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Avian Fecal Collection Protocol
Department of Entomology
University of Illinois at Urbana-Champaign
PI: Professor Sydney A. Cameron
Project Supervisor: John Maddux
Contact information:
John Maddux
320 Morrill Hall
505 S Goodwin Ave
Urbana, IL 61801
maddux2@illinois.edu
(573)880-6700
Introduction
Thank you for agreeing to serve as a research collaborator for this study.
This document is intended to serve as a reference for you throughout the
process. If you have any questions that are not addressed by this document,
please do not hesitate to contact John Maddux using the information above.
The aims of this project are to test the long-held belief that birds are major
predators of bumble bees; this has never been conclusively shown to be the
case. Additionally, we hope to identify the major predatory bird species and
estimate the prevalence of bumble bees in their diet. The results have
implications for the foraging ecology of bumble bees and the birds, mimicry
theory in bumble bees, and conservation efforts for both bumble bees and birds.
As a collaborating bird bander, you will perform the vital sample collection
phase of this study. As you perform your normal bird banding activities, you are
asked to collect avian feces as the opportunities present themselves using the
supplied materials according to the protocol given in this document. All collected
materials will then be returned to the Cameron Lab (postage paid) for DNA
analysis. Your vital contributions will be acknowledged in and provided with the
resulting publications.
The Collection Kit
In March 2014 you should receive your collection kit. Each kit contains the
essential materials needed to collect samples. Your kit should contain the
following:
 200 1.5mL microcentrifuge tubes pre-filled with 100% denatured ethanol
 1 permanent pen for labeling vials
 ZipLock storage bags for containing the samples for the return shipment
 200 Parafilm strips for sealing vials prior to return shipment
 1 hard-copy of this protocol for field use (containing your Collaborator ID#)
 1 USPS pre-addressed postage-paid return box (collapsed)
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If additional tubes are needed, please notify the John Maddux with as much
notice as possible; they will be sent immediately. A return shipping label will be
provided via email at the time of the return shipment.
Target Taxa
The purpose of this study is to screen fecal samples for bumble bee DNA
in an attempt to identify major bumble bee predators and to help estimate the
significance of bumble bees to these birds as a food source. Since very little
evidence exists as to which species may be involved or as to how common
bumble bee predation may be, we ask you to collect as broadly and frequently as
possible. At the end of this study, all collected samples will be deposited at the
Illinois Natural History Survey, where it will be maintained as a research
collection. These samples may provide insight into the foraging habits of
temperate North American birds for years to come.
Field Collection Instructions
During normal mist-netting activities, we ask that you collect avian feces when
the opportunity presents itself. An ideal collection is described below:
1. Mist-netted birds are placed into individual cloth bags prior to processing.
2. As each bird is removed from their bag for measurements, the bag is
turned inside-out and any fecal sample present is scooped into a
collection tube pre-filled with ethanol.
3. Each vial is labeled with
a. Minimally:
i. Your Collaborator ID# ____________
ii. The bird’s band number.
b. Ideally also:
i. The date of collection
ii. The four-letter USGS Alpha Code of the bird contributing
bird species
(http://www.pwrc.usgs.gov/bbl/manual/speclist.cfm)
NOTE: 100% ethanol is very effective at dissolving even permanent
inks. Please use caution in preventing exposure of labeled vials of
ethanol. This is especially important immediately after collection,
when vials may be wet with spilled ethanol.
4. Samples collected in the field should be protected from heat and direct
sunlight. A cooler and ice is ideal (be sure to keep vials dry, not directly in
ice), but a shaded spot is acceptable for a few hours.
5. Multiple samples should not be collected from the same bag until the
bag has been washed in order to prevent cross-contamination of the
samples.
6. A bucket of soapy water should be present so that hands can be quickly
washed after collecting each sample. (Note: Hand Sanitizer is not
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recommended, as it may adversely affect bird feathers during handling of
the animals.)
7. After a day’s collection event, samples should be placed on cold
storage. This can include a refrigerator or a freezer. The colder the better.
8. When you run low on collection vials or storage space, contact John
Maddux at the information above. You will receive replacement tubes and,
periodically, pre-paid shipping labels for sending the samples to the
University of Illinois.
Your sampling procedures may vary slightly from those described above, but
the important elements are 1) preventing cross-contamination of the samples,
2) protecting the samples from heat and sunlight, and 3) labeling the vials in
such a manner that a sample can be related to it’s donor bird. If you have
concerns of would like to discuss your protocol with us, please contact John
Maddux using the information listed above.
Need for Immediate Communication
During the course of your collaboration, it is feasible that you may have
questions or that circumstances may arise in the field that require an
alteration of the protocol in order for you to continue collecting. In these
instances, if you feel that you need to touch consult with us please do not
hesitate to contact John Maddux directly at 573-880-6700. Your call will be
answered or returned shortly.
Running Out of Vials
We do not wish your collection to be obstructed by a lack of collection
supplies. If you find yourself running low or for any other reason expect to run
out of vials in around a week, please contact John Maddux using the above
information to request a resupply shipment.
Returning the Samples
Periodically your accumulated samples will need to be returned to the
University of Illinois for screening. You will return batches approximately 200
samples as you reach that number. Your return shipping materials are
designed to accommodate no more than 200 samples at a time.
A note on records: You must include with your samples a copy of your
station’s records that allows for the individual sample tubes labeled
with band numbers to be linked to the bird measurement data collected
at the time of netting. Records should include all data collected on the bird
at the time of banding. Bird data will ideally include at least the band number,
species, approximate age, sex, and size/weight. Please do not provide only
band numbers.
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Package return procedure:
1. Ensure all vials are securely closed and lock-tabs are engaged.
2. Place a Parafilm strip around the lid of each vial.
3. Assemble the provided Priority Mail box.
4. Place your data sheets into the box.
5. Place the biohazard labeled ZipLock bag into the box. (Do not remove
the paper towels.)
6. Place triple-sealed vials in biohazard-labeled ZipLock bag.
7. Seal the box and mail it at any USPS post office or mailbox.
Communication
You will receive weekly communication via email from John Maddux.
These emails will contain any relevant information regarding changes to
protocols or procedures. These emails may also ask you for updates on your
progress. Please take the time to respond with a general statement of your
progress.
Additionally, please feel free to reach out at any point if you need
assistance or feel that there is an issue with the provided protocols. If you
have special circumstances, for instance a very large banding operation
which makes shipping 200 tubes at a time inconvenient, let us know so that
accommodations can be made.
A Note on Integrity of Research
The use of collaborators greatly increases the potential reach and impact
of this study. However, the participation of so many disparate groups with
very different amounts and types of training and expertise opens the research
to all manner of errors or inconsistencies. Please help us minimize this
possibility by following this protocol as closely as possible. If you feel the
need to deviate, please notify us first, then make careful notes of your
alterations. If you face quandaries such as a tube without a label, mixing up
two unlabeled tubes, or realizing that you have collected 10 samples with the
same unwashed bag, please do not assume the errors are harmless. You
should make careful notes of all such instances and, in most cases, discard
the samples in question.
A Note on Permitting and Ethics:
This study is based on non-invasive methods and asks that you collect feces
only when birds defecate during what would be your normal banding
procedures. At no point should you attempt to force birds to defecate,
invasively retrieve feces, or prolong handling times in an attempt to obtain a
sample. Due to the nature of this collection protocol but the University of
Illinois Institutional Animal Care and Use Committee (IACUC) and the United
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States Department of Fish and Wildlife have determined that permits are not
required for this study by Federal Law or the Migratory Bird Treaty Act.
Written correspondence to this affect can be provided upon request. Any
ethical concerns should be reported to John Maddux immediately.
A Note on Postage and Shipping
This protocol involves shipping samples of animal origin as well as
flammable liquid. As a result, it is very important that all instructions relating to
packaging and labeling the returned shipment are followed exactly. If for any
reason you are unable to use the provided packing materials or need to
modify the return process, contact John Maddux using the above information
to ensure compliance.
Regulatory Details
Ethanol is a hazardous flammable material according to USPS Publication 52, Appendix C,
but is allowed to be shipped in this case according to USPS Publication 52, 3 Hazardous
Materials, 33 Hazard Classes, 334 Small Quantity. Packaging guidance is provided by USPS
Publication 52, Appendix C, Packaging Instruction 10A.
Packaging compliance is ensured by the following conditions:
1. Samples will be sent Priority Mail
2. Tubes are Polypropylene and >0.2mm in thickness
3. Tubes are have redundant lock-tab to ensure positive seal
4. Tubes are protected by absorbent packing
5. Tubes are placed within ZipLock bags as “sealed secondary packaging.”
6. Bagged tubes are enclosed in rigid priority mail box as “strong outer packaging.”
7. Outer packaging is labeled “This package conforms to 49 CRF 1734.”
The samples in this study also constitute exempt animal specimens. These items may be
“transported for routine testing not related to the diagnosis of an infectious disease” pursuant
to USPS Publication 52, Appendix C, Packaging Instruction 6H, Reference DMM
601.10.17.9.
Compliance is ensured by the following conditions:
1. Samples are sent Priority Mail
2. Material is triple sealed by the primary container using 1) closed snap-lid, 2) locking
tab that redundantly secures lid, and 3) sealing with a Parafilm strip
3. Primary container contains <500 mL of liquid and <500g of material
4. <500mL of liquid and <500g of material will be sealed within each ZipLock secondary
container
5. Absorbent/cushioning material surrounds the primary containers
6. The secondary ZipLock container is placed inside a Priority Mail outer box
7. Priority Mail boxes are rigid fiberboard
8. Secondary ZipLock container is marked with the international biohazard identifier
9. The outer Priority Mail box bears the words “Exempt animal specimen.”