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Collaborator ID Number: ___________
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
[email protected]
Project Web Page:
I…………………. Introduction
II………………… The Collection Kit
III…………………Target Taxa
IV…………………Field Collection Instructions
V………………….Need for Immediate Communication
VI…………………Running Out of Vials
VII………………...Returning the Samples
VIII………………..General Communication
IX………………….A Note on Integrity of the Research
X…………………..A Note on Permitting and Ethics
I. 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 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
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analysis. Your vital contributions will be acknowledged (name and institution) in
all resulting publications.
II. 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
1. 1.5mL microcentrifuge tubes pre-filled with 100% ethanol (120)
2. permanent marking pen for labeling vials (1)
o NOTE: Please use care to avoid spilling ethanol onto labeled vials,
as ink may run.
o NOTE: Early versions of the collection kit contained Pigma Micron
pens, which were resistant to ethanol. Do not use Pigma pens to
mark vials, however, as the ink wipes off when abraded.
3. ZipLock storage bag for containing the samples for the return shipment (1)
4. Parafilm strips for sealing vials prior to return shipment (120)
5. hard-copy of this protocol for field use (containing your Collaborator ID#)
6. USPS pre-addressed postage-paid return box (collapsed, 1)
If additional vials are needed, please notify the John Maddux with as much notice
as possible, and they will be sent immediately.
III. 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, along with your
collection notes, as a research collection. These samples may provide insight
into the foraging habits of temperate North American birds for years to come.
IV. Field Collection Instructions
During normal mist-netting activities, we ask that you collect avian feces when
the opportunity presents itself. We understand that all banders work in different
circumstances, but 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.
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3. Each vial is labeled with
a. Minimally:
i. Your Collaborator ID# ____________
ii. Bird’s band number (or other unique identifier, if applicable)
b. Ideally also:
i. The date of collection
ii. The four-letter USGS Alpha Code of the bird contributing
bird species
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. Please use only the
provided pen if possible, as the ink is alcohol-fast
4. Samples collected in the field should be protected from heat and direct
sunlight. A cooler and ice is ideal, but a shaded spot is acceptable for a
few hours while collecting.
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
6. A bucket of soapy water should be present so that hands can be quickly
washed after collecting each sample. This prevents cross-contamination
of the excreta samples (Note: Hand Sanitizer is not 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, contact John Maddux at the
information above. Your next batch of vials will be shipped immediately.
Please do not wait until you are out of empty vials, as samples may be
missed during the shipping lag.
NOTE: 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 at [email protected]
V. 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.
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VI. 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 place a resupply request online or
contact John Maddux directly at [email protected]
VII. Returning the Samples
Periodically your accumulated samples will need to be returned to the
University of Illinois for screening. Vials are sent to you in batches of 120, and
your return shipping material is designed to accommodate that number.
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.
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. Stretch the Parafilm
(like clingwrap) around the circumference of the lid; it will self-adhere
3. Assemble the provided Priority Mail box.
4. Place your data sheets into the box.
5. Place your sealed sample vials into the provided ZipLock bag with the
biohazard indicator. (Do not remove the paper towels.)
6. Remove as much air from the bag as possible and seal it.
7. Place the bag into the Priority Mail Box.
8. Fold and place copies of your collection notes into the remaining space
at the end of the box.
9. Seal the box and mail it at any USPS post office or mailbox.
10. Notify us that the box has been sent using either the by contacting
John Maddux at [email protected]
VIII. General Communication
The website, will serve
as a useful resource for you during your time as a collaborator. New
information, instructions, tutorials, and announcements will be posed
regularly. The website will continue to expand during the coming months.
In addition, you will receive a 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
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updates on your progress. Please take the time to respond with a general
statement of your progress.
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 120 tubes at a time inconvenient, let us know so that
accommodations can be made.
IX. 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.
X. 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
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. Before
shipping your samples to the Cameron Lab, you will be asked to complete a
Permit Disclosure form, declaring your legal permission to handle protected
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 at [email protected]
Regulatory Details
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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
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
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.”
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