The Fischer 344 rat reflects human susceptibility to francisella pulmonary challenge and provides a new platform for virulence and protection studies.

BACKGROUND: The pathogenesis of Francisella tularensis, the causative agent of tularemia, has been primarily characterized in mice. However, the high degree of sensitivity of mice to bacterial challenge, especially with the human virulent strains of F. tularensis, limits this animal model for screening of defined attenuated vaccine candidates for protection studies. METHODS AND FINDINGS: We analyzed the susceptibility of the Fischer 344 rat to pulmonary (intratracheal) challenge with three different subspecies (subsp) of F. tularensis that reflect different levels of virulence in humans, and characterized the bacterial replication profile in rat bone marrow-derived macrophages (BMDM). In contrast to the mouse, Fischer 344 rats exhibit a broader range of sensitivity to pulmonary challenge with the human virulent subsp. tularensis and holarctica. Unlike mice, Fischer rats exhibited a high degree of resistance to pulmonary challenge with LVS (an attenuated derivative of subsp. holarctica) and subsp. novicida. Within BMDM, subsp. tularensis and LVS showed minimal replication, subsp. novicida showed marginal replication, and subsp. holartica replicated robustly. The limited intramacrophage replication of subsp. tularensis and novicida strains was correlated with the induction of nitric oxide production. Importantly, Fischer 344 rats that survived pulmonary infection with subsp. novicida were markedly protected against subsequent pulmonary challenge with subsp. tularensis, suggesting that subsp. novicida may be a useful platform for the development of vaccines against subsp. tularensis. CONCLUSIONS: The Fischer 344 rat exhibits similar sensitivity to F. tularensis strains as that reported for humans, and thus the Fischer 344 ray may serve as a better animal model for tularemia vaccine development.

Detection of Pneumocystis carinii in rats by polymerase chain reaction: comparison of lung tissue and bronchoalveolar lavage specimens.

The purpose of this study was to determine and compare the efficiency of detection of Pneumocystis carinii in rats by polymerase chain reaction (PCR) of DNA extracted from two sampling locations: lung tissue and bronchoalveolar lavage. The study involved naturally infected F344 rats that were allotted to groups to intercollate the investigation of several variables, including nonimmunosuppressed rats, rats subjected to a timed induction sequence of 1 to 4 weeks of immunosuppression, and two immunosuppressants: a corticosteroid and cyclophosphamide. The PCR amplified a 357-base pair region contained within the gene encoding the large ribosomal RNA subunit of P. carinii mitochondrial DNA. The identity of the PCR product was confirmed by Southern blot analysis with an oligonucleotide probe. In a comparison of lung bronchoalveolar lavage specimens after immunosuppression, P. carinii was detected by PCR in 100% of lung tissue but in only 87.5% of the lavage specimens. Lung tissue of three animals was test-positive when the corresponding lavage specimen was negative by PCR analysis. The PCR detected P. carinii in both types of specimens from the same two of three nonimmunosuppressed rats. In all there was 88% agreement of PCR results between the two sampling techniques. The difference in diagnostic outcome for the two specimen types was not statistically significant (Fisher's exact test). It was concluded that both specimen types were adequate for PCR detection of P. carinii in rats.

Karyotypes of Pneumocystis carinii from Korean rats.

Molecular karyotyping was applied to Pneumocystis carinii(Pc) from two strains of experimental rats, Sprague Dawley(SD) and Fisher(F), in Korea. Field inversion gel electrophoresis and contour clamped homogeneous electric field electrophoresis resolved 15 chromosomal bands from the Pc. The size of the bands was estimated 270kb to 684kb from SD rats, and 273kb to 713 kb from F rats. The bands of 283 kb from SD rats and of 273 kb from F rats stained more brightly suggesting duplicated bands. Total number of chromosomes was at least 16, and total genomic size was estimated 7 x 10(6) bp. All of the bands from F rats hybridized to the probe of repeated DNA sequences of Pc and the band of 448 kb size was proved to contain rDNA sequences, but Pc. chromosome bands from SD rats showed no reactions to the probes. The 2 different karyotypes of P. carinii from 2 strains of rats were maintained consistently for 2 years.

Histopathologic observations in weanling B6C3F1 mice and F344/N rats and their adult parental strains.

Weanling Fischer 344/N (F344) rats and the first filial hybrid of C57BL/6 x C3H (B6C3F1) mice and retired breeders from the parental stocks of these strains were monitored over a 5-yr-period by examining the histopathology of selected organs and comparing those results to viral and mycoplasmal serology and the intestinal tract bacterial flora of each animal on an individual basis. Serology gave no evidence of viral infection, but Mycoplasma arthriditis antibodies were detected. Reactivity of serum of adult C57BL/6 female mice with control cells or media (tissue culture, TC) was seen in a significant number of mice. TC reactivity correlated positively with lymphoid perivascular infiltrates, predominantly of the lungs, suggesting an allergic response in development of the lesions. Other lesions of note consisted of Harderian gland inflammation of rats, focal necrotizing lesions of the liver of both species, and thickening of the pleura and adjacent pulmonary interstitium of weanling rats. Embolization of bacteria from the gastrointestinal tract to the liver was considered a possible cause of the liver necrosis in both species. Although lesions of the lung and Harderian gland of the rats are similar to those caused by known viral agents, the cause of the latter could not be determined as these animals were negative for viral antibodies and the former was considered to be related to incomplete pulmonary development in the young rat. Features differentiating the lesions observed in animals of this survey from those caused by viral infection are discussed.

Evaluation of inbred germ-free Fischer 344 albino rats as an experimental model for oral candidiasis.

Inbred germ-free Fischer 344 albino rats were evaluated as models for experimental candidiasis in order to investigate bacterial interaction with Candida albicans. Female rats were exposed to C. albicans in their drinking water and killed at intervals from 2 to 22 days after initial contact with the contaminant. C. albicans was cultured from their mouths from day 2 but from day 12 the number of colonies decreased. From day 2 to 9 all rats showed oral histological signs of candidal infestation, but after 9 days the number declined to 3 out of 9 at 22 days. The dorsal surface of the tongue was the best histological indicator of candidal infestation. All the rats had tongue lesions from day 4 to 9, and from day 6 there was also a concomitant localized loss of filiform papillae. The number of rats with all forms of tongue involvement also decreased after 9 days with only 3 out of 9 affected at 22 days. It is concluded that Fischer 344 inbred germ-free rats can be used on a limited scale as a model for candidiasis and bacterial interaction with C. albicans, the dorsal surface of the tongue would be the best site for studying candidal experimental lesions and it is probable that better results can be achieved with complete standardization of contamination and preparation procedures.

Fecal mutagens and Bacteroides fragilis levels in the feces of dimethylhydrazine-treated rats: influence of diet.

In a study designed to investigate the effects of dietary synergisms on 1,2-dimethylhydrazine (DMH)-induced rat colon carcinogenesis, fecal pellets were examined for the presence of direct-acting fecal mutagens and levels of Bacteroides fragilis group organisms. Intraperitoneal injections of DMH at 10 mg/kg were given for 16 weeks (weeks 3-18) to 160 male F344 rats consuming 4 supplemental dietary factors in all possible combinations. The dietary factors examined were wheat bran (15%), cholesterol (1%), beef tallow (18%) and indole-3-carbinol (IC) (0.1%). Feces were collected 3, 10, 17, 24 and 31 weeks after commencing the dietary treatments and dichloromethane extracts were assayed using the Salmonella typhimurium TA100 without metabolic activation. The numbers of B. fragilis group organisms were enumerated in feces collected at the same time. Most feces samples were negative for mutagens but extracts from weeks 17-31 showed a significant mutagenic response from the IC factor in the diet. The fecal levels of B. fragilis were significantly increased by the inclusion of cholesterol in the diets. The B. fragilis counts and fecal mutagen production were not correlated (r = 0.09), although species of the B. fragilis group have been implicated in the production on human fecal mutagens.

Comparison of methods for detection of Mycoplasma pulmonis in experimentally and naturally infected rats.

Isolation, indirect immunofluorescence, an enzyme-linked immunosorbent assay (ELISA), and histopathological examination of tissues for characteristic lesions were evaluated for their efficiency in detecting Mycoplasma pulmonis infection in rats. Whereas all of the methods were efficient in naturally infected Sprague-Dawley rats, none of the methods consistently detected infection in F344 rats experimentally infected with low doses of the organism. In the experimental infections, however, the success rate of any method was directly related (P less than 0.05) to increasing inoculum dose and time postinoculation. Collectively, the data indicated that isolation of M. pulmonis was the most efficient single detection method and the nasopharyngeal duct was the best single site to culture, although sampling of multiple sites within the respiratory tract increased the rate of isolating the organism. The ELISA was understandably the least sensitive method in the low-dose, experimentally infected rats because of the time required for development of a detectable serum antibody response. Although each of the four methods identified a high percentage of naturally infected rats, the ELISA was the most efficient method in these animals as it was uniformly positive. The use of combinations of methods was found to increase the rate of detection of M. pulmonis infection in both experimentally and naturally infected rats.