Comparative evaluation of establishing a human gut microbial community within rodent models.

The structure of the human gut microbial community is determined by host genetics and environmental factors, where alterations in its structure have been associated with the onset of different diseases. Establishing a defined human gut microbial community within inbred rodent models provides a means to study microbial-related pathologies, however, an in-depth comparison of the established human gut microbiota in the different models is lacking. We compared the efficiency of establishing the bacterial component of a defined human microbial community within germ-free (GF) rats, GF mice, and antibiotic-treated specific pathogen-free mice. Remarkable differences were observed between the different rodent models. While the majority of abundant human-donor bacterial phylotypes were established in the GF rats, only a subset was present in the GF mice. Despite the fact that members of the phylum Bacteriodetes were well established in all rodent models, mice enriched for phylotypes related to species of Bacteroides. In contrary to the efficiency of Clostridiales to populate the GF rat in relative proportions to that of the human-donor, members of Clostridia cluster IV only poorly colonize the mouse gut. Thus, the genetic background of the different recipient rodent systems (that is, rats and mice) strongly influences the nature of the populating human gut microbiota, determining each model's biological suitability.

Coping with parvovirus infections in mice: health surveillance and control.

Parvoviruses of mice, minute virus of mice (MVM) and mouse parvovirus (MPV), are challenging pathogens to eradicate from laboratory animal facilities. Due to the impediment on rodent-based research, recent studies have focused on the assessment of re-derivation techniques and parvoviral potential to induce persistent infections. Summarizing recent data, this review gives an overview on studies associated with parvoviral impact on research, diagnostic methods, parvoviral persistence and re-derivation techniques, demonstrating the complex nature of parvovirus infection in mice and unfolding the challenge of controlling parvovirus infections in laboratory animal facilities.

Presence of Minute virus of mice in immunocompetent mice despite the onset of host immunity.

Infections with the autonomous parvovirus Minute virus of mice (MVM) are generally characterized as acute and self-limiting. However, MVM remains with considerably high prevalence rates in laboratory mouse colonies impeding rodent based research. The objective of this study was to assess whether the immunosuppressive variant of MVM (MVMi) establishes a persistent infection in immunocompetent adult mice. Therefore, we approached the question whether replicating and/or infectious virus is present in mice after the decline of viral shedding and whether immunosuppression might modify the infection. Dissection or induction of immunosuppression of individually housed mice was performed at 8 weeks post inoculation after fecal samples tested negative for viral DNA for at least 2 subsequent weeks as determined by weekly PCR analyses. MVMi mRNA was detected by both, RT-PCR and in situ RT-PCR in spleens at 8 weeks post inoculation with positive cells resembling lymphocytes and macrophages. These findings and the use of explant cultures strongly indicated the presence of replicating virus in spleens at 8 weeks post inoculation. Following immunosuppression (by irradiation), an induction of viral shedding was observed. Additionally, an increase in the amount of viral DNA was detected by real-time qPCR in mesenteric lymph nodes after irradiation. In summary, our data support the notion that MVMi persists in lymphoid tissue of immunocompetent adult mice despite the onset of host immunity.

Risk assessment of minute virus of mice transmission during rederivation: detection in reproductive organs, gametes, and embryos of mice after in vivo infection.

Murine parvoviruses, including minute virus of mice (MVM), represent major infectious disease problems encountered in contemporary laboratory animal research facilities with embryo transfer (ET), one of the most widely used techniques for rederivation. Using an in vivo approach, the objectives of this study were to assess the risk of MVM transmission during rederivation and to provide data that allow recommendation of preventive measures. Therefore, we determined whether immunosuppressive variant MVMi viral DNA is detectable in reproductive organs, gametes (oocytes and spermatozoa), and embryos collected from experimentally infected mice and whether washing as recommended before ET eliminates MVMi sufficiently from gametes and embryos. Fractions of reproductive organs tested positive from Day 5 to Day 30 postinoculation, demonstrating a risk for a minimum period of 4 wk; the highest incidence of positive organs was found between Day 9 and Day 13 postinoculation. Real-time PCR detected viral DNA to a lesser extent in male than in female reproductive organs. MVMi DNA was detected in oocytes and sperm cells derived after in vivo infection but not in two-cell embryos. In vitro contamination studies revealed that the virus firmly adheres to the zona pellucida after 10 wash steps, indicating that even extensive washing might not eliminate MVMi completely from embryos. According to this systematic in vivo approach, recommended measures to prevent transmission of MVM during rederivation include sufficient washing of embryos, accompanying testing using adequate (PCR) methods, and using embryos rather than in vitro fertilization techniques; furthermore, the exchange of gametes should be considered a risk factor.

CpG motifs of bacterial DNA exert protective effects in mouse models of IBD by antigen-independent tolerance induction.

BACKGROUND & AIMS: Prophylactic treatment of mice with CpG motifs of bacterial DNA protects from experimental inflammatory bowel disease, at least partly via induction of inhibitory T-cells. The aim of this study was to elucidate whether these CpG-dependent protective effects require presence of bacterial flora suggesting antigen-specific regulatory activity. METHODS: Germ-free BALB/c and IL-10(-/-) mice were treated with CpG-oligodeoxynucleotides (ODN), control-ODN, or PBS. CD4(+)CD62L(+) cells of these mice were transferred into SCID recipients. CpG-ODN-treated germ-free IL-10(-/-) mice were transferred into colitogenic environment. Monoclonal antibodies were used to neutralize TGF-beta and IFN-alpha/beta during CpG-ODN treatment. CD4(+)CD62L(+) cells of donors were evaluated for cytokine secretion and FOXP3, PD-1, and CD25 expression. RESULTS: Compared to PBS or control-ODN treatment, CpG-ODN application to germ-free donors led to decreased intestinal inflammation as indicated by histology, decreased proinflammatory cytokines, and increased IL-10 secretion. Protection was also observed after cotransfer of cells from PBS and CpG-ODN treated donors. Anti-TGF-beta and anti-INF-alpha/beta partly reversed the protective CpG-ODN effect. CpG-ODN-treated germ-free IL-10(-/-) mice transferred into colitogenic environment developed significantly less colitis than controls but not recipients of IL-10(-/-)CD4(+)CD62L(+)cells. CD4(+)CD62L(+)cells of CpG-treated germ-free animals displayed increased expression of regulatory markers. CONCLUSIONS: Even without pre-existence of bacterial flora CpG-ODN exposition induces tolerance, indicating that CpG-ODN-induced regulatory T-cells are not bacterial antigen specific. TGF-beta and IFN-alpha/beta play major roles in induction of regulatory cells, and although IL10-independent mechanisms play a role in CpG-ODN protection, this cytokine likely is important for the effector mechanism of CpG-ODN-induced regulatory T-cells.

Minute virus of mice: antibody response, viral shedding, and persistence of viral DNA in multiple strains of mice.

Minute virus of mice (MVM) is a major concern for laboratory animal facilities because it remains with considerably high prevalence despite strict barrier systems. The aim of this study was to elucidate potential risks associated with MVM infection by investigating the role of the genetic background on antibody production and persistence as well as viral shedding. Mice of various strains and stocks were inoculated oronasally with the immunosuppressive strain MVMi; in addition, natural infection was modeled through contact exposure. As determined by serology, seroconversion and serum levels of IgG differed considerably among strains and stocks, especially in the contact-exposed group. For example, C57BL/6J mice responded well to exposure in contrast to FVB/N, NMRI, ICR, and C3H/HeN mice. Titration studies indicated that the viral dose necessary to induce seroconversion was strain-dependent. Experiments to dissect the role of the major histocompatibility complex haplotype in the response to MVMi gave inconclusive results. To detect viral persistence, spleens and feces were analyzed by PCR at 16 wk after exposure, and the infectivity of PCR-positive spleens was investigated by IP and oronasal inoculation of naive mice. Although DNA was detected in the spleens of some mice, feces remained negative, and naive mice were not infected by inoculation. In addition, viral shedding declined rapidly after day 20 postinoculation. In summary, the data show that seroconversion and antibody response to MVMi infection depend on the genetic background of mice, with the infective dose being a critical factor. The role of viral DNA in chronically infected mice will require further elucidation.

Sensitivity to Escherichia coli Nissle 1917 in mice is dependent on environment and genetic background.

Escherichia coli Nissle 1917 (EcN) is a well-characterized probiotic bacterium. Although genomic comparisons of EcN with the uropathogenic E. coli strain CFT073 revealed high degrees of similarity, EcN is generally considered a non-pathogenic organism. However, as recent evidence suggests that EcN is capable of inducing inflammatory responses in host intestinal epithelial cells, we aimed to investigate potential pathogenic properties of EcN in an in vivo model using various germ-free (GF) mouse strains. With the exception of C3H/HeJZtm mice, which carry a defective toll-like receptor (TLR)4-allele, no lesions were obvious in mice of different strains orally inoculated with EcN for 1 week, although organ cultures (blood, lung, mesenteric lymph node, pancreas, spleen, liver and kidney) tested positive to various degrees. C3H/HeJZtm mice inoculated with EcN became clinically ill and the majority died or had to be euthanized. Organs of all gnotobiotic C3H/HeJZtm mice were positive for EcN by culture; major histological findings were moderate to severe pyogranulomatous serositis, typhlitis and pancreatitis. Histological findings were corroborated by highly elevated tumour necrosis factor (TNF) serum levels. Lesions were not detected in specified pathogen free maintained C3H/HeJZtm mice, GF C3H/HeJ mice lacking the interleukin-10 gene, or GF C3H/HeJZtm mice that were inoculated with E. coli K12 strain MG1655 as a control. In addition, mild histological lesions were detected in Ztm:NMRI mice 3 months after oral inoculation with EcN. This study shows that EcN is capable of displaying a virulent phenotype in GF C3H/HeJZtm mice. Whether this phenotype is linked to the bacterium's probiotic nature should be the focus of further studies.