Antibiotic Tiamulin Research Articles

Degradation of tiamulin by a packed bed dielectric barrier plasma combined with TiO2 catalyst

Recently, a plasma catalyst was employed to efficiently degrade antibiotic residues in the environment. In this study, the plasma generated in a packed bed dielectric barrier reactor combined with TiO2 catalyst is used to degrade the antibiotic tiamulin (TIA) loaded on the surface of simulated soil particles. The effects of applied voltage, composition of the working gas, gas flow rate and presence or absence of catalyst on the degradation effect were studied. It was found that plasma and catalyst can produce a synergistic effect under optimal conditions (applied voltage 25 kV, oxygen ratio 1%, gas flow rate 0.6 l min−1, treatment time 5 min). The degradation efficiency of the plasma combined with catalyst can reach 78.6%, which is 18.4% higher than that of plasma without catalyst. When the applied voltage is 30 kV, the gas flow rate is 1 l min−1, the oxygen ratio is 1% and the plasma combined with TiO2 catalyst treats the sample for 5 min the degradation efficiency of TIA reached 97%. It can be concluded that a higher applied voltage and longer processing times not only lead to more degradation but also result in a lower energy efficiency. Decreasing the oxygen ratio and gas flow rate could improve the degradation efficiency. The relative distribution and identity of the major TIA degradation product generated was determined by high-performance liquid chromatography–mass spectrometry analysis. The mechanism of TIA removal by plasma and TiO2 catalyst was analyzed, and the possible degradation path is discussed.

Design, synthesis and antibacterial activity of novel pleuromutilin derivatives with thieno[2,3-d]pyrimidine substitution

A series of novel pleuromutilin derivatives with substituted thienopyrimidines were designed, synthesized, and evaluated for antibacterial act ivity. In this study, the activities of these compounds were investigated using the inhibition circle test, the minimum inhibitory concentration (MIC) test, real-time growth curves, time-kill kinetic assays, cytotoxicity assays, and molecular docking. Most of the tested compounds exhibited moderate antibacterial activity against Staphylococcus aureus, Streptococcus agalactiae, and Escherichia coli. Compound A11 was the most active and displayed bacteriostatic activities against methicillin-resistant S. aureus, with MIC values as low as 0.00191 μg/mL, which is 162 and 32 times lower than that of the marketed antibiotics tiamulin and retapamulin, respectively. Furthermore, the mechanism of action of A11 was confirmed by molecular docking studies.

Following the route of veterinary antibiotics tiamulin and tilmicosin from livestock farms to agricultural soils

Veterinary antibiotics (VAs) are not completely metabolized in the animal body. Hence, when animal excretes are used as soil manures, VA residues are dispersed with potential implications for environmental quality and human health. We studied the persistence of tiamulin (TIA) and tilmicosin (TLM) along their route from pig administration to fecal excretion and to agricultural soils. TLM was detected in feces at levels folds higher (4.27–749.6 μg g-1) than TIA (0.55–5.99 μg g-1). Different administration regimes (feed or water) showed different excretion patterns and residual levels for TIA and TLM, respectively. TIA and TLM (0.5, 5 and 50 μg g-1) dissipated gradually from feces when stored at ambient conditions (DT50 5.85–35.9 and 23.5–49.8 days respectively), while they persisted longer during anaerobic digestion (DT90 >365 days) with biomethanation being adversely affected at VA levels > 5 μg g-1. When applied directly in soils, TLM was more persistent than TIA with soil fumigation extending their persistence suggesting microbial degradation, while soil application through feces increased their persistence, probably due to increased sorption to the fecal organic matter. The use of TIA- and TLM-contaminated feces as manures is expected to lead to VAs dispersal with unexplored consequences for the environment and human health.

Improvement of biotreatability of environmentally persistent antibiotic Tiamulin by O3 and O3/H2O2 oxidation processes

ABSTRACT The aim of the work was to assess the efficiency of ozonation and ozonation in combination with H2O2 in jet loop reactor to increase biotreatability of persistent veterinary antibiotic Tiamulin. The efficiency of oxidative processes was monitored by combined approach based on determination of efficiency of wastewater treatment and impact to waste sludge stabilization. Degradation of Tiamulin in model wastewater (100 mg L−1) during oxidation was followed by COD and DOC measurements while changes in biodegradability were determined by respirometric measurements. Biogas production potential was also determined to identify problems related to anaerobic digestion of waste sludge resulted in treatment of Tiamulin-contaminated wastewater. At ozone dose of 69 gozone gCOD −1 and 220 gozone gDOC −1removal for COD and DOC was 26% and 17%, respectively. Better biotreatability was confirmed by respirometric testing. H2O2 addition did not improve removal efficiency (11–13%). The second stage of nitrification was suppressed by the addition of Tiamulin and ozonation again recovered formation. O3/H2O2 treated sample reduced the nitrification, especially formation of in the first phase of the process. Simultaneously, quadratic model was developed to describe the relationship between oxygen uptake rate and changes in ammonium nitrogen concentration due to the oxidative treatment. The positive impact of ozone was also confirmed by ozonation of Tiamulin-contaminated (400 mg L−1) waste sludge where biogas production potential was increased for 6-times. Combination of approaches confirmed, that O3 effectively increase the treatability of Tiamulin in wastewater and sludge while addition of the hydrogen peroxide generally did not improve the performance of the processes.

Radiated temperature of weaned piglets and the response to in-feed antibiotics with correction for ambient conditions

Growth performance, health status, and the radiated thermal temperatures of groups of piglets consuming non-medicated feed and in-feed antibiotics (chlortetracycline hydrochloride and tiamulin) were monitored continuously for up to 35 d, from weaning at 28 d of age until moving to commercial pens at approximately 63 d of age. There was a trend (P = 0.1) for body weight at the time of moving to be higher for the medicated animals. The average daily gain was higher in medicated piglets (P = 0.03). Radiated temperature variables were the maximum (Tmax) and the average (Tmean) pig temperatures, and both were higher for the medicated piglets (P < 0.0001). A threshold temperature was derived to partition the image and derive the background radiated temperature that was used to obtain the expected radiated temperature of the pigs. Temperature data were adjusted for environmental effects by expressing the residual temperature. The residual for Tmax (TmaxRes) and for Tmean (TmeanRes) were higher in medicated piglets (P < 0.0001). In-feed antibiotics were associated with increased growth rates and with an increase in radiated temperature variables, particularly those corrected for environmental conditions, possibly mediated through increased metabolic activity related to growth.

Synthesis, biological activities and docking studies of pleuromutilin derivatives with piperazinyl urea linkage

Antibiotics resistance is becoming increasingly common, involving almost all antibiotics on the market. Diseases caused by drug resistant bacteria, such as MRSA, have high mortality and negatively affect public health. The development of new drugs would be an effective means of solving this problem. Modifications based on bioactive natural products could greatly shorten drug development time and improve success rate. Pleuromutilin, a natural product from the basidiomycete bacterial species, is a promising antibiotic candidate. In this study, a series of novel pleuromutilin derivatives possessing piperazinyl urea linkage were efficiently synthesised, and their antibacterial activities and bactericidal properties were evaluated via MIC, MBC and Time-kill kinetics assays. The results showed that all compounds exhibited potent activities against tested strains, especially MRSA strains with MIC values as low as 0.125 μg/mL; 8 times lower than that of marketed antibiotic Tiamulin. Docking studies indicate substituted piperazinyl urea derivatives could provide hydrogen bonds and initiate π-π stacking between molecules and surrounding residues.

PSV-4 Direct-fed microbial program improved survivability of nursery pigs in commercial production

Abstract A total of 2,304 newly weaned pigs (5.8±0.06 Kg BW, ~21-day old, DNA Genetics) were used in a RCBD to evaluate the effect of a direct-fed microbial (DFM) program on pig performance and survivability, under commercial pig production. Pigs were housed in 2 barns (blocking factor) with 36 pens each (32 pigs/pen, mixed-sex), and pens randomized to treatments: Control or DFM program (36 pens/treatment). The program consisted of a DFM (blend 1) delivered through drinking water (1.32x108 CFU/pig/day) from day 0 to 7, followed by another DFM (blend 2) in feed (1.5x105 CFU/g of feed) from day 7 through 49. Those DFM consisted of spore-forming Bacillus sp. selected to inhibit enterotoxigenic E. coli growth (blend 1) and reduce local inflammation (blend 2). The feeding program included phases 1 and 2 (7 days each) in pellet, and phases 3 (11 days) and 4 (24 days) in meal. Phase 1 feeds included antibiotics Chlortetracycline and Tiamulin; both phase 4 feeds included Carbadox. A severe diarrhea outbreak was observed in week 3; antibiotic therapy (Neomycin, Sulfadiazine or Trimethoprim) throughout drinking water was used in both treatments alike between days 19 and 28. Pig and feed weight were recorded by pen; ADG and ADFI were calculated on pig-days. Pigs removed included mortality and those ill, nonresponsive to antibiotics. Performance data were analyzed by ANOVA, using the GLM procedure. Pigs removed did not follow normal distribution and was analyzed using a parametric survival option with days as time variable (JMP v14.2.0). No differences were detected through day 25. Pigs in DFM consumed less feed (923 vs. 951 g/d; SEM=10.33; P=0.05) and were more efficient (0.61 vs. 0.59 kg/kg; SEM=0.004; P< 0.05) from days 25 to 49. Total pigs removed were reduced (P< 0.05) by 23% in DFM vs. Control (Table 1). In conclusion, the DFM program improved nursery pig survivability and late performance, under commercial pig production that included a diarrhea outbreak and antibiotic therapy.

Bacterial community structures and biodegradation kinetic of Tiamulin antibiotic degrading enriched consortia from swine wastewater

The antibiotic tiamulin (TIA) is common and widely used medication for dysentery eradication in swine productions. Tiamulin persists in livestock manure, and its residues have been found in various environment. This work obtained four tiamulin-degrading enriched bacterial consortia from a covered anaerobic lagoon system and a stabilized pond system of swine farms. Tiamulin was efficiently removed by the enriched cultures at the concentrations between 2.5 and 200mg/L, with a removal of 60.1-99.9% during 16h and a degradation half-life of 4.5-15.7h. The stabilized pond system cultured with taimulin solely could eliminate tiamulin at the highest rates. The logistic substrate degradation model fit most of the experimental data. Next-generation amplicon sequencing was conducted, and it was found that the bacterial community was significantly impacted by the inoculum source, nutrient addition, and high tiamulin concentrations. Principal coordinate analysis (PCoA) indicated the similarity of bacterial communities in the original enriched samples and the 2.5mg/L tiamulin-removed cultures. The 200mg/L consortia were rather different and became similar to the other 200mg/L consortia from different sources and cultures without nutrient supplementation. Shannon and Simpson indices suggested a reduction in bacterial diversity at high concentrations. The microbes that had high growth in the most efficient enriched culture, or which were abundant in all samples, or which increased with higher tiamulin concentrations were likely to be the major tiamulin-degrading bacteria. This is the first report suggested the possible roles of Achromobacter, Delftia, Flavobacterium, Pseudomonas, and Stenotrophomonas in tiamulin degradation.

Occupational inhalation poisoning with the veterinary antibiotic tiamulin

Introduction: Tiamulin is a semisynthetic pleuromutilin diterpene veterinary antibiotic, widely used in farms. We present a case of prolonged QT-interval and ventricular tachyarrhythmia after tiamulin inhalation.Case presentation: A 43-year-old veterinarian without previous medical history was dividing granulated powder of antibiotic gravimetrically without wearing personal protective equipment. Half an hour after exposure, nausea occurred; four hours later he started to vomit and soon after that he experienced syncope. He was unconscious three minutes; afterwards he became somnolent, dizzy and nauseated with sweating and salivation. On admission to hospital five hours after exposure, he was conscious and had heart rate 70 beats/min and blood pressure 140/80 mmHg. Initial laboratory results were normal. Electrocardiography showed a prolonged QTc-interval of 730 ms with numerous polymorphic ventricular extrasystoles and episodes of non-sustained polymorphic ventricular tachycardia that resolved after treatment with lidocaine and magnesium. Subsequent electrocardiography revealed gradual shortening of QTc-interval with QTc-interval normalization (430 ms) between 24 and 32 hours after tiamulin exposure. Laboratory tests, morphologic heart diagnostics and genetic testing excluded other potential causes of QTc-interval prolongation. Subsequent toxicology analysis by LC-MS/MS confirmed tiamulin in his serum samples on admittance (500 ng/mL).Conclusion: Tiamulin inhalation can be associated with prolonged QT-interval and ventricular tachyarrhythmia. QT-interval prolongation could be expected in overdoses of emerging human pleuromutilins.

Ozone as a Pretreatment Method for Antibiotic Contaminated Wastewater and Sludge

ABSTRACTThe capability of ozone to reduce the hazardous impact of environmentally persistent antibiotic tiamulin in term of toxicity reduction and enhancement of biodegradability was investigated. Different ozone doses were applied but ozonation was not effective enough to increase the biodegradability of tiamulin in the aqueous phase. The opposite effect was observed in anaerobic digestion experiments, where ozonation as a pretreatment step of antibiotic-contaminated sludge detoxify tiamulin and improves biogas production for 75%. As confirmed by 1H NMR and HPLC-HRMS analyses, the tiamulin molecule completely reacts with ozone at low ozone/COD molar ratio of 0.03, primarily attacking the vinyl double bond with further oxidation of sulfur and nitrogen atom, and gradual decomposition of tiamulin skeleton.

Mutations in 23S rRNA gene associated with decreased susceptibility to tiamulin and valnemulin in Mycoplasma gallisepticum

Mycoplasma gallisepticum is a major etiological agent of chronic respiratory disease (CRD) in chickens and sinusitis in turkeys. The pleuromutilin antibiotics tiamulin and valnemulin are currently used in the treatment of M. gallisepticum infection. We studied the in vitro development of pleuromutilin resistance in M. gallisepticum and investigated the molecular mechanisms involved in this process. Pleuromutilin-resistant mutants were selected by serial passages of M. gallisepticum strains PG31 and S6 in broth medium containing subinhibitory concentrations of tiamulin or valnemulin. A portion of the gene encoding 23S rRNA gene (domain V) and the gene encoding ribosome protein L3 were amplified and sequenced. No mutation could be detected in ribosome protein L3. Mutations were found at nucleotide positions 2058, 2059, 2061, 2447 and 2503 of 23S rRNA gene (Escherichia coli numbering). Although a single mutation could cause elevation of tiamulin and valnemulin MICs, combinations of two or three mutations were necessary to produce high-level resistance. All the mutants were cross-resistant to lincomycin, chloramphenicol and florfenicol. Mutants with the A2058G or the A2059G mutation exhibited cross-resistance to macrolide antibiotics erythromycin, tilmicosin and tylosin.

The activity and compatibility of the antibiotic tiamulin with other drugs in poultry medicine—A review

Tiamulin hydrogen fumarate is a semisynthetic derivative of the diterpene antibiotic pleuromutilin used in poultry medicine to treat mainly Mycoplasma- and Brachyspira-related diseases. Its use over 30 yr has not generally increased the development of resistance to these pathogens but occasionally resistant isolates are encountered. Tiamulin administered at therapeutic levels is relatively quickly absorbed, metabolized in the liver, and eliminated from the body of the bird after a withdrawal period of 72 h, and as a result, meat products can be safely consumed. A zero withdrawal period for eggs has been granted in several European Union states. When administered with different drugs, tiamulin has been shown to have an enhanced activity with the tetracyclines. There is a strong interaction, even death, with the ionophore anticoccidials monensin, narasin, and salinomycin when tiamulin is used at therapeutic levels, but this is dose-related and low doses do not interact. It is thought to be caused by the preferential metabolism of tiamulin in the liver resulting in a build up of the ionophore leading to clinical signs of overdosage. Tiamulin shows a milder interaction, such as temporary growth depression, with maduramicin and semduramicin but is compatible with lasalocid. Although tiamulin shows small benefits in improving performance in healthy animals, its main production benefit is in the face of infection, as a true therapeutic antibiotic.

Compatibility of a Combination of Tiamulin plus Chlortetracycline with Salinomycin in Feed During a Long-Term Co-Administration in Broilers

The pleuromutilin antibiotic tiamulin (TIA) is known to produce a negative interaction in broilers when administered in combination with several ionophore anticoccidials such as salinomycin (SAL). Chlortetracycline (CTC), when administered simultaneously with TIA, has demonstrated a synergistic antimicrobial effect. A 35-d feeding study was conducted in cages to evaluate the interaction effect of a combination of TIA plus CTC at increasing inclusion levels when administered concurrently with SAL. A total of 200 one-day-old broiler chicks were distributed into 4 groups, and each group consisted of 5 cages containing 10 birds in each. Replicate cages were distributed randomly. Feed for all groups contained 60 ppm SAL, but additionally, 0, 20, 30, and 50 ppm TIA and 0, 60, 90, and 150 ppm CTC were included, respectively. Several enzymes (creatine phosphokinase, lactate dehydrogenase, and aspartate aminotransferase) were determined from blood samples taken at the end of the trial. Blood samples were also collected during d 0, 19, and 35 and were analyzed for antibody titers against Mycoplasma gallisepticum and Mycoplasma synoviae. Necropsy of a few birds (20, 8, 20, 12, and 12 on d 7, 14, 21, 28, and 35, respectively) was conducted at weekly intervals. Results indicated that there was a significant depression of weight gain (P < 0.05) in group 4 (TIA 50 + CTC 150) only. The final weights were 1,809 ± 130, 1,859 ± 52, 1,703 ± 47, and 1,617 ± 98 g for groups 1 (TIA 0 + CTC 0), 2 (TIA 20 + CTC 60), 3 (TIA 30 + CTC 90), and 4 (TIA 50 + CTC 150), respectively. However, feed intake and feed conversion efficiency (g of weight gain/kg of feed intake) were not significantly affected in any of the groups. There was no dose-related adverse effect on mortality or clinical signs exhibited during the trial, and this was supported by necropsy. Maternally derived antibodies against M. gallisepticum were present at the beginning of the trial but disappeared within 19 d. Otherwise, there was no apparent infection by M. gallisepticum or M. synoviae throughout the trial. The results demonstrate that 50 ppm TIA plus 150 ppm CTC along with 60 ppm SAL caused only a depression of growth, but no adverse signs of interaction were detected. Taking into consideration all the aspects of the cost of production, the 20 ppm TIA plus 60 ppm CTC was the most cost-effective level to administer continuously with 60 ppm SAL via the feed, but it would be important to do an additional study using an artificial infection with M. gallisepticum or M. synoviae to know whether this inclusion rate would be sufficient to protect against an infectious challenge.

Tiamulin and semduramicin: effects of simultaneous administration on performance and health of growing broiler chickens

The pleuromutilin antibiotic tiamulin (TIA) is known to produce a variety of negative interactive effects when it is administered in combination with several anticoccidial ionophores. A 35-d growth study was performed in cages to evaluate the compatibility of TIA when it was administered concurrently with the poly-ether ionophore anticoccidial semduramicin (SEM). Tiamulin and SEM, both alone and in combination, were administered to 10 replicates of female broilers arranged in a completely randomized block design. Tiamulin was administered in drinking water (250 mg of TIA/kg of water) from d 15 through 19 of the study, whereas SEM was incorporated in feed (25 mg/kg) from placement to the conclusion of the test. Water consumption was determined during the period of concurrent administration of the drugs and weekly measurements of feed intake and bird performance were recorded. In addition, hematocrit, blood cell counts, serum protein, albumin, glucose, uric acid, electrolytes, and activities of several enzymes were determined from blood samples taken at d 35. Results indicated that simultaneous administration of TIA and SEM during the third week of the trial reduced water and feed intake resulting in a temporary growth depression. Feed efficiency was transiently affected during the period of coadministration. However, during the fourth week of the test, negative effects in body weight were not observed for any treatment and feed conversion improved for birds concurrently receiving TIA + SEM. By the termination of the experiment, no adverse effects were observed in final performance for any treatment. Histopathological and hematological parameters were unaffected by treatment at d 35 of the test. These results demonstrated that simultaneous administration of TIA and SEM produced only temporary impairments of water and feed consumption that transiently influenced performance. Neither mortality nor long-term effects on performance variables occurred in broilers.

Mutations in ribosomal protein L3 and 23S ribosomal RNA at the peptidyl transferase centre are associated with reduced susceptibility to tiamulin in Brachyspira spp. isolates

The pleuromutilin antibiotic tiamulin binds to the ribosomal peptidyl transferase centre. Three groups of Brachyspira spp. isolates with reduced tiamulin susceptibility were analysed to define resistance mechanisms to the drug. Mutations were identified in genes encoding ribosomal protein L3 and 23S rRNA at positions proximal to the peptidyl transferase centre. In two groups of laboratory-selected mutants, mutations were found at nucleotide positions 2032, 2055, 2447, 2499, 2504 and 2572 of 23S rRNA (Escherichia coli numbering) and at amino acid positions 148 and 149 of ribosomal protein L3 (Brachyspira pilosicoli numbering). In a third group of clinical B. hyodysenteriae isolates, only a single mutation at amino acid 148 of ribosomal protein L3 was detected. Chemical footprinting experiments show a reduced binding of tiamulin to ribosomal subunits from mutants with decreased susceptibility to the drug. This reduction in drug binding is likely the resistance mechanism for these strains. Hence, the identified mutations located near the tiamulin binding site are predicted to be responsible for the resistance phenotype. The positions of the mutated residues relative to the bound drug advocate a model where the mutations affect tiamulin binding indirectly through perturbation of nucleotide U2504.

Resistance to the peptidyl transferase inhibitor tiamulin caused by mutation of ribosomal protein l3.

The antibiotic tiamulin targets the 50S subunit of the bacterial ribosome and interacts at the peptidyl transferase center. Tiamulin-resistant Escherichia coli mutants were isolated in order to elucidate mechanisms of resistance to the drug. No mutations in the rRNA were selected as resistance determinants using a strain expressing only a plasmid-encoded rRNA operon. Selection in a strain with all seven chromosomal rRNA operons yielded a mutant with an A445G mutation in the gene coding for ribosomal protein L3, resulting in an Asn149Asp alteration. Complementation experiments and sequencing of transductants demonstrate that the mutation is responsible for the resistance phenotype. Chemical footprinting experiments show a reduced binding of tiamulin to mutant ribosomes. It is inferred that the L3 mutation, which points into the peptidyl transferase cleft, causes tiamulin resistance by alteration of the drug-binding site. This is the first report of a mechanism of resistance to tiamulin unveiled in molecular detail.

In vitro antibiotic susceptibility of field isolates of Mycoplasma synoviae in Argentina.

Minimum inhibitory concentrations (MICs) were determined in vitro for 7 antibiotics (aivlosin, enrofloxacine, tylosin, tiamulin, kitasamycin, chlortetracycline, and oxytetracycline) against eight recent local Argentinean isolates and two standard strains of Mycoplasma synoviae. Aivlosin (3-acetyl-4"-isovaleryl tylosin tartrate), tylosin, and tiamulin showed the lowest MICs with MIC90s of 0.006, 0.012, and 0.05 microg/ml, respectively. Except one strain that showed resistant values to chlortetracycline (> or = 12.5 microg/ml), all the analyzed strains were susceptible in different degrees to all the antibiotics tested. In this study, the improved activity of the tylosin-derived drug, aivlosin, was confirmed because it showed, in most strains, MIC values half those for tylosin.

In vitro complex formation and inhibition of hepatic cytochrome P450 activity by different macrolides and tiamulin in goats and cattle

In humans, clinically relevant drug–drug interactions occur with some macrolide antibiotics via the formation of stable metabolic intermediate (MI) complexes with enzymes of the cytochrome P4503A (CYP3A) subfamily. The formation of such complexes can result in a decreased biotransformation rate of simultaneously administered drugs. In previous studies it was shown that the veterinary antibiotic tiamulin was also able to form a stableMIcomplex in pigs and rats. In the present study the relative CYP3A inhibiting potency andMIcomplex formation of a series of macrolide antibiotics and tiamulin were studied in microsomal fractions of goat and cattle and in a cell-line expressing bovine CYP3A. Tiamulin and triacetyloleandomycin (TAO) were found to be effective inhibitors of CYP450 activity in all systems tested. Erythromycin and tilmicosin were found to be relatively less effective inhibitors of CYP450 activity in microsomes, and their activity in the bovine CYP3A4 expressing cell line was relatively weak. Tylosin was shown to be a weak inhibitor in microsomes and not in the cell line, whereas spiramycin had no effect at all.MI-complex formation measured by spectral analysis was seen withTAO, tiamulin, erythromycin and tylosin, but not with tilmicosin and spiramycin. Although additional factors play a role in vivo, these results may explain potential drug–drug interactions and differences between related compounds in this respect.

In vitro and in vivo reversal of cancer cell multidrug resistance by the semi-synthetic antibiotic tiamulin

A large number of multidrug resistance (MDR) modulators, termed chemosensitizers, have been identified from a variety of chemicals, but most have been proven to be clinically toxic. Low concentrations of the pleuromutilin-derived semi-synthetic antibiotic tiamulin (0.1 to 10 μM) sensitized the three highly resistant P-glycoprotein (Pgp)-overexpressing tumor cell lines P388 (murine lymphoid leukemia), AS30-D (rat hepatoma), CEM (human lymphoblastic leukemia), and the barely resistant AS30-D/S cell lines to several MDR-related anticancer drugs. Flow cytometric analysis showed that tiamulin significantly increased the intracellular accumulation of daunomycin. When compared to reference modulating agents such as verapamil and cyclosporin A, tiamulin proved to be 1.1 to 8.3 times more efficient in sensitizing the resistant cell lines. Moreover, when given i.p. (1.6 μg/mg body weight), tiamulin increased the survival rate of adriamycin-treated mice bearing the P388/ADR25 tumor line by 29%. In the presence of an anticancer drug, tiamulin inhibited both ATPase and drug transport activities of Pgp in plasma membranes from tumor cells. Tiamulin is thus a potent chemosensitizer that antagonizes the Pgp-mediated chemoresistance in many tumor cell lines expressing the MDR phenotype at different levels and displays no toxic effects on contractile tissues at active doses, therefore providing the promise for potential clinical applications.

Cytochrome P-450 complex formation in rat liver by the antibiotic tiamulin.

Tiamulin is a semisynthetic diterpene antibiotic frequently used in farm animals. The drug has been shown to produce clinically important--often lethal--interactions with other compounds. It has been suggested that this is caused by a selective inhibition of oxidative drug metabolism via the formation of a cytochrome P-450 metabolic intermediate complex. In the present study, rats were treated orally for 6 days with tiamulin at two different doses: 40 and 226 mg/kg of body weight. For comparison, another group received 300 mg of triacetyloleandomycin (TAO) per kg, which is equivalent to the 226-mg/kg tiamulin group. Subsequently, microsomal P-450 contents, P-450 enzyme activities, metabolic intermediate complex spectra, and P-450 apoprotein concentrations were assessed. In addition, effects on individual microsomal P-450 activities were studied in control microsomes at different tiamulin and substrate concentrations. In the rats treated with tiamulin, a dose-dependent complex formation as evidenced by its absorption spectrum and an increase in cytochrome P-4503A1/2 contents as assessed by Western blotting (immunoblotting) were found. The effects were comparable to those of TAO. Tiamulin induced microsomal P-450 content, testosterone 6 beta-hydroxylation rate, erythromycin N-demethylation rate, and the ethoxyresorufin O-deethylation activity. Other activities were not affected or decreased. When tiamulin was added to microsomes of control rats, the testosterone 6 beta-hydroxylation rate and the erythromycin N-demethylation were strongly inhibited. It is concluded that tiamulin is a potent and selective inducer-inhibitor of cytochrome P-450. Though not belonging to the macrolides, the compound produces an effect on P-450 similar to those of TAO and related compounds.