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PubMed1976

Multiple-aminoglycoside-resistant mutants of Bacillus subtilis deficient in accumulation of kanamycin.

Three classes of spontaneous multiple-aminoglycoside-resistant (mar) mutants of Bacillus subtilis were isolated by plating on a low (1.2 mug/ml) concentration of kanamycin sulfate and were found to be resistant also to low concentrations of paromomycin, neomycin and gentamicin. The three classes could be distinguished one from another by their degree of cytochrome deficiency, respiration deficiency, and susceptibility to kanamycin lethality. A fluctuation test showed that the mutations were spontaneous and not induced by the conditions of selection. Representative strains from two classes of mutants (mar-2 and mar-3) accumulated aminoglycoside very poorly in comparison with the parent strain, whereas a strain of the third class (mar-1) inactivated aminoglycoside present in the growth medium. The mar-3 strain studied (aroD163) had previously been shown to be a menaquinone auxotroph (Farrand and Taber, 1973) and to be deficient in amino acid uptake (Bisschop et al., 1975). Such mutants, which are resistant to low concentrations of aminoglycosides, may be of use in elucidating the biochemical and genetic bases of certain bacterial transport systems.

Aminoglycosides↗
PubMed1967

Antibiotic control of Mycoplasma in tissue culture.

Seven of eight strains of Mycoplasma (PPLO) were found to be sensitive to the deoxystreptamines, certain macrolides, and the tetracyclines. These antibiotics are relative noncytotoxic. Kanamycin and tetracycline were useful in eliminating PPLO (pleuropneumonia-like organisms) strain Squibb no. 1 from a HeLa cell line which was deliberately contaminated with PPLO. Repeated exposure of M. laidlawii type B cells to neomycin resulted in a 50-fold increase in resistance, and the resistant strain was also resistant to gentamicin, kanamycin, neomycin, and paromomycin. A tetracycline-resistant strain of this culture was found to be resistant to 7-chlortetracycline, 7-chlor-6-demethyltetracycline, and 5-hydroxytetracycline. One PPLO strain, Squibb no. 2, derived from a contaminated HeLa cell culture, was resistant to all antibiotics studied.

Anti-Bacterial Agents↗
PubMed1983

Mapping of the gene specifying aminoglycoside 3'-phosphotransferase II on the Pseudomonas aeruginosa chromosome.

We examined the aminoglycoside inactivation enzymes in Pseudomonas aeruginosa strains, seven clinical isolates and seven laboratory strains without plasmids. All strains were found to possess the enzyme aminoglycoside 3'-phosphotransferase II [APH(3')-II]. We isolated an APH(3')-II-deficient mutant from a PAO strain by mutagenesis with N-methyl-N'-nitro-N-nitrosoguanidine. By plasmid (FP5 or R68.45)-mediated conjugation, we determined the locus of the gene specifying the APH(3')-II between trp-6 and pro-82 on the PAO chromosome and designated this gene aphA. It was concluded that the intrinsic resistance of P. aeruginosa to kanamycins, neomycins, paromomycins, ribostamycin, and butirosins was due to this newly determined gene.

Chromosome Mapping↗
PubMed1988

Ribosomes of the extremely thermophilic eubacterium Thermotoga maritima are uniquely insensitive to the miscoding-inducing action of aminoglycoside antibiotics.

Poly(U)- and poly(UG)-programmed cell-free systems were developed from the extreme thermophilic, anaerobic eubacterium Thermotoga maritima, and their susceptibility to aminoglycoside and other antibiotics was assayed at a temperature (75 degrees C) close to the physiological optimum (80 degrees C) for cell growth and in vitro polypeptide synthesis, using a Bacillus stearothermophilus system as the reference. The synthetic capacity of the Thermotoga assay mixture was abolished by the eubacterium-targeted drugs chloramphenicol, thiostrepton, and kirromycin. However, streptomycin, the disubstituted 2-deoxystreptamines (kanamycin, gentamicin, neomycin, and paromomycin), and the monosubstituted 2-deoxystreptamine (hygromycin) all failed to promote translational misreading of poly(U) on Thermotoga ribosomes; they also failed to block polyphenylalanine synthesis at a low (less than 10(-4) M) concentration and did not inhibit Thermotoga cell growth at a high (10 micrograms/ml) concentration even though Thermotoga ribosomes possess the 16S rRNA sequences required for aminoglycoside action. In contrast to the other eubacteria, Thermotoga elongation factor G was also refractory to the steroid inhibitor of peptidyl-tRNA translocation fusidic acid.

Aminoglycosides↗
PubMed1992

Increase of sensitivity to aminoglycoside antibiotics by polyamine-induced protein (oligopeptide-binding protein) in Escherichia coli.

The sensitivity of Escherichia coli to several aminoglycoside antibiotics was examined with E. coli DR112 transformed by the gene for polyamine-induced protein (oligopeptide-binding [OppA] protein) or polyamine transport proteins. The results clearly showed that sensitivity to aminoglycoside antibiotics (gentamicin, isepamicin, kanamycin, neomycin, paromomycin, and streptomycin) increased due to the highly expressed OppA protein. When the gene for OppA protein was deleted, sensitivity to aminoglycoside antibiotics was greatly decreased. It was also shown that isepamicin could bind to OppA protein with a binding affinity constant of 8.5 x 10(3) M-1 under the ionic conditions of 50 mM K+ and 1 mM Mg2+ at pH 7.5, and isepamicin uptake into cells was greatly stimulated by the OppA protein. These results, taken together, show that the OppA protein increases the uptake of aminoglycoside antibiotics. In addition, the OppA protein increased the transport of spermidine and an oligopeptide (Gly-Leu-Tyr). The uptake of isepamicin into cells was partially inhibited by spermidine, suggesting that the binding site for isepamicin overlaps that for spermidine on the OppA protein. Spermidine uptake activity by the OppA protein was less than 1% of that of the ordinary spermidine uptake system. Aminoglycoside antibiotics neither stimulated the synthesis of OppA protein nor increased spermidine uptake.

Amino Acid Sequence↗
PubMed1997

Fal1p is an essential DEAD-box protein involved in 40S-ribosomal-subunit biogenesis in Saccharomyces cerevisiae.

A previously uncharacterized Saccharomyces cerevisiae gene, FAL1, was found by sequence comparison as a homolog of the eukaryotic translation initiation factor 4A (eIF4A). Fal1p has 55% identity and 73% similarity on the amino acid level to yeast eIF4A, the prototype of ATP-dependent RNA helicases of the DEAD-box protein family. Although clearly grouped in the eIF4A subfamily, the essential Fal1p displays a different subcellular function and localization. An HA epitope-tagged Fal1p is localized predominantly in the nucleolus. Polysome analyses in a temperature-sensitive fal1-1 mutant and a Fal1p-depleted strain reveal a decrease in the number of 40S ribosomal subunits. Furthermore, these strains are hypersensitive to the aminoglycoside antibiotics paromomycin and neomycin. Pulse-chase labeling of pre-rRNA and steady-state-level analysis of pre-rRNAs and mature rRNAs by Northern hybridization and primer extension in the Fal1p-depleted strain show that Fal1p is required for pre-rRNA processing at sites A0, A1, and A2. Consequently, depletion of Fal1p leads to decreased 18S rRNA levels and to an overall deficit in 40S ribosomal subunits. Together, these results implicate Fal1p in the 18S rRNA maturation pathway rather than in translation initiation.

Amino Acid Sequence↗
PubMed2003

Novel methyltransferase for modified uridine residues at the wobble position of tRNA.

We have identified a novel tRNA methyltransferase in Saccharomyces cerevisiae that we designate Trm9. This enzyme, the product of the YML014w gene, catalyzes the esterification of modified uridine nucleotides, resulting in the formation of 5-methylcarbonylmethyluridine in tRNA(Arg3) and 5-methylcarbonylmethyl-2-thiouridine in tRNA(Glu). In intact yeast cells, disruption of the TRM9 gene results in the complete loss of these modified wobble bases and increased sensitivity at 37 degrees C to paromomycin, a translational inhibitor. These results suggest a role for this potentially reversible methyl esterification reaction when cells are under stress.

Amino Acid Sequence↗
PubMed1988

Transformation of Tetrahymena thermophila with hypermethylated rRNA genes.

The extrachromosomal rRNA genes (rDNA) of Tetrahymena thermophila contain 0.4% N6-methyladenine. C3 strain rDNA was isolated, hypermethylated in vitro, and microinjected into B strain host cells. Clonal cell lines were established, and transformants were selected on the basis of resistance to paromomycin, conferred by the injected rDNA. The effects of methylation by three enzymes which methylate the sequence 5'-NAT-3', the dam, EcoRI, and ClaI methylases, were tested. Hypermethylation of the injected rDNA had no effect on transformation efficiency relative to mock-methylated controls. The injected C3 strain rDNA efficiently replaced host rDNA as the major constituent of the population of rDNA molecules. Hypermethylation of the injected DNA was not maintained through 20 to 25 cell generations.

Animals↗
PubMed1989

Accurate processing and amplification of cloned germ line copies of ribosomal DNA injected into developing nuclei of Tetrahymena thermophila.

The ciliate Tetrahymena thermophila contains a chromosomally integrated copy of the rRNA genes (rDNA) in its germinal (micronuclear) genome. These genes are excised from the chromosome through a process involving site-specific DNA breakage, become linear palindromic molecules with added telomeres, and are greatly amplified during development of the somatic nucleus (macronucleus). In this study, we cloned a 15-kilobase segment of the germ line DNA containing these genes and injected it into developing macronuclei of T. thermophila. Up to 11% of injected cells were transformed to the paromomycin-resistant phenotype specified by the injected DNA. Transformation efficiency was dependent on the developmental stages of the injected cells and the integrity of the injected DNA but not the DNA concentration or conformation. The injected DNA was apparently processed and amplified correctly to produce rDNA molecules with the expected linear palindromic structure which carried the appropriate physical markers. Thus, the 15-kilobase DNA contained all cis-acting sequences sufficient for the DNA-processing events leading to rDNA amplification in T. thermophila.

Animals↗
PubMed1976

Chemotherapeutic compounds and Acanthamoebae from eye infections.

The amoebicidal and amoebistatic action in vitro of 24 compounds was tested on two strains of Acanthamoeba, A. polyphaga and A. castellanii, isolated from eye infections in this country. For comparison, the Ryan strain of A. castellanii and Naegleria gruberi L-1 were also examined. Nine compounds showed sufficient activity to merit further consideration, ie, acriflavine, proflavine, hydroxystilbamidine isethionate, paromomycin, miconazole, amphoterin, neomycin, polymyxin, and the last two combined in Neosporin.

Acriflavine↗
PubMed2005

Neamine inhibits xenografic human tumor growth and angiogenesis in athymic mice.

PURPOSE: We have previously shown that the aminoglycoside antibiotic neomycin blocks the nuclear translocation of angiogenin and inhibits its angiogenic activity. However, neomycin has not been considered as a favorable drug candidate for clinical development because of its known nephrotoxicity and ototoxicity. The aim of this study is to determine whether neamine, a nontoxic derivative of neomycin, possesses antitumor activity. EXPERIMENTAL DESIGN: The effect of neamine on the nuclear translocation of angiogenin was examined by means of immunofluorescence and Western blotting. The antitumor activity of neamine was determined with three different animal models. RESULTS: Neamine effectively blocked the nuclear translocation of angiogenin in endothelial cells and inhibited angiogenin-induced cell proliferation. It inhibited the establishment of human tumor xenografts in athymic mice in both ectopic and orthotopic tumor models. It also inhibited the progression of established human tumor transplants, whereas the structurally related antibiotic paromomycin had no effect. Immunohistochemical staining showed that both angiogenesis and cancer cell proliferation are inhibited by neamine. CONCLUSION: These results suggest that the nontoxic aminoglycoside antibiotic neamine is an effective inhibitor of nuclear translocation of angiogenin and may serve as an inhibitor for angiogenin-induced angiogenesis and cancer progression.

Active Transport, Cell Nucleus↗
PubMed2005

Treatment of cryptosporidiosis in immunocompromised hosts.

The most important species of Cryptosporidium in humans, C. parvum and C. hominis, cause severe and chronic life-threatening gastroenteritis in immunocompromised patients. Despite a certain efficacy shown by passive immunotherapy or by some chemotherapeutic agents (e.g. paromomycin and nitazoxanide), no significant benefit has been demonstrated. The use of highly active antiretroviral therapy (HAART) in persons with the acquired immunodeficiency syndrome drastically reduces the prevalence of Cryptosporidium infection. This result seems to be due to aspartyl protease inhibitors of the human immunodeficiency virus included in HAART, which directly interfere with the life cycle of the parasite. The identification of the C. parvum proteases involved in the host-cell interaction could lead to new therapeutic approaches using specific parasite protease inhibitors in immunocompromised persons.

Animals↗
PubMed1980

Inhibitory effects of aminoglycoside antibiotics on reduction of cytochrome c from Candida krusei.

Five aminoglycoside antibiotics (AGAs)--kanamycin (KM), bekamycin (AKM), dibekacin (DKB), ribostamycin (RSM) and paromomycin (PRM)--were studied for their effects on the nonenzymic reduction of cytochrome c by FeSO4 (Yamabe's system). Their inhibitory activity was in the order: DKB greater than AKM greater than KM greater than RSM greater than PRM. As this order correlated closely with that of the antibacterial activity of AGAs, Yamabe's system has proved useful in predicting the latter activity. Divalent metal ions other than Fe2+ enhanced the AGA-dependent inhibition of Yamabe's system in the order: Cu2+ greater than Mn2+ greater than Zn2+ greater than Co2+ greater than Ni2+ greater than. This order was similar to that of stability constants with general chelators except for the low positions of Ni2+ and Co2+. These findings suggested a metal chelation with free or bound Fe2+ for the action mechanism of AGAs on Yamabe's system and the bacterial growing system. The antagonistic effects of exogenous Fe2+ on the antibacterial activity against Staphylococcus aureus and Escherichia coli as measured by the agar dilution method supported this concept. A dual relationship of molecular structure with chelating and antibacterial activities demonstrated the importance of high molecular basicity in a potent AGA. However, the combination effect of pipemidic acid (stimulator on Yamabe's system) with KM was different from that with 1,10-phenanthroline (inhibitor on Yamabe's system) as measured by Dye's method using Pseudomonas aeruginosa.

Aminoglycosides↗
PubMed1981

Cestodes.

A review has been made of the advances in knowledge on the treatment of tapeworm infections of man and animals from the time of the introduction to dichlorophen in 1956. This opened up the era of out-patient treatment for human tapeworm infections. The drugs studied have been compared, where possible, on the basis of estimates of the single dose ED90 or the number of treatments required to reach that efficacy at safe dose levels. During the period under review, niclosamide, introduced in 1960 has been regarded as the drug of choice for the treatment of human tapeworm infections with paromomycin as a possible alternative. The bunamidine salts, introduced in 1966 permitted the treatment of Echinococcus spp in dogs. Several treatments were required to achieve acceptable efficacy. In 1975 praziquantel was introduced and based on the ED90, a single dose at no more than 10mg/kg removed all tapeworms responsible for the cestode zoonoses and for which data are available with the exception of H. nana and D. latum; these require a higher dose rate. In the mid 1970s, several benzimidazoles and praziquantel were shown to have activity against metacestodes. This has opened up a new field of research promising a practical outcome. No progress has been made during the period under review in finding effective ovicides.

Animals↗
PubMed1994

Update on the management of intestinal cryptosporidiosis in AIDS.

OBJECTIVE: To update readers on the pharmacotherapeutic management of cryptosporidiosis in patients with AIDS. DATA SOURCES: A MEDLINE search was used to identify pertinent literature. Additionally, programs and abstracts from the 33rd Interscience Conference on Antimicrobial Agents and Chemotherapy; the 1st International Conference on Macrolides, Azalides, and Streptogramins; the 93rd American Society for Microbiology Meeting; and the 6th and 7th International Conferences on AIDS were used. DATA EXTRACTION: Available data from in vitro, animal, and human experiments were reviewed. DATA SYNTHESIS: Intestinal cryptosporidiosis in patients with AIDS can be a life-threatening opportunistic infection. However, there can be significant variability in disease expression, including spontaneous remission. Supportive care with hydration and nutritional supplementation remains a hallmark of therapy. Unfortunately, there is no proven specific pharmacotherapy of cryptosporidiosis in patients with AIDS. Numerous agents have been analyzed for in vitro activity and efficacy in experimental animal models and actual human cases of the infection, including paromomycin, azithromycin, clarithromycin, octreotide, hyperimmune bovine colostrum, bovine transfer factor, and many others. Because limited numbers of controlled trials have been conducted with potential therapeutic agents, the majority of the information to date is preliminary in nature. CONCLUSIONS: Despite the availability of some evolving and potentially promising treatment modalities, further controlled clinical trials are necessary to evaluate the role of pharmacotherapy for intestinal cryptosporidiosis in patients with AIDS.

AIDS-Related Opportunistic Infections↗
PubMed1995

Treatment of non-HIV cryptosporidial diarrhea with azithromycin.

OBJECTIVE: To report a patient with non-HIV-related cryptosporidial diarrhea who was treated effectively with a regimen of high-dose azithromycin therapy. CASE SUMMARY: A 42-year-old immunocompetent man contracted cryptosporidiosis from an ailing calf that he had purchased. He finally was admitted to the hospital because of excessive weight loss and profuse diarrhea. The patient was started on a course of high-dose azithromycin therapy and symptoms resolved within 48 hours. Follow-up stool cultures were negative for the parasite. DISCUSSION: Although usually associated with immunocompromised patients, cryptosporidiosis occurs in immunocompetent hosts in a significant portion of the reported cases each year. Although self-limiting in most cases in this population, the disease can be severe at times and require treatment. Paromomycin therapy has been used in the past with good results. Although macrolides have had erratic effects against this parasite in the past, azithromycin (an azalide) demonstrated good efficacy in this patient. CONCLUSIONS: Azithromycin has demonstrated that it may be an effective option for the treatment of cryptosporidiosis in immunocompromised patients. Studies involving its use in immunocompromised patients are currently underway.

Adult↗
PubMed1996

Human cryptosporidiosis.

Cryptosporidium parvum is a protozoan which can cause severe debilitating disease in immunocompromised individuals. Animal models have shown that cellular immunity is the most important factor against the development of the disease. Individuals with a humoral immune deficiency are also at risk. In HIV-infected patients there is a clear relationship between disease severity and CD4 cell counts. Insight into the pathogenesis and development of new agents is hampered by the lack of an in vitro culture system. Prevention is of the utmost importance due to the difficulties of therapy and the severity of clinical disease which can develop. Oocysts are highly resistant to commonly used disinfectants. In HIV-infected patients with cryptosporidiosis, antiretroviral therapy should be instituted or modified. Moreover, non-specific therapy with antidiarrhoeal agents should also be instituted. If no effect is seen, therapy with paromomycin 500 mg 4 times daily for 2-3 weeks should be initiated, followed by maintenance therapy with 500 mg twice daily to prevent relapse.

AIDS-Related Opportunistic Infections↗
PubMed2006

Rye (Secale cereale L.).

Rye (Secale cereale L.) is one of the most recalcitrant plant species for tissue culture and genetic transformation. Embryogenic rye callus loses its ability to regenerate plants quickly in response to high density of Agrobacterium and other stressors. The cocultivation of Agrobacterium and rye immature embryos in liquid medium facilitated washing of the cultures to avoid Agrobacterium overgrowth and allowed a high throughput. More than 40 independent transgenic plants were regenerated with one to four Southern-positive, independent events from 100 inoculated immature embryos. Agrobacterium strain AGL0 supported stable integration of a constitutive nptII selectable marker expression cassette into the genome of rye inbred line L22, as indicated by regeneration of plantlets on paromomycin-containing culture medium, Southern blot, Western blot, and the analysis of T-DNA::plant DNA boundary sequences. Transgenic plants were phenotypically normal and fully fertile, which might be a consequence of the short time in tissue culture.

Agrobacterium tumefaciens↗