The etiological agents of the complex of South American arenaviral hemorrhagic fevers are a group of closely related New World mammarenaviruses (NWMs) including Junín, Machupo, Guanarito, Sabiá and Chapare. These viruses are maintained in nature through persistent infections of their respective host rodent species (Salazar-Bravo et al., 2002). Transmission of these agents to humans occurs through inhalation of aerosolized virus particles or contact with virus-containing rodent secreta or excreta (Childs et al., 1995). Junín virus (JUNV) is the most prevalent of pathogenic NWMs and is the etiological agent of Argentine hemorrhagic fever (AHF). AHF is an insidious disease characterized by severe hemorrhagic and/or neurologic manifestations with remittent fever, tremors, vascular leak and shock (de Bracco et al., 1978; Elsner et al., 1973; Enría et al., 1986; Enria et al., 2008). The pathogenesis of AHF is poorly understood; however, a salient feature of severe disease is an elevation in serum interferon-α (IFN-α) and other inflammatory mediators (Marta et al., 1999; Levis et al., 1984; Levis et al., 1985; Heller et al., 1992). The case-fatality rate of AHF can be as high as 30% in untreated individuals and the only countermeasures available for the prevention and treatment of severe JUNV infections are the Candid #1 vaccine and convalescent plasma (Enria et al., 2008). Neither has been approved for use outside of the area of endemicity and recent studies suggest that reversion to virulence by the vaccine virus can occur through a single nucleotide change in the envelope glycoprotein gene (York and Nunberg, 2018; Seregin et al., 2015; Albariño et al., 2011).

Animal models are essential to gaining insights into viral pathogenesis. Several species of non-human primates are susceptible to lethal JUNV disease (Kenyon et al., 1992; McKee et al., 1985; Avila et al., 1987). These models are considered the ‘gold standard’ because JUNV infection in these species more closely recapitulates the human disease. However, these models are costly and require specialized primate housing facilities within maximum biocontainment. Few small-animal models for JUNV exist because most standard laboratory rodent species, such as mice and hamsters, are refractory to severe disease. The most commonly used rodent model for JUNV is the guinea pig, which has served as the primary animal model to investigate pathogenesis and for screening promising therapeutic interventions (Yun et al., 2008; Gowen et al., 2013; Zeitlin et al., 2016). However, the use of this model for investigations into JUNV pathogenesis and countermeasure development has been mired by the lack of commercial reagents for this species.

A feature specific to the pathogenic NWMs is the use of human transferrin receptor 1 (hTfR1) for cellular entry (Radoshitzky et al., 2007; Helguera et al., 2012). In addition, different species known to be susceptible to disease following challenge with pathogenic NWMs express TfR1 orthologs that bind the viral envelope glycoprotein facilitating entry (Helguera et al., 2012; Hickerson et al., 2020). These findings suggest that the use of TfR1 for viral attachment and entry is an important determinant in defining whether a NWM can cause severe disease in a species other than the respective rodent reservoir host. Based on this, we investigated whether the expression of hTfR1 in laboratory mice would render them susceptible to lethal disease following JUNV challenge. Here, we demonstrate that transgenic hTfR1 mice develop a lethal disease course when exposed to the pathogenic Romero strain of JUNV and characterize the natural history and pathogenesis of disease. We also show that the type I IFN response plays a central role in the development of severe JUNV infection and disease in mice expressing hTfR1. The development of the hTfR1 mouse model of JUNV infection provides a novel system to investigate viral pathogenesis and assess promising therapeutics.

The first step in the viral infection process is the interaction between the virion and its host cell receptor. Pathogenic NWMs infect cells of their natural rodent reservoir via the endogenous TfR1 (Zong et al., 2014; Radoshitzky et al., 2008). These viruses have, in addition, adapted to utilize hTfR1 as the principal entry receptor in human cells (Radoshitzky et al., 2007). Related NWMs that enter through TfR1-independent mechanisms do not cause disease in humans (Sarute and Ross, 2017). Moreover, species known to be susceptible to disease when challenged with pathogenic NWMs (guinea pigs, marmosets, macaques) express TfR1 orthologs capable of mediating attachment and cellular entry (Helguera et al., 2012; Hickerson et al., 2020). By contrast, species that are refractory to pathogenic NWM infection (laboratory mice and hamsters) express TfR1 orthologs that do not support virus entry (Hickerson et al., 2020; Radoshitzky et al., 2008). Taken together, these findings suggest that the ability of NWMs to use TfR1 for entry is an important determinant in defining species vulnerability to severe disease (Choe et al., 2011). In the present study, we tested the hypothesis that mice genetically engineered to express hTfR1 would be susceptible to disease following exposure to JUNV. While hTfR1 HET mice were more susceptible to moderate JUNV disease than their WT counterparts, the development of uniformly lethal disease required the homozygous expression of hTfR1. The more severe disease phenotype was likely due to higher levels of hTfR1 in the liver and other tissues in the hTfR1 HOM mice compared to hTfR1 HET littermates (Yu et al., 2014). The susceptibility of hTfR1 HOM mice to lethal JUNV infection was age-dependent as the animals developed resistance as they matured. Several mechanisms may be involved in the development of resilience to JUNV challenge including a reduction in target receptor expression levels as mice age (Hofer et al., 2008; Domellöf et al., 2014; Lönnerdal and Kelleher, 2007; Sciot et al., 1990) and the maturation of the immune response (Coulombié et al., 1986; Blejer et al., 1987; Blejer et al., 1986; Bruyns et al., 1976; Doria et al., 1978; D'Eustachio and Edelman, 1975; Landreth, 2002). Additional studies are needed to investigate the age-dependent susceptibility of hTfR1 mice to JUNV and other New World hemorrhagic fever mammarenaviruses.

The type I IFN response plays an important role in protecting the host against viral pathogens; however, recent studies in mice characterizing the pathogenesis of acute infection by lymphocytic choriomeningitis virus (LCMV), a related mammarenavirus, have shown that the response can also contribute to lethal disease (Oldstone et al., 2018; Baccala et al., 2014). In the natural history and pathogenesis study characterizing JUNV infection and disease in hTfR1 HOM mice, we found that serum levels of IFN-α began to spike on day 8 p.i., just before the onset of clinical disease signs. Notably, patients suffering from AHF have been found to have increased concentrations of circulating IFN-α, which is believed to contribute to the clinical disease manifestations (Levis et al., 1984; Levis et al., 1985). The type I IFN response (IFN-α and IFN-β), which is a critical element of the innate immune response to throttle viral infections prior to the development of the adaptive immune response, can also have a damaging effect on the host when elevated systemic levels of type I IFN persist (Lee and Ashkar, 2018; Sen, 2001). Type I IFN signaling can impair immune system function by enhancing the expression of immunosuppressive molecules and inducing immune cell apoptosis (Stifter and Feng, 2015), which has been shown to affect the development of an effective adaptive immune response and contributes to the inability of the host to resolve persistent LCMV infections (Teijaro et al., 2013; Wilson et al., 2013). In lethal NZB and FVB/N mouse infection models based on intravenous challenge with 2 × 10⁶ plaque-forming units of the Clone 13 (Cl13) LCMV variant, type I IFN was found to underlie the severe disease phenotype (Oldstone et al., 2018; Baccala et al., 2014). Similarly, in hTfR1 HOM mice lacking IFN-α/βR or IFN-α/β and -γ receptors, or hTfR1 HOM mice treated with mAbs that block the IFN-α/βR, we demonstrate the essential role of the type I IFN response in driving JUNV pathogenesis. Deletion of the CD8 T cell response also prevents lethality associated with LCMV Cl13 infection in NZB and FVB/N mice (Oldstone et al., 2018; Schnell et al., 2012). The role of cytotoxic and helper T cells in JUNV pathogenesis in hTfR1 was not defined in the present study, but comparisons to the NZB and FVB/N models are complicated by differences in mouse age and genetics, as well as the virus challenge route and infectious doses administered. With respect to the latter, the outcome of infection (persistence vs. immune-mediated lethality) with LCMV Cl13 in C57BL/6 mice could be manipulated by modifying the virus challenge dose (Waggoner et al., 2012; Cornberg et al., 2013).

Mammarenaviruses are well known for their ability to cause persistent infections in rodents. Despite the lack of observable disease in the absence of IFN-α/β or IFN-α/β and -γ receptors, or following temporary mAb blockade of the IFN-α/βR, JUNV was able to persist in hTfR1 mice for up to 28 days in multiple tissues. It is likely that in the absence of type I IFN signaling JUNV infection would persist for an extended period (60 days), similar to that observed in Cl13 LCMV-infected mice protected from lethal disease through mAb treatment to block IFN-α/βR signaling (Oldstone et al., 2018). Also of note, JUNV persisted in multiple tissues of IFN-α/β and -γR-deficient mice expressing native mouse TfR1, while viral persistence in IFN-α/βR-deficient mice expressing mouse TfR1 was only evident in the brain. Consistent with the antiviral role of IFN-γ (Schroder et al., 2004), the dual type I and type II IFN receptor deficiency was more permissive to persistence regardless of whether or not hTfR1 was present. Further studies investigating long-term persistence and whether JUNV is shed in fecal matter and urine are needed to shed light on the potential virus-host carrier state that may result from infection in hTfR1 mice with compromised type I IFN responses.

In humans, mammarenaviral hemorrhagic fever is generally associated with mild pathological changes (Paessler and Walker, 2013). JUNV infection in hTfR1 mice resulted in limited histologic lesion development observed only during advanced stages of disease. Cell death of splenic mononuclear cells was present in mice at 10 and 12 days p.i. This lesion has been observed in other animal models of JUNV infection and natural infection in humans (Weissenbacher et al., 1975; Kenyon et al., 1985; González et al., 1980; Carballal et al., 1981). In humans, JUNV has been detected in phagocytic cells but not lymphocytes (González et al., 1980). In hTfR1 mice, the presence of JUNV antigen in the spleen was detected mainly in the white pulp suggesting a cytopathic effect on mononuclear cells. Similar findings have also been observed in the guinea pig model for JUNV (Yun et al., 2008), which suggests that targeting of mononuclear phagocytes may be central to pathogenesis. Death of the hTfR1 mice was tentatively attributed to neutrophilic encephalitis with JUNV antigen detected in neurons. Encephalitis has been reported in primates and humans infected with JUNV (González et al., 1983; Weissenbacher et al., 1979). Increased vascular permeability and prominent histopathology are features observed with LCMV Cl13 infection in NZB and FVB/N mice that succumb from respiratory failure and shock (Oldstone et al., 2018; Baccala et al., 2014; Schnell et al., 2012; Puglielli et al., 1999). Additional studies are needed to determine whether vascular leak associated with excess and prolonged proinflammatory cytokine concentrations, including IFN-α, may also be contributing to the demise of the JUNV-infected hTfR1 HOM mice.

The use of hTfR1 as the primary cellular entry receptor renders the pathogenic NWMs vulnerable to the development of countermeasures that interfere with the interaction between the viral envelope glycoprotein GP1 attachment subunit and the host cell receptor. The region of hTfR1 that mediates binding to the ectodomain of GP1 is the apical domain, which is not involved in binding to the known principal ligands, transferrin and hemochromatosis protein (Cheng et al., 2004; Lawrence et al., 1999; Montemiglio et al., 2019), thus representing a potential host-directed therapeutic target to broadly inhibit infection by JUNV and other New World hemorrhagic fever mammarenaviruses. Recent advances to exploit this vulnerability include the identification of a mAb (ch128.1) and an aptamer that bind to the apical domain of hTfR1 and the development of an immunoadhesin with the sequence of white-throated woodrat TfR1 apical domain fused to the Fc region of an IgG which binds to pathogenic NWM GP1 (Helguera et al., 2012; Maier et al., 2016; Cohen-Dvashi et al., 2020). Until now, proof-of-concept experiments to determine whether the ch128.1 strategy would prove to be an effective host-directed therapeutic to broadly treat NWM hemorrhagic fever were not possible due to the lack of an appropriate small-animal model. The novel hTfR1 mouse JUNV infection model that we have developed will be a valuable system in which this type of specific intervention can be investigated due to the requirement of hTfR1. Moreover, the new mouse model also provides an alternative to evaluate experimental therapies that directly target attachment by JUNV and related pathogenic NWMs and benefits from the large number of validated mouse reagents to assess immunological parameters and other physiological processes. By comparison, studies using the guinea pig JUNV infection model are limited by a shortage of reagents, animal costs and the requirement for substantially higher quantities of investigational drugs, which is often a limiting factor during early stages of antiviral drug discovery and development.

In summary, our findings identify hTfR1-mediated entry and the type I IFN response as key factors in the development of lethal JUNV disease in mice. The development of the first mouse model of JUNV infection in immunocompetent mice will be useful for investigating JUNV pathogenesis and early preclinical development of promising therapeutic interventions including approaches that disrupt the pathogenic NWM GP1-hTfR1 apical domain interaction. In addition, the identification of the type I IFN response as a key element in the development of severe JUNV disease in hTfR1 mice further supports investigation and development of immune-modulating agents as potential therapies to limit disease severity in cases of AHF and other NWM hemorrhagic fevers.

All data generated or analyzed during this study are included in the manuscript.

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Acceptance summary:

Current animal models of arenavirus pathogenesis are either not particularly robust or lack broad tractability. This study establishes a new mouse model of lethal Junin virus infection that recapitulates many key features of Junin virus pathogenesis in humans and may have broad utility for the development of vaccines and therapeutics targeting this important human pathogen.

Decision letter after peer review:

Thank you for submitting your article "Type I interferon underlies severe disease associated with Junin virus infection in mice" for consideration by eLife. Your article has been reviewed by three peer reviewers, one of whom is a member of our Board of Reviewing Editors, and the evaluation has been overseen by Karla Kirkegaard as the Senior Editor. The following individuals involved in review of your submission have agreed to reveal their identity: Juan Carlos de la Torre (Reviewer #2).

The reviewers have discussed the reviews with one another and the Reviewing Editor has drafted this decision to help you prepare a revised submission.

Summary:

The New World mammarenavirus Junin (JUNV) is the causative agent of Argentine hemorrhagic fever (AHF), a disease endemic to the Pampas region of Argentina that is associated with severe clinical symptoms including hemorrhagic and neurological manifestations and a case fatality rate of 15-30% in patients who do not receive appropriate supportive treatment. In addition, JUNV possesses features that make it a credible biodefense threat. Therefore, the importance of developing vaccines and therapeutics to counteract JUNV infections and associated disease. The availability of a mouse model of JUNV infection would facilitate the development of therapeutics and vaccines against AHF disease, as well as a better understanding of the molecular and cellular bases underlying JUNV pathogenesis.

Here, the authors use transgenic mice expressing TfnR, a Junin virus entry factor, to establish an immunocompetent mouse model of Junin virus infection that recapitulates several disease outcomes observed in humans. Lethal disease in infected mouse was associated with increased type I interferon responses, demonstrating a key pathological feature of human disease in these mice. The pathogenic effects could be mitigated in mice lacking IFN receptors, or in mice treated with IFN receptor blocking antibody.

Essential revisions:

1) The authors state that type I IFN underlies disease in this mouse model, but this is only demonstrated in young mice. Was viremia and serum IFN levels ever examined in older mice? Are these older mice just not replicating the virus, or is the virus replicating, but the mice are not making IFN? If the latter, would recombinant IFN treatment + Junin virus synergize in older mice to give a lethal phenotype?

2) Tissue viral loads shown in Figure 7D and 8D provide no information regarding the contribution of viral loads to pathogenesis, because they were measured at the late time period and because controls are missing. To be meaningful, viral loads should be measured at 11 or 12 dpi including control groups: hTfR1+/+ (7D) and "Placebo Tx" (8D) groups. Related to this issue, the authors stated "in the absence of a type I IFN response during the acute JUNV infection period hTfR1+/+ mice failed to clear the virus despite no apparent signs of illness". For this to be true, viral loads should be compared to those of control groups. In addition, 6 or 3-4 mice in Figure 7 and 8, respectively, may provide statistically significant survival data, but when population variance is huge like the viral loads in Figure 7D and 8D, 3-6 mice is not nearly sufficient. In order to use right sample size, power and sample size calculation should be redone based on the preliminary data in 7D and 8D.

3) Although hTfR1+/+ mice and IFNa/bR-/- and IFNa/bgR-/- mice are in different genetic background, they were backcrossed only twice. The authors' conclusion of type I IFN response being the key element for JUNV pathogenesis is partially based on the data obtained using these mice with mosaic genetic background (Figure 7). Combined with the data in Figure 8, this conclusion may still be valid, but nonetheless the authors should indicate contaminating 129 genes could have contributed to the susceptibility to viral infection and tissue viral loads.

[Editors' note: further revisions were suggested prior to acceptance, as described below.]

Thank you for resubmitting your article "Type I interferon underlies severe disease associated with Junín virus infection in mice" for consideration by eLife. Your article has been reviewed by two peer reviewers, and the evaluation has been overseen by a Reviewing Editor and Karla Kirkegaard as the Senior Editor.

The reviewers have discussed the reviews with one another and the Reviewing Editor has drafted this decision to help you prepare a revised submission.

We would like to draw your attention to changes in our revision policy that we have made in response to COVID-19 (https://elifesciences.org/articles/57162). Specifically, we are asking editors to accept without delay manuscripts, like yours, that they judge can stand as eLife papers without additional data, even if they feel that they would make the manuscript stronger. Thus the revisions requested below only address clarity and presentation.

Summary:

The responses provided by the authors are overall reasonable, and they have incorporated changes to the text of the revised version of the paper that address many, but not all, the criticisms raised during the review of the originally submitted paper.

Revisions:

In lieu of additional experiments to address outstanding questions, the reviewers request that the authors expand the Discussion along the following lines:

A general suggestion is for the authors to add some useful topic sentences to the Discussion and, most importantly, conclude with a discussion of any other viral infections for which type 1 interferons contribute to pathogenesis.

More specifically, the authors should provide a more elaborated comparison between the mouse model of lethal JUNV infection and the NZB and FVB mouse models of LCMV infection where type I interferon was shown to be a critical factor for fatal outcome of infection.

Essential revisions:

1) The authors state that type I IFN underlies disease in this mouse model, but this is only demonstrated in young mice. Was viremia and serum IFN levels ever examined in older mice? Are these older mice just not replicating the virus, or is the virus replicating, but the mice are not making IFN? If the latter, would recombinant IFN treatment + Junin virus synergize in older mice to give a lethal phenotype?

The questions posed by the reviewers are of great interest and will pursued in future studies. Because our focus was to develop and characterize a lethal disease model of JUNV infection in hTfR1 mice, we did not characterize the infection in older age groups of mice that were refractory to disease. With that said, the weight loss observed in the 5-week-old hTfR1 mice challenged with JUNV suggests that there was viral replication in the older mice, which would most likely be accompanied by type I IFN induction. Studies by Ross and coworkers (Lavanya et al., Sci Transl Med, 2013; Cuevas and Ross, J Virol, 2014) have shown that the attenuated JUNV Candid#1 vaccine strain can replicate in 4 and 8 to 10-week-old mice with spleen infectious virus titers present on days 8 and 7 p.i., respectively, suggesting that the more virulent JUNV Romero strain used in our studies would likely replicate to higher titers and for a longer period of time in older mice expressing hTfR1.

2) Tissue viral loads shown in Figure 7D and 8D provide no information regarding the contribution of viral loads to pathogenesis, because they were measured at the late time period and because controls are missing. To be meaningful, viral loads should be measured at 11 or 12 dpi including control groups: hTfR1+/+ (7D) and "Placebo Tx" (8D) groups.

Ideally, we would have liked to include additional animals in each group that would be euthanized just prior to when the control groups (hTfR1+/+ or Placebo Tx) would be expected to succumb for evaluation of serum and tissue viral loads. Due to our animal and mAb limitations, we were not able to add the additional viral load arms to the studies.

Although the D panels of Figures 7 and 8 do not provide information regarding the contributions of viral loads to the disease process, the data indicate that mice with compromised type I, or compromised type I and type II, IFN responses do not clear JUNV by the end of the 28-day study period. Further studies are needed to investigate long-term persistence which could provide insights into the virus-host reservoir relationship.

Related to this issue, the authors stated "in the absence of a type I IFN response during the acute JUNV infection period hTfR1+/+ mice failed to clear the virus despite no apparent signs of illness". For this to be true, viral loads should be compared to those of control groups.

This statement has been removed from the manuscript as part of the extensive revision. We now include the following statement. “Despite the lack of observable disease in the absence of IFN-α/β or IFN-α/β and γ receptors, or following temporary mAb blockade of the IFN-α/βR, JUNV was able to persist in hTfR1 mice for up to 28 days in multiple tissues.” (fourth to the last sentence of the second paragraph of the Discussion)

In addition, 6 or 3-4 mice in Figure 7 and 8, respectively, may provide statistically significant survival data, but when population variance is huge like the viral loads in Figure 7D and 8D, 3-6 mice is not nearly sufficient. In order to use right sample size, power and sample size calculation should be redone based on the preliminary data in 7D and 8D.

The purpose of the end-of-study sample collection was to qualitatively address the question of whether or not the surviving mice had cleared the JUNV infection. Future studies investigating longer-term persistence will be powered to determine statistical significance comparing titers from the different genotypes based on the findings reported in Panels 7D and 8D.

3) Although hTfR1+/+ mice and IFNa/bR-/- and IFNa/bgR-/- mice are in different genetic background, they were backcrossed only twice. The authors' conclusion of type I IFN response being the key element for JUNV pathogenesis is partially based on the data obtained using these mice with mosaic genetic background (Figure 7). Combined with the data in Figure 8, this conclusion may still be valid, but nonetheless the authors should indicate contaminating 129 genes could have contributed to the susceptibility to viral infection and tissue viral loads.

This is an excellent point. We have addressed this oversight by the addition of the following statement to the Results text. “Because of the hybrid background of the genotypic variants of mice used in the initial experiment to assess the role of the type I IFN response in JUNV pathogenesis, we could not rule out the contribution of genetic variation in susceptibility to JUNV infection, disease and persistence.” (first sentence of last paragraph of the Results section)

[Editors' note: further revisions were suggested prior to acceptance, as described below.]

Revisions:

In lieu of additional experiments to address outstanding questions, the reviewers request that the authors expand the Discussion along the following lines:

A general suggestion is for the authors to add some useful topic sentences to the Discussion and, most importantly, conclude with a discussion of any other viral infections for which type 1 interferons contribute to pathogenesis.

More specifically, the authors should provide a more elaborated comparison between the mouse model of lethal JUNV infection and the NZB and FVB mouse models of LCMV infection where type I interferon was shown to be a critical factor for fatal outcome of infection.

As suggested, we have added useful topic sentences to the Discussion.

Importantly, we have added text to paragraphs two, three and four of the Discussion, specifically elaborating on the comparison between the mouse JUNV infection model and the NZB and FVB mouse models of LCMV infection.

Author details

1. Brady T Hickerson

   Department of Animal, Dairy and Veterinary Sciences, Utah State University, Logan, United States

   Present address

   Division of Biotechnology Review and Research-III, Office of Biotechnology Products, Center for Drug Evaluation and Research, Food and Drug Administration, Silver Spring, United States

   Contribution

   Conceptualization, Data curation, Formal analysis, Funding acquisition, Investigation, Methodology, Writing - original draft, Writing - review and editing

   Competing interests

   No competing interests declared

2. Eric J Sefing

   Department of Animal, Dairy and Veterinary Sciences, Utah State University, Logan, United States

   Contribution

   Investigation

   Competing interests

   No competing interests declared

3. Kevin W Bailey

   Department of Animal, Dairy and Veterinary Sciences, Utah State University, Logan, United States

   Contribution

   Investigation

   Competing interests

   No competing interests declared

4. Arnaud J Van Wettere

   1. Department of Animal, Dairy and Veterinary Sciences, Utah State University, Logan, United States
   2. Utah Veterinary Diagnostic Laboratory, Utah State University, Logan, United States

   Contribution

   Formal analysis, Visualization, Methodology, Writing - review and editing

   Competing interests

   No competing interests declared

5. Manuel L Penichet

   1. Division of Surgical Oncology, Department of Surgery, David Geffen School of Medicine at University of California, Los Angeles (UCLA), Los Angeles, United States
   2. Department of Microbiology, Immunology and Molecular Genetics, David Geffen School of Medicine at UCLA, Los Angeles, United States
   3. UCLA Molecular Biology Institute, Los Angeles, United States
   4. UCLA Jonsson Comprehensive Cancer Center, Los Angeles, United States
   5. UCLA AIDS Institute, Los Angeles, United States

   Contribution

   Conceptualization, Resources, Formal analysis, Funding acquisition, Investigation, Methodology, Writing - review and editing

   Competing interests

   is a shareholder of Klyss Biotech, Inc. The Regents of the University of California licensed Dr. Penichet’s technology to this firm. The author has no other competing interests to declare.

6. Brian B Gowen

   Department of Animal, Dairy and Veterinary Sciences, Utah State University, Logan, United States

   Contribution

   Conceptualization, Data curation, Formal analysis, Supervision, Funding acquisition, Investigation, Methodology, Writing - original draft, Project administration, Writing - review and editing

   For correspondence

   brian.gowen@usu.edu

   Competing interests

   No competing interests declared

   "This ORCID iD identifies the author of this article:" 0000-0001-9113-2575

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