SIST ISO 16000-19:2014
Indoor air - Part 19: Sampling strategy for moulds
Indoor air - Part 19: Sampling strategy for moulds
This part of ISO 16000 describes the measurement strategy for the detection of fungi in indoor environments.It describes suitable sampling and analysis methods together with a description of the applicability and the interpretation of the measurement results to maximize the comparability of the measured data obtained for a given measurement objective. It does not include details on recording building characteristics or field inspections by qualified professionals which have to take place prior to any microbiological measurement.This part of ISO 16000 is not applicable to a detailed description of the building physics- and building-engineering-related procedures applicable to field inspections. The methods and procedures presented do not allow quantitative exposure assessment with regard to the room occupants.The application of this part of ISO 16000 presupposes the knowledge of ISO 16000-1.
Air intérieur - Partie 19: Stratégie d'échantillonnage des moisissures
L'ISO 16000-19:2012 d�crit la strat�gie de mesurage pour d�tecter les champignons dans les environnements int�rieurs.
L'ISO 16000-19:2012 d�crit des m�thodes d'�chantillonnage et d'analyse appropri�es ainsi que l'applicabilit� et l'interpr�tation des r�sultats de mesurage pour maximiser la comparabilit� des donn�es mesur�es obtenues pour un objectif de mesurage donn�. Elle ne contient pas d'indications d�taill�es concernant l'enregistrement des caract�ristiques du b�timent ou les inspections sur le terrain men�es par des professionnels qualifi�s qui doivent �tre effectu�s pr�alablement � tout mesurage microbiologique.
L'ISO 16000-19:2012 ne s'applique pas � une description d�taill�e des modes op�ratoires relatifs � la physique et au g�nie du b�timent applicables aux inspections sur le terrain. Les m�thodes et les modes op�ratoires pr�sent�s ne permettent pas d'�valuer l'exposition quantitative des occupants de la pi�ce.
L'application de l'ISO 16000-19:2012 pr�suppose que l'on ait pris connaissance de l'ISO 16000‑1.
Notranji zrak - 19. del: Strategija vzorčenja gliv
Ta del ISO 16000 opisuje strategijo merjenja za zaznavanje gliv v zaprtih prostorih. Opisuje primerne metode vzorčenja in analize skupaj z opisom uporabnosti in interpretacije rezultatov meritev za povečanje primerljivosti pridobljenih podatkov meritev za dani cilj meritev. Ne vključuje podrobnosti o beleženju lastnosti stavb ali strokovnem pregledu območja, ki se morata izvesti pred mikrobiološkimi merjenji. Ta del standarda ISO 16000 ne velja za podroben opis postopkov, povezanih z gradbeno fiziko ali gradbeno tehniko, ki se uporabljajo pri pregledu območja. Predstavljene metode in postopki ne omogočajo kvantitativnega ocenjevanja izpostavljenosti oseb v prostoru. Uporaba tega dela ISO 16000 predpostavlja poznavanje ISO 16000-1.
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INTERNATIONAL ISO
STANDARD 16000-19
First edition
2012-06-01
Indoor air —
Part 19:
Sampling strategy for moulds
Air intérieur —
Partie 19: Stratégie d'échantillonnage des moisissures
Reference number
ISO 16000-19:2012(E)
©
ISO 2012
---------------------- Page: 1 ----------------------
ISO 16000-19:2012(E)
COPYRIGHT PROTECTED DOCUMENT
© ISO 2012
All rights reserved. Unless otherwise specified, no part of this publication may be reproduced or utilized in any form or by any means,
electronic or mechanical, including photocopying and microfilm, without permission in writing from either ISO at the address below or
ISO's member body in the country of the requester.
ISO copyright office
Case postale 56 CH-1211 Geneva 20
Tel. + 41 22 749 01 11
Fax + 41 22 749 09 47
E-mail copyright@iso.org
Web www.iso.org
Published in Switzerland
ii © ISO 2012 – All rights reserved
---------------------- Page: 2 ----------------------
ISO 16000-19:2012(E)
Contents Page
Foreword . iv
Introduction . vi
1 Scope . 1
2 Normative references . 1
3 Terms and definitions . 1
4 Properties, origin and occurrence of moulds in indoor environments . 4
5 Sampling and detection methods . 5
6 Measurement strategy . 6
6.1 General aspects . 6
6.2 Selection of appropriate procedure . 9
7 Quality requirements and uncertainty considerations . 17
Annex A (informative) Moisture damage indicators . 18
Annex B (informative) Devices for total spore count and detection of culturable fungi . 19
Annex C (informative) Field inspection report to describe sampling procedure and to document
potential mould damage . 21
Bibliography . 27
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ISO 16000-19:2012(E)
Foreword
ISO (the International Organization for Standardization) is a worldwide federation of national standards bodies
(ISO member bodies). The work of preparing International Standards is normally carried out through ISO
technical committees. Each member body interested in a subject for which a technical committee has been
established has the right to be represented on that committee. International organizations, governmental and
non-governmental, in liaison with ISO, also take part in the work. ISO collaborates closely with the
International Electrotechnical Commission (IEC) on all matters of electrotechnical standardization.
International Standards are drafted in accordance with the rules given in the ISO/IEC Directives, Part 2.
The main task of technical committees is to prepare International Standards. Draft International Standards
adopted by the technical committees are circulated to the member bodies for voting. Publication as an
International Standard requires approval by at least 75 % of the member bodies casting a vote.
Attention is drawn to the possibility that some of the elements of this document may be the subject of patent
rights. ISO shall not be held responsible for identifying any or all such patent rights.
ISO 16000-19 was prepared by Technical Committee ISO/TC 146, Air quality, Subcommittee SC 6, Indoor air.
ISO 16000 consists of the following parts, under the general title Indoor air:
Part 1: General aspects of sampling strategy
Part 2: Sampling strategy for formaldehyde
Part 3: Determination of formaldehyde and other carbonyl compounds in indoor air and test chamber
air — Active sampling method
Part 4: Determination of formaldehyde — Diffusive sampling method
Part 5: Sampling strategy for volatile organic compounds (VOCs)
Part 6: Determination of volatile organic compounds in indoor and test chamber air by active sampling on
®
Tenax TA sorbent, thermal desorption and gas chromatography using MS or MS–FID
Part 7: Sampling strategy for determination of airborne asbestos fibre concentrations
Part 8: Determination of local mean ages of air in buildings for characterizing ventilation conditions
Part 9: Determination of the emission of volatile organic compounds from building products and
furnishing — Emission test chamber method
Part 10: Determination of the emission of volatile organic compounds from building products and
furnishing — Emission test cell method
Part 11: Determination of the emission of volatile organic compounds from building products and
furnishing — Sampling, storage of samples and preparation of test specimens
Part 12: Sampling strategy for polychlorinated biphenyls (PCBs), polychlorinated dibenzo-p-dioxins
(PCDDs), polychlorinated dibenzofurans (PCDFs) and polycyclic aromatic hydrocarbons (PAHs)
Part 13: Determination of total (gas and particle-phase) polychlorinated dioxin-like biphenyls (PCBs) and
polychlorinated dibenzo-p-dioxins/dibenzofurans (PCDDs/PCDFs) — Collection on sorbent-backed filters
iv © ISO 2012 – All rights reserved
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ISO 16000-19:2012(E)
Part 14: Determination of total (gas and particle-phase) polychlorinated dioxin-like biphenyls (PCBs) and
polychlorinated dibenzo-p-dioxins/dibenzofurans (PCDDs/PCDFs) — Extraction, clean-up and analysis by
high-resolution gas chromatography and mass spectrometry
Part 15: Sampling strategy for nitrogen dioxide (NO )
2
Part 16: Detection and enumeration of moulds — Sampling by filtration
Part 17: Detection and enumeration of moulds — Culture-based method
Part 18: Detection and enumeration of moulds — Sampling by impaction
Part 19: Sampling strategy for moulds
Part 23: Performance test for evaluating the reduction of formaldehyde concentrations by sorptive
building materials
Part 24: Performance test for evaluating the reduction of volatile organic compound (except
formaldehyde) concentrations by sorptive building materials
Part 25: Determination of the emission of semi-volatile organic compounds by building products —
Micro-chamber method
Part 26: Sampling strategy for carbon dioxide (CO )
2
Part 28: Determination of odour emissions from building products using test chambers
The following parts are under preparation:
Part 21: Detection and enumeration of moulds — Sampling from materials
Part 27: Determination of settled fibrous dust on surfaces by SEM (scanning electron microscopy) (direct
method)
Part 29: Test methods for VOC detectors
Part 30: Sensory testing of indoor air
Part 31: Measurement of flame retardants and plasticizers based on organophosphorus compounds —
Phosphoric acid ester
Part 32: Investigation of constructions on pollutants and other injurious factors — Inspections
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ISO 16000-19:2012(E)
Introduction
Mould spores and metabolites can be inhaled via the air and cause allergic and irritating reactions and/or
complex symptoms in humans. Moreover, mould growth can be associated with severe odour nuisances. In
[14][18][19]
rare cases, some mould species can cause infections (so-called mycoses) in certain risk groups.
There is sufficient epidemiological evidence that damp and mouldy buildings increase the risk of respiratory
[8]
symptoms, respiratory infections and enhances asthma symptoms of the occupants. In addition, there is
some evidence for increased risk of development of allergic rhinitis and asthma. Furthermore, there is clinical
evidence for rare symptoms like allergic alveolitis, chronic rhinosinusitis and allergic sinusitis. Toxicological
studies in vivo and in vitro show irritating and toxic reactions of microorganisms (including spores, cell
[8]
components and metabolites) from damp buildings.
Growth of microorganisms in damp buildings can lead to increased concentrations of spores, cell fragments,
allergens, mycotoxins, endotoxins, -glucanes and MVOC (microbial volatile organic compounds). From the
studies conducted so far it is not clear which compounds are the causative agents of the health effects
observed. Nevertheless, increased concentrations of each of these compounds are considered a potential
[8][18]
health risk and growth of mould in buildings should, therefore, be avoided.
The prime objective of this part of ISO 16000 is to provide assistance in identifying mould sources in indoor
environments.
vi © ISO 2012 – All rights reserved
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INTERNATIONAL STANDARD ISO 16000-19:2012(E)
Indoor air —
Part 19:
Sampling strategy for moulds
1 Scope
This part of ISO 16000 describes the measurement strategy for the detection of fungi in indoor environments.
It describes suitable sampling and analysis methods together with a description of the applicability and the
interpretation of the measurement results to maximize the comparability of the measured data obtained for a
given measurement objective. It does not include details on recording building characteristics or field
inspections by qualified professionals which have to take place prior to any microbiological measurement.
This part of ISO 16000 is not applicable to a detailed description of the building physics- and building-
engineering-related procedures applicable to field inspections. The methods and procedures presented do not
allow quantitative exposure assessment with regard to the room occupants.
The application of this part of ISO 16000 presupposes the knowledge of ISO 16000-1.
2 Normative references
The following referenced documents are indispensable for the application of this document. For dated
references, only the edition cited applies. For undated references, the latest edition of the referenced
document (including any amendments) applies.
ISO 16000-16, Indoor air — Part 16: Detection and enumeration of moulds — Sampling by filtration
ISO 16000-18, Indoor air — Part 18: Detection and enumeration of moulds — Sampling by impaction
3 Terms and definitions
For the purposes of this document, the following terms and definitions apply.
3.1
pre-existing mouldy condition
desiccated “old” mould growth, where additional biomass growth no longer occurs and the indoor air mould
spore concentration gradually decreases with time
3.2
biological preservation efficiency
capacity of the sampler to maintain the viability of the airborne microorganisms during collection and also to
keep the microbial products intact
[6]
[SOURCE: EN 13098:2000 ]
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ISO 16000-19:2012(E)
NOTE The biological collection efficiency considers the sampling stress occurring during sampling and analysis in
addition to the physical collection efficiency.
3.3
identification of moulds
assignment of moulds to spore types or groups on the basis of defined properties (e.g. morphological,
biochemical, molecular-biological properties)
NOTE The term “differentiation” is frequently used instead of identification. The term “differentiation” is, however,
misleading because the intention is not to merely differentiate the moulds but to identify them, i.e. to assign them, e.g. to
genera or species.
3.4
filamentous fungus
fungus growing in the form of filaments of cells known as hyphae
NOTE 1 Hyphae aggregated in bundles are called mycelia.
NOTE 2 The term “filamentous fungi” differentiates fungi with hyphal growth from yeasts.
3.5
filtration
collection of particles suspended in a gas or liquid by flow through a porous medium
[6]
[SOURCE: EN 13098:2000 ]
NOTE In this part of ISO 16000, filtration is understood as the separation of microorganisms or moulds from a
defined volume of air by means of filters.
3.6
total spore count
number of (culturable and non-culturable) spores that are collected and enumerated under the microscope
NOTE For the term “spores”, see 3.19, Note 2.
3.7
yeast
unicellular fungus that does not normally produce a mycelium and reproduce by budding (budding fungi) as
against moulds, which reproduce by sporulation
3.8
impaction
sampling of particles suspended in air by inertial separation on a solid surface (culture medium or adhesive-
coated slides)
NOTE 1 See 16000-18.
NOTE 2 Sampling is carried out using either round-hole or slit impactors, for instance. As the air passes through the
orifices, it is accelerated and the particles are impacted on the medium located directly behind the nozzles as a result of
their inertia, while the air flows around the culture medium and exits the sampler. Impaction samples are only suitable for
direct analysis without further resuspension of the sample.
3.9
colony forming unit
cfu
air quality unit by which the culturable number of microorganisms is expressed
[6]
[SOURCE: EN 13098:2000 ]
NOTE 1 One colony can originate from one single microorganism, from aggregates of many microorganisms as well as
from one or many microorganisms attached to a particle.
NOTE 2 The number of colonies can depend on the cultivation conditions.
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ISO 16000-19:2012(E)
3.10
colony morphology type
group of colonies which due to their morphological appearance seem to belong to a specific species
3.11
colony count
air quality number of all microorganism colonies visible on a culture medium after incubation under the
selected cultivation conditions
3.12
culturable mould
mould that can be cultured under the selected cultivation conditions
NOTE Parameters governing the culturability are, for instance, the type of culture medium and the incubation
temperature.
3.13
cultivation
growing of microorganisms on culture media
3.14
mycotoxin
secondary metabolites of moulds which are toxic to humans and animals
3.15
mycelium
total of fungal hyphae
3.16
non-culturable mould
mould that cannot be cultured under the selected cultivation conditions
3.17
physical sampling efficiency
capacity of the sampler to collect particles with specific aerodynamic diameters suspended in air
[6]
[SOURCE: EN 13098:2000, modified — "aerodynamic diameters" has replaced "sizes".]
3.18
sampling stress
damage suffered by the microorganisms during sampling (e.g. through mechanical and chemical effects or
through water deprivation)
3.19
mould
filamentous fungi from several taxonomic groups, namely Ascomycetes, Zygomycetes, and their anamorphic
states formerly known as Deuteromycetes or fungi imperfecti
NOTE 1 Taxonomically, moulds do not represent a uniform group.
NOTE 2 Moulds form different types of spores depending on the taxonomic group they belong to, namely
conidiospores (conidia), sporangiospores or ascospores. In practice, all these reproductive stages are summarized under
the term “spores”.
3.20
mould damage
damage caused to building materials and surfaces by mould growth
NOTE Mould damage can result in loss in value, health risks and restrict the occupancy of the affected sites.
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ISO 16000-19:2012(E)
3.21
secondary colony
colony that does not originate from the “primary” sampling of airborne spores but from a spore released from a
colony growing on the agar plates
3.22
secondary contamination
mould contamination of surfaces not caused by mould growth but originating from a (contaminated) primary
source after aerial dispersion
3.23
cut-off value
particle size (aerodynamic diameter) for which the sampling efficiency is 50 %
3.24
total sampling efficiency
product of the physical sampling efficiency and the biological preservation efficiency
[6]
[SOURCE: EN 13098:2000 ]
4 Properties, origin and occurrence of moulds in indoor environments
Moulds are ubiquitous on our planet. They are involved in the decomposition of organic material and,
therefore, play an important role in the natural carbon cycle. Their concentration in the ambient air depends,
inter alia, on location, climate, time of the day and season. Airborne mould concentrations are subject to great
[9][10][11]
variability. This is due to the following reasons.
The mould concentration in local ambient air is mainly determined by the location relative to the respective
mould sources, wind direction and wind force. Mould spores are frequently released by specific sources such
as decaying material. Both natural processes and production processes, such as composting, recycling,
animal production facilities, grain and food processing plants as well as horticulture facilities, can be sources
of mould dispersion.
Sporulation, i.e. the production of mould spores occurs discontinuously. It is governed, inter alia, by the mould
lifecycle phase, the environmental conditions, stress factors, humidity as well as substrate composition and
availability.
Factors governing the dispersion of spores, most of which have aerodynamic diameters in the range of 2 µm
to 40 µm, are mechanically or thermally induced air movements, drying phases (leading e.g. to de-
[12][13][14]
agglomeration of deposited dust) and the capability of air dispersal of the mould spores.
Due to the ubiquitous nature of moulds, it can be assumed that they are always present in indoor air. The
presence of moulds in indoor air can be due to spores originating from ambient air on the one hand and to
recent active mould growth, pre-existing mouldy conditions or mould deposits (settled spores) on the other. To
distinguish between sources, it is, therefore, important to perform ambient air measurements for reference
[14][15]
whenever conducting indoor air measurements for moulds. In addition, the collection of a control sample
from a suitable reference room may be helpful.
Possible causes of indoor mould sources are surface moisture on building materials or moisture in the building
structure, but also rotting food, potted plants, biowaste collection, source separation of waste, deposited dust
due to poor cleaning as well as the keeping of animals in residential settings. Moisture damage can be
attributable to building defects, inappropriate ventilation and heating or unfavourable arrangement of furniture
as well as water damage (e.g. plumbing leaks or flooding events). Elevated mould levels in indoor
environments and the occurrence of certain mould species (see Annex A) are indicative of excessive moisture.
When residential environments or occupational settings are infested with moulds, the mould source shall be
located to be able to plan remedial measures.
4 © ISO 2012 – All rights reserved
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ISO 16000-19:2012(E)
Main factors affecting the intensity of mould growth and the mould species developing are moisture,
temperature, nutrient supply and the pH. If environmental conditions are favourable, a great variety of moulds
can develop. Once environmental conditions become less favourable, the species best adapted to the given
[16]
conditions will predominate.
Mould sources can release spores, mycelial fragments, but also cell components and metabolic products such
as -glucans (polysaccharides contained in the cell wall of fungi), ergosterol (steroid compound contained in
the cell membrane of fungi), toxins and MVOCs (microbial volatile organic compounds such as certain
aldehydes, alcohols, esters, ketones). On cultivation, colonies can grow not only from spores, but also from
mycelial fragments.
The number and airborne dissemination of spores released vary with the type of mould damage. For an
assessment of indoor mould sources, it is, therefore, important to differentiate the individual mould species by
their type of spore dispersal. Experience has shown that even minor mould contamination of materials can
result in elevated indoor air mould levels if the species involved have dry spores with good air dispersal
capabilities (e.g. Penicillium and Aspergillus). By contrast, airborne spore concentrations are much lower
when materials are colonized, for instance, by moulds of the genera Acremonium, Fusarium or the species
Stachybotrys chartarum that have relatively large spores embedded in slimy substances and, therefore, have
poor air dispersal capabilities.
Furthermore, it should be taken into account that mould spores are not necessarily present as individual
spores in the air or settled dust, but also occur in the form of spore aggregates or are particle-borne.
Depending on the analysis method, they are determined individually or as spore aggregate. Materials, indoor
air and house dust contain not only culturable but also non-culturable mould spores, some of which can have
the same allergenic and toxic effects as culturable spores. For this reason, techniques have been developed
that allow the microscopic determination of both culturable and non-culturable moulds.
Mould detection and identification are performed either after cultivation based on morphological criteria,
biochemical reactions and/or molecular techniques or by direct microscopic examination. Identification based
on the morphological structure (macroscopic examination, stereo-microscopy and microscopy) either after
prior cultivation or by direct microscopy is still the most prevalent approach for the detection of moulds.
Besides, there are other analytical methods based on the determination of cell components and metabolites of
[17]
moulds such as -glucans, ergosterol, toxins and MVOCs. The determination of these compounds serves,
however, only as supplementary information.
The sampling methods employed for detection of moulds are determined by the objective of the investigation.
Depending on the sampling method, the moulds suffer a sampling stress during sample collection and
preparation, which can lead to their drying-out or dying. Factors affecting the culturability of mould spores are
their physiological state as well as the culture medium employed. Some mould species cannot be cultured at
all under laboratory conditions.
NOTE The genera Stachybotrys and Chaetomium hardly grow and sporulate only poorly, if at all, on DG18 agar. The
use of this culture medium for culture-based analysis of these genera is therefore not recommended (see ISO 16000-17).
For a literature summary see References [8]–[10], [12], and [14]–[18].
5 Sampling and detection methods
Depending on the objective of the investigation, materials (see ISO 16000-21, in preparation), air (see
ISO 16000-16 and ISO 16000-18) and house dust may be sampled and analysed for culturable moulds (see
ISO 16000-17). Moulds can also be quantified and, to some extent, differentiated without prior cultivation. For
this purpose, airborne mould spores are collected on filters or directly on an adhesive-coated microscope slide,
followed by staining and subsequent direct microscopy.
Annex B gives an overview on the most common devices for total spore count measurements as well as for
sampling devices for filtration and impaction and the respective analysis methods.
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ISO 16000-19:2012(E)
6 Measurement strategy
6.1 General aspects
There is no standard procedure for measurement and assessment of mould damage. The type and amount of
measurements as well as the analytical methods employed are determined by the circumstances triggering
the investigation and the investigation objectives. A visual field inspection (walk through) by technically
qualified professionals prior to sampling is a key prerequisite for detecting and assessing mould sources in
indoor environments. Besides a good knowledge of building engineering and building physics, the
professionals conducting the inspection should have a sufficient background in indoor air hygiene and
microbiology.
Investigations are conducted with the objective of locating mould sources in indoor environments. To support
findings from visual observations and confirm suspected mould growth, professionals can draw on a variety of
sampling and analysis methods. These include methods for determining mould concentrations in or on
materials, procedures for the measurement of mould concentrations in indoor air as well as procedures for
determining mould concentrations in house dust. An example for a report accompanying sampling is attached
as Annex C.
Circumstances triggering a microbiological investigation of indoor environments may include the following (see
also Table 1):
visible mould damage;
material dampness without presence of visible mould growth;
structural or non-structural building anomalies without presence of visible mould growth;
health problems without presence of visible mould growth;
odour problems without presence of visible mould growth;
verification measurements during and after remediation.
In the case of visible mould damage with known source, the remediation and elimination of the underlying
causes should be addressed as a priority. In many cases, microbiological investigations is not necessary.
If mould damage is suspected without the presence of visible sources, the indoor environment can be
examined for the presence of elevated mould concentrations. Depending on the circumstances triggering the
investigation, the following media may be sampled and analysed:
a) materials and their surfaces (see 6.1.1);
b) indoor air in comparison with ambient air (see 6.1.2);
c) house dust (see 6.1.3).
The results of the measurements described in the following sections provide only indications on the damage
stage. Assessing the actual age of the mould growth is not feasible as the state of mould growth can change
drastically within very short time intervals.
The inspection of HVAC systems for lack of hygiene is not the subject of this part of ISO 16000.
In planning and performing measurements, the specific field conditions and influencing factors having a major
impact on the investigation results shall be taken into account and documented.
6 © ISO 2012 – All rights reserved
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ISO 16000-19:2012(E)
6.1.1 Analysis of surfaces or materials
For the selection of a suitable sampling method and the definition of the sampling locations, the following
questions have to be clarified.
Is mould growth or secondary contamination expected on the surface or the material?
Is a surface colonization or a colonization of deeper layers expected?
Are the moulds expected or under study culturable?
Are criteria available for an assessment of the analysis result?
Criteria for differentiating between active mould growth and mould deposits on material surfaces or in wall
cavities originating from natural sedimentation are the mould concentration and evidence of mould structures,
e.g. mycelium or spore carriers, in the material or on its surface. The mould concentration in the material or on
the material surface varies with the type of material, especially the density of the material, and the mould
species. Different mould species grow on or in a material depending on moisture, temperature and nutrient
source. Suspected mould contamination of surfaces may be confirmed by surface sampling using the tape-lift
and contact plate methods. The contact plate method presupposes that the moulds are culturable. If the
surface has already been disinfected or if contamination with Stachybotrys chartarum is suspected, the
contact plate method is not applicable. In such cases, it is necessary that tape-lift samples of the surfaces be
examined by direct microscopy.
Surface sampling (contact plate and tape-lift methods) has limited suitability for materials with rough surfaces
(e.g. plaster, insulation materials). Usually, the samples are suspended in a buffer followed by determination
[25]
of the mould concentration by cultivation or direct microscopy.
Where no empirical threshold levels exist for classifying a material as “contaminated” or “not contaminated”,
materials displaying no visible mould growth are sampled as controls for comparison.
6.1.2 Analysis of indoor air
The objective of indoor air sampling and analysis is to determine the concentration of moulds in a
representative air sample in order to ass
...
SLOVENSKI STANDARD
SIST ISO 16000-19:2014
01-junij-2014
1RWUDQML]UDN GHO 6WUDWHJLMDY]RUþHQMDJOLY
Indoor air - Part 19: Sampling strategy for moulds
Air intérieur - Partie 19: Stratégie d'échantillonnage des moisissures
Ta slovenski standard je istoveten z: ISO 16000-19:2012
ICS:
13.040.20 Kakovost okoljskega zraka Ambient atmospheres
SIST ISO 16000-19:2014 en,fr
2003-01.Slovenski inštitut za standardizacijo. Razmnoževanje celote ali delov tega standarda ni dovoljeno.
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SIST ISO 16000-19:2014
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SIST ISO 16000-19:2014
INTERNATIONAL ISO
STANDARD 16000-19
First edition
2012-06-01
Indoor air —
Part 19:
Sampling strategy for moulds
Air intérieur —
Partie 19: Stratégie d'échantillonnage des moisissures
Reference number
ISO 16000-19:2012(E)
©
ISO 2012
---------------------- Page: 3 ----------------------
SIST ISO 16000-19:2014
ISO 16000-19:2012(E)
COPYRIGHT PROTECTED DOCUMENT
© ISO 2012
All rights reserved. Unless otherwise specified, no part of this publication may be reproduced or utilized in any form or by any means,
electronic or mechanical, including photocopying and microfilm, without permission in writing from either ISO at the address below or
ISO's member body in the country of the requester.
ISO copyright office
Case postale 56 CH-1211 Geneva 20
Tel. + 41 22 749 01 11
Fax + 41 22 749 09 47
E-mail copyright@iso.org
Web www.iso.org
Published in Switzerland
ii © ISO 2012 – All rights reserved
---------------------- Page: 4 ----------------------
SIST ISO 16000-19:2014
ISO 16000-19:2012(E)
Contents Page
Foreword . iv
Introduction . vi
1 Scope . 1
2 Normative references . 1
3 Terms and definitions . 1
4 Properties, origin and occurrence of moulds in indoor environments . 4
5 Sampling and detection methods . 5
6 Measurement strategy . 6
6.1 General aspects . 6
6.2 Selection of appropriate procedure . 9
7 Quality requirements and uncertainty considerations . 17
Annex A (informative) Moisture damage indicators . 18
Annex B (informative) Devices for total spore count and detection of culturable fungi . 19
Annex C (informative) Field inspection report to describe sampling procedure and to document
potential mould damage . 21
Bibliography . 27
© ISO 2012 – All rights reserved iii
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SIST ISO 16000-19:2014
ISO 16000-19:2012(E)
Foreword
ISO (the International Organization for Standardization) is a worldwide federation of national standards bodies
(ISO member bodies). The work of preparing International Standards is normally carried out through ISO
technical committees. Each member body interested in a subject for which a technical committee has been
established has the right to be represented on that committee. International organizations, governmental and
non-governmental, in liaison with ISO, also take part in the work. ISO collaborates closely with the
International Electrotechnical Commission (IEC) on all matters of electrotechnical standardization.
International Standards are drafted in accordance with the rules given in the ISO/IEC Directives, Part 2.
The main task of technical committees is to prepare International Standards. Draft International Standards
adopted by the technical committees are circulated to the member bodies for voting. Publication as an
International Standard requires approval by at least 75 % of the member bodies casting a vote.
Attention is drawn to the possibility that some of the elements of this document may be the subject of patent
rights. ISO shall not be held responsible for identifying any or all such patent rights.
ISO 16000-19 was prepared by Technical Committee ISO/TC 146, Air quality, Subcommittee SC 6, Indoor air.
ISO 16000 consists of the following parts, under the general title Indoor air:
Part 1: General aspects of sampling strategy
Part 2: Sampling strategy for formaldehyde
Part 3: Determination of formaldehyde and other carbonyl compounds in indoor air and test chamber
air — Active sampling method
Part 4: Determination of formaldehyde — Diffusive sampling method
Part 5: Sampling strategy for volatile organic compounds (VOCs)
Part 6: Determination of volatile organic compounds in indoor and test chamber air by active sampling on
®
Tenax TA sorbent, thermal desorption and gas chromatography using MS or MS–FID
Part 7: Sampling strategy for determination of airborne asbestos fibre concentrations
Part 8: Determination of local mean ages of air in buildings for characterizing ventilation conditions
Part 9: Determination of the emission of volatile organic compounds from building products and
furnishing — Emission test chamber method
Part 10: Determination of the emission of volatile organic compounds from building products and
furnishing — Emission test cell method
Part 11: Determination of the emission of volatile organic compounds from building products and
furnishing — Sampling, storage of samples and preparation of test specimens
Part 12: Sampling strategy for polychlorinated biphenyls (PCBs), polychlorinated dibenzo-p-dioxins
(PCDDs), polychlorinated dibenzofurans (PCDFs) and polycyclic aromatic hydrocarbons (PAHs)
Part 13: Determination of total (gas and particle-phase) polychlorinated dioxin-like biphenyls (PCBs) and
polychlorinated dibenzo-p-dioxins/dibenzofurans (PCDDs/PCDFs) — Collection on sorbent-backed filters
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Part 14: Determination of total (gas and particle-phase) polychlorinated dioxin-like biphenyls (PCBs) and
polychlorinated dibenzo-p-dioxins/dibenzofurans (PCDDs/PCDFs) — Extraction, clean-up and analysis by
high-resolution gas chromatography and mass spectrometry
Part 15: Sampling strategy for nitrogen dioxide (NO )
2
Part 16: Detection and enumeration of moulds — Sampling by filtration
Part 17: Detection and enumeration of moulds — Culture-based method
Part 18: Detection and enumeration of moulds — Sampling by impaction
Part 19: Sampling strategy for moulds
Part 23: Performance test for evaluating the reduction of formaldehyde concentrations by sorptive
building materials
Part 24: Performance test for evaluating the reduction of volatile organic compound (except
formaldehyde) concentrations by sorptive building materials
Part 25: Determination of the emission of semi-volatile organic compounds by building products —
Micro-chamber method
Part 26: Sampling strategy for carbon dioxide (CO )
2
Part 28: Determination of odour emissions from building products using test chambers
The following parts are under preparation:
Part 21: Detection and enumeration of moulds — Sampling from materials
Part 27: Determination of settled fibrous dust on surfaces by SEM (scanning electron microscopy) (direct
method)
Part 29: Test methods for VOC detectors
Part 30: Sensory testing of indoor air
Part 31: Measurement of flame retardants and plasticizers based on organophosphorus compounds —
Phosphoric acid ester
Part 32: Investigation of constructions on pollutants and other injurious factors — Inspections
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Introduction
Mould spores and metabolites can be inhaled via the air and cause allergic and irritating reactions and/or
complex symptoms in humans. Moreover, mould growth can be associated with severe odour nuisances. In
[14][18][19]
rare cases, some mould species can cause infections (so-called mycoses) in certain risk groups.
There is sufficient epidemiological evidence that damp and mouldy buildings increase the risk of respiratory
[8]
symptoms, respiratory infections and enhances asthma symptoms of the occupants. In addition, there is
some evidence for increased risk of development of allergic rhinitis and asthma. Furthermore, there is clinical
evidence for rare symptoms like allergic alveolitis, chronic rhinosinusitis and allergic sinusitis. Toxicological
studies in vivo and in vitro show irritating and toxic reactions of microorganisms (including spores, cell
[8]
components and metabolites) from damp buildings.
Growth of microorganisms in damp buildings can lead to increased concentrations of spores, cell fragments,
allergens, mycotoxins, endotoxins, -glucanes and MVOC (microbial volatile organic compounds). From the
studies conducted so far it is not clear which compounds are the causative agents of the health effects
observed. Nevertheless, increased concentrations of each of these compounds are considered a potential
[8][18]
health risk and growth of mould in buildings should, therefore, be avoided.
The prime objective of this part of ISO 16000 is to provide assistance in identifying mould sources in indoor
environments.
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INTERNATIONAL STANDARD ISO 16000-19:2012(E)
Indoor air —
Part 19:
Sampling strategy for moulds
1 Scope
This part of ISO 16000 describes the measurement strategy for the detection of fungi in indoor environments.
It describes suitable sampling and analysis methods together with a description of the applicability and the
interpretation of the measurement results to maximize the comparability of the measured data obtained for a
given measurement objective. It does not include details on recording building characteristics or field
inspections by qualified professionals which have to take place prior to any microbiological measurement.
This part of ISO 16000 is not applicable to a detailed description of the building physics- and building-
engineering-related procedures applicable to field inspections. The methods and procedures presented do not
allow quantitative exposure assessment with regard to the room occupants.
The application of this part of ISO 16000 presupposes the knowledge of ISO 16000-1.
2 Normative references
The following referenced documents are indispensable for the application of this document. For dated
references, only the edition cited applies. For undated references, the latest edition of the referenced
document (including any amendments) applies.
ISO 16000-16, Indoor air — Part 16: Detection and enumeration of moulds — Sampling by filtration
ISO 16000-18, Indoor air — Part 18: Detection and enumeration of moulds — Sampling by impaction
3 Terms and definitions
For the purposes of this document, the following terms and definitions apply.
3.1
pre-existing mouldy condition
desiccated “old” mould growth, where additional biomass growth no longer occurs and the indoor air mould
spore concentration gradually decreases with time
3.2
biological preservation efficiency
capacity of the sampler to maintain the viability of the airborne microorganisms during collection and also to
keep the microbial products intact
[6]
[SOURCE: EN 13098:2000 ]
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NOTE The biological collection efficiency considers the sampling stress occurring during sampling and analysis in
addition to the physical collection efficiency.
3.3
identification of moulds
assignment of moulds to spore types or groups on the basis of defined properties (e.g. morphological,
biochemical, molecular-biological properties)
NOTE The term “differentiation” is frequently used instead of identification. The term “differentiation” is, however,
misleading because the intention is not to merely differentiate the moulds but to identify them, i.e. to assign them, e.g. to
genera or species.
3.4
filamentous fungus
fungus growing in the form of filaments of cells known as hyphae
NOTE 1 Hyphae aggregated in bundles are called mycelia.
NOTE 2 The term “filamentous fungi” differentiates fungi with hyphal growth from yeasts.
3.5
filtration
collection of particles suspended in a gas or liquid by flow through a porous medium
[6]
[SOURCE: EN 13098:2000 ]
NOTE In this part of ISO 16000, filtration is understood as the separation of microorganisms or moulds from a
defined volume of air by means of filters.
3.6
total spore count
number of (culturable and non-culturable) spores that are collected and enumerated under the microscope
NOTE For the term “spores”, see 3.19, Note 2.
3.7
yeast
unicellular fungus that does not normally produce a mycelium and reproduce by budding (budding fungi) as
against moulds, which reproduce by sporulation
3.8
impaction
sampling of particles suspended in air by inertial separation on a solid surface (culture medium or adhesive-
coated slides)
NOTE 1 See 16000-18.
NOTE 2 Sampling is carried out using either round-hole or slit impactors, for instance. As the air passes through the
orifices, it is accelerated and the particles are impacted on the medium located directly behind the nozzles as a result of
their inertia, while the air flows around the culture medium and exits the sampler. Impaction samples are only suitable for
direct analysis without further resuspension of the sample.
3.9
colony forming unit
cfu
air quality unit by which the culturable number of microorganisms is expressed
[6]
[SOURCE: EN 13098:2000 ]
NOTE 1 One colony can originate from one single microorganism, from aggregates of many microorganisms as well as
from one or many microorganisms attached to a particle.
NOTE 2 The number of colonies can depend on the cultivation conditions.
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3.10
colony morphology type
group of colonies which due to their morphological appearance seem to belong to a specific species
3.11
colony count
air quality number of all microorganism colonies visible on a culture medium after incubation under the
selected cultivation conditions
3.12
culturable mould
mould that can be cultured under the selected cultivation conditions
NOTE Parameters governing the culturability are, for instance, the type of culture medium and the incubation
temperature.
3.13
cultivation
growing of microorganisms on culture media
3.14
mycotoxin
secondary metabolites of moulds which are toxic to humans and animals
3.15
mycelium
total of fungal hyphae
3.16
non-culturable mould
mould that cannot be cultured under the selected cultivation conditions
3.17
physical sampling efficiency
capacity of the sampler to collect particles with specific aerodynamic diameters suspended in air
[6]
[SOURCE: EN 13098:2000, modified — "aerodynamic diameters" has replaced "sizes".]
3.18
sampling stress
damage suffered by the microorganisms during sampling (e.g. through mechanical and chemical effects or
through water deprivation)
3.19
mould
filamentous fungi from several taxonomic groups, namely Ascomycetes, Zygomycetes, and their anamorphic
states formerly known as Deuteromycetes or fungi imperfecti
NOTE 1 Taxonomically, moulds do not represent a uniform group.
NOTE 2 Moulds form different types of spores depending on the taxonomic group they belong to, namely
conidiospores (conidia), sporangiospores or ascospores. In practice, all these reproductive stages are summarized under
the term “spores”.
3.20
mould damage
damage caused to building materials and surfaces by mould growth
NOTE Mould damage can result in loss in value, health risks and restrict the occupancy of the affected sites.
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3.21
secondary colony
colony that does not originate from the “primary” sampling of airborne spores but from a spore released from a
colony growing on the agar plates
3.22
secondary contamination
mould contamination of surfaces not caused by mould growth but originating from a (contaminated) primary
source after aerial dispersion
3.23
cut-off value
particle size (aerodynamic diameter) for which the sampling efficiency is 50 %
3.24
total sampling efficiency
product of the physical sampling efficiency and the biological preservation efficiency
[6]
[SOURCE: EN 13098:2000 ]
4 Properties, origin and occurrence of moulds in indoor environments
Moulds are ubiquitous on our planet. They are involved in the decomposition of organic material and,
therefore, play an important role in the natural carbon cycle. Their concentration in the ambient air depends,
inter alia, on location, climate, time of the day and season. Airborne mould concentrations are subject to great
[9][10][11]
variability. This is due to the following reasons.
The mould concentration in local ambient air is mainly determined by the location relative to the respective
mould sources, wind direction and wind force. Mould spores are frequently released by specific sources such
as decaying material. Both natural processes and production processes, such as composting, recycling,
animal production facilities, grain and food processing plants as well as horticulture facilities, can be sources
of mould dispersion.
Sporulation, i.e. the production of mould spores occurs discontinuously. It is governed, inter alia, by the mould
lifecycle phase, the environmental conditions, stress factors, humidity as well as substrate composition and
availability.
Factors governing the dispersion of spores, most of which have aerodynamic diameters in the range of 2 µm
to 40 µm, are mechanically or thermally induced air movements, drying phases (leading e.g. to de-
[12][13][14]
agglomeration of deposited dust) and the capability of air dispersal of the mould spores.
Due to the ubiquitous nature of moulds, it can be assumed that they are always present in indoor air. The
presence of moulds in indoor air can be due to spores originating from ambient air on the one hand and to
recent active mould growth, pre-existing mouldy conditions or mould deposits (settled spores) on the other. To
distinguish between sources, it is, therefore, important to perform ambient air measurements for reference
[14][15]
whenever conducting indoor air measurements for moulds. In addition, the collection of a control sample
from a suitable reference room may be helpful.
Possible causes of indoor mould sources are surface moisture on building materials or moisture in the building
structure, but also rotting food, potted plants, biowaste collection, source separation of waste, deposited dust
due to poor cleaning as well as the keeping of animals in residential settings. Moisture damage can be
attributable to building defects, inappropriate ventilation and heating or unfavourable arrangement of furniture
as well as water damage (e.g. plumbing leaks or flooding events). Elevated mould levels in indoor
environments and the occurrence of certain mould species (see Annex A) are indicative of excessive moisture.
When residential environments or occupational settings are infested with moulds, the mould source shall be
located to be able to plan remedial measures.
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Main factors affecting the intensity of mould growth and the mould species developing are moisture,
temperature, nutrient supply and the pH. If environmental conditions are favourable, a great variety of moulds
can develop. Once environmental conditions become less favourable, the species best adapted to the given
[16]
conditions will predominate.
Mould sources can release spores, mycelial fragments, but also cell components and metabolic products such
as -glucans (polysaccharides contained in the cell wall of fungi), ergosterol (steroid compound contained in
the cell membrane of fungi), toxins and MVOCs (microbial volatile organic compounds such as certain
aldehydes, alcohols, esters, ketones). On cultivation, colonies can grow not only from spores, but also from
mycelial fragments.
The number and airborne dissemination of spores released vary with the type of mould damage. For an
assessment of indoor mould sources, it is, therefore, important to differentiate the individual mould species by
their type of spore dispersal. Experience has shown that even minor mould contamination of materials can
result in elevated indoor air mould levels if the species involved have dry spores with good air dispersal
capabilities (e.g. Penicillium and Aspergillus). By contrast, airborne spore concentrations are much lower
when materials are colonized, for instance, by moulds of the genera Acremonium, Fusarium or the species
Stachybotrys chartarum that have relatively large spores embedded in slimy substances and, therefore, have
poor air dispersal capabilities.
Furthermore, it should be taken into account that mould spores are not necessarily present as individual
spores in the air or settled dust, but also occur in the form of spore aggregates or are particle-borne.
Depending on the analysis method, they are determined individually or as spore aggregate. Materials, indoor
air and house dust contain not only culturable but also non-culturable mould spores, some of which can have
the same allergenic and toxic effects as culturable spores. For this reason, techniques have been developed
that allow the microscopic determination of both culturable and non-culturable moulds.
Mould detection and identification are performed either after cultivation based on morphological criteria,
biochemical reactions and/or molecular techniques or by direct microscopic examination. Identification based
on the morphological structure (macroscopic examination, stereo-microscopy and microscopy) either after
prior cultivation or by direct microscopy is still the most prevalent approach for the detection of moulds.
Besides, there are other analytical methods based on the determination of cell components and metabolites of
[17]
moulds such as -glucans, ergosterol, toxins and MVOCs. The determination of these compounds serves,
however, only as supplementary information.
The sampling methods employed for detection of moulds are determined by the objective of the investigation.
Depending on the sampling method, the moulds suffer a sampling stress during sample collection and
preparation, which can lead to their drying-out or dying. Factors affecting the culturability of mould spores are
their physiological state as well as the culture medium employed. Some mould species cannot be cultured at
all under laboratory conditions.
NOTE The genera Stachybotrys and Chaetomium hardly grow and sporulate only poorly, if at all, on DG18 agar. The
use of this culture medium for culture-based analysis of these genera is therefore not recommended (see ISO 16000-17).
For a literature summary see References [8]–[10], [12], and [14]–[18].
5 Sampling and detection methods
Depending on the objective of the investigation, materials (see ISO 16000-21, in preparation), air (see
ISO 16000-16 and ISO 16000-18) and house dust may be sampled and analysed for culturable moulds (see
ISO 16000-17). Moulds can also be quantified and, to some extent, differentiated without prior cultivation. For
this purpose, airborne mould spores are collected on filters or directly on an adhesive-coated microscope slide,
followed by staining and subsequent direct microscopy.
Annex B gives an overview on the most common devices for total spore count measurements as well as for
sampling devices for filtration and impaction and the respective analysis methods.
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6 Measurement strategy
6.1 General aspects
There is no standard procedure for measurement and assessment of mould damage. The type and amount of
measurements as well as the analytical methods employed are determined by the circumstances triggering
the investigation and the investigation objectives. A visual field inspection (walk through) by technically
qualified professionals prior to sampling is a key prerequisite for detecting and assessing mould sources in
indoor environments. Besides a good knowledge of building engineering and building physics, the
professionals conducting the inspection should have a sufficient background in indoor air hygiene and
microbiology.
Investigations are conducted with the objective of locating mould sources in indoor environments. To support
findings from visual observations and confirm suspected mould growth, professionals can draw on a variety of
sampling and analysis methods. These include methods for determining mould concentrations in or on
materials, procedures for the measurement of mould concentrations in indoor air as well as procedures for
determining mould concentrations in house dust. An example for a report accompanying sampling is attached
as Annex C.
Circumstances triggering a microbiological investigation of indoor environments may include the following (see
also Table 1):
visible mould damage;
material dampness without presence of visible mould growth;
structural or non-structural building anomalies without presence of visible mould growth;
health problems without presence of visible mould growth;
odour problems without presence of visible mould growth;
verification measurements during and after remediation.
In the case of visible mould damage with known source, the remediation and elimination of the underlying
causes should be addressed as a priority. In many cases, microbiological investigations is not necessary.
If mould damage is suspected without the presence of visible sources, the indoor environment can be
examined for the presence of elevated mould concentrations. Depending on the circumstances triggering the
investigation, the following media may be sampled and analysed:
a) materials and their surfaces (see 6.1.1);
b) indoor air in comparison with ambient air (see 6.1.2);
c) house dust (see 6.1.3).
The results of the measurements described in the following sections provide only indications on the damage
stage. Assessing the actual age of the mould growth is not feasible as the state of mould growth can change
drastically within very short time intervals.
The inspection of HVAC systems for lack of hygiene is not the subject of this part of ISO 16000.
In planning and performing measurements, the specific field conditions and influencing factors having a major
impact on the investigation results shall be taken into account and documented.
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6.1.1 Analysis of surfaces or materials
For the selection of a suitable sampling method and the definition of the sampling locations, the following
questions have to be clarified.
Is mould growth or secondary contamination expected on the surface or the material?
Is a surface colonization or a colonization of deeper layers expected?
Are the moulds expected or under study culturable?
Are criteria available for an assessment of the analysis result?
Criteria for differentiating between active mould growth and mould deposits on material surfaces or in wall
cavities originating from natural sedimentation are the mould concentration and evidence of mould structures,
e.g. mycelium or spore carriers, in the material or on its surface. The mould concentration in the material or on
the material surface varies with the type of material, especially the density of the material, and the mould
species. Different mould species grow on or in a material depending on moisture, temperature and nutrient
source. Suspected mould contamination of surfaces may be confirmed by surface sampling using the tape-lift
and contact plate methods. The contact plate method presupposes t
...
2003-01.Slovenski inštitut za standardizacijo. Razmnoževanje celote ali delov tega standarda ni dovoljeno.Air intérieur - Partie 19: Stratégie d'échantillonnage des moisissuresIndoor air - Part 19: Sampling strategy for moulds13.040.20Kakovost okoljskega zrakaAmbient atmospheresICS:Ta slovenski standard je istoveten z:ISO 16000-19:2012oSIST ISO 16000-19:2013en,fr01-april-2013oSIST ISO 16000-19:2013SLOVENSKI
STANDARD
oSIST ISO 16000-19:2013
Reference numberISO 16000-19:2012(E)© ISO 2012
INTERNATIONAL STANDARD ISO16000-19First edition2012-06-01Indoor air — Part 19: Sampling strategy for moulds Air intérieur — Partie 19: Stratégie d'échantillonnage des moisissures
oSIST ISO 16000-19:2013
ISO 16000-19:2012(E)
COPYRIGHT PROTECTED DOCUMENT
©
ISO 2012 All rights reserved. Unless otherwise specified, no part of this publication may be reproduced or utilized in any form or by any means, electronic or mechanical, including photocopying and microfilm, without permission in writing from either ISO at the address below or ISO's member body in the country of the requester. ISO copyright office Case postale 56 CH-1211 Geneva 20 Tel.
+ 41 22 749 01 11 Fax
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copyright@iso.org Web
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ii © ISO 2012 – All rights reserved
oSIST ISO 16000-19:2013
ISO 16000-19:2012(E) © ISO 2012 – All rights reserved iii Contents Page Foreword . iv Introduction . vi 1 Scope . 1 2 Normative references . 1 3 Terms and definitions . 1 4 Properties, origin and occurrence of moulds in indoor environments . 4 5 Sampling and detection methods . 5 6 Measurement strategy . 6 6.1 General aspects . 6 6.2 Selection of appropriate procedure . 9 7 Quality requirements and uncertainty considerations . 17 Annex A (informative)
Moisture damage indicators . 18 Annex B (informative)
Devices for total spore count and detection of culturable fungi . 19 Annex C (informative)
Field inspection report to describe sampling procedure and to document potential mould damage . 21 Bibliography . 27
oSIST ISO 16000-19:2013
ISO 16000-19:2012(E) iv © ISO 2012 – All rights reserved Foreword ISO (the International Organization for Standardization) is a worldwide federation of national standards bodies (ISO member bodies). The work of preparing International Standards is normally carried out through ISO technical committees. Each member body interested in a subject for which a technical committee has been established has the right to be represented on that committee. International organizations, governmental and non-governmental, in liaison with ISO, also take part in the work. ISO collaborates closely with the International Electrotechnical Commission (IEC) on all matters of electrotechnical standardization. International Standards are drafted in accordance with the rules given in the ISO/IEC Directives, Part 2. The main task of technical committees is to prepare International Standards. Draft International Standards adopted by the technical committees are circulated to the member bodies for voting. Publication as an International Standard requires approval by at least 75 % of the member bodies casting a vote. Attention is drawn to the possibility that some of the elements of this document may be the subject of patent rights. ISO shall not be held responsible for identifying any or all such patent rights. ISO 16000-19 was prepared by Technical Committee ISO/TC 146, Air quality, Subcommittee SC 6, Indoor air. ISO 16000 consists of the following parts, under the general title Indoor air: Part 1: General aspects of sampling strategy Part 2: Sampling strategy for formaldehyde Part 3: Determination of formaldehyde and other carbonyl compounds in indoor air and test chamber air — Active sampling method Part 4: Determination of formaldehyde — Diffusive sampling method Part 5: Sampling strategy for volatile organic compounds (VOCs) Part 6: Determination of volatile organic compounds in indoor and test chamber air by active sampling on Tenax TA® sorbent, thermal desorption and gas chromatography using MS or MS–FID Part 7: Sampling strategy for determination of airborne asbestos fibre concentrations Part 8: Determination of local mean ages of air in buildings for characterizing ventilation conditions Part 9: Determination of the emission of volatile organic compounds from building products and furnishing — Emission test chamber method Part 10: Determination of the emission of volatile organic compounds from building products and furnishing — Emission test cell method Part 11: Determination of the emission of volatile organic compounds from building products and furnishing — Sampling, storage of samples and preparation of test specimens Part 12: Sampling strategy for polychlorinated biphenyls (PCBs), polychlorinated dibenzo-p-dioxins (PCDDs), polychlorinated dibenzofurans (PCDFs) and polycyclic aromatic hydrocarbons (PAHs) Part 13: Determination of total (gas and particle-phase) polychlorinated dioxin-like biphenyls (PCBs) and polychlorinated dibenzo-p-dioxins/dibenzofurans (PCDDs/PCDFs) — Collection on sorbent-backed filters oSIST ISO 16000-19:2013
ISO 16000-19:2012(E) © ISO 2012 – All rights reserved v Part 14: Determination of total (gas and particle-phase) polychlorinated dioxin-like biphenyls (PCBs) and polychlorinated dibenzo-p-dioxins/dibenzofurans (PCDDs/PCDFs) — Extraction, clean-up and analysis by high-resolution gas chromatography and mass spectrometry Part 15: Sampling strategy for nitrogen dioxide (NO2) Part 16: Detection and enumeration of moulds — Sampling by filtration Part 17: Detection and enumeration of moulds — Culture-based method Part 18: Detection and enumeration of moulds — Sampling by impaction Part 19: Sampling strategy for moulds Part 23: Performance test for evaluating the reduction of formaldehyde concentrations by sorptive building materials Part 24: Performance test for evaluating the reduction of volatile organic compound (except formaldehyde) concentrations by sorptive building materials Part 25: Determination of the emission of semi-volatile organic compounds by building products — Micro-chamber method Part 26: Sampling strategy for carbon dioxide (CO2) Part 28: Determination of odour emissions from building products using test chambers The following parts are under preparation: Part 21: Detection and enumeration of moulds — Sampling from materials Part 27: Determination of settled fibrous dust on surfaces by SEM (scanning electron microscopy) (direct method) Part 29: Test methods for VOC detectors Part 30: Sensory testing of indoor air Part 31: Measurement of flame retardants and plasticizers based on organophosphorus compounds — Phosphoric acid ester Part 32: Investigation of constructions on pollutants and other injurious factors — Inspections
oSIST ISO 16000-19:2013
ISO 16000-19:2012(E) vi © ISO 2012 – All rights reserved Introduction Mould spores and metabolites can be inhaled via the air and cause allergic and irritating reactions and/or complex symptoms in humans. Moreover, mould growth can be associated with severe odour nuisances. In rare cases, some mould species can cause infections (so-called mycoses) in certain risk groups.[14][18][19] There is sufficient epidemiological evidence that damp and mouldy buildings increase the risk of respiratory symptoms, respiratory infections and enhances asthma symptoms of the occupants.[8] In addition, there is some evidence for increased risk of development of allergic rhinitis and asthma. Furthermore, there is clinical evidence for rare symptoms like allergic alveolitis, chronic rhinosinusitis and allergic sinusitis. Toxicological studies in vivo and in vitro show irritating and toxic reactions of microorganisms (including spores, cell components and metabolites) from damp buildings.[8] Growth of microorganisms in damp buildings can lead to increased concentrations of spores, cell fragments, allergens, mycotoxins, endotoxins, -glucanes and MVOC (microbial volatile organic compounds). From the studies conducted so far it is not clear which compounds are the causative agents of the health effects observed. Nevertheless, increased concentrations of each of these compounds are considered a potential health risk[8][18] and growth of mould in buildings should, therefore, be avoided. The prime objective of this part of ISO 16000 is to provide assistance in identifying mould sources in indoor environments. oSIST ISO 16000-19:2013
INTERNATIONAL STANDARD ISO 16000-19:2012(E) © ISO 2012 – All rights reserved 1 Indoor air — Part 19: Sampling strategy for moulds 1 Scope This part of ISO 16000 describes the measurement strategy for the detection of fungi in indoor environments. It describes suitable sampling and analysis methods together with a description of the applicability and the interpretation of the measurement results to maximize the comparability of the measured data obtained for a given measurement objective. It does not include details on recording building characteristics or field inspections by qualified professionals which have to take place prior to any microbiological measurement. This part of ISO 16000 is not applicable to a detailed description of the building physics- and building-engineering-related procedures applicable to field inspections. The methods and procedures presented do not allow quantitative exposure assessment with regard to the room occupants. The application of this part of ISO 16000 presupposes the knowledge of ISO 16000-1. 2 Normative references The following referenced documents are indispensable for the application of this document. For dated references, only the edition cited applies. For undated references, the latest edition of the referenced document (including any amendments) applies. ISO 16000-16, Indoor air — Part 16: Detection and enumeration of moulds — Sampling by filtration ISO 16000-18, Indoor air — Part 18: Detection and enumeration of moulds — Sampling by impaction
3 Terms and definitions For the purposes of this document, the following terms and definitions apply. 3.1 pre-existing mouldy condition desiccated “old” mould growth, where additional biomass growth no longer occurs and the indoor air mould spore concentration gradually decreases with time 3.2 biological preservation efficiency capacity of the sampler to maintain the viability of the airborne microorganisms during collection and also to keep the microbial products intact [SOURCE: EN 13098:2000[6]] oSIST ISO 16000-19:2013
ISO 16000-19:2012(E) 2 © ISO 2012 – All rights reserved NOTE The biological collection efficiency considers the sampling stress occurring during sampling and analysis in addition to the physical collection efficiency. 3.3 identification of moulds assignment of moulds to spore types or groups on the basis of defined properties (e.g. morphological, biochemical, molecular-biological properties) NOTE The term “differentiation” is frequently used instead of identification. The term “differentiation” is, however, misleading because the intention is not to merely differentiate the moulds but to identify them, i.e. to assign them, e.g. to genera or species. 3.4 filamentous fungus fungus growing in the form of filaments of cells known as hyphae NOTE 1 Hyphae aggregated in bundles are called mycelia. NOTE 2 The term “filamentous fungi” differentiates fungi with hyphal growth from yeasts. 3.5 filtration collection of particles suspended in a gas or liquid by flow through a porous medium [SOURCE: EN 13098:2000[6]] NOTE In this part of ISO 16000, filtration is understood as the separation of microorganisms or moulds from a defined volume of air by means of filters. 3.6 total spore count number of (culturable and non-culturable) spores that are collected and enumerated under the microscope NOTE For the term “spores”, see 3.19, Note 2. 3.7 yeast unicellular fungus that does not normally produce a mycelium and reproduce by budding (budding fungi) as against moulds, which reproduce by sporulation 3.8 impaction sampling of particles suspended in air by inertial separation on a solid surface (culture medium or adhesive-coated slides) NOTE 1 See 16000-18. NOTE 2 Sampling is carried out using either round-hole or slit impactors, for instance. As the air passes through the orifices, it is accelerated and the particles are impacted on the medium located directly behind the nozzles as a result of their inertia, while the air flows around the culture medium and exits the sampler. Impaction samples are only suitable for direct analysis without further resuspension of the sample. 3.9 colony forming unit cfu air quality unit by which the culturable number of microorganisms is expressed [SOURCE: EN 13098:2000[6]] NOTE 1 One colony can originate from one single microorganism, from aggregates of many microorganisms as well as from one or many microorganisms attached to a particle. NOTE 2 The number of colonies can depend on the cultivation conditions. oSIST ISO 16000-19:2013
ISO 16000-19:2012(E) © ISO 2012 – All rights reserved 3 3.10 colony morphology type group of colonies which due to their morphological appearance seem to belong to a specific species 3.11 colony count air quality number of all microorganism colonies visible on a culture medium after incubation under the selected cultivation conditions 3.12 culturable mould mould that can be cultured under the selected cultivation conditions NOTE Parameters governing the culturability are, for instance, the type of culture medium and the incubation temperature. 3.13 cultivation growing of microorganisms on culture media 3.14 mycotoxin secondary metabolites of moulds which are toxic to humans and animals 3.15 mycelium total of fungal hyphae 3.16 non-culturable mould mould that cannot be cultured under the selected cultivation conditions 3.17 physical sampling efficiency capacity of the sampler to collect particles with specific aerodynamic diameters suspended in air [SOURCE: EN 13098:2000,[6] modified — "aerodynamic diameters" has replaced "sizes".] 3.18 sampling stress damage suffered by the microorganisms during sampling (e.g. through mechanical and chemical effects or through water deprivation) 3.19 mould filamentous fungi from several taxonomic groups, namely Ascomycetes, Zygomycetes, and their anamorphic states formerly known as Deuteromycetes or fungi imperfecti NOTE 1 Taxonomically, moulds do not represent a uniform group. NOTE 2 Moulds form different types of spores depending on the taxonomic group they belong to, namely conidiospores (conidia), sporangiospores or ascospores. In practice, all these reproductive stages are summarized under the term “spores”. 3.20 mould damage damage caused to building materials and surfaces by mould growth NOTE Mould damage can result in loss in value, health risks and restrict the occupancy of the affected sites. oSIST ISO 16000-19:2013
ISO 16000-19:2012(E) 4 © ISO 2012 – All rights reserved 3.21 secondary colony colony that does not originate from the “primary” sampling of airborne spores but from a spore released from a colony growing on the agar plates 3.22 secondary contamination mould contamination of surfaces not caused by mould growth but originating from a (contaminated) primary source after aerial dispersion 3.23 cut-off value particle size (aerodynamic diameter) for which the sampling efficiency is 50 % 3.24 total sampling efficiency product of the physical sampling efficiency and the biological preservation efficiency [SOURCE: EN 13098:2000[6]] 4 Properties, origin and occurrence of moulds in indoor environments Moulds are ubiquitous on our planet. They are involved in the decomposition of organic material and, therefore, play an important role in the natural carbon cycle. Their concentration in the ambient air depends, inter alia, on location, climate, time of the day and season. Airborne mould concentrations are subject to great variability.[9][10][11] This is due to the following reasons. The mould concentration in local ambient air is mainly determined by the location relative to the respective mould sources, wind direction and wind force. Mould spores are frequently released by specific sources such as decaying material. Both natural processes and production processes, such as composting, recycling, animal production facilities, grain and food processing plants as well as horticulture facilities, can be sources of mould dispersion. Sporulation, i.e. the production of mould spores occurs discontinuously. It is governed, inter alia, by the mould lifecycle phase, the environmental conditions, stress factors, humidity as well as substrate composition and availability. Factors governing the dispersion of spores, most of which have aerodynamic diameters in the range of 2 µm to 40 µm, are mechanically or thermally induced air movements, drying phases (leading e.g. to de-agglomeration of deposited dust) and the capability of air dispersal of the mould spores.[12][13][14] Due to the ubiquitous nature of moulds, it can be assumed that they are always present in indoor air. The presence of moulds in indoor air can be due to spores originating from ambient air on the one hand and to recent active mould growth, pre-existing mouldy conditions or mould deposits (settled spores) on the other. To distinguish between sources, it is, therefore, important to perform ambient air measurements for reference whenever conducting indoor air measurements for moulds.[14][15] In addition, the collection of a control sample from a suitable reference room may be helpful. Possible causes of indoor mould sources are surface moisture on building materials or moisture in the building structure, but also rotting food, potted plants, biowaste collection, source separation of waste, deposited dust due to poor cleaning as well as the keeping of animals in residential settings. Moisture damage can be attributable to building defects, inappropriate ventilation and heating or unfavourable arrangement of furniture as well as water damage (e.g. plumbing leaks or flooding events). Elevated mould levels in indoor environments and the occurrence of certain mould species (see Annex A) are indicative of excessive moisture. When residential environments or occupational settings are infested with moulds, the mould source shall be located to be able to plan remedial measures. oSIST ISO 16000-19:2013
ISO 16000-19:2012(E) © ISO 2012 – All rights reserved 5 Main factors affecting the intensity of mould growth and the mould species developing are moisture, temperature, nutrient supply and the pH. If environmental conditions are favourable, a great variety of moulds can develop. Once environmental conditions become less favourable, the species best adapted to the given conditions will predominate.[16] Mould sources can release spores, mycelial fragments, but also cell components and metabolic products such as -glucans (polysaccharides contained in the cell wall of fungi), ergosterol (steroid compound contained in the cell membrane of fungi), toxins and MVOCs (microbial volatile organic compounds such as certain aldehydes, alcohols, esters, ketones). On cultivation, colonies can grow not only from spores, but also from mycelial fragments. The number and airborne dissemination of spores released vary with the type of mould damage. For an assessment of indoor mould sources, it is, therefore, important to differentiate the individual mould species by their type of spore dispersal. Experience has shown that even minor mould contamination of materials can result in elevated indoor air mould levels if the species involved have dry spores with good air dispersal capabilities (e.g. Penicillium and Aspergillus). By contrast, airborne spore concentrations are much lower when materials are colonized, for instance, by moulds of the genera Acremonium, Fusarium or the species Stachybotrys chartarum that have relatively large spores embedded in slimy substances and, therefore, have poor air dispersal capabilities. Furthermore, it should be taken into account that mould spores are not necessarily present as individual spores in the air or settled dust, but also occur in the form of spore aggregates or are particle-borne. Depending on the analysis method, they are determined individually or as spore aggregate. Materials, indoor air and house dust contain not only culturable but also non-culturable mould spores, some of which can have the same allergenic and toxic effects as culturable spores. For this reason, techniques have been developed that allow the microscopic determination of both culturable and non-culturable moulds. Mould detection and identification are performed either after cultivation based on morphological criteria, biochemical reactions and/or molecular techniques or by direct microscopic examination. Identification based on the morphological structure (macroscopic examination, stereo-microscopy and microscopy) either after prior cultivation or by direct microscopy is still the most prevalent approach for the detection of moulds. Besides, there are other analytical methods based on the determination of cell components and metabolites of moulds such as -glucans, ergosterol, toxins and MVOCs.[17] The determination of these compounds serves, however, only as supplementary information. The sampling methods employed for detection of moulds are determined by the objective of the investigation. Depending on the sampling method, the moulds suffer a sampling stress during sample collection and preparation, which can lead to their drying-out or dying. Factors affecting the culturability of mould spores are their physiological state as well as the culture medium employed. Some mould species cannot be cultured at all under laboratory conditions. NOTE The genera Stachybotrys and Chaetomium hardly grow and sporulate only poorly, if at all, on DG18 agar. The use of this culture medium for culture-based analysis of these genera is therefore not recommended (see ISO 16000-17). For a literature summary see References [8]–[10], [12], and [14]–[18]. 5 Sampling and detection methods Depending on the objective of the investigation, materials (see ISO 16000-21, in preparation), air (see ISO 16000-16 and ISO 16000-18) and house dust may be sampled and analysed for culturable moulds (see ISO 16000-17). Moulds can also be quantified and, to some extent, differentiated without prior cultivation. For this purpose, airborne mould spores are collected on filters or directly on an adhesive-coated microscope slide, followed by staining and subsequent direct microscopy. Annex B gives an overview on the most common devices for total spore count measurements as well as for sampling devices for filtration and impaction and the respective analysis methods. oSIST ISO 16000-19:2013
ISO 16000-19:2012(E) 6 © ISO 2012 – All rights reserved 6 Measurement strategy 6.1 General aspects There is no standard procedure for measurement and assessment of mould damage. The type and amount of measurements as well as the analytical methods employed are determined by the circumstances triggering the investigation and the investigation objectives. A visual field inspection (walk through) by technically qualified professionals prior to sampling is a key prerequisite for detecting and assessing mould sources in indoor environments. Besides a good knowledge of building engineering and building physics, the professionals conducting the inspection should have a sufficient background in indoor air hygiene and microbiology. Investigations are conducted with the objective of locating mould sources in indoor environments. To support findings from visual observations and confirm suspected mould growth, professionals can draw on a variety of sampling and analysis methods. These include methods for determining mould concentrations in or on materials, procedures for the measurement of mould concentrations in indoor air as well as procedures for determining mould concentrations in house dust. An example for a report accompanying sampling is attached as Annex C. Circumstances triggering a microbiological investigation of indoor environments may include the following (see also Table 1): visible mould damage; material dampness without presence of visible mould growth; structural or non-structural building anomalies without presence of visible mould growth; health problems without presence of visible mould growth; odour problems without presence of visible mould growth; verification measurements during and after remediation. In the case of visible mould damage with known source, the remediation and elimination of the underlying causes should be addressed as a priority. In many cases, microbiological investigations is not necessary. If mould damage is suspected without the presence of visible sources, the indoor environment can be examined for the presence of elevated mould concentrations. Depending on the circumstances triggering the investigation, the following media may be sampled and analysed: a) materials and their surfaces (see 6.1.1); b) indoor air in comparison with ambient air (see 6.1.2); c) house dust (see 6.1.3). The results of the measurements described in the following sections provide only indications on the damage stage. Assessing the actual age of the mould growth is not feasible as the state of mould growth can change drastically within very short time intervals. The inspection of HVAC systems for lack of hygiene is not the subject of this part of ISO 16000. In planning and performing measurements, the specific field conditions and influencing factors having a major impact on the investigation results shall be taken into account and documented. oSIST ISO 16000-19:2013
ISO 16000-19:2012(E) © ISO 2012 – All rights reserved 7 6.1.1 Analysis of surfaces or materials For the selection of a suitable sampling method and the definition of the sampling locations, the following questions have to be clarified. Is mould growth or secondary contamination expected on the surface or the material? Is a surface colonization or a colonization of deeper layers expected? Are the moulds expected or under study culturable? Are criteria available for an assessment of the analysis result? Criteria for differentiating between active mould growth and mould deposits on material surfaces or in wall cavities originating from natural sedimentation are the mould concentration and evidence of mould structures, e.g. mycelium or spore carriers, in the material or on its surface. The mould concentration in the material or on the material surface varies with the type of material, especially the density of the material, and the mould species. Different mould species grow on or in a material depending on moisture, temperature and nutrient source. Suspected mould contamination of surfaces may be confirmed by surface sampling using the tape-lift and contact plate methods. The contact plate method presupposes that the moulds are culturable. If the surface has already been disinfected or if contamination with Stachybotrys chartarum is suspected, the contact plate method is not applicable. In such cases, it is necessary that tape-lift samples of the surfaces be examined by direct microscopy. Surface sampling (contact plate and tape-lift methods) has limited suitability for materials with rough surfaces (e.g. plaster, insulation materials). Usually, the samples are suspended in a buffer followed by determination of the mould concentration by cultivation or direct microscopy.[25] Where no empirical threshold levels exist for classifying a material as “contaminated” or “not contaminated”, materials displaying no visible mould growth are sampled as controls for comparison. 6.1.2 Analysis of indoor air The objective of indoor air sampling and analysis is to determine the concentration of moulds in a representative air sample in order to assess the likelihood of mould sources in the indoor environment. Depending on the investigation objective, this requires a more or less complete identification of the moulds (see 6.2). In analysing air samples, special attention is given to differences in the species spectrum present in the indoor compared to ambient air. Moreover, the presence of moisture-in
...
NORME ISO
INTERNATIONALE 16000-19
Première édition
2012-06-01
Air intérieur —
Partie 19:
Stratégie d'échantillonnage des
moisissures
Indoor air —
Part 19: Sampling strategy for moulds
Numéro de référence
ISO 16000-19:2012(F)
©
ISO 2012
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ISO 16000-19:2012(F)
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© ISO 2012
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ii © ISO 2012 – Tous droits réservés
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ISO 16000-19:2012(F)
Sommaire Page
Avant-propos . iv
Introduction . vi
1 Domaine d'application . 1
2 Références normatives . 1
3 Termes et définitions . 1
4 Propriétés, origine et occurrence des moisissures dans les environnements intérieurs . 4
5 Méthodes d'échantillonnage et de détection . 6
6 Stratégie de mesurage . 6
6.1 Aspects généraux . 6
6.2 Sélection d'un mode opératoire approprié . 10
7 Exigences de qualité et considérations relatives à l'incertitude . 19
Annexe A (informative) Indicateurs de dommage de moisissure . 20
Annexe B (informative) Dispositifs de détermination du nombre total de spores et de détection
des champignons cultivables . 21
Annexe C (informative) Rapport d'inspection sur le terrain pour décrire un mode opératoire de
prélèvement et pour documenter un dommage de moisissure potentiel . 23
Bibliographie . 29
© ISO 2012 – Tous droits réservés iii
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ISO 16000-19:2012(F)
Avant-propos
L'ISO (Organisation internationale de normalisation) est une fédération mondiale d'organismes nationaux de
normalisation (comités membres de l'ISO). L'élaboration des Normes internationales est en général confiée
aux comités techniques de l'ISO. Chaque comité membre intéressé par une étude a le droit de faire partie du
comité technique créé à cet effet. Les organisations internationales, gouvernementales et non
gouvernementales, en liaison avec l'ISO participent également aux travaux. L'ISO collabore étroitement avec
la Commission électrotechnique internationale (CEI) en ce qui concerne la normalisation électrotechnique.
Les Normes internationales sont rédigées conformément aux règles données dans les Directives ISO/CEI,
Partie 2.
La tâche principale des comités techniques est d'élaborer les Normes internationales. Les projets de Normes
internationales adoptés par les comités techniques sont soumis aux comités membres pour vote. Leur
publication comme Normes internationales requiert l'approbation de 75 % au moins des comités membres
votants.
L'attention est appelée sur le fait que certains des éléments du présent document peuvent faire l'objet de
droits de propriété intellectuelle ou de droits analogues. L'ISO ne saurait être tenue pour responsable de ne
pas avoir identifié de tels droits de propriété et averti de leur existence.
L'ISO 16000-19 a été élaborée par le comité technique ISO/TC 146, Qualité de l'air, sous-comité SC 6, Air
intérieur.
L'ISO 16000 comprend les parties suivantes, présentées sous le titre général Air intérieur:
Partie 1: Aspects généraux de la stratégie d'échantillonnage
Partie 2: Stratégie d'échantillonnage du formaldéhyde
Partie 3: Dosage du formaldéhyde et d'autres composés carbonylés dans l'air intérieur et dans l'air des
chambres d'essai — Méthode par échantillonnage actif
Partie 4: Dosage du formaldéhyde — Méthode par échantillonnage diffusif
Partie 5: Stratégie d'échantillonnage pour les composés organiques volatils (COV)
Partie 6: Dosage des composés organiques volatils dans l'air intérieur des locaux et chambres d'essai
®
par échantillonnage actif sur le sorbant Tenax TA , désorption thermique et chromatographie en phase
gazeuse utilisant MS ou MS-FID
Partie 7: Stratégie d'échantillonnage pour la détermination des concentrations en fibres d'amiante en
suspension dans l'air
Partie 8: Détermination des âges moyens locaux de l'air dans des bâtiments pour caractériser les
conditions de ventilation
Partie 9: Dosage de l'émission de composés organiques volatils de produits de construction et d'objets
d'équipement — Méthode de la chambre d'essai d'émission
Partie 10: Dosage de l'émission de composés organiques volatils de produits de construction et d'objets
d'équipement — Méthode de la cellule d'essai d'émission
iv © ISO 2012 – Tous droits réservés
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ISO 16000-19:2012(F)
Partie 11: Dosage de l'émission de composés organiques volatils de produits de construction et d'objets
d'équipement — Échantillonnage, conservation des échantillons et préparation d'échantillons pour essai
Partie 12: Stratégie d'échantillonnage des polychlorobiphényles (PCB), des polychlorodibenzo-p-dioxines
(PCDD), des polychlorodibenzofuranes (PCDF) et des hydrocarbures aromatiques polycycliques (HAP)
Partie 13: Dosage des polychlorobiphényles (PCB) de type dioxine et des polychlorodibenzo-p-dioxines
(PCDD)/polychlorodibenzofuranes (PCDF) totaux (en phase gazeuse et en phase particulaire) —
Collecte sur des filtres adsorbants
Partie 14: Dosage des polychlorobiphényles (PCB) de type dioxine et des polychlorodibenzo-p-dioxines
(PCDD)/polychlorodibenzofuranes (PCDF) totaux (en phase gazeuse et en phase particulaire) —
Extraction, purification et analyse par chromatographie en phase gazeuse haute résolution et
spectrométrie de masse
Partie 15: Stratégie d'échantillonnage du dioxyde d'azote (NO )
2
Partie 16: Détection et dénombrement des moisissures — Échantillonnage par filtration
Partie 17: Détection et dénombrement des moisissures — Méthode par culture
Partie 18: Détection et dénombrement des moisissures — Échantillonnage par impaction
Partie 19: Stratégie d'échantillonnage des moisissures
Partie 23: Essai de performance pour l'évaluation de la réduction des concentrations en formaldéhyde
par des matériaux de construction sorptifs
Partie 24: Essai de performance pour l'évaluation de la réduction des concentrations en composés
organiques volatils (sauf formaldéhyde) par des matériaux de construction sorptifs
Partie 25: Dosage de l'émission de composés organiques semi-volatils des produits de construction —
Méthode de la micro-chambre
Partie 26: Stratégie d'échantillonnage du dioxyde de carbone (CO )
2
Partie 28: Détermination des émissions d'odeurs des produits de construction au moyen de chambres
d'essai
Les parties suivantes sont en cours d'élaboration:
Partie 21: Détection et dénombrement des moisissures — Échantillonnage à partir de matériaux
Partie 27: Détermination de la poussière fibreuse déposée sur les surfaces par microscopie électronique
à balayage (MEB) (méthode directe)
Partie 29: Méthodes d'essai pour détecteurs de composés organiques volatils (COV)
Partie 30: Essai sensoriel de l'air intérieur
Partie 31: Mesurage des ignifugeants basés sur des composés organophosphorés — Ester d'acide
phosphorique
Partie 32: Investigation de polluants et autres facteurs nocifs dans les constructions — Inspections
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ISO 16000-19:2012(F)
Introduction
Les spores de moisissures et les métabolites peuvent être inhalés par voie aérienne et provoquer des
réactions allergiques et d'irritation et/ou des symptômes complexes chez l'homme. En outre, la formation de
moisissures peut s'accompagner de sérieuses nuisances olfactives. Dans de rares cas, certaines espèces de
[14][18][19]
moisissures peuvent être à l'origine d'infections (appelées mycoses) dans certains groupes à risque .
Il existe suffisamment de données épidémiologiques montrant que les bâtiments humides et moisis
augmentent le risque de symptômes et d'infections respiratoires et accentuent les symptômes asthmatiques
[8]
des occupants . Le risque accru de développement de rhinites allergiques et d'asthme est en outre démontré.
Il existe par ailleurs des preuves cliniques de symptômes rares tels que l'alvéolite allergique, la rhinosinusite
chronique et la sinusite allergique. Les études toxicologiques in vivo et in vitro montrent les effets irritants et
toxiques des micro-organismes (incluant les spores, les composants cellulaires et les métabolites) des
[8]
bâtiments humides .
Le développement de micro-organismes dans les bâtiments humides peut donner lieu à des concentrations
accrues de spores, de fragments cellulaires, d'allergènes, de mycotoxines, d'endotoxines, de -glucanes et de
COVM (composés organiques volatils microbiens). Les études menées jusqu'à présent n'ont pas pu montrer
clairement quels composés sont les agents responsables des effets observés sur la santé. Les fortes
concentrations de chacun de ces composés n'en sont pas moins considérées comme un risque potentiel pour
[8][18]
la santé et il convient par conséquent d'éviter la formation de moisissures dans les bâtiments.
Le principal objectif de la présente partie de l'ISO 16000 est d'aider à l'identification des sources de
moisissures dans les environnements intérieurs.
vi © ISO 2012 – Tous droits réservés
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NORME INTERNATIONALE ISO 16000-19:2012(F)
Air intérieur —
Partie 19:
Stratégie d'échantillonnage des moisissures
1 Domaine d'application
La présente partie de l'ISO 16000 décrit la stratégie de mesurage pour détecter les champignons dans les
environnements intérieurs.
Elle décrit des méthodes d'échantillonnage et d'analyse appropriées ainsi que l'applicabilité et l'interprétation
des résultats de mesurage pour maximiser la comparabilité des données mesurées obtenues pour un objectif
de mesurage donné. Elle ne contient pas d'indications détaillées concernant l'enregistrement des
caractéristiques du bâtiment ou les inspections sur le terrain menées par des professionnels qualifiés qui
doivent être effectués préalablement à tout mesurage microbiologique.
La présente partie de l'ISO 16000 ne s’applique pas à une description détaillée des modes opératoires relatifs
à la physique et au génie du bâtiment applicables aux inspections sur le terrain. Les méthodes et les modes
opératoires présentés ne permettent pas d'évaluer l'exposition quantitative des occupants de la pièce.
L'application de la présente partie de l'ISO 16000 présuppose que l'on ait pris connaissance de l'ISO 16000-1.
2 Références normatives
Les documents de référence suivants sont indispensables pour l'application du présent document. Pour les
références datées, seule l'édition citée s'applique. Pour les références non datées, la dernière édition du
document de référence s'applique (y compris les éventuels amendements).
ISO 16000-16, Air intérieur — Partie 16: Détection et dénombrement des moisissures — Échantillonnage par
filtration
ISO 16000-18, Air intérieur — Partie 18: Détection et dénombrement des moisissures — Échantillonnage par
impaction
3 Termes et définitions
Pour les besoins du présent document, les termes et définitions suivants s'appliquent.
3.1
état moisi pré-existant
formation de moisissures «ancienne» desséchée où la croissance d'une biomasse additionnelle a cessé, la
concentration des spores de moisissures dans l'air intérieur diminuant avec le temps
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ISO 16000-19:2012(F)
3.2
efficacité de conservation biologique
capacité de l'échantillonneur à conserver la viabilité des micro-organismes en suspension dans l'air pendant
la collecte et à conserver les produits microbiens intacts
[6]
[EN 13098:2000 ]
NOTE L'efficacité de collecte biologique tient compte du stress de prélèvement intervenant pendant le prélèvement
et l'analyse, en plus de l'efficacité de collecte physique.
3.3
identification des moisissures
affectation des moisissures à des types ou des groupes de spores sur la base de propriétés définies (par
exemple des propriétés morphologiques, biochimiques, moléculaires-biologiques)
NOTE Le terme «différenciation» est fréquemment utilisé à la place du terme «identification». Cependant, le terme
«différenciation» est trompeur car il ne suffit pas seulement de distinguer les moisissures mais il faut les identifier,
c'est-à-dire les affecter, par exemple, à un genre ou à une espèce.
3.4
champignon filamenteux
champignon poussant sous la forme de filaments de cellules appelés hyphes
NOTE 1 Les hyphes agrégées en faisceaux sont appelées mycélium.
NOTE 2 Le terme «champignons filamenteux» distingue les champignons à hyphes des levures.
3.5
filtration
prélèvement de particules en suspension dans un gaz ou un liquide par passage à travers un milieu poreux
[6]
[EN 13098:2000 ]
NOTE Dans la présente partie de l'ISO 16000, la filtration désigne la séparation des micro-organismes ou des
moisissures d'un volume défini d'air au moyen de filtres.
3.6
nombre total de spores
nombre de spores (cultivables et non cultivables) qui sont prélevées et dénombrées au microscope
NOTE Pour le terme «spores» voir 3.19, Note 2.
3.7
levure
champignon unicellulaire qui ne produit normalement pas de mycélium et se reproduit par bourgeonnement
(champignons bourgeonnants), contrairement aux moisissures qui se reproduisent par sporulation
3.8
impaction
prélèvement de particules en suspension dans l'air par séparation inertielle sur une surface solide (milieu de
culture ou lames à revêtement adhésif)
NOTE 1 Voir l'ISO 16000-18.
NOTE 2 Le prélèvement est réalisé en utilisant soit des impacteurs à trous ronds, soit des impacteurs à fente, par
exemple. L'air est accéléré lors de son passage à travers les orifices et les particules s'impactent sur le milieu placé
directement derrière les buses sous l'effet de leur inertie; l'air, quant à lui, contourne le milieu de culture et sort de
l'échantillonneur. Les échantillons obtenus par impaction ne conviennent que pour les analyses directes, sans remise en
suspension supplémentaire de l'échantillon.
2 © ISO 2012 – Tous droits réservés
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ISO 16000-19:2012(F)
3.9
unité formant colonie
ufc
qualité de l'air unité dans laquelle est exprimé le nombre de micro-organismes cultivables
[6]
[EN 13098:2000 ]
NOTE 1 Une colonie peut provenir d'un seul micro-organisme, d'agrégats de nombreux micro-organismes ou d'un ou
plusieurs micro-organismes liés à une particule.
NOTE 2 Le nombre de colonies peut dépendre des conditions de culture.
3.10
type morphologique des colonies
groupe de colonies qui, de par leur apparence morphologique, semblent appartenir à une espèce spécifique
3.11
nombre de colonies
qualité de l'air nombre de toutes les colonies de micro-organismes visibles sur un milieu de culture après
incubation dans les conditions de culture choisies
3.12
moisissure cultivable
moisissure qui peut être cultivée dans les conditions de culture choisies
NOTE Les paramètres déterminant l'aptitude à la culture sont, par exemple, le type de milieu de culture et la
température d'incubation.
3.13
culture
développement de micro-organismes sur des milieux de culture
3.14
mycotoxine
métabolites secondaires des moisissures qui sont toxiques pour l'homme et pour les animaux
3.15
mycélium
ensemble des hyphes d'un champignon
3.16
moisissure non cultivable
moisissure qui ne peut pas être cultivée dans les conditions de culture choisies
3.17
efficacité de prélèvement physique
capacité de l'échantillonneur à recueillir des particules de diamètres aérodynamiques spécifiques en
suspension dans l'air
[6]
[EN 13098:2000 modifiée — «tailles» a été remplacé par «diamètres aérodynamiques».]
3.18
stress de prélèvement
dommages subis par les micro-organismes pendant le prélèvement (par exemple résultant d'effets
mécaniques et chimiques ou de la privation d'eau)
© ISO 2012 – Tous droits réservés 3
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ISO 16000-19:2012(F)
3.19
moisissure
champignons filamenteux appartenant à plusieurs groupes taxonomiques, notamment les Ascomycètes, les
Zygomycètes et leurs états anamorphiques auparavant connus sous le nom de Deutéromycètes ou
«champignons imparfaits» (fungi imperfecti)
NOTE 1 Les moisissures ne représentent pas un groupe taxonomique uniforme.
NOTE 2 Les moisissures forment différents types de spores selon le groupe taxonomique auquel elles appartiennent, à
savoir des conidiospores (conidies), des sporangiospores ou des ascospores. Dans la pratique, toutes ces étapes de la
reproduction sont rassemblées sous le terme «spores».
3.20
dommage de moisissure
dommage causé aux matériaux de construction et aux surfaces des bâtiments par la formation de moisissures
NOTE Un dommage de moisissure peut entraîner une dépréciation des lieux et des risques pour la santé et
restreindre l'occupation des sites affectés.
3.21
colonie secondaire
colonie qui ne provient pas du prélèvement «initial» de spores en suspension dans l'air mais d'une spore
libérée par une colonie se développant sur les boîtes de gélose
3.22
contamination secondaire
contamination de surfaces par des moisissures, causée non par la formation de moisissures mais par une
source primaire (contaminée) après dispersion aérienne
3.23
valeur seuil
taille des particules (diamètre aérodynamique) pour laquelle l'efficacité du prélèvement est de 50 %
3.24
efficacité totale de prélèvement
produit de l'efficacité de prélèvement physique par l'efficacité de conservation biologique
[6]
[EN 13098:2000 ]
4 Propriétés, origine et occurrence des moisissures dans les environnements
intérieurs
Les moisissures existent partout sur la planète. Elles interviennent dans la décomposition de la matière
organique et jouent par conséquent un rôle important dans le cycle naturel du carbone. Leur concentration
dans l'air ambiant dépend, entre autres, du lieu, du climat, de l'heure et de la saison. Les concentrations de
[9][10][11]
moisissures en suspension dans l'air sont très variables , pour les raisons suivantes.
La concentration de moisissures dans l'air ambiant local est déterminée principalement par la position par
rapport aux sources de moisissures respectives, ainsi que par la direction et la force du vent. Les spores de
moisissures sont souvent libérées par des sources spécifiques telles que la matière en décomposition. Les
sources de dispersion des moisissures résident aussi bien dans des processus naturels que dans des
processus de production tels que le compostage, le recyclage, les installations de production animale, les
usines de traitement des céréales et des aliments ou encore les installations d'horticulture.
La sporulation, c'est-à-dire la production de spores de moisissures, est un processus discontinu qui est régi,
entre autres, par la phase du cycle de vie des moisissures, les conditions environnementales, les facteurs de
stress, l'humidité, la composition et la disponibilité du substrat.
4 © ISO 2012 – Tous droits réservés
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ISO 16000-19:2012(F)
Les facteurs influençant la dispersion des spores, dont la plupart ont des diamètres aérodynamiques d'un
ordre compris entre 2 µm et 40 µm, sont les mouvements de l'air induits mécaniquement ou thermiquement,
les phases de sécheresse (entraînant, par exemple, le soulèvement des poussières déposées) et la capacité
[12][13][14]
de dispersion aérienne des spores des moisissures .
Les moisissures faisant preuve d'ubiquité, on peut partir du principe qu'elles sont toujours présentes dans l'air
intérieur. La présence de moisissures dans l'air intérieur peut être due à des spores provenant de l'air ambiant,
d'une part, et à la formation récente de moisissures actives, à des conditions favorables à la moisissure pré-
existantes ou à des dépôts de moisissures (spores déposées), d'autre part. Pour faire la différence entre ces
sources, il est donc important de procéder à des mesurages de référence de l'air ambiant à chaque fois que
[14][15]
des mesurages de l'air intérieur sont effectués pour détecter les moisissures . Le prélèvement d'un
échantillon témoin dans une pièce de référence appropriée peut en outre s'avérer utile.
Les causes possibles des sources de moisissures à l'intérieur sont l'humidité présente à la surface des
matériaux de construction ou l'humidité présente dans la structure du bâtiment, mais également la nourriture
en putréfaction, les plantes en pot, la collecte des déchets organiques, le tri des déchets à la source, la
poussière déposée en raison d'un mauvais nettoyage, de même que la présence d'animaux dans les lieux de
résidence. Les dommages de moisissures peuvent être attribués à des défauts de construction, une
ventilation et un chauffage inappropriés, un agencement inadapté du mobilier ou encore un dégât des eaux
(par exemple des fuites de plomberie ou les suites d'une inondation). Des niveaux de moisissures élevés
dans les environnements intérieurs et l'occurrence de certaines espèces de moisissures (voir Annexe A) sont
indicateurs d'un excès d'humidité. Lorsque des environnements résidentiels ou des locaux professionnels
sont infestés par des moisissures, la source de moisissures doit être localisée pour permettre la mise en place
de mesures correctives.
Les principaux facteurs influençant l'intensité de la formation de moisissures et le développement des
espèces de moisissures sont l'humidité, la température, l'apport d'éléments nutritifs et le pH. Si les conditions
environnementales sont favorables, une grande diversité de moisissures peut se développer. Lorsque les
conditions environnementales deviennent moins favorables, les espèces les mieux adaptées aux conditions
[16]
données vont être prédominantes .
Les sources de moisissures peuvent libérer des spores, des fragments de mycélium, mais également des
composants cellulaires et des produits métaboliques tels que les -glucanes (polysaccharide contenu dans la
paroi cellulaire des champignons), l'ergostérol (composé stéroïdien contenu dans la paroi cellulaire des
champignons), les toxines et les COVM (composés organiques volatils microbiens tels certains aldéhydes,
alcools, esters, cétones). En culture, les colonies peuvent se développer non seulement à partir de spores,
mais également à partir de fragments de mycélium.
Le nombre et la dissémination aérienne des spores libérées peuvent varier en fonction du type de dommage
de moisissure. Pour évaluer les sources de moisissures intérieures, il est important, par conséquent, de
différencier les espèces de moisissures individuelles d'après le mode de dispersion de leurs spores.
L'expérience a montré qu'une contamination, même mineure, de matériaux par des moisissures peut donner
lieu à des niveaux de moisissures élevés dans l'air intérieur si les espèces impliquées ont des spores sèches
présentant de bonnes capacités de dispersion aérienne (par exemple Penicillium et Aspergillus). Au contraire,
les concentrations de spores en suspension dans l'air sont beaucoup plus faibles lorsque les matériaux sont
colonisés, par exemple, par des moisissures des genres Acremonium, Fusarium ou de l'espèce Stachybotrys
chartarum, qui ont des spores relativement grandes noyées dans des substances gluantes et possèdent par
conséquent de faibles capacités de dispersion aérienne.
En outre, il convient de prendre en compte le fait que les spores de moisissures ne se présentent pas
nécessairement sous forme de spores individuelles dans l'air ou dans la poussière sédimentée, mais
également sous forme d'agrégats de spores ou de spores portées par des particules. Selon la méthode
d'analyse, elles sont déterminées en tant que spores isolées ou en tant qu'agrégats de spores. Les matériaux,
l'air intérieur et la poussière domestique contiennent non seulement des spores de moisissures cultivables,
mais également des spores de moisissures non cultivables dont certaines ont les mêmes effets allergènes et
toxiques que les spores cultivables. Pour cette raison, des techniques permettant de déterminer au
microscope aussi bien les moisissures cultivables que non cultivables ont été mises au point.
La détection et la différenciation des moisissures sont effectuées soit après culture, en se basant sur des
critères morphologiques, des réactions biochimiques et/ou des techniques moléculaires, soit par examen
© ISO 2012 – Tous droits réservés 5
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ISO 16000-19:2012(F)
direct au microscope. La différenciation basée sur la structure morphologique (examen macroscopique,
stéréo-microscopie et microscopie), soit après une culture préalable, soit par microscopie directe, demeure
l'approche la plus fréquente pour la détection des moisissures.
Il existe également d'autres méthodes d'analyse basées sur le dosage des composants cellulaires et des
[17]
métabolites des moisissures, tels que les -glucanes, l'ergostérol, les toxines et les COVM . Le dosage de
ces composés ne sert toutefois que d'information complémentaire.
Les méthodes d'échantillonnage utilisées pour la détection des moisissures sont déterminées par l'objectif de
l'investigation. Selon la méthode d'échantillonnage, les moisissures subissent un stress de prélèvement
pendant le prélèvement et la préparation des échantillons, ce qui peut entraîner leur dessèchement ou leur
mort. Les facteurs influençant l'aptitude à la culture des spores de moisissures sont leur état physiologique
ainsi que le milieu de culture utilisé. Certaines espèces de moisissures ne peuvent pas du tout être cultivées
dans des conditions de laboratoire.
NOTE Les genres Stachybotrys et Chaetomium se développent difficilement et leur sporulation sur la gélose DG18
est médiocre, voire inexistante. Par conséquent, l'utilisation de ce milieu de culture pour une analyse basée sur la culture
de ces genres est déconseillée (voir l'ISO 16000-17).
Pour un résumé de la documentation, voir les Références [8] à [10], [12] et [14] à [18].
5 Méthodes d'échantillonnage et de détection
Selon l'objectif de l'investigation, on peut échantillonner et analyser les matériaux (voir l'ISO 16000-21, en
préparation), l'air (voir l'ISO 16000-16 et l'ISO 16000-18) et la poussière domestique pour détecter des
moisissures cultivables (voir l'ISO 16000-17). Les moisissures peuvent également être quantifiées et, dans
une certaine mesure, différenciées sans culture préalable. Pour ce faire, les spores de moisissures en
suspension dans l'air sont recueillies sur des filtres ou directement sur une lame de microscope à revêtement
adhésif; après quoi elles sont colorées puis examinées par microscopie directe.
L'Annexe B fournit une vue d'ensemble des dispositifs de mesurage du nombre total de spores les plus
courants, ainsi que des dispositifs de prélèvement par filtration et par impaction et des méthodes d'analyse
respectives.
6 Stratégie de mesurage
6.1 Aspects généraux
Il n'existe pas de mode opératoire normalisé pour le mesurage et l'évaluation d'un dommage de moisissure.
Le type et le nombre de mesurages, de même que les méthodes d'analyse utilisées, sont déterminés par les
circonstances qui ont déclenché l'investigation et par les objectifs de cette dernière. Une inspection visuelle
sur le terrain (visite de diagnostic) par des professionnels techniquement qualifiés avant le prélèvement est
une condition préalable indispensable pour la détection et l'évaluation des sources de moisissures dans les
environnements intérieurs. Outre de bonnes connaissances en génie et en physique du bâtiment, il convient
que les professionnels chargés de l'inspection disposent d'une formation suffisante en matière d'hygiène de
l'air intérieur et de microbiologie.
Les investigations sont menées dans le but de localiser les sources de moisissur
...
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