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Macrolides are unique among the various classes of antimicrobial agents because of the manner in which they interact with pathogens and the host immune system to produce a clinical response. Conventional efficacy explanations require the serum or tissue concentration of the macrolide to exceed the MIC of the macrolide to account for bacteriostatic activity. However, at concentrations below the MIC, there are other effects on the expression of proteinaceous virulence factors which could prevent the pathogen from establishing or expanding an infection. The purpose of this review is to describe these effects and to provide an in-vivo scenario delineating the role of macrolides and leucocytes as they contribute to clinical efficacy. [ABSTRACT FROM AUTHOR]

A letter to the editor is presented in response to the article "Presence of Multidrug Resistant Enteric Bacteria in Dairy Farm Topsoil," by J. M. Burgos, B. A. Ellington and M. F. Varela in a 2005 issue.

Much effort has been invested in the development and implementation of international recommendations to manage the risk of foodborne antimicrobial resistance, and monitoring programmes to measure bacterial antimicrobial resistance and antimicrobial product volumes. A variety of approaches have been recommended for various stakeholders in the food animal and food production sectors. Interestingly, much less consideration has been given to the establishment of success criteria for the individual interventions and even less for the cumulative effects, when all interventions are considered together as consecutive 'hurdles' along the food chain. The author explores the outcome and unforeseen consequences of these various interventions and appropriate methods that could provide data to assess their impact, as well as key learning experiences that should lead to refinements of such interventions in the future.

Many studies and meeting reports have suggested that the use of some antibiotics in food animals can compromise the treatment of some infectious diseases in humans. Although the studies and reports are timely and important, it is difficult to assess the relative value of the conclusions in relationship to the overall situation concerning antibiotic resistant foodborne bacteria because the data used in the analyses are often of disparate origin. The studies have attempted to establish a cause and effect relationship between the use ('consumption') of antibiotics in food animals and treatment failures in human disease on the basis of [1] antibiotic usage data; [2] in vitro determinations of antibiotic susceptibility of animal and human isolates, [3] results obtained from controlled animal experiments or [4] epidemiological data. Each approach has sought to associate bacterial antibiotic resistance data with it's own immediate focus area of investigation. However, a true assessment of the degree of contribution to human antibiotic resistance problems from animal use can only be facilitated by comprehensively organizing these different approaches into a concerted, coordinated effort. Concurrently, the implementation of a multinational programme aimed at monitoring antibiotic usage in food animals and resistance in specific bacteria associated with those animals should be instituted. In parallel with this endeavour is the implementation of new prudent use guidelines for antibiotic use by veterinarians. Through the use of science-based approaches like these, the development and spread of antibiotic resistant bacteria associated with food animals could be minimized and contained.

Tilmicosin is a semisynthetic macrolide antibiotic currently approved for veterinary use in cattle and swine to combat respiratory disease. Because the concentrations of tilmicosin are generally low in bovine serum, the interaction of tilmicosin with three types of bovine phagocytes (monocyte-macrophages, macrophages, and neutrophils from blood, lungs, and mammary gland, respectively) and mammary gland epithelial cells was evaluated to provide an understanding of potential clinical efficacy. After incubation with radiolabeled tilmicosin, uptake was determined and expressed as the ratio of the intracellular to the extracellular drug concentration. Accumulation of tilmicosin at 4 h of incubation by the alveolar macrophages (Cc/Ce 193) was 4 to 13 times more than that observed in monocyte-macrophages (Cc/Ce 43), neutrophils, (Cc/Ce 13), or mammary epithelial cells (Cc/Ce 20). Subcellular distribution showed that 70 to 80% of tilmicosin was localized in the lysosomes. Uptake in mammary gland cells was dependent on cell viability, temperature, and pH, but was not influenced by metabolic inhibitors or anaerobiosis. However, lipopolysaccharide exposure increased tilmicosin uptake by the bovine mammary macrophages and epithelial cells. When neutrophils and epithelial cells were incubated in the presence of tilmicosin and extracellular tilmicosin was then removed, 40% of the intracellular tilmicosin remained cell associated after 4 h of incubation (i.e., 60% effluxed), but only 25% remained in macrophages. These in vitro interactions of tilmicosin with bovine phagocytes and epithelial cells suggest an integral role in effecting clinical efficacy.

Forty-four cows (26 Jerseys and 18 Holsteins) that had at least 1 mammary quarter that was naturally (n = 12) or experimentally (n = 84) infected with Staphylococcus aureus were allotted to three treatment groups of approximately equal number at the end of lactation. Cows were dried off by abrupt cessation of milking, and dry cow therapy was administered as an intramammary infusion of cephapirin benzathine at 10 ml per quarter, an intramammary infusion of tilmicosin (solution containing 300 mg/ml) at 5 ml per quarter, or a subcutaneous injection of tilmicosin at 5 mg/kg of body weight on the day of drying off and another injection 4 d later. Mammary secretions were monitored during the dry period and postpartum for antimicrobial residues, intramammary infection (IMI) status, and somatic cell counts. Results demonstrated the following percentage cures for IMI caused by Staph. aureus at 28 d postcalving based on individual mammary quarters: cephapirin benzathine, 78.1%; tilmicosin infused, 74.2%; and tilmicosin injected, 9.1%. During the first 4 wk after drying off, the mean concentration of tilmicosin in mammary secretions from cows infused with the antibiotic remained approximately 10-fold higher than that in secretions from cows injected with the antibiotic (3.43 vs. 0.32 ppm), and, by the time of calving, concentrations for cows treated with both methods were below the dilution limit of the assay (< 0.1 ppm). Results demonstrated that intramammary infusion of tilmicosin was equally as effective as cephapirin benzathine in curing IMI caused by Staph. aureus at drying off; however, the subcutaneous injection of tilmicosin at the dose used was not effective as a dry cow therapeutic against Staph. aureus.

In order to compare the intracellular concentration of antimicrobial agents in phagocytic and nonphagocytic cells, the knowledge of their cell volume is essential. For the first time, the determination of the avian, porcine, and bovine polymorphonuclear neutrophils (PMN), monocyte-derived macrophages, macrophages, and bovine mammary epithelial cell volume was performed using [3H]-water and [14C]-carboxyinulin. The comparison of all the cells showed that the PMN have a size range between 3.58 and 4.04 microL per mg of protein, and are smaller than the monocyte-derived macrophages and mammary epithelial cells (4.32-5.01 microL per mg of protein). The macrophages show the largest size (5.84-6.57 microL per mg of protein). The cellular uptake of tilmicosin in these cells was then determined. The examination of the intracellular/extracellular concentration ratios (Ci/Ce) after 4 h of incubation with 10 mg/mL of [14C]-labelled tilmicosin revealed that tilmicosin was well accumulated and showed a ratio of 137, 169 and 193 in avian PMN, porcine PMN, and bovine alveolar macrophages, respectively. The cellular uptake data also demonstrated that tilmicosin accumulated in nonphagocytic bovine mammary epithelial cells. The importance of the use of the appropriate species and cell type specific cell volume values for calculations was exemplified by calculating the Ci/Ce of tilmicosin using cell volume data found in the literature for human and mouse cells. The subsequent comparison of these data with the Ci/Ce calculated with the actual cell volume appropriate for the species tested revealed an under evaluation of 3-13% in monocyte-macrophages, an over evaluation of 7-18%, 16-31% and 69% in PMN, macrophages, and epithelial cells, respectively. This study highlights the importance of the proper cell volume in order to determine the Ci/Ce. Moreover, the cell volumes determined here for avian, porcine and bovine cells should facilitate further in vitro and in vivo cellular studies by veterinary researchers.

Tilmicosin is a semi-synthetic macrolide antibiotic, currently approved for veterinary use in cattle and swine respiratory disease, and is in development for use in poultry mycoplasma air sacculitis. In order to provide an understanding of clinical efficacy, the in vitro interaction of tilmicosin with three types of chicken phagocytes (MQ-NCSU macrophages, monocyte-macrophages, and heterophils) was evaluated. After incubation with radiolabeled tilmicosin, uptake was determined and expressed as the ratio of the cellular (Cc) to the extracellular (Ce) drug concentration (Cc:Ce). Tilmicosin was avidly accumulated by heterophils (Cc: Ce 138 at 4 h incubation vs 32 and 66, respectively, in MQ-NCSU and monocyte-macrophages) with 61 to 88% localized in the lysosomes. Uptake was dependent on cell viability, temperature, and pH, but was not influenced by metabolic inhibitors. However, phagocytosis of Pasteurella multocida and lipopolysaccharide exposure increased tilmicosin uptake by the chicken phagocytes. Upon removal of extracellular tilmicosin, 50% of the intracellular tilmicosin was effluxed within the first 30 min, but after 4 h of incubation in antibiotic-free medium, 30% remained cell-associated. Opsonized P. multocida significantly enhanced the release of tilmicosin from all three types of chicken phagocytes. Tilmicosin uptake was observed to increase lysosomal enzyme (acid phosphatase, lysozyme, avidin, and beta-glucuronidase) production. Finally, neutrophils were shown to transport and efflux bioactive tilmicosin in a test system measuring both neutrophil chemotaxis under agarose and a bioassay measuring inhibition of bacterial growth in the presence of antibiotic in agar. These in vitro observations of cellular pharmacology suggest a complex interaction between phagocytes and tilmicosin that contribute to clinical efficacy.

A non-invasive imaging modality, computed tomography (CT), was used to visualize changes in nasal turbinates of anesthetized pigs over a 12-week observation period (pigs were 14 wk of age at study week 0). Normal, non-infected pigs were compared to pigs with mild challenge-induced atrophic rhinitis (AR) in order to detect subtle differences in morphology. To determine feasibility for time course studies in future experiments, morphometric quantitation at the level of the 2nd premolar (turbinate area ratio or TAR) in cross-section CT images at multiple timepoints was done. Additionally, at study termination, the TAR determined from CT images, magnetic resonance imaging (MRI), and wet tissue (WT), were compared to each other and to the standard subjective measure, visual scoring. There were no statistically significant differences between the control and AR groups at CT imaging dates of 0, 3, 6, 9, or 12 wk (P = 0.182). However, a statistically significant decrease in TAR measurements over time (P = 0.015) was observed in both groups, with lower mean values observed on Weeks 3 and 6 before rebounding to baseline values at study termination. At Week 12 (termination of the study), the TAR measurements derived from CT, MRI, and WT were not statistically different from one another (P = 0.220) and the treatment group-by-method interaction was not significant (P = 0.800). This provided evidence of equivalency of the techniques. Mean values for normal and infected groups were not significantly different based on either TAR imaging methods (P = 0.552) or visual scores (P = 0.088). Thus, the current study demonstrated that CT was an acceptable alternative imaging modality which could be used for quantitation of turbinate changes in snouts of live pigs to provide data comparable to tissue taken at necropsy. Computed tomographic imaging would allow non-invasive tracking of disease or treatment responses within individual animals over time. Morphometric analysis of the TAR was equivalent between the CT, MRI, and WT specimens.

Tilmicosin is a semi-synthetic macrolide antibiotic, currently approved for veterinary use in cattle and swine respiratory disease. As the concentrations of tilmicosin are generally low in swine lung tissue, the interaction of tilmicosin with three types of swine phagocytes (monocyte-macrophages, alveolar macrophages, and neutrophils) was evaluated to provide an understanding of clinical efficacy. After incubation with radiolabelled tilmicosin, uptake was determined and expressed as the ratio of the intracellular (Ci) to the extracellular (Ce) drug concentration (Ci/Ce). Tilmicosin was avidly accumulated by the swine phagocytes (Ci/Ce 48-69 at 4 h incubation) with 51 to 85% localized in the lysosomes. Uptake was dependent on cell viability, temperature and pH, but was not influenced by the metabolic inhibitors, sodium cyanide or potassium fluoride. However, lipopolysaccharide (LPS) exposure increased tilmicosin uptake by the swine phagocytes. In neutrophils, upon removal of extracellular tilmicosin, 60% of the intracellular tilmicosin was effluxed within the first 30 min, but after 4 h of incubation in drug-free medium, 25% remained cell-associated. In contrast, after 4 h of incubation in drug-free medium, 60% and 45% of tilmicosin remained cell-associated, within alveolar macrophages and monocyte-derived macrophages, respectively. Tilmicosin uptake was observed to increase lysosomal enzyme (acid phosphatase, lysozyme and beta-glucuronidase) production. Finally, neutrophils were shown to transport and efflux bioactive tilmicosin in a test system measuring both neutrophil chemotaxis under agarose and a bioassay measuring inhibition of bacterial growth in the presence of antibiotic in agar. These in vitro interactions of tilmicosin with swine phagocytes suggest an integral role in effecting clinical efficacy.

Various environmental conditions likely to be encountered at a nidus of infection were evaluated for their effect on selected classes of antimicrobial agents. The minimum inhibitory concentration (MIC) of several aminoglycosides (apramycin, kanamycin, gentamicin, tobramycin, amikacin), tetracycline, and chloramphenicol for five strains of E. coli were unchanged by temperature (35 degrees-39.5 degrees C), atmosphere (aerobic to anaerobic), pH > 7, NaCl concentration (up to 150 mM), zinc concentration (up to 50 mM), and manganese (up to 10 mM). However, the aminoglycoside MICs were increased up to fivefold at pH < 6.5. Magnesium and calcium ion concentrations >10 mM and ferric iron concentrations >/=10 mM increased aminoglycoside MICs from 3.66- to 8-fold. Tetracycline MICs were increased 1.2- to 6.5-fold when the concentration of magnesium or calcium was >/=10 mM. The results of this in vitro study might provide insight into the effects of local in vivo environmental conditions on several classes of antimicrobial agents.

Tilmicosin is a novel macrolide antibiotic developed for exclusive use in veterinary medicine. The first tilmicosin-containing product was approved to treat bovine respiratory disease associated with pasteurellae. The development of antimicrobial susceptibility testing guidelines for tilmicosin was predicated on the relationship of clinical efficacy studies that demonstrated a favorable therapeutic outcome, on pharmaco-kinetic data, and on in vitro test data, as recommended by the National Committee for Clinical Laboratory Standards (NCCLS). The NCCLS-approved breakpoints for the MIC dilution testing are resistant > or = 32 micrograms/ml, intermediate 16 micrograms/ml, and susceptible < or = 8 micrograms/ml. The zone of inhibition interpretive criteria for disk diffusion testing with a 15-micrograms disk are resistant < or = 10 mm, intermediate 11-13 mm, and susceptible > or = 14 mm.

The acute phase reaction, in association with progressive atrophic rhinitis (AR), was monitored for 3 wk using serum haptoglobin (HPT) quantification in thirty-six, 15 kg swine after intranasal challenge with varying doses of Pasteurella multocida type D (toxigenic strain) and Bordetella bronchiseptica. The challenge doses were administered alone or in combination with pigs divided into 9 isolated treatment groups. Increasing doses of B. bronchiseptica were associated with lower serum HPT (P < 0.05), whereas increasing doses of P. multocida tended to increase serum HPT (0.05 < P < 0.10). Significant and positive correlation of mean HPT and AR score was found in these pigs; increased AR scores were associated with elevated mean HPT concentration (r = 0.41, P < 0.01). A significant interaction between P. multocida and B. bronchiseptica dose indicated that increasing the dose of B. bronchiseptica, for a fixed P. multocida dose, was associated with less AR (P < 0.05). The AR scores were greater in pigs given P. multocida, than B. bronchiseptica alone. These results indicate that a complex interaction between Pasteurella multocida and Bordetella bronchiseptica causes progressive atrophic rhinitis and alters serum HPT concentration in swine.

The two-fold purpose of this study was to establish a useful image analysis technique for quantitation of turbinate atrophy and to determine an optimum bacterial dose for inducing atrophic rhinitis (AR). Two morphometric analysis methods were compared to determine a turbinate area ratio (TAR) and a turbinate perimeter ratio (TPR); the ratios of turbinate area to total nostril area and of turbinate perimeter to total nostril perimeter, respectively. Our first image analysis method differed from Collins et al (1) in that we used direct image capture (digitalization) via a video camera and a Macintosh microcomputer, rather than photographs and a digitizer tablet. The tracing techniques were the same as those used by Collins et al. The second morphometric method was modified from the first by exclusion of dorsal turbinate when tracing the nostril area and directly tracing only the ventral turbinate to get a turbinate measurement without subtracting. Area and perimeter ratios, for both methods, were compared to conventional visual snout scores, ventral measurements, and to each other. The results of the two image analysis methods correlated well, both with each other and with the visual scores. Doses of Pasteurella multocida (Pm) at a constant level, and Bordetella bronchiseptica (Bb) at various concentrations, were administered to 36 Hampshire-Duroc F1 SPF pigs to determine the best dose and frequency for inducing AR. Although the dose selection may have been somewhat affected by the pre-existing presence of Bb, the optimal dose per naris in this study was 2 mL Bb at 10(7) cfu/mL combined with 2 mL Pm at 10(9) cfu/mL inoculum. The frequency of administration (1 x or 2 x) did not greatly affect results. Turbinate area ratio was the best tool for quantitating gross morphological turbinate changes associated with atrophic rhinitis in this study. Our simplified modification of Collins et al image analysis method (exclusion of dorsal turbinates and direct measurement of ventral turbinates) correlated well with visual scores, and, when compared to Collins et al method, required less data manipulation and labour.

A new fluorinated analog of tilmicosin was synthesized by the reductive amination of desmycosin with 3,5-bis(trifluoromethyl)piperidine. Despite an apparently small change in structure, the fluorinated analog had much less in vitro antimicrobial activity than tilmicosin and it failed to protect 3-day old chicks against a Pasteurella multocida challenge at 64 mg/kg sc. In a preliminary in vivo fluorine NMR experiment in a female Sprague-Dawley rat, a 19F NMR signal was detected in the liver one hour after ip administration of the fluorinated compound. Therefore, although this fluorinated derivative had less antimicrobial activity than tilmicosin, it may nevertheless provide a suitable model of tilmicosin for pharmacokinetic studies using in vivo fluorine NMR.

Five strains of Actinobacillus pleuropneumoniae serotype 1 were used to intranasally infect 5 groups of pigs. Using each bacterial strain, infected pigs (termed seeder pigs) were commingled for 48 hours with 5 groups of noninfected test pigs, then were removed. Seeder and test pigs were maintained in isolation and were observed for 14 days. Seeder pigs had mortality that was threefold greater than that of test pigs (24% vs 8%). Rectal temperature in excess of 40.3 C was achieved for 84% of test pigs and 88% of seeder pigs. Neither of these 2 variables was statistically different between the 2 groups of pigs. Clinical impression scores > or = 2 (on a 0 to 3 scale) were three-fold (64% vs 20%) greater for seeder than for test pigs (P < 0.05). The total number of bacterial isolations or nonrecoverable isolates was tabulated for test and seeder pigs' lungs at necropsy, irrespective of the amount of lesions. The number of A pleuropneumoniae isolations was not statistically different between test and seeder pig populations. Recovery of Pasteurella multocida or other bacteria was greater from the seeder pigs (P < 0.05), whereas the number of non-recoverable isolates was greater from test pigs than from seeder pigs (P < 0.05). Assessment of lung lesions at necropsy by either visual estimation or on a weight basis were in agreement.(ABSTRACT TRUNCATED AT 250 WORDS)

A factorial arrangement of tilmicosin and bentonite was evaluated for efficacy in broiler chickens infected with Mycoplasma gallisepticum and correlated to tilmicosin recovery in a feed assay method. Tilmicosin at 300-500 g/ton prevented development of airsacculitis. The addition of 2% bentonite to the ration caused tilmicosin at 300 g/ton to be ineffective in controlling air-sac lesions, whereas 400 and 500 g/ton were moderately effective. Six percent bentonite rendered tilmicosin completely ineffective at all dose levels. There was a direct correlation between the percentage of bentonite in the feed rations, the percentage of the tilmicosin recovered in the assay procedure, and the increased incidence of air-sac lesions.