Author: Terrence A. Lynch
Specializing in the study of fireflies and rearing of bioluminescent small animals
Homepage: (http://members.aol.com/terrylynch/fireflies.htm)
Email: terrylynch@aol.com
Published on-line at: http://www.geocities.com/bioluminus/anurida_granaria.html
ABSTRACT: Anurida granaria (Nicolet, 1847), a bioluminescent species of Collembola, is described, along with its history and global distribution which make the species an excellent organism for international transport which may NOT require permitting as is often required of endangered or threatened species, etiologic agents or other organisms which may be harmful to agriculture or trees. Methodology for handling, rearing and suggested applications as an educational tool is presented defining A. granaria as an ideal candidate for teaching and demonstration of basic entomology techniques throughout the world academic community.
Introduction
Anurida granaria (Nicolet, 1847), is a pseudachorutinae Springtail occurring globally in the arctic and sub-arctic, Europe, N. Eurasia, Mediterranean, Micronesian, and Central Australia. The N. North America and Pacific North America are "doubtful regions" of occurrence, this biogeographical regions for A. granaria as per Christiansen and Bellinger (1995)(See extensive on-line biography of Collembola at http://www.collembola.org/doc/bibliogr.htm#0628).
Specimens were collected by Graham East of Christchurch, New Zealand and identified by Wanda M. Wiener who was selected to make this identification, as an unknown species was collected and observed to be exhibiting bioluminescence by Graham East. This was originally thought to be an Onychiuridae; hence, from review of W. M. Wiener (1996) and other publications by the same author, it was concluded Wierner would be ideally qualified to make an identification. Specimens under consideration were identified by W. M. Wiener to be cosmopolitan and to include A. granaria. Additional isolation and identification of only bioluminescent individuals proved A. granaria was the species of Collembola Graham East observed to exhibit bioluminescence. (Lynch 2001a)(http://www.geocities.com/bioluminus/introduction.html). Lynch confirmed bioluminescence of A. granaria in live specimens he view on 3 Dec. 2001 (Anurida granaria exhibits unique bioluminescent display). Graham East had originally contacted Lynch as an expert on the bioluminescence of small animals, namely due to Lynch's extensive pioneering studies of the firefly Photinus pyralis (Lynch 2000).
Specimens were reared according to methodology detailed and adapted by Michael L. Draney (2000)(http://www.collembola.org/publicat/culture.htm). Draney states that this was basically the same methodology used by Louis Lipovsky, a graduate student at The University of Kansas, who reared Sinella curviseta Brooks 1882 to use as food for postlarval chiggers (Acari: Trombiculidae)(Lipovsky 1951, 1954).
Methodology
This methodology for the collection, rearing and handling of Anurida granaria (Nicolet, 1847) as applicable in the enhancement of the educational and research process was originally posted at http://www.geocities.com/bioluminus/methodology.html by Terrence A. Lynch 2001.
Purpose
Anurida granaria (Nicolet, 1847) is a cosmopolitan species of Collembola. Specimen collected by Graham East in and around Christchurch, New Zealand, were observed to exhibit bioluminescence when disturbed. In order to enable this behavior to be observed and to make the species available to schools and universities throughout New Zealand, methods were develop for their collection, rearing and handling. Although these techniques may be utilized by experts, they are equally adaptable to students and teachers who would like to use this species in the classroom. The fact that A. granaria is a small, harmless and relatively easy to rear species, one which does not transmit disease to crops or trees and which is not a vector for human disease, this makes it an ideal bioluminescent species for classroom study. Given also that A. granaria has a wide distribution, pure stock may be safely be distributed throughout New Zealand without any harm to the ecosystem. Add to this the fact this species exhibits bioluminescence, and it becomes evident that it is an excellent species to offer teachers, students and researchers. It is therefore hoped this methodology will stimulate an interest in the species and its wide usage throughout the scientific and educational community.
Collection and Handling
Initial collection of A. granaria was made by Graham East in the Christ Church area on South Island, New Zealand. East had known of the species for many years, having chanced to observe large glowing colonies under bricks, logs and other ground cover.
Figure 1 (http://www.geocities.com/bioluminus/collecting_ag.jpg). Initial field collection of A. granaria (see inset). Specimens are brushed off the underside of a brick onto a black plastic tray, then examined with a magnifier. Later in a darkened room bioluminescent individuals are separated from specimens which fail to glow when stimulated with a gentle puff of air. Photo by Graham East
When specimens were collected and sent for analysis it was found that these natural colonies consisted of a mixed group of species. Therefore it became necessary to individually and carefully pick out those individuals which were exhibiting bioluminescence. This was done with great patience and skill using mouth operated aspirator (http://www.geocities.com/~fransjanssens/images/collect/aspirat.gif ) to secure glowing specimens from those of a group. Selection from among cosmopolitan Collembola was made by placing singular individuals in small test tubes or vial under light and setting aside to rest; then in absolute darkness test tubes were picked up. Any vibration as by tapping or jarring the test tube was sufficient to result in a display of bioluminescence and enable selection from non-bioluminescent individuals.
Figure 2 (http://www.geocities.com/bioluminus/mouth_aspirator.jpg). Mouth aspirator of style used to collect A. granaria or other small insects. The aspirator shown here was made using an empty spice bottle and tubing obtained from a hardware store. A very fine nylon mesh was used around the tubing on the inside of the aspirator to prevent sucking Collembola into the mouth. Photo (c) 2001 by Terry Lynch
Specimens so collected were preserved in alcohol and sent for analysis. It was thus established that A. granaria was the species which was bioluminescent. This determination was made by Wanda M. Weiner Ph.D., Head of the Laboratory of Invertebrates at the Institute of Systematics and Evolution of Animals in Krakow, Poland (http://www.isez.pan.krakow.pl/staff/wwei.htm).
Live specimens were placed in containers for rearing and maintaining pure stock.
Pure Stock
Because A. granaria is cosmopolitan, specimens collected in the field were a mix of different species. Once this was established, A. granaria was carefully isolated, selecting only for bioluminescent individuals. Then from these individuals pure stock was established.
Although this may sound like an easy task, it certainly was not. A. granaria is a very tiny animal measuring only about 1.7 mm in length. The only way to distinguish A. granaria from other species is careful microscopic examination of preserved specimens by an expert. That is, in daylight one Collembola looks very much like another and it is impossible to tell which one is bioluminescent. Thus selection had to be done in darkness, using an aspirator to collect only those Collembola producing light.
Figure 3 (http://www.geocities.com/bioluminus/ag_rearing15nov2001.jpg). When rearing A. granaria it is important to maintain pure stock. Here A. granaria are seen being reared upon moist peat moss. Yeast has formed a mycelium network upon which the A. granaria feed. Photo by Graham East
Over time significant numbers of bioluminescent individuals were collected and isolated from which pure stock was obtained. This became very apparent for when one has a container of A. granaria it only takes a gentle blowing upon them to cause bioluminescence. Also one then finds that all individuals are similar in every aspect of their appearance and expert analysis of a representative sample shows that only one species is present.
Because obtaining a pure stock in this manner is a difficult, time consuming process which requires the assistance of an expert Collembola taxonomist, it is recommended that those wishing to obtain A. granaria simply order a pure stock. This avoids duplication of effort, saves time and money and is really the most sensible way to obtain A. granaria now that a pure stock has been established.
Rearing
As reported by Michael L. Draney (daneym@uwgb.edu) of the Dept. Natural and Applied Sciences, University of Wisconsin-Green Bay, Collembola can be reared on a moist substrate (http://www.collembola.org/publicat/culture.htm). Moist soil or peat moss works adequately. For better results, Draney says one may use a plaster of Paris/powdered charcoal substrate kept moist with aged tap water. Small plastic or glass jars provide with tight fitting lids are the recommended rearing vessel. Collembola may be fed powdered baker's yeast and watered added at least once per month.
Figure 4 (http://www.geocities.com/bioluminus/ag_rearing_kit.jpg). Basic materials recommended for rearing A. granaria. Place sterile peat moss or other substrate in a one-half pint jar. Add baker's yeast. Use eye dropper or red sable brush in handling fragile A. granaria. Use filter paper or cut double folded paper towel to provide a porous lid or use regular canning lid. Add distilled water as necessary to maintain moisture. Photo (c) 2001 by Terry Lynch
In keeping with these recommendations it was found that A. granaria do indeed thrive upon moistened peat moss or a plaster of Paris/powdered charcoal substrate. Rearing at room temperature as has worked well with Sinella curiseta Brook 1882 (Gist, Crossley, and Merchant, 1974) has proven adequate for A. granaria. A primary consideration may be maintaining a pure stock free of species of Collembola which are normally found with A. granaria. Also pure stock must be kept free of mites. In order to establish and maintain pure stock care must be used to begin only with specimens which are of a single species, i.e., A. granaria. In this case that was achieved by selecting only bioluminescent individuals to use in establishing stock. To insure that stock was only of a single species, a single large, bioluminescent adult was selected, washed and introduced into a rearing vessel with substrate. This was repeated to provide greater odds of isolating a single adult fertile female. Once these females produced young, they were reared. The stock was checked to insure that no mites were present and that only A. granaria was present. Over time this process was repeated to insure that the resulting stock was a singular species and free of mites.
When rearing A. granaria or any species of Collembola for that matter, it is best to use great care to avoid introduction of mites from the very start of one's rearing program. Because mites may occur in samples of soil where cosmopolitan Collembola occur, one should NOT use a Berlese funnel to establish stock, as obviously mites would be collected. The better method is careful collection of singular individuals as above stated. Yet even after establishing a culture on a sterile substrate one should check for mites. This may be done using floatation. When a sample of substrate is flooded with water and left 24-48 hours, mites float to the top and may easily be seen, collected and observed under a microscope. When one uses this method it is easy to establish whether or not a stock has been established which is free of mites. It is recommended that in effort to maintain pure cultures this method be used to regularly check for mites and insure that stock does not become contaminated.
To avoid the introduction or A. granaria into areas where it does not occur naturally all material, such as peat moss and other substrate used in rearing should be disposed of by burning. Glassware should be sterilized in an autoclave. The following methods of sterilization are recommended by the US Department of Agriculture, APHIS as a means for killing pathogens associated with import/export of animal products, namely pet chews (http://www.aphis.usda.gov/NCIE/lhpetch.shtml) and would certainly be adequate to sterilize equipment or materials used in the rearing of Collembola.
Dry heat at 180 degrees Fahrenheit (82.2 C) for 30 minute
Soaking in 0.1% chlorine bleach for 2 hours;
Soaking in either 5% acetic acid or 5% hydrogen peroxide solution for 2 hours; or
Soaking in boiling water for 20 minutes.
Safe for Export/Import
For a species to be acceptable for export/import in the United States it must meet a number of requirements or else require special permitting from government agencies:
(1) The species must not be an endangered or threatened species defined by CITES (Convention on International Trade in Endangered Species) or be likely to become a threatened or endangered species if not regulated by USDA, Fish & Wildlife Service (http://international.fws.gov/permits/dmapermits.html).
(2) The species must not be an etiologic agent (microorganism which causes disease in humans) as defined by Centers for Disease Control (CDC) and Prevention (Importation Permits for Etiologic Agents, Centers for Disease Control and Prevention, Office of Health and Safety)(http://www.cdc.gov/od/ohs/biosfty/imprtper.htm).
(3) The species must not be injurious to agricultural crops or trees or be a vector for disease causing organisms which may infect animals or plants as restricted by the United States Department of Agriculture (USDA) Animal and Plant Health Inspection Service (APHIS)(http://www.aphis.usda.gov).
(4) Specimens must be easily reared in vitro, free of contaminants; field collection must not be likely to endanger existing wildlife populations occurring in the natural environment.
A. granaria meets each of these provisions. Its wide global distribution means that it is not an endangered or threatened species nor is it likely to become one anytime soon. Specimens collected in preparation of this report occurred in an urban environment so did not even impact wildlife populations.
A. granariais NOT an etiologic agent as it does not transmit any diseases to humans or animals and is not a vector for transmission of disease. In fact A. granaria is quite harmless and lives a gentle life, retired under bricks, logs and other ground cover eating yeast mycelium.
A. granaria does not damage crops or trees and is NOT a vector for any diseases injurious to plants or animals. In fact because A. granaria feeds upon mycelium it is a beneficial insect which aids in the breakdown of organic material vital to the healthy and vigorous growth of agricultural crops. In fact, quite often Collembola are added to compost piles to aid in the breakdown of organic material.
As indicated in this report, A. granaria is certainly easy to rear and maintain in vitro; hence pure stock free of mites, flies or other soil contaminates are easy to establish and maintain in the laboratory upon sterile substrate. The collection of small numbers of specimens required to establish a stock in not likely to have any impact whatsoever upon colonies occurring in the wild which will regenerate themselves from eggs occurring in soil and individuals in the surrounding environment.
Each of these factors contributes to making A. granaria an ideal species for rearing and teaching purposes, especially with respect the study and demonstration of bioluminescence in insects. Unlike other species of small bioluminescent animals which may be impacted by human population growth, as is the case with fireflies (Lynch 2001c, d), A. granaria occurs in such great abundance around the globe, even in urban areas where it takes up residence in yards and gardens, that it will likely remain a species forever a resident of the planet Earth. A. granaria is therefore a completely safe and harmless species and should be adopted by the world academic community for general research, teaching and other educational purposes.
These factors combine to make pure stock of A. granaria reared upon sterile substrate a safe species for export/import.
Beneficial Insect Species
The harmless nature of A. granaria combines with other factors which distinguish A. granaria as a beneficial insect species. These include the following applications and usage:
(1) Pet Food -- Collembola which are easy to rear, as are A. granaria, have been used as pet food for a variety of small live animals including: spiders, predacious mites, chiggers, (Lipovsky 1951, 1954) mantids and reptiles and amphibians.
(2) Instructional Aid -- A. granaria may be used in the teaching of basic entomology techniques and to demonstrate bioluminescence in insects.
(3) Organic Farming and Terra Farming -- Given A. granaria aid in the decomposition of organic material they may be a valuable composite in compost as to organic farmers. This is especially true in closed systems such as biodomes and terra farming, indicating that A. granaria may have a vital role to play in development of artificial living environments in space habitats and upon the Moon, Mars and other extra terrestrial bodies where humans may wish to establish habitates in the future.
(4) Research Tool and Biological Indicator -- Because A. granaria is a bioluminescent Collembola the light it produces may enable a way to measure the health of living organisms in soil using photometric devices. Hence A. granaria have great potential as a research tool and biological indicator such that a healthy A. granaria population may indicate a healthy living environment for other species as well.
Together these factors combine to establish A. granaria as a beneficial insect species which universities and agricultural institutions throughout the world may wish to add to their permanent collections of small live animals. Certainly A. granaria has great potential as a beneficial insect which demands that A. granaria be further studied and distributed throughout the academic community.
Because A. granaria is a safe, harmless and beneficial insect species pure stock established upon sterile substrates should be permitted expoxt/import between all nations without any prohibition, restriction, quarantine or other delay in shipment which might cause injury to specimens in transport. Customs agents and special inspectors may be reasonably certain that pure stock of A. granaria represent a permitted species by all definitions of the term.
Suggested Experiments
A. granaria are an ideal species for teachers to use in the classroom to demonstrate bioluminescence in insects. All that is required is to observe A. granaria in a perfectly darkened room. By gently blowing upon A. granaria they will briefly glow. When this is seen for the first time by young and old alike, it results in an impressive feeling of awe and amazement. What better way to encourage an interest in science and learning that using A. granaria to demonstrate bioluminescence and stimulate the learning process!
A. granaria are also ideal for a variety of experiments. For example, one may maintain stock under different periods of light and dark to determine how this effects bioluminescence. It should be found that when kept under a 12/12 daylight/dark period, A. granaria will only exhibit bioluminescence during the dark phase. This circadian rhythm may prove to be an interesting topic for further investigation.
A. granaria is also ideal for studying the effect of diet or other variables upon bioluminescence. Simply add different chemicals to the substrate upon which A. granaria feed and see what happens. For example does the pH of a substrate, variation in diet or addition of other chemicals effect bioluminescence?
Figure 5 (http://www.geocities.com/bioluminus/jewel_box_inset.jpg). Bioluminescent A. granaria being reared in a ring box for display. A. granaria (see inset at http://www.geocities.com/bioluminus/jewel_box_inset.jpg) successfully adapt to being reared and maintained upon a sterile substrate and yeast when housed in a variety of different glassware or vials. Photo by Graham East
Students just learning how to maintain pure cultures may wish to experiment to discover which type of glassware or other vials are most useful in rearing stock. It has been reported that Collembola do well when reared in numerous small containers and that in large containers a reproductively inhibitory chemical may build up (Waldorf 1971). Also carbon dioxide may build up in large, sealed containers, due to fermentation of yeast and this may kill colonies. Additional considerations here are how to handle A. granaria so as to maintain pure stock free of such pest as mites or fruit flies.
Students and researchers alike may wish to investigate the mechanism of bioluminescence. Can simple experiments be devised to prove the nature of bioluminescence in A. granaria? This certainly represent a challenge, especially if presented to students giving them no clue as to the nature of bioluminescence. Let students dwell upon the mystery and see what they suggest in the way of experiments to seek an answer to this perplexing question.
Conclusion
A. granaria represents a great addition to the teacher's arsenal of species to use in the classroom and in the laboratory. For this reason alone schools and universities far and wide would be wise to obtain and maintain pure stock of A. granaria. If you are interested in adding A. granaria to your repository of teaching tools, please place your order today. Order forms are available on-line for New Zealand residents only at http://www.geocities.com/bioluminus/geo_order_form.html.
Given that A. granaria is easy to rear in vitro and is NOT an endangered or threatened species, and has no possibility of becoming an endangered or threaten species in the near future, owing to the fact that it has a wide global occurrence, this is an ideal species for universal distribution and usage by the academic community. It is anticipated that A. granaria will be an excellent species for easy export/import, given this is a harmless small animal that is NOT an etiologic agent, is NOT harmful to animals, agricultural crops or trees and is NOT a vector for any disease agents. Hence A. granaria does NOT represent a biohazard and may be freely distributed and used in the academic community without risk to people, animals or agriculture.
Because A. granaria meets all requirements for a species with respect to being safe and ideal for research, teaching, demonstration of bioluminescence and export/import, it is highly recommended that the global community adopt A. granaria for these purposes and establish pure rearing stock to enhance the educational experience, stimulate the learning process and generally serve as an asset to the scientific community. The beneficial nature of A. granaria, combined with the fact that it exhibits bioluminescence, promises to make this Collembola an ideal species for safe and easy export/import and general usage in academic, research and even commercial communities throughout the world.
Links
The following on-line sites related to A. granaria may be of interest to students and will be updated regularly:
A. granaria Homepage (Project GEO): http://www.geocities.com/bioluminus/geo.html
Introduction: http://www.geocities.com/bioluminus/introduction_geo.html
Frequently Asked Questions (FAQ) : http://www.geocities.com/bioluminus/faq_geo.html
Gallery: http://www.geocities.com/bioluminus/gallery_geo.html
Links: http://www.geocities.com/bioluminus/links_geo.html
Save the Planet: A. granaria has been adopted by the GEO Project as an international symbol to use to preserve and protect the environment and is featured at: http://www.geocities.com/bioluminus/save_planet.html
Emporium: http://www.geocities.com/bioluminus/emporium_geo.html
Experiments: http://www.geocities.com/bioluminus/experiments_geo.html
Acknowledgments: http://www.geocities.com/bioluminus/credits_geo.html
Order Form: http://www.geocities.com/bioluminus/geo_order_form.html
Contact: terrylynch@aol.com (Terry Lynch)
Hyper Links. The above hyperlinks are provides for on-line reprint of this publication:
Home | Introduction | FAQ | Gallery | Links | Save the Planet | Emporium | Experiments | Acknowledgments | Order Form | Contact
Additional contact information:
Graham East, Collector
soupbonesurfer@xtra.co.nz (soupbonesurfer)
Terrence A. Lynch
System Engineer, Naturalist and Photographer
PO Box 241035
Montgomery, AL 36124-1035
Email: terrylynch@aol.com
Homepages:
Blink and Links: http://members.aol.com/terrylynch/fireflies.htm
Firefly Mysteries: http://home.att.net/~y2kvault/firefly_mysteries.html
Project GEO: http://www.geocities.com/bioluminus/geo.html
Wanda M. Weiner Ph.D.
Head of the Laboratory of Invertebrates
Institute of Systematics and Evolution of Animals
Polish Academy of Sciences
Slawkowska 17, Pl- 31 016 Krakow, Poland
e-mail: weiner@isez.pan.krakow.pl
www.isez.pan.krakow.pl
Draney, Michael L.
Dept. Natural and Applied Sciences
University of Wisconsin-Green Bay
2420 Nicolet Drive
Green Bay, WI 54311
draneym@uwgb.edu
Bibliography
Draney, Michael L. 2000. Checklist of the Collembola: Culturing Sinella curviseta Brooks. Department of Natural and Applied Sciences, University of Wisconsin-Green Bay, Green Bay, WI 54311. Posted at http://www.collembola.org/publicat/culture.htm - Last updated on 2000.09.15 by Frans Janssens.
Gist, C. S., D. A. Crossley, Jr., and V. A. Merchant. 1974. An analysis of life tables for Sinella curviseta (Collembola). Environmental Entomol. 3(5): 840-844.
Lipovsky, L. J. 1951. Collembola as food for chiggers. J. Parasit. 37: 324-326.
Lipovsky, L. J. 1954. Studies of the food habits of postlarval stages of chiggers (Acarina, Trombiculidae). Univ. Kansas Sci. Bull. 36pt.II(14): 943-958.
Lynch, Terrence A. Firefly Notebooks: How to rear fireflies: Techniques with respect to the collection, observation and rearing of the firefly Photinus pyralis and Photuris sp. with notes upon the feeding behavior of Photinid and Photuris larvae and the implications these contrasting behaviors have on evolution of the species in fireflies. Presents photomicrographs, drawings, diagrams and methodology with respect to original research related to the rearing of fireflies. (http://home.att.net/~y2kvault/firefly_notebooks.html) See also Blink and Links at http://members.aol.com/terrylynch/fireflies.htm
Lynch, Terrence A. 2001a Identification of Anurida granaria for the GEO Project by Dr. Wanda Weiner. http://www.geocities.com/bioluminus/introduction.html.
Lynch, Terrence A. 2001b Methodology for the collection, rearing and handling of Anurida granaria (Nicolet, 1847) as applicable in the enhancement of the educational and research process. http://www.geocities.com/bioluminus/methodology.html.
Lynch, Terrence A. 2001c Firefly Safe Zones: Strategies for Reintroduction, Preservation and Maintenance of Vigorous Firefly Populations (http://home.att.net/~y2kvault/safezone.html)
Lynch, Terrance A. 2001d Fireflies vs. Humans: More Alien than Alien: Which is the more alien species homo sapiens or P. pyralis? (http://home.att.net/~y2kvault/more_alien.html).
Waldorf, E. S. 1971. Oviposition inhibition in Sinella curviseta (Collembola: Entomobryidae). Trans. Am. Micros. Soc. 90(3): 314-325.
Weiner, W.M. 1996. Generic Revision of Onychiurinae (Collembola : Onychiuridae) with a Cladistic Analysis., Ann. Soc. Entomol. Fr. (N.S.), 1996, 32 (2), p.163-200.
Copyright © 2001 by Terry Lynch. All Rights Reserved. The background picture of bioluminescent Anurida granaria (Nicolet, 1847)(http://www.geocities.com/bioluminus/geo.html) was made by Terry Lynch and is a photo composite and enhanced image created from a photograph provided by Graham East.