Because of their strong suppressive effects, bacterial contamination must be excluded before attempting mycoplasma detection in pharmaceutical testing.
Bacteria suppress mycoplasma via nutrient competition, space occupation, pH shifts, and by secreting antibiotics or lytic enzymes.
Therefore, early detection is critical. If bacterial contamination is suspected or confirmed and previous storage conditions were acceptable, proceed by first filtering out the bacteria and then diluting the sample to reduce concentrations of inhibitory substances.
Even if the mycoplasma concentration becomes very low, repeated passaging and dilution can reduce the impact of inhibitors and potentially allow recovery of viable mycoplasma.

For biopharmaceutical samples that are confirmed to be bacteria-free, you can simply filter through a 0.1 µm membrane to collect mycoplasma. The membrane can then be placed onto mycoplasma culture medium.
For clinical research samples, it is recommended to first filter out bacteria using a 0.45 µm membrane, followed by capturing mycoplasma with a 0.1 µm filter.

Theoretically, antibiotics such as tetracyclines, macrolides, fluoroquinolones, lincosamides, aminoglycosides, amphenicols, and spectinomycin are effective against mycoplasma.
However, field and clinical data from various continents and animal species over the past decade have shown widespread emergence of resistance among many mycoplasma strains.

Bacteria exert a strong suppressive effect on mycoplasma. They not only outcompete for nutrients and space, but their metabolic byproducts can shift pH levels (either too low or too high) to lethal levels for mycoplasma.
Additionally, bacterial enzymes may directly degrade mycoplasma cells—even before any change in pH is detected, microscopic observation might already reveal active destruction.
Bacteria also secrete antibiotics and lytic enzymes that can suppress or kill mycoplasma. Even if the bacteria are later eliminated by sterilization, the residual inhibitory substances may remain and suppress mycoplasma, causing them to become latent. Once the environment becomes favorable—such as entry into a human host—these inhibitory substances are quickly diluted, potentially triggering explosive mycoplasma growth.

Although nucleic acid technologies (e.g., PCR) are now highly advanced, the overall research on mycoplasma remains limited. This has resulted in poorly optimized primers, and most commercial mycoplasma PCR kits have a significantly higher risk of false positives and false negatives compared to those for bacteria.