Effects of sialidase knockout and complementation on virulence of Mycoplasma gallisepticum.

Reannotation of the pathogenic Mycoplasma gallisepticum strain R(low) genome identified the hypothetical gene MGA_0329 as a homolog of the sialidase gene MS53_0199 of Mycoplasma synoviae strain MS53. Potent sialidase activity was subsequently quantitated in several M. gallisepticum strains. Because sialidase activity levels correlate significantly with differing M. synoviae strain virulence, we hypothesized this enzyme may also influence the virulence of M. gallisepticum. MGA_0329 was disrupted in strain R(low) to create mutants 6, 358 and P1C5, which resulted in the loss of sialidase activity in all three mutants. Chickens infected with the knockout mutants had significantly less severe (P<0.05) tracheal lesions and tracheal mucosal thickening than chickens infected with equal doses of strain R(low). Significantly fewer (P<0.05) CCU especially of strains 6 and P1C5 were recovered at necropsy. Mini-Tn4001tet plasmid pTF20 carrying a wild-type copy of MGA_0329 with its native promoter was used to complement the genetic lesion in strain P1C5. Three clones derived from P1C5, each having one copy of MGA_0329 stably transposed into a different site in its genome, expressed sialidase restored to wild-type activity levels (1.58×10(-8)U/CFU). Complementation of P1C5 with MGA_0329 did not restore it to wild-type levels of virulence, indicating that the contribution of sialidase to M. gallisepticum virulence is not straightforward.

In vitro antibiotic susceptibility of Mycoplasma iguanae proposed sp. nov. isolated from vertebral lesions of green iguanas (Iguana iguana).

Mycoplasma iguanae proposed species nova was isolated from vertebral abscesses of two feral iguanas (Iguana iguana) from Florida. Three strains were evaluated for sensitivity to a variety of antibiotics. The minimum inhibitory concentrations for M. iguanae, assessed by broth dilution methods, of clindamycin, doxycycline, enrofloxacin, oxytetracycline, and tylosin (all <1 microg/ml) were lower than those of chloramphenicol (32 micro/ml) and erythromycin (64 microg/ml). The profile was identical to that of Mycoplasma alligatoris, previously isolated from American alligators (Alligator mississippiensis). M. iguanae strain 2327T was subcultured without antibiotics to assess mycoplasmacidal activity. Clindamycin, doxycycline, oxytetracycline, and tylosin were bacteriostatic from 0.1 to 0.5 microg/ml, whereas enrofloxacin was bactericidal at 20 ng/ml. An enrofloxacin dosage of 5-10 mg/kg achieves peak plasma concentrations >1 microg/ml, with an elimination half-life of 6-20 hr, in alligators. Although concentrations achieved in the vertebrae by i.m. or i.v. injection are probably lower than those in plasma, these data suggest that enrofloxacin may be useful to treat M. iguanae mycoplasmosis in iguanas.