Sampling Soil and Roots in Citrus Groves: Principles and Methods
Larry Duncan UF, IFAS, CREC Lake Alfred Observation Measurement Why sample? How many? When and where? Then what? Observation Never miss opportunities to examine the root systems of trees. Conventional APS, microjet APS, drip
Schumann APS trial Auburndale Severely affected by HLB by year 4. Exacerbated by other pests?
Measurement
What is there? Are there enough to be damaging? How can we know what is there with reasonable certainty at an affordable cost? (sample optimization) . From the APS survey, how confident can we be that pests caused greater damage to roots in the APS plots than in the conventional plots? Confidence interval – if we had sampled this block the same way 100 times, 95% of the 100 mean estimates would be in the range shown by the error bars. P=0.003 If there is actually no difference between these treatments in how much pests damaged the roots, we could expect to find this difference in our sample results just 3 times in one thousand trials. Confidence interval = x ± 2 s / √n
P=0.003 Length of error bars depends on the natural variation that occurs among samples and the number of samples taken.
This is an example of what we P=0.62 might find if we had sampled just 6 instead of 16 trees in each treatment. Optimizing sample size
P=0.003 Standard error = d x = s / √n where d=some proportion of x
√n = s / dx
n = (s / dx)2
So if we know the likely mean and P=0.62 standard deviation we can estimate an appropriate sample number. Optimal sample numbers have been estimated for many insect and nematode pests and most methods are based on some variation of this formula:
b-2 2 Number samples = zἀ/2ax /(0.5 CI/x) Which is based on a remarkable relationship described as Taylor’s Power Law. All organisms display this property. The sample variability depends on the population density.
To estimate a sample size that will deliver a certain accuracy we need only to know the expected mean and variance.
Taylor’s Power Law allows us to select a mean density (e.g., economic threshold) and predict the variance and an optimal sample size. First principle of sampling. Sample size decreases as population size increases. We can increase sample accuracy by sampling when and where population density is highest (e.g. mid-way between trunk and dripline during spring or late autumn for most nematodes. Stratify site base on properties affecting organisms). Second principle of sampling. Organisms are aggregated, clumped. Use a systematic pattern, don’t sample randomly. The most effective sampling pattern is one in which each sample is equally distant from the others. This is what a clump of potato cyst nematodes looks like. The dimensions of the clumps are related to the population density. By knowing the dimensions of the smallest detectable clump sampling plans were Development and evaluation of sampling methods for fields with infestation foci of potato cyst nematodes. Been and Schomaker (2000) Phytopathology. developed that detected The demarcation of cyst nematode infestations in field for seed potatoes using a monte carlo approach. Schomaker and Been (2010) OEPP/EPPO Bulletin. this nematode with 95% reliability. Polk County Indian River County
Osceola County S. diaprepesi >>>DRW 53% sand 16% mortality 3% EPN H. indica
H. indica A zigzag pattern is an effective method of spacing soil samples at regular intervals.
The figure is based on the spatial pattern of soybean cyst nematode in a Missouri soybean field. APS Press, 1999 Two samples of 15 cores each (12 inch depth, 2-5 acres) adequate to estimate root and nematode density. Stratify area based on physical properties likely to affect target. Systematic, not random sampling pattern. Under canopy, midway between trunk and dripline. Best to sample in spring and late autumn. Break 1
20 The HLB Era: “manageable problems have become intolerable”
Larry Duncan CREC D. abbreviatus Citrus root weevil refers to a complex of species. Diaprepes abbreviatus is the most damaging to citrus, but blue green weevils (Pachnaeus litus) are abundant and damaging in some parts of Florida. Little leaf notcher P. litus (Artipus floridanus) is not thought to be economically important. A newly introduced species, Sri Lanka weevil (Myllocerus undatus) looks similar to little leaf notcher and its economic importance is unknown.
A. floridanus M. undatus Diaprepes abbreviatus life cycle
Eggs
Adults
Neonates
Pupae Larvae + Phytophthora The abundance of weevil populations and their damage to trees varies regionally
0.6 0.25 Weevils are occasional
Flatwoods -1 Ridge 0.20 and sporadic pests on week 0.5 -1 0.15 the central ridge
-1 0.10 0.4 0.05 Weevils are major and Weevils trap Weevils week 0.00 chronic pests in the
-1 Lake DeSotoHendryn River Polk 1Polk 2 0.3 India flatwoods
0.2 Consequently, another consideration is that Weevils trap Weevils 0.1 weevil management should be considered 0.0 to be habitat specific MJJASONDJFMAMJJASONDJFMAMJJASON 2001 2002 2003
Futch et al., Fla. St. Hort. Proc. 2004 Typical seasonal pattern of adult weevil abundance in citrus orchards
In this Polk County grove, weevils emerged from soil in April-May each year (with a 2nd Fall emergence in one year). The timing varies from grove to grove.
So monitoring weevil abundance is another important managemet tactic. Adult weevil management should be coordinated to follow peak adult emergence by treating with adulticides and ovicides.
Duncan et al., 2001, Environmental Entomology Citrus root weevils
Why so hard to manage? Basically… Modern pesticides have little residual activity
Different weevil stages live in both the tree canopy and the soil. They serve as source of new individuals to replace those killed by management practices
Damage to roots is cumulative. Yearly damage by just a few larvae eventually kills trees. The fact that weevils comprise part of a pest-disease complex is the basis for an important management tactic…
While there are currently no rootstocks that are resistant or tolerant to weevils, they do exist for Phytophthora control. Diaprepes- Phytophthora complex evident on the sour orange trees, but not the Swingle citrumelo resets, in this DeSoto County grove.
Symptoms of the pest-disease complex on roots of sour orange Rootstocks for central ridge (Phytophthora nicotianae) Swingle citrumello Carrizo citrange C-35 C-32 Rootstocks for flatwoods (Phytophthora palmivora) Cleopatra mandarin US-802 US-897 Relationship Between Tree Decline and Phytophthora Incidence
Chemical Treatment, r2 = 0.78 Cleo 60 No Treatment, r2 = 0.65 50 40 C-22 C-32 30 Cleo 20 Swingle C-35 C-22 10 C-32 C-35
Percent Tree declinePercent Tree 0 Swingle 10 15 20 25 30 35 40 45 50 Percent Phytophthora Incidence Plot I, Treated, 2005
C22 C35 Swingle C32 Cleo Plot 2, Untreated, 2005
C32 C22 Swingle Cleo C35 Weevil management Rootstocks Regional considerations Cultural practices Soil drainage pH Physical barriers Insecticides (monitoring weevil abundance ) EPNs Management of soilborne insects
Diaprepes Task Force website has biliography of all research papers, general information and dichotomous management key for 8 common grove situations Management of soilborne insects
Each of 8 common scenarios provides links to resources from which recommendations were derived Management of soilborne insects
Each of 8 common scenarios provides links to resources from which recommendations were derived Cultural practices Soil drainage is critically important because weevil abundance and therefore root damage is greatest in wet soil. Trees stressed by wet soil are less tolerant of weevil damage. Some natural enemies of weevils are less abundant in wet soil.
S. diaprepesi H. indica EPNs EPNs
Low area of Diaprepes- infested grove
Campos-Herrera et al., 2013 Soil Biology and Biochemistry Cultural practices Low pH High pH
Soil pH is critically important because many rootstocks are less tolerant of stress in 1.0 )
high pH soil (>6.5) and 3 0.8 because some important 0.6 natural enemies of 0.4 weevils are intolerant 0.2
of high pH soil. (mg/cm density root Fibrous 0.0
4.5 5.0 5.5 6.0 6.5 7.0 7.5 Soil pH
Graham et al., 2013. Plant Disease Low300 pH High pH Cultural practices Raised natural pH 250 Soil pH is critically pH 5.5 important because some 200 pH 7.0 entomopathogenic 150 nematodes are intolerant of 100
high pH soil, whereas some 50 plant parasitic nematodes 0 are favored by high pH. 180 Reduced natural pH 1200 160 140 1000 soil 120 -3 800 100 Recovered IJ Steinernema diaprepesi 80 600 60 400 40
200 20 Citrus nemas x 100 cm Citrus x nemas 0 0 Ck Ad Ca Control Biocontrol Manure Low pH
Treatment Cultural Practices Landscape fabric can be installed as a barrier to prevent larvae from entering soil and adults from exiting soil McKenzie et al. 2001 Duncan et al 2008
The integrity of these Neonate barriers was maintained for 5- larva 6 years. Herbicide savings equaled the fabric cost.
Typical effect of landscape fabric mulch on tree growth
3500 Bare soil Landscape fabric In an experiment measuring 3000 Culture P=0.001 effects of Advanced Citrus Mulch P=0.001 CxM P=0.79 Production System (ACPS) versus conventional citriculture (CC), we also installed landscape 2500 fabric under some trees. ACPS trees were 25% larger than CC trees, but only 9% 2000 larger than mulched CC trees. Trunk cross-sectional area (mm) area cross-sectional Trunk
0 Mulched ACPS trees were 35% Conventional ACPS larger than conventional trees. Particle film of kaolin clay
These trees are the same age!
Protective tree covers that prevent psyllid feeding will also prevent weevil egg laying. These covers also increase tree growth tremendously. The effect is not simply due to prevention of HLB…. How does it work? Lab studies @ Optimum T and % RH Photosynthesis rate does not increase beyond 1/3 full sunlight – light is not limiting
Syvertsen & Albrigo 1980 + 9o June Citrus leaves in sun can be as much as 9-10oC higher temperature than ambient. Most o fruit trees regulate Mar. + 5 their temperatures to not exceed ambient. But citrus evolved as an understory plant in the subtropics so full sun was not a problem.
Syvertsen & Albrigo, 1980. Photosynthesis is maximum At 330C. Hence the beneficial effect of particle films and covers is due to increased 0.2 production of carbohydrate.
Particle films and covers also interfere with parasitoids and result in major disruption of 0.1 natural control of scales Photosynthesis
which become severe conductance Stomatal secondary pests.
Rainfastness of particle films needs improvement.
(Syvertsen and Lloyd, 1994) Chemical management of aboveground weevil stages
Monitoring peak abundance is important for timing pesticide application Adulticide combined with ovicide is most effective Egg laying begins 7-10 days post emergence of adult from soil Choice of adulticide should be integrated with rotation schedule for psyllid resistence management Chemical management of aboveground weevil stages
90 Chemical Treatment NoTreatment d 80 25 Yield 2007 70 c c 60 32 31 50 b b b b 40 21 24 b 21
(Mean + + (Mean SE) 34 30 24 Yield Per Tree (Kg) Tree Per Yield a a 20 19 20 10 0 C22 C32 C35 Cleo Swingle
Remember, rootstock is also critical and choice is based on Phytophthora species present at site. Search for enemies in center of origin of pest – Classical biological control Quadrastichus hatiensis
Aprostocetus vaquitarum
Parasitoids of D. abbreviatus introduced into Florida from Caribbean countries.
Some establishment has occurred, particularly in south Florida, but effective control has not.
Currently searching in highland areas to Ceratogramma etiennei find wasps that are more cold tolerant. Egg parasitoids, Ceratogramma etiennei, Quadrastichus haitiensi, from Puerto Rico/Guadeloupe (putative center of origin).
No Larval, pupal or adult parasitoids known.
Past failures attributed to widespread use of carbaryl, lack of correlation of life cycles of pest and prey and winter temperatures.
Strains of Bacillus thurengiensis recently discovered / GMO possiblities for exotoxin.
Ants prey on larvae (particularly fire ants; see handout showing >98% mortality from ants, nematodes and other natural enemies). Mortality Rate for Diaprepes Larvae from Neonate Drop to Adult Emergence
Mean No. Mean No. Mortality Neonates/m2 Adults/m2 Rate Year 2000 376.25 6.05 98.39% Year 2001 945.51 6.73 99.29%
Neonates Cone Trap Funnel Trap Adults Predation on Diaprepes Neonates: Field Assay
- Assay dish (4 mm x 48 mm dia) - 20 neonates per dish - 20 minute observation periods - 3 groves - 199 replicates, 3980 neonates Predators of Neonates on the Soil Surface % Predation Red Imported Fire Ant 10 20 30 40 Ants: Solenopsis invicta Buren Solenopsis invicta 140 (Hymenoptera; Formicidae) Pheidole moerens 132
Dorymyrmex reginicula 46
Brachymyrmex obscurior 42
Dorymyrmex bureni 41 Cardiocondyla emeryi 38 Paratrechina bourbonica 23 Pheidole morrisi 12
Hemipteran: 1 Geocoris floridanus N = 475 Break 2
56 IT TAKES A VILLAGE: CONSERVATION OF ENTOMOPATHOGENIC NEMATODES
Larry Duncan. University of Florida, IFAS Citrus Research and Education Center Lake Alfred FL, USA. Conservation biological control
Critical in canopy Unknown in rhizosphere
What natural enemies? Where are they? How effective are they? Studying Subterranean Good old days… Food Webs Today… Sucrose centrifugation to Sucrose centrifugation to recover nematodes. recover nematodes, fungi and Dilution plating to recover microarthropods. fungi and bacteria. Either qPCR or next Berlese funnels to recover generation sequencing. microarthropods. Lower detection limit, greater quantitative Quantitation was either accuracy. inexact or tedious. Can detect fungi and Could not discriminate bacteria associated with nematophagy from nematodes as a better saprophagy. measure of nematophagy. Culture media biased recovery of microorganisms. Diaprepes abbreviatus Pachnaeus litus Life cycle of steinernematid and heterorhabditid EPNs Baiting EPNs with caged, buried Earliest trials used this sentinel D. abbreviatus larvae technique to assess efficacy of Steinernema riobrave formulated and sold as BioVector®.
However, depending on the habitat, endemic, known and undescribed EPNs were frequently more abundant than the commercially formulated and augmented S. riobrave Weevils are more abundant and damaging to citrus in the flatwoods than on the central ridge.
EPNs kill weevils at a greater rate Citrus is grown in on the central ridge than in the two ecoregions flatwoods. Central ridge Do EPNs regulate geospatial Flatwoods pattern of weevil?
What causes geospatial patterns of EPN species? Citrus tree
Herbivore Pest
Endemic Entomopathogenic Nematodes
Soil Physical Natural Properties Enemies Citrus tree Current evidence-based conservation tactics focus on the effects of: Herbivore 3 1. Soil moisture Pest 2. Soil pH 4 3. Root HIPVs 4. Non-native EPN introductions Endemic Entomopathogenic Nematodes 1 Soil Physical Natural Properties 2 Enemies Florida EPN Foodweb Surveys Citrus orchards (53) Geospatial Natural areas (91) • Citrus orchards (4) Temporal qPCR primers/probes to measure EPNs and natural enemies EPNs (13 species) Natural enemies of EPNs Free living nematode (FLN) competitors (Acrobeloides spp.) Ectoparasitic bacteria (Paenibacillus, 2 species) Nematophagous fungi (NF, 7 species; nematophagy assay) Soil properties Texture Soil physico-chemistry (15 elements, pH, CEC, AWC, etc.) Pesticide residues (6 classes) Elevation Groundwater depth Florida EPN Foodweb Surveys Citrus orchards (53) Geospatial Natural areas (91) • Citrus orchards (4) Temporal qPCR primers/probes to measure EPNs and natural enemies
Sampled sites using soil probes to obtain 15 cores/acre composited into a single sample. Repeat to obtain 2 samples per site.
Extract nematodes from a 500 cm3 subsample.
Extract DNA, quantify target species with species-specific primers-probe sets to measure nematodes and organisms closely associated with nematodes. Variables that determine soil moisture were those that explained significant amounts of variation in the EPN spatial patterns at the landscape scale.
Depth to groundwater Water holding capacity Organic matter Clay content Sentinel wax moth larvae killed by the four most commonly encountered EPN species in Florida citrus orchards
now S. khuongi 6% 18 % 6% 10% 18% 30% Glycerol Conserving EPNs In addition to augmenting soils with the exotic Steinernema riobrave to manage Diaprepes root weevil, there are options to enhance and exploit naturally occurring EPN communities:
Employ composted animal manure mulch to increase biological control by EPNs Coarsen soil texture in tree planting holes to increase EPN efficacy Reduce soil pH to inhibit bacterial ectoparasites of EPNs Introduce Steinernema khuongi into depauperate flatwoods orchards Plant germplasm that respond to herbivory with semiochemical attractants for EPNs Polk County Indian River County 96% sand 88% sand H. indica 52% mortality (35-80%) 28% mortality 42% EPN (29-78%) 14% EPN
H. zealandica
Osceola County S. diaprepesi 53% sand 16% mortality S. riobrave H. indica 3% EPN
H. indica Conservation Serendipity
Sand or loam in planting hole Augment Sd, Sx, Sr, Hi, Hz Fewer weevils in sand More active EPNs in sand >Tree survival in sand Sd , Sr and Hi dominant for 6 yrs Paper published… …Two years later… EPN community succession in site with shallow depth to groundwater 2011 2013 S Sr Sd r Sx Sx Sandy Hi Sand favored soil Hi EPNs
Sx displaced Sd Sd Sx in site with Native Sr shallow water loamy soil Hi Hi table Conserving EPNs In addition to augmenting soils with the exotic Steinernema riobrave to manage Diaprepes root weevil, there are options to enhance and exploit naturally occurring EPN communities:
Employ composted animal manure mulch to increase biological control by EPNs Coarsen soil texture in tree planting holes to increase EPN efficacy Reduce soil pH to inhibit bacterial ectoparasites of EPNs Introduce Steinernema khuongi into depauperate flatwoods orchards Plant germplasm that respond to herbivory with semiochemical attractants for EPNs Infective (3rd stage) juvenile Steinernema diaprepesi with oval spores adhering to the 3rd stage cuticle along the length of the body. Development
Cast 3rd stage S. diaprepesi cuticles from insects infected by IJ that were either unencumbered (above) or spore-encumbered (right). Infective (3rd stage) juvenile Heterorhabditis spp. with spindal-shaped spores adhering the length of the body to the 2nd stage cuticle (sheath). Isolated and characterized bacterium associated with S. diaprepesi Morphology Stain FAME Partial 16S rDNA (536 bp) P. thiaminolyticus P. popilliae Entomopathogens P. lentimorbus Paenibacillus sp. Not entomopathogens P. nematophilus P. polymyxa Not entomopathogenic B. sphaericus Specific to S. diaprepesi G. stearothermophilus Did not inhibit EPN T. candidus reproduction T. vulgaris A. cycloheptanicus Impaired EPN host- A. acidocaldarius finding in bioassays P. ramosa Pasteuria S1 Do they regulate P. penetrans P20 EPN populations in
5 nature? Conventional Advanced Production System Citriculture (CC) Higher density planting Daily fertigation Drippers rather than microjets 19 months Greater resource efficiency. Earlier return on investment Advanced Harvestable yield by 2nd year Production System Less ground water pollution (APS) Very different soil physico-chemical
19 months properties Conventional Advanced Production System Citriculture (CC) Very different soil physico-chemical properties
19 months Compared EPN communities and natural enemies of EPNs (NF, FLN, Advanced Paenibacillus spp.) isolated from Production System nematode samples using qPCR. (APS)
19 months Among many soil properties, APS raised soil pH APS increased the 0.9 infestation rate 0.8 of Paenibacillus sp. 0.7
0.6
APS decreased the 10 M 1.0 0.5 50 M abundance of Steinernema 0.8 100 M diaprepesi – 0.4 0.6 Did this affect IJs with spores 0.4
0.3 0.2 weevil abundance? Encumbered % 0.0 0.2 5.0 5.5 6.0 6.5 7.0 7.5 8.0 Laboratory studies showed pH 0.1 that pH is directly related to 5.0 5.5 6.06.5 7.0 7.5 8.0 the ability of Paenibacillus pH sp. to adhere to the nematode cuticle APS increased the abundance of Diaprepes abbreviatus and Artipus floridanus root weevils.
Phytophthora nicotianae responded to increased root herbivory by infecting the roots at a higher rate. Temporal Food Web Surveys
0-15 cm 4 Orchard sites
15-30 cm Sampled monthly for two years
0-15 cm 10 trees/orchard
15-30 cm 3 Samples/orchard each month
0-15 cm Samples divided between ‘deep’ 0-15 cm 15-30 cm and ‘shallow’ Extracted 15-30 cm Temporal Food Web Surveys
0-15 cm Extracted nematodes 15-30 cm from 500 cm3 soil
0-15 cm Recovered DNA from 15-30 cm nematode sample
Measured target 0-15 cm EPNs, FLNs, bacteria 0-15 cm 15-30 cm and fungi.
15-30 cm Temporal Foodweb Surveys qPCR primers/probes for EPNs and natural enemies • EPNs (13 species) Natural enemies of EPNs Free living nematode (FLN) competitors (Acrobeloides spp.) Ectoparasitic bacteria (Paenibacillus, 2 species) Nematophagous fungi (NF, 7 species) Soil properties Texture Soil physico-chemistry (15 elements, pH, CEC, AWC, etc.) Pesticide residues (6 classes) Elevation Groundwater depth 4.0 Circle - Shallow 3.5 Triangle - Deep 3.0 2.5 As seen in APS trial, the abundance 2.0 of S. diaprepesi was inversely S. diaprepesiS. 1.5 n related to soil pH and to spore 1.0 Log per half liter of soil) encumbrance. ( r = 0.92 0.5 P = 0.01 Steinernema diaprepesi 0.0 infested with r = 0.97 Paenibacillus sp. 0.5 P = 0.001 0.0 As seen both in the lab and in the -0.5
encumbrance APS trial, encumbrance of S. -1.0 diaprepesi by Paenibacillus sp. was -1.5 directly proportional to solution or -2.0
Paenibacillus soil pH. -2.5 5.6 5.8 6.0 6.2 6.4 6.6 Soil pH Lotka-Voltera phase-space model of predator-prey interaction predicts counterclockwise dynamic for the asynchronous populations
2
a7
a22
0 a10
a11 a18 a13 a12 a8 a9
a14 a15 -2 a16 a17 a19 a23 a21 a20 Spore infection Sporerate infection -4 a25 a24
Nematode population size Logn (Sd + 1) 0 1 2 3 4 5 6 0 1 2 3 4 0 1 2 3 4 5 White=Encumbrance Black=EPN A. CREC A. A. N40 A. A. PCD A. OD MMJJAOD MMJJAS J J FMAM ASONDJ J J FMAM SONDJ Month -4 -2 -5 -4 -3 -2 -1 0 2 4 -4 -2 0 1 2 3 0 2 4 6
logn (Psp + 1) - logn (Sd + 1) 5 4 3 2 1 0 a17 a16 a20 a24 P=0.001 88% counterclockwise a11 a10 P=0.01 80% counterclockwise B. CREC B. PCD B. a23 a9 a14 a13 B. N40 B. a2 a25 a24 a23 a18 a15 a17 a18 a16 a22 a11 a12 a8 a19 a20 a7 a15 a7 a21 a4 a10 Log a18 a19 a16 a22 a15 a3 a21 a14 a13 a1 a10 a9 a4 a19 a12 a6 n a14 a13 a25 a11 (Sd+1) a23 a17 a2 a8 a7 a20 a1 a8 a5 a12 a3 a25 a5 a24 a21 a22 a9 a6 -5 -4 -3 -2 -1 -4 -2 -4 -2 0 1 2 0 2 4 0 2 4 6
logn (Psp + 1) - logn (Sd + 1) Orchard 3 Orchard 2 Orchard 1 Voltera consistent Lotka with encumbrance were in Periodsof major change Major cyclesrequired 3 model. months. S. diaprepesi phase - space and sporeand - -4 -4 Temporal Food Web Surveys
The surveys also revealed evidence for density dependent regulation by other natural enemies of EPNs. qPCR finds only what we seek. What if we could measure everything? Metagenomics using high throughput sequencing and do that.
Material and Methods cont’d
• s300 Ij / cm2 , 12M Ijs per Tree • Sprayed with nitrogen tanks at 20psi • 8 cores each sample • 96 samples • Sucrose Centrifugation • DNA extraction
2m
2m Populations of H. bacteriophora Declined much more rapidly Hb and Sf population than those of S. feltiae. fluctuation through the time S. feltiae
• Figure shows much faster decline of he lowest abundance of the species are on 14th day after application
H. bacteriophora 100% Relative Abundance of microarthropod families Acaridae Analgidae Histiostomatidae 80% Hypoderatidae Kramerellidae Lobalgidae Proctophyllodidae Psoroptidae Psoroptoididae Pyroglyphidae Trouessartiidae Japygidae Parajapygidae Alicorhagiidae Alycidae Nanorchestidae Pachygnathidae Entomobryidae 60% Isotomidae Seiridae Tomoceridae Argasidae Ixodidae NA AmeroseiidaeTullbergiidaeAscidae Blattisociidae Laelapidae Leptolaelapidae Melicharidae OlogamasidaeScheloribatidaePachylaelapidae Parasitidae 40% ParatennulidaeOppiidae Parholaspididae Phytoseiidae Rhodacaridae Uropodidae Zerconidae Neelidae CeratozetidaeAchipteriidae Brachychthoniidae Cepheidae AchipteriidaeCeratokalummidae Ceratoppiidae Ceratozetidae Chamobatidae Crotoniidae 20% CymbaeremaeidaeParajapygidaeDamaeidae Eremaeidae Euphthiracaridae Galumnidae Haplozetidae Hemileiidae Hermanniellidae Hydrozetidae Lohmanniidae Malaconothridae Mochlozetidae Mycobatidae Nothridae Oppiidae 0% Oribatellidae Oribatulidae Oripodidae Phenopelopidae Punctoribatidae Scheloribatidae Sf Hb Control Potential Predators of EPN Potential Prey of EPN
Treatments Control Treatments Hb Control Sf Hb Sf Intra-family variability of a potential predator of EPN
39.3 %
22.7 %
20%
10% Conserving EPNs In addition to augmenting soils with the exotic Steinernema riobrave to manage Diaprepes root weevil, there are options to enhance and exploit naturally occurring EPN communities:
Reduce soil pH to inhibit bacterial ectoparasites of EPNs Coarsen soil texture in tree planting holes to increase EPN efficacy Employ composted animal manure mulch to increase biological control by EPNs Introduce Sx into depauperate flatwoods orchards Plant germplasm that respond to herbivory with semiochemical attractants for EPNs Olfactometers Cage with Diaprepes larva + or - Pregeijerene Bury cages beneath citrus trees
100 billion neurons in the human brain 302 neurons in the body of C. elegans Conserving EPNs Soil is vastly more complex than are habitats aboveground. Conservation of arboreal natural enemies is routine, whereas conservation biocontrol is rare in soil. Does the endemic EPN community have potential to provide significant pest management? Is Florida citrus an exception? Same methodology revealed orchards in Portugal have much lower species diversity and detection frequency lower than Florida. Metagenomic methods will facilitate understanding food web/habitat dynamics. EPN-microarthropod barcode work is ongoing. Tarasco, et al. (2014). Journal of Helminthology, 1- 8. 10 Broadleaf forests (408) 9 Pine forests (302) A 10-year survey of Italian 8 Grasslands (88) habitats revealed a gradient of Uncultivated fields (217) EPN detection frequency 7 related to habitat disturbance. 6 Orchards (355) However the detection 5 Sea coasts (182) frequency is quite low. 4 EPN detection frequency (%) EPNdetection frequency Cultivated fields (329) 3 High Low Habitat Disturbance (number samples in parentheses) (53) 1.0 Some recent (5 years) EPN
(19) surveys 0.8 (91) Typical of previous reports 0.6 Wide range in detection 0.4 (280) (149) EPNdetected frequency 0.2
(proportionsamples) of (791) (288) (90) (570) EPNs detected in all citrus 0.0 orchards sampled in Florida
BeninChina Mexico EthiopiaLebanon PortugalTanzania Florida NA Florida Citrus Conserving EPNs Soil is vastly more complex than are habitats aboveground. Conservation of arboreal natural enemies is routine, whereas conservation biocontrol is rare in soil. Does the endemic EPN community have potential to provide significant pest management? Is Florida citrus an exception? Same methodology revealed orchards in Portugal have much lower species diversity and detection frequency lower than Florida. Metagenomic methods will facilitate understanding food web/habitat dynamics. EPN-microarthropod barcode work is ongoing. Questions?
“Everything you can imagine is real”
--Pablo Picasso Termites Identification of the pest
Order Isoptera Family Rhinotermitidae
• Small to medium size, white to yellow
• Social groups and have a highly developed caste system • Most common Reticulitermes flavipes Termites Biology and ecology
• Pine-palmetto woodlands are their natural habitat • When planted with citrus, termites were maintained in the buried wood • Feed on living bark of young trees above and belowground areas, girdling the tree • P. nicotinae often infects trees attacked by termites
McCoy (1999) Citrus Health Management Termites IPM
• Removal of wood residues, especially before planting
• Shallow planting to expose upper scaffold roots
• Avoid tree wraps for cold protection
• Monitoring regularly with wood bait blocks or wooden stakes
McCoy (1999) Citrus Health Management Ants Identification of the pest
Order Hymenoptera Family Formicidae
• Ubiquitous social insects
• Ants may play an important roll in the dynamics of citrus production and successful pest management
Predator Omnivorous Foliar damage Humans
• Solenopsis invicta, fire ant Ants Biology and ecology
• S. invicta builds nests in disturbed areas
• Those might be close to surface irrigation lines or beneath wraps used for cold protection
• Active feeding on the bark and tender new leaf flush of young trees that can kill by trunk girdling
Ants might play different rolls in the citrus agroecosystem, some will be beneficial and others not. Ecological consideration might be included when managing this organism Where the authors are right now
mulch on EPN efficacy Effects of compostedmanure results? measurable Can produce conservation Duncan etal.,JON (2007)
Weevils plot-1 0.00 0.25 0.50 0.75 1.00 1.25 from months during soil 12 caughtweevils emerging adult of EPNs and reducednumbersthe sentinelweevilsby nativekilled increased the mortality of Manure decreased mulches NF, Control Sr Mulch Mulch P=0.06 P=0.02 Sr Mulch Sr +