Mycoplasma Flashcards
Mycoplasma sources
Humans, primates, bovine, porcine
Respiratory tract
Mycoplasma definition
Parasitic bacterium that are slow growing and lack a cell wall
Features
Small genome, AT rich
Reproduce by binary fission
Antibiotics are ineffective
Mycoplasma alter
Cell function, growth, metabolism
Morphology
Attachment
Membranes
Alter nucleic acid and protein synthesis
Affect cell antigenicity
Interfere with viral propagation and yield
Mimic viral actions
Induce chromosomal aberrations and damage
Cytotoxic effects
Transfection efficiency
High contamination rates
Absence of visible signs of contamination
Use if antibiotics
Pass through filters for sterilization
Methods for detecting mycoplasma
Microbiological culturing
Biochemical
DNA fluorescent staining
PCR
Microbiological culturing
Sensitive
Must use mycoplasma specific agar and broth
Many mycoplasma can not be cultured
Can take up to 4 weeks to get results
Biochemical
Mycoplasma produce adenosine phosphorylase which converts 6MPDR into 2 products that are cytotoxic to cells
Cultures and 6MPDR are incubated then stained with crystal violet
Strong stringing=no mycoplasma
DNA fluorescent staining
Use DAPI or Hoechst stain
Intermediate sensitivity
PCR
Use primers to conserved and mycoplasma specific 16S rRNA spacer regions
High sensitivity
Rapid
Can amplify multiple species in a single reaction
Mycoplasma positive band at 450-550 bps
Ways to deal with contaminated cell lines
Discard cultures and reagents used for cultivation
Try removal agents
Culture in the presence of specific antibiotics
Mycoplasma removal agent
Mynox it integrates into the mycoplasmic membrane and causes osmotic influx
Antibiotics for mycoplasma
Ciorofloxacin, MRA and novobiocin, plasmocin
Mycoplasma testing program
Test all cells in culture Quarantine new cell lines Test routinely Test all cells before freezing Test new lots of sera
