F. T. W. Jordan

Demonstration of sites of latency of infectious laryngotracheitis virus using the polymerase chain reaction.

Mature laying chickens were inoculated intratracheally with a field strain of infectious laryngotracheitis (ILT) virus. Tracheal swabs were collected regularly from all birds for virus culture. At various times post-inoculation, pairs of birds were killed and tissues removed for detection of virus products using conventional tissue homogenization and culture, organ culture, indirect immunofluorescence (IF) and also the polymerase chain reaction (PCR). The latter was used to detect a DNA sequence from the ILT virus thymidine kinase gene. Following inoculation the birds developed mild respiratory disease with clinical signs characteristic of ILT from 3 to 10 days post-inoculation. Trachea and turbinate tissues were virus-positive as determined by virus isolation, organ culture, IF and PCR on day 4 post-inoculation. After recovery from the acute phase, virus shedding initially ceased, then intermittent, low level shedding was recorded for five of the six remaining birds. In an attempt to locate sites of latency, pairs of birds were sampled at 31, 46 and 61 days post-inoculation. Virus was not detected in upper respiratory tract or ocular tissues by conventional techniques, or in the trigeminal, proximal and distal ganglia. All tissues were also negative by PCR, except for the trigeminal ganglia of five of the six birds. All PCR-positive birds had previously shed ILT virus intermittently between days 19 and 59 post-inoculation. As we did not detect viral DNA in any of the other tissues sampled from clinically recovered birds, we conclude that the trigeminal ganglion is the main site of latency of ILT virus.

Latency and reactivation of infectious laryngotracheitis vaccine virus.

SummaryLatency and reactivation of a commercial infectious laryngotracheitis virus vaccine were demonstrated in live chickens. Virus was re-isolated at intervals between seven and fourteen weeks post-vaccination and this may be of epizootiological significance.

Studies on the adsorption of certain medium proteins to Mycoplasma gallisepticum and their influence on agglutination and haemagglutination reactions

Serum proteins from Mycoplasma gallisepticum culture medium could be detected on the organisms as a result of incubation at low pH. Only certain of the serum proteins, including IgG and IgM, were found, and the adsorption appears to depress the haemagglutinating activity of the organisms. There was no obvious effect on slide agglutination (SA) sensitivity but an incidental finding was that brief acid treatment enhanced the SA sensitivity of an antigen prepared from a young culture.

Effects of Baytril, Tylosin and Iamulin on avian mycoplasmas

The minimum inhibitory concentration (MIC) of Baytril, Tylosin and Tiamulin for strains of Mycoplasma gallisepticum (MG) M. synoviae (MS), M. meleagridis (MM) and M. iowae (MI) and serovars were compared. In general the lowest MIC for MG, MS and MI was obtained with Baytril, while for MM both Baytril and Tiamulin gave the lowest MICs. Protection against mortality was best attained with Baytril for broiler chicks and poults but against prevention of growth depression Baytril was best for broiler chicks whilst Baytril and Tylosin were equally effective in poults. MG was recovered from fewer birds following treatment with Tiamulin and then Baytril in that order. Baytril also had a suppressive effect on the natural infection of MI in poults. Fewer poults showed rapid serum agglutination reactions to MG antigen following Baytril treatment than with the other antimicrobials. The tolerance of turkey embryos for Baytril was between 1 and 2 mg/ egg. Baytril reduced MG infection of turkey embryos.

Avian mycoplasma and pathogenicity ‐ A review

M. gallisepticum, M. synoviae and M. meleagridis are pathogenic strains of avian mycoplasma. M. gallisepticum and M. synoviae affect the chicken and turkey while M. meleagridis is associated with disease in the turkey. The tissues affected and the lesions are described and the influence on disease production of certain factors is considered. They include the age of the host, route of infection, numbers and virulence of organisms and predisposing and debilitating factors. The mechanism of pathogenesis is also indicated and reference made to a method of determining virulence.

The influence of pH of the culture medium on the sensitivity of Mycoplasma gallisepticum antigens for use in certain serological tests.

Mycoplasma gallisepticum antigens were prepared from organisms cultured in broth medium with glucose. The influence of period of growth, pH of the medium and duration of incubation at low pH (5.0) on the sensitivity of these antigens was determined in certain tests. The most sensitive antigen for the serum plate test was harvested after no more than 8 hr. incubation at pH 5.0. Sensitivity in serum plate, haemagglutination and gel diffusion tests was impaired if organisms were incubated at pH 5.0 for long periods. Antigens prepared from buffered broth medium were found to be at least as sensitive as those from unbuffered medium for the haemagglutination and gel precipitation tests, but considerably less so for the serum plate test.

The minimum inhibitory concentration of kitasamycin, tylosin and tiamulin for Mycoplasma gallisepticum and their protective effect on infected chicks

The minimum inhibitory concentration (m.i.c.) of kitasamycin, tylosin and tiamulin for Mycoplasma gallisepticum (Mg) were compared with 10(6), 10(4), and 10(2) CFU/ml of the organisms with the drug incorporated in mycoplasma agar. The lowest m.i.c. was obtained with tiamulin and the highest with kitasamycin and, in general, the m.i.c.s were directly influenced by the concentration of mycoplasmas. Chick embryos at 19 days of incubation were infected with Mg and the hatched infected chicks were used for comparing the protective effect of the three drugs. They were given in the drinking water when the chicks were 2, 3 and 4 days of age. Weight gains for the infected treated birds were similar for all three drugs; they were significantly lower than for those of uninfected chicks and significantly higher than for those of untreated infected chicks. However Mg could be isolated from a high proportion of chicks in the infected treated and untreated groups at 32 days of age.

Molecular epidemiology of a reproductive tract-associated colibacillosis outbreak in a layer breeder flock associated with atypical avian pathogenic Escherichia coli.

The molecular epidemiology of 70 Escherichia coli isolates from an infection outbreak in a layer breeder flock was examined by pulsed-field gel electrophoresis and for a range of virulence factors by polymerase chain reaction. Pulsed-field gel electrophoresis showed 35 of 45 isolates from eight disease cases were associated with a single clonal group that was the exclusive strain associated with reproductive tract. A second unrelated group was found in environmental isolates and healthy birds. The remaining isolates were unrelated to each other or either clonal group. Polymerase chain reaction virulotyping indicated the “epidemic” clonal group contains virulence factors including iss, sfa, tsh, iucC, ibeA, and sitA associated with avian pathogenic E. coli plus several virulence factors more normally associated with human urinary tract infection. Significantly, the “epidemic” clone was also found in an environmental sample, suggesting it may have been transmitted to the flock via the environment.

Mycoplasma anseris sp. nov. Found in Geese

A mycoplasma designated strain 1219T (T, type strain) was isolated from the phallus of a gander in Hungary. It was assigned to the class Mollicutes, order Mycoplasmatales, on the basis of morphological, cultural, and physical studies. The base composition of its deoxyribonucleic acid was 25 mol% guanine plus cytosine. It was dependent on sterol for growth, and its growth was inhibited by digitonin. The organism was assigned to the genus Mycoplasma since it did not hydrolyze urea and there was no evidence of helical forms. It hydrolyzed arginine, but other biochemical tests were negative. Strain 1219T could not be identified as any of 77 accepted Mycoplasma species by growth inhibition, immunofluorescence, or metabolism inhibition tests and thus appeared to be a new species. We propose the name Mycoplasma anseris for this organism, for which the type strain is strain 1219.

The response of chickens to experimental infection with strains of M. gallisept1cum of different virulence and M. gallinarum

Three strains of M. gallisepticum comprising S6 of low passage (virulent), S6 of high passage and A514 of high passage in artificial media and one M. gallinarum strain of high passage, were inoculated into 18-day-old chick embryos. Four groups of infected chicks hatched, one corresponding to each mycoplasma strain; these were investigated during the first 28 days of life and again for a period in lay. During the first 28 days of life, the virulent S6 strain was recovered more frequently from a wider variety of tissues for a longer time than were the other two strains of M. gallisepticum; this strain also produced more severe clinical disease including nervous signs, respiratory lesions and swollen hocks. Although nervous signs were confined to birds infected with the virulent S6, both S6 strains were isolated from brains and hocks, suggesting this strains proclivity for these sites. One A514 infected chick developed unilateral eye enlargement from which mycoplasma were isolated. All strains of mycoplasma were isolated from respiratory tissues, indicating a proclivity common to all. Recoveries of mycoplasma were often associated with signs and/or lesions but not always, and recoveries also occurred in the absence of signs and lesions. M. gallinarum caused no clinical signs or lesions in young chicks and was rarely isolated. In mature birds, mycoplasma were isolated from the respiratory tissues of some infected with either of the two S6 strains or with M. gallinarum, but not from A514 infected chickens. In both younger and mature chickens, mycoplasma were most likely to be recovered from respiratory tissues and the infra-orbital sinus. Most chicks infected with the virulent S6, showed positive haemagglutination inhibition (HAI) reactions from 4 days of age which preceded the rapid serum agglutination (RSA) reactions. In contrast, chicks infected with other strains of M. gallisepticum either showed no positive HAI reactions, or developed them after 21 days. In either case, positive RSA preceded HAI reactions. The numbers of mature birds examined were small, but, comparing isolation of M. gallisepticum with serological results, it appears that false positives occur with the RSA and both false positives and negatives with the HAI test. No egg transmission of mycoplasma was observed.