The host range of SARS-CoV-2 and the susceptibility of animal species to the virus are topics of great interest to the international scientific community. The angiotensin I converting enzyme 2 (ACE2) protein is the major receptor for the virus, and sequence and structural analysis of the protein has been performed to determine its cross-species conservation. Based on these analyses, cattle have been implicated as a potential susceptible species to SARS-CoV-2 and have been reported to have increased ACE2 receptor distribution in the liver and kidney, and lower levels in the lungs. The goal of the current study was to determine the susceptibility of cattle to SARS-CoV-2 utilizing inoculation routes that facilitated exposure to tissues with increased ACE2 receptor distribution. For this, colostrum-deprived calves approximately 6 weeks of age were inoculated via the intratracheal or intravenous routes. Nasal and rectal swab samples, as well as blood and urine samples, were collected over the course of the study to evaluate viral shedding, viremia, and seroconversion. Pyrexia was used as the primary criteria for euthanasia and tissue samples were collected during necropsy. Importantly, SARS-CoV-2 RNA was detected in only two nasal swab samples collected on days 3 and 10 post-inoculation (pi) in two calves; one calf in the intratracheal group and the other calf in the intravenous group, respectively. Additionally, the calf in the intratracheal group that was positive on the nasal swab on day 3 pi also had a positive tracheobronchial lymph node on day 9 pi. Viral nucleic acid load on these samples, based on PCR cycle threshold values, were low and infectious virus was not recovered from the samples. These results suggest that there was no productive replication of SARS-CoV-2 in calves following intratracheal and intravenous inoculation.

Antigenic differences between bovine viral diarrhea virus (BVDV) vaccine strains and field isolates can lead to reduced vaccine efficacy. Historically, antigenic differences among BVDV strains were evaluated using techniques based on polyclonal and monoclonal antibody activity. The most common method for antigenic comparison among BVDV isolates is determination of virus neutralization titer (VNT). BVDV antigenic comparisons using VNT only account for the humoral component of the adaptive immune response, and not cell mediated immunity (CMI) giving an incomplete picture of protective responses. Currently, little data is available regarding potential antigenic differences between BVDV vaccine strains and field isolates as measured by CMI responses. The goal of the current paper is to evaluate two groups of cattle that differed in the frequency they were vaccinated, to determine if similar trends in CMI responses exist within each respective group when stimulated with antigenically different BVDV strains. Data from the current study demonstrated variability in the CMI response is associated with the viral strain used for stimulation. Variability in IFN-γ mRNA expression was most pronounced in the CD4⁺population, this was observed between the viruses within each respective BVDV subgenotype in the Group 1 calves. The increase in frequency of CD25⁺cells and IFN-γ mRNA expression in the CD8⁺and CD335⁺populations were not as variable between BVDV strains used for stimulation in the Group 1 calves. Additionally, an inverse relationship between VNT and IFN-γ mRNA expression was observed, as the lowest VNT and highest IFN-γ mRNA expression was observed and vice versa, the highest VNT and lowest IFN-γ mRNA expression was observed. A similar trend regardless of vaccination status was observed between the two groups of calves, as the BVDV-1b strain had lower IFN-γ mRNA expression. Collectively, data from the current study and previous data support, conferring protection against BVDV as a method for control of BVDV in cattle populations is still a complex issue and requires a multifactorial approach to understand factors associated with vaccine efficacy or conversely vaccine failure. Although, there does appear to be an antigenic component associated with CMI responses as well as with humoral responses as determined by VNT.

Theileria orientalis Ikeda genotype, a parasite causing a disease in cattle that leads to significant economic challenges in Asia, New Zealand, and Australia, has been identified in seven U.S. States since 2017. Two previously validated PCR tests for Theileria followed by DNA sequencing were performed to test blood samples collected from 219 cattle in Alabama, USA, during the period of 2022–2023. Bidirectional Sanger sequencing revealed that the MPSP gene sequences (639–660 bp) from two cattle in Lee and Mobile Counties of Alabama exhibited a 100% match with those of recognized T. orientalis Ikeda strains, and showed similarities ranging from 76% to 88% with ten other T. orientalis genotypes. A high copy number of T. orientalis Ikeda was detected in the blood of infected cattle (ALP-1: 1.7 × 10⁵ and 1.3 × 10⁶/mL whole blood, six months apart; ALP-2: 7.1 × 10⁶/mL whole blood). Although the confirmed competent vector for T. orientalis Ikeda, Haemaphysalis longicornis tick, has not yet been identified in Alabama, the persistent nature of T. orientalis Ikeda infection and the detection of a high pathogen burden in seemingly healthy cattle in this study suggest that other tick species, as well as shared needles and dehorning procedures, could facilitate pathogen transmission within the herd. Continued investigations are necessary for the surveillance of T. orientalis Ikeda and Haemaphysalis longicornis ticks in Alabama and other U.S. states, along with assessing the pathogenicity of T. orientalis Ikeda infections in cattle.

Bovine serum has been widely used as a universal supplement in culture media and other applications, including the manufacture of biological products and the production of synthetic meat. Currently, commercial bovine serum is tested for possible viral contaminants following regional guidelines. Regulatory agencies’ established tests focused on detecting selected animal origin viruses and are based on virus isolation, immunofluorescence, and hemadsorption assays. However, these tests may fail to detect new or emerging viruses in biological products. High-throughput sequencing is a powerful option since no prior knowledge of the viral targets is required. In the present study, we evaluate the virome of seven commercial batches of bovine serum from Mexico (one batch), New Zealand (two batches), and the United States (four batches) using a specific preparation and enrichment method for pooled samples and sequencing using an Illumina platform. A variety of circular replicase-encoding single-stranded (CRESS) DNA families (Genomoviridae, Circoviridae, and Smacoviridae) was identified. Additionally, CrAssphage, a recently discovered group of bacteriophage correlated with fecal contamination, was identified in 85% of the tested batches. Furthermore, sequences representing viral families with single-stranded DNA (Parvoviridae), double-stranded DNA (Polyomaviridae and Adenoviridae), single-stranded RNA (Flaviviridae, Picornaviridae, and Retroviridae), and double-stranded RNA (Reoviridae) were identified. These results support that high-throughput sequencing associated with viral enrichment is a robust tool and should be considered an additional layer of safety when testing pooled biologicals to detect viral contaminants overlooked by the current testing protocols.