Symbiotic microbes may mediate songbird chemical signals

Danielle Whittaker

Kevin Theis

Photo by Marine Drouilly

Symbiotic hypothesis for chemical communication

Symbiotic microbes in spotted hyena scent pouches are responsible for odors in scent marks

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Structure (Bray-Curtis)

ANOSIM: R = 0.36, P = 0.002

Fig Tree

SouthernComfort

Mara River

Emarti Hill

Bacterial community structure varies with clan membership

(Theis et al. 2012)

Preen oil is an odor source in birds

Oil secreted from the uropygial gland contains volatile compounds that give birds an odor

Photo by Danielle Whittaker

Volatile compounds in preen oil vary with:

Species

Sex

Population

Relatedness

Breeding condition

Quality …and could play a role in mate choice.

Photo by Marine Drouilly

Do avian preen glands harbor odor-producing bacteria?

Do adults transmit these bacteria to their offspring during the nestling phase?

Dark-eyed junco (Junco hyemalis)Mountain Lake Biological Station, VA

Dark-eyed Juncos

Seasonal breeders

Socially monogamous

~30% extra-pair fertilization (EPF) rate

Females incubate eggs, both males and females feed nestlings

Nestlings fledge at day 12

Photo by Marine Drouilly

Photo by Dawn O’Neal

Field Methods

13 nests, 64 juncos

Swab samples from preen glands:• all nestlings (2-4,

mean = 2.9) at age 11-12 days

• both parents at all but 2 nests

Photo by Marine Drouilly

Extracted DNA from swabs using MO BIO PowerSoil Kits

Illumina MiSeq platform, targeting the V4 region of bacterial 16S rRNA gene

Processed using mothur

Each sample subsampled to 5000 sequences

Bacterial sequences clustered based on 97% nucleotide similarity to define OTUs

Sequencing Methods

Photo by Flickr user Super Bay

Bacterial communities in junco preen glands have very high levels of diversity

Top 20 OTUs account for ~45% of sequences, with no OTU accounting for more than 5%

In hyena scent glands, the top 20 OTUs account for ~90% of sequences, with the top OTU accounting for ~45%

Top 20 OTUs in junco preen glands:

Burkholderiaceae, Burkholderia

Burkholderiaceae, Ralstonia

Clostridiaceae, Clostridium

Clostridiales Family XI, Incertae Sedis XI, Anaerococcus

Comamonadaceae, unknown

Enterobacteriaceae, unknown

Enterobacteriaceae, unknown

Enterococcaceae, Enterococcus

Halomonadaceae, Kushneria

Halomonadaceae, Salinicola

Kineosporiaceae, Kineococcus

Micrococcaceae, unknown

Moraxellaceae, Alkanindiges

Moraxellaceae, unknown

Moraxellaceae, unknown

Unknown, unknown

Pseudomonadaceae, Pseudomonas

Rhodobacteraceae, unknown

Sphingomonadaceae, Sphingomonas

Sphingomonadaceae, Sphingomonas

Other prominent OTUs associated with odor:

Corynebacterium: human axillary odor

Porphyromonas: malodorous breath

Bacteroides, Finegoldia & Fusobacterium: common volatile fatty acid producers associated with many vertebrates

Bacterial community structure varied by nest

Nests significantly different (70/78 pairwise comparisons)

NPMANOVA, all: F = 3.751, P = 0.0001 (Bray-Curtis)

Nestlings only: F = 6.946, P = 0.0001

Photo by Nicole Gerlach

all individuals, NPMANOVA, F = 3.751, P = 0.0001

nestlings only, NPMANOVA, F = 6.946, P = 0.0001

Nestlings were more similar to mother than father

Wilcoxon’s test, N = 34, W = 564, P < 0.0001

Junco preen gland bacterial communities

Highly diverse

Cluster by nest

Nestlings closely resemble each other and mother

Reliable transmission across generations via physical contact

Acknowledgments

Photo by Nicole Gerlach

Tracy Teal

Arvind Venkataraman

Ellen Ketterson

Samuel Slowinski