Overall Trends in the Efficacy of Allied Group/CleanFlo’s ... · Benefits and Limitations of the...

Overall Trends in the Efficacy of Allied Group/CleanFlo’s Laminar Flow

Aeration and Bioaugmentation as a Treatment for Sediment Floc Reduction,

Nuisance Algae Reduction, Water Quality Improvements, and Other Observed

Ecological Benefits, in Lake Apopka, Florida

January, 2016

Notice: This report is © copyrighted and property of Restorative Lake Sciences and Allied

Group. No portion of this report can be duplicated or used without written consent from the

Water Resources Director at Restorative Lake Sciences (Effective January 4, 2016).

Prepared by: Restorative Lake Sciences

Jennifer L. Jermalowicz-Jones, Ph.D. Candidate

Water Resources Director

18406 West Spring Lake Road

Spring Lake, Michigan 49456

www.restorativelakesciences.com

http://www.restorativelakesciences.com/

Restorative Lake Sciences

Lake Apopka Aeration Interim Report

January, 2016

TABLE OF CONTENTS

SECTION PAGE

1.0 INTRODUCTION & SUMMARY........................................................................................................ 6

1.1 Introduction to Inversion Oxygenation and Bioaugmentation ................................................. 7

1.2 Summary of Lake Apopka Aeration Operations ....................................................................... 9

1.3 Summary of Lake Apopka Aeration Operation Purpose/Goals ............................................... 9

2.0 LAKE APOPKA WATER QUALITY SAMPLING METHODS ....................................................... 9

2.1 Summary of Equipment/Sampling Devices/Replicates/Parameters ......................................... 9

2.2 Sampling Dates and Locations ............................................................................................... 10

3.0 LAKE APOPKA WATER QUALITY DATA ANALYSIS AND DISCUSSION ............................ 11

3.1 Dissolved Oxygen .................................................................................................................... 11

3.2 Water Temperature ................................................................................................................. 11

3.3 Conductivity ............................................................................................................................ 11

3.4 Total Suspended Solids ........................................................................................................... 12

3.5 Water Column Total Phosphorus and Total Nitrogen ........................................................... 12

3.6 pH ............................................................................................................................................ 13

3.7 Turbidity .................................................................................................................................. 13

3.8 Oxidative Reduction Potential ................................................................................................ 14

3.9 Algal Composition and Chlorophyll-a .................................................................................... 14

3.10 Organic Matter........................................................................................................................ 15

3.11 Sediment Nutrients (Nitrogen and Phosphorus) ..................................................................... 16

4.0 LAKE APOPKA SEDIMENT BOTTOM HARDNESS MAPS ........................................................ 20

5.0 LAKE APOPKA AQUATIC VEGETATION BIOVOLUME MAPS .............................................. 23



January, 2016

6.0 LAKE APOPKA INTERIM PROJECT CONCLUSIONS & RECOMMENDATIONS .................. 24

7.0 SCIENTIFIC LITERATURE CITED .................................................................................................. 27



January, 2016

FIGURES

FIGURE PAGE

Figure 1. Map of Lake Apopka Water Quality Sampling Locations .......................................................... 10

Figure 2. Lake Apopka Rapid Assay Phytoplankton Diagram (May, 2015) .............................................. 15

Figure 3. Lake Apopka Depth Contour Map (May, 2015) ......................................................................... 21

Figure 4. Lake Apopka Depth Contour (December, 2015) ......................................................................... 21

Figure 5. Lake Apopka Bottom Hardness Map (May, 2015) ..................................................................... 22

Figure 6. Lake Apopka Bottom Hardness Map (December, 2015) ............................................................ 22

Figure 7. Lake Apopka Aquatic Vegetation Biovolume Map (May, 2015) ............................................... 23

Figure 8. Lake Apopka Aquatic Vegetation Biovolume Map (December, 2015) ...................................... 23

Figure 9. Conceptual Diagram of Lake Apopka Bottom Sediment Layers (Allied Group) ....................... 25

Figure 10. Conceptual Diagram of Water/Floc Flow in Lake Apopka (Allied Group) .............................. 25



January, 2016

TABLES

TABLE PAGE

Table 1. Lake Apopka Mean Baseline Treatment WQ Physical Data (May, 2015) ................................... 17

Table 2. Lake Apopka Mean Baseline Control WQ Physical Data (May, 2015) ....................................... 17

Table 3. Lake Apopka Mean Baseline Treatment WQ Chemical Data (May, 2015) ................................. 18

Table 4. Lake Apopka Mean Baseline Control WQ Chemical Data (May, 2015)...................................... 18

Table 5. Lake Apopka Mean Post-Aeration Treatment WQ Physical Data (December, 2015) .................. 19

Table 6. Lake Apopka Mean Post-Aeration Control WQ Physical Data (December, 2015) ...................... 19

Table 7. Lake Apopka Mean Post-Aeration Treatment WQ Chemical Data (December, 2015) ................ 20

Table 8. Lake Apopka Mean Post-Aeration Control WQ Chemical WQ Data (December, 2015) ............ 20

Table 9. Lake Apopka Sediment Loss Statistics (BioBase®) ..................................................................... 22

Table 10. Lake Apopka Aquatic Vegetation Biovolume Loss Statistics (BioBase®) ................................ 24



January, 2016

Overall Trends in the Efficacy of Allied Group/CleanFlo Laminar Flow Aeration

and Bioaugmentation as a Treatment for Sediment Floc Reduction, Nuisance

Algae Reduction, Water Quality Improvements, and Other Observed Ecological

Benefits, in Lake Apopka, Florida

January, 2016

1.0 INTRODUCTION AND SUMMARY:

Lake Apopka is a 12,500 ha (30,888 acre) hypereutrophic lake located in central Florida approximately

32 km (20 miles) northwest of Orlando, Florida. The lake has an approximate mean depth of 1.6 meters

(5.2 feet) and has been the focus of intense restoration efforts over the past few decades due to excessive

nutrient inputs that have devastated the aquatic ecosystem and have transitioned the lake from a clear

macrophyte-dominated system to a turbid phytoplankton-dominated system.

The most prominent phytoplankton communities consist of blue-green algae (cyanobacteria) such as

Planktolyngbya spp., among others. These blue-green algae are capable of secreting neurotoxins which

are known saxotoxins. In addition to the turbidity issue caused by the high density of phytoplankton,

there is an overabundance of soft organic flocculent muck on the lake bottom. This material becomes

easily suspended during wave activity or when boats make contact with the lake bottom. As a result of

these factors, the dissolved oxygen concentration of the lake water is often compromised due to the

lack of light penetration into the water column and the high water temperatures that are increased both

by seasonality and the high amount of suspended solids that are continually circulated throughout the

water column.

When the lake was in a previous macrophyte-dominated state, previous technologies such as

mechanical harvesting were used. Water Hyacinth and lake drawdown were also historically used to

reduce the nutrient pool present in the water column and sediments of the lake. Eventually, there arose

a need for a technology or a set of technologies that would increase the dissolved oxygen of the water

column, reduce the flocculent organic muck layer on the bottom, and reduce the nuisance blue-green

algae in the water column, along with the excess nutrients in the lake water column. This report analyzes

the implementation of a 250-acre proprietary Allied Group/CleanFlo laminar flow aeration system

supplemented with microbes (bioaugmentation) near the area known as Magnolia Park in northern Lake

Apopka. In four months of operation, the system has significantly reduced sediment floc and nutrients

in the treatment area in both the water and sediments as well as increased dissolved oxygen

concentrations. Aquatic vegetation biovolume has also significantly declined.



January, 2016

1.1 Introduction to Laminar Flow Aeration and Bioaugmentation (In General):

Allied Group/CleanFlo laminar flow aeration systems are retrofitted to a particular site and account for

variables such as water depth and volume, contours, water flow rates, and thickness and composition

of lake sediment. The systems are designed to completely mix the surrounding waters and evenly

distribute dissolved oxygen throughout the lake sediments for efficient microbial utilization.

An Allied Group/CleanFlo laminar flow aeration system utilizes diffusers which are powered by

onshore air compressors. The diffusers are connected via extensive self-sinking airlines which help to

purge gasses such as H2S from the sediments. In addition to the placement of the diffuser units, the

concomitant use of bacteria and enzymatic treatments to facilitate the microbial breakdown of organic

sedimentary constituents is also used as a component of the treatment.

Beutel (2006) found that lake oxygenation eliminates release of (ammonia) NH3+ from sediments

through oxygenation of the sediment-water interface. Allen (2009) demonstrated that NH3+ oxidation

in aerated sediments was significantly higher than that of control mesocosms with a relative mean of

2.6 ± 0.80 mg N g dry wt. day-1 for aerated mesocosms and 0.48 ± 0.20 mg N g dry wt. day-1 in controls.

Although this is a relatively new area of research, recent case studies have shown promise on the

positive impacts of laminar flow aeration systems on aquatic ecosystem management with respect to

organic matter degradation and increase in water depth and rooted aquatic plant management in

eutrophic ecosystems (Restorative Lake Sciences, 2009-present).

Toetz (1981) found evidence of a decline in Microcystis algae (a toxin-producing blue-green algae) in

Arbuckle Lake in Oklahoma. Other studies (Weiss and Breedlove, 1973; Malueg et al., 1973) have

also shown declines in overall algal biomass.

Conversely, a study by Engstrom and Wright (2002) found no significant differences between aerated

and non-aerated lakes with respect to reduction in organic sediments. This study was however limited

to one sediment core per lake and given the high degree of heterogeneous sediments in inland lakes

may not have accurately represented the conditions present throughout much of the lake bottom. The

philosophy and science behind the inversion oxygenation aeration system is to reduce the organic

matter layer in the sediment so that a significant amount of nutrient is removed from the sediments and

excessive sediments are reduced to yield a greater water depth.



January, 2016

Benefits and Limitations of the Laminar Flow Aeration System (In General):

In addition to the reduction in toxic blue-green algae (such as Microcystis sp.) as described by Toetz

(1981), aeration and bioaugmentation in combination have been shown to exhibit other benefits for the

improvements of water bodies. Laing (1978) showed that a range of 49-82 cm of organic sediment was

removed annually in a study of nine lakes which received aeration and bioaugmentation. It was further

concluded that this sediment reduction was not due to re-distribution of sediments since samples were

collected outside of the aeration “crater” that is usually formed. A detailed study by Restorative Lake

Sciences of Indian Lake (Cass County, Michigan) during 2010-2015 also indicated a significant

reduction of organic sediments in bio augmented/aerated regions, as well as a decline in the relative

proportion of blue-green algae and the presence of the rooted, submersed, exotic aquatic plant, Eurasian

Watermilfoil (Myriophyllum spicatum). A study by Turcotte et al. (1988) analyzed the impacts of

bioaugmentation on the growth of M. spicatum and found that during two four-month studies, the

growth and re-generation of this plant was reduced significantly with little change in external nutrient

loading.

Furthermore, bacteria are the major factor in the degradation of organic matter in sediments (Fenchel

and Blackburn, 1979) so the concomitant addition of microbes to lake sediments will accelerate that

process. A study by Verma and Dixit (2006) evaluated aeration systems in Lower Lake, Bhopal, India,

and found that the aeration increased overall dissolved oxygen, and reduced biochemical oxygen

demand (BOD), chemical oxygen demand (COD), and total coliform counts.

The Allied Group/CleanFlo laminar flow aeration system has some limitations including the inability

to degrade mineral sediments, the requirement of a constant Phase I electrical energy source to power

the units, and unpredictable response by various species of rooted aquatic plants. Thus, the main

objective of inversion oxygenation system use should be related primarily to sediment parameters and

not in generalized aquatic plant control.

Expectations of Lake Apopka Basin Allied Group/CleanFlo Laminar Flow Aeration System:

The scientific research supports inversion oxygenation for the decline in sediment organic matter and

ammonia nitrogen which fuels primary production as well as reductions in water column nutrients and

chlorophyll-a. This phenomenon is also related to an observed increase in water clarity among aeration

lakes. The water clarity of Lake Apopka can expect to improve with implementation of an Allied

Group/CleanFlo laminar flow aeration system if it significantly reduces the planktonic algae and water

column nutrients that fuel algae growth. Since baseline data has been collected, post-installation data

offers definitive conclusions on the aeration program efficacy to assist in future decision-making.



January, 2016

1.2 Summary of Lake Apopka Aeration Operations:

Allied Group/CleanFlo laminar flow aeration was installed in the area known as Magnolia Park of

northern Lake Apopka between mid-July and mid-August of 2015. Baseline evaluation of this area

began just prior to installation in early May of 2015 in the pilot project area. The official start-up of the

Allied Group/CleanFlo laminar flow aeration technology began in mid-August of 2015. The pilot

project area consisted of a 250-acre area consisting of 96 ceramic diffusers with 156,000 feet of airline

and two 23 hp compressors. An additional three diffusers were added to the canal leading to the pilot

project area in September of 2015 and this area is being evaluated and will be analyzed by May of

2016. Microbial application (bioaugmentation) was added to the pilot project area two months apart

with the dosing of 1,000 lbs. of custom microbes and 400 gallons of custom concentrated enzymes.

1.3 Summary of Lake Apopka Aeration Operation Purpose/Goals:

The Fisheries and Wildlife Commission (FWC) desired a lake restoration strategy that would make the

lake healthier and accomplish the following objectives:

1) Increase dissolved oxygen concentrations in water column

2) Reduce the organic “floc” layer and muck layers on the lake bottom

3) Reduce blue-green algae in the water column

4) Reduce sediment and water column nutrients

5) Reduction of aquatic vegetation biovolume on the lake bottom

2.0 LAKE APOPKA WATER QUALITY SAMPLING METHODS:

2.1 Summary of Equipment/Sampling Devices/Replicates/Parameters:

Restorative Lake Sciences (RLS) and Allied Group (AG) sampled 15 diffuser and 8 control sites both

before implementation of aeration in May of 2015 and again in December of 2015 for sediments and

for water column parameters in the pilot project area.

All chemical water samples (such as total phosphorus, total nitrogen, and total suspended solids) were

collected at the specified depths (one each at the middle depths of each of the sampling sites) using a

4-liter VanDorn horizontal water sampler with weighted messenger (Wildco® brand). Water physical

parameters (such as water temperature, dissolved oxygen, conductivity, turbidity, oxidative reduction

potential and pH) were measured with a calibrated YSI® 6920 V2 multi-probe meter. All water and

sediment samples were sent to Flowers Laboratories (Altamonte Springs, Florida) for analysis.

All data were analyzed with an independent samples t-test at the 95% level of significance (p<.05).



January, 2016

2.2 Sampling Dates and Locations:

Water quality samples were collected on May 5, 2015 by Restorative Lake Sciences and again on

December 14, 2015 by Allied Group staff. The map in Figure 1 shows the treatment area border in

green. The small white squares are the diffuser locations and the flags (1D through 15D) are the

sampling points. The red lines delineate which of the compressors provides air to which the diffusers.

There are also eight sampling locations that exist ½ mile outside the boundary to the west and south.

Figure 1. Aeration diffuser sampling locations on Lake Apopka, Florida (2015). Note: There were 8

control sites sampled outside of the project area which were marked with GPS.



January, 2016

3.0 LAKE APOPKA WATER QUALITY DATA ANALYSIS AND DISCUSSION:

3.1 Dissolved Oxygen:

Dissolved oxygen (DO) is a measure of the amount of oxygen that exists in the water column. In

general, DO levels should be greater than 5 mg L-1 to sustain a healthy warm-water fishery or higher

for better overall water quality. DO is generally higher in colder waters. DO at the surface is generally

higher due to the exchange of oxygen from the atmosphere with the lake surface, whereas DO is lower

at the lake bottom due to decreased contact with the atmosphere and increased biochemical oxygen

demand (BOD) from microbial activity. A decline in DO may cause increased release rates of

phosphorus (P) from lake bottom sediments if DO levels drop to near zero milligrams per liter. Under

low DO conditions, the imminent accumulation of NH3+ (ammonia) may result in toxicity to aquatic

organisms (Camargo et al., 2005; Beutel, 2006).

The mean dissolved oxygen (DO) was 6.0±0.8 mg L-1 in the treatment area and 4.8±0.9 mg L-1 in the

control area in May of 2015. By December of 2015 the mean treatment DO was 9.1±1.4 mg L-1 and

the mean control DO was 9.0±0.6 mg L-1. This increase in dissolved oxygen in both the treatment and

control regions is due to the Allied Group/CleanFlo laminar flow aeration system.

3.2 Water Temperature:

The water temperature of Lake Apopka is quite stable and warm. In general, shallow lakes will not

stratify while deeper lakes may experience single or multiple turnover cycles. Water temperature was

measured in degrees Celsius (°C) with the use of a submersible thermometer. Differences in water

temperatures are due to seasonal effects of sunlight penetration, water level changes, and degree of

particles and color in the water that absorb heat. Typically, a challenge with whole-lake aeration is to

aerate the entire waterbody without causing thermal stress throughout the entire water column. The

May, 2015 mean water temperatures for both the control and treatment areas were around 23°±0.4C.

In December of 2015, the mean water temperatures for the control area were around 22°±0.1C and the

water temperatures in the treatment area were around 20°±0.7C. The changes were not significant,

however.

3.3 Conductivity:

Conductivity is a measure of the amount of mineral ions present in the water, especially those of salts

and other dissolved inorganic substances. Conductivity generally increases as the amount of dissolved

minerals and salts in a lake increases, and also increases as water temperature increases. Conductivity

is measured in micro-Siemens per centimeter (µS cm-1) with the use of a conductivity probe and meter.

The overall conductivity values of Lake Apopka have traditionally been moderate for a large inland

lake. The mean conductivity (measure of electrical current in the water) was 449±16 µS cm-1 in the



January, 2016

treatment area and 462±46 µS cm-1 in the control area in May of 2015. By December of 2015, the

treatment area had dropped to 404±8.6 µS cm-1 and the control area to 417±2.6 µS cm-1. These

significant reductions in conductivity of both the treatment and control regions are due to the Allied

Group/CleanFlo laminar flow aeration system.

3.4 Total Suspended Solids:

Total Suspended Solids (TSS) is a measure of the amount of suspended particles in the water column.

Particles suspended in the water column absorb heat from the sun and raise the water temperature. Total

suspended solids is often measured in mg L-1 and analyzed in the laboratory. The lake bottom contains

many fine sediment particles that are easily perturbed from winds and wave turbulence. Total

Suspended Solids (TSS) refers to the amount of solids in the water column. The mean TSS in the

treatment area was 93±9.0 mg L-1 and the control area was 97±5.0 mg L-1 in May of 2015. By

December of 2015, the TSS was 75±8.4 mg L-1 in the treatment area and 82±6.2 mg L-1 in the control

area. This is a significant reduction in TSS in the treatment area. The control region shows reductions

as well and will be monitored and analyzed in the final report.

3.5 Water Column Total Phosphorus and Total Nitrogen:

Total phosphorus (TP) is a measure of the amount of phosphorus (P) present in the water column.

Phosphorus is the primary nutrient necessary for abundant algae and aquatic plant growth. Lakes which

contain greater than 20 µg L-1 (or 0.020 mg L-1) of TP are defined as eutrophic or nutrient-enriched.

TP concentrations are usually higher at increased depths due to higher release rates of P from lake

sediments under low oxygen (anoxic) conditions. Phosphorus may also be released from sediments as

pH increases. The pH of Lake Apopka is high enough and the DO is low enough to create an

environment for continuous release of TP. Total Phosphorus (TP) is a measure of the amount of

phosphorus in the water column that can cause algae and plant growth. The mean TP in the treatment

area was 0.090±0.01 mg L-1 and the mean TP in the control area was 0.100±0.00 mg L-1 in May of

2015. By December of 2015, the mean TP in the treatment area was 0.280±0.06 mg L-1 and the control

area was 0.240±0.09 mg L-1. These numbers increased possibly due to mobilization of TP from

breakdown of sediment floc which is loosely consolidated and rich in phosphorus.

Total Kjeldahl Nitrogen (TKN) is the sum of ammonia (NH4+) and organic nitrogen forms of nitrogen

in freshwater systems. Much nitrogen (amino acids and proteins) also comprises the bulk of living

organisms in an aquatic ecosystem. Nitrogen originates from atmospheric inputs (i.e. burning of fossil

fuels), wastewater sources from developed areas (i.e. runoff from fertilized lawns), agricultural lands,

septic systems, and from waterfowl droppings. It also enters lakes through groundwater or surface

drainage, drainage from marshes and wetlands, or from precipitation (Wetzel, 2001). In lakes with an

abundance of nitrogen (N: P > 15), phosphorus may be the limiting nutrient for phytoplankton and

aquatic macrophyte growth. Alternatively, in lakes with low nitrogen concentrations (and relatively

high phosphorus), the blue-green algae populations may increase due to the ability to fix nitrogen gas



January, 2016

from atmospheric inputs. Lakes with a mean TKN value of 0.66 mg L-1 may be classified as

oligotrophic, those with a mean TKN value of 0.75 mg L-1 may be classified as mesotrophic, and those

with a mean TKN value greater than 1.88 mg L-1 may be classified as eutrophic. The mean (TKN) in

the treatment area was 3.6±0.1 mg L-1 and control area was 3.7±0.2 mg L-1 in May of 2015. By

December of 2015, the mean TKN decreased slightly to 3.2±03 mg L-1 in the treatment area and 3.1±0.2

mg L-1 in the control area. This represents a significant reduction on TKN in both the treatment and

control regions due to the Allied Group/CleanFlo laminar flow aeration system.

3.6 pH:

pH is the measure of acidity or basicity of water. The standard pH scale ranges from 0 (acidic) to 14

(alkaline), with neutral values around 7. Most Florida lakes have pH values that range from 6.3 to 8.4.

Acidic lakes (pH < 7) are rare in Florida and are most sensitive to inputs of acidic substances due to a

low acid neutralizing capacity (ANC). pH is measured with a pH electrode and pH-meter in Standard

Units (S.U). The pH of Lake Apopka water has ranged from 7.7-9.2 S.U. The mean baseline pH of

both the treatment and control area was 8.8±0.2 S.U. By mid-December of 2015, the mean post-

aeration pH in the aeration zone was 8.9±0.2 S.U. and the mean pH in the control zone was 9.1±0.1

S.U. There were no statistically significant differences in pH in the treatment or control zones.

3.7 Turbidity:

Turbidity is a measure of the loss of water transparency due to the presence of suspended particles.

The turbidity of water increases as the number of total suspended particles increases. Turbidity may

be caused by erosion inputs, phytoplankton blooms, storm water discharge, urban runoff, re-suspension

of bottom sediments, and by large bottom-feeding fish. Particles suspended in the water column absorb

heat from the sun and raise water temperatures. Since higher water temperatures generally hold less

oxygen, shallow turbid waters are usually lower in dissolved oxygen. Turbidity is measured in

Nephelometric Turbidity Units (NTU’s) with the use of a turbidimeter. The World Health Organization

(WHO) requires that drinking water be less than 5 NTU’s; however, recreational waters may be

significantly higher than that. The mean baseline turbidity in the treatment area was 28.1±4.2 NTU’s

and the mean turbidity in the control area was 36.6±14 NTU’s. Turbidity data was collected in the

control area in mid-December of 2015 and was determined to have a mean of 27.6±2.2 NTU’s which

is marginally significantly lower than in May of 2015. Turbidity data was collected in the treatment

area in mid-December of 2015 and was determined to have a mean of 29.7±3.8 NTU’s which is not

sigfnicantly different than the baseline values. This indicates that the Allied Group/CleanFlo laminar

flow aeration system is marginally reducing turbidity in the control region.



January, 2016

3.8 Oxidative Reduction Potential:

The oxidation-reduction potential (ORP) of lake water describes the effectiveness of certain atoms to

serve as potential oxidizers and indicates the degree of reductants present within the water. In general,

the Eh level (measured in millivolts) decreases in anoxic (low oxygen) waters. Low Eh values are

therefore indicative of reducing environments where sulfates (if present in the lake water) may be

reduced to hydrogen sulfide (H2S). The ORP values in Lake Apopka fluctuate substantially within a

single day, seasonally, and annually. The mean ORP (measure of oxidation-reduction potential in the

water) in the treatment area in was -107.5±51 µohms and in the control area was -145.1±27 µohms in

May of 2015. By December of 2015, the mean ORP was +186.5±5.4 µohms in the treatment area and

+221.6±6.1 µohms in the control area. ORP values were strongly negative during the baseline sampling

and strongly positive during the mid-December 2015 sampling. This indicates significant oxidative

effects of the Allied Group/CleanFlo laminar flow aeration system at the sediment water interface in

both the treatment and control regions.

3.9 Algal Composition and Chlorophyll-a:

Water samples for phytoplankton (algae) and chlorophyll-a analysis were collected at all sites via a

composite sample from above the sediment to the surface using a composite sampler as described by

Nicholls (1979). Samples were placed in dark brown polyethylene bottles and maintained at 4°C until

microscopic analysis could be executed. All samples were preserved with buffered glutaraldehyde and

analyzed within 48 hours of collection.

Chlorophyll-a is a measure of the amount of green plant pigment present in the water, often in the form

of planktonic algae. High chlorophyll-a concentrations are indicative of nutrient-enriched lakes.

Chlorophyll-a concentrations greater than 6 µg L-1 are found in eutrophic or nutrient-enriched aquatic

systems, whereas chlorophyll-a concentrations less than 2.2 µg L-1 are found in nutrient-poor or

oligotrophic lakes. Chlorophyll-a is measured in micrograms per liter (µg L-1) with the use of an

acetone extraction method and a spectrometer. Chlorophyll-a (chl-a) is a measure of the algal pigment

in the water. The mean chl-a in the treatment area was 15.5±23 µg L-1 and the control area was 7.0±8.8

µg L-1 in May of 2015. By December of 2015, the chl-a in the treatment was 46.3±29 µg L-1 and the

chl-a in the control was 12.5±18 µg L-1. The reason for this may be the same as for the increased TP

in that there is increased mobilization of soluble TP which will temporarily increase chl-a until the

sediment floc is further biodegraded.

Algal samples collected on May 13, 2015 were submitted to PhycoTech, Inc. (St. Joseph, Michigan)

for a rapid assay determination of algal species present and their relative abundance (see Figure 2). The

analysis determined that the sample contained predominantly Cyanophyta (blue-green algae) with the

genera Planktolyngbya spp., Aphanocapsa spp., and Aphanothece spp. There were very few

Chlorophyta (green algae) or Basicllariophyta (diatoms). An additional two samples collected on

October 8, 2015 which was sent to GreenWater Laboratories (Palatka, FL) also revealed that the

dominant alga was the blue-green Planktolyngbya spp. which was present in a concentration of



January, 2016

17,048,242 µg L-1 in the control area and a concentration of 14,908,984 µg L-1 in the treatment area.

Planktolyngbya spp. is capable of secreting saxotoxins which are nerve toxins. A goal of the Allied

Group/CleanFlo laminar flow aeration system is to reduce the overall presence of blue-green algae in

treatment areas since it is not a desirable component of the food chain for zooplankton in an aquatic

ecosystem.

Figure 2. Various alga taxa found in Lake Apopka May, 2015 samples (note the high dominance of

blue-green algae relative to more favorable green algae and diatoms).

3.10 Organic Matter:

Organic matter (OM) contains a high amount of carbon that is derived from biota such as decayed plant

and animal matter. Detritus is the term for all dead organic matter which is different than living organic

and inorganic matter. OM may be autochthonous or allochthonus in nature where it originates from

within the system or external to the system, respectively. Sediment OM is usually expressed in a

percentage (%) of total bulk volume. Lake Apopka sediment samples were collected at the sampling

locations in May of 2015 with the use of an Ekman hand dredge. The baseline mean percentage OM in

the treatment area was very high at 94±4% and the baseline OM mean percentage in the control area

was 96±0.4%. This indicates that the top sediment layers of Lake Apopka are almost entirely organic

and are capable of being degraded by microbes.



January, 2016

3.11 Sediment Phosphorus and Sediment Nitrogen:

Sediment phosphorus (TP) is a measure of the amount of phosphorus present in the lake sediment.

Phosphorus is the primary nutrient necessary for abundant algae and aquatic plant growth. The TP

concentrations in lake sediments are often up to several times higher than those in the water column

since phosphorus tends to adsorb onto sediment particles and sediments thus act as a “sink” or reservoir

of nutrients. TP concentrations are usually higher at increased depths due to higher release rates of

phosphorus from lake sediments under low oxygen (anoxic) conditions. Sediment TP is measured in

milligrams per kilogram (mg kg-1). These TP concentrations are high and anoxic sediment conditions

at the lake bottom likely result in higher release rates of phosphorus from lake sediments. A study by

Krogerus and Ekholm (2003) measured the release rates of phosphorus from sediment in shallow,

agriculturally impacted lakes and found that the mean daily rate of gross sedimentation was 0.04-0.18

g m-2 day-1 of phosphorus. Sediment Total Phosphorus (TP) is a measure of the amount of sediment

phosphorus in the sediments. The mean TP in the treatment area was 79.17±16 mg kg-1 and control

area was 87.31±18mg kg-1. By December of 2015 the TP in the treatment area was 66±41 mg kg-1 and

the control area was 59±17 mg kg-1. These numbers represent significant TP reductions in both the

treatment and control regions due to the Allied Group/CleanFlo laminar flow aeration system.

Sediment nitrogen consists of nitrate, nitrite, and ammonia forms. The total nitrogen is measured in

milligrams per kilogram (mg kg -1) of total sediment mass. In general, the nitrite form is very low and

the nitrate form is slightly higher but typically less than ammonia or other forms. Denitrifying bacteria

in the water and sediment then convert most of the nitrate to nitrogen gas. Nitrogen in the sediments

is derived primarily from decaying plant matter. The redox potentials in the lake sediments must be

sufficient for the ammonia in the sediment to be converted to nitrite. The conversion of ammonia

(NH3+) to organic nitrogen is necessary for the formation of lake bottom sediment and adds bulk

density. The release of NH3+ under anoxic conditions follows the accumulation of NH3+ in sediments

where nitrification cannot occur, and NH3+ assimilation by anaerobic microbes declines. The imminent

accumulation of NH3+ then results in toxicity to aquatic organisms (Camargo et al., 2005; Beutel,

2006). Beutel (2006) found that NH3+ release rates in anoxic sediment were > 15 mg N m-2 day-1, but

were nearly absent in oxic conditions.

Sediment Total Nitrogen (TN) and Ammonia Nitrogen (NH3) are measures of the amount of nitrogen

in the sediments. The mean sediment TN in the treatment area was 1241.73+360 mg kg-1 while the

control area registered 918.71+349 mg kg-1 in May 2015. By December of 2015, the mean sediment

TN in the treatment area increased to 1461+576 mg kg-1 and the control area increased to 1397+436

mg kg-1. Although both increased in this time period, it is interesting to note that the treatment area

increased by about half as much as the control area. With ammonia nitrogen the results were different.

The NH3 measurement in the treatment area in May of 2015 was 11.34+6 mg kg-1 while the control

area registered 17.9+8 mg kg-1. By December of 2015 the NH3 in the treatment area reduced to

10.9+6.9 mg kg-1 while the control area increased to 18.4+7.1mg kg-1. These numbers represent

nitrification of the NH3 into nitrates and nitrites. This is the first step toward denitrification. Although

both areas increased in TN, this shows a denitrification of TN in the treatment area over the control



January, 2016

area since the increase in the treatment area was less than half what occurred in the control area.

Tables 1-4 below show the summary baseline data for the measured water quality parameters. Tables

5-8 below show the summary post-aeration data for the measured water quality parameters.

Table 1. Baseline treatment water quality physical data (May 5, 2015) in Lake Apopka pilot

project area.

Table 2. Baseline control water quality physical data (May 5, 2015) in Lake Apopka pilot

project area.

Water

Column

Descriptive

Statistic

ORP

µohms

Water

Temp.

°C

pH

S.U.

Diss.

Oxygen

mg L-1

Turb.

mg L-1

Cond.

µS cm-1

TSS

mg L-1

Mean ± SD

Max

Min

-107.5± 51

-1.5

-157

22.9±0.4

23.2

21.8

8.8 ± 0.2

9.1

8.4

6.0 ± 0.8

7.3

4.0

28.1±4.2

39.1

20.4

449.3±16

503

435

92.7±9

106

72

Water

Column

Descriptive

Statistic

ORP

µohms

Water

Temp.

°C

pH

S.U.

Diss.

Oxygen

mg L-1

Turb.

mg L-1

Cond.

µS cm-1

TSS

mg L-1

Mean ± SD

Max

Min

-145.1± 27

-97

-172

22.8±0.4

23.3

22.3

8.8 ± 0.5

9.2

7.7

4.8± 0.9

6.1

3.7

36.6±13.8

69.0

25.6

462.4±46

566

428

96.6±5

104

88



January, 2016

Table 3. Baseline treatment water quality chemical data (May 5, 2015) in Lake Apopka pilot project area.

Table 4. Baseline control water quality chemical data (May 5, 2015) in Lake Apopka pilot project area.

Water

Column

Descriptive

Statistic

Water

Col

TKN

mg L-1

TP

mg L-1

Chl-a

µg L-1

Sediment

TN

mg kg-1

Sediment

OM

%

Sediment

NH3

mg kg -1

Sediment

TP

mg kg -1

Mean ± SD

Max

Min

3.6± 0.1

3.8

3.4

0.090 ± 0.01

0.119

0.082

15.5±23

88.6

1.0

1,242±360

2,140

468

94± 4

99

85

11.3±6

6.1

3.9

79.1±16

107

53

Water

Column

Descriptive

Statistic

Water

Col

TKN

mg L-1

TP

mg L-1

Chl-a

µg L-1

Sediment

TN

mg kg-1

Sediment

OM

%

Sediment

NH3

mg kg -1

Sediment

TP

mg kg -1

Mean ± SD

Max

Min

3.7± 0.2

4.0

3.5

0.100 ± 0.00

0.103

0.093

7.0±8.8

26.7

1.0

918±349

1,220

117

96± 0

96

95

18±7

8.2

30

66±16

167

4.0



January, 2016

Table 5. Post-aeration treatment water quality physical data (December, 2015 data) in

Lake Apopka pilot project area.

Table 6. Post-aeration control water quality physical data (December, 2015) in Lake Apopka pilot

project area.

Water

Column

Descriptive

Statistic

ORP

µohms

Water

Temp.

°C

pH

S.U.

Diss.

Oxygen

mg L-1

Cond.

µS cm-1

TSS

mg L-1

Turb.

NTU

Mean ± SD

Max

Min

186.5±5.4

195.6

176.3

20.0±0.7

22.3

19.4

8.9 ± 0.2

9.1

8.3

9.1 ± 1.4

10.5

5.1

404.6±8.6

433

397

74.8±8.4

96

62

29.7±3.8

25

38

Water

Column

Descriptive

Statistic

ORP

µohms

Water

Temp.

°C

pH

S.U.

Diss.

Oxygen

mg L-1

Cond.

µS cm-1

TSS

mg L-1

Turb.

NTU

Mean ± SD

Max

Min

220.6±6.1

230

211

22.0±0.1

22.1

21.9

9.1±0.1

9.1

8.9

9.0± 0.6

9.6

7.6

417.6±2.6

424

415

81.5±6.2

92

72

27.6±2.2

31

25



January, 2016

Table 7. Post-aeration treatment water quality chemical data (December, 2015) in Lake

Apopka pilot project area.

Table 8. Post-aeration control water quality chemical data (December, 2015) in Lake

Apopka pilot project area.

4.0 LAKE APOPKA DEPTH CONTOUR AND BOTTOM HARDNESS MAPS:

On May 4, 2015, and December 14, 2015 a benthic scan using a Lowrance® 50-satellite GPS HDS 8

WAAS-enabled unit (accuracy within 2 feet) was conducted to analyze a section of the bottom of Lake

Apopka in the pilot project area near Magnolia Park and record all of the bottom attributes which included

sediment bottom contours, bottom hardness and aquatic vegetation biovolume.

Water

Column

Descriptive

Statistic

Water

Col

TKN

mg L-1

TP

mg L-1

Chl-a

µg L-1

Sediment

TN

mg kg-1

Sediment

NH3

mg kg -1

Sediment

TP

mg kg -1

Mean ± SD

Max

Min

3.2±0.3

4.0

2.9

0.28 ± 0.06

0.41

0.20

46.3±29

105

1.7

1,461±576

2,730

544.0

10.9±6.9

26.0

1.4

65.9±41

167.0

4.0

Water

Column

Descriptive

Statistic

Water

Col

TKN

mg L-1

TP

mg L-1

Chl-a

µg L-1

Sediment

TN

mg kg-1

Sediment

NH3

mg kg -1

Sediment

TP

mg kg -1

Mean ± SD

Max

Min

3.0± 0.2

3.3

2.8

0.24 ± 0.09

0.33

0.07

12.5±18

59.3

1.0

1,397±436

1,990

735

18.4±7.1

29.8

8.2

59.4±17

80.1

33.1



January, 2016

The depth contour maps shown below represent the increase in water depth that has occurred between May

and December (Figures 3 and 4, respectively). An increase in depth can be seen where the organic matter

has been consumed by the microbes. Each line in these scans represents an increase of one foot in depth

from the surface of the lake with the dark blue representing the deepest areas in the treatment zone. By

December, some areas that were ≥ five feet were forming as well as expansion of the 2, 3 and 4 foot depth

contours. This area will be re-scanned again in June of 2016 and data will be compared to May of 2015 to

determine the amount of change in one year of operation.

Figure 3. May, 2015 depth contours Figure 4. December, 2015 depth contours

Figure 5 shows the baseline sediment bottom hardness of the pilot project area in May of 2015. The

statistics generated for this area (Table 9 below) report that there was a loss of 1.5% in total area of soft

bottom from May of 2015 to December of 2015 (Figure 6) due to effects of the Allied Group/CleanFlo

laminar flow aeration system. This means that the Allied Group/CleanFlo laminar flow aeration system

has effectively reduced sediment unconsolidated “floc” and consolidated organic sediment and is

converting the organic layers to into mineral or consolidated layers. This area will be scanned again in

June of 2016 and will once again be statistically compared to the baseline values so that more calculations

on sediment muck loss can be made. On the sediment maps, the red and orange colors indicate firmer

substrate types whereas the grey colors denote softer, organic substrate.



January, 2016

Figure 5. May, 2015 bottom hardness Figure 6. December, 2015 bottom hardness

Table 9. Changes in Lake Apopka sediment hardness before (May, 2015) and after (December, 2015)

aeration began. The percentages represent categories of bottom hardness. There has been a slight loss

of soft bottom in the treatment zone since the aeration program began.

Parameter May 2015 December 2015 Net Loss or Gain

Total Sonar Points 2095 2206 -- Total Acres Scanned 301.3 295.5 -- Soft Bottom (<.25) 1468 (70.1%) 1513 (68.6%) (1.5%) Medium Soft (.25 to .35) 514 (24.5%) 598 (27.1%) 2.6% Medium Hard (.35 to .40) 75 (3.6%) 74 (3.4%) (0.2%) Hard (.40 to .50) 17 (0.8%) 21 (0.9%) 0.1%



January, 2016

5.0 LAKE APOPKA AQUATIC VEGETATION BIOVOLUME MAPS:

Although aquatic vegetation species was not specifically studied as a component of this pilot project, the

aquatic vegetation biovolume is incidentally scanned as a component of the benthic scanning and adds

considerable evidence that the Allied Group/CleanFlo laminar flow aeration technology is influencing the

sediment water interface. The maps below show the changes from red (dense vegetation growth) to blue

(no vegetation growth) from five months of operation by the Allied Group/CleanFlo laminar flow aeration

system. Additionally Table 10 below quantifies the changes in aquatic vegetation biovolume. There has

been a substantial reduction in the highest percent cover categories (60-80%) from May to December of

2015 as a result of the Allied Group/CleanFlo laminar flow aeration system. This is due to the ability of the

Allied Group/CleanFlo laminar flow aeration system to affect the sediment pore water chemistry and drive

the denitrification process which limits sediment nutrients (such as ammonia) to many rooted macrophytes.

It is also possible that this scanning technology may have also detected loss of unconsolidated “floc” on the

lake bottom. This is being further investigated and is supported with observations such as the visual loss of

“floc” suspension after a boat enters the lake and creates a small wake relative to pre-aeration conditions

where “floc” was readily visible with any boat activity.

Figure 7. May, 2015 biovolume Figure 8. December, 2015 biovolume



January, 2016

Table 10. Changes in Lake Apopka aquatic vegetation biovolume before (May, 2015) and after

(December, 2015) aeration began. The percentages represent categories of aquatic plant cover relative

to the various heights of aquatic vegetation above the lake bottom. There has been a significant loss of

aquatic vegetation loss in the treatment zone since the aeration program began.

6.0 LAKE APOPKA INTERIM PROJECT CONCLUSIONS AND

RECOMMENDATIONS:

There exists a very high biochemical oxygen demand (BOD) in the pilot project area as the microbes

actively break down the existing sediment “floc” on the Lake Apopka bottom. Despite this process,

the Allied Group/CleanFlo laminar flow aeration system is able to produce an ample enough dissolved

oxygen concertation to compensate for losses in the water column. This means that the existing design

and blend of proprietary microbes are formulated correctly to break down the organic matter on the

lake bottom while allowing for the primary production to slowly transition over time to a green algae-

dominated phytoplankton community (and eventually diatom-dominated community) while not

causing the BOD to measurably reduce dissolved oxygen levels and affect aquatic life. A diagram of

the lake bottom sediments (provided to RLS from Allied Group) is shown below in Figure 9.

Additionally, benefits from the Allied Group/CleanFlo laminar flow aeration treatment zone may be

realized by the control zone. This means that the once confined 250-acre pilot project aeration

treatment area may be affecting an area of Lake Apopka much further out into the lake. This is an

unexpected but positive scientific discovery since it may lend support to further expansion of the Allied

Group/CleanFlo Laminar flow aeration system throughout other areas of the lake. Figure 10

demonstrates the movement of water that occurs relative to “floc” and also water that flows

perpendicular to shore. This may explain how water that is processed by the Allied Group/CleanFlo

laminar flow aeration system ends up in the measured “control” regions. In a sense, the aeration zone

acts as a continuous “bioreactor” site.

% Aquatic Plant Bio-Volume May 2015 December 2015 Net Loss or Gain

0%-5% 26.09% 85.93% 59.84%

5%-20% 8.69% 10.87% 2.18%

20%-40% 12.64% 1.29% (11.35%)

40%-60% 17.76% 0.99% (16.77%) 60%-80% 19.34% 0.55% (18.79%)

>80% 15.48% 0.37% (15.11%)



January, 2016

Figure 9. Conceptual diagram of the bottom sediment layers of Lake Apopka

(Allied Group, 2015).

Figure 10. Conceptual diagram of the flow of water and floc in the aeration zone in Lake

Apopka (Allied Group, 2015).



January, 2016

The Allied Group/CleanFlo laminar flow aeration system is reducing the presence of excessive rooted

submersed aquatic vegetation on the lake bottom and thus would possibly keep nuisance species such

as Hydrilla under control in the future since it is rooted via shallow tubers and the Allied

Group/CleanFlo laminar flow aeration technology readily influences sediment pore water nutrient

levels.

From a restoration perspective, the process of converting Lake Apopka from a “hyper-eutrophic” lake

to a “eutrophic” lake is possible but will take years to accomplish. Implementation of this Allied

Group/CleanFlo laminar flow aeration technology on the Lake Apopka pilot project area has already

shown that it can significantly reduce water column ammonia, TKN, sediment phosphorus and organic

matter due to the activity of unique proprietary blends of beneficial microbes. Restorative Lake

Sciences recommends continued prolonged use of the Allied Group/CleanFlo laminar flow aeration

system with microbes in the pilot project area as well as expansion of the Allied Group/CleanFlo

laminar flow aeration system into other regions of Lake Apopka. Additionally, we recommend

continued scientific monitoring such as that which has been executed and possibly more with increased

funding opportunities.

In summary, this interim study has demonstrated the existing Allied Group/CleanFlo laminar flow

aeration technology and unique blend of microbes has significantly increased dissolved oxygen,

reduced total suspended solids, reduced water column total nitrogen, reduced sediment ammonia and

phosphorus, oxidized lake sediments, and reduced conductivity. Other parameters such as chlorophyll-

a and total phosphorus have temporarily increased, possibly due to increased biological activity of

microbes and the increasing availability of light that will create a shift from cyanobacteria to

chlorophyta with time and temporarily increase the chlorophyll-a concentration. Eventually, a shift in

phytoplankton composition with diatoms and green algae in higher relative abundance will create a

healthier food chain base for the zooplankton community and will lead to overall improved water

quality.



January, 2016

7.0 SCIENTIFIC LITERATURE CITED

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bench-scale oxygen-activated nitrification wetlands. MS thesis, Washington State University,

Department of civil and Environmental Engineering.

Beutel, M.W. 2006. Inhibition of ammonia release from anoxic profundal sediments in lakes

using hypolimnetic oxygenation. Ecological Engineering 28(3): 271-279.

Camargo, J.A., A. Alonso, and A. Salmanca. 2005. Nitrate toxicity to aquatic animals: a review with

new data for freshwater invertebrates. Chemosphere 58: 1255-1267.

Engstrom, D.R., and D.I. Wright. 2002. Sedimentological effects of aeration-induced lake circulation.

Lake and Reservoir Management, 18(3):201-214.

Fenchel, T., and T.H. Blackburn. 1979. Bacteria and mineral cycling. Academic.

Krogerus, K., and P. Ekholm. 2003. Phosphorus in settling matter and bottom sediments in lakes

loaded by agriculture. Hydrobiologia 429: 15-28.

Laing, R.L. 1978. Pond/Lake Management organic waste removal through multiple inversion. In

house report. Clean-Flo Lab, Inc.

Malueg, K., J. Tilstra, D. Schults, and C. Powers. Effect of induced aeration upon stratification and

eutrophication processes in an Oregon farm pond. Geophysical Monograph Series, 17: 578-

587. American Geophysical Union. Washington DC.

Nicholls, K.H. 1979. A simple tubular phytoplankton sampler for vertical profiling in lakes.

Freshwater Biology 9:85-89.

Prescott, G.W. 1970. Algae of the western great lakes areas. Pub. Cranbrook Institute of Science

Bulletin 33:1-496.

Toetz, D.W., 1981. Effects of whole lake mixing on water quality and phytoplankton. Water

Research, 15: 1205-1210.

Turcotte, A.C., C.V. Déry, and K.F. Ehrlich. 1988. Manipulation of microbial ecosystems to control

the aquatic plant Eurasian Watermilfoil. Preprint paper. Département de Biologie, Université

de Sherbrooke, Sherbrooke, Québec, CANADA J1K 2R1.

Verma, N. and S. Dixit. 2006. Effectiveness of aeration units in improving water quality of Lower

Lake, Bhopal, India. Asian Journal of Experimental Science 20(1): 87-95.

Weiss, C., and B. Breedlove. 1973. Water quality changes in an impoundment as a consequence of

artificial destratification. 216 pp. Water Resources Research Institute. University of North

Carolina. Raleigh.

Wetzel, R. G. 2001. Limnology: Lake and River Ecosystems. Third Edition. Academic Press, 1006

pgs.

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