New York City (NYC) Medium: Composition, Principle, Uses, and Colony Characteristics

A 26-year-old woman presents with vaginal discharge and lower abdominal pain. Her clinician suspects pelvic inflammatory disease (PID) — a condition that can be caused by Neisseria gonorrhoeae, Mycoplasma hominis, or Ureaplasma urealyticum, either alone or in combination. An endocervical swab is sent to the laboratory with a request for culture for gonococci and genital mycoplasmas.

Modified Thayer-Martin agar — the most widely used gonococcal culture medium — cannot support Mycoplasma or Ureaplasma growth. Culturing these organisms requires a completely different medium. New York City medium was designed precisely for this scenario: a single selective plate that simultaneously recovers N. gonorrhoeae, M. hominis, and U. urealyticum from the same genital specimen, allowing the laboratory to address all three pathogens in one culture setup.

New York City agar base was developed initially by Fauer, Weisburd, and Wilson at the New York City Department of Health for selective isolation of pathogenic Neisseria species from clinical specimens.

New York City (NYC) medium is primarily designed to isolate pathogenic Neisseria. It also supports the growth of genital mycoplasmas (Mycoplasma hominis and Ureaplasma urealyticum). New York City (NYC) medium is helpful in the diagnosis of gonorrhea and recognition of active or asymptomatic mycoplasma infections. It is a transparent medium.

Figure: Neisseria gonorrhoeae in New York City Medium (Imagesource)

Principle

NYC medium is both selective (suppressing normal flora while allowing Neisseria and genital mycoplasmas to grow) and transparent (enabling observation of the distinctive small colony types produced by these organisms).

Why NYC medium supports Mycoplasma and Ureaplasma when MTM does not:

Mycoplasma hominis and Ureaplasma urealyticum are the smallest self-replicating bacteria and lack a cell wall entirely. They have two distinctive growth requirements that standard Neisseria media cannot meet:

1. Cholesterol and lipids — Mycoplasmas incorporate exogenous cholesterol into their cell membranes (they cannot synthesise their own). Horse plasma in NYC medium is a rich source of cholesterol and phospholipids that Mycoplasmas require. GC agar base in MTM contains no plasma.
2. Enriched peptide and nucleotide environment — Mycoplasmas have highly reduced genomes with limited biosynthetic capacity, making them dependent on preformed nucleotides and amino acids in the medium. The combination of horse plasma, horse hemoglobin, and yeast autolysate in NYC medium provides this enriched nutritional environment.

Component roles:

The yeast autolysate CO2 mechanism — a distinctive feature of NYC medium: N. gonorrhoeae is capnophilic — it requires elevated CO2 (5–10%) for optimal growth. Yeast autolysate contains oxaloacetic acid, which gonococci metabolise intracellularly to produce CO2. This means NYC medium partially self-supplies the CO2 requirement, reducing (though not eliminating) the dependence on a CO2 incubator or candle jar compared to MTM. This is particularly advantageous in resource-limited settings where CO2 supply is inconsistent.

Transparent medium — why it matters: Unlike the opaque red-brown appearance of MTM or chocolate agar, NYC medium is transparent to translucent. This transparency allows direct microscopic examination of colonial morphology on the plate and makes it easier to identify the tiny "fried-egg" colonies of genital mycoplasmas alongside larger gonococcal colonies.

Uses of New York City Medium

1. Primary isolation of Neisseria gonorrhoeae from urogenital specimens NYC medium is used for the same clinical indications as MTM: urethral swabs, endocervical swabs, vaginal swabs, rectal swabs, and pharyngeal swabs from patients with suspected gonorrhoea.

2. Simultaneous isolation of genital mycoplasmas alongside Neisseria The unique advantage of NYC medium is its ability to support Mycoplasma hominis and Ureaplasma urealyticum growth from the same plate. This is clinically relevant in:

• Pelvic inflammatory disease (PID) — where both N. gonorrhoeae and M. hominis are implicated
• Non-gonococcal urethritis (NGU) — where U. urealyticum is a recognised cause
• Postpartum fever and neonatal infections — where M. hominis transmission from mother to neonate occurs
• Infertility investigations — where genital mycoplasma colonisation is under study

3. Neisseria meningitidis isolation from nasopharyngeal specimens NYC medium supports meningococcal carrier screening from nasopharyngeal swabs, where a mixed upper respiratory flora requires selective suppression.

4. STI clinic settings requiring multi-pathogen coverage In sexual health clinics processing high volumes of genital specimens, NYC medium offers the advantage of detecting both gonococcal and mycoplasmal infections from a single plate, potentially reducing the number of media required per specimen.

Practical note — NYC medium availability: NYC medium requires horse plasma and horse hemoglobin, making in-house preparation more complex than MTM (which uses sheep blood). In settings where these components are unavailable, MTM remains the standard. Commercially prepared NYC medium plates are available from major manufacturers but at higher cost than MTM.

Composition of New York City medium agar base

Final pH ( at 25°C) 7.4±0.2

Procedure for the preparation of New York City medium

1. Suspend 25.50 grams in 320 ml distilled water.
2. Heat to boiling to dissolve the medium completely.
3. Sterilize by autoclaving at 15 lbs pressure (121°C) for 15 minutes. Avoid overheating.
4. Cool to 45-50°C and add aseptically 100 ml of sedimented horse blood cells and 60 ml of citrated horse plasma along with rehydrated contents of 1 vial of NYC Supplement and 1 vial of yeast autolysate supplement.
5. Mix well and pour into sterile Petri plates.

Colony Characteristics on NYC Medium

Neisseria gonorrhoeae: Colonies appear at 24–48 hours. They are small (0.5–1 mm), grey-white to colourless, raised, convex, smooth, and glistening — identical in appearance to colonies on MTM. The transparent medium background makes these colonies easier to visualise than on the opaque background of MTM. Colony types T1 and T2 (piliated, virulent) predominate in fresh clinical isolates; T3 and T4 (non-piliated) emerge on subculture. All colony types are oxidase-positive — confirmed by tetramethyl-p-phenylenediamine reagent turning colonies dark purple within 10 seconds.

Neisseria meningitidis: Larger (1–2 mm), grey to bluish-grey, mucoid colonies due to polysaccharide capsule. Slightly larger and more mucoid than gonococci. Oxidase-positive. Maltose-positive (unlike gonococci — the key biochemical differentiator).

Genital mycoplasmas on NYC medium: Mycoplasma and Ureaplasma colonies are very small and require careful examination — often magnification with a stereomicroscope or hand lens:

Identifying mycoplasma colonies on NYC medium: Because mycoplasma colonies are so small relative to gonococcal colonies, they can be missed during routine plate reading. After 48–72 hours, hold the NYC plate up to oblique light and scan the plate surface systematically with magnification. The fried-egg morphology of M. hominis is distinctive once recognised — the flat, transparent periphery growing out from the opaque centre is unlike any standard bacterial colony.

Choosing Between NYC, MTM, and Martin-Lewis for Neisseria Isolation

Decision rule:

• Need both Neisseria and genital Mycoplasma/Ureaplasma covered: NYC medium
• Standard gonorrhoea workup, no Mycoplasma concern: MTM
• MTM nystatin batches showing variability in selectivity: Martin-Lewis (anisomycin is more stable)
• Sterile site specimens (joint fluid, blood) or culture-negative cases: Add non-selective chocolate agar alongside any selective medium — vancomycin-sensitive gonococci will be recovered.

How to Remember

NYC medium's defining advantage over MTM is horse plasma + yeast autolysate = Mycoplasma support + CO2 supply.

Two ingredients do the work that distinguishes NYC from all other Neisseria media:

1. Horse plasma → cholesterol for Mycoplasma membrane → enables M. hominis and U. urealyticum growth
2. Yeast autolysate → oxaloacetic acid → gonococci metabolise it to CO2 → partial self-supply of the capnophilic requirement → reduces CO2 incubator dependence + reduces lag phase

The transparent medium as a mnemonic: NYC medium is transparent; MTM and Martin-Lewis are opaque. The transparency was deliberate — tiny mycoplasma colonies that would disappear against a dark background become visible against a transparent background. If a medium is clear or translucent and used for Neisseria, think NYC.

The four-antibiotic parallel with MTM: Both NYC and MTM use vancomycin + colistin + trimethoprim for selectivity. The antifungal is the differentiator:

• NYC: Amphotericin B (A for America's NYC)
• MTM: Nystatin
• Martin-Lewis: Anisomycin

Clinical scenario anchor: Any time a patient has symptoms that could involve both N. gonorrhoeae AND genital Mycoplasma (PID, NGU, postpartum fever), NYC medium is the logical choice because it covers both pathogens on a single plate. MTM would miss the Mycoplasma entirely.

Limitation and recommendation

1. Some strains of N. gonorrhoeae are inhibited by the concentration of vancomycin in the selective media, so the addition of nonselective chocolate agar is recommended, especially in suspect cases that are culture negative or for sterile specimens (e.g., joint fluid)

References

1. Faur, Y. C., Weisburd, M. H., Wilson, M. E., & May, P. S. (1974). A new medium for the isolation of pathogenic Neisseria from clinical material. Applied Microbiology, 27(6), 1041–1045. https://doi.org/10.1128/am.27.6.1041-1045.1974
2. Tille, P. M. (2017). Bailey and Scott's Diagnostic Microbiology (14th ed.). Elsevier.
3. Greenwood, J. R., Voss, J., Smith, R. F., Wallace, H., Peter, C., Nachtigall, M., Maier, T., Wilber, J., & Butsumyo, A. (1986). Comparative evaluation of New York City and modified Thayer-Martin media for isolation of Neisseria gonorrhoeae. Journal of Clinical Microbiology, 24(6), 1111–1112.
4. World Health Organization. (2016). WHO Guidelines for the Treatment of Neisseria gonorrhoeae. Geneva: WHO.