Escherichia coli O78 isolated from septicemic lambs shows high pathogenicity in a zebrafish model.

The pathogenicity of Escherichia coli O78 strain K46, originally isolated from an outbreak of septicemia in neonatal lambs, was investigated in zebrafish embryo and murine models of infection. Its biofilm potential, cellulose production, and the expression of type 1 pili and curli fimbriae were measured by in vitro assays. The strain was highly pathogenic in the zebrafish embryo model of infection, where it killed all embryos within 24 h post inoculation (hpi) at doses as low as 1000 colony forming units. Zebrafish embryos inoculated with similar doses of commensal E. coli strains showed no signs of disease, and cleared the bacteria within 24 h. E. coli K46 colonized the murine gut at the same level as the uropathogenic E. coli (UPEC) reference strain CFT073 in CBA/J mice after oral inoculation, but infected the murine bladder significantly less than CFT073 after transurethral inoculation. Type 1 pili were clearly expressed by E. coli K46, while curli fimbriae and cellulose production were weakly expressed. The ability to produce biofilm varied in different growth media, but overall E. coli K46 was a poorer biofilm producer compared to the reference strain E. coli UTI89. In conclusion, the zebrafish lethality model provides further evidence that E. coli K46 is highly pathogenic and might be useful in future studies to identify bacterial virulence factors.

A phyletically rare gene promotes the niche-specific fitness of an E. coli pathogen during bacteremia.

In bacteria, laterally acquired genes are often concentrated within chromosomal regions known as genomic islands. Using a recently developed zebrafish infection model, we set out to identify unique factors encoded within genomic islands that contribute to the fitness and virulence of a reference urosepsis isolate-extraintestinal pathogenic Escherichia coli strain CFT073. By screening a series of deletion mutants, we discovered a previously uncharacterized gene, neaT, that is conditionally required by the pathogen during systemic infections. In vitro assays indicate that neaT can limit bacterial interactions with host phagocytes and alter the aggregative properties of CFT073. The neaT gene is localized within an integrated P2-like bacteriophage in CFT073, but was rarely found within other proteobacterial genomes. Sequence-based analyses revealed that neaT homologues are present, but discordantly conserved, within a phyletically diverse set of bacterial species. In CFT073, neaT appears to be unameliorated, having an exceptionally A+T-rich composition along with a notably altered codon bias. These data suggest that neaT was recently brought into the proteobacterial pan-genome from an extra-phyletic source. Interestingly, even in G+C-poor genomes, as found within the Firmicutes lineage, neaT-like genes are often unameliorated. Sequence-level features of neaT homologues challenge the common supposition that the A+T-rich nature of many recently acquired genes reflects the nucleotide composition of their genomes of origin. In total, these findings highlight the complexity of the evolutionary forces that can affect the acquisition, utilization, and assimilation of rare genes that promote the niche-dependent fitness and virulence of a bacterial pathogen.

[The evaluation of the degree of attenuation of cold-adapted influenza A virus strains in CBA-strain mouse and Syrian hamster models]. / Otsenka stepeni attenuatsii kholodo-adaptirovannykh shtammov virusa grippa A na modeliakh myshei linii CBA i siriiskikh khomiachkov.

The pattern of the infectious process induced by the epidemic A/Leningrad/134/57 (H2N2) virus and its cold-adapted (CA) variants in CBA mice and Syrian hamsters was studied. The strains under study inoculated into the animals under a mild ether anesthesia differed by virulence, reproductive capacity in the nasopharynx, trachea and lungs, as well as by the isolation rate from extrarespiratory organs of both mice and hamsters. Upon intranasal inoculation of mice without anesthesia, the CA strains were found to be incapable of dissemination into the lower parts of the respiratory tract with distinguished these viruses from the original epidemic strain A/Leningrad/134/57 as well as from the mouse-adapted strain A/PR/8/34 (H1N1) used as control. The experimental results show that both models are suitable for laboratory evaluation of the attenuation degree of human influenza viruses.

Use of cycloheximide on intracellular growth of Mycobacterium leprae in cultured murine macrophages.

Mycobacterium leprae is an obligate intracellular parasite and grows within mononuclear phagocytes. When murine macrophages derived from the peritoneal cavities of CBA mice were infected in vitro with M. leprae (Thai-53 strain), intracellular multiplication was observed three weeks after infection. On the other hand, there was no increase in the number of heat-killed M. leprae at the same times after infection. Morphological studies showed that the growth rate of the bacteria increased by about 20-30% in medium supplemented with cycloheximide. With the addition of cycloheximide to the culture medium, metabolic activity of macrophages was decreased but infected cells were maintained in good condition and seldom floated off from the culture flask.

Intracellular fate of Mycobacterium leprae in cultured murine macrophages in 24-well trays.

Mycobacterium leprae is an obligate intracellular parasite which grows within mononuclear phagocytes. In clinical cases, M. leprae reaches enormous numbers in the macrophage-rich granulomas of leprosy. Peritoneal macrophages from CBA mice were cultured in Waymouth medium containing fresh horse serum in Costar 3424 trays (24 wells, 16 mm in diameter) each containing 9 x 12 mm cover slips. This medium was supplemented with 0.5 micrograms/ml of cycloheximide. These cells were infected with M. leprae Thai-53 strain obtained by nude mice inoculation. Significant multiplication of the acid-fast bacilli in the macrophages was observed three weeks after inoculation. This experiment showed M. leprae mainly multiplied in cells and not by rephagocytization of M. leprae derived from destroyed cells.

The cutaneous microbiology of haired and hairless mice.

The cutaneous microflora of the mid-dorsal area of hairless and haired mice was studied by processing skin biopsies. In both C3H and CBA hairless genotype animals the prevalence of colonization and the bacterial density were significantly greater than in the haired animals. The dominant bacteria were staphylococci and aerobic coryneforms. No propionibacteria were isolated. Temporal studies with C3H mice showed that from 0 to 9 days after birth the cutaneous microflora reduced and from then on the haired genotype animals maintained a low cutaneous microflora, whilst hairless genotype animals gradually lost hair from head to tail and the microflora density increased. Reciprocal skin grafting between haired and hairless animals showed that the donor skin acquired the microflora characteristics of the recipient animal after 15 d post-grafting even though the donor skin remained morphologically true to genotype.

Doubling time of Mycobacterium lepraemurium in mouse footpads.

The doubling time of Mycobacterium lepraemurium (MLM) was measured in CBA/Ca mice. In eight experiments, 5 X 10(3) MLM were inoculated into the hind footpads of groups of mice, and the organisms were harvested from 1-45 days later. The harvested organisms were enumerated, and the doubling time was calculated assuming that MLM had multiplied without a lag phase and that multiplication continued at a constant rate from inoculation to harvest. Simultaneously, the proportions of viable organisms in the inocula were determined by inoculation of serially diluted suspensions into the footpads of other mice, harvesting 4 months later and calculating the most probable number. MLM were observed to multiply rapidly during the first several days, and more slowly thereafter; the mean initial doubling time was determined to be 0.5 days, a value much smaller than those previously reported by other workers.

Observations on the occurrence of mycoplasmas in the central nervous system of some laboratory animals.

Mycoplasma pulmonis was isolated from the brains of 6 (23%) of 26 mice which had a naturally-occurring respiratory infection with this mycoplasma, and from the brains of 6 (8%) of 71 mice which had been inoculated intranasally or intravenously. The incidence of natural infection was greater in older mice, but there was no obvious mouse strain difference except for higher incidence in athymic nudes. There was no evidence that the organisms passed the blood-brain barrier. Some isolations, especially from nudes, may have been extraneous contaminants, as these were fewer when the mouse skulls were sterilized with ignited methanol. M. pneumoniae was not isolated from the brains of 14 hamsters which had a respiratory infection after intranasal inoculation nor were ureaplasmas isolated from the cerebrospinal fluids of 12 marmosets with a natural oropharyngeal infection. The aetiology of M. pneumoniae encephalitis in man is discussed.

[Biological properties of antibiotic-resistant strains of lactic acid bacteria]. / Izuchenie biologicheskikh svoistv antibiotikoustoichivykh shtammov molochnikislykh bakterii.

Lactic-acid bacteria (L. fermenti, L. acidophilus, L. delbruecki), when developing resistance to antibiotics, did not change their main biochemical, antagonistic properties and did not lose capacity for acid production. Only a decrease in their growth rate and a change in their sensitivity to the action of ultraviolet radiation were observed. Both initial and antibiotic-resistant strains were capable of taking on the mucous membrane of the large and small intestines in CBA mice.