YM agar preparation (Nymphalidae)

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Method synopsis

Method to quantify microbial load on different butterfly body parts by direct plating on agar media. Samples tarsi/proboscis by dragging across plate and thorax by washing technique.

Synthesized from method descriptions across 2 papers using this protocol.

Procedure from a recent peer-reviewed implementation

Steps below were extracted from the most recent peer-reviewed implementation of this protocol in the corpus — The dispersal of microbes among and within flowers by butterflies (2023), Ecological Entomology. The protocol was originally introduced by Butterfly foraging and the dispersal of microbes among & within flowers (2020). Implementations in other papers (listed below) may differ.

1. Collect wild butterflies
   Collected 10 Glaucopsyche lygdamus and 31 Speyeria mormonia adults from the field site using individual autoclave envelopes to prevent contamination. Recorded species, sex, and wing wear as a proxy for age using a 1-5 scale with 0.5 gradations (1.0 being fresh, 5.0 being worn).
   Quantities: 41 total butterflies (10 G. lygdamus, 31 S. mormonia)Duration: Not specifiedConditions: Field conditions at Rocky Mountain Biological Laboratory site
   Equipment: autoclave envelopes
2. Culture microbes from butterfly body parts
   Used sterile technique to plate three samples from each butterfly by: (1) walking the butterfly across YM agar plate, (2) plating a wash of the thorax, and (3) extending the proboscis to touch the plate. Held butterflies so body was perpendicular to agar surface while dragging proboscis length across plate. Counted microbial colonies that grew after 2 days at room temperature.
   Quantities: 3 samples per butterfly, incubated for 2 daysDuration: 2 days incubation at room temperatureConditions: Room temperature incubation
   Equipment: YM agar plates, sterile needle-tip forceps

Typical Equipment

• YM agar plates
• sterile needle-tip forceps
• autoclave envelopes

Output Measurements

• microbial colony counts by body part

Papers Using This Protocol (2)

Species Studied With This Protocol

Places Where Used