The rising incidence of serious fungal diseases represents a growing threat to human health. Cryptococcus neoformans, an encapsulated yeast saprophyte with global distribution, has been recognized as an important emerging pathogen. Humans frequently develop asymptomatic or mild infection with C. neoformans, but individuals with impaired host defense systems may develop severe pneumonia and potentially fatal meningoencephalitis. Insight into the biology and virulence of C. neoformans is advancing rapidly and will be propelled even further by the recently completed and published genome sequences for two related strains of C. neoformans serotype D. Several mammalian model hosts including the guinea pig, rabbit, rat, and mouse have been developed for the study of cryptococcosis. The combination of microbial genomics with well-characterized model hosts that are amenable to immunologic and genetic manipulation represents a powerful resource for comprehensive study of cryptococcal disease pathogenesis as well as vaccine and antifungal drug therapy. This review provides an introduction to each mammalian model host and briefly highlights the advantages, limitations, and potential of each system for future research involving cryptococci.

The Centers for Disease Control and Prevention Category A infectious agents include Bacillus anthracis (anthrax), Clostridium botulinum toxin (botulism), Yersinia pestis (plague), variola major virus (smallpox), Francisella tularensis (tularemia), and the filoviruses and arenaviruses that induce viral hemorrhagic fevers. These agents are regarded as having the greatest potential for adverse impact on public health and therefore are a focus of renewed attention in infectious disease research. Frequently rodent models are used to study the pathobiology of these agents. Although much is known regarding naturally occurring infections in humans, less is documented on the sources of exposures and potential risks of infection to researchers and animal care personnel after the administration of these hazardous substances to laboratory animals. Failure to appropriately manage the animals can result both in the creation of workplace hazards if human exposures occur and in disruption of the research if unintended animal exposures occur. Here we review representative Category A agents, with a focus on comparing the biologic effects in naturally infected humans and rodent models and on considerations specific to the management of infected rodent subjects. The information reviewed for each agent has been curated manually and stored in a unique Internet-based database system called HazARD (Hazards in Animal Research Database, http://helab.bioinformatics.med.umich.edu/hazard/) that is designed to assist researchers, administrators, safety officials, Institutional Biosafety Committees, and veterinary personnel seeking information on the management of risks associated with animal studies involving hazardous substances.

Human immunodeficiency virus (HIV), the causative agent for acquired immune deficiency syndrome, was described over 25 y ago. Since that time, much progress has been made in characterizing the pathogenesis, etiology, transmission, and disease syndromes resulting from this devastating pathogen. However, despite decades of study by many investigators, basic questions about HIV biology still remain, and an effective prophylactic vaccine has not been developed. This review provides an overview of the viruses related to HIV that have been used in experimental animal models to improve our knowledge of lentiviral disease. Viruses discussed are grouped as causing (1) nonlentiviral immunodeficiency-inducing diseases, (2) naturally occurring pathogenic infections, (3) experimentally induced lentiviral infections, and (4) nonpathogenic lentiviral infections. Each of these model types has provided unique contributions to our understanding of HIV disease; further, a comparative overview of these models both reinforces the unique attributes of each agent and provides a basis for describing elements of lentiviral disease that are similar across mammalian species.

Epstein-Barr virus (EBV) is a ubiquitous human gammaherpesvirus (GHV) that causes acute infection and establishes life-long latency. EBV is associated with the development of B-cell lymphoproliferative disorders, several malignant cancers, the syndrome of infectious mononucleosis, and chronic interstitial lung disease. Although the molecular biology of EBV has been characterized extensively, the associated disease conditions and their pathogenesis are difficult to study in human populations because of variation in human environments and genetics, the well-documented effect of stressors on pathogenesis, and the chronic and latent properties of the virus. GHV are highly species-specific, and suitable animal models for EBV are not available. However, in 1980, a murine gammaherpesvirus (MuGHV, also known as MHV68 and γHV68) was identified as a natural pathogen of bank voles and wood mice. Experimental MuGHV infections in laboratory mice share many features of EBV infections in humans, including facets of the clinical human syndrome known as infectious mononucleosis. These features make MuGHV a valuable experimental model for studying the pathophysiology of a GHV in a natural host.

We used primary and nested polymerase chain reaction (PCR) assays to determine the presence of mouse parvovirus (MPV) in mouse sperm, oocytes, preimplantation embryos, and ovarian tissues collected from MPV-infected mice. The primary PCR assay detected MPV in 56% of the sperm samples. MPV was not eliminated by passing sperm samples through a Percoll gradient. After Percoll treatment, MPV was still present in 50% of the samples according to primary PCR assay. Oocyte samples that did not undergo extensive washing procedures had detectable MPV in 7% of the samples based on the primary PCR assay, but nested PCR assay detected higher (28%) infection rate. However, MPV was not detected in oocytes that underwent extensive washing procedures, as assessed by either primary or nested PCR assay. Although primary PCR did not detect MPV in embryos, a nested PCR assay determined that 50% of the embryos were positive for the virus. In addition, ovarian tissues were collected from 3 different mouse colonies with enzootic MPV infection. Ovarian tissue collected from 129CT, 101/R1, and Sencar mice had high incidence (38%, 63%, and 65%, respectively) of MPV infection on the basis of nested PCR amplification. These results demonstrate that mouse gametes, embryos, and ovarian tissues may be contaminated with MPV and therefore caution is necessary when infected germplasm is used for assisted reproductive technologies such as embryo transfer, establishing embryonic stem cell lines, in vitro fertilization, ovary transplantation, and intracytoplasmic sperm injection.

Comparison of experimental groups by microscopic examination is a common and useful method for evaluating animal models of disease. Quantification of lesions is challenging, however, and differences in scoring systems hinder comparison of results from different laboratories. The purpose of this study was to validate a simple and reproducible scoring system for Helicobacter pylori-associated gastric disease in mice. The system is based on quantification of the percentage of microscopic fields in which lesions are present, rather than on subjective estimates of lesion severity. Linear regression analyses revealed good agreement between investigators in scoring of all 3 histologic criteria examined. The range of correlation coefficients between individual readers' scores and mean scores for the 3 histologic criteria examined were: neutrophilic inflammation, 0.845 to 0.935; gastritis sufficient to displace glands, 0.919 to 0.943; and epithelial metaplasia, 0.650 to 0.799. Comparison of scores in different experimental groups by analysis of variance and Fisher least significant difference tests revealed significant differences between infected and uninfected groups and between immunodeficient and immunocompetent groups. We propose that this system may be useful in standardizing the morphologic evaluation of rodent models of H. pylori and that a similar system could be devised for evaluation of other animal models of enteric disease.

Fecal shedding and transmission of mouse parvovirus 1 (MPV) to naïve sentinels, breeding mates, and progeny were assessed. Neonatal SCID and BALB/c mice inoculated with MPV were evaluated over 24 wk; several mice from each strain were mated once during this period. Fecal MPV loads for each cage were determined weekly by quantitative polymerase chain reaction (PCR) analysis, and all mice were evaluated by quantitative PCR analysis of lymphoid tissues and seroconversion to MPV antigens in immunocompetent mice. Results indicated persistently high fecal shedding of MPV in SCID mice throughout the evaluation period sufficient to allow transmission to sentinels, naïve breeding partners, and the progeny of infected male mice and naïve partners. Lymphoid tissue viral loads in the progeny of infected female SCID mice were high at weaning but low at 6 wk of age. Infected BALB/c mice shed high levels of MPV in feces for 3 wk postinoculation, with seroconversion only in sentinels exposed during the first 2 wk postinoculation. Thereafter the feces of infected BALB/c mice and the lymphoid tissues of sentinels, naïve breeding partners, and progeny intermittently contained extremely low levels of MPV DNA. Although pregnancy and lactation did not increase viral shedding in BALB/c mice, MPV exposure levels were sufficient to induce productive infection in some BALB/c progeny. These data indicate that the adaptive immune response suppresses, but does not eliminate, MPV shedding; this suppression is sufficient to inhibit infection of weanling and adult mice but allows productive infection of some progeny.

Two natural outbreaks of mouse minute virus (MMV) are described. Observations during management of the naturally infected colonies led to a study in which 4-wk-old C57BL/6NCr and C57BL/6Tac mice were inoculated oronasally with an immunosuppressive variant of MMV (MMVi), as were adult C57BL/6NCr lactating dams or their pups (age, 10 d). By day 28 postinoculation, 100% of the 4-wk-old male C57BL/6NCr and C57BL/6Tac mice, 56.2% of 4-wk-old C57BL/6NCr female and 62.5% of 4-wk-old C57BL/6Tac female mice, 100% of adult lactating C57BL/6NCr dams, and 100% of inoculated pups (10 d) had seroconverted. Serologically positive nursing dams did not infect their nursing pups. In contrast, when nursing pups were inoculated, 100% of their dams seroconverted by 28 d postinoculation. Only 1 of 4 facility sentinels (Tac:SW female mice) seroconverted to MMVi and none of the 4 research sentinels (Tac:SW female mice) seroconverted under a once-weekly bedding transfer program. Consequently, 4 new research Tac:SW sentinels of each gender (n = 8) were placed in known-positive cages at cage-change; 100% of the male mice but 0% of the females seroconverted by day 48. Study results suggest gender influences both infectivity and the ability to detect subclinical infections of MMVi. Other factors that may influence detection of MMV include mouse strain or stock, short shedding period, and prolonged time between cage changes. In light of the data from both the natural infections and the experimental cases, cessation of breeding likely will be beneficial when trying to eradicate this virus.

Mortality after influenza is often due to secondary bacterial pneumonia with Streptococcus pneumoniae, particularly in the elderly. The reasons for the high fatality rate seen with this disease are unclear. To further characterize the pathogenesis of pneumonia after influenza in a mouse model, we examined the pathology and immunology that leads to fatal infection. Influenza-infected mice were either euthanized 24 h after secondary infection with S. pneumoniae for determination of pathology, bacterial cultures, and levels of immune effectors or were followed by use of a live imaging system for development of pneumonia. Influenza-infected mice challenged with each of 3 serotypes of pneumococcus developed a severe, necrotic pneumonia and met endpoints for euthanasia in 24 to 60 h. Strikingly elevated levels of both pro- and anti-inflammatory molecules including interleukins 6 and 10, macrophage inflammatory protein 1α, and chemokine KC were present in the blood. High levels of these cytokines and chemokines as well as tumor necrosis factor α, interleukin 1β, and heme oxygenase 1 were present in the lungs, accompanied by a massive influx of neutrophils. Mortality correlated with the development of pneumonia and lung inflammation but not with bacteremia. This model has the potential to help us understand the pathogenesis of severe lung infections.

Serologic monitoring of sentinel mice exposed to soiled bedding is a common method of detecting viral infections in mice. Because bedding transfer protocols vary, the sensitivity of this method has not been documented sufficiently. We examined the reliability of bedding transfer during various stages of infection with mouse parvovirus (MPV) and mouse hepatitis virus (MHV). Most mice exposed to bedding contaminated with MPV 0, 3, or 7 d previously seroconverted, whereas only mice exposed to bedding contaminated with MHV 4 h previously seroconverted, thus confirming the differing stabilities of these viruses. Index mice were inoculated with 30 times the infectious dose 50 (ID₅₀) of MPV or 300 ID₅₀ of MHV. At 3 d, 1 wk, and 2 wk postinoculation (PI), we transferred 25, 50, or 100 ml of bedding to cages of sentinel mice. Viral infection and shedding by index mice was confirmed by serology and fecal polymerase chain reaction assay. Transfer of soiled bedding between mice in static cages induced seroconversion of sentinel mice most reliably during peak viral shedding (1 wk PI for MPV and 3 d PI for MHV). Soiled bedding transfer between mice in individually ventilated cages induced a higher prevalence of sentinel seroconversion to MPV and MHV than that after transfer between mice in static cages. Our findings indicate that although soiled bedding transfer is an effective method for detecting MHV and MPV under optimal conditions, the method is less than 100% reliable under many conditions in contemporary mouse facilities.

The University of Massachusetts Medical School maintains 3 separate research colonies of Xenopus laevis, with each colony located in a separate building on campus. After a 5-wk in-house quarantine period, 34 wild-caught X. laevis were transferred into one of the existing colonies. As a result, this colony grew from 51 to 85 frogs. All animals were housed in a recirculating frog housing system. During the first 2 mo, 6 frogs died suddenly, and health reports were generated for another 10 frogs in this colony. The majority of health reports were written in response to acute coelomic distention. These patterns continued until, after 1 y, only 25 of the original 85 animals remained. Necropsies performed showed large accumulations of serosanguinous fluid in the subcutaneous space or body cavity. Granulomatous inflammatory lesions with acid-fast bacilli were generally present in the liver, lung, or spleen. Culture of affected tissues grew Mycobacterium sp. within 40 d. Polymerase chain reaction analysis confirmed the isolated organism to be the same species of Mycobacterium (provisionally named M. liflandii) recently reported by 2 other groups. However, previous clinical publications suggested that this bacterium originated only from X. tropicalis. The cases we present highlight the rapidly lethal effects of M. liflandii in a colony of wild-caught X. laevis and illustrate the need to dedicate further attention to this emerging Xenopus disease.

We investigated the prevalence, distribution, and transmission of simian T-lymphotropic virus type 1 (STLV1) in a baboon breeding colony over a 4-y period. We used polymerase chain reaction amplification of the proviral tax gene to assess the infection status of 272 animals housed in 4 separate corrals. Sequencing the proviral envelope gene from individual baboons detected several molecular subtypes (genotypes) of STLV1. At the start of the study, 31% (54 of 176) of all baboons were infected; the majority of infections (91%) were in mature females, with only 3 of 12 mature males and 2 of 48 infants and juveniles being infected. Over the next 4 years, 41 new infections were diagnosed. Of these, 83% occurred in sexually mature female baboons (at least 3 y of age), 17% in infants and juveniles (younger than 3 y), and 0% in mature males. The 7 infections in juveniles were probably derived from mother-to-infant transmission because mother–infant pairs consistently were infected with the same viral genotype. Of the 34 new infections in sexually mature female baboons, the genotyping data showed that 25 (73%) originated from other infected females as opposed to males. Male-to-female sexual transmission may have accounted for the remaining 9 new infections. There was no evidence of female-to-male sexual transmission. The high percentage of female-to-female transmission of STLV1 in our baboons was unexpected; we speculate that transmission may have occurred due to blood contamination from biting during aggressive behavior between females in establishing hierarchical dominance.

West Nile virus (WNV) surfaced as an emerging infectious disease in the northeastern United States in 1999, gradually spread across the continent, and is now endemic throughout North America. Outdoor-housed nonhuman primates at the Tulane National Primate Research Center (TNPRC) in Louisiana were documented with a relatively high prevalence (36%) of antibodies to West Nile virus. We examined the prevalence of antibodies to WNV in a nonhuman primate population housed in outdoor colonies at the Yerkes National Primate Research Center Field Station located near Atlanta, Georgia. We screened rhesus macaques (Macaca mulatta) and sooty mangabeys (Cercocebus atys) that were at least 3 y old by serum neutralization for antibodies to WNV and confirmed these results by hemagglutination-inhibition assay. None of the 45 rhesus monkeys had antibodies to WNV, but 3 of the 45 mangabeys (6.6%) were positive by both serum neutralization and hemagglutination-inhibition tests. The ratio of seroprevalences in the TNPRC and Yerkes primate populations was similar to the ratio of WNV incidences in people in Louisiana and Georgia from 2002 to 2004. The difference in the exposure of nonhuman primates (and possibly humans) to WNV between these 2 regions is consistent with the difference in the abundance of mammal-biting WNV-infectious mosquitoes, which was 23 times lower near Yerkes than around TNPRC in 2003 and 33 times lower in 2004.

Changes in levels of antibody to B virus (Cercopithecine herpesvirus 1; BV) were examined in BV-positive macaques by ELISA. We observed increases in anti-BV IgG titers in a BV-infected cynomolgus monkey after overseas transportation by air and in a rhesus monkey after transfer from an outdoor group cage to an indoor individual cage. Although shedding of infectious virus was not examined, the increase in antibody titer suggested reactivation of BV. Interestingly, we also found an increase in anti-BV IgG levels during the breeding season in male but not female Japanese macaques kept in an enclosed outdoor colony. Further studies should be performed to investigate whether reactivation of BV led to the observed increase in the anti-BV antibody titer.

We noted naturally occurring infection with Clostridium piliforme (Tyzzer's disease) in 2 captive-reared cotton-top tamarins (Saguinus oedipus). Spontaneous Tyzzer's disease has been reported in multiple species of laboratory, domestic, and wild animals but is extremely rare in humans and nonhuman primates. Distinct from idiopathic colitis, which is common in cotton-top tamarins, these 2 tamarins had severe, transmural, necrotizing typhlocolitis accompanied by myocarditis and hepatitis. Abundant bacteria compatible with C. piliforme, the etiologic agent of Tyzzer's disease, were present adjacent to lesions in the cecum–colon, liver, and heart. Therefore, colitis caused by C. piliforme, although rare, should be included as a differential diagnosis in cotton-top tamarins and as a cause of postnatal mortality in this species.