J F Timoney

University of Kentucky, Lexington, Kentucky, United States

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Publications (15)42.73 Total impact

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    ABSTRACT: Previous use of repeated nasopharyngeal swabbing and culture of Streptococcus equi showed that healthy carriers developed in more than 50% of 'strangles' outbreaks. The guttural pouches were the only detectable site of S. equi colonisation on endoscopic examination of horses during one of these outbreaks and S. equi was sometimes not detected by culture of nasopharyngeal swabs from carriers for up to 2 or 3 months before nasal shedding resumed sporadically. A more sensitive way of detecting S. equi on swabs from established guttural pouch carriers was therefore required. Conveniently selected 'strangles' outbreaks were investigated in detail using endoscopy, in order to develop and assess a suitable polymerase chain reaction (PCR) test. We report here 3 protracted 'strangles' outbreaks on different kinds of establishments in which between 29 and 52% of sampled horses were infected as detected by culture and/or PCR. Of the infected horses, between 9 and 44% were identified as carrying S. equi after clinical signs had disappeared and the predominant site of carriage was the guttural pouch. Prolonged carriage of S. equi, which lasted up to 8 months, did not cease spontaneously before treatment was initiated to eliminate the infections. The detection and isolation of the carriers, in conjunction with strict hygiene measures, apparently resulted in the control of the outbreaks and allowed the premises to return to normal activity. Comparing PCR and culture, many more swabs were found to be positive using PCR (56 vs. 30% of 61 swabs). Similar results were obtained for guttural pouch samples from 12 established carriers (PCR 76% and culture 59%). These results from repeated samples from relatively few animals need confirming using more long-term carriers. PCR can also detect dead organisms and is, therefore, liable to yield false positive results. Despite this drawback, it is argued that PCR provides a potentially useful adjunct to culture of nasopharyngeal swabs in the detection of asymptomatic carriers of S. equi following outbreaks of 'strangles'.
    Equine Veterinary Journal 12/2000; 32(6):515-26. DOI:10.2746/042516400777584721 · 2.37 Impact Factor
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    ABSTRACT: To determine whether repetitive sequence-based polymerase chain reaction (rep-PCR) could be used to differentiate Streptococcus equi isolates, to examine S equi isolates from throughout the world, and to determine whether a horse had > 1 subtype of S equi during an outbreak of disease. An initial group of 32 S equi isolates, 63 S equi isolates from various geographic areas, and 17 S equi isolates obtained during outbreaks of disease. An aliquot of S equi genomic DNA was amplified, using enterobacterial repetitive intergenic consensus primers. Gel electrophoresis was performed on 1.5% agarose gels, and a computed-assisted program was used to compare rep-PCR results. Use of these primers to analyze 100 ng of S equi genomic DNA resulted in patterns of 6 to 14 bands. The 32 initial isolates were separated into 7 rep-PCR subtypes. There were 30 rep-PCR subtypes found among 29 S equi isolates obtained from Minnesota, Michigan, Canada, and Australia and 34 S equi isolates obtained from Kentucky and other sources. Furthermore, the same clone was identified in several horses during an outbreak of disease. Infected horses on the same farm all had a single clone of S equi. Analysis of these results suggests that rep-PCR is useful for delineating S equi into rep-PCR subtypes. Results revealed that isolates with the same geographic source or similar date of collection did not always have the same rep-PCR subtype. A single clone of S equi usually predominated during an outbreak of disease.
    American Journal of Veterinary Research 06/2000; 61(6):699-705. DOI:10.2460/ajvr.2000.61.699 · 1.34 Impact Factor
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    ABSTRACT: Magnetic resonance (MR) scan technique and lesion detectability were evaluated using a newly developed spinal abscess model in the New Zealand White rabbit. To create the lesion, an epidural needle was inserted under fluoroscopic guidance in the lumbar region and advanced to penetrate the ligamentum flavum. Next, polyethylene tubing was fed through the needle into the epidural space. A mixed suspension of Staphylococcus aureus (Cowan I) and blue polystyrene microspheres then was injected. Lesions were evaluated by MR imaging in four animals at multiple time points (3, 6, and 9 days). Imaging was performed at 1.5 tesla using a surface coil. Precontrast T2-and T1-weighted scans were first obtained. The T1-weighted scans were acquired both with and without fat saturation, and were repeated after intravenous contrast administration. The contrast agent used was gadoteridol (gadolinium HP-DO3A or ProHance) at a dose of 0.3 mmol/kg. On prospective film review, postcontrast scans proved superior for lesion detection. A spinal abscess could be identified postcontrast in all cases, irrespective of the use of fat saturation. The next best imaging technique for lesion detection was the T2-weighted scan, with 5 of 8 lesions noted thereon. Visualization of lesion margins proved to be a primary factor in prospective lesion identification. Region of interest image analysis demonstrated the postcontrast scans to be superior to all precontrast scan techniques for conspicuity of the interface between the abscess and the compressed spinal cord, with these results statistically significant. The lesions were characterized histologically by infiltrates of heterophils into the meninges and outer spinal cord with accompanying mild hemorrhage, fibrin exudation, and bacterial colonies. The lesions in three animals were confirmed to be in the epidural space, with the lesion in one animal in the subdural space. The current animal model was developed to study spine infection and, specifically, imaging characteristics and lesion detectability on MR. With the increased use of epidural catheters for pain management and the large number of acquired immunodeficiency syndrome cases, epidural infection is becoming an increasingly important clinical problem. Imaging technique, in particular the use of intravenous contrast, is critical for lesion detection and evaluation.
    Investigative Radiology 05/1998; 33(4):246-55. DOI:10.1097/00004424-199804000-00008 · 4.44 Impact Factor
  • J Flanagan · N Collin · J.F. Timoney · T Mitchell · J.A Mumford · N Chanter ·
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    ABSTRACT: The haemolytic activity of Streptococcus equi, the cause of equine strangles, was characterized. Production of haemolysin in Todd Hewitt broth was dependent on an equine serum supplement and the logarithmic phase of growth after which activity declined sharply. RNA core also induced haemolysin production from cells harvested at the end of the logarithmic phase of growth. Haemolysis was not affected by cholesterol, was only slightly increased in reducing conditions and was completely inactivated by trypan blue, identifying the haemolytic activity as streptolysin S-like (SLS-like). Purification by hydroxyapatite and Sephacryl column chromatography yielded proteins of molecular weights of approximately 6000 and 17 000-22 000 Da with a 64-fold increase in specific activity. Low molecular weight proteins from the RNA core were still present in the purified toxin. Two non-haemolytic mutants were derived by conjugation with an Enterococcus faecalis-carrying transposon Tn916. Southern blots of HindIII digests of DNA revealed that one of the mutants contained three transposon insertions and the other just one. A lambda phage library of S. equi contained plaques whose haemolytic activity was enhanced by reducing conditions and inhibited by cholesterol, suggesting a streptolysin O-like (SLO-like) activity. However, haemolysin in culture sonicates of host E. coli in which the lambda phage insert was subcloned into plasmid (pUC18), was not affected by these conditions. Seven isolates of S. equi in medium without SLS-like inducers showed no SLO-like activity and no evidence for an SLO-like toxin could be found by immunoblotting with pneumolysin antiserum and monoclonal antibodies or by polymerase chain reaction with primers derived from sequences conserved between the SLO genes of Lancefield group A, C and G streptococci. S. equi does not appear to possess a streptolysin O but does make a streptolysin S-like toxin whose production can be interrupted at just one genetic locus.
    Microbial Pathogenesis 04/1998; 24(4):211-21. DOI:10.1006/mpat.1997.0190 · 1.79 Impact Factor
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    ABSTRACT: An experimental model of acute focal pyelonephritis was developed in the New Zealand White rabbit, with initial imaging evaluation performed using high-field contrast-enhanced magnetic resonance (MR) in six animals. The left kidney was visualized fluoroscopically after intravenous injection of iodinated contrast. A 0.1 mL mixture of agarose/Streptococcus faecalis was then injected percutaneously into the kidney at the corticomedullary junction with a 25-gauge needle. The animals were imaged at 1.5 tesla on day 5 after injection of bacteria. Breathhold T2-weighted and T1-weighted scans were acquired prior to contrast injection. Using an MR-compatible power injector, 0.3 mmol/kg gadoteridol was then administered, with both dynamic and delayed postcontrast scans obtained. The lesion was confirmed in each animal by gross and microscopic exam. The area of acute focal pyelonephritis was somewhat difficult to identify on precontrast scans. The lesion was mildly hyperintense on T2-weighted scans and slightly hypointense to isointense on T1-weighted scans relative to normal surrounding renal parenchyma. On early dynamic turbo-fast low-angle shot scans, the lesion could be identified due to differential tissue enhancement. Direct visualization of the lesion, with increased conspicuity relative to precontrast scans, was possible because of delayed positive enhancement at 60 seconds postcontrast. Lesion conspicuity, assessed by signal difference/noise ratio, increased from 0 +/- 5 precontrast to a peak of 12 +/- 6 at 60 seconds postcontrast (P = 0.003; n = 6). On dynamic contrast-enhanced MR, acute focal pyelonephritis can demonstrate transient positive contrast enhancement (using a contrast dose of 0.3 mmol/kg) relative to surrounding normal renal parenchyma. This appearance is due to hyperconcentration of the contrast agent in normal parenchyma and T2 effects. The pattern is opposite that seen in spiral computed tomography.
    Investigative Radiology 12/1997; 32(11):696-704. DOI:10.1097/00004424-199711000-00008 · 4.44 Impact Factor
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    ABSTRACT: The use of gadolinium (Gd)-BOPTA as a magnetic resonance contrast agent for central nervous system disease was studied in a canine brain abscess model. A Streptococcus faecalis brain abscess was evaluated in five dogs at 1.5T. Imaging was performed during the late cerebritis stage, at 5 to 7 days after surgery. Magnetic resonance scans were acquired before and at 1, 5, 15, 30, 45, and 60 minutes after contrast administration, using a dose of 0.1 mmol/kg. Scans also were acquired both before and after contrast injection with the implementation of magnetization transfer. Lesion enhancement, quantified by region-of-interest measurement, peaked at 5 minutes after contrast injection. Both the increase in lesion enhancement from 1 to 5 minutes after injection and the decrease from 5 to 15 minutes after injection, although small, were statistically significant (P < 0.004 and P < 0.03, respectively). The application of magnetization transfer improved lesion enhancement, as measured by signal difference/noise, by 39%. This result also was statistically significant (P < 0.001). In intraparenchymal brain infection, Gd-BOPTA provides effective lesion enhancement when used at a dose of 0.1 mmol/kg. Further research is needed to compare the magnitude of enhancement achieved with Gd-BOPTA, which has weak protein binding and both hepatobiliary and renal excretion, with that with Gd chelates, which have pure renal excretion.
    Investigative Radiology 06/1996; 31(5):294-9. DOI:10.1097/00004424-199605000-00009 · 4.44 Impact Factor
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    ABSTRACT: The ability of high-field (1.5 T) magnetic resonance imaging (MRI) to detect early brain meningitis was evaluated in a canine model. Contrast dose, timing postinjection, and imaging technique (specifically the use of magnetization transfer) were assessed. Imaging of five canines was performed at 1.5 T 24 hours after injection of Cowans staphylococcus into the cisterna magna. Two control animals also were imaged using the same protocol, with one animal receiving a cisternal injection of nutrient broth only and the other no injection. Contrast doses of 0.1, 0.3, and 0.8 mmol/kg gadoteridol (Gd HP-DO3A or Pro-Hance) were compared. Scans were performed at 2, 12, and 22 minutes after an initial injection of 0.1 mmol/kg. At each time point, paired T1-weighted scans with and without magnetization transfer (MT) were acquired. Thirty minutes after the initial injection of contrast, a supplemental dose of 0.2 mmol/kg was given (for a cumulative dose of 0.3 mmol/kg). Scans were then repeated at 2, 12, and 22 minutes after this dose was administered. A second supplemental contrast injection of 0.5 mmol/kg (for a cumulative dose of 0.8 mmol/kg) was given at 70 minutes, and immediate postinjection scans with and without MT were acquired. In the animals receiving a cisternal injection of bacteria, the degree of meningeal enhancement was greatest at 0.8 mmol/kg, intermediate at 0.3 mmol/kg, and least at 0.1 mmol/kg. These conclusions were constant whether imaging was performed with or without MT. Scans in control studies did not demonstrate abnormal meningeal enhancement. High-contrast dose, MT, and acquisition of immediate postcontrast scans all resulted in statistically significant improvement. On masked film review, abnormal meningeal enhancement was noted in only 2 of 5 experimental dogs at a dose of 0.1 mmol/kg (regardless of the use of MT) compared with all animals at a dose of 0.3 mmol/kg. In 18 of 37 dogs (paired scans with and without MT), when abnormal enhancement was noted, the use of MT improved the visualization of abnormal meningeal enhancement. In early brain meningitis, high-contrast dose (0.3 mmol/kg), MT, and scanning immediately after injection improve detection of abnormal meningeal enhancement, thus facilitating the diagnosis of meningitis. Of these factors, contrast dose is the most important.
    Investigative Radiology 09/1995; 30(8):484-95. DOI:10.1097/00004424-199508000-00006 · 4.44 Impact Factor
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    J F Timoney · J Walker · M Zhou · J Ding ·
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    ABSTRACT: Streptococcus equi subsp. zooepidemicus, a Lancefield group C streptococcus, is a frequently isolated opportunist pathogen from a variety of animal hosts, including the horse. Previous studies have indicated that equine strains carry antigens with characteristics of the antiphagocytic M proteins on the Lancefield groups A and G streptococci. We have cloned a protective M-like protein gene (SzPW60) of an equine strain of S. equi subsp. zooepidemicus W60 and determined its sequence. This gene encodes a protein with a molecular weight of 40,123 which protects mice against subsp. zooepidemicus but not subsp. equi, stimulates antibodies which opsonize subsp. zooepidemicus but not equi, and reacts with antiserum to the protein of the parent strain. The predicted amino acid structure shows significant homology with the carboxy termini of groups A and G M proteins but no other homology. The M-like protein, although showing an extensive region of alpha helix, lacks the A, B, and C repeats found in group A M proteins and has a shorter signal sequence. A proline-rich region upstream from the LPSTGE motif contains 20 repeats of the tetrapeptide PEPK. The presence of this repeat region may account for the slow migration of the M-like protein in sodium dodecyl sulfate-polyacrylamide gel electrophoresis.
    Infection and Immunity 05/1995; 63(4):1440-5. · 3.73 Impact Factor
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    A Efstratiou · G Colman · G Hahn · J F Timoney · J M Boeufgras · D Monget ·
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    ABSTRACT: Pyogenic streptococci of Lancefield group C or group G from human or animal sources were examined with a view to increasing the number of diagnostic tests useful for their differentiation. Human strains of group G produced L-prolyl-L-arginine aminopeptidase but isolates of Streptococcus equisimilis (group C) did not. Tests for alpha-L-glutamate aminopeptidase together with fermentation of glycogen or sorbitol distinguished S. dysgalactiae from strains of S. equisimilis isolated from animals. It was confirmed that fermentation tests were helpful in the study of S. equi and S. zooepidemicus and that enzyme reactions helped distinguish between S. canis and the human strains of group G.
    Journal of Medical Microbiology 09/1994; 41(2):145-8. DOI:10.1099/00222615-41-2-145 · 2.25 Impact Factor
  • John F. Timoney · Maowia M. Mukhtar ·
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    ABSTRACT: The group C streptococci are the most commonly isolated bacteria from disease states in the horse. Important virulence factors of S. equi and S. zooepidemicus are the hyaluronic acid capsule and the antiphagocytic fibrillar M protein located on the surface of the cell wall and extending into and through the capsule. The hyaluronic acid capsule is non-antigenic and so is not involved in protective immunity. The M protein, a superantigen, elicits very strong B and T cell responses that may result in protective immunity mediated by opsonic antibodies in plasma and by locally synthesized IgG and IgA on the pharyngeal mucosa. However, vaccines based on acid or mutanolysin extracted M protein do not confer a high level of protection against field exposure. Protective antibodies to S. equi or S. zooepidemicus can in part be assayed by the bactericidal test that measures opsonization for equine neutrophils. A mouse-challenge model has also been used to test immunizing potency of streptococcal extracts and in a passive protection test for protective antibody. There is as yet no means of distinguishing protective opsonic or mucosal antibodies from other antibodies produced against the many epitopes on the M molecule.
    Veterinary Microbiology 12/1993; 37(3-4):389-95. DOI:10.1016/0378-1135(93)90037-8 · 2.51 Impact Factor
  • John F. Timoney · Martin M. Groschup ·
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    ABSTRACT: Erysipelothrix rhusiopathiae is a widely distributed mucosal commensal of the alimentary tracts of vertebrates. Antibodies to a 66-64 kDa protein released from the cell surface have been shown to be involved in protective immunity. Mice immunized with the purified 66-64 kDa protein from strain T28, serotype 2b were protected against challenge by the United States challenge strain E1-6P (serotype 1a) and by the official German challenge strain Frankfurt 1 (serotype N). Thus, protection is not serotype specific, a result consistent with previous observations that polysaccharide, non-proteinaceous antigens are the type specific antigens useful in serotyping. The 66-64 kDa protein appears to be most immunogenic when complexed to glycolipid. This may be due to an adjuvant effect of polysaccharide antigens. Further studies on the correlation between antibody titers to the 66-64 kDa protein and protection in pigs, turkeys and mice in vivo should be helpful in developing a basis for an in vitro assay to replace the mouse protection test in vaccine testing.
    Veterinary Microbiology 12/1993; 37(3-4):381-7. DOI:10.1016/0378-1135(93)90036-7 · 2.51 Impact Factor
  • J F Timoney ·
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    ABSTRACT: No abstract is available for this article.
    Equine Veterinary Journal 12/1988; 20(6):392-4. DOI:10.1111/j.2042-3306.1988.tb01555.x · 2.37 Impact Factor
  • J F Timoney · Diana Eggers ·
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    ABSTRACT: An indirect test based on horse blood was used to study bactericidal responses of the horse to Streptococcus equi following infection or vaccination. Bactericidal antibody appeared in convalescent sera between two and four weeks and high titres were usually attained by eight weeks. Infection without clinical evidence of abscessation was also effective in eliciting strong bactericidal responses. Serum bactericidal activity of horses either recovered from strangles or immunised with commercial bacterin had declined eight months after vaccination. However, horses that developed strangles eight to 10 months after vaccination exhibited rapid and substantial increases in serum bactericidal activity. Groups of yearlings immunised with commercial S equi vaccines consisting either of M protein or bacterin developed clinical strangles within six months of vaccination although the majority of the animals had exhibited strong serum bactericidal activity a few weeks before occurrence of the disease. Similarly, a group of seven yearling ponies hyperimmunised with experimental vaccine, rich in M protein, were found to be highly susceptible to an intranasal challenge of 5 X 10(8) colony forming units of S equi, although their sera exhibited strong bactericidal activity at the time of challenge. These observations suggest that the role of serum bactericidal antibody in protection of the horse against strangles has been overrated.
    Equine Veterinary Journal 08/1985; 17(4):306-10. DOI:10.1111/j.2042-3306.1985.tb02505.x · 2.37 Impact Factor
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    J F Timoney · J Trachman ·
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    ABSTRACT: Immunologically reactive proteins in acid extracts and culture supernatants of Streptococcus equi were recognized through a combination of chromatographic and immunologic procedures. Both high- and low-molecular-weight components of each of these protein preparations were protective for mice and were, therefore, presumed to contain a variety of hydrolytic products or fragments of the M protein of S. equi. Convalescent horse sera that exhibited strong bactericidal activity for S. equi always reacted with polypeptides in the molecular weight range of 24,000 to 29,000, whereas preinfection sera did not. Rabbit antisera to affinity-purified S. equi protein also reacted with these polypeptides, as well as with a polypeptide of about 36,000 to 37,000 molecular weight. M protein in acid extract and culture supernatant did not cross-react in immunodiffusion, but rabbit antiserum to affinity-purified M protein from an acid extract of S. equi reacted strongly with culture supernatant proteins of approximate molecular weights of 67,000, 58,000, and 43,000. We suggest, therefore, that the M protein in culture supernatant is masked by other sequences that are removed by hot acid during preparation of acid extracts. Polypeptides common to acid extracts of S. equi and Streptococcus zooepidemicus were also identified. These polypeptides had molecular weights of about 55,000 and 31,000.
    Infection and Immunity 05/1985; 48(1):29-34. · 3.73 Impact Factor
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    ABSTRACT: Mucosal nasopharyngeal immunoglobulin A (IgA) andIgGresponsestoproteins ofStreptococcus equi were studied inhorses after theexperimental production ofstrangles. S.equi-specific IgAandIgGtiters in nasopharyngeal mucus weremuchhigher insamples fromanimals 1 to2weeksafter challenge thaninsamples fromcontrol animals. Although IgAwas themajorimmunoglobulin innasal mucus,there was more antibody activity associated withIgGasmeasured byradioimmunoassay. Greatdifferences between thespecificities of antibodies innasal mucusandinserumweredetected. IgAandIgGofmucusorigin recognized onlytwomajor proteins withmolecular weights ofabout 41,000 and46,000 inacidextracts ofS.equiandgaveno detectable reaction withculture supernatant proteins. Onlyoneprotein ofabout62,000 molecular weight was recognized inacid extracts ofan equine strain ofS.zooepidemicus. Incontrast, immunoglobulins inserum recognized a greatvariety ofproteins inculture supernatants andacidextracts ofS.equiandS.zooepidemicus whichdid notinclude those proteins recognized byimmunoglobulins inmucus.Thesefindings provide goodevidence for theindependence ofthelocal andsystemic immuneresponsesofthehorsetoS.equi. Horsesrechallenged shortly after recoveryfromthefirst infection were resistant tochallenge withan inoculum ofS.equi10times greater thanthat towhichthey were originally susceptible. Thisresistance appeared tobeindependent ofthe levels ofbactericidal antibody inserum.We therefore suggest thatimmunity toS.equiinfection ismediated bylocally produced nasopharyngeal antibodies.

Publication Stats

350 Citations
42.73 Total Impact Points


  • 1993-2000
    • University of Kentucky
      • Magnetic Resonance Imaging and Spectroscopy Center
      Lexington, Kentucky, United States
  • 1985-1988
    • Cornell University
      • Department of Microbiology
      Ithaca, New York, United States