1 EVALUATION OF INORGANIC AND ORGANIC SUBSTRATES ON THE GROWTH OF Zamia pumila By VICKIE LYNN MURPHY A THESIS PRESENTED TO THE GRADUATE SCHOOL OF THE UNIVERSITY OF FLORIDA IN PARTIAL FULFILLMENT OF THE REQUIREMENTS FOR THE DEGREE OF MASTER OF SCIENCE UNIVERSITY OF FLORIDA 2012
2 2012 Vickie Lynn Murphy
3 To my mentor Dr. George Fitzpatrick
4 ACKNOWLEDGMENTS I would like to thank Montgomery Botanical Center for supporting my education and funding my research project. Thank you to Francisco Jimenez and Alberto Veloz, from Jardin Botanico Nacional Dr. Rafael Ma. Moscoso of the Dominican Rebublic and Michael C a l o nje for collecting and shipping the seed. I would especially like to thank Dr. Chad Husby for his in finite patience in helping to design this experiment and process ing the results. I would like to acknowledge Dr. Patrick Griffith for his constant guidance, mentoring, and serving on my committee. Special t hanks to Dr. Kimberly Moore for being my graduat e chai r. I greatly appreciate her generosity with her time and her dedication to her students. I would like to thank Dr. George Fitzpatrick and Dr. Wagner Vendrame for serving on my committee and inspiring me in their classes. Additionally, I would like to thank Joanne Korv i ck and Luc i Fisher for their assistance in furthering my education. It has be en my honor to be a student at the U niversity of Florida Fort Lauderdale Research and Education Center Last, but not least, I would like to thank the following people for helping with data collection and horticultural care: Albert Diaz, Xavier Gratacos Trish Hicks, Lynn Leveritt, Margaret Martin, Lane Park, Ce celia Thornton, and Bill Walker
5 TABLE OF CONTENTS page ACKNOWLEDGMENTS ................................ ................................ ................................ .. 4 LIST OF TABLES ................................ ................................ ................................ ............ 6 LIST OF FIGURES ................................ ................................ ................................ .......... 7 LIST OF ABBREVIATIONS ................................ ................................ ............................. 8 ABSTRACT ................................ ................................ ................................ ..................... 9 CHAPTER 1 INTRODUCTION ................................ ................................ ................................ .... 11 2 LITERATURE REVIEW ................................ ................................ .......................... 13 A Brief Review of Cycads ................................ ................................ ....................... 13 Conservation ................................ ................................ ................................ ........... 13 Growing Substrates ................................ ................................ ................................ 15 3 MATERIALS AND METHODS ................................ ................................ ................ 18 4 RESULTS AND DISCUSSION ................................ ................................ ............... 22 Zamia Growth ................................ ................................ ................................ ......... 22 Growing Substrates ................................ ................................ ................................ 23 5 CONCLUSIONS ................................ ................................ ................................ ..... 33 LI ST OF REFERENCES ................................ ................................ ............................... 34 BIOGRAPHICAL SKETCH ................................ ................................ ............................ 37
6 LIST OF TABLES Table page 4 1 Final fresh Zam ia pumila tissue growth measurements of plants grown in nine diffe rent media for eighteen months. ................................ .......................... 26 4 2 Final Zamia pumila dry tissue growth measurements of plants grown in nine different media fo r eighteen months.. ................................ ................................ 27 4 3 Initial Zamia pumila leaf tissue nutrient concentration prior to transplanting into nine different growing media.. ................................ ................................ ...... 28 4 4 Initial chemical properties of the nine growing media used to grow Zamia pumila for eighteen months. ................................ ................................ ............... 29 4 5 Initial physical properties of nine growing media used to grow Zamia p umila for eighteen months. ................................ ................................ ........................... 30 4 6 Particle size distribution for nine substrates used to grow Zamia pumila for eighteen months. Values are percentage s retained at each sieve size. ............ 30
7 LIST OF FIGURES Figure page 4 1 Final Zamia pumila dry tissue growth measurements of plants grown in nine different media for eighteen months ................................ ................................ .. 32
8 LIST OF ABBREVIATION S Al Chemical element symbol for Aluminum B Chemical element symbol for Boron Ca Chemical element symbol for Calcium CEC Cation Exchange Capacity measured as meg/100 grams. The sum of to tal exc hangeable cations that a soil can absorb expressed in cent i moles of charge per kilograms. Cu Chemical element symbol for C opper EC Electrical conductivity measured in mmhos / cm. Fe Chemical element symbol for Iron K Chemical element symbol for Potassiu m MBC Montgomery Botanical Center Mg Chemical element symbol for Magnesium Mn Chemical element symbol for Manganese N Chemical element symbol for Nitrogen Na Chemical element symbol for Sodium P Chemical element symbol for Phosphorus pH The negative logari thm of the Hydrogen concentration of a soil. ppm C oncentration as measured in parts per million P1 The P1 (weak Bray) test measures phosphorus which is readily available to the plants. S Chemical element symbol for Sulfur
9 Abstract of Thesis Presente d to the Graduate School of the University of Florida in Partial Fulfillment of the Requirements for the Degree of Master of Science EVALUATION OF INORGANIC AND ORGANIC SUBSTRATES ON THE GROWTH OF Zamia pumila By Vickie Lynn Murphy Dece mber 2012 Chair: Kimberly Moore Major: Horticulture Science Cycads comprise the most threatened group of plants on earth. Ex situ cultivation is an essential tool in rare cycad conservation. Appropriate container media properties especially excellent aeration, are crucial to successful cultivation of most cycads. Typical cycad media include substantial portions of organic materials that will decompose over time, reducing aeration At Montgomery Botanical Center, the use of novel inorganic media ha s improved survival and growth of several very rare and challenging Zamia species suggesting the need for a rigorous evaluation of different inorganic container media. E ffects of : 1) silica sand ; 2) Fafard ( a peat/perlite mix ); 3) perlite (expanded volcani c glass); 4) pumice (volcanic rock); 5) Turfac e (montmorrillinite clay); 6) Profile (calcined clay ); 7) a 50% sand: 50% Profile mix ; 8) Permatil (calcined slate); or 9) Axis (cal c in ed diatomaceous earth) on growth of Zamia pumila seedlings were evaluated. At transplanting 35 g of Nutricote fertilizer 18N 6P 2 0 5 8K 2 O slow release 360 (Fl orikan E.S.A. Sarasota, FL) was incorporated into the top one inch of all media Plants were fertilized at the same rate after one year of growth. Plants were watered to p oint of saturation three times a week. Media physical properties and media nutrient analysis were measured. Nutri ti onal analysis of seedlings was measured at
10 t ransplant ing Plants were harvested and g rowth parameters were measured 18 months after planting. The s and medium produced significantly higher total dry weight and leaf area in Z pumila than all other substrates. We suspect that lower water holding capacity and higher air filled space of sand, perlite and pumice contributed to better growth in th ese substrates. Although P ermatil had the lowest water holding capacity, it also had the highest air filled space and this might have been too dry for optimum Zamia growth. Although Fafard had a high water holding capacity, the percentage of larger particl es (#10 and #35) was similar to sand. Under conditions similar to this experiment, we would recommend growing Z pumila in silica sand.
11 CHAPTER 1 INTRODUCTION Cycads are the oldest living seed plants, originating approximately 300 million years ago. Acco rding to the 2010 International Union for Conservation of Nature (IUCN) ( I U CN, 2010) This is a 16% increase from 46% in 1978. Cycads are the most threatened group of plant species on Earth due to loss of habitat a nd over collection in the wild. The cycad specialist group of IUCN recommends concerted efforts to improve propagation and cultivation of cycads (Donaldson et al ., 2003). Although progress has been made in cycad horticulture, c yca d conservation depends on successful cultivation of ex situ collections (Chavez and Litz, 2007). All Zamia are considered New World plants. The majority of Zamia species are from Central America but species can be found in the Bahamas, Cuba, and Columbia Historically the range of Z. pumila was Central Cuba, Southern Puerto Rico and Hi spaniola and Dominican Republic. Field research in the native habit of Z pumila found plants growing in limestone, a hard rock formed by marine coralline depo sit, and sand near the coastal area (Zanoni, 1982). Commercially produced organic and inorganic substrates would not necessarily be readily available in these countries. The ability to use whatever inorganic substrates that are naturally occurring in each country could greatly benefit conservation efforts. To successfully grow cycads good drainage is crucial (Whitelock, 2002). Aeration, water retention, nutrient holding capacity, and decomposition rate are vital in cycad substrates. Drainage of organic subs trates can decline over time as decomposition occurs (Bilde r back, et al., 2005). Inorganic substrates could prove to be the ideal long
12 term cycad substrate. Several substrates may prove to be successful in sustaining wild collected Z pumila in container p roduction Recent work includes research on evaluating the effects of inorganic substrates versus organic substrates on germination and early seedling growth of 3 rare Zamia sp ecies : Z. fairchildiana, Z. cunaria, and Z. portorsensis ( Calonje et al ., 201 1 ) In this study, three substrates were e valuated over a period of 14 months. Turface, silica sand, and one mixed substrate (organic and inorganic components) were tested. A ll three substrates performed adequately for germination and growth of these 3 Zamia species. A number of inorganic media and media components, such as perlite, vermiculite, T urface, rockwool and baked clay balls have long been used in the US and Europe and are being increasingly used in Asia (Kang et al., 2004). A growing variety of co ntainer media are being used worldwide by nursery growers, including Turface and Profile for several species including Cycas, Dioons, Encephalartos, Macrozamia Microcycas, and Zamia However, little research has been published on from this work Hypoth esis and Objectives: Our hypothesis is that Z pumila growth will be greater in inorganic media compared to growth in an organic medium The objective was to compare Z pumila growth in eight inorganic media and o ne organic medium used as a control.
13 CHAPT ER 2 LITERATURE REVIEW A Brief Review of Cycads The first true cycads are represented in leaf fossils from the late Paleozoic period, during the Upper Permian Period, about 230 million years ago ( Stewart and Rothwell, 2001 ) About 180 million years ago, du ring the Mesozoic, the supercontinent of Pangea began to divide. Because cycads evolved before this separation, cycad fossils have been discovered on every modern continent, including Antarctica ( Hubbach, 1 994 ). A motile sperm cell is one characteristic fo und only in cycads and the ginkgo tree of China, among all modern seed bearing plants ( Vaughn and Regzaglia 2006). This is considered by some to be a primitive feature in nature, shared with ferns, mosses, and other spore bearing plants (Witte, 1977) Ada ptations that have enabled cycads to survive into modern times include insect pollination, highly toxic defensive compounds, and symbioses with nitrogen fixing bacteria (Hall, 2011). These unique characteristics help to make cycad conservation very importa nt to understanding plant evolution. Conservation Cycads present many conservation challenges and there are tremendous research opportunities for these living fossils. In a recent letter to the editor of the Cycad newsletter, Dr. Tang quoted Dr. Dennis Ste venson as having mentioned a shipment of 25,000 plants of a newly described species, Ceratozamia norstogii Stevenson, into the US (Dehgan, 1983). This resulted in complete decimation of 1 of only 2 known populations of this taxon. Tang also stated that du ring the same time period 20,000 Zamia plants were exported from the Dominican Republic (Dehgan, 1983)
14 In the 1980s over 80 tons of the Mexican cycad Zamia furfuracea were extracted from the wild for the horticultural industry (Donaldson et al., 2003). No w this cycad is widely cultivated and the horticultural demand is met entirely by nursery propagated plants (Calonje et al. 2011). At least one species of cycad in South Africa, Encephalartos woodii is known only to exist as a single clone. It is conside red to be extirpated from the wild ( Gorelick and Osborne, 1998) S everal individuals from the same source have been reproduced from vegetative growth and maintained in cultivation. Seed propagated cycads reintroduced into native habitats help to prevent th e genetic erosion and extinction of wild populations (Walters, 1999). Proper documentation of parent plants and detailed provenance is necessary to prevent genetic contamination. All cycads are dioecious having male and female rep roductive structures on se parate plants ( Walters and Osborne 2004) Natural pollination is by insects, usually beetles It is essential to grow cycads in containers in ex situ collections to the point of maturity in order to develop seed bearing population plant s of known gender. Male cones typically develop first and female cones are produced later Research by C a lonje et al. (2011) report several successful methods for hand pollinating and determining viability of cycad seeds. When attempting to germinate cycad seeds it is of cr itical importance that the embryos are fully developed. The same heat and humidity that encourages germination in ripe seeds may result in the death of seeds with immature embryos (Calonje et al 2011). It is widely believed that seed germination in most cycads can be improved by germinating under higher than ambient temperatures. Several experts recommend temperatures between 27 C and 43 C.
15 Silica sand substrates tend to maintain higher temperature s than other organic and inorganic substrates (Calonje et al., 2011). Tang (1990) reported that seed size in cycads can be an indicator of the care required by the grower for successful survival rates of seedlings. Zamia seeds are small in comparison to other species. There is a 90% or more mortality rate in t he wild. This makes an excellent case for ex situ collections to ensure conservation of Zamia plants. C y cad conservation also can be done using vegetative propagation. This method of propagation is especially important for rare plants and when seeds are no t available (Osborne and Walkley 1997 ) Inorganic substrates are recommended for vegetative propagation. Vegetative propagation provides a predictable quantity of male and female plants. This increases the ability for seed production. Several species can be cultivated this way Zamia species have branched subterranean stems that can be divided and propagated. Offsets called pups are removed from mature plants and rooted. It is essential to prevent rot while waiting for new root systems and consequent l eaf development to occur. Growing S ubstrates Soilless mixtures, such as a 1:1:1 ratio of sharp sand, perlite, and pine bark have been commonly used to grow Zamia plants Many growing media mixes have a more or less pronounced hydrophobic character in rel ation to their moisture level Although no significant change is revealed in terms of water retention with or without clay, we t tability measurements clearly showed large modifications in relatively dry conditions. We can conclude that clay can be character ized by its ability to improve the wet t ability and spee d of the growing media to rewet (Michel, 2009).
16 Organic mixtures containing peat, pine bark and /or sphagnum have the disadvantage of decomposing rapidly and resulting in a compressed substrate that in hibits air circulation to roots (Rischer, 2000) Rischer experimented with 100% inorganic substrates to grow carnivorous plant previously grown in organic media. H e discovered that plants previously potted in organic media and transplanted into in org anic media showed no signs of stress. Similarly, Turface, an inorganic substrate, was used in the successful propagation of Cycas micronesica from Guam, Zamia decumbens from Belize, and Caribbean Zamia species (Griffith et al., 2010). Inorganic substrates also provide a sterile growing environment in comparison to organic substrates. Inorganic substrates are recommended for germinating cycad seeds (Broom e 2012) because a sterile environment helps to prevent fungus from invading the vulnerable seeds and see dlings. Perlite and sand have been used for successful cycad germination. The nursery at MBC is currently using Profile, an inorganic media, to germinate Zamia seedlings. Inorganic substrates also aid in weed control in container production. Fewer weeds d evelop and are more easily removed without disturbance or loss of substrate. This is important because the fertilizer is sown in the top one inch of container. Traditional weeding with organic substrates often causes loss of media and fertilizer. An insect pest of cycads, the zamia borer Eubulus sp. (Order: Coleoptera ; family : Curculionidae) can devastate a Zamia population Zamia from the Greater Antilles and NE Florida are especially vu l nerable and the ex situ collection at MBC have been decimated by it ( O b erprieler, 1995). The beetle lays its eggs at the base of the plant and the larvae tunnel into the trunk just above the root zone. Their feeding causes the plant
17 to rot and separate from the roots. Inorganic substrates can play important role s in rehabil itating these damaged plants. They can be saved if returned to containers with inorganic substrates to recuperate as their r oot systems will regenerate if kept in a well drain ed substrate.
18 CHAPTER 3 MATERIALS AND METHODS Approximately 1600 Z pumila seed s were collected from several mature mother plants on February 08, 2010 at the Jardin Botanico Nacional Dr. Rafael Ma. Moscoso of the Dominican Republic and shipped to MBC. Seeds were sown in Fafard Super Fine Germinating Mix (Conrad Fafard, Inc. Aguwam, M A ) on July 20, 2010 in one quart size Ziploc baggies (S.C. Johnson & Son, Inc. Racine, WI ) and placed in a dark closet for germination at ambient temperatures ranging from 21 to 27 C Seeds were checked weekly for signs of germination. By September 1, 2010, 905 seeds had germinated. Seedlings were transplanted into Deepot D27L (Stuewe & Sons, Inc. Tangent, Oregon) seven inch tubes with a volume of 444 m L filled with Profile. Profile (Profile Products, LLC. Buffalo Grove, IL) is a fine porous ceramic s ubstrate. POLY FIL a polyester fiber (Fairfield Processing. Danbury, CT) was placed in the bottom of the tube to hold P rofile in the pot. D27L tubes were placed into Nova 25 trays (Stuewe & Sons, Inc.) for support. On October 1, 2010, 135 seedlings for the substrate experiment were randomly selected with 45 seedlings from three different mother plants and plants from each mother were labeled with a common accession number. Five individual seedlings from each accession were randomly assigned a treatment t o provide a total of 15 seedlings per treatment. The seedlings were transplanted into one gallon Poly Tainer PTC700 (Nursery Supplies, Chambersburg, PA) containers filled with( 1) Sand : coarse silica sand (Florida Silica Sand Company, Fort Lauderdale, F L ); ( 2) Perlite : heat expanded volcanic glass (W.R. Grace & Co., Cambridge, MA) ; (3) Pumice: light weig ht volcanic rock ( Hess Pumice Products, Malad City, Idaho); (4) Turface: calcined montmorillinite
19 clay ( PROFILE Products LLC, Buffalo Grove, IL ) ; ( 5) P rofile : calcined montmorillinite clay ( PROFILE Products LLC, Buffalo Grove, IL ); ( 6) 50% sand/ 50% P rofile (1:1) ; ( 7) PermaTill : calcined slate ( Carolina Stalite Company, Salisbury, NC ); ( 8) A xis: calcined dia to maceous earth ( EnviroTech Soil Solutions, O regon City OR ); or the organic control medium ( 9) Farfard Potting mix #2 : Sphagnum peat moss (65%), perlite, vermiculite, starter nutrients, wetting agent and dolomitic limestone ( Conrad Fafard Inc. Agawam, MA ). The pots were double potted to prevent leak age of the substrate, with one pot placed inside the other. At transplanting, 35 g of Nutricote fertilizer 18 N 6 P 2 0 5 8 K 2 O slow release 360 (Florikan E.S.A. Sarasota, FL) was incorporated into the top one inch of the substrate. Plants were fertilized at the same rate after one year of growth on October 1, 2011. Plants were placed in a 20 ft. by 60 ft. glass greenhouse. The experiment was arranged as a r andomized c omplete b lock d esign. Temperature was maintained between 15.5 and 32 C Plants were ha nd watered three times a week to the point of saturation using municipal water. Leaf and leaflet counts were recorded periodically during the experiment. All plants were harvested on May 1, 2012, 18 months after planting. Immediately before harvest, each plant was examined and given a quality rating from 0 to 5. Plants were evaluated visually and rated by growth and overall appearance. Plants alive with no leaves were rated 0. Poor quality plants with a few leaves were rated 1. Fair quality plants were rat ed 2. Acceptable landscape plants were rated 3 and 4 accordingly. Robust plants with the most leaves were rated 5.
20 At harvest, the leaves were separated from the rest of the plant and l eaflets were separated from the ir petiole s Leaf area was measured us ing a LI 3000 portable area meter console and head (LI COR, INC. Lincoln, Nebraska). All leaflets and petioles from each individual plant were then dried to determine dry weight. Each individual plant was removed from the pot and all substrate was rinsed off using gentle water pressure. The diameter of the stem was measured using an electronic digital caliper (Fred V. Fowler Co., Newton, MA ) The stem was separated from the root system and dried separately. A total of 405 samples ( 135 plants, 3 samples per plant) were placed in to a Grieve (The Grieve Corporation, Round Lake, IL) forced air d rying oven at 63 C for seven days until constant dry weight was achieved. Leaf, shoot, and root dry weight was recorded. A comparative analysis of overall growth was done to determine which inorganic substrates were best suited for cycad container production Particl e size distribution was measured using a Hubbard Scientific #548 six screen sieve set (Hubbard Scientific Inc. Chippawa Falls, WI). Particle size distribution percentages were determined by weight using a model 720 4A digital scale (Fisher Scientific Pitt sburgh, PA ). Airspace, water holding capacity, and t otal porosity was measure d on a volume basis The container used for testing was the Poly Tainer PTC700 ( Nursery Supplies, Chambersburg, PA) and its internal, 2450 m L volume of container was determined by placing a one gallon Z iploc freezer bag inside container to hold water. The c ontainer was then filled with substrate and weighed to determine dry weight. Water was added until all pore spaces were filled. Water was then drained off and measured to deter mine water holding capacity, air space and total porosity.
21 The following physical and chemical properties were determined by A & L Southern Agricultural Laboratories, LLC. (Deerfield Beach, F L ) : cation exchange capacity ( CEC meq/100g); electrical conduc tivity ( EC mmhos/cm); pH; nitrogen ( N ppm,) available phosphorus ( P1 ppm), potassium ( K ppm), magnesium ( Mg ppm), and calcium ( Ca ppm) Nutritional analysis of leaf tissue was measured at transplant ing Sixty leaflets from different plants were tested to g et a comprehensive analysis. N, sulphur ( S ) P, K, Mg, Ca, and sodium ( Na ) were reported as a percentage. The other nutrients iron ( Fe ) aluminum ( Al ) manganese ( Mn ), boron ( B ) copper (C u ) and zinc ( Zn ) were reported as ppm. All variables except for qua lity rating were analyzed by a one way analysis of variance (ANOVA) using JMP p.0.2. (SAS Institute, Inc., NC). Subsequently a post hoc multiple comparisons test, Tukey's honestly significant difference (HSD), was used to perform pairwise comparisons betw een treatments. The plant quality rating variable was analyzed via contingency table analysis testing for non homogeneity using a chi square goodness of fit test.
22 C HAPTER 4 RESULTS AND DISCUSSION Zamia pumila Growth Leaf area, leaf number, leaflet numb er and stem diameter were greatest in containers filled with sand and lowest in containers filled with Axis (Table 4 1). Plants grown in s and also achieved the greatest leaf area (1886.81 cm 2 ) while Axis produced the lowest leaf area ( 13.88 cm 2 ) On averag e, plants grown in sand produced 25 times as many leaves as Axis and had the highest leaflet count at 111.00 leaflets. Plants grown in sand achieved the largest stem diameter (46.54 mm) while those grown in Permatil produced the lowest stem diameter (16. 4 mm). All plants grown in sand received the highest q uality rating of 5 .00 (Table 4 1). Plants were evaluated visually and rated by growth and overall appearance. Plants alive with no leaves were rated 0. Poor quality plants with a few leaves were rated 1 Fair quality plants were rated 2. Acceptable landscape plants were rated 3 and 4 accordingly. Robust plants wit h the most leaves were rated 5. There was significant non homogeneity among substrates for quality rating (p<0.0001). Research measuring the e ffects of shade and controlled release fertilizer showed a direct correlation between stem size and number of leaves : t he larger the stem size, the greater number of leaves despite treatment s of shade and fertilizer (Dehgan et al., 2004). Shoot dry weight leaf dry weight, root dry weight, and total plant dry weight also were greatest for plants grown in sand (Table 4 2). Plants grown in Fafard, perlite and pumice ha d greater shoot, leaf, root and total dry weight compared to plants grown in Turface, Prof ile, Sand/Profile, Permatil or Axis (Figure 4 1). Plants grown in Axis had
23 the least shoot, leaf, root and total dry weight (Table 4 2). It is interesting to note that overall growth was consistent between the nine media (Figure 4 1). However, sand, perl ite, and pumice produced the largest ratio of stem dry weight to leaf dry weight while Fafard produced a greater leaf to stem ratio. Previous research indicated that modification of irrigation frequency and controlled release fertilizer application result ed in exceptionally rapid cycad growth and elimination of deficiencies (Dehgan, 1999) Additionally, soil moisture should be cont rolled by irrigation frequency rather than modifying water holding capacity (Dehgan, 1 999) During 18 months in production, we did not observe any nutritional deficiencies There are no published guidelines for nutritional analysis of cycads including Z pumila but t here are established guidelines for wo o dy ornamentals (Yeager, 2010) and using these standards, the leaf tissue a nalysis for Z. pumila did not reflect any nutritional deficiency (Table 4 3) Cycads have the unique ability to fix nitrogen from the atmosphere with collaroid roots (Thoumire, 2000). This is another advantage of cycads reducing the need for a nutrient ric h substrate. Growing S ubstrates Although sand had the lowest initial pH (4.4) and a low initial EC, no apparent nutrient deficiencies were observed (Table 4 4). We do not believe that the chemical properties of sand contributed to improved plant growth in this substrate. All plants were fertilized, irrigated and treated the same. Furthermore, perlite and pumice produced acceptable plant growth and the pH of pumice was the highest while perlite had the lowest EC (Table 4 4).
24 Seven out of eleven common symptoms of poor plant health in indoor plants are caused by overwatering and poor drainage (Taylor et al., 2010). It is well established that Zamia plant s prefer well drained media (Culbert, 2010). We suspect that lower water holding capacity and higher a ir filled space of sand, perlite and pumice contributed to better growth in these substrates (Table 4 5). The coarse particles of sand have little surface area per unit of volume compared to the smaller particles of clay. However, sand provides excellent gas exchange and physical support for the plant (Nelson, 2003). Although P ermatil had the lowest water holding capacity it also had the highest air filled space and this might have been too dry for optimum Z pumila growth Fafard did not have the same a ir filled porosity as sand, but the relative amount of air space to total pore space was high. T he percentage of larger particles (#10 and #35) in Fafard was similar to sand, 89.34 and 99.1, respectively. Permatil was the only substrate with particle size significantly larger than 4 mm (98.32%). Perlite (72.63%), pumice (85.04%), and Turface (67.42%) had the majority of particles greater than 2 mm. Sand (97.52 %), Fafard (68.0%), Profile (89.99%), and sand/ Profile (94.12%) had the majority of particles gr eater than 0.5 mm. The majority of Axis particles were divided between greater than 2 mm (49.62%) and greater than 0.5 mm (47.76%). Particle size distribution for sand and Permatil will not change over time. However, the particle size distribution of the o ther substrates could change due to leaching and weathering. Particle size did not appear to be a limiting factor for the top five substrates.
25 The poorest performing substrates had similar particle size which may explain the ir poor perform ance. Sand, per lite and pumice provided a more uniform distribution of moisture in the soil. Turface tends to dry out at the top of the pot and remain quite wet at the bottom. Therefore, it is misleading in terms of water requirements. Turface must be monitored closely t o avoid water stress (Sard, 1989). Water release curves for i norganic amendments added to the root zone were studied for calcined clay, calcined diatomaceous earth, calcined volcanic ash, and zeolite (Curtis, 2 008). Th os e authors found that the inorganic a mendments did not release appreciable amounts of internally held water until exposed to matric potentials more negative than usual for turf grass management. The internal porosity of inorganic amendments is very different than the media they are used to am end. We believe that the smaller particle size of the poorest performing media did not allow for proper drainage. For example, the 50% sand / 50% Profile mix was in the bottom three substrates. We believe that the small particles of Profile filled all th e air spaces and prevented water flow and aeration. Although plants grown in Fafard grew well, the rapid breakdown of standard organic substrates can lead to poor drainage, which can contribute to problems with root health and foliar deterioration (Taylor et al., 2010). Because cycads have a long growing cycle, organic substrates may not be a good choice. Typical container substrates are 75% or more organic matter and breakdown of the substrate typically leads to a deficient level of O2 and an excess of CO2 in the root zone especially when plants are saturated with water (Whitcomb, 2003).
26 Table 4 1. Final fresh Zamia pumila tissue growth measurements of plants grown in nine different media for eighteen months. Growing M edium z Mean l eaf area (cm 2 ) Mean l eaf number Mean l eaflet number Mean c audex diameter (mm) Mean q uality rating y Sand 1886.81 a 10.73 a 111.00 a 46.54 a 5.00 Fafard x 1543.67 b 9.60 ab 97.20 a 38.45 b 4.67 Perlite 1069.39 c 8.13 b 64.53 b 39.31 a b 4.60 Pumice 729.77 d 6.00 c 44.53 b 37.23 b 3.73 Turface 202.25 e 3.13 d 18.20 c 27.32 c 2.27 Profile 163.34 e 3.13 d 19.20 c 26.60 cd 2.20 Sand/Pro 130.87 e 2.86 d 15.93 c 24.85 cd 2.13 Permatil 85.17 e 1.66 de 8.93 c 16.4 e 1.33 Axis 13.88 e 0.67 de 3.00 c 19.35 de 0.67 z Within columns, d ifferent letters beside the values denote statistically significant ( <0.05) y Plant quality was rated on a scale of one to five based on size, number and a ppearance of leaves There was significant non homogenei ty among substrates for quality rating ( p<0.0001). x Control media
27 Table 4 2. Final Zamia pumila dry tissue growth measurements of plants grown in nine different media for eighteen months. Growing M edium z Mean l eaf dry weight (g) Mean s hoot dry weight (g) Mean r oot dry weight (g) Mean t otal dry weight (g) Sand 21.95 a 22.47 a 10.00 a 54.41 a Fafard y 19.88 a 12.21 b 6.77 b 38.86 b P erlite 12.04 b 12.47 b 6.95 b 31.47 bc Pumice 7.82 b 9.19 bc 6.74 b 23.75 c Turface 2.07 c 4.62 cd 2.25 c 8.93 d Profile 1.57 c 4.37 cd 2.10 c 8.046 d Sand/Pro 1.33 c 4.14 d 1.94 c 7.40 d Permatil 0 .79 c 1.84 d 1.17 c 3.80 d Axis 0.12 c 1.38 d 0.61 c 2.11 d z Within columns, d ifferent letters beside the values denote statis tically significant ( <0.05) y Control media
28 Table 4 3. Initial Zamia pumila leaf tissue nutrient concentration prior to transplanting into nine different growing media. Nutrient z Mean Concentration (ppm) Nitrogen (N) 28000 Sulphur (S) 2300 Phosphorus ( P ) 700 Potassium ( K ) 27100 Magnesium ( Mg ) 3800 Calcium ( Ca ) 7100 Sodium ( Na ) 900 Iron ( Fe ) 2 111 Aluminum ( Al ) 157 Manganese ( Mn ) 369 Boron ( B ) 47 Copper ( Cu ) 45 Zinc ( Zn ) 73 z Analysis was conducte d by A&L Southern Laboratories in Deerfield Beach FL.
29 Table 4 4. Initial chemical properties of the nine growing media used to grow Zamia pumila for eighteen months. Growing m edium z N (ppm) P1 (ppm) K (ppm) Mg (ppm) Ca (ppm) CEC (meq/ 100 g) pH EC (mmhos/ cm) Sand 56 1 7 1 34 1.8 4.4 0.21 Fafard y 94 4 132 159 346 4.6 5.5 0.39 Perlite 4 1 11 1 1 0.1 6.4 0.03 Pumice 8 1 35 1 207 1.1 7.3 0.12 Turface 21 1 316 24 104 1.7 6.1 0.14 Profile 78 1 367 40 120 2.7 5.4 0.28 Sand/Pro 62 1 254 60 273 3.5 5.4 0.44 Permatil 7 1 21 1 54 0.5 5.8 0.16 Axis 0 1 79 14 102 0.8 6.9 0.04 z Samples were analyzed by A&L Southern Laboratories in Deerfield Beach, FL. These values are from single determinations. y Control media
30 Table 4 5. Initial physical p roperties of nine growing media used to grow Zamia pumila for eighteen months. z Within columns, d ifferent letters beside the values denote statistically significant ( <0.05) y Control substrate Growing Medium z Mean Water Holding Capacity (%) Mean Airspace (%) Mean Total Porosity (%) Sand 15.50 ef 18.50 bc 34.00 g Fafard y 43.46 b 28.22 b 71.69 a Perlite 16.73 e 41.49 a 58.22 bcd Pumice 19.86 de 28.5 b 48.41 ef Turface 26.59 d 26.87 b 53.46 cde Profile 54.42 a 6.12 d 60.54 bc Sand/Pro 35.23 c 9.18 cd 44.41 f Permatil 9.59 f 42.78 a 52.37 de Axis 38.23 bc 23.39 b 61.69 b
31 Table 4 6 Particle siz e distribution for nine substrates used to grow Zamia pumila for eighteen months. Values are percentages retained at each sieve size. Sieve opening diameter (mm) Growing Medium z 4 2 0.5 0.25 0.125 Sand <0.01 d 1.58 e 97.52 a 0.50 d 0.10 b Fafard y 2.46 c 21.34 d 68.0 b 5. 73 b 2.46 a Perlite 6.42 b 72.63 b 20.95 e <0.01 d <0.01 b Pumice 4.53 b c 85.04 a 8.55 f 1.07 cd <0.01 b Turface <0.01 d 67.42 b 31.91 d <0.01 d <0.01 b Profile <0.01 d <0.01 e 89.99 a 10.01 a <0.01 b Sand/Pro <0.01 d 2.04 e 94.12 a 3.83 bc 0.10 b Per matil 98.32 a 1.69 e 0.22 f <0.01 d <0.01 b Axis <0.01 d 49.61 c 47.76 c 1.13 cd <0.01 b z Within columns, d ifferent letters beside the values denote statistically significant ( <0.05) y Control media
32 Figure 4 1 Final Zamia pumila d ry tissue growth measurements of plants grown in nine different media for eighteen months 0 5 10 15 20 25 Mean dry weight (g) Leaf Stem Root
33 CHAPTER 5 CONCLUSIONS Because cycads are slow growing and extremely long lived perennials, off site collections require a long term commitment from botanical gardens. Based on the results from our experiments, sand appeared to be the most opti mum growing medi um for Zamia pumila. Choice of medium greatly influences growth of Zamia pumila as well as other cycads Sand provides a viable alternative to standard organic mixes for growth of Zamia species Future research in container media could gr eatly improve the efficiency and effectiveness of conservation horticulture of rare cycads Although we did not measure this, we suspect that one reason that Zamia might grow well in sand is because Zamia have colloroid roots containing cyanobacteria th at can fix nitrogen (Thoumire, 2000) These roots are known to grow and appear on the surface of the media. This would be something to investigate in future studies. In the end, w e will conserve only what we love; w e will only love what we u nderstand ; and we will understand only what we are taught Baba Dioum, Propagation and Culture of Cycads
34 LIST OF REFERENCES Bilde r back, T.E., S. Warren, J. Owen, and J. Albano. 2005. Healthy s ubstrates need p hysicals too. HortTechnology 15(4) : 747 751. Broom e T 2012. Optimizing Cycad Seed Germination. 01 October 2012 < www. plantapalm.com/vce / horticulture/seeds.html >. Calonje M., J. Kay, M.P. Griffith 2011. Propagation of cycad c ollections from seed: appli ed reproductive biology for conservation. Sibbaldia 9:77 96 Chavez V.M. and R.E. Litz. 2007 R ecovery and difficult to regenerate species: the cycad example. Acta Hort. 738:51 61 Culbert, D.F. 2010. Florida coonties and Atala butterflies. Fact Sheet ENH 117. Institute of Food and Agricultural Sciences. University of Florida Gainesville, FL. Curtis, M.J., 2008. An alternate method for measuring plant available water in inorganic amendments. Crop Sci 48:2447 2 452. Dehgan, B 1983 Propagation and gro wth of cycads: A conservation strategy. Proc Florida State Hort. Soc 96 : 137 139 Dehgan, B 19 99 Propagation and culture of cycads: a practical approach. Acta Hort. 486:123 131. Dehgan, B F.C. Almira, A.E. Duddeck, and B. Shultzman, 2004. Effects of v arying shade and fertilizer on the growth of Zamia floridana The Bot Rev 70(1):79 85. Donaldson J.S., B. Dehgan, A.P. Vovides and W.Tang. 2003 .Cycads in the trade and sustainable use of cycad populations,. p. 39 47. In: Donaldson, J.S.(ed.) Cycads: st atus survey and action plan. IUCN,Gland, Switzerland. Griffith, M.P., C.E. Husby, and M. C a lonje, 2010. Cycad collections in the modern context: challenges, opportunities, investments, and outcomes. Proc of the 4th Global Bot Congress 1 10. Gorelick, R. and R Osborne. 2002. I nducing sex change and organogenesis from tissue culture in endangered African cycad Encephalartos woodii South African J Sci 98 :114 117. Hall, J. 2011. The Ecology of Cycads. Living Representatives of an Ancient Plant Linea ge and their interactions with animals. PhD Thesis, School of Biological Sciences, The University of Queensland Brisbane, Australia. Hill, K.D. 2000.The Genus (Cycadaceae) in China. Telopea 12(1) : 71 118.
35 Hubb u ch,C. 1994. A brief review of cycads. Fairc hild Tropi cal Gardens. Unpublished manuscript. Montgomery Botanical Center archives, Coral Gables, FL IUCN Red List of Threatened Species. 2010. International Union for Conservation of N ature 01 October 2012.< http://cmsdata.iucn.org/downloads/cycad/factsheet/final.pdf > Kang, J.Y., H. H. Lee & K. H. Kim. 2004. Physical and chemical properties of inorganic S ubstrates used in Korea. Acta Hort 644:237 242. Michel, J.C., 2009. Influence of clay addition on physical properties and wettability of peat growing media. HortScience 44(6):1694 1 697. Nelson, P. 2003. Root Substrate. P. 198. Greenhouse operations and management 6th ed. Printice Hall. Upper Saddle River, N.J. Oberpri eler, R.1995 The weevils (Coleoptera: Curculionoidea) associated with cycads 1. Classification, relationships, and biology. Pp. 295 334 in P. Vorster (Ed.) Proc Third International Conf Cycad Bio Cycad Society of South Africa, Stellenbosch South Africa Osborne, R. and S. Walkley,1997. Cycad sucker propagation. Encephalartos 51: 9 10. Rischer, H. 2000. Growing Nepenthes in a Completely Inorganic Substrate. ICPS Newsletter. CPN 29(2):50 53. Sard, E.L. 1989. An experiment with Turface. J Bromeliad So c 39(6) : 274 275. Stewart, W.N. and W. Rothwell, 2001. Cycads: origin and relationships P p. 338 349. Paleobotany and the evolution of plants. 2th ed. Cambridge University Press. New York, NY. Tang, W 19 9 5 A guide to pollinating and collecting seed s of cycads. Handbook of Cycad Cultivation and Landscaping. Miam i, FL. Published by the author. Taylor N. J., S. Nameth, and J. Chatfield. 2010 Diagnosing Problems on Indoor Plants. HYG 3068 96. 01 October 2012.< http://ohio.osu.edu/hyg fact/3000/3068.html >. Thoumire, E .2000.Collaroid Roots of Cycads. www.plantapalm.com 01 October 2012. < www plantapalm.com /vce/biology/corraloid.htm >. Vaughn K.C. and K. S. Renzaglia .2006. Structural and immunocytochemical characterization of the Ginkgo biloba L.sperm motility app aratus. Protoplasma 227:165 173 Walters, T.W. 1999 Off si te Cycad conservation: What is off site preservation? The Cycad Newsletter 22(1) : 110 1 12.
36 Walters, T.W. and R. Osborne.(ed) 2004. Cycad classification : concepts and recommendations. CABI Publicat ions. Cambridge, Mass. Whitcomb, C.E. 2003. Plant Production in Containers II. Lacebark, Inc., Stillwater, OK. Whitelock, L.M., 2002 The Cycads. Timber Press. Portland, OR. Witte, W.T., 1977. Storage and Germination of Zamia Seed. Pro c. Florida State Hort Soc 90:89 91. Yeager, T.H., 2010.Use of Tissue Analysis in Woody Ornamental Nurseries. Fact Sheet EDIS OCH16. Institute of Food and Agricultural Sciences, University of Florida Gainesville, FL. Zanoni, T.A., 1982. Guayiga ( Zamia) in Hi spaniola The Cycad Newsletter 111:5 13.
37 BIOGRAPHICAL SKETCH Vickie Lynn Murphy is currently employed as Nursery Curator at the Montgomery Botanical Center in Coral Gables, Florida. She has worked in several capacities at MBC since 1998 Vickie earned her Associ ate of Science in a griculture at Miami Dade Community College, Miami Florida, in 1997. She later completed her Bachelor of Science in Landscape and Nursery Management at the College of Agricultural and Life Science, Univers ity of Florida in Dec ember 2009 and then graduated with her MS in Horticulture from the University of Florida in December 2012.