T-cell responses to SARS-CoV-2 have been described in recovered patients, and may be important for immunity following infection and vaccination as well as for the development of an adoptive immunotherapy for the treatment of immunocompromised individuals. In this report, we demonstrate that SARS-CoV-2-specific T-cells can be expanded from convalescent donors, and recognize immunodominant viral epitopes in conserved regions of membrane, spike, and nucleocapsid. Following in vitro expansion using a GMP-compliant methodology (designed to allow the rapid translation of this novel SARS-CoV-2 T-cell therapy to the clinic), membrane, spike, and nucleocapsid peptides elicited IFN-γ production, in 27 (59%), 12 (26%), and 10 (22%) convalescent donors (respectively), as well as in 2 of 15 unexposed controls. We identified multiple polyfunctional CD4-restricted T-cell epitopes within a highly conserved region of membrane protein, which induced polyfunctional T cell responses, which may be critical for the development of effective vaccine and T cell therapies. Hence, our study shows that SARS-CoV-2 directed T-cell immunotherapy targeting structural proteins, most importantly membrane protein, should be feasible for the prevention or early treatment of SARS-CoV-2 infection in immunocompromised patients with blood disorders or after bone marrow transplantation to achieve anti-viral control while mitigating uncontrolled inflammation.

Research has shown that changes in nutrition both before and after weaning can affect mammary development. Additionally, estrogen is known to be a potent mammogenic stimulant. Our objectives were to determine effects of altered preweaning feeding and exogenous estradiol postweaning on growth, intake, and health. Thirty-six Holstein heifer calves were reared on (1) a restricted milk replacer (MR) diet fed at 0.44kg powder dry matter (DM)/day [R; 20.9% crude protein (CP), 19.8% fat, DM basis], or (2) an enhanced MR fed at 1.08kg powder DM/d (EH; 28.9% CP, 26.2% fat, DM basis). The MR feeding was reduced 50% during wk 8 to prepare for weaning. Starter was offered after wk 4 but balanced between treatments. Body weight and frame were measured weekly with intakes and health monitored daily. At weaning, a subset of calves were slaughtered (n=6/diet). Enhanced-fed calves had greater carcass, thymus, liver, spleen, and mammary gland (parenchyma and mammary fat pad) weights. The EH calves also had greater average daily gain (ADG) starting during wk 1 (0.36 vs. -0.06kg/d) and lasting through wk 7 (1.00 vs. 0.41kg/d). Remaining calves received estrogen implants or placebo and were slaughtered at the end of wk 10, creating 4 treatments: (1) R, (2) R + estrogen (R-E2), (3) EH, and (4) EH + estrogen (EH-E2). Postweaning ADG was similar between R, EH, and EH-E2 calves, but greater in R-E2 calves than E calves. The EH-E2 calves had the heaviest mammary glands, and R-E2 calves had heavier mammary glands than R calves. The EH calves consumed more MR DM, CP, and fat preweaning. The R-fed calves consumed more starter DM preweaning. Fecal score was greater for EH calves (1.74 vs. 1.50) preweaning, but days medicated did not differ. Fecal scores were lower for R-E2 calves postweaning. Improved preweaning feeding of calves increased body weights and frame measures. Differences in body weights remained postweaning. Enhanced-fed calves showed greater ADG during the preweaning period but not postweaning. Exogenous estrogen may elicit diet-dependent growth responses. Analysis of collected samples will allow determination of cellular and molecular processes responsible for the marked differences in mammary development observed.

Feeding heifers a higher plane of nutrition postweaning but before puberty can negatively affect mammary gland development and future milk yield. However, enhanced nutrition preweaning may promote development and future production. Our objectives were to determine the effects of enhanced feeding preweaning and exogenous estrogen immediately postweaning on mammary gland development and the composition of the mammary parenchyma (PAR) and mammary fat pad (MFP). Thirty-six Holstein heifer calves (<1 wk old) were reared on 1 of 2 dietary treatments for 8 wk: (1) a restricted milk replacer fed at 0.45 kg/d (R; 20% crude protein, 20% fat), or (2) an enhanced milk replacer fed at 1.13 kg/d (EH; 28% crude protein, 25% fat). Upon weaning, calves from each diet (n=6) were given either a placebo or estrogen implant for 2 wk, creating 4 treatments: R, R + estrogen (R-E2), EH, and EH + estrogen (EH-E2). Calves were housed individually with ad libitum access to water. Starter feeding began at wk 5 and was balanced between treatments. Udders were evaluated by palpation and physical measurements weekly. Subsets of calves were killed at weaning (n=6 per diet) and at the conclusion of the trial (n=6 per treatment). Udders were removed, dissected, and weighed. At wk 8, EH calves had longer front and rear teats. Providing estrogen to EH calves increased the length of rear teats during wk 9 and 10. Enhanced-fed calves had 5.2-fold more trimmed mammary gland mass than R calves. Providing estrogen to EH calves further increased mammary gland weight. Masses of PAR and MFP were markedly greater for EH calves than for R calves (e.g., 7.3-fold greater PAR tissue). Estrogen increased the mass of both PAR and MFP in EH calves. Feeding a higher plane of nutrition increased total protein, DNA, and fat in the MFP and total protein and DNA in the PAR. Dual-energy x-ray absorptiometry estimates of mammary fat mass were highly correlated with biochemical analyses of fat content. From histological study, we observed that the degree of expansion of epithelium into the adjacent stromal tissue and the complexity of ductal development were minimal in R, increased in EH, and increased by estrogen in both dietary treatments. Results provide compelling evidence that preweaning nutrition and estrogen administration immediately postweaning markedly increase mammary gland development in dairy calves. Cellular and molecular mechanisms responsible for these differences are currently under study.

Commercially available colostrum replacers (CR) are commonly used when maternal colostrum (MC) is unavailable, for managerial convenience, to ensure quality consistency at first feeding, or in disease control and eradication programs. The objective of this study was to determine the efficacy of feeding First Day Formula (Accelerated Genetics, Baraboo, WI) CR versus pooled MC on immunological status, growth, and health of preweaned dairy calves. A total of 1,220 Jersey and Jersey × Holstein calves born on a California Central Valley dairy farm were assigned after birth to receive either CR or MC following a systematic allocation procedure. Calves assigned to MC were tube fed 2.8 L of MC, and calves assigned to CR were tube fed a total of 500 g of CR (150 g of immunoglobulin G; IgG) mixed into 1.9 L of water at 1 h ± 5 min after the calf was born. A subset of calves was selected for passive transfer (n = 592) and growth (n = 268) analyses. Although both coliform count and total bacteria count were low for MC and CR fed to calves during the study, the predicted probability of calves receiving contaminated liquid feed (coliform count >10,000 cfu/mL) at first feeding was reduced for calves fed CR (1.5%) compared with calves fed MC (6.1%). The mean blood concentration of IgG was lower for calves fed CR than for calves fed MC (19.6 vs. 23.4 mg/mL). However, the apparent efficiency of absorption of IgG did not differ between treatments (34.4 and 35.9% for CR and MC, respectively). Total proteins were lower in calves fed CR compared with MC at 24 h (5.16 vs. 5.84 g/dL, respectively). Calves fed CR were 1.5 kg lighter at weaning and gained 0.03 kg less per day (0.30 vs. 0.33 kg/d, respectively) than calves fed MC before weaning. Height at weaning did not differ between the 2 treatment groups. Calves fed CR tended to have a higher predicted probability of not being treated for diarrhea than calves fed MC (0.142 vs. 0.110, respectively). However, when the disease was present, CR had a higher number of treatment days compared with MC (11.6 vs. 10.8 d, respectively). The hazard ratio of dying did not differ between MC and CR; however, CR calves had a numerically higher risk (hazard ratio = 1.347) of dying compared with calves that received MC. In conclusion, IgG absorption and serum concentration of calves were adequate when calves were fed either CR or MC. The CR-fed calves had a lower probability of receiving contaminated liquid feed and performed similar in terms of health compared with calves receiving high-quality MC, although they were slightly lighter at weaning. Therefore, the CR evaluated in this study is a valid alternative to high-quality (>50 mg of IgG/mL) MC.

Successful passive transfer of antibodies in neonatal calves can be achieved by feeding an adequate quantity and quality of maternal colostrum (MC) or colostrum replacer (CR). An alternative could be feeding lowquality maternal colostrum (LMC) with added IgG from a CR. The objective of this study was to determine if a commercial whey-based CR product containing low levels of casein (Premolac PLUS Bovine IgG; Zinpro Corporation, Eden Prairie, MN) fed to replace MC or supplement LMC could lead to adequate serum IgG levels and apparent efficiency of absorption (AEA) in neonatal dairy calves. Holstein calves (n = 20 per treatment) were separated from their dam after birth and randomly assigned to be fed 3.79 L of MC (106 g/L of IgG; 401 g of IgG fed), LMC (30 g/L IgG) supplemented with CR (41 g/L IgG; 154 g of IgG total fed; LMC-CR), or 1.3 L of 1 of 2 levels of CR (110 or 150 g of IgG fed; CR-110 or CR-150) within 1.5 h of birth. Colostrum was obtained from the first (MC) or second and third milkings (LMC) of cows from Pennsylvania State University dairy and pooled by source into large batches. Blood samples were taken from calves before colostrum feeding and 24 h after birth and were analyzed for serum total protein, total IgG, hematocrit, and Brix percentage. Calves fed MC had higher 24-h IgG values (means ± SEM) than calves fed LMC-CR (27.04 ± 1.07 vs. 22.33 ± 1.08 mg/mL, respectively). Feeding 150 g of IgG from CR led to higher 24-h serum IgG values than feeding 110 g of IgG (16.90 ± 1.09 vs. 12.79 ± 1.08 mg/mL). Serum IgG levels were different between the CR-fed calves and the calves fed LMC-CR and MC, but all had average values >10 mg/mL IgG. Calves fed LMC-CR had greater AEA than calves fed MC (54.58 ± 2.39 vs. 24.38 ± 2.36%, respectively). Among calves fed CR-110 or CR-150, AEA did not differ. Serum total protein and Brix percentage had strong correlations with actual IgG values across the entire study. We found no differences in average daily gain or health variables measured, and no differences in final hip width, withers height, or body weight for calves fed MC, LMC-CR, CR-150, or CR-110. These results indicate that CR can be fed successfully as an alternative to MC or as a supplement to colostrum with low IgG.