RESUMO
Chlamydia muridarum (Cm) has reemerged as a prevalent bacterial contaminant of academic research mouse colonies. A study was conducted to assess the effectiveness of husbandry and cage sanitization methods in preventing intercage transmission of Cm. To assess intercage transmission during cage change, a cage housing 2 Cm-free Swiss Webster (SW; Tac:SW) sentinel mice was placed randomly on each of 12 individually ventilated cage racks, housing cages with Cm-shedding mice, located in one of 2 animal holding rooms. Husbandry staff blinded to the study cages changed all cages in the animal holding rooms weekly using a microisolation cage technique. PCR testing performed at 180 d postplacement confirmed all mice remained negative for Cm. To assess the effectiveness of cage sanitization to eliminate Cm, we investigated transmission of Cm to a naive Cm-free SW and NOD.Cg-Prkdcscid Il2rgtm1Wjl/SzJ (NSG) mouse cohoused for 7 d (repeated weekly for 4 wk) in cages assigned to one of 3 groups (n = 10 pairs of mice/group). Cages that previously housed 2 Cm-shedding BALB/c mice were either washed in a tunnel washer (82.2 °C [180 °F] final rinse for an average of 16 s per run; n = 10) with and without postwashing autoclaving (121 °C for 20 min; n = 10), or were untreated (bedding change only; n = 10). Pre- and postsanitization swabs of each cage were assayed for Cm by PCR. All pretreatment swabs tested positive, while posttreatment swabs from all cages (excluding bedding change) tested negative. All SW and NSG mice, irrespective of group, remained negative for Cm as determined by PCR. These findings suggest that infectious Cm does not persist in untreated cages or after mechanical washing with and without autoclaving. Collectively, these findings suggest that neither our husbandry protocols nor inadequate cage sanitization methods likely contributed to the observed prevalence of Cm in contemporary research mouse colonies.
RESUMO
Chlamydia muridarum (Cm) has reemerged as a moderately prevalent infectious agent in research mouse colonies. Despite its' experimental use, there are limited studies evaluating Cm's effects on immunocompetent mice following its natural route of infection. A Cm field isolate was administered (orogastric gavage) to 8-week-old female BALB/cJ (C) mice. After confirming shedding (through 95d), these mice were cohoused with naïve C57BL/6J (B6), C, and Swiss (J:ARC[S]) mice (n=28/strain) for 30 days. Cohoused mice (n=3-6 exposed and 1-6 control/strain) were evaluated 7, 14, 21, 63, 120, and 180 days post-cohousing (DPC) via hemograms, serum biochemistry analysis, fecal qPCR, histopathology, and Cm MOMP immunohistochemistry. Immunophenotyping was performed on spleen (B6, C, S; n=6/strain) and intestines (B6; n=6) at 14 and 63 DPC. Serum cytokine concentrations were measured (B6; n=6 exposed and 2 control) at 14 and 63 DPC. All B6 mice were shedding Cm by 3 through 180 DPI. One of 3 C and 1 of 6 S mice began shedding Cm at 3 and 14 DPC, respectively, with the remaining shedding thereafter. Clinical pathology was nonremarkable. Minimal-to-moderate enterotyphlocolitis and gastrointestinal associated lymphoid tissue (GALT) hyperplasia was found in numerous infected mice. Cm antigen was frequently detected in GALT-associated surface intestinal epithelial cells. Splenic immunophenotyping revealed increased monocytes and shifts in T cell population subsets in all strains/timepoints. Gastrointestinal immunophenotyping (B6) revealed sustained increases in total inflammatory cells and elevated cytokine production in innate lymphoid cells and effector T cells (large intestine). Elevated concentrations of pro-inflammatory cytokines were detected in the serum (B6). These results demonstrate that while clinical disease was not appreciated, 3 commonly utilized strains of mice are susceptible to chronic enteric infections with Cm which may alter various immune responses. Considering the widespread use of mice to model GI disease, institutions should consider excluding Cm from their colonies.
RESUMO
Chlamydia muridarum (Cm) is a moderately prevalent, gram-negative, intracellular bacterium that affects laboratory mice, causing subclinical to severe disease, depending on the host's immune status. The effectiveness of various antibiotic regimens aimed at eradicating Cm in both immunodeficient and immunocompetent laboratory mice was evaluated. NSG mice were cohoused with Cm-shedding BALB/cJ mice for 14 days to simulate natural exposure. Four groups of 8 infected NSG mice were treated for 7 days with either 0.08% sulfamethoxazole and 0.016% trimethoprim (TMS) in water, 0.0625% doxycycline in feed, 0.124%/0.025% TMS in feed, or 0.12% amoxicillin in feed. A control group was provided standard water and feed. The impact of treatment on gastrointestinal microbiota (GM) was performed through shotgun sequencing on the last day of treatment. TMS and Amoxicillin had negligible effects on GM, while doxycycline had the largest effect. All antibiotic treated NSG mice exhibited clinical disease, including dehydration, hunched posture, >20% weight loss, and dyspnea, leading to euthanasia 21-40 days post-treatment (32.6 ± 4.2 days; mean ± SD). Untreated controls were euthanized 14-33 days post-exposure (23.75 ± 5.9 days). All mice were fecal PCR positive for Cm at euthanasia. Histological evaluation revealed multifocal histiocytic and neutrophilic bronchointerstitial pneumonia and/or bronchiolitis featuring prominent intralesional chlamydial inclusion bodies in all mice. Subsequently, groups of 8 C57BL/6J, BALB/cJ, NOD.SCID, and NSG mice infected with Cm were treated with 0.124%/0.025% TMS in feed for 7 (BALB/cJ and C57BL/6J) or 21 days (NSG and NOD.SCID). All immunocompetent and NOD.SCID mice were negative for Cm by PCR 14 days post-treatment, remained clinically normal and had no evidence of Cm infection at necropsy, all NSG mice remained Cm positive and were euthanized. While these findings highlight the difficulties in eradicating Cm from highly immunodeficient mice, eradication of Cm from immunocompetent or moderately immunocompromised mice with antibiotics is feasible.
RESUMO
Chlamydia muridarum (Cm), an intracellular bacterium of historical importance, was recently rediscovered as moderately prevalent in research mouse colonies. Cm was first reported as a causative agent of severe pneumonia in mice about 80 y ago, and while it has been used experimentally to model Chlamydia trachomatis infection of humans, there have been no further reports of clinical disease associated with natural infection. We observed clinical disease and pathology in 2 genetically engi- neered mouse (GEM) strains, Il12rb2 KO and STAT1 KO, with impaired interferon-γ signaling and Th1 CD4+ T cell responses in a colony of various GEM strains known to be colonized with and shedding Cm. Clinical signs included poor condition, hunched posture, and poor fecundity. Histopathology revealed disseminated Cm with lesions in pulmonary, gastrointestinal, and urogenital tissues. The presence of Cm was confirmed using both immunohistochemistry for Cm major outer membrane protein-1 antigen and in situ hybridization using a target probe directed against select regions of Cm strain Nigg. Cm was also found in association with a urothelial papilloma in one mouse. These cases provide additional support for excluding Cm from research mouse colonies.
Assuntos
Infecções por Chlamydia , Chlamydia muridarum , Camundongos Knockout , Fator de Transcrição STAT1 , Animais , Infecções por Chlamydia/patologia , Infecções por Chlamydia/veterinária , Infecções por Chlamydia/microbiologia , Camundongos , Fator de Transcrição STAT1/genética , Fator de Transcrição STAT1/metabolismo , Feminino , Receptores de Interleucina-12/deficiência , Receptores de Interleucina-12/genética , Masculino , Pneumopatias/microbiologia , Pneumopatias/patologia , Pneumopatias/veterináriaRESUMO
Chlamydia muridarum (Cm) has reemerged as a prevalent bacterial contaminant of academic research mouse colonies. A study was conducted to assess the effectiveness of husbandry and cage sanitization methods in preventing intercage transmission of Cm. To assess intercage transmission during cage change, a cage housing 2 Cm-free Swiss Webster (Tac:SW; SW) sentinel mice was placed randomly on each of 12 individually ventilated cage racks, housing cages with Cm-shedding mice, located in 1 of 2 animal holding rooms. Husbandry staff blinded to the study cages, changed all cages in the animal holding rooms weekly using microisolator cage technique. PCR testing performed 180 days post-placement confirmed all mice remained negative for Cm. To assess the effectiveness of cage sanitization to eliminate Cm, we investigated transmission of Cm to a naïve Cm-free SW and NOD.Cg- Prkdc scid Il2rg tm1Wjl /SzJ (NSG) mouse co-housed for 7 days (repeated weekly for 4 weeks) in cages assigned to 1 of 3 groups (n=10 pairs of mice/group). Cages that previously housed 2 Cm-shedding BALB/c mice were either washed in a tunnel washer (82.2°C [180°F] final rinse for an average of 16 seconds per run; n=10) with and without post-washing autoclaving (121°C for 20 minutes; n=10), or were untreated (bedding change only; n=10). Pre- and post-sanitization swabs of each cage were assayed for Cm by PCR. All pre-treatment swabs tested positive, while post-treatment swabs from all cages (excluding bedding change) tested negative. All SW and NSG mice, irrespective of group, remained negative for Cm as determined by PCR. These findings suggest that infectious Cm does not persist in untreated cages nor after mechanical washing with and without autoclaving. Collectively, these findings suggest that neither our husbandry protocols nor inadequate cage sanitization methods likely contributed to the observed prevalence of Cm in contemporary research mouse colonies.
RESUMO
The murine bacterial pathogen Chlamydia muridarum (Cm) has been used to study human Chlamydia infections in various mouse models. CD4+ T-cells, natural killer cells, and interferon-gamma (IFN-γ)-mediated immunity are important to control experimentally induced Cm infections. Despite its experimental use, natural infection by Cm has not been documented in laboratory mice since the 1940s. In 2022, the authors reported the discovery of natural Cm infections in numerous academic institutional laboratory mouse colonies around the globe. To evaluate the impact of Cm infection in severely immunocompromised mice, 19 NOD.Cg-PrkdcscidIl2rgtm1Wjl/SzJ (NSG) mice were cohoused with Cm shedding, naturally infected immunocompetent mice and/or their soiled bedding for 4 weeks and subsequently euthanized. Clinical disease, characterized by lethargy, dyspnea, and weight loss, was observed in 11/19 NSG mice, and 16/18 NSG mice had neutrophilia. All mice exhibited multifocal to coalescing histiocytic and neutrophilic bronchointerstitial pneumonia (17/19) or bronchiolitis (2/19) with intraepithelial chlamydial inclusions (CIs). Immunofluorescence showed CIs were often associated with bronchiolar epithelium. CIs were frequently detected by immunohistochemistry in tracheal and bronchiolar epithelium (19/19), as well as throughout the small and large intestinal epithelium without lesions (19/19). In a subset of cases, Cm colonized the surface epithelium in the nasopharynx (16/19), nasal cavity (7/19), and middle ear canal (5/19). Endometritis and salpingitis with intraepithelial CI were identified in a single mouse. These findings demonstrate that Cm infection acquired through direct contact or soiled bedding causes significant pulmonary pathology and widespread intestinal colonization in NSG mice.
Assuntos
Infecções por Chlamydia , Chlamydia muridarum , Pneumonia , Feminino , Animais , Camundongos , Humanos , Camundongos Endogâmicos NOD , Camundongos SCID , Infecções por Chlamydia/veterinária , Infecções por Chlamydia/microbiologia , Pneumonia/veterinária , Proteínas de Ligação a DNA , Proteína Quinase Ativada por DNA , Subunidade gama Comum de Receptores de InterleucinaRESUMO
Rotifers, Brachionus plicatilis, are a valuable first exogenous feed for zebrafish because they can provide continuous nutrition for growing zebrafish larvae when used in a rotifer-zebrafish polyculture. Typically cultured at high salinities (>10 ppt), B. plicatilis are temporarily immobilized when moved to lower salinities (5 ppt) used for polycultures, decreasing their accessibility and attractiveness to the larvae. The nutritional value of rotifers varies based on their diet, typically live algae, which has limited nutritional value and may pose biosecurity risks. After confirming that rotifers consume and can reproduce when fed an irradiated, processed larval fish diet (PD), they were reared at 5 or 15 ppt, and fed various combinations of an algae mix and/or PD. Population densities and percentages of egg-bearing rotifers were quantified daily until the population density plateaued, and then their nutritional value was assessed. Results indicated that rotifers thrived at both salinities. Those fed PD were successfully maintained at >500 rotifers per mL and contained a greater ω-6/ω-3 fatty acid ratio. Our findings indicate that enriching rotifers with PD raised at 5 ppt can potentially eliminate rotifer immobilization in polyculture, while providing a nutritious, attractive diet for zebrafish larvae and decreasing biosecurity risks.