Abigail L. Smith

A SEROLOGIC SURVEY FOR VIRUSES AND MYCOPLASMA PULMONIS AMONG WILD HOUSE MICE (MUS DOMESTICUS) IN SOUTHEASTERN AUSTRALIA

Plasma samples from 267 wild house mice (Mus domesticus) trapped at 14 sites in southeastern Australia were screened for antibody to 14 viruses normally associated with laboratory-reared rodents and to Mycoplasma pulmonis. Serologic prevalence was high for murine cytomegalovirus (99%, n = 94), murine coronavirus (95%), and murine rotavirus (74%). Samples from mice collected at all sites contained antibody to these viruses. The serologic prevalence was lower for mouse adenovirus, strain K87 (37%), parvovirus (33%), and reovirus type 3 (28%), with substantial site-to-site variation. Plasma from mice collected at 12 sites contained mouse adenovirus or reovirus antibody, and samples from mice at eight sites contained parvovirus antibody. Parvovirus-antibody positive mice were typically from high density populations or from low density populations that had recently declined from high density. Antibody to lymphocytic choriomeningitis virus (LCMV) and Sendai virus occurred at only three sites, and the serologic prevalence was very low (9.6% and 1.8%, respectively). All of the LCMV-positive mice were from northeastern New South Wales. The presence of this zoonotic virus in a mouse plague-prone region raises questions about human health risks resulting from cohabitation with large numbers of mice. It appeared that mouse populations at high density or declining from high density had higher prevalence of viral antibody than populations that had been at low or moderate density for some time. Thus, viral epizootics may occur among high-density populations and may be responsible for or precipitate declines in mouse density. These data raise the possibility of rodent viruses having potential as biological control agents.

Mouse parvovirus infection potentiates allogeneic skin graft rejection and induces syngeneic graft rejection

BACKGROUND The recently identified autonomous mouse parvovirus designated mouse parvovirus-1 (MPV-1) persists in adult BALB/c mice for at least 9 weeks, infects lymphoid tissues, interferes with the ability of cloned T cells to proliferate, and exhibits immunomodulatory properties. As a consequence of these findings, the present studies were undertaken to characterize further the inmunomodulatory effects of MPV-1 on T cell-mediated immune responses in vivo and in vitro. METHODS To evaluate the effect of MPV-1 infection on CD8+ T cell-mediated responses, BALB/c-H2dm2 mice were infected after transplantation of allogeneic BALB/c skin. RESULTS MPV-1 potentiated the rejection of allogeneic skin grafts. This potentiation was not a result of virus infecting the cellular or vascular component of the graft as determined by in situ hybridization, but was mediated by T cells. However, the proliferative capacity of alloantigen-reactive lymphocytes from graft-sensitized infected mice was diminished. MPV-1 also induced the rejection of syngeneic skin grafts, and T cells from these infected graft-sensitized mice lysed syngeneic P815 target cells. CONCLUSIONS These results suggest that MPV-1 infection of skin-grafted mice may disrupt normal mechanisms of peripheral tolerance and provide a unique model to study virus-induced autoimmunity.

MURINE VIRUSES IN AN ISLAND POPULATION OF INTRODUCED HOUSE MICE AND ENDEMIC SHORT-TAILED MICE IN WESTERN AUSTRALIA

House mice (Mus domesticus) were recently introduced to Thevenard Island, off the northwest coast of Western Australia. This island is also habitat for an endangered native rodent, the short-tailed mouse (Leggadina lakedownensis). Concerns have been raised that house mice may pose a threat to L. lakedownensis both through competition and as a source of infection. To assess the threat to L. lakedownensis posed by viral pathogens from M. domesticus, a serological survey was conducted from 1994 to 1996 of both species for evidence of infection with 14 common murine viruses (mouse hepatitis virus, murine cytomegalovirus, lymphocytic choriomeningitis virus, ectromelia virus, mouse adenovirus strains FL and K87, minute virus of mice, mouse parvovirus, reovirus type 3, Sendai virus, Theilers mouse encephalomyelitis virus, polyoma virus, pneumonia virus of mice, and encephalomyocarditis virus) and Mycoplasma pulmonis. Despite previous evidence that populations of free-living M. domesticus from various locations on the Australian mainland were infected with up to eight viruses, M. domesticus on Thevenard Island were seropositive only to murine cytomegalovirus (MCMV). Antibodies to MCMV were detected in this species at all times of sampling, although seroprevalence varied. Infectious MCMV could be isolated in culture of salivary gland homogenates from seropositive mice. In contrast, L. lakedownensis on Thevenard Island showed no serological evidence of infection with MCMV, any of the other murine viruses, or M. pulmonis, and no virus could be isolated in culture from salivary gland homogenates. Although MCMV replicated to high titers in experimentally infected inbred BALB/c laboratory mice as expected, it did not replicate in the target organs of experimentally inoculated L. lakedownensis, indicating that the strict host specificity of MCMV may prevent its infection of L. lakedownensis. These results suggest that native mice on Thevenard Island are not at risk of MCMV infection from introduced house mice, and raise interesting questions about the possible selective survival of MCMV in small isolated populations of M. domesticus.

Modulation of lethal and persistent rat parvovirus infection by antibody

SummaryTwo day-old athymic (rnu/rnu) and euthymic (rnu/+) rat pups nursing immune or non-immune dams were inoculated oronasally with the Yale strain of rat virus (RV-Y). All athymic and euthymic pups (57/57) from immune dams remained clinically normal, whereas 51 of 66 athymic and euthymic pups from non-immune dams died within 30 days. Infectious RV was detected by explant culture in 12 of 15 surviving pups of both genotypes from non-immune dams 30 days after inoculation, but in none of the 57 surviving pups from immune dams. RV-Y DNA was detected by Southern blotting in kidneys of surviving athymic pups from non-immune dams but was not detected in pups from immune dams. Euthymic pups from immune dams appeared not to produce endogenous antibody to RV after virus challenge, whereas euthymic pups from non-immune dams produced high-titered RV immune serum. Pups of both genotypes given immune serum prior to or with RV were fully protected from disease and persistent infection, whereas pups given immune serum 24 hours after RV were partially protected. These studies show that RV antibody offers significant protection against lethal and persistent RV infection.

Microbiological Quality Control for Laboratory Rodents and Lagomorphs

Mice (Mus musculus), rats (Rattus norvegicus), other rodent species, and domestic rabbits (Oryctolagus cuniculus) have been used in research for over 100 years. During the first half of the 20th century, microbiological quality control of lab animals was at best rudimentary as colonies were conventionally housed and little or no diagnostic testing was done. Hence, animal studies were often curtailed and confounded by infectious disease (Mobraaten and Sharp, 1999; Morse, 2007; Weisbroth, 1999). By the 1950s, it became apparent to veterinarians in the nascent field of comparative medicine that disease-free animals suitable for research could not be produced by standard veterinary disease control measures (e.g., improved sanitation and nutrition, antimicrobial treatments) in conventional facilities. Henry Foster, the veterinarian who founded Charles River Breeding Laboratories in 1948 and a pioneer in the large-scale production of laboratory rodents, stated in a seminar presented at the 30th anniversary of AALAS, “After a variety of frustrating health-related problems, it was decided that a major change in the company’s philosophy was required and an entirely different approach was essential”. Consequently, he and others developed innovative biosecurity systems to eliminate and exclude pathogens (Allen, 1999). In 1958, Foster reported on the Cesarean-originated barrier-sustained (COBS) process for the large-scale production of specific pathogen-free (SPF) laboratory rodents (Foster, 1958). To eliminate horizontally transmitted pathogens, a hysterectomy was performed on a near-term dam from a contaminated or conventionally housed colony. The gravid uterus was pulled through a disinfectant solution into a sterile flexible film isolator where the pups were removed from the uterus and suckled on axenic (i.e., germ-free) foster dams. After being mated to expand their number and associated with a cocktail of nonpathogenic bacteria to normalize their physiology and prime their immune system, rederived rodents were transferred to so-called barrier rooms for large-scale production. The room-level barrier to adventitious infection entailed disinfection of the room, equipment, and supplies, limiting access to trained and properly gowned personnel, and the application of new technologies such as high-efficiency particulate air-filtration of incoming air (Dubos and Schaedler, 1960; Foster, 1980; Schaedler and Orcutt, 1983; Trexler and Orcutt, 1999). The axenic and associated rodents mentioned in the COBS process are collectively classified as gnotobiotic to indicate that they have a completely known microflora. By contrast, barrier-reared rodent colonies are not gnotobiotic because they are housed in uncovered cages and thus acquire a complex microflora from the environment, supplies, personnel, and other sources. Instead, they are described as SPF to indicate that according to laboratory testing, they are free from infection with a defined list of infectious agents, commonly known as an ‘exclusion’ list.

Lymphocytic Choriomeningitis Virus

Publisher Summary Lymphocytic choriomeningitis virus (LCMV) is an important virus of the laboratory mouse from a number of perspectives. Foremost is its well-documented zoonotic potential for humans. Mus musculus and its various aboriginal and commensal species or subspecies represent the natural reservoir hosts for LCMV, with an intimate host-virus relationship. This relationship may involve subclinical persistent infection that is associated with minimal or undetectable levels of circulating antibody; thus, detection of infection can be a challenge. LCMV infects a wide variety of tissues, and infection of mice can have protean effects upon normal physiology and immune response, with deleterious impact upon research. The polytropism of LCMV and its noncytolytic course of infection contribute to a well-deserved reputation as a cryptic contaminant of tumors and cell lines. This chapter emphasizes the biology of LCMV as a naturally occurring infection of laboratory mice and the practical consequences of infection. Experimental studies provide insight into understanding the biology of natural infection and are therefore reviewed.

Management of Rodent Viral Disease Outbreaks: One Institution's (R)Evolution

Abstract At first blush, an outbreak of mouse hepatitis virus or epizootic diarrhea of infant mice virus in a research colony of laboratory mice may not seem like a “disaster.” However, irrespective of magnitude, such an outbreak at an academic institution is disruptive for researchers at all levels. It can be a disaster for the graduate student who may have just a few experiments to finish before writing the thesis or for the postdoctoral fellow who is in the lab for only 1 or 2 years. Infectious disease outbreaks also limit the ability of principal investigators to share their animals with collaborators at their home institution as well as with those at extramural sites, thereby thwarting the expectation that research materials supported by federal funds will be made readily available to colleagues. This article traces the evolution of a change in “culture” at a large, well-funded academic institution with over 1,800 active IACUC protocols, more than 1,000 of which include mice. During a period of less than 5 years, the institution evolved from virtual paralysis in the face of such outbreaks to the implementation of policies and practices that enable effective outbreak management and the timely resumption of research functionality. This evolution required not only support from the highest levels of leadership in the university and its school of medicine but also a huge outlay of financial resources.

Oral ivermectin as an unexpected initiator of CreT2-mediated deletion in T cells.

197 Our breeding strategy segregates breeders with loxP-flanked alleles from those transgenic for Cre; specifically, we intercrossed R26RYFP/YFP with mice heterozygous for the transgene encoding UBC-CreT2 to generate R26RYFP/+UBC-CreT2+ offspring (Supplementary Methods). After initiating the administration of feed containing ivermectin (12 p.p.m.), we unexpectedly found that approximately 60% of the R26RYFPUBC-CreT2+ offspring expressed YFP in peripheral blood cells (Fig. 1a). Deletion was not confined to the Rosa26 locus; many additional loxP-flanked loci present in our colony underwent recombination in UBC-CreT2+ mice treated with ivermectin (data not shown). Breeding cages continued to generate offspring with YFP+ cells more than 15 weeks after the cessation of ivermectin treatment. Notably, adult R26RYFP/+UBC-CreT2+ mice weaned before the initiation of treatment responded very differently and remained YFP– despite direct ingestion of ivermectin-containing feed (data not shown). Neither the R26RYFP knock-in allele nor the transgene encoding UBC-CreT2 alone was sufficient for YFP expression induced by ivermectin (data not shown). To determine if CreT2-mediated deletion induced by ivermectin occurred in all tissues or only a subset of tissues, we evaluated YFP expression in histology sections of thymuses, spleens and kidneys from R26RYFP/+UBC-CreT2+, R26RYFP/+UBC-CreT2– and tamoxifentreated mice. Oral treatment with tamoxifen resulted in YFP expression regardless of cell or tissue type (data not shown). In contrast, ivermectin treatment resulted in limited distribution of YFP expression, with YFP detected in spleen and thymus but not kidney tissue in ivermectin-treated To the Editor: Ivermectin is a broad-spectrum antihelmintic drug used to treat pinworms and fur mites in laboratory mouse colonies. Here we report activation of a tamoxifen-regulated Cre recombinase fusion protein as an unintended but immunologically relevant side effect of oral treatment with ivermectin. Ectoparasites and their treatment represent a growing dilemma in laboratory mouse colonies1,2. Topical or oral ivermectin is an effective treatment for fur mites and pinworms because ivermectin selectively binds to the glutamate-gated chloride ion channels in muscle and nerve cells, leading to hyperpolarization of the cells, followed by paralysis and death of the parasite3. Ivermectin is generally considered to be safe for rodent colonies but may alter immune responses4 and has neurotoxicity in susceptible strains5. During a facility-wide treatment for mite infestation, we noted unexpected deletion of the loxP-flanked ‘stop’ cassette of a reporter for the expression of Cre recombinase from the ubiquitously expressed Rosa26 promoter with knock-in of sequence encoding yellow fluorescent protein (R26RYFP)6 in two independent colonies of mice with transgenic tamoxifen-regulated expression of Cre (UBC-CreT2)7. Normally the CreT2 fusion protein, transcriptionally regulated here by the ubiquitin promoter, is inactive unless bound specifically to 4-hydroxytamoxifen or estrogen antagonist ICI 182780 (ref. 8). In over 5 years of experience with the UBCCreT2 and R26RYFP strains, we have not noted CreT2-mediated deletion in hematopoietic lineages without tamoxifen exposure9 (data not shown). Oral ivermectin as an unexpected initiator of CreT2mediated deletion in T cells

Reducing exposure to laboratory animal allergens

Laboratory animal allergy is a serious health problem. We examined several possible allergen-reducing strategies that might be effective in the working mouse room. Ambient allergen concentrations were measured when mice were maintained under several conditions: conventional housing versus ventilated cage racks operated under negative or positive pressure. We found that housing mice in ventilated cages operated under negative pressure and using ventilated changing tables reduced ambient mouse allergen (Mus m 1) concentrations tenfold, compared with values when mice were housed in conventional caging and using a conventional (non-ventilated) changing table. Housing mice in positively pressurized cages versus conventional cages did not reduce ambient allergen values. Cleaning mouse rooms at an accelerated frequency also did not reduce ambient Mus m 1 concentration. We also quantified ambient allergen values in several areas of The Jackson Laboratory. A facility-wide survey of Mus m 1 concentrations indicated that allergen concentrations were undetectable in control areas, but ranged from a mean (+/- SEM) 0.11 +/- 0.02 ng/m3 to 5.40 +/- 0.30 ng/m3 in mouse rooms with different cage types. The percentage of animal caretakers reporting allergy symptoms correlated significantly with ambient allergen concentrations: 12.9% reported symptoms in the rooms with the lowest allergen concentration (0.14 +/- 0.02 ng/m3), but 45.9% reported symptoms in rooms with the highest concentration (2.3 +/- 0.4 ng/m3). These data indicate that existing technology can significantly reduce exposure to laboratory animal allergens and improve the health of animal caretakers.