Limited Environmental Sampling - Martin

6278 North Federal Highway #450, Fort Lauderdale, FL 33308 | Phone 954.978.8839 | Fax 954.973.0530 www.enviroteam.com

Limited Environmental Sampling Martin County Engineering Department

Stuart, Florida

Martin County Engineering Department Dianne K. Hughes, Senior Ecosystem Specialist Ecosystem Restoration & Management Division 2401 SE Monterey Road, Stuart, Florida 34996

Prepared By:

Enviro Team North America, LLC 6278 North Federal Highway, #450

Ft. Lauderdale, FL 33308

July 25, 2016

http://www.enviroteam.com/

6278 North Federal Highway #450, Fort Lauderdale, FL 33308 | Phone 954.978.8839 | Fax 954.973.0530 www.enviroteam.com

July 25, 2016 Martin County Engineering Department Dianne K. Hughes, Senior Ecosystem Specialist Ecosystem Restoration & Management Division 2401 SE Monterey Road, Stuart, Florida 34996 Office: 772-219-4980 Cell: 772-260-0410 Reference: Limited Environmental Sampling Four Outdoor Locations Martin County, Florida Project No. 160185 Dear Ms. Hughes: Enclosed is the final report for the above referenced project. If you have any questions concerning this project or need further assistance, please feel free to contact me at your convenience.

Patrick O’Donnell, CIEC Managing Member


Martin County Engineering Department Limited Environmental Sampling Four Outdoor Locations Project No. 160185 Martin County, FL Page 3 of 11

6278 North Federal Highway #450, Fort Lauderdale, FL 33308 | Phone 954.978.8839 | Fax 954.973.0530

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INTRODUCTION

The purpose of our environmental sampling was to obtain outdoor ambient air samples in response to a harmful algal bloom (HAB). Client reports that the HAB developed throughout the St. Lucie Estuary, and has accumulated in dead-canals, or enclosed basins. The airborne microalgae of concern in this study are the cyanobacteria (also known as blue-green algae) and microcystins/nodulans (MC) toxins. Droplet formation is a major factor of passive emission of microorganisms from aqueous ecosystems. Drops (6 to 100 µm diameter) are generated from bubble bursting which may occur due to, e.g., waves, rainfall, boat traffic, or supersaturation of gases in the water. Drop formation is therefore a major way for microorganisms to become airborne since the water surface microlayer is enriched with biological material.1 The parameters included in the scope of work included the following: (1) outdoor ambient air temperature, relative humidity and dew point, (2) hydrogen sulfide, (3) respirable particulates, (4) water samples for toxins, (5) total volatile organic compounds, and (6) total algal airborne samples for toxin analysis and fungal spore traps for direct microscopic exam. It should be noted that our testing was performed at approximately week six of the HAB. We conducted a brief interview with Mary Radabaugh and her husband Dutch Radabaugh, the managers of Central Marine. Mary reported that sometime on, or around June 3, 2016 she had initially noticed blue-green algae in the water basin at the facility. Over time, she noticed that the algal blooms continued to worsen. Sometime around June 23 she noticed that the algal bloom manifested a bluish color. Dutch reported that during the time period when the algae had a bluish color they experienced the most significant respiratory irritant affects. Mary reported that on July 1 the algal bloom color transitioned into a mostly brown color and then on July 3 a new green algal bloom was observed. At the time of sampling Outboards Only (also referred to as Outboard Marine) was working with a local contractor that had initiated a pilot demonstration project to reduce algal concentrations in their Marina basin. This pilot demonstration utilized a water pump (or water pump systems) to draw in water and algal matter from the Marina basin, followed by a treatment process and then discharged the treated water back into the basin. 1Applied and Environmental Microbiology, “Airborne Microalgae: Insights, Opportunities, and Challenges”, Sylvie V. M. Tesson, et. al, April, 2016, Vol. 82, Number 7, p. 1978-1991.




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POTENTIAL CONTAMINANT CONCERNS

There is currently no regulatory agency, federal, state or local, which has set standards for acceptable airborne levels of MC or HAB air contaminants. Our water quality references identify blue green algae with Microcystis aeruginosa and a least 3 other cyanobacteria that release exotoxins. Additionally, secondary effects of these algal masses reportedly release methane, hydrogen sulfite and ammonia ions. These constituents are likely associated with the malodors. Exposure routes include skin, ingestion and inhalation2. The airborne exposure would occur by breathing in tiny airborne droplets or mist contaminated with the HAB toxins. Algae are a natural part of our waterways. There are many species of algae, and most do not produce toxins. However, all blue-green algae, or cyanobacteria, can produce skin irritants under certain conditions, and some can produce multiple types of the more harmful toxins. Inhalation of water droplets that have toxins in them may cause allergic-like reactions, runny noses, or sore throats3. Hydrogen sulfide gas causes a wide range of health effects. Individuals are primarily exposed to hydrogen sulfide by breathing it. The effects depend on the concentration and duration of exposure of hydrogen sulfide inhaled. Longer term health effects affect some people who breathed in levels of hydrogen sulfide high enough to become unconscious continue to have headaches and poor attention span, memory, and motor function after waking up. Problems with the cardiovascular system have also been reported at exposures above permissible exposure limits5. People who have asthma may be more sensitive to hydrogen sulfide exposure. The National Institute of Occupational Safety and Health (NIOSH) have established an exposure guideline of 10 ppm as a 10-minute ceiling concentration.

Total volatile organic compound (TVOC) sampling for exposure to chemical contaminants is useful when there is no specific chemical source such as typically found in industrial settings. However, there is insufficient evidence that TVOC measurements can be used to predict health or comfort effects because odor and irritation responses to organic compounds are highly variable between individuals.

SAMPLING METHODOLOGY The Enviro Team North America scope of work for this project was developed with assistance from Dianne Hughes, Senior Ecosystem Specialist, Ecosystem Restoration & Management Division, Martin County FL., Mr. Andrew Reich, MS, MSPH, RRT, Administrator- Public Health Toxicology Section, Bureau of Environmental Health, Florida Department of Health & Lorraine C. Backer, PhD, MPH with the Centers for Disease Control. 2 World Health Organization Guidelines for safe recreational water environments 2003 3 Ohio Department of Health – Harmful Algal Blooms 5 OSHAFactSheet - Hydrogen Sulfide - DSG 10/2005




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• Air collection for Algal Taxomy analysis was collected at four outdoor locations using a

Buck Bio-Aire air pump calibrated to 15 liters per minute (LPM) and Zefon Air-O-Cell spore trap cassettes containing a slide coated with a sticky transparent acrylic substrate to concentrate particles in a longitudinal trace for optical microscopy analysis. Four samples were collected and retained dry in the cassette. Our initial discussions with Green Water Laboratories indicated that there was a possibility of the algal particles becoming desiccated after the collection process which could make the analysis difficult. To address this concern, we also obtained four additional samples as a backup in case the dry samples were compromised. We placed the backup cassettes in a 100 ML sealed container inoculated with a small amount of sterile saline water. However upon examination in the lab, Green Water Laboratories found the dry sample condition satisfactory.

• Air Collection for Algal Toxin analysis was collected at four outdoor locations using Gast air pumps. Two pumps were calibrated to 20 LPM and two calibrated to 25 LPM (6 hour sample) and 37mm 0.8 micron Mixed Cellulose Ester (MCE) cassettes. Samples were prepared & analyzed by Green Water Laboratories via enzyme linked immunosorbent assay (ELISA) method based on L.C. Backer Study – Toxicon 55 (2010) 909-921.

• Air samples for Hydrogen Sulfide (H2S) were procured with a Jerome Hydrogen Sulfide Analyzer model 631-X with 30 minute averages at a total four outdoor locations.

• Air samples for respirable particles were procured with a Fluke model 983 laser particle counter at four locations.

• Air samples for air temperature, relative humidity and dew point were obtained at four locations with a Fluke model 971 digital hygrometer.

• Water sample analyses for Algal Toxin analysis were collected at two locations using a 500 ML sample container. Samples were analyzed via by Green Water Laboratories via ELISA method.

• Air samples for total volatile organic compounds (TVOC) were collected at four locations with a ppbRAE model PGM-7240 VOC Monitor.

The Florida Department of Health (FL DOH) Public Health Toxicology Section recommended the following protocol for sampling hydrogen sulfide in air: 1) report 30 minute averages for testing lasting less than a day or 2) report levels every 30 minutes for testing lasting more than a day. FL DOH further commented that “since there is no standard for outdoor air, we will probably compare the results directly to a sentential study of asthmatics breathing 2,000 ppb H2S for 30 minutes who reported headaches and nose/throat irritation.” All water samples were analyzed by Green Water Laboratories located in Palatka, Florida and air samples analyzed at their lab in Gainesville Florida. Established in 2001, GreenWater Laboratories/CyanoLab is the only private full-service laboratory of its kind in the United States, with a strict focus on cyanobacteria/algae and the toxins they produce.




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RESULTS AND DISCUSSION Airborne microalgae are recognized as allergens and antigens. They are the cause of severe medical issues, including respiratory allergies (e.g., hay fevers), asthmatic attacks, dermatitis and skin lesions, rhinitis, and disturbances in lymphatic systems or vital organs. Alive or not, inhaled airborne microalgae are potentially harmful to animals and humans. Their small size facilitates their inhalation and deposition in the respiratory tract. Secondary metabolites produced by certain microalgae are the causes of further human illnesses (e.g., aerosolized algal toxins). The sensitivity of the target can increase when coupled with high temperature or pollutant concentrations.1 Deposition and subsequent colonization have an impact on the environment. Depending on their ecological strategy, viable deposited microalgae can form seed banks or can proliferate rapidly in a suitable environment, colonizing empty niches, increasing community diversity, or supporting the development of organisms in pioneer environments. Certain microalgae can be harmful, forming blooms that cause public health, economic, and recreational issues. The settlement of airborne microalgae in aquatic/terrestrial environments constitutes a threat for environmental, economic, and sanitation issues. Harmful and noxious microalgae are able to produce toxins and extracellular compounds that are accumulated in the water column, causing recreational disturbances (e.g., skin irritations or change in the water color) and deteriorating water supplies.1 The results of the site surveys conducted on July 11, 2016 are summarized in the Tables below. Table 1

Location Temp (F)

Relative Humidity

(%)

Dew Point (F)

Hydrogen Sulfide (ppm)

TVOC (ppb)

Comments

Central Marine 93 55 74 0.033-0.081 ND

0.081-0.113 Agitating Water 2.5’ above water

0.21-1.39 Agitating Water on Floating Dock 1’ above water

Outboards Only 93 51 71 0.19-0.32 ND

0.11-0.45 4.5’ above water

1.3-4.3 40 feet From pilot treatment

discharge hose

Residence 1976 NE River Ct

Jenson Beach 88 65

75 0.001-0.11

ND

6’ from water fish kill observed

0.006-0.013

Dock Basin

762 NW Sunset Drive 86 80

78 0.11-0.29 ND

2.5’ above water




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Table 2 – Toxins & Microcystis Cells Location Microcystins

in water (ug/L)

Microcystins in aira & b (ng/m3)

Microcystis Cells

in air*

Comments

Central Marine 33100 0.6361a ND

Outboards Only Not collected 0.0311a ND

Residence 1976 NE River Ct

Jenson Beach Not collected 0.0132b

ND

Dock Basin

762 NW Sunset Drive 13700

0.0813b ND

a -Total sample 9,000L+ of air (360 minutes @ 25 lpm) b -Total sample 7,200L+ of air (360 minutes @ 20 lpm - Note lower flow rate was because of the pumps capability) *Total sample 150L of air (10 minutes @ 15 lpm) ND – None Detected Airborne Microcystin cells were not identified in the short term samples (150L). Airborne long term air samples (9,000L & 7,200L respectively) - Microcystin was detected and quantitated at (ng/m3) levels. Water samples levels are 100-1000 times higher than the recreational guidance levels for microcystins in many states (6-10 ppb). At this level, microcystins are considered to present a significant risk and contact with the bloom should be avoided. Microcystin levels in the algal samples present a significant health threat upon direct exposure or ingestion.

Microcystis sp. colonies at 40x Microcystis sp. at 400x

The airborne sampling for total volatile organic compounds (TVOC’s) did not identify any detectable concentrations.




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The airborne concentrations of hydrogen sulfide were generally below 1 ppm if the algal matter was undisturbed. With agitation and disturbance of the algal matter, the hydrogen sulfide levels exceed 1 ppm. The levels from the treatment pilot project were measured at 4.3 ppm approximately 40 feet away from the water discharge point. This data indicates the potential for significant levels of hydrogen sulfide from HAB’s if disturbed and aerosolized which can occur with wind/wave action or actions such as boat movement. Table 3 - Respirable particle concentrations Location 0.3 um 0.5 um 1.0 um 2.0 um 5.0 um 10 um Comments Central Marine 55,369 10,764 5,138 2,420 123 1 Outboards Only 36,921 14,254 7,337 3,535 293 12 142,511 52,832 18,112 7,554 929 40 By Water Pump

Discharge 23,429 11,572 6,073 2,977 440 31 Upwind of Water

Pump 1976 NE River Ct 43,286 12,033 5,946 2,451 140 2 762 NW Sunset Dr 23,884 9,042 4,130 1,719 100 3 Respirable particle concentrations indicate decreasing numbers with increase in particle size which we would expect in an outdoor environment and most indoor environments. At the Outboards Only location a comparative analysis of upwind conditions and conditions measured near the water pump discharge demonstrate higher concentrations associated with the water discharge process which would be expected. Weather conditions obtained from Stuart Airport METAR data. Note that no rain occurred at any site locations during sampling procedure.

Time of

day Temp F Relative Humidity

Dew Point

Wind Direction

Wind Speed

9:47 AM 90° 62% 75° SSE 9 mph 10:47 AM 90° 62% 75° SSE 12 mph 11:47 AM 90° 66% 77° ESE 14 mph 12:47 PM 93° 59% 77° E 12 mph 1:47 PM 93° 56% 75° E 14 mph 2:47 PM 91° 59% 75° ESE 14 mph 3:47 PM 91° 63% 77° ESE 16 mph 4:47 PM 91° 59% 75° ESE 14 mph 5:57 PM 88° 70% 77° ESE 14 mph




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CONCLUSIONS AND RECOMMENDATIONS Based on the results of the data obtained from the limited environmental sampling obtained on July 11, 2016 the following conclusions and recommendations are provided: CONCLUSIONS

(1) Hydrogen Sulfide - Levels were discernable at levels which may be objectionable for individuals with respiratory conditions and asthmatics.

(2) Water Toxins – Microcystin/nodularins levels in the algal samples present a significant health threat upon direct exposure or ingestion.

(3) Airborne Toxins - Microcystin/nodularins levels in the air were identified, however in absence of no set standards by any federal, state or local regulatory agency, we defer to our public health authorities for inhalation avoidance or inhalation risk concerns.

(4) Particulates - Measured respirable particle concentrations in our experience are within an expected range however increased particle concentrations measured by water pump discharge suggest that further study should be performed to determine what impact this may have on human, animal exposure and/or the environment.

(5) Volatile Organic Compounds- The total volatile organic compounds (TVOC’s) were Not Detectable.

HEALTH RELATED RECOMMENDATIONS FROM PUBLIC HEALTH AUTHORITIES4

• To protect yourself, your family, and your pets from exposure to cyanobacterial HABs (CyanoHABs):

• Avoid areas with visible algal concentrations and/or algal scums in the sea as well as on the shore. Direct contact and swallowing appreciable amounts are associated with the highest health risk.

• Don't swim, water ski, or boat in areas where the water is discolored or where you see foam, scum, or mats of algae on the water. If you do swim in water that might have a CyanoHAB, rinse off with fresh water as soon as possible.

• On the beach, avoid sitting downwind of any and of the material drying on the shore, which could form an aerosol can be inhaled particularly in areas with Gymnodinium breve blooms.

• Don't let pets or livestock swim in or drink from areas where the water is discolored or where you see foam, scum, or mats of algae on the water. If pets (especially dogs) swim in scummy water, rinse them off immediately—do not let them lick the algae (and toxins) off their fur.

• Respect any water-body closures announced by local public health authorities. • If any health effects are subsequently experienced in whatever the nature of the exposure,

seek medical advice.

4 Ref: World Health Organization, United States Centers for Disease Control, and Florida Department of Health:




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Limitations: This report was prepared in a manner consistent with that level of care and skill ordinarily exercised by members of the profession currently practicing under similar conditions subject to the time limits and financial and physical constraints applicable to the services requested. This report was prepared for the exclusive use of the client and assigned agents and is not intended for any other purposes. Our report is based on the information available to us at the time of our investigation and limited in scope to the stated purpose and/or the areas inspected. Due to the limited ambient air sampling performed, the data may or may not be statistically valid, therefore the opinions rendered based on limited data may or may not be representative of the actual conditions. Other conditions elsewhere in the subject building(s) may differ from those in the inspected/surveyed locations and such conditions are unknown, may change over time and have not been considered. This report does not claim to identify all potential hazards and/or contaminants that may be present, nor does it imply any medical opinion on the relationship of potential health effects with any reported hazards and/or contaminants. Our opinions are based on our findings and upon professional expertise with no warranty or guarantee implied herein. The data obtained in this report does not establish the habitability of the building(s) nor does it determine if a building is safe or unsafe. Should additional information become available, we reserve the right to determine the impact, if any, of the new information on our opinions, conclusions, and recommendations, and to revise our opinions, conclusions, and recommendations if necessary as warranted by the discovery of the additional information. Enviro Team North America, LLC accepts no responsibility for interpretation of this report by others. Its contents shall not be used or relied upon by other parties without prior written authorization of Enviro Team North America, LLC.


ANALYTICAL RESULTS

Microcystin Report Enviro Team North America

Sample ID Site Collection Date Time Liters Per

Minute Matrix

ATW-1 Central Marine 7/11/16 10:15 Water ATW-2 Dock Basin 762 NW Sunset 7/11/16 12:15 Water 91255 Central Marine 7/11/16 6 hours 25 LPM Air Sample 91238 Outboard Marine 7/11/16 6 hours 25 LPM Air Sample 91203 1976 NE River Court Jenson 7/11/16 6 hours 20 LPM Air Sample 91224 762 NW Sunset 7/11/16 6 hours 20 LPM Air Sample

Dominant Genera

The ATW-1 & -2 samples were dominated by the potentially toxigenic (PTOX) cyanobacteria Microcystis sp.

Microcystis sp. colonies at 40x

Microcystis sp. at 400x

Toxins – microcystins/nodularins (MC)

Sample Preparation

Water

The samples were diluted 100x (2 mL into 200 mL) and ultra-sonicated to lyse cells and release toxins. Additional dilutions were conducted to achieve absorbance values within range of the standard curve and allow for quantitation.

Air Filters

The filters were removed from the filter cassettes and cut into small pieces. The filters were placed in 15 mL glass vials and 75% acidified methanol was added. Samples were vortexed continuously for 1 minute, manually mixed using a stirrer and sonicated for 25 minutes in a sonicating bath. The samples were centrifuged for 10 minutes at 3,000 RPM and supernatants retained. The pellets were rinsed with 5 mLs of 75% acidified methanol and the pooled supernatants were subjected to N2 and heat (60°C) to remove the methanol. The samples were diluted with DI water prior to Strata X SPE for sample clean up. The samples were reconstituted in DI water (0.5 mL) and analyzed using ELISA.

Analytical Methodology

MC

The Adda (Abraxis) microcystins enzyme linked immunosorbent assay (ELISA) was utilized for the quantitative and sensitive congener-independent detection of MCs. The current assay is sensitive to down to a LOD/LOQ of 0.15 µg/L for total MCs in water. The average recoveries of laboratory fortified blanks (LFB) spiked with 1 µg/L MCLR were 111% and 114%.

Summary of Results

Water

Sample MC levels (µg/L)

ATW-1 33100

ATW-2 13700

Detection Limit (µg/L) 0.15

Microcystin was detected and quantitated at ppm (mg/L) levels. These levels are 100-1000 times higher than the recreational guidance levels for microcystins in many states (6-10 ppb). At this level, microcystins are considered to present a significant risk and contact with the bloom should be avoided.

Air Filters

Sample MC levels (ng/m3)

91255 0.6361

91238 0.0311

Detection Limit (ng/m3) 0.0083

91203 0.0132

91224 0.0813

Detection Limit (ng/m3) 0.0104

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Enviro Team North America Qualitative Microscopic Analysis Report Prepared: July 14, 2016 Prepared By: GreenWater Laboratories Samples: 4 (Collected on 7/11/2016)

1. Central Marine (22618258) 2. Outboard Marine (22618251) 3. 1976 NE River Court Jensen (22618250) 4. 762 NW Sunset (22618257)

Sample 1: Central Marine (22618258) The glass cover slip was removed from the Air-O-Cell CSI cassette and analyzed directly at 400X using a Nikon Eclipse TE200 Inverted Microscope equipped with phase contrast optics and epi-fluorescence. Microscopic observations revealed that the Central Marine sample contained fungal hyphae (Fig. 1) and fungal spores (Fig. 2) as well as a single pollen grain (Fig. 3), one green algal unicell (Chlorophyta; Fig. 4) and one empty pennate diatom frustule (Bacillariophyceae; Fig. 5). No Microcystis cells or colonies were observed in the sample.

Fig. 1 fungal hypha 400X (scale bar = 10 μm)

Fig. 2 fungal spores 400X (scale bar = 10 μm)

Fig. 3 pollen grain 400X (scale bar = 10 μm)

Fig. 4 chlorophyte unicell 400X (scale bar = 10 μm)

Fig. 5 empty diatom frustule 400X (scale bar = 10 μm)

Sample 2: Outboard Marine (22618251) The glass cover slip was removed from the Air-O-Cell CSI cassette and analyzed directly at 400X using a Nikon Eclipse TE200 Inverted Microscope equipped with phase contrast optics and epi-fluorescence. Microscopic observations revealed that the Outboard Marine sample contained fungal spores (Fig. 6) and fungal hyphae (Fig. 7) as well as a single colony of the green alga Pseudodidymocystis (Fig. 8) and one dividing blue-green algal unicell (Cyanobacteria; Fig. 9). No Microcystis cells or colonies were observed in the sample.

Fig. 6 fungal spore 400X (scale bar = 10 μm)


Fig. 8 Pseudodidymocystis 400X (scale bar = 10 μm)

Fig. 9 dividing cyanophyte unicell 400X (scale bar = 10 μm)

Sample 3: NE River Court Jensen (22618250) The glass cover slip was removed from the Air-O-Cell CSI cassette and analyzed directly at 400X using a Nikon Eclipse TE200 Inverted Microscope equipped with phase contrast optics and epi-fluorescence. Microscopic observations revealed that the NE River Court Jensen sample contained relatively less fungal hyphae (Fig. 10) and fungal spores (Fig. 11) than other samples. No Microcystis cells or colonies were observed in the sample.


Fig. 11 fungal spore 400X (scale bar = 10 μm)

Sample 4: 762 NW Sunset (22618257) The glass cover slip was removed from the Air-O-Cell CSI cassette and analyzed directly at 400X using a Nikon Eclipse TE200 Inverted Microscope equipped with phase contrast optics and epi-fluorescence. Microscopic observations revealed that the 762 NW Sunset sample contained a large number and variety of fungal spores (Figs. 12-14) than was found in the other samples. Also observed was a fungal thallus (Fig. 15) and butterfly wing scales (Fig. 16). No Microcystis cells or colonies were observed in the sample.



Fig. 14 possible fungal spores 400X (scale bar = 10 μm)

Fig. 15 fungal thallus 400X (scale bar = 10 μm)

Fig. 16 butterfly wing scale 400X (scale bar = 10 μm)

GEOGRAPHICAL SITE SAMPLE LOCATIONS

Enviro Team North America 6278 North Federal Highway, #450, Fort Lauderdale, FL 33308 (954) 978-8839 Project #160185

Sample Site/Locations Page 1

Central Marine



Outboards Only



1976 NE River Ct



762 NW Sunset Dr.

SITE PHOTOS


Photos Page 1

Central Marine

Central Marine - agitating the algal matter


Photos Page 2

Outboards Only

Outboards Only - downstream water discharge


Photos Page 3

NE River Court

Basin on NW Sunset Drive

TIMELINE IMAGES Courtesy of Mary Radabaugh Central Marine


Photos Courtesy Mary Radabaugh Central Marine Page 1

June 3, 2016



June 3, 2016



June 21, 2016



June 22, 2016



June 23, 2016



June 27, 2016



June 27, 2016



June 27, 2016



June 27, 2016



June 27, 2016



June 27, 2016



June 28, 2016



June 28, 2016



June 28, 2016



June 29, 2016



July 1, 2016

July 1, 2016



July 1, 2016



July 3, 2016



July 3, 2016



July 6, 2016



July 6, 2016



July 7, 2016

July 7, 2016



July 8, 2016

July 8, 2016



July 8, 2016

July 8, 2016



July 11, 2016

July 12, 2016



July 13, 2016

July 13, 2016



July 13, 2016



July 15, 2016



July 18, 2016



July 20, 2016

July 20, 2016

APPENDIX A

FLUKE iCalibration Review Coversheet

The calibration referenced below was subcontracted to a supplier who is

currently on the Fluke approved suppliers list. This subcontracting process is in

accordance with the Fluke quality management system registered to ISO

9001:2008 and accredited to ISO 17025:2005. The subcontracted work has been

reviewed and meets the requirements of the PO listed below. The calibration

report includes this supplementary information page and the calibration package

from the subcontracted calibration supplier. Each section may be numbered

separately.

Customer Name: ENVIRO TEAM

RMA Number: 30827338

PO Number: GCS ODONELL

Model Number: FLUKE 983

Serial Number: 9020005

Date of Calibration: 24-AUG-2015

Feel free to contact us if any questions.

Best Regards,

Fluke Corporation

Form 104.3 Calibration Review Coversheet

06/28/2012

Fluke Corporation Telephone Facsimile Internet1420 75 SW Everett, Wa 98203

888-993-5853 425.446.6390 www.fluke.com

BECKMANCOULTER

CERTIFICATE OF CALIBRATION

Instrument Infortnation

Counter Model 983 Station ID GP Depot Temperature 22.0 °C

Part Number 2414646 Calibration Date 2015-Aug-24 Relative Humidity 41.0%

Counter Serial 009020005 Nominal Flow 2.83 LPM

Sensor Model AIR Calibration Due 2016-Aug-24 Laser Current 91.4 mA

Sensor Serial 8123 RA Number 25270821 (Reference Only)Procedure C088149-1 REV E

Performance InformationTest Result Criteria

0.03 m'

Pass/FailTest Name

ISO-21501 Flow

Noise

Peak to Valley

False Count Rate

2.840 LPM

10.0 mV

1:1

0.0 particles in 0.03 m'

±5%

Reference Only

N/A

2.0 particles in

Pass

N/A

N/A

Pass

Calibration EquipmentModel Serial Cal Due DateType

Flow Meter

Thermometer

DMM

TSI

Fluke 971

Fluke 196B

40430740003

93491349

DM9540221

2016-Feb-27

2015-Sep-12

2015-Oct-28

Calibration InformationChannel Size (um) Cal (mV) Threshold (mV) Particle Size (urn) Mfq Lot Number Expiration Date

1 0.3 26.34 29.41 0.296 Thermo 41479 2015-Dec-30

2 0.5 193.66 187.0 0.508 Thermo 44115 2018-Jan-31

3 1.0 670.7 653.48 1.019 Thermo 41321 2015-Oct-31

4 2.0 1501.61 1502.25 1.999 Thermo 41569 2016-Jan-30

5 5.0 2546.11 2547.6 4.993 Thermo 42613 2016-Oct-31

6 10.0 3367.76 3324.09 10.3 Thermo 44199 2018-Feb-28

Standard Calibration

009020005.08.24.2015 , Calibrator : Lawrence Meyer

Reproduction of this Certificate except in full is strictly forbidden without the written approval of Beckman Coulter Pg 1 of 2

BECKMANCOULTER

SUPPLEMENTAL DATA

Instrument Information

Counter Model 983 Station ID GP Depot Temperature 22.0 °C

Part Number 2414646 Calibration Date 2015-Aug-24 Relative Humidity 41.0 %

Counter Serial 009020005 Calibration Due 2016-Aug-24 Nominal Flow 2.83 LPM

Sensor Model AIR RA Number 25270821 Laser Current 91.4 mA

Sensor Serial 8123 Cal Voltage 0.9 V

Procedure C088149-1 REV E

Supplemental Information

Standard Calibration

009020005.08.24.2015

Reproduction of this Certificate except in full is strictly forbidden without the written approval of Beckman Coulter P9 2 of 2

Channel Information

Channel Size Calibrated (mV1 Pulse (mV) As Received (mV) As Received Size Deviation(%) Criteria(%1 Pass/Fail

1 0.3 29.41 26.34 26.34 0.296 -1.33 15.00 Pass

2 0.5 187.0 193.66 199.45 0.515 3.00 15.00 Pass

3 1.0 653.48 670.7 680.76 1.03 3.00 15.00 Pass

4 2.0 1502.25 1501.61 1460.22 1.934 -3.30 15.00 Pass

5 5.0 2547.6 2546.11 2551.63 5.013 0.26 15.00 Pass

6 10.0 3324.09 3367.76 3148.82 8.861 -11.39 15.00 Pass

BECKMANCOULTER

AS RECEIVED DATA

Instrument InformationCounter Model 983 Station ID GP Depot Temperature 22.0 °C

Part Number 2414646 Calibration Date 2015-Aug-24 Relative Humidity 41.0%

Counter Serial 009020005 Calibration Due 2016-Aug-24 Nominal Flow 2.83 LPM

Sensor Model AIR RA Number 25270821 Laser Current 91.4 mA

Sensor Serial 8123 Procedure C088149-1 REV E

&RtniMgEnfr.ffnalW.Test Name

As Received

As Calibrated

Deviation (%)

Criteria (%1 Pass/Fail

Flow

2.805

2.840 -1.23

±10% Pass

Noise

9.3

10.0 -7

N/A

Laser Current

90.8

91.4 -0.66

N/A

Zero Count

PASS

N/A

QUALITY SYSTEMS LAB Certificate No ET018:99,163

AN ISO 17025 ACCREDITED LAB

CERTIFICATE OF CALIBRATION

Performed for: Test Results:

PassEnviro Team6278 N Federal Hwy, #450Ft. Lauderdale, FL 33308

Instrument Details:

Calibration Details:Description: Temperature Humidity Meter Procedure Used: GIDEP/MFQ's/QSL ProcedureManufacturer: Fluke Temp: 74°FModel Number: 971 Humidity: 41 % RHSerial Number: 99440789 Calibrated: 07/31/2015Equipment ID: N/A Frequency: AnnuallyLocation: Calibrated by: Z - Jorge RamirezPO Number: Cornments:

Quality Systems Laboratory (QSL) certifies that the above listed instrument meets or exceeds all specifications as stated in the referencedprocedure unless otherwise noted in the Calibration Results or Comments. It has been calibrated using measurement standards traceable to theNational Institute of Standards and Technology (NIST), or to NIST accepted intrinsic standards of measurement, or derived by the ratio type ofself-calibration techniques. This calibration complies with ISO/IEC 17025:2005. The test results published in this report were obtained usingequipment capable of producing results that are traceable through NIST to the International System of Units (SI). If applicable, QSL follows the USPharmacopeia 37-NF32 Standard, chapter USPC Balance (41), and Weighing on an Analytical Balance (1251) to test Analytical Balances.

Uncertainty evaluation includes the instrument under test and is calculated in accordance with the ISO "Guide to the Expression of Uncertainty inMeasurement". The uncertainty represents an expanded uncertainty using a coverage factor k =2 to approximate a 95% confidence level. Whenstatements of compliance with a specification are made, the measurement uncertainty of the calibration result is accounted for in its entirety.

QSL's responsibility shall in no event, nor for any cause whatsoever, exceed the purchase price of this certification. This report may not bereproduced, except in full, unless permission for the publication of an approved abstract is obtained in writing from the calibration organizationissuing this report. Calibration results are only valid for the time of calibration and cannot be guaranteed due to unforeseen causes that may arisepost-calibration. QSL will not be held liable for calibrated equipment performance once it leaves QSL.

Calibration Standards(As Of Cal Entry Date)

Test Instrument ID Description Manufacturer Serial Number Cal Date Due DateQSL003 Thermometer, Chub-E4 Hart Scientific A65172 10/30/2014 10/30/2015QSL057 Humidity Generator Rense 059973 1/21/2015 1/21/2016QSL062 Platinum Resistance Thermometer Hart Scientific 772740 2/25/2015 2/25/2016

Calibration Results

Group # 1 Range Acc % 0.0000Group Name Temperature Calibration Reading Acc ()/0 0.0000Stated Accy Plus / Minus Plus/Minus 0.5

Nom In Val / In Val In Type Out Val Out Type Fnd As Lft As Low High Pass/Fail150 / 15 1 °C 15.1 °C 15.3 15.3 14.6 15.6 Pass

2a0/2t0 °C 21.0 °C 20.6 20.6 20.5 21.5 Pass

3a0/30.8 °C 30.8 °C 30.5 30.5 30.3 31.3 Pass

Group # 2 Range Acc (1/0 0.0000Group Name Humidity Calibration Reading Acc % 0.0000Stated Accy Plus / Minus Plus/Minus 2.5

7604 NW 6th Ave • Boca Raton, FL 33487 • TEL: 561.999.1896 • FAX: [email protected] • www.QSL-Inc.com

Page 1 of 2

Certificate No: ET018:99,163

Nom In Val/ In Val In Type Out Val Out Type Fnd As Lft As Low Hiqh Pass/Fail

20.0 / 19.1 %RH 19.1 °/0RH 20.3 20.3 16.6 21.6 Pass

50.0 / 49.5 %RH 49.5 %RH 51.1 51.1 47.0 52.0 Pass

80.0 / 77.2 %RH 77.2 %RH 75.4 75.4 74.7 79.7 Pass

Laboratory Manager: Pablo Millan Quality Manager: Catherine SturgisSignature: Vag° Wan Signature: C414444 5/toli4

7604 NW 6th Ave • Boca Raton, FL 33487 • TEL. 561.999.1896 • FAX: [email protected] • www.QSL-Inc.com

Page 2 of 2

Documents

Limited Environmental Sampling - Martin