model for chronic HEV infection, we demonstrate infectious HEV in mature sperm cells altering the sperm motility and morphology. HEV isolated from sperm cells remained infectious in human hepatoma cells. Spermatic fluid contained lower virus titers than the sperm cells from chronically infected pigs highlighting that the sperm cells themselves can associate with the virus. Evaluation of the male accessory glands demonstrated viral replication, infiltration of CD45 leukocytes, and apoptosis associated with HEV infection. A decrease in serum testosterone levels was evident in the HEV infected pigs. Even though a lower viral RNA titer was seen in serum and feces of chronically infected, immunosuppressed and ribavirin treated pigs, high viral RNA and infectious particles in sperm is a concern. Our findings necessitate further studies defining the mechanism of sperm cell invasion by HEV, length of HEV survival in sperm cells during chronic HEV infection, and risk of sexual transmission of HEV during both acute and chronic phases of infection. Author Summary Hepatitis E virus, a leading cause of acute viral hepatitis, causes both acute and chronic infection in humans. Recent advances within the HEV field have demonstrated extrahepatic diseases associated with HEV. More recent findings have revealed infectious HEV in the vagina, Sertoli cells, and ejaculate of humans, and sperm cells of pigs. We demonstrate that the male accessory sex glands may have a role in the persistence of HEV infection during chronic infections. We utilized an established immunosuppressed pig model and treated pigs with ribavirin to study the presence of virus in the sperm cells. We demonstrated high viral RNA loads and infectious particles associated with sperm cells. Our study further highlights the importance of the testis, as an immune privileged site, in the maintenance of chronic HEV infection. New studies to evaluate the mechanisms by which HEV associates with sperm cells, the length of HEV survival in sperm cell fractions, and consideration of the testes as a potential HEV reservoir are necessary.

Strains of Rocahepevirus ratti, an emerging hepatitis E virus (HEV), have recently been found to be infectious to humans. Rats are a primary reservoir of the virus; thus, it is referred to as “rat HEV”. Rats are often found on swine farms in close contact with pigs. Our goal was to determine whether swine may serve as a transmission host for zoonotic rat HEV by characterizing an infectious cDNA clone of a zoonotic rat HEV, strain LCK-3110, in vitro and in vivo. RNA transcripts of LCK-3110 were constructed and assessed for their replicative capacity in cell culture and in gnotobiotic pigs. Fecal suspension from rat HEV-positive gnotobiotic pigs was inoculated into conventional pigs co-housed with naïve pigs. Our results demonstrated that capped RNA transcripts of LCK-3110 rat HEV replicated in vitro and successfully infected conventional pigs that transmit the virus to co-housed animals. The infectious clone of rat HEV may afford an opportunity to study the genetic mechanisms of rat HEV cross-species infection and tissue tropism.

Rocahepevirus ratti, an emerging hepatitis E virus (HEV), has recently been found to be infectious to humans. Rats are a primary reservoir of the virus; thus, it is referred to as “rat HEV”. Rats are often found on swine farms in close contact with pigs. Our goal was to determine whether swine may serve as a transmission host for rat HEV by characterizing an infectious cDNA clone of a zoonotic rat HEV, strain LCK-3110, in vitro and in vivo. RNA transcripts of LCK-3110 were constructed and assessed for their replicative capacity in cell culture and in gnotobiotic pigs. Fecal suspension from rat HEV-positive gnotobiotic pigs was inoculated into conventional pigs cohoused with naïve pigs. Our results demonstrated that capped RNA transcripts of LCK-3110 rat HEV replicated in vitro and successfully infected conventional pigs that transmit the virus to cohoused animals. The infectious clone of rat HEV may afford an opportunity to study the genetic mechanisms of rat HEV cross-species infection and tissue tropism. Significance Statement: New zoonotic strains of Rocahepevirus ratti (rat HEV) have emerged infecting both immunocompetent and immunosuppressed people through unknown transmission sources. Pigs are a primary source of transmission for human HEV strains and could be serving a similar role for rat HEV transmission as rats are a common pest found on swine farms worldwide. Rats could be transmitting rat HEV to pigs which could then be transmitted to humans. Determining susceptibility of pigs to emerging zoonotic rat HEV strains can define potentially new transmission routes to inform public health policy and could provide pathology models for rat HEV disease.

After parturition, dairy cows mobilize AA from skeletal muscle to meet metabolizable protein (MP) requirements. High mobilization may compromise cow health and longer-term milk production. Postpartum diets with higher MP concentrations, improved AA profiles, or MP increased at the expense of forages rather than nonforage fiber sources may attenuate muscle catabolism; however, the molecular mechanisms responsible need investigation. We evaluated mRNA expression in the longissimus dorsi of cows fed postpartum diets differing in MP concentration, AA profile, and fiber source. From 0 to 25 d after parturition, 40 multiparous cows received the following diets: (1) 13% deficient in MP (D-MP), (2) adequate in MP using primarily soy protein (A-MP), (3) adequate in MP using blends of proteins and individual AA to improve the AA profile (Blend), or (4) similar to Blend except additional protein replaced forage (Blend-fNDF). Biopsies were taken approximately -5, 7, and 25 d relative to parturition. Greater dietary MP concentration (D-MP vs. A-MP and Blend) decreased expression of genes related to protein synthesis (MTOR, RPS6KB1) and degradation (FOXO1), inflammation (IFNG, TLR4), and endoplasmic reticulum (ER) stress (HSPA5, DDIT) and increased genes associated with lipogenesis (PPARG) and glucose oxidation (LDH, MB). In Blend versus A-MP (i.e., effect of AA profile), expression related to apoptosis (CASP8) and inflammation (TNFA) decreased and genes associated with cell cycle progression (E2F1) and fast-twitch glycolytic muscle fiber type (MYH4) increased. Less forage (Blend-fNDF vs. Blend) decreased genes associated with lipogenesis (PPARG, ACACA) and ER stress (BCL2, DDIT3, EIF2AK3, PPP1R15A) and increased genes associated with inflammation (TNF), inhibition of myogenesis (MSTN), and autophagy (PEBP1). In summary and based on mRNA expression, increasing MP supply may attenuate muscle turnover and ER stress. However, an unbalanced AA supply reduced cell cycle progression and protein synthesis. Lower energy supplies may reduce cell growth and cause autophagy.

Mammary blood flow is central to mammary growth, development, and productivity, but the development of the vasculature network is poorly understood. The objective of this study was to determine how the vascular system adapts to mammary growth by inducing different levels of mammary growth and examining 2 regions of mammary parenchymal tissue. Holstein heifer calves (n = 12) received daily injections on the days immediately preceding euthanasia at 82 d of age. Treatments were control (CON), short-term estradiol (STE), and long-term estradiol (LTE). The CON calves received corn oil injections, the STE calves received 9 injections of corn oil followed by 3 injections of estradiol, and the LTE calves received 12 estradiol injections. Mammary tissues were collected from the center and edge parenchymal regions of all right rear mammary glands to quantify the tissue area of various tissue structures, the percentage of proliferating epithelial cells, and the number and form of blood vessels. Results showed that LTE calves had a greater tissue area occupied by epithelium than CON and STE calves, and the epithelial area in CON and STE calves was similar. Edge parenchyma had a greater percentage of proliferating epithelial cells than center parenchyma across all treatment groups. In the edge region, LTE calves had the greatest percentage of proliferating epithelial cells, coinciding with greater epithelial area. The number of blood vessels per unit of tissue area was greater in center than in edge parenchyma; the corresponding vessel surface area per unit of tissue area followed the same pattern. Mammary blood vessel measures were not markedly influenced by estradiol treatment. These results highlight the marked difference in the number and organization of blood vessels in different mammary parenchyma regions but indicate that the effects of estradiol on stimulating mammary epithelial proliferation does not directly translate to increasing numbers of blood vessels.

Experimental efforts to measure neutrinos by radio-frequency (RF) signals resulting from neutrino interactions in-ice have intensified over the last decade. Recent calculations indicate that one may dramatically improve the sensitivity of ultra-high energy ("UHE"; >EeV) neutrino experiments via detection of radio waves trapped along the air-ice surface. Detectors designed to observe the "Askaryan effect" currently search for RF electromagnetic pulses propagating through bulk ice, and could therefore gain sensitivity if signals are confined to the ice-air boundary. To test the feasibilty of this scenario, measurements of the complex radio-frequency properties of several air-dielectric interfaces were performed for a variety of materials. Two-dimensional surfaces of granulated fused silica (sand), both in the lab as well as occurring naturally, water doped with varying concentrations of salt, natural rock salt formations, granulated salt and ice itself were studied, both in North America and also Antarctica. In no experiment do we observe unambiguous surface wave propagation, as would be evidenced by signals travelling with reduced signal loss and/or superluminal velocities, compared to conventional EM wave propagation.