Occurrence of Helicobacter species other than H. hepaticus in laboratory mice and rats in Sweden.

The aim of this study was to determine which Helicobacter species other than H. hepaticus colonize laboratory mice and rats in Sweden. We analyzed 63 intestinal samples from mice and 42 intestinal samples from rats by partial 16S rDNA sequence analysis. Previously these samples had been found positive for Helicobacter species but negative for H. hepaticus in a polymerase chain reaction screening assay at the National Veterinary Institute in Sweden. H. ganmani, H. typhlonius, H. rodentium, an uncharacterized Helicobacter species ('hamster B'), and a possibly novel species were detected in mice. The possibly novel species was most closely related to H. apodemus strain YMRC 000216 (98.3% sequence similarity). Two different Helicobacter species were detected in rats: H. ganmani and H. rodentium. H. ganmani colonization of rats has not previously been reported.

Health surveillance of specific pathogen-free and conventionally-housed mice and rats in Korea.

The present study contains information about proper microbiological monitoring of laboratory animals' health and the standardization of microbiological monitoring methods in Korea. Microbiological quality control for laboratory animals, composed of biosecurity and health surveillance, is essential to guard against research complications and public health dangers that have been associated with adventitious infections. In this study, one hundred and twenty-two mice and ninety rats from laboratory animal breeding companies and one animal facility of the national universities in Korea were monitored in 2000-2003. Histopathologically, thickening of the alveolar walls and lymphocytic infiltration around the bronchioles were observed in mice and rats from microbiologically contaminated facilities. Cryptosporidial oocysts were observed in the gastric pits of only conventionally-housed mice and rats. Helicobacter spp. infection was also detected in 1 of 24 feces DNA samples in mice and 9 of 40 feces DNA samples in rats by PCR in 2003, but they were not Helicobacter hepaticus. This paper describes bacteriological, parasitological, and virological examinations of the animals.

Amylase-binding proteins A (AbpA) and B (AbpB) differentially affect colonization of rats' teeth by Streptococcus gordonii.

Streptococcus gordonii produces two alpha-amylase-binding proteins, AbpA and AbpB, that have been extensively studied in vitro. Little is known, however, about their significance in oral colonization and cariogenicity (virulence). To clarify these issues, weanling specific pathogen-free Osborne-Mendel rats, TAN : SPFOM(OM)BR, were inoculated either with wild-type strains FAS4-S or Challis-S or with strains having isogenic mutations of abpA, abpB, or both, to compare their colonization abilities and persistence on the teeth. Experiments were done with rats fed a sucrose-rich diet containing low amounts of starch or containing only starch. The mutants and wild-types were quantified in vivo and carious lesions were scored. In 11 experiments, S. gordonii was a prolific colonizer of the teeth when rats were fed the sucrose (with low starch)-supplemented diet, often dominating the flora. Sucrose-fed rats had several-fold higher recoveries of inoculants than those eating the sucrose-free, starch-supplemented diet, regardless of inoculant type. The strain defective in AbpB could not colonize teeth of starch-only-eating rats, but could colonize rats if sucrose was added to the diet. Strains defective in AbpA surprisingly colonized better than their wild-types. A double mutant deficient in both AbpA and AbpB (abpA/abpB) colonized like its wild-type. Wild-types FAS4-S and Challis-S had no more than marginal cariogenicity. Notably, in the absence of AbpA, cariogenicity was slightly augmented. Both the rescue of colonization by the AbpB- mutant and the augmentation of colonization by AbpA- mutant in the presence of dietary sucrose suggested additional amylase-binding protein interactions relevant to colonization. Glucosyltransferase activity was greater in mutants defective in abpA and modestly increased in the abpB mutant. It was concluded that AbpB is required for colonization of teeth of starch-eating rats and its deletion is partially masked if rats eat a sucrose-starch diet. AbpA appears to inhibit colonization of the plaque biofilm in vivo. This unexpected effect in vivo may be associated with interaction of AbpA with glucosyltransferase or with other colonization factors of these cells. These data illustrate that the complex nature of the oral environment may not be adequately modelled by in vitro systems.

Oral colonization and cariogenicity of Streptococcus gordonii in specific pathogen-free TAN:SPFOM(OM)BR rats consuming starch or sucrose diets.

The significance of Streptococcus gordonii in dental caries is undefined, as is that of other alpha-amylase-binding bacteria (ABB) commonly found in the mouth. To clarify the ecological and cariological roles of S. gordonii our specific pathogen-free Osborne-Mendel rats, TAN:SPFOM(OM)BR, were fed either diet 2000 (containing 56% confectioner's sugar, most of which is sucrose) or diet 2000CS (containing 56% cornstarch, in lieu of confectioner's sugar) and inoculated with S. gordonii strains. Uninoculated rats were free of both indigenous mutans streptococci (MS) and ABB, including S. gordonii, as shown by culture on mitis salivarius and blood agars of swabs and sonicates of dentitions after weanlings had consumed these diets for 26 days. ABB were detected by radiochemical assay using [125I]-amylase reactive to alpha-amylase-binding protein characteristic of the surface of S. gordonii and other ABB. No ABB were detected (detection limit < 1 colony-forming units in 10(6) colony-forming units). Thus the TAN:SPFOM(OM)BR colony presents a 'clean animal model' for subsequent study. Consequently, S. gordonii strains Challis or G9B were used to inoculate weanling rat groups consuming either the high-sucrose diet 2000 or the cornstarch diet 2000CS. Two additional groups fed each of these diets remained unioculated. Recoveries of inoculants were tested 12 and 26 days later by oral swabs and sonication of the molars of one hemimandible of each animal, respectively. Uninoculated animals were reconfirmed to be free of ABB and mutans streptococci, but inoculated ones eating diet 2000CS had S. gordonii recoveries of 1-10% or, if eating diet 2000, 10-30% of total colony-farming units in sonicates. There were no statistically significant differences among the inoculated and uninoculated animal groups' caries scores when they ate the cornstarch diet. Lesion scores for sucrose-eating rats were, however, from 2.4-5.1-fold higher than for cornstarch-eating rats, P < 0.001, and were still higher if animals had been inoculated with either Challis (1.41-fold) or G9B (1.64-fold), than if uninoculated, both P < 0.001, so long as the rats ate the sucrose diet. Therefore, TAN:SPFOM(OM)BR rats do not harbour ABB or S. gordonii but can be colonized by S. gordonii. Colonization levels of S. gordonii on the teeth are higher in the presence of high sucrose than with high starch-containing diets. Caries scores are augmented by sucrose compared with starch, and are further augmented by S gordonii colonization. S. gordonii is thus cariologically significant in the presence of sucrose, at least in this rat.

Distribution of Corynebacterium renale among apparently healthy rats.

We examined the distribution of Corynebacterium renale, a causative agent of urinary calculus, in clinically normal rats at 6 animal facilities in Japan. Swabs of the vulva and vaginal vestibule or prepuce of the rats were cultured for isolation of the organisms. C. renale has been isolated at only one animal facility, where cases of urinary calculus were reported several years ago. In this facility, 32% of female (43/135) and 22% of male (18/82) rats, 4-28 weeks old, were positive for C. renale. In contrast, 92 female and 169 male rats at other facilities without a history of the disease were negative for the organisms.

Detection of Pasteurella pneumotropica in laboratory mice and rats by polymerase chain reaction.

A 16S rDNA-based polymerase chain reaction (PCR) method specific for Pasteurella pneumotropica was developed. The PCR product, a 395-base pair DNA fragment, was amplified from P. pneumotropica and not from 42 other bacterial species tested, including four other Pasteurella species and Actinobacillus ureae. The PCR method was used to identify 13 previously isolated strains that had been identified as P. pneumotropica by conventional methods: 12 were confirmed by PCR; one that was PCR-negative was re-examined by biochemical methods and determined to be A. ureae. The PCR detection of P. pneumotropica in nasopharyngeal swab specimens from 121 surveillance animals (15 inbred mice and 5 inbred rats from 20 animal rooms) had a high carrier state in healthy laboratory animals; for example, rat swab specimens were 89.6% (43/48) positive by PCR, 8.3% were positive by the direct culture-biochemical method, and 16.7% were positive by the enrichment culture-biochemical method. The positive rate for mice (21.9% [16/73]) was lower than that for rats.

Chronic active hepatitis and associated liver tumors in mice caused by a persistent bacterial infection with a novel Helicobacter species.

BACKGROUND: In the autumn of 1992, a novel form of chronic, active hepatitis of unknown etiology was discovered in mice at the National Cancer Institute-Frederick Cancer Research and Development Center (NCI-FCRDC), Frederick, Md. A high incidence of hepatocellular tumors occurred in affected animals. The disease entity was originally identified in A/JCr mice that were untreated controls in a long-term toxicologic study. PURPOSE: Our original purpose was to determine the origin and etiology of the chronic hepatitis and to quantify its association with hepatocellular tumors in mice of low liver tumor incidence strains. After a helical microorganism was discovered in hepatic parenchyma of diseased mice, we undertook characterization of the organism and investigation of its relationship to the disease process. METHODS: Hepatic histopathology of many strains of mice and rats, as well as guinea pigs and Syrian hamsters, in our research and animal production facilities was reviewed. Steiner's modification of the Warthin-Starry stain and transmission electron microscopy were used to identify bacteria in the liver. We transmitted the hepatitis with liver suspensions from affected mice and by inoculation with bacterial cultures. Bacteria were cultivated on blood agar plates maintained under anaerobic or microaerophilic conditions and characterized morphologically, biochemically, and by 16S rRNA sequence. RESULTS: We report here the isolation of a new species of Helicobacter (provisionally designated Helicobacter hepaticus sp. nov.) that selectively and persistently colonizes the hepatic bile canaliculi of mice (and possibly the intrahepatic biliary system and large bowel), causing a morphologically distinctive pattern of chronic, active hepatitis and associated with a high incidence of hepatocellular neoplasms in infected animals. CONCLUSIONS: The novel Helicobacter is a likely candidate for the etiology of hepatocellular tumors in our mice. The Helicobacter-associated chronic active hepatitis represents a new model to study mechanisms of carcinogenesis by this genus of bacteria. IMPLICATIONS: Adenocarcinoma of the stomach, the second most prevalent of all human malignancies world-wide, is associated with infection at an early age with Helicobacter pylori. Infection leads to several distinctive forms of gastritis, including chronic atrophic gastritis, which is a precursor of adenocarcinoma. H. hepaticus infection in mice constitutes the only other parallel association between a persistent bacterial infection and tumor development known to exist naturally. Study of the H. hepaticus syndrome of chronic active hepatitis and liver tumors in mice may yield insights into the role of H. pylori in human stomach cancer and gastric lymphoma.

Infection of rats with human T-cell leukemia virus type-I: susceptibility of inbred strains, antibody response and provirus location.

The susceptibilities of different strains of inbred rats to infection with the human T-cell leukemia virus (HTLV-I) after inoculation of human HTLV-I producer cell lines were compared. The Fisher F344 and Brown Norway strains developed the highest antibody response to HTLV-I, while the Lewis and BB strains were low responders. Antibodies against the HTLV-I gag proteins, and env gp21 but not env gp46, were detected in Western blots with sera from HTLV-I-infected Fischer F344 and Brown Norway rats. These sera were inactive in an in vitro syncytium-formation inhibition test. The HTLV-I provirus was detected by polymerase chain reaction in all Fischer F344, and some Lewis and Brown Norway rats, but not in the BB, which lack CD8+ T lymphocytes. The most frequent locations of the HTLV-I provirus in the Fischer F344, Lewis and Brown Norway rats at 12 weeks after infection were the peripheral blood mononuclear cells (PBMC) and spinal cord. In a second experiment in Brown Norway rats, the provirus was again detected in the PBMC of rats at 12 weeks, but not at 22 weeks, and among the other organs tested at 22 weeks the sympathetic nerve ganglia were positive. It is concluded that HTLV-I infection occurs in adult rats, but is suppressed with time.

Rederivation of rat colonies seropositive for Bacillus piliformis and the subsequent screening for antibodies.

Latent infection of rats in a breeding colony with Bacillus piliformis detectable by antibodies to the agent in an immunofluorescence assay was eliminated by a combination of traditional rederivation techniques, using animal units not previously used for rat breeding, and the use of specific disinfection procedures. The success rate was apparently correlated with the use of peracetic acid instead of aldehyde products to decontaminate the animal unit.

Pulmonary clearance of Mycoplasma pulmonis in rats with respiratory viral infections or of susceptible genotype.

We sought to determine whether or not increased severity of bronchopulmonary disease due to Mycoplasma pulmonis infection in rats with respiratory viral infections and in rats of susceptible genotype could result from altered pulmonary clearance. Pathogen-free rats were exposed to aerosols of radiolabeled M. pulmonis and the numbers of M. pulmonis colony-forming units, and amounts of radiolabel in the lungs were determined immediately after exposure or 4 hours later. Intrapulmonary killing of M. pulmonis during the 4-hour interval was determined from decreases in ratios of colony-forming units to radiolabel, and physical clearance was determined from decreases in radiolabel. Neither intrapulmonary killing nor physical clearance differed between control F344 rats and F344 rats inoculated with Sendai virus or sialodacryoadenitis virus, or between F344 and LEW rats. Rates of intrapulmonary killing and physical clearance were 64 +/- 3% and 44 +/- 2%, respectively (overall means +/- standard error).

[The establishment efficiency of S. sobrinus OMZ 176 in a model caries experiment on the Osborne-Mendel rat]. / Etablierungseffizienz von S. sobrinus OMZ 176 im Kariesmodellversuch an der Osborne-Mendel Ratte.

S. sobrinus OMZ 176 was inoculated in young OM rats during tooth eruption in two experiments and the microbial composition of dental plaque as well as the caries decay was estimated after 25 days on cariogenic sucrose rich diet. Germs were reproducibly established in the oral cavity by such way. The resulting caries decay correlated with the streptococci in the dental plaque. Due to this fact it seems to be necessary to identify not only the inoculated microorganisms at the end of each animal experiment but also to estimate the qualitative composition of the whole dental flora which may have an essential cariogenic effect.

Susceptibility of laboratory and wild rodents to Rattus or Apodemus-type hantaviruses.

Adult Wistar rats (Rattus norvegicus), Apodemus agrarius, Meriones unguiculatus, Clethrionomys rufocanus, and Apodemus argenteus were inoculated with Rattus-type (SR-11, KI-262, and TB-314) or Apodemus-type (Hantaan 76-118) hantaviruses. Production of serum antibody to the inoculated virus (IAHA titres of 1:32 to 1:4 096) was obsersved in all rodent species 10 weeks after virus inoculation. Rattus-type virus was detected in some organs of all the rodent species employed except of Apodemus agrarius. Apodemus-type virus was found only in some organs of Apodemus agrarius. Newborn Wistar rats induced antibody in high titres to both Rattus- and Apodemus-type hantaviruses. Rattus-type virus was detected in all the organs examined for up to 6 weeks after inoculation, whereas Apodemus-type virus disappeared from all organs except of brain and lung tissues. The virulence of the three Rattus-type viruses to newborn rats was different. These findings indicate that the susceptibility of rodents may vary depending on the combination of rodent species and virus strains; they also suggest that the various species of rodents may be the reservoir animals of hantavirus infection in nature.

Estudio de la composición microbiógica de la placa dental de ratas Spragley Dawley como modelo experimental / Study of the microbiologic composition of the dental plaque in Spragley Dawley rats as experimental models

En este estudio se analizó la composición microbiológica de la placa dental de 12 ratas machos de la raza Spragley-Dawley de 28 días de nacidas. La observación microscópica de las bacterias presentes en cada una de las 9 colonias demostró morfología sugerente de cocos en 6 de los casos de Diplococos en 1, de bacilos en 2. Todas las bacterias fueron grampositivas a excepción de las que presentaban morfología de bacilos y Dilpococos en cuyo caso resultaron gramnegativas. La morfología observada para las bacterias de las diferentes colonias es sugerente de Actinomyces en el caso de bacilos grampositivos, de Fusobacterias en el caso de bacilos gramnegativos, de Neisseria y Veillonela en el caso de cocos gramnegativos y de diferentes especies de Estreptococos en las colonias restantes. Se realizaron pruebas de caracterización bioquímica sólo para Estreptococos mutans. Asimismo, se pudo comprobar ausencia de este microorganismo y una baja relación acidogénica en la placa dental de esta raza de animales

Differences in susceptibility to peroral inoculation with Bacillus piliformis spores in rats and mice.

The median liver lesion producing doses of peroral inoculation with the spores of Tyzzer's organism RJ strain were 10(4. 3) in rats and 10(2. 7) in rats receiving prednisolone treatment for the provocation of Tyzzer's disease. In contrast to rats, liver lesions were detected in few mice inoculated perorally with 10(7) spores. In mice inoculated perorally with 10(7) spores, excretion of infective spores in the feces was detected only on day 1 postinoculation. On the other hand, no difference in susceptibility between rats and mice was detected upon intravenous inoculation with vegetative cells of the RJ strain. These results suggest that germination of the spores in the intestinal tract causes the difference in the susceptibility in rats and mice.

Duration of protection from reinfection following exposure to sialodacryoadenitis virus in Wistar rats.

Wistar rats [Cr1:(WI)BR] were inoculated intranasally with approximately 10(3) median mouse lethal infective doses of sialodacryoadenitis (SDA) virus. Animals were subsequently selected at random, removed to a separate isolation room, and reinfected with SDA virus at 3, 6, 9, 12 or 15 months. Pre- and postinoculation serum samples were collected from all animals during the course of the study and evaluated for antibody titers to SDA virus. All experimental, control and sentinel animals, following inoculation with SDA virus, were necropsied and examined for lesions consistent with SDA. Salivary gland lesions were minimal to absent in rats reinfected with SDA virus for up to 12 to 15 months after the initial exposure and minimal to moderate in the respiratory tract at 12 or 15 months. SDA-associated lesions were extensive in age matched control animals examined at each time period of reinfection with SDA virus. Thus, prior exposure to SDA virus did protect against the development of typical salivary gland lesions for up to 15 months. Recovered animals were evaluated for their ability to transmit the virus following reinfection. Rats reinfected at 6 or 9 months were infectious to their naive cage mates. The results indicate that reinfection with homologous rat coronavirus can occur as early as 6 months after the initial infection, and such rats can transmit the infection to contact controls.

Isolation of Mycoplasma moatsii from the intestine of wild Norway rats (Rattus norvegicus).

The intestinal tracts of twenty inbred SPF rats (LEW, BN, WKY, DA) and six wild Norway rats (Rattus norvegicus Berkenhout 1769) were investigated for mycoplasmas. Cultivation was in three different media. Mycoplasmas were not isolated from the intestine of the inbred SPF rats but were found in the epithelia and contents of caecum, colon and jejunum as well as in fecal samples of all of the wild Norway rats investigated. The mycoplasmas isolated all belonged to the same species, and were identified as Mycoplasma moatsii, originally isolated from grivit monkeys (Cercopithecus aethiops) and thought to be specific for this host.

[Leukocyte agglutination test for the determination of microbial burden]. / Leukozyten-Agglomerationstest zur Feststellung mikrobieller Belastung.

Transfer of laboratory animals from the breeder to the experimental unit includes in most cases a change of the microbial environment even under SPF-conditions. Many experimental treatments may also disturb the balance between host and microbial load and provoke infections. With the aim to detect such undesired effects in the course of the experiment the suitability of the leukergy test was investigated. SPF-rats, endotoxin-treated rats, and conventionalized germ-free rats were checked for leukocyte agglomeration, and the findings were compared with total leukocyte number, blood picture, endotoxin content in the blood and body temperature. A rise of leukergy from 10 to more than 20% was recorded in endotoxin-treated and conventionalized rats.

Experimental induction of arthritis in LEW rats and antibody response to four Mycoplasma arthritidis strains.

Four Mycoplasma arthritidis strains were examined for differences in virulence for LEW rats and elicitation of antibody responses in the immunoglobulin (Ig) M and G classes and in the four IgG subclasses. Two strains were highly arthritogenic and two were relatively avirulent. When the latter strains did induce arthritis, it was significantly less severe (P less than 0.05) and developed significantly later (P less than 0.001) than in rats injected with the two virulent strains, suggesting that the low-virulence organisms are able to persist asymptomatically in rats for several weeks. None of the M. arthritidis-injected rats developed metabolism-inhibiting (MI) antibodies at any time during the 6-week observation period. Responses to other M. arthritidis antigens from all four strains were measured by enzyme immunoassay (ELISA); they were similar qualitatively but differed quantitatively. Rats injected with the two avirulent strains showed significantly lower titers of IgM antibodies (P less than 0.01) throughout the 6-week observation period and significantly lower early titers of IgG antibodies (P less than 0.05) than rats injected with the two virulent strains. In addition, peak IgM antibody titers, IgM titers measured 1 and 6 weeks after injection and IgG antibody titers measured 1 week after injection all correlated significantly with peak arthritis scores (P less than 0.05). The IgG antibody response against all four strains appeared mostly in the IgG2a and IgG2b fractions, with very little in the IgG1 and IgG2c fractions. Using immunoblotting, the immunodominant antigens of the two virulent strains appeared very similar, but the avirulent strains differed slightly from each other and from the other two. This study indicates that immune responses of rats to virulent and avirulent strains are similar but not identical and that immunogenicity for LEW rats may be a strain-specific characteristic for M. arthritidis.

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.