16S rRNA-based amplicon analysis of changes in the bacterial population in the lesions of papillomatous digital dermatitis in dairy cattle after topical treatment with allyl isothiocyanate.

Papillomatous digital dermatitis (PDD) is a foot disease causing lameness in dairy cattle. It is regarded as a polymicrobial infection, although its etiology is not fully understood. PDD is treated by the topical or systemic administration of antibiotics such as lincomycin (LCM); however, the milk of the cows cannot be marketed during the treatment and withdrawal period due to the residual antibiotics in milk. Allyl isothiocyanate (AITC), an extract of Wasabia japonica (known as wasabi or Japanese horseradish) widely employed as a food additive, can be used as an alternative antimicrobial agent that overcomes this problem. We previously showed that AITC is as effective as LCM in PDD treatment. Here, using the samples obtained in the previous clinical study, we analyzed changes in the bacterial population in the PDD-associated microbiota after AITC treatment and compared those with that following LCM treatment by 16S ribosomal RNA (rRNA)-based amplicon analysis. Both treatments induced major changes in the bacterial population, and Treponema species, which have been regarded as the major causative agents of PDD, were efficiently eliminated by both agents. However, the AITC-treated samples exhibited higher diversity compared with pretreatment samples, but this trend was not observed for LCM treatment, probably reflecting different antibacterial activities of the two agents. Importantly, this analysis detected population changes before morphological changes in PDD lesions (clinical signs of healing) became evident, indicating that 16S rRNA-based amplicon analysis represents an efficient strategy for analyzing and monitoring the treatment efficiency of PDD as well as other polymicrobial diseases.

Removal of bovine digital dermatitis-associated treponemes from hoof knives after foot-trimming: a disinfection field study.

BACKGROUND: Bovine digital dermatitis (BDD) is an infectious foot disease found commonly in dairy herds. Foot-trimming is an important husbandry procedure for reducing the ensuing lameness; however, epidemiological, and microbiological studies have identified this as a risk activity for transmitting BDD. Three disinfectants have previously been identified in laboratory work as effective for removing viable BDD-associated Treponema spp., from hoof knife blades. The present study enrolled 133 dairy cattle with BDD lesions, and swabbed hoof knife blades before and after foot-trimming, and after knife disinfection with one of three disinfectants (1:100 FAM30®, 2% Virkon® and 2% sodium hypochlorite) to assess their efficacy under field conditions. RESULTS: Detection of BDD treponeme phylogroup DNA was undertaken by direct PCR of swabs, and viable treponemes were detected by PCR of swab cultures after 6 weeks' incubation. Where hoof knives did not contact the lesion, BDD-associated treponemes were detected after foot-trimming in 12/22 (54.5%) cases by direct PCR and 1/22 (4.5%) cases by PCR of cultured organisms. Where contact was made with the lesion, 111/111 (100%) samples taken after trimming were positive by direct PCR and 47/118 (39.8%) were positive by culture PCR. Viable organisms were identified in cultures from lesion stages M2, M3, M4 and M4.1. No viable organisms were detected after disinfection of hoof knives. CONCLUSIONS: Hoof knives post-trimming were frequently contaminated with BDD-associated treponeme DNA. Viable organisms were identified in cultures whether contact had been made between hoof knife and lesion or not, although contact clearly increased the frequency of detection of viable organisms. The three disinfectants tested were effective for removing viable organisms. The disinfection protocol used in this study should therefore be considered reliable for adoption as standard industry practice.

Antimicrobial susceptibility profiles of oral Treponema species.

Treponemes occur in the microflora of the dental plaque. Certain Treponema species that are frequently isolated from chronic periodontitis lesions are involved in its initiation and progression. In addition to mechanical instrumentation, antimicrobial agents are used as an adjunctive treatment modality for periodontitis. Despite its importance for successful antimicrobial treatment, information about susceptibility is limited for Treponema species. The aim of this study was to assess the susceptibility of Treponema denticola strains, Treponema socranskii, and Treponema vincentii to eleven antimicrobial agents. The minimum inhibitory and minimum bactericidal concentrations of these antimicrobial agents revealed strain-specific variation. Doxycycline, minocycline, azithromycin, and erythromycin were effective against all Treponema species tested in this study, whereas fluoroquinolones only exhibited an equivalent effectiveness on T. socranskii. The susceptibility of one T. denticola strain, T. socranskii, and T. vincentii to kanamycin was influenced by prior exposure to aerobic conditions. The susceptibility to quinolone drugs varied among strains of T. denticola, although they share an amino acid sequence identity of greater than 99% for DNA gyrase (type II topoisomerase) subunit A. In addition, an ATP-binding cassette (ABC) transporter inhibitor assay for T. denticola indicated that the transport of quinolone drugs is partially related to this transporter, although there may be parallel transport mechanisms. Our results provide important insights into antimicrobial agent-Treponema dynamics and establish a basis for developing an appropriate adjunctive therapy for periodontal disease.

In vitro susceptibility of contagious ovine digital dermatitis associated Treponema spp. isolates to antimicrobial agents in the UK.

BACKGROUND: Contagious ovine digital dermatitis (CODD) is an important cause of infectious lameness in sheep in the UK and Ireland and has a severe impact on the welfare of affected individuals. The three treponemal phylogroups Treponema medium/Treponema vincentii-like, Treponema phagedenis-like and Treponema pedis spirochaetes have been associated with clinical CODD lesions and are considered to be a necessary cause of disease. There are scant data on the antimicrobial susceptibility of the treponemes cultured from CODD lesions. OBJECTIVE: The aim of this study was to determine in vitro the miniumum inhibitory concentration/ minimum bactericidal concentration (MIC/MBC) of antimicrobials used in the sheep industry for isolates of the three CODD associated treponeme phylogroups T. medium/T. vincentii-like, T. phagedenis-like and T. pedis. ANIMALS: Twenty treponeme isolates; from 19 sheep with clinical CODD lesions. METHODS: A microdilution method was used to determine in vitro the MIC/MBC of 10 antimicrobial agents for 20 treponeme isolates (five T. medium/T. vincentii-like, 10 T. phagedenis-like and five T. pedis). The antimicrobials tested were penicillin G, amoxicillin, oxytetracycline, tilmicosin, lincomycin, spectinomycin, tylosin, tildipirosin, tulathromycin and gamithromycin. RESULTS: The treponeme isolates tested showed low MICs and MBCs to all 10 antimicrobials tested. They were most susceptible to gamithromycin and tildipirosin (MIC90: 0.0469 mg/L), and were least susceptible to lincomycin, spectinomycin and oxytetracycline (MIC90: 48 mg/L, 24 mg/L and 3 mg/L, respectively). CONCLUSIONS: These data are comparable to in vitro antimicrobial susceptibility data for treponemes cultured from bovine digital dermatitis lesions. Dependent on local licensing, penicillin and tilmicosin appear to be the best candidates for future in vivo studies.

Short communication: minimum bactericidal concentration of disinfectants evaluated for bovine digital dermatitis-associated Treponema phagedenis-like spirochetes.

The bacterial spirochetes, Treponema spp., are thought to be a major contributor to the etiology of bovine digital dermatitis (DD), a skin disease with worldwide economic impact. Hoofbath strategies are commonly used in an attempt to control and prevent the development of DD and continuing research has been done to develop an optimal hoofbath strategy for this purpose. The aim of this study was to develop a protocol that can be used as part of the screening process for candidate hoofbath disinfectants. This protocol allows an accurate determination of the in vitro minimum inhibitory concentration and minimum bactericidal concentration of a series of disinfectants for Treponema microorganisms. Assays were performed in triplicate for each of the disinfectants at 30-s and 10-min exposure times and exposed to 10 and 20% manure (vol/vol). The results of this study can be used to categorize disinfectants based on the effect of exposure and manure concentration regarding their ability to inhibit Treponema growth. This information can then aid in optimizing strategies for hoofbath-based control of DD development and spread.

Antimicrobial susceptibility of Treponema phagedenis-like spirochetes isolated from dairy cattle with papillomatous digital dermatitis lesions in Japan.

The minimal inhibitory concentrations (MICs) of 23 Treponema phagedenis-like spirochetes isolated from dairy cattle with papillomatous digital dermatitis (PDD) lesions in Japan were investigated by a broth microdilution method using 15 antimicrobial agents. Although all MIC values showed a monomodal distribution, the MICs of the antimicrobial agents for 90% (MIC(90)) of the isolates tested varied among the agents examined. The MIC(90) values for penicillin G, ampicillin, and erythromycin were <0.06 microg/ml. In contrast, the MIC(90) values for kanamycin, streptomycin, rifampicin, sulfamethoxazole, trimethoprim, and colistin were >128 microg/ml. Oxytetracycline, lincomycin, enrofloxacin, chloramphenicol, ceftiofur, and gentamicin showed intermediate values, i.e., 0.5~32 microg/ml. The present study suggested that no isolate had acquired resistance to the antimicrobial agents examined, although they may have natural resistance to some agents. Furthermore, the in vitro antimicrobial susceptibility data would provide helpful information for PDD treatment and the development of a selective medium for isolating the organism effectively.

In vitro susceptibility of bovine digital dermatitis associated spirochaetes to antimicrobial agents.

Bovine digital dermatitis (BDD) is an infectious lameness in cattle, which has a large global impact in terms of animal welfare and cost. The majority of evidence suggests that spirochaetes are the aetiological agent of this disease. The aim of this study was to identify the susceptibility of BDD associated spirochaetes to a range of antimicrobial agents with a view to potential usage in vivo to treat this widespread cattle disease. A microdilution method was adapted to determine the in vitro susceptibilities of 19 UK digital dermatitis spirochaetes (6 Treponema medium/Treponema vincentii-like, 8 Treponema phagedenis-like and 5 Treponema denticola/Treponema putidum-like) to eight relevant antimicrobials. The BDD spirochaetes exhibited the highest susceptibility to penicillin and erythromycin and this information may now be used to aid development of efficacious treatments. This study has also identified that BDD spirochaete T167 is spectinomycin resistant and that the likely biological basis is a point mutation in the 16S rRNA gene. Interestingly, nearly all Brachyspira isolate 16S rRNA gene sequences in Genbank have this substitution, suggesting it may be responsible for the characteristic spectinomycin resistance reported for the Brachyspira genus.

A randomised clinical study to determine the effect of a toothpaste containing enzymes and proteins on plaque oral microbiome ecology.

The numerous species that make up the oral microbiome are now understood to play a key role in establishment and maintenance of oral health. The ability to taxonomically identify community members at the species level is important to elucidating its diversity and association to health and disease. We report the overall ecological effects of using a toothpaste containing enzymes and proteins compared to a control toothpaste on the plaque microbiome. The results reported here demonstrate that a toothpaste containing enzymes and proteins can augment natural salivary defences to promote an overall community shift resulting in an increase in bacteria associated with gum health and a concomitant decrease in those associated with periodontal disease. Statistical analysis shows significant increases in 12 taxa associated with gum health including Neisseria spp. and a significant decrease in 10 taxa associated with periodontal disease including Treponema spp. The results demonstrate that a toothpaste containing enzymes and proteins can significantly shift the ecology of the oral microbiome (at species level) resulting in a community with a stronger association to health.

Molecular characterization of the gyrB region of the oral spirochete, Treponema denticola.

The nucleotide (nt) sequence of the Treponema denticola (Td) DNA gyrase beta-subunit gene (gyrB) has been determined. Southern blot analysis of Td chromosomal DNA indicated that gyrB is present as a single copy. Approximately 3.2kb of the nt sequence 5' and 0.7kb of nucleotide sequence 3' of gyrB were obtained. Analysis of the deduced amino acid (aa) sequence revealed two complete open reading frames (ORFs) (ORF1 and ORF3) and a truncated ORF (ORF4'). ORF1 has no homology to sequences in the databases, whereas ORF3 and ORF4' have significant homology to several bacterial DnaA (replication initiator) and DnaE (DNA polymerase III) proteins respectively. RT-PCR data showed that orf1-gyrB are co-transcribed, while dnaA-dnaE are co-transcribed but in the opposite direction. These data indicated that the gene organization of the Td gyrB region is unique compared with that of other bacteria. Eighteen putative DnaA boxes with several AT-rich regions were identified in the dnaA-dnaE intergenic region, and three putative DnaA boxes were identified in the gyrB-dnaA intergenic region. Spontaneous coumermycin A(1)-resistant Td mutants were isolated and characterized. The mutants have a >20-fold higher resistance to coumermycin A(1) than wild-type Td. A single point mutation in gyrB that changed GyrB Lys(136) to Glu or Thr appears to be responsible for the coumermycin A(1) resistance.

Synthesis and antibacterial activity of some 3-[(alkylthio)methyl]quinoxaline 1-oxide derivatives.

Some 3-[(alkylthio)methyl]quinoxaline 1-oxide derivatives (1) have been synthesized and screened for antibacterial activity. 2-Acetyl-3-[(methylsulfonyl)methyl]quinoxaline 1-oxide (7a) was found to possess good in vitro activity against some pathogens important to veterinary medicine including Treponema hyodysenteriae, a causative agent in swine dysentery. In an in vivo experiment, this compound (7a) completely protected pigs against a swine dysentery challenge over a 21-day period.

23S rRNA point mutation associated with erythromycin resistance in Treponema denticola.

Mechanisms and occurrence of macrolide resistance in the periodontal pathogen Treponema denticola have received little attention. In this study, erythromycin resistance due to mutations in the genes encoding T. denticola 23S rRNA was investigated. The T. denticola genome was shown to contain two copies of 23S rDNA. 23S rRNA genes of nine erythromycin-resistant isolates derived from T. denticola were amplified and sequences were analyzed. All the erythromycin-resistant strains had at least one A-->G transition mutation at the 23S rRNA gene sequence cognate to position A2058 in Escherichia coli 23S rDNA. This suggests that antibiotic pressure is sufficient to select for point mutations that confer resistance in this organism.

Stimulation of in vitro growth of Treponema denticola by extracellular growth factors produced by Porphyromonas gingivalis.

Cell-free culture filtrates of Porphyromonas gingivalis grown in Wilkins-Chalgren broth stimulated the growth of six strains of Treponema denticola in 1186 broth when compared with the effect of uninoculated WC. The pH of the 1186 broth was not altered by the addition of either culture filtrate or WC, and all media were fully reduced prior to inoculation with T. denticola. Growth was also stimulated by factors precipitated from the culture filtrate with 90% (NH4)2SO4, 50% cold ethanol, or 50% cold acetone, and by factors retained after dialysis of the culture filtrate through a membrane with a molecular weight cut-off of 50 kDa. Growth factor activity was eliminated by heating of the culture filtrate at 55 degrees C for 4 h. An ether extract of the culture filtrate containing acetic, butyric, isobutyric, isovaleric, propionic, and phenylacetic acids did not stimulate growth. Since subgingival plaque from periodontal pockets colonized with T. denticola also contains P. gingivalis, certain extracellular proteins with molecular weights greater than 50 kDa produced by P. gingivalis may act as growth factors for T. denticola in the microenvironment of the periodontal pocket.

The utility of the BANA test for monitoring anaerobic infections due to spirochetes (Treponema denticola) in periodontal disease.

Treponema denticola, Porphyromonas gingivalis, and Bacteroides forsythus each possesses an enzyme(s) that hydrolyzes the synthetic substrate benzoyl-DL-arginine-naphthylamide (BANA). The presence of these organisms in a subgingival plaque sample can be determined by the ability of the plaque to hydrolyze BANA. In the present study, we describe the usefulness of the BANA test at various stages of a clinical trial of the efficacy of metronidazole in the treatment of periodontal disease. A BANA-positive test was significantly associated with high levels and proportions of spirochetes in the plaque, so that it provided information comparable with that which could be obtained by a microscopic examination of the plaque. Patients with such anaerobic spirochetal infections were randomly assigned to a group receiving either metronidazole or placebo (250 mg, three times a day) for one week and whose teeth were scaled and root-planed. The advantages of the decision that metronidazole be used were apparent from the comparison with the results obtained in the patients who received only the scaling and root planing. The initially BANA-positive teeth in the patients treated with metronidazole, scaling, and root planing gained attachment and exhibited a significant reduction in the need for periodontal surgery, when compared with the BANA-positive teeth in the patients who received only placebo, scaling, and root planing. After the conclusion of this therapy, those teeth with persistent BANA-positive plaques had significantly higher proportions and levels of spirochetes than did the teeth with BANA-negative plaques.(ABSTRACT TRUNCATED AT 250 WORDS)

Effects of the inorganic salts sodium chloride, sodium bicarbonate, and magnesium sulfate upon the growth and motility of Treponema vincentii.

The use of inorganic salt solutions as chemotherapeutic agents in the control of periodontal disease has received considerable attention in the past few years. Although some research has been published on their clinical effectiveness, little is known about their therapeutic activity and bactericidal effects upon oral spirochetes. The present study investigated the effects of varied concentrations of NaCl, NaHCO3, and MgSO4 upon the in vitro growth and motility of Treponema vincentii. Growth determinations were performed using a turbidiometric analysis at 545 nm. Motility was qualitatively studied by direct examination of 200 treponemes in a wet mount specimen. Samples were taken at 24, 48, 72, and 96 hours following inoculation with the treponemes. Concentrations of 0.5 M NaCl, NaHCO3, or MgSO4 totally inhibited the growth and motility of T. vincentii over a 96-hour period. Salt concentrations less than or equal to 0.10 M had little if any effect upon growth and motility. The data support the hypothesis that the bactericidal and antimotility effects of these salts are related more to their concentrations than to the presence of a specific inorganic ion. They also suggest that motility may be a valid indicator of bacterial viability. Before the clinical significance of the results can be ascertained, human studies are needed to establish sulcular salt concentrations which can be achieved with local irrigation and to determine how long bactericidal concentrations can be maintained.

A 15-month evaluation of the effects of repeated subgingival minocycline in chronic adult periodontitis.

BACKGROUND: A double-blind, randomized, parallel, comparative study was designed to evaluate the long-term safety and efficacy of subgingivally administered minocycline ointment versus a vehicle control. METHODS: One hundred four patients (104) with moderate to severe adult periodontitis (34 to 64 years of age; mean 46 years) were enrolled in the study. Following scaling and root planing, patients were randomized to receive either 2% minocycline ointment or a matched vehicle control. Study medication was administered directly into the periodontal pocket with a specially designed, graduated, disposable applicator at baseline; week 2; and at months 1, 3, 6, 9, and 12. Scaling and root planing was repeated at months 6 and 12. Standard clinical variables (including probing depth and attachment level) were evaluated at baseline and at months 1, 3, 6, 9, 12, and 15. Microbiological sampling using DNA probes was done at baseline; at week 2; and at months 1, 3, 6, 9, 12, and 15. RESULTS: Both treatment groups showed significant and clinically relevant reductions in the numbers of each of the 7 microorganisms measured during the entire 15-month study period. When differences were detected, sites treated with minocycline ointment always produced statistically significantly greater reductions than sites which received the vehicle control. For initial pockets > or =5 mm, a mean reduction in probing depth of 1.9 mm was seen in the test sites, versus 1.2 mm in the control sites. Sites with a baseline probing depth > or =7 mm and bleeding index >2 showed an average of 2.5 mm reduction with minocycline versus 1.5 mm with the vehicle. Gains in attachment (0.9 mm and 1.1 mm) were observed in minocycline-treated sites, with baseline probing depth > or =5 mm and > or =7 mm, respectively, compared with 0.5 mm and 0.7 mm gain at control sites. Subgingival administration of minocycline ointment was well tolerated. CONCLUSIONS: Overall, the results demonstrate that repeated subgingival administration of minocycline ointment in the treatment of adult periodontitis is safe and leads to significant adjunctive improvement after subgingival instrumentation in both clinical and microbiologic variables over a 15-month period.

Local metronidazole application in maintenance patients. Clinical and microbiological evaluation.

The purpose of this investigation was to evaluate the clinical and microbiological effect of local antibiotic therapy in comparison with subgingival scaling and root planing in a randomized semi-masked study. Forty-six recall patients who completed systematic periodontal therapy 6 to 24 months prior to the study were enrolled. The inclusion requirements were at least one site with probing depth > or = 5 mm in each quadrant, no scaling, and no antibiotic therapy during the last 6 months. After randomization each patient received 2 different treatments: in 2 quadrants metronidazole 25% dental gel was applied subgingivally to the pockets at day 0 and day 7; scaling and root planing was carried out in the 2 other quadrants, one at day 0 and in the remaining quadrant at day 7. Subgingival microbiological samples were taken from each patient before treatment and on days 21, 91, and 175 after the treatment. The analyses were carried out by indirect immunofluorescence assay. At all treated sites probing depth (PD), clinical attachment level (CAL), and bleeding on probing (BOP) were recorded on days 0, 21, 91, and 175. Both treatments resulted in PD reduction and CAL gain. PD reduction was statistically significant (P < 0.01) for both treatment modalities after 6 months. The CAL gain was not significant for either treatment. There was no statistical significance between scaling and antibiotic therapy. Treponema denticola, Porphyromonas gingivalis, and Prevotella intermedia were significantly reduced after therapy; however, there were no statistically significant differences between treatments. If Actinobacillus actinomycetemcomitans was present before therapy, it was also present after treatment in both groups. The conclusion is that, in recall patients, local application of metronidazole and scaling and root planing showed similar clinical and microbiological effects without statistically significant differences.

Short-term microbiological and clinical effects of subgingival irrigation with an antimicrobial mouthrinse.

Fifty chronic adult periodontitis patients completed a 6-week controlled, double-blind, split mouth clinical study to determine the effects of subgingival irrigation with an antimicrobial mouthrinse on periodontal microflora, supragingival plaque, and gingivitis when used as an adjunct to normal oral hygiene. Qualifying subjects had at least four sites, two on each side of the mouth, with probing depths between 4 and 6 mm, which bled on gentle probing. Following baseline examinations, subjects received a half mouth scaling and prophylaxis and full mouth subgingival irrigation with either the antimicrobial mouthrinse or sterile colored water control professionally delivered. Subjects continued irrigation at home once daily for 42 days with their assigned rinse delivered via a subgingival delivery system. All sites in the mouth were scored at baseline and at day 42 for supragingival plaque, bleeding on probing, and redness. For the four selected periodontitis sites, probing depth and attachment level were measured at baseline and on day 42; additionally, supragingival plaque and gingival redness were scored on days 7 and 21. Subgingival plaque samples for microbiological analysis were harvested from the selected periodontal sites at baseline and on days 7, 21, and 42. Microbiologically, irrigation with the antimicrobial mouthrinse resulted in statistically significant reductions compared to control in putative periodontopathogens, including black pigmenting species, which persisted at 42 days. Clinically, subgingival irrigation with the antimicrobial mouthrinse produced a significant reduction in supragingival plaque (P < 0.001), bleeding on probing (P = 0.019), and redness (P = 0.017) compared to the control, whether or not the area irrigated received a prophylaxis.(ABSTRACT TRUNCATED AT 250 WORDS)

Inhibition of protease activities of periodontopathic bacteria by extracts of plants used in Kenya as chewing sticks (mswaki).

Extracts from five plants used as chewing sticks, and tannic acid, gallic acid and methyl ester of gallic acid, were tested for their ability to inhibit proteolytic activities of three strains of Bacteroides gingivalis, three strains of Bacteroides intermedius and two strains of Treponema denticola. Aqueous extract from the plants Rhus natalensis and Euclea divinorum were the most inhibitory of those tested, inhibiting by 50% the proteolytic activity of the test organisms, at concentrations of up to 200 micrograms/ml. Tannic and gallic acids had similar effects at concentrations of less than 10 micrograms/ml, while the methyl ester of gallic acid was less inhibitory. These findings suggest that extracts from plants used as chewing sticks may possess enough inhibitory components to interfere with the virulence and growth of periodontopathic bacteria in vivo, provided they are able to gain access to the subgingival sites such bacteria preferentially inhabit.

Isolation and characterization of a novel polyether antibiotic of the pyrrolether class, antibiotic X-14885A.

Antibiotic X-14885A is a polyether antibiotic belonging to the class of these natural acid ionophores known as pyrrolethers. The structure of the antibiotic was elucidated by X-ray crystallographic analysis of the hydrated sodium salt, which crystallized as a tetramer containing four antibiotic and water molecules and four atoms of sodium. Antibiotic X-14885A differs from the most well-known member of the class, A-23187, in two respects: the aromatic N-methylamino group present in the latter is replaced by a phenolic hydroxyl, and one of the four aliphatic methyls is replaced by a proton. Antibiotic X-14885A is active against Gram-positive bacteria and the spirochete, Treponema hyodysenteriae.

Minimal inhibitory concentrations of five antimicrobials against Treponema hyodysenteriae and Treponema innocens.

The minimal inhibitory concentrations of carbadox, dimetridazole, lincomycin, ronidazole, and tiamulin against isolates of Treponema hyodysenteriae and Treponema innocens were determined by an agar-dilution method. The results obtained indicated that tiamulin was the most effective antimicrobial in vitro against T. hyodysenteriae, followed by carbadox. Dimetridazole, lincomycin, and ronidazole had poor efficacy in vitro against the T. hyodysenteriae isolates. Isolates of T. innocens were more sensitive to the various antimicrobials. Carbadox and tiamulin were the most effective in vitro, followed by ronidazole, dimetridazole, and lincomycin.