Antigenicity of ovine strains of Anaplasma phagocytophilum grown in tick cells and ovine granulocytes.

Antigens prepared from ovine granulocytes and tick cells infected with ovine strains of Anaplasma phagocytophilum, the causative agent of tick-borne fever, were tested in respect of their suitability for the assay of antibodies in ovine sera by enzyme-linked immunosorbent assay (ELISA) and by indirect immunofluorescent antibody test (IFAT). Antigens prepared from tick cells were as sensitive and specific as those expressed in ovine granulocytes for the detection of specific antibodies by ELISA, but they failed to react in the IFAT with immune sera obtained from sheep previously infected with ovine strains of A. phagocytophilum.

The minimum inhibitory concentration of tilmicosin and tylosin for mycoplasma gallisepticum and Mycoplasma synoviae and a comparison of their efficacy in the control of Mycoplasma gallisepticum infection in broiler chicks.

The minimum inhibitory concentrations of tylosin tartrate and a new macrolid antimicrobial agent, tilmicosin, were assessed for six strains of Mycoplasma gallisepticum (MG) and three strains of Mycoplasma synoviae (MS) in vitro by the microbroth method. For four of the strains of MG, tilmicosin showed a slightly lower minimum inhibitory concentration (MIC) than did tylosin at both the initial reading (when pH 7.0 is first seen in the dilutions under test) and the final reading at 14 days of incubation. For one of the remaining strains, the MIC for tilmicosin was equal to or less than that for tylosin at the initial reading but greater at the final reading. For the other strain, the MIC for tilmicosin was greater than for tylosin, and for both of them the MICs were very much higher than for other strains. For the three strains of MS, there was little difference between the two drugs for one strain whereas the MIC for tilmicosin was slightly less for the other two groups. Groups of 30 chicks were infected with a virulent strain of MG and treated with either tylosin (0.5 g/liter) or tilmicosin (at concentrations of 0.125, 0.25 or 0.5 g/liter). One infected group was untreated and another group was uninfected and untreated. Clinical signs, mainly depression and nervous signs, were seen in two to five birds in the infected treated groups. In contrast, in the infected untreated group, 16 of 30 birds showed clinical signs. Mortality was significantly less in the infected treated groups compared with the infected untreated group (P < 0.001), and following infection there were significantly (P < 0.05) greater weight gains in the infected medicated groups. At necropsy the prevalence of gross lesions of the airsac walls was similar in all the infected medicated groups and was less than that for the infected unmedicated group. For the group on tylosin, MG was recovered from five chicks during life and from six dead chicks. The corresponding figures for the group receiving the lowest dose of tilmicosin were four for each; however, the organism was not recovered from the groups on the higher doses of tilmicosin either during life or from dead chicks. Serological results were negative for all groups except the infected untreated group, in which all three birds that were tested were positive.

A model for testing the efficacy of enrofloxacin (Baytril) administered to turkey hens in the control of Mycoplasma iowae infection in eggs and embryos.

In a preliminary experiment, a field infection with Mycoplasma iowae was simulated by inoculating turkey eggs with various doses of two strains of M. iowae immediately before incubation. The strain and dose chosen for further study were those that best multiplied and resulted in infection of embryos from which the organism could be isolated after 25 days of incubation. Ten turkey hens free from infection with mycoplasmae were housed in isolation. The hens were given enrofloxacin in the drinking water at a concentration of 50 ppm on 3 successive days, on two occasions at intervals of 14 days. Within 48 hours of lay, their eggs were each inoculated with 0.1 ml of the selected strain and dose (10(5) colony-forming units/ml) of M. iowae. M. iowae was recovered from almost all eggs laid by hens before the initial medication but not from any of the eggs laid for several days after each period of medication. Thereafter, the organism could be recovered from a high proportion of inoculated eggs. The treatment of infected turkey laying flocks with enrofloxacin at strategic periods might be helpful in the control of this Mycoplasma by limiting both vertical and horizontal transmission.

Cultivation of an ovine strain of Ehrlichia phagocytophila in tick cell cultures.

Ehrlichia phagocytophila (previously known as Cytoecetes phagocytophila) which causes tick-borne fever (TBF) in sheep and pasture fever in cattle in the UK and mainland Europe is transmitted by the temperate hard tick Ixodes ricinus. The disease in sheep is characterized by fever, leucopenia and immunosuppression. Studies on the pathogenesis and other aspects of the disease have been hampered because the organism has not been cultivated in continuous or primary cell culture systems. This paper describes the first successful cultivation of a European isolate of E. phagocytophila in two continuous cell lines, IDE8 and ISE6, derived from the temperate hard tick Ixodes scapularis. Once adapted to tick cell cultures the organism was serially sub-cultured in new cells by transferring small portions of infected cell suspension every 2 to 3 weeks. The identity of the organism was confirmed by polymerase chain reaction (PCR), with primers specific to the granulocytic ehrlichiae. Sequence analysis of the PCR products amplified from infected tick cells were shown to be identical with those amplified from the blood of sheep infected with the same strain of E. phagocytophila. A susceptible sheep inoculated with a third passage of the tick cell-adapted E. phagocytophila reacted with fever and rickettsiaemia 5 days later, thus satisfying Koch's postulates.

A comparison of Baytril, Tylosin and Tiamulin in the control of Mycoplasma iowae infection of turkey poults.

Comparison was made of the protection afforded by Baytril, Tylosin and Tiamulin for turkey poults infected with Mycoplasma iowae (MI) by injection of the organism into the lungs. Poults were infected at 2 days of age and treatment commenced 3 days later by including the appropriate drug in the drinking water. Following treatment with Baytril at 50 mug/ml for 5 days, MI was not isolated at 7, 14 and 19 days after infection from the oropharynx or cloaca, during life, but was recovered from five of 23 poults at necropsy. With the same concentration of Baytril, but for only 3 days, isolations of MI were made at 19 days post infection from the oropharynx and cloaca, from 2 of 23 live birds, but at necropsy at this time from 19/23 birds. From poults given Tylosin or Tiamulin (at 500 and 250 mug/ml respectively for 5 days) MI was isolated from a proportion of birds on all occasions during life, after infection, and from 19/21 and 18/21, respectively, at necropsy.

The production of Mycoplasma iowae infection of turkey poults suitable for monitoring antimicrobials.

In an attempt to produce a persistent infection with M. iowae (Mi) three separate trials were conducted using strain B 11/80, a virulent strain, strain M 012-118, a recent isolate of unknown virulence, and strain Iowae 695 (I 695), the type strain. In each trial groups of 2-day-old poults were infected via the oesophagus, trachea, cloaca and directly into the lungs. Isolation during life was attempted from the oropharynx and the cloaca, and at necropsy at the end of the experiment (21 days after infection) from the trachea, lungs and airsacs, and the brain. The highest proportion of isolations were made at necropsy from the lungs and air sacs, and trachea, from birds infected with B 11/80 or M 012-118 via the lungs or B 11/80 given via the trachea. During life the proportion of isolations was lower than at necropsy but highest, overall, with B 11/80 given via the lungs or trachea and isolated from the oropharynx, or administered via the cloaca and isolated form this site. Strain I 695 was rarely isolated whatever the route of infection. There were few recoveries following infection via the oesophagus with any strain and no mycoplasmas were isolated from the brain. For the production of MI infection suitable for monitoring antimicrobials in young poults we would recommend infection with a pathogenic strain of the organism directly into the lungs.

Granulocytic Ehrlichia infection in ixodid ticks and mammals in woodlands and uplands of the U.K.

The prevalence of infection with Ehrlichiae of the Ehrlichia phagocytophila genogroup (the granulocytic Ehrlichiae), in questing Ixodes ricinus ticks of U.K. upland and woodland habitats, was investigated by PCR. The prevalence of infection in the three feeding stages of I. ricinus indicated that granulocytic Ehrlichiae are transmitted transstadially with no, or inefficient, transovarial transmission. The presence of infected ticks in both habitats indicates that endemic cycles of granulocytic Ehrlichia (GE) infection are maintained by both domesticated sheep and by wild reservoirs, and coexist with endemic cycles of Borrelia burgdorferi infection. Moreover, demonstration, for the first time, of GE infection in engorged Ixodes trianguliceps ticks and blood collected from wild rodents, suggests that European wild rodents are competent reservoirs. GE infection prevalence in nymphal and adult I. ricinus was significantly greater in uplands than woodlands, which is consistent with ticks of all three feeding stages feeding on reservoir-competent sheep in uplands. In one woodland studied, pheasants are important hosts for nymphal I. ricinus but are incompetent or inefficient reservoirs, so reducing GE infection prevalence in I. ricinus ticks in this habitat. 16S rRNA sequences of GE from ticks of these U.K. habitats, showed a high degree of homology with those of granulocytic Ehrlichiae isolated from humans, but also showed some evidence of genetic diversity of granulocytic ehrlichiae in the U.K. The implications of these findings, for the taxonomy of granulocytic ehrlichiae and the potential for human infections to occur in the U.K., is discussed.

A comparison of the efficacy of danofloxacin and tylosin in the control of Mycoplasma gallisepticum infection in broiler chicks.

Groups of chicks were infected with a virulent strain of Mycoplasma gallisepticum (MG) and treated with either danofloxacin or tylosin while one infected group was left untreated and a further group was uninfected and untreated. Control of clinical signs and mortality was better with danofloxacin than tylosin and there was significantly (P < 0.05) greater weight gain with danofloxacin at 21 days after infection. However at necropsy the prevalence of lesions of the airsac walls was similar in both groups. MG was recovered from fewer live chicks for the first week following treatment with danofloxacin, but at 2 weeks and at necropsy, at the termination of the experiment, it was recovered from a similar proportion of birds in both treated groups. This was reflected also in the serological results at the end of the trial.