SAMPLING FOR CONTAMINANTS OF BIOLOGICAL ORIGIN PRESENTED BY

TOXIC MOLD! OH MY!

BIOLOGICAL CONTAMINANTS DEFINING THE ISSUE

BIOLOGICAL CONTAMINATION As defined by ACGIH Aerosols, gases and vapors of biological origin of a type and concentration likely to cause disease or predispose persons to adverse health effects. Inappropriate levels indoors of bioaerosols typically found outdoors. Biological growth indoors of particles that may become airborne and have an adverse effect on exposed individuals.

BIOLOGICAL CONTAMINANTS AEROSOLS Airborne particles of biological origin bacteria, fungi, pollen, viruses and their by-products endotoxins and mycotoxins and other fragments insect parts and excreta, skin scales, hair

BIOLOGICAL CONTAMINANTS VAPORS Fungi and bacteria in indoor environments produce microbial volatile organic compounds or MVOCs as a by-product of their metabolism. The odors of MVOCs are good indicators of microbial growth even when growth is not visible. Knowledge is limited. MVOC research will attempt to answer the questions: What role do MVOCs play in health effects? Do certain microorganisms produce a typical MVOC fingerprint?

BACTERIA FOUND IN THE WORKPLACE Legionella bacteria can grow in water systems including cooling towers and air conditioners. Pseudomonas bacteria can grow in water- based metal working fluids. Staphylococcus aureus including the methicillin-resistant superbug (MRSA) can be transmitted by direct skin-to-skin contact with infected individuals.

BACTERIA BY-PRODUCTS Gram-negative bacteria may contain harmful substances in their outer membranes called endotoxins. Endotoxins are referred to as pyrogens as they will induce fever. They will also cause respiratory distress and even death at high levels. In the workplace, the most common route of endotoxin exposure is through inhalation of aerosolized bacteria including Pseudomonas in metalworking fluids.

ENDOTOXIN EXPOSURES WORKPLACE Can be found in sewage treatment plants, cotton textile mills, fiberglass production plants, poultry/swine facilities, and in industries using metal-working fluids. OTHER Have been found in air conditioning units, spa water, and swimming pools. They can also be found on water damaged material following a water intrusion event.

FUNGI THE USUAL SUSPECT IN IAQ Primary biological contaminant implicated in indoor air complaints. Fungi found most often in the indoor environment include Penicillium, Aspergillus, and Cladosporium. Described as saprophytic in that they can grow on any nonliving organic material if adequate moisture is present.

PRODUCES MYCOTOXINS Mycotoxins are natural by-products of fungal metabolism. They are produced by some species of Aspergillus (versicolor), Fusarium (moniliforme), and Stachybotrys (chartarum). Chemical structures and health effects of mycotoxins are quite diverse.

MYCOTOXIN EXPOSURES WORKPLACE Inhalation of aerosolized fungal spores or other fungal structures in water- damaged building materials. OTHER Ingestion of moldy food products by animals and people. Aflatoxin and trichothecenes have been found in mold- contaminated animal feed and cereal grains.

BIOLOGICAL CONTAMINANTS WHO’S EXPOSED

WORKERS POTENTIALLY EXPOSED TO BIOAEROSOLS Agricultural Workers Grain Handlers Tobacco and Cotton Handlers Farmers Livestock Producers Food handlers and processors Meat packing plants Poultry processors

WORKERS POTENTIALLY EXPOSED TO BIOAEROSOLS Industrial workers Pulp and Paper Mills Textile Mills Wastewater and Sewage Treatment Plants Machinists Industries with cooling towers

WORKERS POTENTIALLY EXPOSED TO BIOAERSOLS Healthcare workers Military personnel Construction and maintenance personnel including remediation workers Office workers in humidified indoor air

BIOLOGICAL CONTAMINANTS HEALTH EFFECTS

SKIN INFECTIONS Staph bacteria, including MRSA, can cause skin infections that look like a pimple or boil. Serious cases can lead to bloodstream infections or pneumonia. Mold may also induce skin infections like ringworm or rashes.

EYE, NOSE, THROAT IRRITATION ACGIH reports that most health complaints in indoor environments are due to eye, nose and throat irritation, headache and fatigue from unknown causes. Sick Building Syndrome

INHALATION FEVERS HUMIDIFIER FEVER Flu-like symptoms that arise 4-8 hours after exposure and subside within 24 hours. Possibly related to endotoxins. PONTIAC FEVER A self-limited, flu-like illness caused by contamination of water systems with Legionella bacteria.

HYPERSENSITIVITY DISEASES Result from exposure to specific antigens in the environment that trigger an immunological response. Dust-mite and animal allergens are common causes in residences.

HYPERSENSITIVITY DISEASES IN WORKPLACE Hypersensitivity pneumonitis characterized by acute, recurrent pneumonia with fever, cough, chest tightness with progression of symptoms. Building-related asthma characterized by chest tightness, wheezing, cough, and shortness of breath that is worse on work days and improves on weekends.

INFECTIOUS DISEASES Influenza (H1N1) and SARS-viral illnesses Legionnaires’ Disease-pneumonia caused by Legionella pneumophila bacteria contaminated water sources. Tuberculosis-lung disease caused by Mycobacterium tuberculosis and spread from person to person.

HEALTH EFFECTS FROM MYCOTOXINS Mycotoxins will reduce the effectiveness of the immune system by interfering with or killing macrophages. This results in increased susceptibility to infectious diseases and a reduction in defense against other contaminants.

WIDE-RANGING EFFECTS FROM MYCOTOXINS VASCULAR SYSTEM-increased vascular fragility; hemorrhage DIGESTIVE SYSTEM-vomiting; intestinal hemorrhage; liver effects RESPIRATORY SYSTEM-respiratory distress; bleeding from lungs NERVOUS SYSTEM-tremors; lack of coordination; depression, headache.

BIOLOGICAL CONTAMINANTS CONDUCTING AN INVESTIGATION

ACGIH RECOMMENDS A STEPWISE APPROACH 1. Gather information through occupant interviews, surveys, and building inspections. 2. Formulate a hypothesis on the cause of the complaints/illness by using the information gathered. 3. Test the hypothesis by collecting samples. 4. Make recommendations for controls by using sampling data & professional judgment.

INFORMATION GATHERING OCCUPANT INTERVIEWS Before embarking on a program of air measurements, survey both the area and the people involved in the complaint. Use this time for information gathering. Look around. Listen. Complaint vs non- complaint areas. Date when problem was first noted. Days or times when problem is noted more and less. Seek input so you can formulate a hypothesis on the root cause of the complaints.

INFORMATION GATHERING BUILDING INSPECTION 1. Examine the physical structure, maintenance activities, and occupancy patterns. 2. Look for potential sources of biological contaminants and evidence of water damage.

BUILDING INSPECTION: MOISTURE INDICATORS Water marks on ceiling tiles and other surfaces Visual presence of mold Musty smell of microbial VOCs White, powdery or crystalline substance on the surface of concrete, plaster and masonry which are soluble salts dissolved from the building materials

BUILDING INSPECTION: MOISTURE METERS Used to survey moisture in any non- conductive porous material to which the probes can be applied. Ceiling Tiles Gypsum Board Carpeting Wood Plaster Concrete

MOISTURE METER FROM SKC Operates by measuring the electrical conductance between two probes inserted into the material to be tested. Useful for construction or renovation projects or other situations when the test surface can be punctured by the probes. SKC

DATA INTERPRETATION MOISTURE METERS Moisture levels can be compared from wall to wall to determine where moisture intrusion is occurring. Once the location of the moisture is found, an investigation can be made as to the cause and a control strategy can be developed.

DATA INTERPRETATION MOISTURE The Western Wood Products Association (WWPA) has prepared a technical guide on preventing and controlling mold in lumber. See WWPA recommends that the moisture content of the wood be kept below 20%.

DATA INTERPRETATION MOISTURE Greenguard Environmental Institute (GEI) has received ANSI approval for a standard covering the management of moisture (and mold growth) during building construction. See

BUILDING INSPECTION: HVAC SYSTEM CHECKS OUTDOOR AIR SUPPLY Inadequate amounts of outdoor air often leads to building-related complaints and health-related symptoms. Ensure outdoor air supply meets ASHRAE or other appropriate standards.

BUILDING INSPECTION: HVAC SYSTEM CHECKS LOCATION OF AIR INTAKES Air intakes on rooftops can draw in bioaerosols from cooling towers, sanitary vents, building exhausts and animal waste. Air intakes at street level can draw in moisture, vehicle emissions, and odors.

BUILDING INSPECTION: HVAC SYSTEM CHECKS CONDITION OF AIR FILTERS HVAC filters are not designed to protect equipment or occupants from heavily contaminated air. Filters may themselves promote the growth of microorganisms if they become damp.

BUILDING INSPECTION: HVAC SYSTEM CHECKS SUPPLY AIR Ductwork should not be coated with excessive debris. Dirt mixed with moisture can support microbial growth. Ensure that cold air leaving a diffuser does not produce condensation and the potential for microbial growth.

CHOOSING A SAMPLING METHOD TO TEST THE HYPOTHESES Data Interpretation Tips

SAMPLING FOR BIOLOGICAL CONTAMINANTS WHY: To test your hypothesis on the cause of the problem To positively confirm the absence/presence of contaminant To identify the type of microbe (genus/species) To confirm the effectiveness of decontamination. HOW: Bulk samples Surface samples Air samples Followed by analysis at a qualified environmental microbiology laboratory.

BULK SAMPLING Portions of materials in the building can be tested for mold or other biological contaminants. Typical test materials include sections of wallboard/wallpaper, carpet pieces, return-air filters, duct lining, and settled dust. These are very useful as air sampling may miss some contaminants due to temporal variations.

BULK SAMPLING Portions of the test material are typically placed in a sealable plastic bag for transport to the lab. In some cases, sterile jars for dry items or sterile bottles for water or metalworking fluid samples may be required. Settled dust can be collected using conventional vacuum cleaners and a new vacuum cleaner bag for each sample.

SURFACE SAMPLING MICROVACUUM CASSETTES Carpeting is an effective reservoir for fungal spores and sampling this surface can reveal the history of mold in the building. To sample fungal spores in carpeting, a vacuum- style cassette is available with a 0.45 um polycarbonate filter loaded into a 3-piece styrene cassette with 2-inch tubing nozzle. Sample at flows up to 16 L/min to vacuum a defined area. Work the inlet tube as deep as possible into the carpeting to collect a good sample of the dust.

CARPET SAMPLING KIT SKC

DATA INTERPRETATION CARPET SAMPLES An 2003 AIHCE paper by MidWest Microbiology gave some numerical guidelines for fungal spores on surfaces like carpet using microvacuum cassettes: Normal- <5,000/1000 cm 2 Borderline- 25,000/1000 cm 2 Elevated- >75,000/1000 cm 2

SURFACE SAMPLING STERILE WIPES A swab or filter wetted with sterile water or wash solution is used to wipe a specified area. Typically, the swab is then used to inoculate an agar plate for growth culture. This technique is often used for MRSA testing. SKC

DATA INTERPRETATION SWAB SAMPLES The November 2001 AIHA Synergist guidelines for fungal spores in swab samples: Normal: <1,500 cfu/cm 2 Probable Contamination: >1,500 cfu/cm 2

SURFACE SAMPLING LIFT TAPE Collected by placing clear adhesive or packing tape or commercially available sampling strips onto a surface and removing it with slow, steady force Following collection, the tape is attached to glass slides and examined using light microscopy to view mold spores.

SURFACE SAMPLING LIFT TAPE ON A SLIDE Flexible plastic microscopic slides with a sticky adhesive sample area can be used like lift tape. Press on the test surface, place the slide in the provided mailer and send to a qualified laboratory. Stick-to-It Slides SKC /9

DATA INTERPRETATION LIFT TAPE SAMPLES The November 2001 AIHA Synergist guidelines for fungal spores in tape samples: Normal: No significant fungal material or biomass; 1-5% spores Probable Contamination: % spores

AIR SAMPLING: WHY AND HOW Like with chemical sampling, air sampling for biological contaminants is done for the purpose of evaluating actual human exposures. Air Sampling for Bioaerosols involves the use of: Impactors Filters or Liquid-based (impinger-type) devices

AIR SAMPLING SPORETRAP CASSETTES Easy, inexpensive screening device. Use with a pump at L/min for up to 10 minutes. Spores impact onto a microscopic slide with a sticky surface. Slide is stained and analyzed microscopically.

VERSATRAP CASSETTES SKC /1

AIR SAMPLING PUMPS FOR USE WITH SPORETRAPS Constant flows from L/min User selectable sampling times Lithium-ion battery powered up to 4 hrs Indefinite run time from AC adapter Optional sampling wand SKC

DATA INTERPRETATION SPORE TRAPS Spore trap analysis will provide the total number of spores and the genus of the spores found. This information can be used to compare the complaint area to non- complaint areas of the building and to the outdoors. The genus of the spores should be similar inside and out. The numbers should be lower inside.

DATA INTERPRETATION SPORE TRAPS November 2001 AIHA Synergist guidelines for Air samples: Residential Buildings: Normal: <5,000 spores/m 3 Probable Contamination: >10,000 spores/m 3 Commercial Buildings: Normal: <2,500 spores/m 3 Probable Contamination: >10,000 spores/m 3

AIR SAMPLING VIABLE CASCADE IMPACTOR Specified in NIOSH Methods 0800 and 0801 Used with a pump at 28.3 L/min for typical sample times of 2-5 minutes Bioaerosols impact onto growth medium (agar). Agar plates are shipped to a microbiology laboratory for growth culture. SKC Biostage /11

SKC BIOSTAGE SAMPLER PREPARATION

SKC BIOSTAGE WITH QUICKTAKE 30 PUMP Sample Assurance Tip: Impactor must be cleaned with isopropyl alcohol before each use. Evaluate blank samples of agar plates.

AIR SAMPLING FILTERS Collection of bioaerosols is achieved by passage of air through a porous medium, typically a membrane filter. Polycarbonate, mixed cellulose ester, or polyvinyl chloride filters may be used depending upon the application. Gelatin filters will help to maintain viability by minimizing dehydration of the microbes.

GELATIN FILTERS WITH SKC BUTTON SAMPLER An AIHA Journal article reported that 25-mm filters used with the SKC Button Sampler provided collection efficiencies close to 100% for enumeration of airborne spores. SKC offers sterile gelatin filters in 25 or 37-mm diameters. (Cat. No /2). SKC INHALABLE 4 L/min

VIABLE AIR SAMPLING FILTERS Filters, support pads and cassettes should be sterile. Samples are collected with a portable pump at 1-4 L/min for 5-30 minutes. After sampling, the material collected on the filters is inoculated onto agar plates.

AIR SAMPLING COLLECTION INTO LIQUID Bioaerosols are often collected into a liquid medium, typically a dilute buffer solution or mineral oil. Portions of the collection liquid can be placed onto nutrient agar and incubated or analyzed using other methods. Biosampler SKC ADVANTAGE: 8-hr sampling

OPERATION OF THE BIOSAMPLER Particle collection is achieved by drawing the bioaerosols through 3 nozzles directed at an angle toward the inner sampler wall. The collection liquid swirls upward on the inner surface and removes the collected particles.

ANALYSIS OPTIONS Growth culture Microscopic Bioassay Immunoassay PCR (using water as collection media )

2 COMMON QUESTIONS 1. Why is the Sonic Flow Pump required for the Biosampler? Can we use the QT30 instead? 2. Can the Biosampler be used for PCR (DNA based) analysis?

DATA INTERPRETATION VIABLE AIR SAMPLES November 2001 AIHA Synergist guidelines: Residential Buildings: Normal: <500 cfu/m 3 Probable Contamination: >1,000 cfu/m 3 Commercial Buildings: Normal: <250 cfu/m 3 Probable Contamination: >1,000 cfu/m 3

AIR SAMPLING FOR MVOCs Sorbent Tubes/Pumps Stainless Steel Canisters

AIR SAMPLING FOR MVOCs Photoionization Detectors (ppb RAE) Man’s best friend See

DATA INTERPRETATION GENERAL GUIDELINES

WHAT ABOUT OELs? Few established guidelines/standards for biological contaminants except for wood dust, cotton dust, etc. ACGIH indicates that a “general exposure limit for concentrations or countable biological agents is not scientifically supportable”.

1. Biological contamination typically is a complex mixture of many types of microorganisms. 2. The health effects for various microorganisms vary greatly between individuals. 3. There is no single sampling method that is appropriate for all types of biological contamination. 4. There is insufficient scientific evidence to support a dose-response relationship from which an exposure standard could be derived.

ACGIH RECOMMENDS “Gather the best data possible and use knowledge, experience, expert opinion, logic, and common sense to interpret information, design control and remediation strategies.”

GENERAL GUIDELINES FOR DATA INTERPRETATION 1. Fungi found in indoor air should be the same species as that found in outdoor air, but the levels should be lower. 2. If there is a dominant fungal species in indoor air that is not present in outdoor air, it is probably growing from biological contamination within the building and is reducing air quality.

GENERAL GUIDELINES FOR DATA INTERPRETATION 3. Comparisons of indoor/outdoor air or problem/non-problem areas should be made at the species level, not just at the genus level. 4. Some species may be considered indicator organisms indicating specific problems such as E. Coli as an indicator of sewage contamination.

GENERAL GUIDELINES FOR DATA INTERPRETATION 5. In order to compare sampling results from indoor/outdoor areas or from different zones, an identical sampling protocol for each zone including the sample media, sampling duration, sampler type and laboratory analysis is required.

HOW DO YOU KNOW WHEN YOU HAVE FINISHED REMEDIATION? 1. Water problem is fixed. 2. Moldy materials removed. 3. Mold similar in type/number inside and outside. 4. No new growth or water damage. 5. No health problems on reentry.

THANK YOU FOR YOUR ATTENTION! YOU ARE NOW AN OFFICIAL SKC MOLDBUSTER! SKC