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We evaluated the pathogenicity of three live Mycoplasma gallisepticum (MG) vaccine candidates by infection via aerosol of 3-wk-old chickens with log phase broth cultures (trial 1). Two of the candidates (K3020 and K4649A) colonized only 10% and 20% of the chickens, respectively, unlike K2101 (K-strain), which was reisolated from all of the vaccinated chickens tested. K-strain inoculation did not result in significant air sac or tracheal lesions in chickens at 10 and 39 days postinfection (P < or = 0.05). The efficacy of K-strain as a live vaccine was evaluated in trial 2, by challenge of vaccinated chickens with virulent R-strain via aerosol at 6 wk postvaccination. K-strain vaccination resulted in significant protection from air sac and tracheal lesions (P < or = 0.05). The K-strain was further investigated to evaluate transmissibility (trial 3), colonization and persistence of infection following aerosol administration (trial 4), genetic and phenotypic stability following back passage through chickens (trial 5), and vertical transmission (trial 6). The K-strain had a low rate of horizontal transmission; it remained primarily in the respiratory system of inoculated birds and persisted in the upper respiratory tract for the duration of the trial 4 (5 mo). There was no increase in virulence of K-strain when it was back passaged five times through chickens, and no vertical transmission of K-strain was detected. K-strain showed great potential as a safe and effective live MG vaccine.

Mycoplasmas are pathogens of different avian species, and they are able to be vertically transmitted. Even detected, Mycoplasma prevalence in raptor eggs is very low. In contrast to poultry, raptor eggs submitted for investigations are usually incubated. To investigate the influence of incubation length on the recovery of mycoplasmas from eggs, infertile specific-pathogen-free chicken eggs and embryos were infected with Mycoplasma lipofaciens (strain ML64), which had previously been isolated from an egg of a northern goshawk (Accipiter gentilis), in two different dosages. The eggs were investigated up to 12 days after infection (infertile eggs) or embryonic death. Mycoplasmas were recovered over the entire period after embryonic death by isolation. It was possible to re-isolate M. lipofaciens (strain ML64) from infertile eggs infected with 10(6) colony-forming units (CFUs) up to 12 days, but only up to 7 days if infected with 10(2) CFUs, which may be closer to the situation after natural infection. This study demonstrates that incubation of infertile eggs does have an influence on the recovery rate of mycoplasmas. This influence must be considered if interpreting results of Mycoplasma investigations in eggs of nonpoultry species. Additionally, it is recommended to use dead in shell embryos rather than infertile eggs for Mycoplasma detection.

The use of controlled, horizontal-transmission experiments provides detailed information on the spread of disease within fixed social groups, which informs our understanding of disease dynamics both in an empirical and theoretical context. For that reason, we characterized in 2002, horizontal transmission of Mycoplasma gallisepticum (MG) in two flocks of 11 wild-caught house finches housed in outdoor aviaries over a 6-mo period. All birds were initially free of MG by a polymerase chain reaction (PCR)-based test, rapid plate agglutination (RPA), and the scoring of physical signs. We inoculated one flock member bilaterally in the palpebral conjunctiva and reintroduced it into its cage. Index birds developed conjunctivitis within 3 to 5 days but died 13 and 20 days postinfection (PI) possibly because of very severe weather. The proportion of birds with physical signs increased gradually, reached 40% at 6 wk PI, and fluctuated around 40% until 21 wk PI. By the time our experiment ended at 24.5 wk PI, 28% of the birds still exhibited physical signs. Across both flocks, 80% of the birds developed unilateral or bilateral conjunctivitis, and several birds relapsed. The appearance of physical signs in new individuals occurred between 10 and 144 days PI (median 41 days PI). Physical signs lasted 1-172 days (median 42 days). Birds that became infected earlier during the experiment developed more severe conjunctivitis, and there was a tendency for birds that developed bilateral conjunctivitis to develop physical signs earlier. Most birds that developed physical signs of MG were also PCR- and RPA-positive, although we detected a single asymptomatic carrier and a single symptomatic false negative. No birds died as a result of secondary MG infection.

To determine the Mycoplasma gallisepticum (MG) rapid serum plate agglutination (RSPA) test response of broiler breeders after ts-11 strain vaccination, 55 Cobb pullets derived from a nonvaccinated, MG-negative, commercial, broiler breeder grandparent flock were monitored from 8 to 20 wk of age (over a 12-wk trial period). To evaluate the effect of lateral spread of the ts-11 vaccine strain on RSPA test results from commingled and adjacently penned birds, treatment groups included (A) birds vaccinated with ts-11strain MG at 8 wk of age, (B) commingled nonvaccinates in the same pen as the vaccinated birds, (C) nonvaccinates in a second pen separated from the first pen by a distance of 2 m, and (D) birds vaccinated with ts-11 strain MG at 8 wk of age and kept in a separate room. Rapid serum plate agglutination tests were performed once a week for 6 wk and then every 2 wk for 6 more wk, postvaccination. A polymerase chain reaction (PCR) assay specific fbr ts-11 strain MG was used to confirm vaccination, and a second PCR specific for non-ts-11 strain MG was used to confirm the absence of field infection. Seroconversion was first detected by the RSPA test 2 wk postvaccination and attained maximum positive rates of 58% at 12 wk postvaccination in treatment A and 60% at 8 wk postvaccination in treatment D. Seroconversion rates in nonvaccinated, commingled pullets was 10% at 5 wk and 30% at 12 wk after the vaccination of pen mates. The ts-11-specific PCR detected the vaccine strain in 80%-100% of the vaccinated birds 2 wk after vaccination. One of 15 nonvaccinated birds penned 2 m from vaccinated birds yielded ts-11 by PCR assay 12 wk after vaccination, which indicates that the spread of ts-11 over short distances may be possible in situations in which there is a common caretaker. PCR on tracheal swabs taken 12 wk postvaccination detected ts-l1 in 50% and 60% of the vaccinated birds in treatments A and D, respectively; in 30% of the commingled nonvaccinates; and in 6.6% of the separately penned nonvaccinates. In contrast, choanal swabs collected from vaccinated birds at 12 wk were 21% and 40% PCR positive for ts-11 strain MG, while those from nonvaccinates were negative. All samples were PCR negative for field strain MG. The pattern of seroconversion as measured by RSPA test in small groups of broiler breeders was different from that previously reported for leghorns. Lateral spread of the ts-11 strain to commingled pen mates occurred rapidly, causing RSPA seroconversion patterns that mimicked those of the vaccinated pen mates.

Long known as a pathogen of poultry, Mycoplasma gallisepticum (MG) was first detected in house finches in 1994. The disease rapidly spread throughout the eastern United States and Canada and was associated with debilitating disease and high mortality in house finches. However, in the late 1990s, the proportion of infected finches dying as a result of infection with MG decreased, and asymptomatic infection was more common among wild birds than in the past. We documented MG infections in breeding house finches and concluded that adults of both sexes transmit the infection to dependent young, probably after hatch. MG infections of breeding adults occurred late in the breeding season and were found in birds completing significantly more nests than birds that never tested positive for MG, implying that higher rates of reproduction carry a cost in the form of increased risk of infection. We found evidence of an MG-induced delay in dispersal of nestlings from their natal area and demonstrated a significant impact of infection on nestling growth.

An outbreak of Mycoplasma gallisepticum (MG) in commercial turkeys involving very mild clinical signs was difficult to confirm by routine methods. In the first part of this study (trial A), we conducted a bioassay to increase the likelihood of detecting MG. Susceptible turkeys were inoculated with sinus exudates from four different affected commercial turkey flocks. Turkeys were evaluated for clinical signs, as well as by serology and culture of tracheal swabs, at 21 and 42 days postchallenge. An MG isolate from one of the sinus exudates used for inoculation, designated K5054, was very similar to isolates from house finches when characterized by random amplified polymorphic DNA analysis as well as DNA sequence analysis of portions of the phase-variable putative adhesin protein (pvpA) gene, a lipoprotein gene, and the cytadhesin gapA/mgc1 gene. The turkeys inoculated with the K5054 sinus exudate seroconverted in the absence of severe clinical signs. There was a single reisolation of K5054 from these turkeys 42 days postchallenge. Susceptible contact turkeys were commingled with the K5054-inoculated turkeys at 49 days postchallenge. We found no evidence of transmission of MG to the contacts by culture or serology at 7, 21, or 35 days after commingling. In the second part of this study (trial B), we challenged the contacts and K5054 sinus exudate-inoculated turkeys from trial A with virulent R strain 88 days after the K5054 sinus exudate inoculation. On necropsy 10 days postchallenge, the evaluation of gross and microscopic lesions, serology, and culture showed that the turkeys previously inoculated with K5054 sinus exudate were protected against disease and reinfection.

Since 1994, an epidemic of conjunctivitis caused by Mycoplasma gallisepticum (MG) has spread throughout the eastern population of house finches (Carpodacus mexicanus). The adaptation of MG to a free-flying avian species presents potential problems for the control of mycoplasmosis in commercial poultry. To evaluate risks associated with this emerging problem, a field survey was conducted to assess prevalence of MG infection in house finches and other passerine birds associated with poultry farms. Between November 1997 and March 1999, 1058 birds were captured by mist net or trap at 17 farms and at 10 feeder stations in northeast Georgia. Birds were bled and screened by serum plate agglutination (SPA) for antibodies to MG. Birds with negative or weak positive SPA results were released at capture sites, and those with strong positive SPA reactions were kept for further evaluation. Necropsies were performed on selected house finches and individuals of 11 other passerine species, and samples were collected for MG testing by culture, polymerase chain reaction (PCR), hemagglutination inhibition, and histopathology. Testing revealed 19.1% of 671 birds caught at farms and 11.6% of 387 birds caught at feeder sites were SPA positive for MG. Three house finches captured on farms were positive for MG by culture and PCR, whereas three from feeder sites were positive only by PCR. No MG isolates were made from tufted titmice (Baeolophus bicolor), but 40% were positive by PCR. Individuals from 10 additional species were SPA positive only. Results suggest that MG persists at low levels in house finches in northeast Georgia and that tufted titmice may be nonclinical carriers of MG or a related mycoplasma. Positive SPA reactions in other species may be caused by nonspecific reactions or contact exposure. Current biosecurity recommendations should be sufficient to minimize risks of transmission between wild and domestic birds.

Mycoplasma gallisepticum (MG) has been isolated from wild house finches. The pathogenic effects of MG finch strain (K4058) and MG R-strain were compared after exposure of chickens and turkeys. Gross and histologic lesions, reisolation of the organism, serology, and clinical disease were evaluated. Milder histologic and gross lesions, in addition to lower serologic titers, occurred in birds inoculated with the finch strain. Mortality, concurrent with clinical and gross respiratory signs and lesions, was observed only in chickens challenged with R-strain. Both the MG finch strain and MG R-strain were recovered from the respective challenge groups at 14 and 28 days postexposure. The results show that MG isolated from wild house finches may infect domestic poultry species but causes only mild disease and is less virulent than MG R-strain. Commercial enzyme-linked immunosorbent assay kits best detected the serologic response of chickens and turkeys to the MG finch strain.

Although Mycoplasma gallisepticum (MG) is established in house finch (Carpodacus mexicanus) populations in at least 33 states, the potential risk of MG introduction to domestic poultry by infected finches currently is unknown. The objectives of this study were to determine if chickens could be infected with the finch strain of MG via direct, across-wire, and proximity (across-room) contact with naturally infected house finches and to determine if house finches could be infected through direct contact with experimentally infected chickens. Chickens were infected with the finch strain of MG through direct contact with naturally infected house finches, a determined by seroconversion (80%), polymerase chain reaction (PCR) (20%), and culture of MG (30%). Clinical disease was not observed in infected chickens. Isolates from chickens were identified as the original finch strain by arbitrary primed PCR. Transmission required an extended period of direct contact (10 wk) with infected finches, and no evidence of MG infection was detected in chickens exposed to infected finches across wire or across the room. Evidence of contact transmission of MG from infected chickens to house finches was limited to positive serum plate agglutination results, and infection could not be confirmed by PCR or culture. Results suggest that minimal biosecurity measures that restrict direct contact between chickens and house finches should significantly reduce the potential for MG transmission between these species.

Fifteen mycoplasma-free chickens were contact exposed to five chickens that had been experimentally infected with one of three different strains (two field strains and one laboratory strain) of Mycoplasma synoviae (MS). Culture and polymerase chain reaction (PCR) were positive by 3 days postinoculation (PI) in the experimentally infected birds. Lateral transmission was found by 7-14 days postexposure. Positive serum plate agglutination (SPA) results were detected 3-4 wk after positive culture and/or PCR in individual birds. By 42 days PI, all the birds in the groups exposed to field strain K1858 or K3344 had become infected as determined by culture and PCR, whereas only half of the birds in the group exposed to laboratory strain WUV1853 had become infected. Because of the unanticipated lack of seroconversion to hemagglutination inhibition (HI) and enzyme-linked immunosorbent assay (ELISA) in infected chickens, the study was extended. Each group was split into two groups of 10 birds each, one of which was vaccinated with a live B1/LaSota Newcastle disease (ND) vaccine virus to determine if a viral respiratory challenge might incite a stronger antibody response to the mycoplasma infection. All the birds were tested for seroconversion 14 and 21 days later. Of the birds vaccinated for ND, a slightly greater number were MS positive by SPA than the nonvaccinated birds. This effect was not present 21 days after vaccination, and there was no significant difference in the MS HI results from these groups, suggesting that the viral respiratory infection had little direct impact on seroconversion. The virulent field strain (K3344) elicited a stronger MS antibody response than the other strains. All results from the MS ELISA were negative in all groups through 9 wk. Positive results from PCR analysis correlated well with culture results, whereas serologic tests did not detect MS infection for several weeks. Monitoring programs solely dependent on seroconversion may be inadequate for diagnosis and control of mycoplasma infections.

In separate trials, layer pullets were vaccinated with Mycoplasma gallisepticum (MG) strain 6/85 or strain ts-11 commercially produced live vaccines. For a 15-wk postvaccination (PV) period, vaccinates were commingled with unvaccinated pullets and were in indirect contact with sentinel groups of pullets, broiler breeders, turkey breeders, or meat turkeys in adjoining pens. Infectivity and transmissibility of vaccine strains were determined by tracheal culture and serology at 1 wk followed by 3-wk intervals PV. Strain 6/85 was recovered from 0%-20% of vaccinates, but not from commingled pullets or sentinel birds. Strain ts-11 was recovered from 60%-90% of vaccinates and 0%-40% of commingled pullets but not from any of the sentinel birds. No birds in the 6/85 vaccine trial tested positive for MG antibodies by serology. MG enzyme-linked immunosorbent assays detected positive responses in ts-11 vaccinates (range = 10%-70%) at 42, 63, 84, and 105 days PV, and commingled pullets (10%) at 84 and 105 days PV. MG serum plate agglutination tests detected positive responses in 90% and 20% of ts-11 vaccinates at 42 and 105 days PV, respectively, and commingled pullets (10%) at day 42 PV. Clinical signs, morbidity, or mortality suggestive of pathogenic MG infection were not observed in any bird during either trial, and no gross lesions were observed at necropsy. Random amplified polymorphic DNA analysis was capable of distinguishing each of the vaccinal strains 6/85 and ts-11 from each other by their distinct DNA banding patterns.

Five groups of 20 commercial leghorn hens near peak production were challenged with Mycoplasma gallisepticum, and medicated with enrofloxacin in the drinking water at 7-11, 21-25, or 7-11 and 21-25 days postchallenge (PC), a combination of lincomycin/spectinomycin at 7-11 and 21-25 days PC, or left as untreated controls. Egg production records were maintained, and all eggs produced during the 63 days following challenge were incubated for 18 days and then cultured for M. gallisepticum. All groups experienced marked egg production drops beginning about 1 week PC; production returned to near normal levels after approximately 3 weeks. Under the conditions of this study water medication did not prevent egg production losses, probably because of the delay in the onset of medication, but medication may have resulted in a more rapid recovery of egg production. Enrofloxacin was highly effective in reducing the level of egg transmission of M. gallisepticum, although a small number of eggs (0-0.3%) in the treated groups were culture positive after medication was begun. Only 1 of 2463 live embryos was culture positive after the onset of treatment. In the unmedicated control group, 12.8% of all eggs were culture positive. Lincomycin/spectinomycin also reduced egg transmission (3.4% of eggs were culture positive after treatment was initiated), but not as effectively as enrofloxacin. Most of the egg transmission occurred before the medication could have had an effect. Both enrofloxacin and lincomycin/spectinomycin reduced the number of dead-in-shell embryos.

The effects of Mycoplasma gallisepticum (MG) vaccination on egg transmission of MG and egg production were evaluated. Leghorn hens vaccinated with live MG (strain F), with strain F plus MG bacterin, with one dose of MG bacterin, or with two doses of MG bacterin all transmitted MG through the egg at a significantly lower level than unvaccinated controls. Hens vaccinated with two doses of MG bacterin had the longest lag before detectable transmission of MG through the egg. All vaccinated groups were protected against the egg-production drop seen in unvaccinated hens challenged with virulent MG.

The egg transmissibility of strains F and R of Mycoplasma gallisepticum (MG) was studied in 53 pullets by isolating MG from the eggs and embryos. MG was isolated at rates of 3.19% and 1.59% from the eggs of hens exposed by aerosol to the R or F strain, respectively. However, MG was isolated at rates of 0.19% and 0.00% from the eggs of chickens exposed by eye drop to the R or F strain, respectively. A statistically significant reduction in egg production was observed during the first 4 weeks postexposure in the group exposed to the R strain of MG by aerosol. An unexpected finding was significantly lower fertility of eggs from all MG-infected groups.

A Mycoplasma gallisepticum bacterin was prepared and used in Mycoplasma gallisepticum (MG)-positive primary breeders to control vertical transmission of MG. Two generations were vaccinated, but the third generation was not vaccinated and was monitored serologically. Results showed no evidence of MG at 1 day, 6 weeks, 11 weeks, 16 weeks, or 31 weeks of age. This procedure may offer small breeder organizations and showbird fanciers a way to eliminate MG.

Leghorn pullets free of Mycoplasma gallisepticum (MG) were infected with the F strain of MG at approximately 18 weeks of age. At various times up to 27 weeks postinfection, infected chickens were placed in a pen with uninfected controls. Infected chickens remained tracheal carriers up to 49 weeks postinfection. Infection was readily transmitted to penmates during the first 4 weeks postinfection; in most instances from 6 through 27 weeks postinfection, transmission to penmates became progressively slower. Five groups of broiler chickens were reared in the same house as the Leghorn chickens up to 8 weeks of age. Only one of the 5 groups became infected. Birds in this group were reared in a pen adjacent to infected pullets from 1 day of age and were apparently not infected until they were nearly 8 weeks old. At no time did transmission occur between groups separated by an aisle or empty pen.

Lateral spread of S6 strain Mycoplasma gallisepticum (MG) was studied in small populations of chickens. One experimentally exposed bird served as the source of infection, and the presence of MG-agglutinating antibody was evidence of infection in individuals. The results were subjected to survival data analysis. In the seven experiments, four similar but not identical phases of lateral spread were observed: phase 1, a generally long latent phase (median 15, range 12-21 days) before antibody was first detected in the MG-inoculated bird; phase 2, a generally short period (median 1, range 1-21 days) in which infection gradually appeared in 5-10% of the population; phase 3, a fairly constant characteristic phase (median 24, range 7-32 days) in which 90-95% of the remaining population developed MG antibody; phase 4, a generally short terminal phase (median 4, range 3-19 days) in which the remainder of the population became positive. Increasing the population density increased the rate at which lateral spread occurred.

House sparrows were infected by aerosol with Mycoplasma gallisepticum (MG) or M. synoviae (MS). MG was reisolated from 5 to 11 sparrows 10 days postinfection, but infection appeared to be temporary. Mycoplasma-free chickens reared in the experimental house became infected with MG during the trial. MS was recovered from only one sparrow. Serological tests were unsatisfactory for diagnosing infected birds. The results suggest that house sparrows may be temporary biological carriers of MG.

An infection with Mycoplasma synoviae that spread slowly within and among pens of chickens was observed in a brown-egg breeding population that had long been free of M. gallisepticum and M. synoviae. There were unusually long periods of time between the first serological evidence of infection in a pen of chickens and the time that most of the group became positive. Transmission to adjacent pens was also slow.

Turkey eggs infected naturally and experimentally and treated with 1 mg of tylosin or gentamicin before incubation produced progeny with less infection with Mycoplasma meleagridis than did infected but untreated eggs. Higher doses of gentamicin eliminated infection, as indicated by an absence of M. meleagridis antibodies in poults tested at 3 weeks and retested at 5 weeks of age. The rate of reduction or elimination of egg transmission was correlated negatively with the incidence of M. meleagridis infection in untreated controls. Hatchability was reduced by gentamicin, especially at doses of 1.75 and 2 mg, whereas tylosin at 1 mg/egg improved hatchability.

Twenty-four 70-week-old and sixteen 27-week-old white leghorn hens were challenged with R strain Mycoplasma gallisepticum (MG) by injection into the caudal thoracic air sac and infraorbital sinus. Eggs were collected daily and cultured within 7 days or incubated for 18 days. Vitelline membranes of eggs were cultured directly; in 18-day-old embryos, cultures were taken from the yolk sac, air sacs, and oral cavity. Culture of vitelline membrane of eggs within 2 days was compared with culture of eggs stored 10 days post oviposition. The first MG-positive egg was laid 2 days postinfection (PI). Hens continued to lay positive eggs to the end of the experiments. There was no significant difference in MG recovery between eggs cultured within 2 days and those cultured 10 days post oviposition. MG was isolated at a significantly higher rate from eggs than from 18-day-old embryos. MG was isolated at a higher rate from the yolk sac of 18-day-old embryos than from the air sacs or oral cavity of the same embryos.

Groups of white leghorn hens were vaccinated twice with a Mycoplasma gallisepticum (MG) bacterin, once with bacterin, or left unvaccinated. Four weeks after vaccination, they were challenged with virulent R strain MG. Egg production was significantly higher in challenged vaccinated groups than in the challenged control group. Four challenged control hens went out of production, whereas only one twice-vaccinated hen did. MG was first isolated directly from eggs 5 days postchallenge (PC) in twice-vaccinated hens, 4 days PC in once-vaccinated hens, and 2 days PC in controls, and the hens continued to lay positive eggs till the end of the experiment 7 weeks PC. MG was found in 17.65%, 38.55%, and 45.90% of eggs cultured in twice-vaccinated, once-vaccinated, and control groups, respectively. Nine of 16 twice-vaccinated hens were found to be shedding MG through their eggs, whereas 15 of 17 once-vaccinated hens and 14 of 16 controls were shedding MG through their eggs.