Cell Line Contamination: Causes, Detection, and Prevention
By Dr. Zubair Khalid, DVM, MS, PhD ·
Key Takeaways
- Cell line contamination, encompassing microbial agents (bacteria, fungi, mycoplasma, viruses) and cross-contamination with other cell lines, fundamentally compromises experimental integrity and reproducibility.
- Mycoplasma contamination is particularly insidious due to its lack of visible indicators, yet it significantly alters host cell metabolism, gene expression, and drug responses by depleting essential nutrients like arginine and inducing chromosomal aberrations.
- Contamination is primarily introduced via breaches in aseptic technique, contaminated reagents (especially fetal bovine serum), incubator reservoirs, and human error in handling and labeling, spreading through aerosols and shared laboratory equipment.
- Definitive detection necessitates molecular methods such as PCR for mycoplasma and bacteria/fungi, and Short Tandem Repeat (STR) profiling for cell line authentication to identify cross-contamination.
- Prevention hinges on stringent aseptic practices, rigorous quarantine and testing of new cell lines (including mycoplasma PCR and STR profiling), and judicious use of antibiotics, which do not eliminate mycoplasma and can mask low-level infections.
- Routine monitoring, including visual inspection at each passage, periodic mycoplasma testing (every 1-3 months), and regular STR authentication (every 6 months or 20 passages), is critical for early detection and mitigation of contamination.
Introduction to Cell Line Contamination
What is Cell Line Contamination?
Cell line contamination refers to the unintended introduction of foreign biological material—microorganisms or cells from another line—into a cultured cell population. In practical terms, a contaminated culture is one that no longer contains only the intended cells in a defined, controlled environment. The contaminating agents fall into two broad categories: microbial contaminants (bacteria, fungi, yeast, mycoplasma, and viruses) and cellular contaminants (cells from a different species or from a different tissue or individual of the same species).
Contamination is not a rare event. It is estimated that between 5% and 30% of cell lines in use worldwide are cross-contaminated with another cell line, and mycoplasma infection rates in routine culture laboratories have historically ranged from 15% to 80% depending on the lab and detection method used. These figures underscore that contamination is not an occasional mishap but a systemic problem in cell culture work.
Why It Matters in Research
The consequences of contamination extend far beyond a ruined flask. A contaminated culture produces data that are unreliable, unreproducible, and potentially misleading. If a researcher believes they are studying the response of a lung epithelial cell line to a drug, but the culture is actually a mixture of HeLa cells and mycoplasma, every downstream measurement—gene expression, protein levels, proliferation rates, apoptosis—reflects the biology of the contaminant, not the intended system.
The scientific literature contains numerous examples of retracted papers and invalidated conclusions traced back to contaminated cell lines. The most infamous case involves HeLa cells, which have been shown to contaminate and overgrow hundreds of other cell lines over decades of research. Similarly, mycoplasma contamination can alter cellular metabolism, induce chromosomal aberrations, and change cell surface antigen expression without producing visible turbidity or pH change, making it a silent but potent confounder.
For an undergraduate student learning cell culture, understanding contamination is not merely an academic exercise. It is the foundation of experimental rigor. Every protocol you learn—from Cell Passaging to Mtt Assay Cell Viability—assumes a pure, uncontaminated culture. Without that assumption, the results are meaningless.
Types of Cell Line Contamination
Microbial Contamination
Microbial contamination is the introduction of bacteria, fungi, yeast, mycoplasma, or viruses into a cell culture. Each class has distinct characteristics, detection methods, and implications.
Bacteria. Bacterial contamination is the most common and most visible form. Contaminated cultures often become turbid within 24 to 48 hours, and the pH of the medium drops as bacteria produce lactic acid and other metabolic byproducts, turning the phenol red indicator from red to yellow. Common contaminants include species of Escherichia coli, Staphylococcus, Streptococcus, and Pseudomonas. Under an inverted microscope, bacteria appear as small, motile or non-motile granules between or on top of cells. Rapidly growing bacteria can outcompete mammalian cells for nutrients and produce toxins that kill the culture within days.
Fungi and Yeast. Fungal contamination, most commonly by species of Aspergillus, Penicillium, and Candida, is slower to develop than bacterial contamination. Fungal hyphae appear as thin, branching filaments visible under low magnification. Yeast, particularly Candida albicans and Saccharomyces cerevisiae, appear as oval or spherical budding cells. Fungal contamination often produces a visible mycelial mat on the surface of the culture medium and can be accompanied by a musty odor. Because fungal spores are resistant to many disinfectants and can survive in air currents, they are particularly difficult to eliminate once introduced into a laboratory environment.
Mycoplasma. Mycoplasmas are the smallest self-replicating prokaryotes, lacking a cell wall and measuring only 0.2 to 0.3 micrometers in diameter. They are the most insidious contaminants because they do not produce turbidity, pH change, or visible growth under a light microscope. Mycoplasma contamination is typically detected only through specialized staining, PCR, or enzymatic assays. The most common species contaminating cell cultures are Mycoplasma arginini, M. hyorhinis, M. orale, and Acholeplasma laidlawii. These organisms deplete arginine and other amino acids from the medium, alter host cell metabolism, induce chromosomal abnormalities, and can modulate viral replication. Because they pass through 0.22-micrometer filters used for sterilizing media, they are notoriously difficult to eliminate.
Viruses. Viral contamination is less common but potentially more dangerous. Endogenous retroviruses are present in many rodent cell lines and can be shed into the culture medium. Bovine viral diarrhea virus (BVDV) can contaminate fetal bovine serum (FBS), a common medium supplement. Viral contamination is typically detected through PCR, electron microscopy, or infectivity assays. Unlike bacterial or fungal contamination, viral contamination does not usually kill the culture or produce visible changes, making it a hidden variable that can profoundly affect experiments involving viral infection, immune responses, or gene expression.
Cross-Contamination with Other Cell Lines
Cross-contamination occurs when cells from one line are inadvertently introduced into a culture of another line. This can happen through shared media, mislabeled flasks, or the use of the same pipette or tip across multiple cell lines. The contaminating cells often grow faster than the intended line and can overtake the culture within a few passages.
The most notorious example is HeLa, the first immortal human cell line, derived from Henrietta Lacks in 1951. HeLa cells grow aggressively, attach firmly to surfaces, and can be aerosolized during pipetting. They have been found contaminating hundreds of other cell lines, including many that were believed to be unique. Other fast-growing lines, such as T24 (bladder carcinoma) and HT-29 (colorectal adenocarcinoma), have also been implicated in widespread cross-contamination.
Cross-contamination is particularly dangerous because it is invisible. The culture looks healthy, grows normally, and appears identical under the microscope. Only molecular authentication methods, such as STR profiling, can reveal that the cells are not what the label claims. This problem is so pervasive that the International Cell Line Authentication Committee (ICLAC) maintains a registry of known cross-contaminated cell lines, and many journals now require STR authentication before publishing results.
Sources and Routes of Contamination
Common Sources in the Lab
Contamination enters cell cultures through several well-defined routes. Understanding these routes is the first step in prevention.
Poor Aseptic Technique. The most common source of contamination is the researcher. Touching the inside of a flask neck, allowing a pipette to contact a non-sterile surface, or failing to flame the mouth of a bottle before opening can introduce microorganisms directly into the culture. Even experienced researchers make these errors, particularly when working quickly or multitasking.
Contaminated Reagents. Media, serum, trypsin, and other liquid reagents can arrive contaminated or become contaminated during use. FBS is a particular concern because it is a rich growth medium for mycoplasma and viruses. Water baths, which are used to warm media, are common reservoirs of bacteria and fungi; if the necks of bottles are submerged, contaminated water can wick into the bottle when the cap is loosened.
Incubator Issues. CO₂ incubators maintain a warm, humid environment that is ideal for microbial growth. If the incubator is not cleaned regularly, or if the water pan is contaminated, mold and bacteria can proliferate and be carried into cultures by air currents. Fungal spores, in particular, can survive in incubator crevices and contaminate cultures over weeks.
Human Error and Mislabeling. Cross-contamination often results from mislabeling flasks, sharing media between cell lines, or using the same pipette for multiple lines. A single moment of inattention—picking up the wrong tube, using a shared bottle of medium, or writing an incorrect label—can compromise an entire experiment.
How Contamination Spreads
Once introduced, contamination spreads through the laboratory in predictable ways. Aerosols generated during pipetting or centrifugation can carry bacteria, fungi, or cells into the air. These particles can settle on surfaces, be inhaled by researchers, or enter other cultures. Mycoplasma, in particular, is easily aerosolized and can survive on surfaces for weeks.
Shared equipment is another major vector. Laminar flow hoods, if not properly cleaned and UV-irradiated between uses, can harbor contaminants. Centrifuges, water baths, and even refrigerator door handles can transfer microorganisms between researchers and cultures. Cell lines themselves can spread contamination: a contaminated culture placed in a shared incubator can release mycoplasma into the air, which then infects neighboring cultures.
Detecting Cell Line Contamination
Visual and Microscopic Indicators
The first line of detection is visual inspection. A contaminated culture often shows one or more of the following signs:
- Turbidity: The medium becomes cloudy, indicating bacterial or yeast growth.
- pH change: The medium turns yellow (acidic) due to bacterial metabolism or purple (alkaline) due to cell death and media exhaustion.
- Visible particles: Floating debris, clumps, or a film on the surface of the medium may indicate fungal growth.
- Microscopic features: Under an inverted microscope, bacteria appear as small granules between cells; fungi appear as long, branching hyphae; yeast appear as oval budding cells. Mycoplasma, however, are too small to be seen with a standard light microscope.
Visual inspection is necessary but not sufficient. Many contaminants, especially mycoplasma and viruses, produce no visible changes. Relying solely on visual inspection is a common pitfall that leads to undetected contamination.
Molecular Detection Methods
For definitive detection, molecular methods are required.
Mycoplasma Detection. The most common methods are PCR, direct DNA staining, and enzymatic assays. PCR amplification of the 16S rRNA gene, which is conserved across mycoplasma species, can detect as few as 10 to 100 copies of mycoplasma DNA. A typical protocol uses primers that amplify a 280–300 base pair fragment of the 16S rRNA gene, with 35 to 40 cycles of amplification. The PCR product is then visualized on a 1.5% to 2% agarose gel. Alternatively, the Hoechst 33258 stain binds to DNA and can be used to visualize mycoplasma as small, fluorescent dots around the nucleus of cultured cells. This method is less sensitive than PCR but does not require specialized equipment.
Bacterial and Fungal Detection. Standard microbiological culture—plating a sample of the culture medium on blood agar or tryptic soy agar and incubating at 37°C—can detect bacterial and fungal contaminants within 24 to 48 hours. However, many contaminants are slow-growing or fastidious, so a negative culture does not rule out contamination. PCR-based methods targeting the 16S rRNA gene (for bacteria) or the internal transcribed spacer (ITS) region (for fungi) are more sensitive and faster.
Viral Detection. Detection of viral contaminants typically requires PCR or reverse transcription-PCR (RT-PCR) targeting conserved viral sequences. For example, BVDV can be detected using primers specific to the 5' untranslated region of the viral genome. For endogenous retroviruses, assays may measure reverse transcriptase activity in the culture supernatant.
Cell Line Authentication
Cross-contamination can only be detected through authentication methods that confirm the identity of the cells. The gold standard is short tandem repeat (STR) profiling, which is analogous to human forensic DNA fingerprinting.
STR profiling analyzes the number of repeats at specific loci in the genome. Human cell lines are typed at a set of 8 to 16 core STR loci (such as D5S818, D13S317, D7S820, and the amelogenin locus for sex determination). Each cell line has a unique combination of repeat numbers at these loci, producing a DNA fingerprint. The profile is compared against a reference database (such as the ATCC or DSMZ databases) to confirm the cell line identity.
The process involves extracting genomic DNA from the cells, amplifying the STR loci by PCR using fluorescently labeled primers, and analyzing the fragment sizes by capillary electrophoresis. The resulting electropherogram shows peaks corresponding to each allele. A match between the sample and the reference profile confirms the cell line identity; a mismatch indicates either cross-contamination or misidentification.
STR profiling is required by many journals and funding agencies before publication of results involving human cell lines. For non-human cell lines, species-specific PCR or isoenzyme analysis can be used, but these methods are less discriminating than STR profiling.
Consequences of Contamination
Effects on Experimental Data
Contamination corrupts experimental data in multiple ways. Microbial contaminants consume nutrients, produce toxic metabolites, and alter the pH of the medium, all of which change the physiology of the cultured cells. A cell line stressed by bacterial or fungal contamination will show altered gene expression, reduced proliferation, and increased apoptosis, even if the contamination is not visible.
Mycoplasma contamination is particularly damaging because it is often undetected. Mycoplasmas compete with host cells for amino acids, particularly arginine, which is essential for protein synthesis. They also produce nucleases that degrade host DNA and RNA, and they can induce chromosomal aberrations and morphological changes. A mycoplasma-contaminated culture may show reduced growth rates, altered enzyme activities, and changed responses to drugs or growth factors—all without any visible sign of contamination.
Cross-contamination has even more profound effects. If a culture labeled as a slow-growing fibroblast line is actually overgrown by HeLa cells, every experiment performed on that culture is actually studying HeLa biology. The published results may be internally consistent and reproducible, but they are entirely invalid for the intended cell type. This is why cross-contamination is sometimes called the "invisible fraud" of cell culture.
Broader Scientific Impact
The impact of contamination extends beyond individual experiments. Contaminated cell lines have been used in thousands of published studies, leading to a substantial body of literature that is unreliable or outright wrong. The financial cost is enormous: reagents, time, and salaries are wasted on experiments that cannot be replicated. The scientific cost is even greater: false conclusions can misdirect entire research fields, and clinical applications based on contaminated cell lines can harm patients.
The problem is compounded by the fact that contamination is often not reported. Researchers may discard contaminated cultures without documenting the event, or they may not detect the contamination at all. This lack of transparency means that the true scale of the problem is unknown, and contaminated cell lines continue to circulate in the research community.
Prevention Strategies
Aseptic Technique and Workflow
The most effective prevention strategy is rigorous aseptic technique. This is not a single action but a set of habits that must be practiced consistently.
- Work in a laminar flow hood. All cell culture work should be performed in a Class II biological safety cabinet that provides a sterile, particle-free environment. The hood should be turned on at least 15 minutes before use, and the work surface should be wiped with 70% ethanol before and after each session.
- Wear appropriate personal protective equipment. Gloves, a lab coat, and safety glasses are essential. Gloves should be sprayed with 70% ethanol before placing hands in the hood.
- Use sterile supplies. All pipettes, tips, flasks, and media should be sterile. Never use a pipette that has touched a non-sterile surface, and never insert a non-sterile object into a bottle of medium.
- Minimize aerosol generation. Pipette slowly and gently to avoid creating aerosols. Centrifuge tubes should be capped before centrifugation, and tubes should be opened only inside the hood.
- Clean up spills immediately. Any spill of culture medium or cells should be wiped up with 70% ethanol or a suitable disinfectant, and contaminated materials should be disposed of in biohazard waste.
- Maintain separate media for each cell line. Do not share bottles of medium between cell lines. Each cell line should have its own dedicated bottle of medium, trypsin, and phosphate-buffered saline (PBS).
Use of Antibiotics and Antimycotics
Antibiotics are commonly added to cell culture media to prevent bacterial contamination. Penicillin (100 U/mL) and streptomycin (100 μg/mL) are the most widely used combination, often supplemented with amphotericin B (0.25 μg/mL) as an antimycotic.
However, antibiotics are not a substitute for aseptic technique. They do not eliminate mycoplasma, which lacks a cell wall and is therefore resistant to penicillin and other cell-wall-active antibiotics. They can also mask low-level contamination, allowing it to persist undetected. Moreover, prolonged antibiotic use can select for resistant strains and alter cellular metabolism.
The best practice is to culture cells without antibiotics whenever possible. If antibiotics are used, they should be added only to media for routine maintenance, not to media used for experiments, because antibiotics can interfere with cellular responses to drugs and other treatments.
Quarantine Procedures
New cell lines, whether received from another laboratory or from a cell bank, should be quarantined until they are confirmed to be free of contamination. The quarantine period should include:
- Culture in a separate incubator or a clearly designated area of the lab.
- Mycoplasma testing by PCR or Hoechst staining within the first week.
- STR profiling to confirm the cell line identity.
- Bacterial and fungal testing by plating a sample of the medium on appropriate agar plates.
Only after the cell line passes all tests should it be introduced into the main culture area.
Routine Monitoring and Testing
Regular testing is essential for early detection of contamination. A recommended schedule includes:
- Visual inspection of every culture at each passage.
- Mycoplasma testing every 1 to 3 months, or whenever a new cell line is introduced.
- STR profiling at the time of receipt, at the establishment of a master cell bank, and periodically thereafter (e.g., every 6 months or every 20 passages).
- Bacterial and fungal testing whenever a culture shows signs of contamination or when a new reagent lot is introduced.
Common Pitfalls and Misconceptions
Over-Reliance on Antibiotics
A common mistake among students and even experienced researchers is to treat antibiotics as a safety net. The reasoning is that if the medium contains penicillin and streptomycin, then occasional lapses in aseptic technique will not matter. This is false for several reasons.
First, antibiotics do not kill mycoplasma. Mycoplasma lack a cell wall, so penicillin and other β-lactam antibiotics are ineffective. Second, antibiotics do not eliminate fungal contaminants; amphotericin B is fungistatic rather than fungicidal at typical concentrations, meaning it inhibits growth but does not kill the fungi. Third, antibiotic use selects for resistant bacterial strains, which can then become established in the laboratory and be even harder to eliminate.
The most serious problem with antibiotic use is that it masks contamination. A culture that would otherwise become visibly turbid within 24 hours may remain clear for days or weeks when antibiotics are present, allowing the contaminant to spread to other cultures in the meantime. By the time the contamination is detected, it may be too late to save the experiment or to prevent the spread to other cell lines.
Underestimating Mycoplasma
Mycoplasma contamination is frequently overlooked because it produces no visible signs. Students are taught to look for turbidity, pH change, and microscopic particles, but mycoplasma produce none of these. A culture can be heavily contaminated with mycoplasma and still look perfectly healthy under the microscope.
The prevalence of mycoplasma contamination is high—some surveys suggest that 15% to 35% of cell cultures in routine use are contaminated—yet many laboratories do not test for it regularly. This is a serious oversight. Mycoplasma contamination can alter virtually every aspect of cell biology, from gene expression to drug sensitivity, and it can invalidate experimental results without the researcher ever knowing.
The solution is simple: test for mycoplasma regularly, using PCR or Hoechst staining, and treat any positive culture immediately. Because mycoplasma can be transmitted through shared media and equipment, a single contaminated culture can infect an entire laboratory.
Assuming Cell Lines Are Authentic
Many researchers assume that a cell line received from another laboratory is exactly what the label claims. This assumption is often wrong. Studies have shown that a significant fraction of cell lines in use are cross-contaminated or misidentified. The ICLAC database lists over 500 known cross-contaminated cell lines, and the true number is likely higher.
The consequences of using a misidentified cell line are severe. If a researcher believes they are studying a breast cancer cell line but are actually using a cervical cancer line (HeLa), their results are invalid for breast cancer research. The problem is compounded when the misidentified line is used in multiple studies, leading to a cascade of unreliable publications.
The solution is to authenticate every cell line upon receipt and periodically thereafter. STR profiling is the gold standard for human cell lines, and it should be performed before starting any major experiment. Many journals now require STR authentication as a condition for publication, and funding agencies are increasingly demanding it as well.
Practical Summary and Best Practices
Quick Checklist for Cell Culture
- [ ] Work in a laminar flow hood, wiped with 70% ethanol before and after use.
- [ ] Wear gloves, lab coat, and safety glasses; spray gloves with 70% ethanol before handling cells.
- [ ] Use sterile pipettes, tips, and media; never touch a non-sterile surface with a pipette.
- [ ] Keep separate media and reagents for each cell line.
- [ ] Inspect every culture visually at each passage for turbidity, pH change, or visible particles.
- [ ] Test for mycoplasma by PCR or Hoechst staining every 1 to 3 months.
- [ ] Authenticate new cell lines by STR profiling before use.
- [ ] Quarantine new cell lines until they pass contamination tests.
- [ ] Avoid antibiotics in experimental media; use them sparingly, if at all, in maintenance media.
- [ ] Clean the incubator and water bath regularly; use sterile water in the incubator pan.
- [ ] Document all contamination events and report them to the laboratory supervisor.
Frequently Asked Questions
What is cell line contamination?
Cell line contamination is the unintended introduction of foreign biological material into a cultured cell population. It includes microbial contamination (bacteria, fungi, yeast, mycoplasma, viruses) and cross-contamination with cells from another cell line. Contamination compromises the purity and identity of the culture, making experimental results unreliable.
How can I detect cell line contamination?
Detection methods range from visual inspection (turbidity, pH change, microscopic particles) to molecular methods. Mycoplasma is detected by PCR amplification of the 16S rRNA gene or by Hoechst 33258 DNA staining. Bacteria and fungi can be detected by plating on agar or by PCR. Cross-contamination is detected by STR profiling, which produces a DNA fingerprint unique to each cell line.
What are the most common types of cell line contamination?
The most common types are bacterial contamination (often E. coli, Staphylococcus, or Pseudomonas), fungal contamination (species of Aspergillus, Penicillium, or Candida), mycoplasma contamination (especially M. arginini, M. hyorhinis, and M. orale), and cross-contamination with fast-growing cell lines such as HeLa.
Why is mycoplasma contamination a serious problem?
Mycoplasma contamination is serious because it is invisible—it produces no turbidity, pH change, or visible growth—yet it alters host cell metabolism, depletes nutrients, induces chromosomal abnormalities, and changes cellular responses to drugs and growth factors. Because it is undetected, it can invalidate experiments without the researcher's knowledge. Mycoplasma is also resistant to many antibiotics and can spread easily between cultures.
How can I prevent cell line contamination?
Prevention requires rigorous aseptic technique, including working in a laminar flow hood, using sterile supplies, and minimizing aerosols. New cell lines should be quarantined and tested before use. Antibiotics should be used sparingly, as they do not eliminate mycoplasma and can mask contamination. Routine testing for mycoplasma and periodic STR authentication are essential.
What is cell line cross-contamination?
Cross-contamination occurs when cells from one cell line are inadvertently introduced into a culture of another line. The contaminating cells often grow faster and overtake the intended culture. Cross-contamination is invisible and can only be detected by authentication methods such as STR profiling. It is a major cause of invalid research results.
What is STR profiling and how does it help?
Short tandem repeat (STR) profiling is a DNA fingerprinting method that analyzes the number of repeats at specific loci in the genome. Each human cell line has a unique combination of repeat numbers, producing a characteristic profile. STR profiling confirms the identity of a cell line and detects cross-contamination by comparing the sample profile against reference databases.
Can antibiotics prevent all cell line contamination?
No. Antibiotics are effective against some bacteria but do not kill mycoplasma (which lack a cell wall) and are not reliably fungicidal. Antibiotic use can also mask low-level contamination, allowing it to spread undetected, and can select for resistant strains. The best prevention is rigorous aseptic technique, not antibiotics.
Key Takeaways
- Cell line contamination includes microbial contaminants (bacteria, fungi, mycoplasma, viruses) and cross-contamination with other cell lines; both invalidate experimental results.
- Mycoplasma contamination is the most dangerous because it is invisible and alters host cell biology without producing visible signs.
- Contamination enters cultures through poor aseptic technique, contaminated reagents, incubator issues, and human error; it spreads through aerosols, shared equipment, and shared media.
- Detection requires visual inspection, molecular methods (PCR, staining), and authentication (STR profiling); visual inspection alone is insufficient.
- Prevention relies on rigorous aseptic technique, quarantine of new lines, limited antibiotic use, and routine testing for mycoplasma and cell line identity.
- Antibiotics do not prevent all contamination and can mask low-level infections; they are not a substitute for good technique.
- Every cell line should be authenticated by STR profiling upon receipt and periodically thereafter to prevent the use of misidentified or cross-contaminated cells.
Further Reading
- Gorphe P. A comprehensive review of Hep-2 cell line in translational research for laryngeal cancer. American journal of cancer research. 2019. PubMed 31105993
- Gazdar AF et al. Lung cancer cell lines as tools for biomedical discovery and research. Journal of the National Cancer Institute. 2010. PubMed 20679594
- Gazdar AF, Gao B, Minna JD. Lung cancer cell lines: Useless artifacts or invaluable tools for medical science?. Lung cancer (Amsterdam, Netherlands). 2010. PubMed 20079948
- Canny G. Cell line contamination and misidentification. Biology of reproduction. 2013. PubMed 24078722
- Mohammad TA, Chen Y. Approaching RNA-seq for Cell Line Identification. Bio-protocol. 2020. PubMed 33163581
- Carrillo-Ávila JA et al. Identification of cell culture contamination by an unusual species of Mycoplasma related to the M. mycoides cluster. Cytotechnology. 2023. PubMed 36969572