Detection of Corynebacterium kutscheri in the faeces of subclinically infected mice.

Mice were infected experimentally and subclinically with Corynebacterium kutscheri to recover the organism from mice faeces. The faeces were then cultured using selective furazolidone-nalidixic acid-colimycin agar. The number of C. kutscheri per gram of fresh faeces varied from mouse to mouse, but once established in the intestine, the organism was excreted in the faeces for at least five months. Viable bacteria were detected in most of the faecal samples, including those stored in the animal room for five days. The number of organisms in the stored faeces decreased gradually but did not differ significantly from those in the fresh faeces until they had been stored for more than three days. Many infected mice excreted between 10(4.77) and 10(5.37) colony forming units (CFU) of C. kutscheri per day in their faeces, and one mouse even excreted 10(3.74) CFU at eight weeks postinfection. These values showed little daily variation. Our present study showed that subclinically infected mice discharged the organism continuously and persistently in their faeces. Therefore, faecal samples would be useful for monitoring infection with C. kutscheri in living mice in a manner that is not stressful for the animals.

Isolation of a thermophile degrading poly(butylene succinate-co-butylene adipate).

A thermophile, strain 73, which degrades poly(butylene succinate-co-butylene adipate) (PBSA) film was isolated from 95 soil samples obtained from different locations by cultivation using an enrichment culture technique at 60 degrees C. At this temperature, the strain grew on PBSA and the dissolved total organic carbon (TOC) concentration in the medium changed according to the growth stage, i.e., after the TOC concentration rapidly reached the minimum, it increased rapidly until it reached a peak and then decreased thereafter. During cultivation, PBSA was almost eliminated and the solution viscosity of the residual PBSA decreased markedly. Gel permeation chromatograms of the residual PBSA showed a significant decrease in the main peak and the appearance of a new peak at the low molecular weight region. The strain was identified as Bacillus stearothermophilus, which has an optimum growth temperature of approximately 63 degrees C.

Sex differences in susceptibility of ICR mice to oral infection with Corynebacterium kutscheri.

Sex difference in susceptibility to oral infection with Corynebacterium (C.) kutscheri was experimentally studied in ICR mice. Immature (4-week-old) and adult (14-week-old) mice were inoculated with two infecting doses of C. kutscheri, and necropsied for bacteriological and serological survey 4 weeks after the bacterial infection. No macroscopic lesions at necropsy were demonstrated, except for one adult male given 10(9) bacteria. In immature mice, C. Kutscheri isolated from the oral cavity and cecum with FNC agar, were recovered in only 40.0% of female mice but in 90.0% of male mice given 10(6) bacteria (p < 0.05), and in only 55.6% of female mice but in 80.0% male mice given 10(8) bacteria. In adult mice given 10(9) bacteria, the organism were recovered in only 45.5% of female mice but in 90.9% of male mice (p < 0.05), furthermore, the mean number of organisms in the cecum of male mice harboring the organism was significantly higher than that in females (p < 0.01). Castration caused an increase in host resistance in adult male mice. These results indicated that ICR male mice were more susceptible than females, in terms of bacterial colonization in the cecum and the oral cavity, to oral infection with C. kutscheri.

Conversion of the coenzyme specificity of isocitrate dehydrogenase by module replacement.

The coenzyme specificity of isocitrate dehydrogenase from an extreme thermophilic bacterium was converted from NADP-dependent to NAD-dependent by replacing a "module" involved in the coenzyme binding site. The conversion was not possible with the replacement of a few residues that interact with the coenzyme. In addition, the module-replaced mutant dehydrogenase was as stable as the original, wild type enzyme. The results support a previous hypothesis that a module is a structural and functional unit of a protein.

Pathogenicity of Corynebacterium kutscheri in the Syrian hamster.

The pathogenicity of Corynebacterium kutscheri isolated for the first time from Syrian hamster was experimentally studied in hamsters. In hamsters given intramuscular (i.m.) or subcutaneous (s.c.) inoculation with 10 or 10(3) bacteria, neither clinical signs nor gross lesions were found. In those given 10(5) bacteria i.m., moderate proliferation of granulation tissue was found in the muscle of the inoculation region at necropsy. In the animals given 10(5) bacteria s.c., a nodular lesion was observed at the inoculation site 2 days post-inoculation (p.i.), but the nodules subsided gradually from 6 days p.i. and were unclear 10 days p.i. At necropsy, small abscesses were found in all the animals in this group. In those given 10(7) bacteria either i.m. or s.c., lesions were clearly observed at the inoculation site 1 to 10 days p.i., and a large abscess was noted at necropsy. The organisms were isolated only from the lesions in the groups. Agglutinating antibody in the sera was detected only in the animals given 10(5) or 10(7) bacteria. This suggests that 10(5) of C. kutscheri are needed to form localized nodular abscesses in Syrian hamsters.

Natural and subclinical Corynebacterium kutscheri infection in rats.

Distribution of Corynebacterium kutscheri was determined in 41 rats housed in a conventionally managed colony that were infected naturally and subclinically. At 2, 5, 10, 20 and 25 months after initial isolation of C. kutscheri, attempts were made to isolate C. kutscheri from 17 sites, with a new selective medium, FNC agar. In total, the prevalence (97.6%) of C. kutscheri isolation was significantly (P < 0.001) higher than the frequency (70.7%) of antibody detection. None of the rats manifested any distinct clinical signs of disease and macroscopic lesions caused by C. kutscheri were not detected. In 40 rats with subclinical infection, the organisms were most frequently isolated from the oral cavity, esophagus, cecal contents, and colon and rectum (> 95.0%). The isolation rate was next highest in the trachea, submaxillary lymph nodes, and nasal cavity (47.5 to 52.5%). The organisms hardly colonized the lung, liver, and kidney. Mean numbers of organisms found in the esophagus, cecal contents, and colon and rectum ranged from 10(3.9) to 10(4.2) CFU/g, and were significantly (P < 0.05, P < 0.01) high in comparison with those in the lung. These results indicated that many healthy rats in the naturally infected colony harbored C. kutscheri, and the organisms colonized the oral cavity, esophagus, cecal contents, and colon and rectum most frequently.

Natural habitats of Corynebacterium kutscheri in subclinically infected ICGN and DBA/2 strains of mice.

Subclinically infected mice of ICGN and DBA/2 strains housed in a conventionally managed colony were examined to determine natural habitats of Corynebacterium kutscheri. At 5, 7, 9, 12 and 13 months after initial isolation of the organism from oral cavity and cecal contents of five ICGN mice, attempts were made to isolate C. kutscheri from 19 sites using a new selective medium, furazolidone-nalidixic acid-colimycin agar. From the initial survey to 13 months, C. kutscheri was isolated from 27 of 29 ICGN mice (93.1%) and 9 of 10 DBA/2 mice (90%). In contrast, antibody against C. kutscheri was detected in only 3 of 29 ICGN mice (10.3%). None of the mice manifested distinct clinical signs of infection, and only 1 ICGN mouse had macroscopic lesions such as hepatic abscess and large spleen. In 21 ICGN and 9 DBA/2 mice that harbored the organism without macroscopic lesions, the organisms were most frequently isolated from the oral cavity (ICGN:100%, DBA/2:66.7%), cecum (ICGN:95.2%, DBA/2:100%), and colon and rectum (ICGN:95.2%, DBA/2:100%). Remarkable differences between the two mouse strains were observed in colonization of the nasal cavity (ICGN:85.7%, DBA/2:0%) and trachea (ICGN:71.4%, DBA/2:33.3%). In mice of both strains, the organisms rarely colonized the lung, liver, and kidney. Mean numbers of organisms in the cecum, and colon and rectum ranged from 10(4.1) to 10(4.6) colony-forming units/g and were significantly higher in comparison with those in the small intestine (P < 0.01, P < 0.001).(ABSTRACT TRUNCATED AT 250 WORDS)

[Differences in susceptibility of mice among various strains to oral infection with Corynebacterium kutscheri].

Differences in susceptibility of female mice among 10 strains to Corynebacterium kutscheri infection were studied pathologically and bacteriologically. Twenty mice of each strain were inoculated orally with 4 x 10(5.0) CFU of the bacteria. The gross lesions were observed in 60.0% of BALB/c-nu/nu mice, 25.0% of CBA/N mice, 10.0% of MPS mice, and 5.0% of A/J and C3H/He mice, while BALB/cCr, C57BL/6Cr, B10.BR/SgSn, ddY and ICR mice showed neither clinical signs nor gross lesions. Six BALB/c-nu/nu, two CBA/N and one MPS mice died within 15 days after inoculation. C. kutscheri was recovered from 95.0% of BALB/c-nu/nu mice, followed by 75.0% of A/J mice, 65.0% of CBA/N mice, 55.0% of MPS and BALB/cCr mice, and 30% of C3H/He mice. On the other hand, from C57BL/6Cr, B10.BR/SgSn and ddY mice, the bacteria were recovered at less than 15.0% of the mice. No bacteria were recovered from ICR mice. C. kutscheri colonized most frequently in the cecum, colon and rectum. Number of mice having agglutinating antibodies were less than 20% and their antibody titers ranged from 1:10 to 1:80. These results indicated that there were differences in susceptibility of mice among the strains to oral infection of C. kutscheri. Namely, BALB/c-nu/nu, A/J, CBA/N, MPS and BALB/cCr mice appeared to be susceptible, and C3H/He mice intermediate, while C57BL/6Cr, B10.BR/SgSn, ddY and ICR resistant. BALB/c-nu/nu mice were most susceptible and exhibited markedly severe disease by the infection.

A gene homologous to chloroplast carbonic anhydrase (icfA) is essential to photosynthetic carbon dioxide fixation by Synechococcus PCC7942.

To understand the CO2-concentrating mechanism in cyanobacteria, a genomic DNA fragment that complements a temperature-sensitive high-CO2 (5%)-requiring mutant of Synechococcus PCC7942 has been isolated. An open reading frame (ORF272) encoding a polypeptide of 272 amino acids (Mr, 30,184) was found within the genomic region located 20 kilobases downstream from the genes for ribulose-1,5-bisphosphate carboxylase/oxygenase (rbcLS). Insertion of a kanamycin-resistance gene cartridge within the ORF272 in wild-type cells led to a high-CO2-requiring phenotype. Strains carrying a gene disabled by insertional mutagenesis accumulated inorganic carbon in the cells, but they could not fix it efficiently, even though ribulose-1,5-bisphosphate carboxylase activity was comparable to that of the wild-type strain. Therefore, the ORF272 was designated as a gene icfA, which is essential to inorganic carbon fixation. Furthermore, the predicted icfA gene product shared significant sequence similarities with plant chloroplast carbonic anhydrases (CAs) from pea (22%) and spinach (22%) and also with the Escherichia coli cynT gene product (31%), which was recently identified to be E. coli CA. These results indicate that the putative CA encoded by icfA is essential to photosynthetic carbon dioxide fixation in cyanobacteria and that plant chloroplast CAs may have evolved from a common ancestor of the prokaryotic CAs, which are distinct from mammalian CAs and Chlamydomonas periplasmic CAs.