In vitro inhibition of mouse dental development by tetracycline

Embryonic molars and incisors were dissected from mandibles of 15-day post-fertilization C57BL/10 mouse embryos and were cultured in vitro for six days on agar-solidified Eagle's basal medium. Experimental explants were cultured on medium which was the same as the control except that 50, 75 or 100 microgram/ml tetracycline was added. Treated explants of both incisors and molars were suppressed in development and reduced in size. Enamel organs and dental papillae of all tooth germs subjected to higher tetracycline concentrations were abnormal in structure and differentiation of ameloblasts and odontoblasts was inhibited. Explants treated with higher dosage levels of the drug were more severely affected than those exposed to lower concentrations. Recovery from the suppression induced by tetracycline was observed in explants transferred to control medium for four days of growth following treatment. Differentiated ameloblasts and odontoblasts observed in the recovering tooth germs indicated that the inhibition in development was temporary. The results of this study showed that tetracycline can alter dental development in vitro prior to mineralization. The observed inhibition may be related to a disruption of collagen biosynthesis which is thought to play a role in the controlling epithelial-mesenchymal interaction involved in tooth germ morphogenesis.     

Quantitative proteome profiling of C. burnetii under tetracycline stress conditions

The recommended antibiotic regimen against Coxiella burnetii, the etiological agent of Q fever, is based on a semi-synthetic, second-generation tetracycline, doxycycline. Here, we report on the comparison of the proteomes of a C. burnetii reference strain either cultured under control conditions or under tetracycline stress conditions. Using the MS-driven combined fractional diagonal chromatography proteomics technique, out of the 531 proteins identified, 5 and 19 proteins were found significantly up- and down-regulated respectively, under tetracycline stress. Although the predicted cellular functions of these regulated proteins did not point to known tetracycline resistance mechanisms, our data clearly reveal the plasticity of the proteome of C. burnetii to battle tetracycline stress. Finally, we raise several plausible hypotheses that could further lead to more focused experiments on studying tetracycline resistance in C. burnetii and thus reduced treatment failures of Q fever.     

Specificity and mechanism of tetracycline resistance in a multiple drug resistant strain of Escherichia coli

Izaki, Kazuo (University of Tokyo, Tokyo, Japan), Kan Kiuchi, and Kei Arima. Specificity and mechanism of tetracycline resistance in a multiple drug resistant strain of Escherichia coli. J. Bacteriol. 91:628-633. 1966.-A decrease in the uptake of tetracycline occurred concurrently with a rise in the level of resistance of a multiple drug resistant strain of Escherichia coli grown in the presence of tetracycline. Although the strain was also resistant to streptomycin and chloramphenicol, growth in the presence of these two antibiotics did not influence the uptake of tetracycline. The induction of resistance, or decreased uptake of tetracycline, was dependent on growth of the organism in the presence of the drug. Decreased uptake of tetracycline could not be induced in a sensitive strain of the same organism under conditions suitable for induction of the resistant strain. The decrease in accumulating power of the resistant organism cultured in the presence of tetracycline does not appear to be due to selection of a resistant strain from cultures containing both resistant and sensitive strains.     

Detection of R plasmids in naturally occurring fish-pathogenic bacteria, Edwardsiella tarda.

The conjugative R plasmids were detected from fish-pathogenic Edwardsiella tarda isolated from cultured eels. These R plasmids were resistant to sulfonamide and tetracycline, or sulfonamide, streptomycin, chloramphenicol, tetracycline and kanamycin. There were no inhibitions in growth of phages W-31, P1, T1, T3, T7, and lambda. They were classified into incompatibility group A.     

Resistance of Escherichia coli to tetracyclines

1. A strain of Escherichia coli highly resistant to chlortetracycline and partially cross-resistant to tetracycline has been isolated. 2. The nitro-reductase system of the resistant cells was inhibited to a smaller extent by chlortetracycline than was the corresponding enzyme of sensitive cells. 3. The incorporation of leucine in vitro into the ribosomal protein of cell-free preparations from sensitive and resistant cells was equally inhibited by chlortetracycline. 4. Resistant cells accumulated much less chlortetracycline and tetracycline than did sensitive cells when both were cultured in the presence of these drugs. 5. The uptake of tetracycline by both sensitive and resistant E. coli was dependent on the presence of glucose in the medium. 6. Fractionation of cells cultured in medium containing [¹⁴C]chlortetracycline indicated that the largest proportion of radioactivity in sensitive cells was in the fraction consisting mainly of cell-wall material. There was no concentration of radioactivity in any one fraction of the resistant cells. 7. No evidence could be obtained for a specific tetracycline-excretion system in the resistant cells. 8. The significance of these results in relation to current theories of the antibiotic action of and resistance to the tetracycline drugs is discussed.     

Effect of osmotic shock on tetracycline resistance in Escherichia coli bearing an R-factor

1. Escherichia coli with an R-factor conferring resistance to tetracycline was induced to high-degree resistance by pre-exposure to the antibiotic. The degree of resistance was drastically lowered by subjecting the cells to osmotic shock. 2. Resistance to tetracycline was rapidly restored by incubating the shocked cells in a glucose-salts medium containing shock proteins prepared from tetracycline-sensitive and -resistant cells. Resistance was also restored by incubating the cells in a complex medium without shock protein. 3. The initial recovery of resistance was followed by a secondary fall in resistance when the cells were cultured in complex medium; this secondary fall was largely prevented by the addition of a low concentration (10mug/ml) of tetracycline to cells. The secondary fall was significantly less in shocked E. coli cells harbouring a mutant R-factor in which tetracycline resistance is largely constitutive. 4. Tetracycline resistance was also transiently depressed by treating R-factor-bearing cells with EDTA in tris buffer. 5. The significance of these results in relation to the mechanism of tetracycline resistance in R-factor-bearing cells is discussed.     

牙菌斑培养菌群宏基因组文库构建及抗生素耐药基因筛选

[目的]构建牙菌斑培养菌群宏基因组文库,筛选牙菌斑生物膜中细菌的抗生素耐药基因.[方法]采集20例无龋健康人的集合牙菌斑并进行厌氧培养.提取牙菌斑培养菌群宏基因组构建Fosmid文库.用卡那霉素、四环素及氨苄西林对文库进行筛选,并对筛选到的抗性Fosmid克隆进行末端测序、亚克隆构建、亚克隆测序和序列分析.[结果]构建了牙菌斑培养菌群宏基因组Fosmid文库,插入片段长度在36-48 kb间约有15 120个克隆,插入片段长度小于36 kb的约有3 360个克隆.筛选获得一个氨基糖苷类双功能修饰酶AacA-AphD基因、一个核糖体保护蛋白型四环素耐药基因tet (M)及一个C家族β-内酰胺酶基因.[结论]证实了可以通过构建宏基因组文库的方法来筛选牙菌斑培养菌群中的抗生素耐药基因.     

Positive influence of tetracycline on human fetal kidney in serum-free organ culture

Summary Human fetal kidney explants can be maintained during 5 days in Leibovitz’s L15, a basic serum-free medium. Because culture conditions are minimal for growth and differentiation, DNA synthesis drastically decreases during the first 48 h, but stabilizes thereafter. The addition of insulin plus transferrin significantly restores this important cellular function in kidneys of fetuses younger than 16 wk. However, renal explants from older fetuses are more difficult to culture: they respond less to growth factors and are more prone to necrosis. The objective of this study was to verify the influence of tetracycline, an antibiotic with anti-collagenase potential, on cultured kidney explants aged 17 to 20 wk. The addition of 20μg/ml tetracycline did not influence DNA synthesis nor the effectiveness of insulin plus transferrin on cell proliferation. Nor did it change the activities of alkaline phosphatase and γ-glutamyltransferase, two enzymic markers of brush border differentiation. After 5 days in L15 alone, explants often showed necrosis and an important reduction in both weight and volume. Insulin plus transferrin significantly restored these parameters to control values observed at Day 0, but evidence of necrosis was still present. Tetracycline alone markedly reduced explant necrosis resulting in a significant increase in weight and volume. The effectiveness of insulin plus transferrin on explant morphometry was not improved when tetracycline was added as third factor. These results indicate that insulin plus transferrin restores explant mass through cell proliferation, whereas tetracycline does so possibly through a reduction in extracellular matrix degradation. The two effects are not additive in cultured mid-term fetal kidneys.     

Contribution of gut bacteria to digestion and development of the velvetbean caterpillar, Anticarsia gemmatalis

Bacteria colonies from gut homogenates of fifth instar velvetbean caterpillars (Lepidoptera: Noctuidae) were subjected to antibiotic sensitivity experiments using discs containing 22 antibiotics. The antibiotic tetracycline provided the best results, followed by chloramphenicol. Tetracycline also provided higher inhibition of colony forming units than chloramphenicol and was therefore provided to the caterpillars in increasing diet concentrations to assess the contribution of gut bacteria to their digestion and development. The activity of proteases (general), serine-proteinases and lipases were significantly suppressed by tetracycline. Concentration-inhibition curves were successfully established for tetracycline and this antibiotic was effective in suppressing them, particularly serine-proteinases, suggesting that gut bacteria may significantly contribute with lipid- and mainly protein-digestion in velvetbean caterpillars. Increased diet concentrations of tetracycline led only to mild increase in insect mortality (ca. 20%), with the surviving insects showing faster development (≤4 days) and higher pupa weight (<0.04 mg) with increased concentrations of tetracycline. Therefore, the gut bacteria inhibited by tetracycline does not seem to play a crucial role in the survival and development of the velvetbean caterpillar, but may be important in the adaptation of this pest species to hosts rich in protease inhibitors, such as soybean.     

Effects of tetracycline on the filarial worms Brugia pahangi and Dirofilaria immitis and their bacterial endosymbionts Wolbachia

Wolbachia endosymbiotic bacteria have been shown to be widespread among filarial worms and could thus play some role in the biology of these nematodes. Indeed, tetracycline has been shown to inhibit both the development of adult worms from third-stage larvae and the development of the microfilaraemia in jirds infected with Brugia pahangi. The possibility that these effects are related to the bacteriostatic activity of tetracycline on Wolbachia symbionts should be considered. Here we show that tetracycline treatment is very effective in blocking embryo development in two filarial nematodes, B. pahangi and Dirofilaria immitis. Embryo degeneration was documented by TEM, while the inhibition of the transovarial transmission of Wolbachia was documented by PCR. Phylogenetic analysis on the ssrDNA sequence of the Wolbachia of B. pahangi confirms that the phylogeny of the bacterial endosymbionts is consistent with that of the host worms. The possibility that tetracycline inhibition of embryo development in B. pahangi and D. immitis is determined by cytoplasmic incompatibility is discussed.     

Transmission of Salmonella enterica serotype Typhimurium in poultry with and without antimicrobial selective pressure

AIMS: To determine the effect of antimicrobial selective pressure on the transmission of antimicrobial resistant and sensitive strains of Salmonella in poultry. METHODS AND RESULTS: Eight pens housed 12 broiler chicks each. Two chicks in four of the pens were inoculated with a Salm. Typhimurium strain resistant to 12 antimicrobials (including tetracycline), and two chicks in each of the four other pens were inoculated with a strain sensitive to all antimicrobials tested. Two pens inoculated with each strain were treated with chlortetracycline and two were not. Chicks were killed on day 7 and caeca were cultured for Salmonella. Experiments were performed independently twice. Chicks exposed to pen mates inoculated with the resistant strain and treated with tetracycline were 90% positive for Salmonella; whereas 60% of chicks given no antimicrobials were positive. Chicks exposed to the sensitive strain were 95% positive with tetracycline treatment and 90% positive without treatment. CONCLUSIONS: A multidrug-resistant Salm. Typhimurium strain had significantly increased transmission when chicks were treated with tetracycline. Transmission of a sensitive strain was not inhibited by antimicrobial selective pressure at recommended therapeutic dose. SIGNIFICANCE AND IMPACT OF THE STUDY: This study demonstrates that antimicrobial usage may influence the transmission of antimicrobial-resistant pathogens in poultry.     

Makeup and properties of E. coli cells with a varying level of resistance to tetracycline]

Lipopolysaccharide composition of tetracycline sensitive and resistant strains of E. coli was studied comparatively. It was shown that that resistance of E. coli to tetracycline was probably due to the differences in the lipopolysaccharide component composition of the outer membrane. On the basis of the activity comparison of the Mg2+- and Ca2+-activated ATP-ase of the membrane fraction of the tetracycline sensitive and resistance strains of E. coli it was concluded that the resistance development in the strains tested to tetracycline was not associated with the changes in the ATP-ase activity.     

Drug Resistance of Staphylococci

It was reported that resistance to tetracycline (TC) in some staphylococcal strains was lost irreversibly without loss of resistance to other drugs when cultured at high temperature, and that the determinant for TC resistance exists on a plasmid. According to the transductional analysis of TC resistance in Staphylococcus aureus E169 it was found that the genes which govern resistance to tetracycline and streptomycin are located close together on a single genetic element. When TC resistance in MS146, one of our stock cultures, was transduced to E169S which had lost TC resistance, TC resistance in transductants was found to be unstable at elevated temperatures. By contrast, TC resistance in transductants MS353 TC^r, to which TC resistance was transduced from E169, was indicated to be stable even when cultured at high temperature. From these results, it is strongly suggested that instability of TC resistance in E169 is not accounted for by the genetic properties of the genes which govern TC resistance but those of host cell (E169) itself.     

Selection of tetracycline inducible self-cleaving ribozymes as synthetic devices for gene regulation in yeast

Synthetic regulatory devices are key components for the development of complex biological systems and the reprogramming of cellular functions and networks. Here we describe the selection of tetracycline inducible hammerhead ribozymes. A tetracycline aptamer was fused to the full-length hammerhead ribozyme via a variable linker region. 11 rounds of in vitro selection were applied to isolate linker sequences that mediate tetracycline dependent hammerhead cleavage. We identified allosteric ribozymes that cleave in the presence of 1 μM tetracycline as fast as the full-length ribozyme whereas cleavage is inhibited up to 333-fold in the absence of tetracycline. Reporter gene assays indicate that the allosteric ribozymes can be employed to control gene expression in yeast.     

Uncultured soil bacteria are a reservoir of new antibiotic resistance genes

Antibiotic resistance genes are typically isolated by cloning from cultured bacteria or by polymerase chain reaction (PCR) amplification from environmental samples. These methods do not access the potential reservoir of undiscovered antibiotic resistance genes harboured by soil bacteria because most soil bacteria are not cultured readily, and PCR detection of antibiotic resistance genes depends on primers that are based on known genes. To explore this reservoir, we isolated DNA directly from soil samples, cloned the DNA and selected for clones that expressed antibiotic resistance in Escherichia coli. We constructed four libraries that collectively contain 4.1 gigabases of cloned soil DNA. From these and two previously reported libraries, we identified nine clones expressing resistance to aminoglycoside antibiotics and one expressing tetracycline resistance. Based on the predicted amino acid sequences of the resistance genes, the resistance mechanisms include efflux of tetracycline and inactivation of aminoglycoside antibiotics by phosphorylation and acetylation. With one exception, all the sequences are considerably different from previously reported sequences. The results indicate that soil bacteria are a reservoir of antibiotic resistance genes with greater genetic diversity than previously accounted for, and that the diversity can be surveyed by a culture-independent method.     

两株四环素降解菌的分离鉴定及降解条件优化

为发掘四环素高效降解菌株,本研究从以四环素为唯一生长碳源的养鸡场粪便堆肥样品中分离筛选到2株四环素降解菌TC-04和TC-09,并通过形态特征、生理生化特征和16SrRNA基因序列分析,对其进行鉴定.采用单因素试验分别探究不同碳源、氮源、微量元素这3个培养基成分和培养时间、接种量和装液量这3个培养条件对两株菌降解四环素...     

Development of Resistance to Novobiocin, Tetracycline, and a Novobiocin-Tetracycline Combination in Staphylococcus aureus Populations

The antibiotic sensitivity of the individual organisms of a bacterial population was determined to study the comparative rates of development of resistance of Staphylococcus aureus to novobiocin, tetracycline, and to a combination of these antibiotics. Serial subculture of S. aureus with the combination of novobiocin-tetracycline (N-T 2.5:1; the ratio in serum of patients dosed with Panalba) showed a significant retardation of resistance outgrowth compared with subculture in the presence of the antibiotics individually. Increase in organisms resistant to novobiocin seen after one N-T subculture was related to the “concentration gap” between novobiocin and tetracycline. Two additional subcultures with N-T caused little or no increase in organisms resistant to novobiocin, tetracycline, or to the combination. The data suggest that the retardation of further development of resistance was the result of tetracycline inhibition of novobiocin-resistant strains and vice versa.