32873458_PhDThesisE.M.Coetzee.docxfor shipping containers and their in-transit fumigation
This thesis is presented for the degree of
Doctor of Philosophy at Murdoch University
By
Murdoch University
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Declaration
I declare that this thesis is my own account of my research and has not been submitted for
a degree at any tertiary education institution.
Signature: Eugene Marco Coetzee Date: 30/11/2020
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Abstract
Fumigation is required by most governments as an appropriate biosecurity measure to
exterminate pests in shipping containers. The use of shipping containers for cargo
transportation has the potential to transport pests from infested to non-infested areas. Initially
fumigation trials were conducted in stationary 20ft shipping containers. Four species of
stored product insect adults were used for bioassays namely, the Cigarette beetle,
Lasioderma serricorne (F.), Rice weevil, Sitophilus oryzae (L.), Warehouse beetle,
Trogoderma variabile (Ballion) and the Lesser grain borer, Rhyzopertha dominica (F.).
Ethyl formate (90 g/m3) was purged with nitrogen (99.5%) into the containers. Ethyl formate
concentration inside containers and the surrounding environment was monitored at timed
intervals. Fumigation achieved target concentration × time (Ct) products of 437.54 - 449.19
g h/m3 in the containers, which can exterminate all life stages of most common insect pests.
Ethyl formate distributed evenly in the containers within 0.5 hours after application with a
variation <3%. This study demonstrated that onsite generation of a non-flammable ethyl
formate + nitrogen fumigant can be achieved and that this new ethyl formate + nitrogen
application can be used as a quarantine pre-shipment treatment for controlling insect pests
in shipping containers. Ventilation of shipping containers after fumigation to remove ethyl
formate from containers was successful with a threshold limit value (TLV) of zero achieved
after approximately 15 minutes. The levels of ethyl formate in the surrounding workspace
were <0.5 parts per million (ppm) during application, fumigation and aeriation which is 200
times below the required TLV level of 100ppm.
In-transit ethyl formate and nitrogen fumigation trials were conducted in 20ft shipping
containers during a two-day road journey in both September and December 2017. Ethyl
formate (90 g/m3) was purged with nitrogen into the containers. Ethyl formate concentrations
inside the containers and the surrounding environment were monitored at timed intervals
throughout the journey. Fumigation achieved concentration × time (Ct) products in the
containers during the journey, which can exterminate all life stages of most common insect
pests. Levels of ethyl formate in the environment between 1-15 meters downwind from the
containers and truck driver's cabin were less than 0.5ppm at each of the timed intervals. The
study confirms that ethyl formate can safely be used as an in-transit fumigant.
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Further trials were conducted in 20ft shipping containers during a two-day road journey and
a one-day sea journey. Ethyl formate concentrations inside the containers and the
surrounding environment on the vessel were monitored at timed intervals throughout the
overnight sea voyage. This research adds to the previous research for in-transit fumigation
with ethyl formate and nitrogen via road and has successfully demonstrated that in-transit
fumigation with ethyl formate and nitrogen via the marine sector is effective and also safe
with no detectable risk to the public, crew members on the barge or workers on Barrow
Island throughout the journey. In addition, all tested containers were ready to be opened and
unloaded with 5-10 minutes aeration or without aeration upon arrival.
In conclusion, this study indicates that in-transit ethyl formate and nitrogen technology has
the potential to deliver cost savings in the fumigation process through reduction of the labour
cost, elimination of the time a container and cargo must remain stationary in a fumigation
yard and a significant decrease in total supply chain time (between container packing and
receival).
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1.1 Barrow Island ................................................................................................................ 23
1.1.2 Biosecurity to protect Barrow Island ................................................................... 24
1.1.3 Barrow Island pathways for non-indigenous species .......................................... 26
1.2 Invertebrate pests and non-indigenous invasive species............................................ 27
1.2.1 Invertebrate pests ................................................................................................... 27
1.3 Fumigants and fumigation – as mitigation measure .................................................. 29
1.3.1 Fumigants .............................................................................................................. 29
1.3.2.1 In-transit fumigation of shipping containers............................................ 38
1.4 Ethyl formate ................................................................................................................. 39
1.4.2 Toxicity of ethyl formate to insect pests .............................................................. 42
1.4.3 Determination of concentration x time (Ct) ........................................................ 44
1.4.3.2 Well sealed enclosures that have passed a pressure test………………...45
1.4.3.2 Under Gas proof sheeting where gas losses are high……………………45
1.4.4 Residues .................................................................................................................. 46
1.6 Fumigants and synergism ............................................................................................. 48
1.7 Ventilation, safety and the environment ..................................................................... 49
1.8 Aims of this PhD thesis ................................................................................................. 50
Aims of this study in chapters ....................................................................................... 51
Chapter Two
Evaluation of ethyl formate + nitrogen as a suitable treatment fumigant
for 20ft shipping containers loaded with general freight…………………….52
Chapter 2. Declaration of contribution ............................................................................. 53
Abstract ................................................................................................................................ 54
2.2.1 Insect species for bioassays .................................................................................... 58
2.2.2 Ethyl formate and apparatus ................................................................................ 59
2.2.3 Shipping containers ................................................................................................ 59
2.2.5 Installation of gas sampling lines and insect cages in containers....................... 61
2.2.6 Measurement of temperature and relative humidity .......................................... 63
2.2.7 Measurement of in-container and environmental concentrations of ethyl
formate ..................................................................................................................... 63
2.2.8 Analysis of ethyl formate, ethanol and formic acid in drinks ............................ 64
2.2.9 Generation of a non-flammable ethyl formate fumigant formulation .............. 64
2.2.10 Insect bioassay ...................................................................................................... 66
2.2.13 Statistical Analysis ............................................................................................... 68
2.3.1 Variation of temperature and relative humidity ................................................. 68
2.3.2 Reliability of GC method for analysis of ethyl formate ...................................... 68
2.3.3 Concentration of ethyl formate in the container and environment ................... 69
2.3.4 Residues and natural levels of ethyl formate in drinks and food ...................... 72
2.3.5 Efficacy of ethyl formate fumigation .................................................................... 73
Conclusion ............................................................................................................................ 73
Chapter Three
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Commercial trials evaluating the novel use of ethyl formate for in-transit
fumigation of shipping containers…………………………………………………75
Chapter 3. Declaration of contribution ............................................................................. 76
Abstract ................................................................................................................................ 78
3.2.1 Shipping containers ................................................................................................ 81
3.2.4 Gas sampling from container and environment .................................................. 82
3.2.5 Monitoring of ethyl formate in container and environment .............................. 83
3.2.6 Preparation of gas standard for DPS portable GC ............................................. 85
3.2.7 Comparison of the different equipment used for analysis of ethyl formate ..... 85
3.2.8 Determination of Concentration × time (Ct) ........................................................ 87
3.2.9 Statistical analysis. ................................................................................................. 87
3.3 Results ............................................................................................................................ 87
3.3.1 Sensitivity and accuracy of the G460 ethyl formate monitor and the DPS
portable GC ............................................................................................................. 87
3.3.2 Variation of temperature and relative humidity ................................................. 88
3.3.3 Safety of ethyl formate fumigation during the in-transit journey ..................... 89
3.3.4 Concentration of ethyl formate in the stationary container and containers
during the in-transit journey ................................................................................. 92
3.3.5 Efficacy of ethyl formate fumigation during the in-transit journey.................. 96
3.4 Discussion ....................................................................................................................... 99
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3.4.1 Efficacy of in-transit fumigation on ethyl formate concentration and its Ct
products ................................................................................................................. 100
3.4.2 Efficacy of Ct products achieved during in-transit fumigation ....................... 101
3.4.3 Effect of in-transit fumigation with ethyl formate on environment and worker
safety ...................................................................................................................... 101
3.4.4 The advantages of using in-transit ethyl formate+nitrogen fumigation ......... 101
Conclusion .......................................................................................................................... 102
Chapter Four
In transit fumigation of shipping containers with ethyl formate + nitrogen
by road and continued by sea .................................................................................. 103
Chapter 4. Declaration of contribution .......................................................................... 104
Abstract .............................................................................................................................. 106
4.2.1 Shipping containers .............................................................................................. 108
4.2.2 Gas tightness ......................................................................................................... 109
4.2.4 Installation of gas sampling lines in containers ................................................. 110
4.2.5 Installation of gas sampling lines on the vessel .................................................. 110
4.2.6 Fumigant and fumigation .................................................................................... 111
4.2.7 Gas sampling from containers and the environment ........................................ 112
4.2.7.1 Gas sampling from containers ......................................................................... 112
4.2.8 Gas sampling during voyage ............................................................................... 112
4.2.9 Gas sampling during aeration ............................................................................. 113
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4.2.10 Analysis of ethyl formate in container and environment ............................... 113
4.2.11 Statistical analysis .............................................................................................. 114
4.3.2 Concentration of ethyl formate in the containers ............................................. 115
4.3.3 Ambient ethyl formate concentrations surrounding containers during the
voyage on the barge .............................................................................................. 117
4.3.4 Ambient ethyl formate concentrations surrounding containers during aeration
at Barrow Island ................................................................................................... 118
5.1 General discussion ....................................................................................................... 123
BIOMOD - Biomolecular Design
BWI - Barrow Island
°C - Degrees Celsius
CBD - Convention on Biological Diversity
CFTRI - Central Food Technological Research Institute
cm - centimetre
Ct - Concentration by time
DPAW - Department of Parks and Wildlife
EF - Ethyl Formate
EPA - Environment Protection Agency
FID – Flame Ionization Detector
GC-MS - Gas Chromatography - Mass Spectrometer
GRAS - Generally Regarded As Safe
g/m3 - gram per cubic meter
g m-³ - gram per cubic meter
g h/m3 - gram hour per cubic meter
g h m-³ - gram hour per cubic meter
GP - General Purpose
ISPM - International Standards for Phytosanitary Measures
ISSG - The Invasive Species Specialist Group
JEFCA - Joint FAO/WHO Expert Committee on Food Additives
Kc - reaction equilibrium constants
mL - milli Litre
MNS - Membrane Nitrogen Separator
MRL - Maximum Residue Limits
N2 - Nitrogen
NTIS - National Technical Reports Library
NOHSC - National Occupational Health and Safety Commission
NZMAF- New Zeeland Ministry of Agriculture and Forestry
NZME - New Zealand Ministry for the Environment
o.d. - outer diameter
Pa - Pascal
SOP - Standard operating Procedure
TEU - 20-foot Equivalent Unit
UNEP - United Nations Environment Programme
USA - United States of America
US$ - United States Dollar
USBC - Environmental and economic costs of nonindigenous species in the U.S.
USBS - United States Bureau of the Census
USDA - United States Department of Agriculture
USEPA - United States Environmental Protection Agency
UK - United Kingdom
Table 2-1. Expanded gas sample location abbreviations.
Table 2-2. The variation of ethyl formate concentrations and concentration × time (Ct)
product (g h/m3) compared at different location in containers and environmental
levels of ethyl formate at different locations downwind during application,
fumigation and aeration.
Table 2-3. Ethyl formate and its break down compounds (ethanol and formic acid) for the
fumigated drinks and food, as well as the unfumigated control.
Table 3-1. Comparison of sensitivity and working range for different instruments for the
analysis of ethyl formate gas.
Table 3-2. Environmental levels of ethyl formate at different locations downwind during
truck stops for September 2017 trials.
Table 3-3. Environmental levels of ethyl formate at different locations downwind during
truck stops for December 2017 trials.
Table 3-4. The variation of ethyl formate concentrations compared at different locations in
containers, between two containers during the same trip in September and
December trials and between September and December trials.
Table 3-5. Comparison of Concentration ×time (Ct) product (g h/m3) for different exposure
times (hours) and mortality of common stored product insect pests in our study
and previous studies.
Table 4-1. Concentration of ethyl formate in the containers at different locations upon arrival
at Dampier and during the voyage to Barrow Island.
Table 4-2. Environmental gas levels of ethyl formate at different locations during the voyage
to Barrow Island.
Table 4-3. Environmental gas levels of ethyl formate at different locations during aeration
at Barrow Island.
List of Figures
Figure 1-1. Barrow Island is located about 56 kilometres west off the mainland.
Figure 1-2. Barrow Island is a Class A reserve and is one of the most valuable biodiversity
conservation areas in the world.
Figure 2-1. Insect colonies were established at the Postharvest Biosecurity and Food Safety
Laboratory, Murdoch University Laboratory, Perth, Australia.
Figure 2-2. Container (20ft) loaded with palletised drinks and singles being placed randomly
throughout the container.
Figure 2-3. Schematic representation of a placement of fumigant monitoring ports ( ),
temperature and relative humidity monitors ( ) and insect cages ( )in
containers.
Figure 2-4. The concentrations of ethyl formate in the fumigated containers were analysed
with a Gas Chromatograph (GC) equipped with a Flame Ionisation Detector
(FIP).
Figure 2-5. Schematic representation of an ethyl formate vaporiser coupled with a nitrogen
generator for onsite generation of a non-flammable ethyl formate + nitrogen
fumigant formulation.
Figure 2-6. Ethyl formate vaporiser coupled to a 20ft shipping container.
Figure 2-7. GC spectra shows that ethyl formate (at 27.306), ethanol (at 24.123) and formic
acid (at 21.860) were detected and completely separated.
Figure 3-1. Placement of fumigant monitoring ports (- -), and temperature and relative
humidity monitors (- -) in containers.
Figure 3-2. Sampling and monitoring of ethyl formate in containers and environment, during
in-transit fumigation by road.
Figure 3-3. Temperature (C) and relative humidity (r.h.) data from containers 1 and 2
during the a) September and b) December trials.
Figure 3-4. In transit fumigation of cargo container using ethyl formate showing
concentrations (g h/m3) in various parts of the container during the September
trial in containers 1 and (LRB = Left rear bottom, RRT = Right rear top, RMB
= Right middle bottom, MMM = Middle middle middle, LFT = Left front top,
FBR = Front bottom right, Cargo = cargo).
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Figure 3-5. In transit fumigation of cargo container using ethyl formate showing
concentrations (g h/m3) in various parts of the container during the December
trial in containers 1 and 2 (LRB = Left rear bottom, RRT = Right rear top,
RMB = Right middle bottom, MMM = Middle middle middle, LFT = Left
front top, FBR = Front bottom right, Cargo = cargo).
Figure 3-6. Non in-transit stationary container fumigation during the December 2017 trial.
Figure 4-1. Placement of fumigant monitoring ports (- -), and temperature and relative
humidity monitors (- -) in containers.
Figure 4-2. TOLL Astrolabe vessel (Toll Astrolabe) were used for the trial.
Figure 4-3. Schematic representation of the shipping container stack on the barge with gas
sampling ports (1-13).
• Gas sample ports 1-8 (- -) located at head height around the stack
• Gas sample ports 9 and 10 (- -) located in between containers
• Gas sample ports 11 (- -) located in barge tunnel
• Gas sample ports 12 and 13 (- -) located in cabin
• Two containers at the front and two containers at the rear (lowest level) were
the fumigated containers.
Figure 4-4. Temperature in the headspace of the containers during the period of fumigation
(Container #1–blue; #2-orange; #3-yelow and #4-black)
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Publications
E. Marco Coetzee, James Newman, Bob Du, YongLin Ren and Simon J. McKirdy. 2020.
Evaluation of ethyl formate + nitrogen as a suitable treatment fumigant for 20ft
shipping containers loaded with general freight. (Submitted for publication: Journal of
Environmental Science and Health, Part B).
E. Marco Coetzee, James Newman, Grey T. Coupland, Melissa Thomas, Johann van der
Merwe, YongLin Ren and Simon J. McKirdy, 2019. Commercial trials evaluating the
novel use of ethyl formate for in-transit fumigation of shipping containers. (Published:
Journal of Environmental Science and Health, Part B).
DOI: 10.1080/03601234.2019.1631101
Eugene M. Coetzee, Bob Du, Melissa L. Thomas, YongLin Ren and Simon J. McKirdy,
2020. In-transit fumigation of shipping containers with ethyl formate + nitrogen on road
and continued journey on sea. (Published: Journal of Environmental Science and Health,
Part B). DOI: 10.1080/03601234.2020.1786328
Is in-transit fumigation of shipping containers using ethyl formate safe and effective? The
12th International Working Conference on Stored Product Protection (IWCSPP), Berlin,
Germany, 7 -11 October 2018.
The evaluation of ethyl formate for biosecurity treatment of invertebrate pests in shipping
containers. Chevron presentation. Harry Butler Institute, Murdoch University, Perth,
Australia, 2017.
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Acknowledgments
I thank Chevron for support to pursue my research on the fumigation of shipping containers.
I thank Murdoch University and the Harry Butler Institute for providing research materials,
laboratories and technical support.
I express my sincere gratitude to my principle supervisor, Professor Simon McKirdy, for his
encouragement, guidance, commitment and continued support in many ways throughout my
study. I thank my co-supervisor, Professor YongLin Ren, for sharing much of his time
explaining and discussing the various aspects of my study. His extensive and expert
knowledge on both the subjects of ethyl formate and fumigation made it possible for me to
make a valuable contribution towards a safe and efficient technology to fumigate shipping
containers.
Many thanks to Bob Du for data analysis and technical support in the laboratory with the
GC-MS, GC-FID and LC-MS equipment, as well as for his help with the collection of
samples on the TOLL Astrolabe. A very helpful perfectionist.
I am thankful to James Newman and Supaporn Yamaungmorn for their help with the
collecting of samples during the September trial and James Newman and Graeme George
for their help with the collection of samples during the December trial.
I thank the Gorgon Gas Development and its contractors for the use of their supply chain
that enabled these trials to be undertaken. I thank Toll for the provision of the containers and
TOLL Astrolabe for the provision of the barge and accommodation. I thank Sadliers for the
provision of the trucks and drivers.
I thank my wife Yvette for her patience, encouragement and support throughout my studies
during the years.
General introduction and literature review
The focus of this study was the biosecure transport of cargo in shipping containers to Barrow
Island, Western Australia (Chevron, 2014b, Scott et al., 2017). All cargo, including food,
linen, tools, machinery, etc., that can fit into a container are being shipped to Barrow Island
in shipping containers. These containers must be quarantine compliant to prevent any
contamination of any invertebrates, vertebrates or plant material to the pristine environment
of Barrow Island (Chevron, 2014a and b).
This research investigated the use of ethyl formate and nitrogen as a fumigant to replace
existing expensive, toxic and environmentally unfriendly fumigants such as methyl bromide,
sulfuryl fluoride and phosphine in shipping containers (Bond, 1984, Banks, 1988, Banks,
1994, Reichmuth, 1998, Banks et al., 1979, USDA, 2016, Armstrong et al., 2014).
The research investigated the required concentration of ethyl formate and over what period
of exposure this fumigant will be effective to kill most invertebrate pests. Furthermore, the
study investigated if ethyl formate and nitrogen would be a safe fumigant for workers from
loading and unloading the containers and if in-transit fumigation in shipping containers
could save costs. Currently, there are no studies in the literature that address fumigation of
ethyl formate and nitrogen in shipping containers. There are also no studies that have been
done on in-transit fumigation of shipping containers by road or by sea with ethyl formate
and nitrogen.
The gaps identified in the current knowledge showed that there is not currently a fumigant
that is safe, environmentally friendly and cost effective, but still effective to kill a wide range
of invertebrate pests (Muthu et al., 1984, Banks et al., 1996, Damcevski et al., 2002, Haritos,
2005). Some fumigants are being phased out for being toxic, environmentally unfriendly and
expensive (Banks, 1994, Annis, 1987, Damcevski et al., 2002, Simpson et al., 2007,
Armstrong et al., 2014). Most of the above-mentioned studies regarding ethyl formate and
invertebrates have been done on stored grain insects pests to protect the grain industry, but
none on other invertebrate pests that could be harmful to biodiversity.
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1.1.1 Biodiversity of Barrow Island
Barrow Island is situated on the north-west coast of Australia (56 km from the coast) (Figure
1-1) and is one of the world’s most significant conservation areas with a rich diversity of
plants and animals (Jarrad et al., 2010, Chevron, 2015, Chevron, 2014a) and the fauna and
flora has been described in detail (Butler, 1987). The island is well known for its abundant
mammals and rich bird and reptile fauna with a unique composition of subterranean animals
as well as vegetation communities (Scott et al., 2017). It is also classified as a Class A nature
reserve having been classified in 1910 (Dudley, 2008) (Figure 1-2).
Figure 1-1. Barrow Island is located about 56 kilometres west off the mainland.
Barrow Island Nature Reserve is one of the largest land masses in the world that has no
introduced vertebrates (Conservation Commission of Western Australia, 2003). The island
has an area of approximately 23,000 ha and is the second largest island in Western Australia
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(Chevron, 2014a). It is one of the oldest and most valuable biodiversity conservation reserves
in the world and only a few non-indigenous species (NIS) are present on the island (Jarred
et al., 2010, Scott et al., 2017, Dudley, 2008). Fortunately, the island has retained its high
conservation values (DPAW, 2015, Butler, 1987, Scott et al., 2017, Chevron, 2014a).
Figure 1-2. Barrow Island is a Class A reserve and is one of the most valuable biodiversity
conservation areas in the world.
1.1.2 Biosecurity to protect Barrow Island
The Western Australian State and Commonwealth Governments approved the Gorgon
Project in 2009 (Chevron, 2014a). At the time it was one of the world’s largest natural gas
projects and the largest single-resource development in Australia’s history (Chevron, 2015).
A Liquefied Natural Gas (LNG) plant, Australia’s largest infrastructure development, was
built on the island on condition that no non-indigenous species become established on the
island (Scott et al., 2017, DPAW, 2015, Butler, 1987, Chevron, 2014a, Caley et al., 2006,
EPA, 2007). The lack of data on the invasions of introduced organisms to the island posed a
biosecurity threat to the biodiversity of Barrow Island (Smith et al., 1999, Caley et al., 2006,
Sahlin et al., 2011, Williamson et al., 1996). This threat mandated that the Gorgon Project
implement an exceptionally high level of biosecurity (Scott, et al., 2017).
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During the construction phase of the LNG project, different modes of introduction were
identified for cargo and personnel, which included personnel and luggage, and domestic
vessels and food and perishables (Chevron, 2014c). Therefore, detecting, intercepting and
identifying potential non-indigenous species arriving on the island became a priority in
biosecurity surveillance and management of the island (Government of Western Australia,
2009, Chevron, 2014c).
A Quarantine Management System (QMS) was developed to protect the environmental
values and unique ecosystem of Barrow Island. The QMS is governed by the Australian
Biosecurity Act of 2015 from which the Barrow Island Act (2003) emanated (Government
of Western Australia, 2003a, Chevron, 2014c). The QMS is a risk-based quarantine approach
with more than 300 procedures, specifications, checklists and guidelines (Stockloza, 2005,
Chevron, 2014b).
According to McKirdy et al., (2014), quarantine is “the official confinement of regulated
articles for observation and research or for further inspection, testing, or treatment” and
biosecurity is “a set of measures that protect the economy, environment, and community
from harm from pests and diseases”. The protection of the biodiversity of Barrow Island is
more than just “official confinement of regulated articles for observation and research or
for further inspection, testing, or treatment” but more like a “set of measures that protect
the economy, environment, and community from harm from pests and diseases” and
quarantine is an integral part of biosecurity (McKirdy et al., 2014, Sharma et al., 2008).
According to Beale et al., (2000), biosecurity has a more proactive approach which focus
more on risk management and border management. The term biosecurity is therefore a much
more comprehensive term which describe the protection of the unique biodiversity of
Barrow Island better.
Since the Gorgon Project began, there have been zero introductions or proliferations of non-
indigenous species on Barrow Island or in its surrounding waters. The QMS applies to all
Gorgon Project operations whether in Australia, on Barrow Island or at any of the overseas
construction facilities (Chevron, 2015). The QMS addresses invasion by invertebrates,
26
vertebrates, plants and marine invasive pests and all methods are well documented
(Stockloza, 2005, Chevron, 2014b, Chevron, 2014c, Chevron, 2015).
Chevron has preserved the integrity of Barrow Island’s environment by maintaining natural
habitats and preventing the introduction of non-indigenous plants, animals and micro-
organisms by implementing the QMS as invasive species are one of the greatest threats to
Barrow Island biodiversity (Chevron, 2015, Scott et al., 2017).
1.1.2 Barrow Island pathways for non-indigenous species
A pathway can be defined as the route from a source region to the region of destination
(Lockwood et al., 2007). Numerous studies have investigated non-indigenous species
invasions by identifying the manners and routes (pathways) to their destination (Hulme et
al., 2008, Pysek et al., 2011).
Horticulture, landscaping and agriculture are all pathways for non-indigenous species to
islands (Wasowics, 2014) and globalization has increased the frequency by which non-
indigenous species are introduced, which can change natural biogeographical patterns
(Lockwood et al., 2007). The potential for non-indigenous species to invade remote islands
is often limited due to the nature of their remoteness and pathways of entry (Wasowics,
2014).
The operators of the Gorgon Project, Chevron Australia Pty Ltd, identified thirteen pathways
by which non-indigenous species might enter Barrow Island, including food, luggage,
marine vessels and aerial transfers. These pathways are part of the QMS protocol of
interventions that span pre-border (before goods and personnel reach the island), border (on
arrival) and post-border (outside the development site) (Scott et al., 2017, Chevron, 2014 a
and b, Chevron, 2015).
All passengers, cargo and equipment are screened during the pre-border stage prior to arrival
at Barrow Island. Detector dogs screen passengers and their luggage for any non-compliant
items such as fruit, insects, seeds, etc. Pest control measures are applied to oversized
machinery and equipment, including small buildings. All fresh food destined for Barrow
27
shipped. Anything non-compliant is remediated or returned to the mainland (Chevron, 2015,
Scott et al., 2017, Stockloza, 2005, Chevron, 2014c).
1.2 Invertebrate pests and non-indigenous invasive species
1.2.1 Invertebrate pests
It is important to understand the biology and behaviour of invertebrate pests, as well as the
kinds of damage they can cause in order to implement an integrated pest management plan
to achieve a system that optimizes the use of natural resources, protects the environment and
maximizes output in a sustainable way (Garcia-Lara et al., 2016).
Invertebrate pests are also destructive to fruits and vegetables and have a big impact on the
international trade market as infestations reduce the economic value of the product (Johnson,
2013). As populations grow there is an ever increasing demand on food resources and
environmental changes has resulted in an increase in the number of species that competes
for food resources (Barkai et al., 2017). There are various methods to control these pests,
which include cultural control, physical control, physical barriers, as well as chemical and
biological control (Saxena et al., 2003, Garcia-Lara et al., 2016, Hagstrum et al., 2006,
Havila, 2011, Keshavareddy et al., 2016, Armstrong et al., 2014).
1.2.2 Non-indigenous invasive species
Invertebrates form a large part of the non-indigenous fauna worldwide, but invasive
invertebrate pests appear to have received disproportionately less attention regarding their
effects on the environment compared to plants, vertebrates, or aquatic organisms (Parker et
al., 1999, Levine et al., 2003, Long, 2003). The ecological impacts by these species can have
genetic effects, effects on individuals and populations, as well as effects on ecosystem
processes (Parker et al., 1999).
Invasive species have been referred to as exotic species, non-indigenous species
(Williamson, 1996, Wittenberg et al., 2001) and invasive alien species (IAS) (Pimentel et
al., 2000). According to Pimentel et al., (2000), invasive pest species are one of the real
threats to native biodiversity and if they become locally dominant, they invade natural
communities and are referred to as invasive alien species. Invasive alien species constitute
the second most serious threat to biodiversity habitat destruction and include introduced
plants, animals and other organisms. The establishment and spread of these organisms
threaten ecosystems, habitats, and other species (CBD, 2001, Pimental et al., 2000, Lowe et
al., 2000). Alien and exotic species will be referred to as non-indigenous species hereafter.
Invasion by non-indigenous species has been recognized as a major threat to global
biodiversity (Williamson, 1996, Wittenberg et al., 2001, Pimentel, 2002) and the economic
impact of these species is a major concern and can cause major environmental damage
(Pimentel et al., 2000). The management and control of these species is a big challenge for
the agricultural sector, as well as for conservation biologists. Invasive species can change
the structure and species diversity of ecosystems by repressing native species, either directly
by out-competing them for resources or indirectly by changing the way nutrients are cycled
through the system (Drake et al., 1999).
An invasive non-indigenous species spreads from one geographic region to another where it
does not occur naturally and can be a major cause of crop loss and can adversely affect food
security (Cook et al., 2011). In the United States crop and forest production losses from
invasive insects and pathogens have been estimated at almost US$40 billion per year
(Pimentel et al., 2005). Biotic invaders are species that establish a new range in which they
proliferate, spread and persist to the detriment of the environment and are the most important
ecological outcomes from alterations in the distribution of the environmental biota through
human transport and commerce. Few, if any, areas in the world remain sheltered from these
invasions (Mack et al., 2000).
A few studies have been conducted on the arrival and the establishment of invasive species,
often with a focus on the threat from individual species to a specific country (Bourdot et al.,
2012, Hill et al., 2012). Various studies have been conducted on the establishment of broader
species assemblages (Thuiller, 2003, Thuiller, 2005, Bomford et al., 2009, Paini et al., 2010).
29
According to Lowe et al., (2000), there are more than 100 non-indigenous species which
include micro-organisms, macro fungi, plants, invertebrates, amphibians, fishes, birds,
reptiles, and mammals that are recognised as most problematic. In addition to the invasion
of these invasive species from different countries, these invasions can also occur from one
geographic location to another within the same country (Ananthakrishnan, 2009, Gallardo et
al., 2019, Kumschick et al., 2015). The increase of non-indigenous species can cause
homogenizing the world’s flora and fauna and such bio-invasion may be regarded as a form
of biological pollution, as well as having a significant component on global change, which
can cause species extinction (Drake et al., 1999).
Most plant and vertebrate introductions have been intentional, while most invertebrate
introductions were accidental (Pimentel et al., 2005). The rate of introductions with invaders
have increased significantly because of population growth, movement of people and the
movement of goods and materials. Trading amongst nations have increased, which created
increased opportunities for unintentional introductions (Bryan, 1996, USBC, 1998).
The prediction of impacts of invasion by invasive species in natural environments is more
difficult than in agricultural systems as natural systems have greater ecological complexity,
which affects the prediction of the direct impacts on the environment. It is also more difficult
to anticipate the pathways of invasion, the range of possible invaders or the geographic
boundaries of potential impacts (Myers et al., 2000).
1.3 Fumigants and fumigation as mitigation measure
1.3.1 Fumigants
A fumigant is a chemical that will exist as a lethal gas at certain temperatures and pressures
(Davis, 2003). A fumigant can also be described as a chemical which, at a required
temperature and pressure, will exist as a gas in enough concentration to be lethal to a given
pest organism. (FAO, 2007, Bond, 1984) and can diffuse through air and permeate most
products which includes food and non-foods, as well as most packaging materials, except
for metals. This permeation disrupts the biological processes of the organisms, which is
lethal to most living organisms, both pests and non-pests (Davis, 2003).
30
According to the USDA, (2016), the ideal fumigant would be easily and cheaply generated,
easily detected by human senses, easily diffuses and rapidly penetrates commodities,
harmless to foods and commodities, highly toxic to the target pest, inexpensive, insoluble in
water, non-explosive, non-flammable, non-persistent, non-toxic to plants and vertebrates
and stable in the gaseous state. Unfortunately, not one fumigant has all the above properties.
According to Davies, (2003), the ideal fumigant should have a low cost, should be highly
toxic to all stages of the target pests and not hazardous to humans and domestic animals. It
should be highly volatile with good penetration, but not be excessively sorbted on and in the
commodity. It should also be easily detected and be non-corrosive, non-flammable and non-
explosive under practical conditions. It should be of low molecular weight to aid diffusion
and penetration, possess a good storage life and be easily removed by aeration. It should also
not affect seed viability adversely, produce no quality changes in raw and processed foods
and be easy and safe to apply. Again, not one fumigant has all the above properties.
Price, (1985), describes a true fumigant as a toxic chemical that controls the target pest while
the chemical is in the gaseous state. The fumigation process involves a lethal concentration
of toxic gas for a long enough period to kill the target pests (Stark, 1994). Bond, (1984),
believes that although the toxicity of some insecticides can be attributed to vapour action,
they are not considered true fumigants. The lethal effect of an insecticidal fumigant is
dependent on the extent of the total uptake of the fumigant during the exposure period
(Outram, 1967a).
Fumigants vary in their mode of action, where some will kill rapidly, while others will kill
slowly (Armstrong et al., 2014). Some fumigants may have a paralysing effect on the pest
while others will kill the pest. Some fumigants have no effect on commodities, while others
are detrimental, even at low concentrations. Some commodities absorb the fumigants and
needs to be aerated after fumigation (USDA, 2016, Armstrong et al., 2014). Fumigants may
possess fungicidal, insecticidal, bactericidal and nematicidal properties (Xin et al., 2008).
Numerous fumigants have been eliminated as commercial fumigants because of their
unfavourable properties such as residues, work safety and their impact on the environment
(Banks et al., 1979, Bond, 1984, Banks, 1988).
31
Fumigants are seldom commercially available as gasses and is usually available in cylinders
of pressurized liquids that become a gas as they are released at normal atmospheric pressure
(Armstrong et al., 2014). Phosphine is one of the examples of a fumigant that is available in
a solid form, as well as a gas form and requires exposure to atmospheric moisture to activate
the gas (Davis, 2003). Fumigants are efficacious and important pesticides for those involved
in producing, storing, transporting and processing of products. Although many chemical
compounds are efficient to act as fumigants, some are corrosive to metals, dissolve plastics,
are destructive to plant tissues, or leave unacceptable residues in the treated commodity
(Bond, 1984, Armstrong et al., 2014). One of the main reasons for the wide acceptance of
fumigants has been their ease of application and the availability of different application
methodologies (Davis, 2003, Banks, 1994, Banks, 2002).
Some fumigants with excessive hazard potential have been restricted or prohibited so that
they are no longer widely used for pest control in some countries (Bond, 1984, Banks, 1994,
Reichmuth, 1998). The Montreal Protocol limited the use of methyl bromide because it is
ozone depleting (UNEP, 2010). The development of new fumigants may not be realistic
because the production of toxicology and residue data for registration of a new compound is
expensive and time consuming (Reichmuth, 1998, Armstrong et al., 2014).
Only two fumigants are left in widespread use, which is phosphine and methyl bromide
(Banks, 1994, Annis, 1987, Armstrong et al., 2014), while sulphuryl fluoride is still used as
a structural fumigant (mainly against termites) and for stored products and museum pests
(Bond, 1984, Banks, 1994). Modern technology and research brought to light certain
problems with fumigants that were previously unknown and have shown that some of these
materials have serious effects on human health (Armstrong et al., 2014, Svedberg et al.,
2013).
1.3.1.1 Methyl Bromide
Methyl bromide is colourless, odourless, non-flammable, boils at 3.56°C (38.5°F) and has
a very low solubility in water. For ease in transportation and handling, methyl bromide is
compressed and stored in metal cylinders as a liquid. Methyl bromide is an effective
32
fumigant for treating a wide variety of plant pests and is the most frequently used fumigant
in quarantine and pre-shipment purposes (USDA, 2016, Armstrong et al., 2014).
As mentioned, methyl bromide is being phased out as it is listed as an ozone depleting
substance under the Montreal Protocol (UNEP, 2006), but is still being used for postharvest
treatment of non-perishables (13%), as well as perishables (8.6%) and for quarantine
purposes (<1%) to a certain extent (Belova et al., 2013, Qiao et al., 2015., UNEP, 1992).
The 2008 International Plant Protection Convention recommended the reduction and
replacement of methyl bromide for phytosanitary measures. This stressed the need for
continued research to find alternative quarantine treatments to methyl bromide fumigation
(UNEP, 2010).
The use of methyl bromide as a pre-shipment quarantine measure has also been restricted by
the Montreal Protocol (UNEP, 2010). The restrictions in New Zealand requiring complete
recapture or destruction by 2020 (NZMAF, 2007a, NZME, 2011), which creates additional
pressure to identify alternative treatments to methyl bromide. The use of methyl bromide in
populated areas may come under further scrutiny and restrictions based on research on
pregnant women which indicated that residential proximity to methyl bromide use during
the second trimester was associated with markers of restricted foetal growth (Gemmill et al.,
2013).
Exposure may occur when Methyl bromide is used as a fumigant or pesticide during
fumigation activities. Methyl bromide is highly toxic and studies in humans showed that the
lungs may be severely injured by the acute (short-term) inhalation of methyl bromide. Acute
and chronic (long-term) inhalation of methyl bromide can lead to neurological effects in
humans (EPA, 1999).
Symptoms of acute exposure in humans include fainting, headaches, dizziness, weakness,
apathy, confusion, speech impairment, numbness, visual effects and tremors; Paralysis and
convulsions are possible in severe cases. Kidney damage and liver damage has been
observed in humans who have inhaled high levels of methyl bromide (ATSDR, 1992).
1.3.1.2 Phosphine
33
Phosphine was patented 80 years ago for crop protection and continued to be used in many
forms for fumigation because of its value to agriculture. It is relative safe to handle, easy to
use, an effective alternative to ozone depleting methyl bromide and with virtually no
residues. In recent years, concerns have been raised regarding insect resistance, but research
has shown that this can be successfully managed with correct application techniques
(Armstrong et al., 2014).
In later years the use of phosphine has been extended for treatment of pests in animal fodder,
fresh flowers, vegetables and fruits which brought new challenges to agricultural research.
Insects infesting fresh produce require much higher doses of phosphine, respond in relatively
shorter times and are killed at far lower temperatures than insects in stored grain (Ryan et
al., 2014). According to Banks, (2002), phosphine’s weaknesses are flammability,
corrosiveness, poor action at low temperatures, slow and resistant, whereas its strength is
that it is a cheap fumigant.
Although phosphine has genotoxic effects to humans and increasing environmental and
workspace restrictions (Bond, 1984), phosphine is still widely available as a registered
fumigant around the world, but requires very long exposure periods (Bond, 1984, Xin et al.,
2008).
1.3.1.3 Sulphuryl fluoride
Sulfuryl fluoride is a compressed gas fumigant, which is used primarily against insects that
attack wood and is effective at very low dosages on dry wood termites (Stewart, 1956) and
commodity insect pests (Kenaga, 1957). Control of the adult stage is the only concern for
stored product and museum pests (Bond, 1984, Banks, 1994). Higher dosages are required
for control of the egg stage in insects (USDA, 2016). Sulphuryl fluoride is used as a structural
fumigant and can control a wide range of insects, but the efficacy of this fumigant is
dependent on the concentration reaching the target pest and the time of exposure (Nitschke
et al., 2001, Misumi et al., 2010).
Sulphuryl fluoride is toxic to insects under all temperature and exposure conditions. It is also
easily dispersed, non-flammable, non-explosive, non-reactive with a wide range of
34
materials, non-sorbtive and penetrates rapidly (Kenaga, 1957). Sulphuryl fluoride is stable
at very high temperatures (400°C) and is non-corrosive to equipment and electronics (Bell,
2006, Thoms et al., 2010b). Sulphuryl fluoride requires high concentrations and long
exposure periods to be effective (Xin et al., 2008). According to Banks, (2002), the
weaknesses of sulphuryl fluoride are registration and low effectiveness against egg stages of
some insects, while the strengths are good penetration and little sorbtion.
Environmental issues and the lack of efficacy against insect eggs cannot be overlooked
(Armstrong et al., 2014) and this lack of efficacy in controlling insect eggs using sulphuryl
fluoride as a fumigant is considered its major weakness as an alternative treatment (Nitschke
et al., 2001, TEAP, 2010). Unlike methyl bromide, sulphuryl fluoride does not impact the
atmospheric ozone layer (Thoms et al., 1994, USEPA, 1996b), but recent studies showed
that sulphuryl fluoride is a greenhouse gas (Hunt, 2009, Mühle et al., 2009, Papadimitriou
et al., 2008) and is 4800 times worse than CO as a climate warming gas (Hunt, 2009, Mühle
et al., 2009).
Tsai, (2010), reported that replacing methyl bromide with sulphuryl fluoride could create
significant impacts on human health. The United States Environmental Protection Agency
(USEPA, 2011), revised the amount of fluoride allowed in drinking water and limited the
amount of fluorine exposure for children, while Tiemann, (2013), suggested the banning of
sulphuryl fluoride on edible commodities.
1.3.1.4 Treatments of quarantine pests
Treatments for biosecurity pests can be chemical or non-chemical. There are various
approved chemical treatments which include fumigants, drenches and spray (aerosols or
fogging) (Armstrong et al., 2014). Chemicals applied as aerosols, smokes, mists, and fogs
are suspensions of particulate matter in air and are not fumigants (USDA, 2016). Pesticide
drenches were used against insect pests in quarantine programs (USDA, 2010) and nursery
industries (Dennis et al., 2004), but are hazardous to labour and time consuming (USDA,
2010).
35
The fumigants currently used includes methyl bromide, phosphine and sulfuryl fluoride
(Armstrong et al., 2014) and according to Bond, (1984) and Reichmuth, (1998), finding a
new fumigant will be difficult because of the increased scrutiny of the environmental and
health issues associated with fumigants as well as the amount of time required to formulate
new fumigants. Furthermore, the investment and costs involved with registering new
fumigants is significant (Adam et al., 2010) and that we may have to rely less on the true
fumigants and more on alternative methods of control, such as non-chemical treatment with
irradiation, controlled atmospheres, heat, and cold (Armstrong et al., 2014).
Non-chemical treatments include cold treatment, microwave, hot water immersion, vapor
heat treatment, steam sterilization and irradiation (USDA, 2020, Thornhill et al., 1995,
Lindgren, 1969, Blickenstaff, 1973, Dcstan, 1963, Hooper, 1970, USEPA, 2014a, Wang et
al., 2001a, Wang et al., 2001, Mangan et al., 1997, Tang et al., 2007, Armstrong et al., 2007),
but will not be discussed further as these treatments were not used in any way during this
study.
1.3.2 Fumigation
Fumigation is the treatment that will be discussed and there are several definitions for
fumigation which include, “Fumigation is the process of application, exposure and
dissipation of a toxic chemical in its gaseous state with the purpose of control of target pests
in the product and its enclosure” (GAFTA, 2012). “Fumigation is a treatment with a
chemical agent that reaches the commodity wholly or primarily in a gaseous state” (FAO,
1990, FAO, 1995, Price, 1985). “Fumigation is the act of releasing and dispersing a toxic
chemical and reaches the target organism in a gaseous state” (USDA, 2016). “Fumigation
is a process that creates a lethal concentration of toxic gas for a period that will kill the
target pest” (Stark, 1994).
As stated above according to Armstrong et al., (2014), phosphine and methyl bromide
remain the two most universally used fumigants followed by sulphuryl fluoride for structural
fumigation and that the testing of new fumigants can be problematic and costly. Banks,
(1994), stated that fumigation is becoming an “endangered technology”, while Annis,
(1987), calculated that the “development of a new fumigant would require approximately
36
106 individual treatment tests involving 109 test insects, representing the life stages of a
single target species, to identify the time, temperature and concentration parameters for an
efficacious treatment”.
Fumigation with toxic gases has been widely used to control invertebrates and other target
pests and fumigants can often provide effective economical control where other forms of
pest control are not feasible (Will, 2009, Bond, 1984, Armstrong et al., 2014). Fumigation
techniques are very adaptable to different commodities such as grain, perishables and
durables and in different situations, such as ship holds, warehouses, food processing plants,
under tarpaulins, or in specifically designed fumigation chambers (Bond, 1984) and in this
study, shipping containers.
Modern technology and new research on fumigants have brought certain problems to light
with fumigants, previously unknown and numerous investigations made on the adverse
effects of fumigants have shown that some of these fumigants can produce serious effects
on human health (Armstrong, et al., 2014, Banks, 1994). Some fumigants with excessive
hazard potential have been restricted or prohibited and are no longer used for pest control in
some countries (Bond, 1984, UNEP, 2010, Corey, 2013).
Banks, (2002), argues that the selection of the best alternative for methyl bromide will have
to be made on a case-by-case basis because they will be situation specific and the
development of a single, direct replacement for methyl bromide is most unlikely. Except for
ethane dinitrile (Brash et al., 2013), ethyl formate (Jamieson et al., 2009b) and sulphuryl
fluoride (Zhang, 2006) there have been no major successes in finding a new alternative
fumigant for methyl bromide.
1.3.2.1 Fumigation of shipping containers
A freight container (hereafter referred to as a shipping container) is “an article of transport
equipment intended to facilitate the carriage of goods by one or more modes of transport,
without intermediate loading” (ISO, 1992). During the 1970’s ISO standards were
37
(Rajendran, 2004).
Globalization of trade has increased the volume of goods transported by shipping containers
and more than 500 million shipping containers units are shipped annually between countries
and continents (Svedberg et al., 2013). In 2010, 540 million units (number of containers)
were handled by ports (UNCTAD, 2012). These 20ft or 6.1 m shipping containers referred
to as TEU’s (twenty-foot equivalent unit), are the most commonly used containers for the
transportation of goods (Rajendran, 2004). Shipping containers are high-risk carriers of
invasive non-indigenous pests, which may occur in the cargo, on packaging materials or
anywhere else in the container itself (Rajendran, 2004). Biosecurity invaders have also been
found on the exterior of containers (Crowe, 2001).
It is a requirement to fumigate containers before shipment and these containers are mostly
fumigated with phosphine and methyl bromide and sometimes with sulfuryl fluoride (FAO,
2004, P&O Nedlloyd, 2004). According to Rajendran, (2004), containerized transport in
shipping containers eliminates the risk of cross-infestation of commodities during transport.
It also facilitates in-transit fumigation and ensures an insect-free environment if disinfested
properly before shipment.
There are limited studies on fumigation of containers and containerized agricultural products
or any other commodities. De Lima et al., (1994), conducted fumigation experiments on
containers with agricultural products using phosphine to treat oaten hay in Australia.
Carbonyl sulphide was used to fumigate oaten hay in Australia (Weller et al., 2002) and
sulfuryl fluoride was used to fumigate an unspecified commodity in the USA (Schneider et
al., 2001). Carbon dioxide (CO2) atmosphere for fumigation of wheat was investigated in
Australia by Banks et al., (1979) and malt in Australia (Banks et al., 1981). Taylor, (2000),
investigated phosphine in CO2 to fumigate coffee and wood products in Indonesia as well as
methyl bromide to fumigate coffee and wood products in Indonesia. Ethyl formate was
investigated to fumigate dried fruits in Australia (Annis, 2002).
According to Rajendran, (2004), gas-tightness of shipping containers is an essential
parameter for a successful fumigation and is checked by pressure tests. Shipping containers
that are used for controlled atmosphere treatments and fumigation must be gastight for the
process to be effective and should not loose gas through apertures, such as ventilators, open
eaves and imperfections in the fabric. Changes in temperature, atmospheric pressure and
wind forces also contribute to gas loss from the storage structure (Banks et al., 1979).
According to Banks et al., (1975), leakage appears to be the main cause of fumigation
failures. In containers under fumigation during transit, several environmental forces are
known to cause gas loss from the container (Banks et al., 1986, Banks, 1988), which include
wind and transport velocity, rate of ascent and descent, headspace changes in temperature
and variation in barometric pressure.
De Lima et al., (1994), also stressed the significance of gas-tightness tests on empty or loaded
shipping containers to decide whether fumigation of the containers should occur with or
without tarpaulins. Gas tightness is also very important to maintain Ct product for long
periods and especially for in-transit fumigation (Ren 2018, pers. comm.). The pressure decay
test is the more convenient test for testing gas-tightness of the containers where the time for
an imposed pressure difference to decay from one value (e.g. 200 Pa) to 50% of it (100 Pa)
is taken into consideration. An automatic pressure-decay timer for rapid measurement of
gas-tightness known as contesters has been developed (Sharp et al., 1982).
Ball et al., (1997), carried out tests on 6,000 40ft containers loaded with hay over a period
of three years, with contester pressure decay timer for a standard of pressure decay time of
≥ 10 sec from 200 to 100 Pa. The tests revealed that plywood-floored containers were more
gas-tight than plank-floored containers and was recommended that containers failing in the
pressure decay must be fumigated under a gas-proof sheet (Rajendran, 2004).
1.3.2.2 In-transit fumigation of shipping containers
There have been no studies found in the literature that have investigated the use of ethyl
formate and nitrogen for in-transit fumigation of shipping containers. In transit fumigation
are mostly used in ships and the fumigants that are the most widely used in vessels are
phosphine-evolving gases such as aluminium phosphide, magnesium phosphide, gas toxin
or mangotoxin. These fumigants are solid pellets and are usually placed on the surface of the
39
stow or inserted just beneath it. Methyl bromide is applied in gaseous form but is not allowed
for in-transit fumigations (IMO, 2003).
There have been some studies that investigated in-transit fumigation of containers carrying
agricultural products and commercial cargo using fumigants such as methyl bromide, CO2
(Banks et al., 1981, Banks et al., 1986) and phosphine (Donahaye et al., 2000, Leesch et al.,
1986). However, the studies conducted have been limited due to the toxicity of these two
pesticides. Ethyl formate has not been investigated as an in-transit fumigant for shipping
containers.
1.4 Ethyl formate
Ethyl formate is a colourless natural plant volatile that has a pleasant aromatic fruity odour
and is highly flammable (Chemical Book, 2010) and occurs naturally in a variety of products,
including essential oils of grasses, beer, rice, beef, and cheese (Desmarchelier et al., 1999).
Ethyl formate breaks down to formic acid and ethanol through the process of hydrolysis and
are further metabolised by many organisms to be incorporated into cellular components or
used as sources of energy (Haritos et al., 2003). Formic acid itself can be oxidised to carbon
dioxide and water. It can also be excreted unchanged or partly metabolised in the tissues, or
it can be incorporated into proteins, lipids and nucleic acids (Liesi-vuori et al., 1991), while
ethanol is endogenously produced as a metabolite in small quantities in the human body and
are not considered to be of toxicological significance (JEFCA, 1997).
Using volatiles, such as ethyl formate, has a big advantage for fumigation because residues
found on treated commodities are found only in trace amounts (Desmarchelier et al., 1999,
Muthu et al., 1984). Plant volatiles such as ethyl formate have been shown to have
insecticidal properties (Vincent et al., 1972, Aharoni et al., 1980).
Ethyl formate is used as a food additive worldwide (FDA, 1979) and is registered in Australia
as a fumigant for dried fruit (Banks et al., 1996). Ethyl formate is a low molecular weight,
volatile compound which is produced in some fruits and vegetables and is also a flavouring
40
agent and aroma component (Ryan et al., 2014). Ethyl formate and formic acid are present
in fruits, vegetables, beer, wine and spirits, tuna, meat, mussels, cheese and bread
(Desmarchelier, 1999) and is commercially used in the manufacturing of Rum, as a flavour
agent for lemonade and essences and also as a fungicide and an organic solvent (Merck
Index, 1989). Ethyl formate has low toxicity to mammals and the environment with a
threshold limit value (TLV) of 100ppm (Lee et al., 2007) and readily breaks down to ethanol
and formic acid with no residues (Haritos et al., 2003).
Ethyl formate’s effectiveness against dried fruit pests was first reported in 1925 (Ryan et al.,
2012) and the insecticidal properties of ethyl formate were demonstrated by Vincent et al.,
(1971,1972) and tested as a fumigant by Aharoni et al., (1980) and Rohitha et al., (1993).
An historical overview of the use of ethyl formate was published by Ryan et al., (2012).
Ethyl formate provide rapid mortality in insects, but high concentrations are required to
obtain complete control (Haritos et al., 2003).
Temperature appears to be a limiting factor in ethyl formate fumigations and according to
the literature (Hilton et al., 1996, Aharoni et al., 1980 Stewart et al., 1983, Aharoni et al.,
1981, Vincent et al., 1972, Griffin et al., 2013, Harein, 1962, Stewart et al., 1984, Rohitha et
al., 1993), ethyl formate should be applied at temperatures between 18-27°C. According to
Ren 2020, pers. comm.) ethyl formate in a vaporized state can be applied at temperatures as
low as 10°C. The Methyl Bromide Technical Options Committee (MBTOC) assessment
stated that ethyl formate is highly sorbed by commodities and even more in high humidity
and it is difficult to attain adequate distribution which means longer exposure times may be
needed to ensure adequate penetration of bulk commodities (MBTOC, 2002, Armstrong et
al., 2014).
Ethyl formate is “generally regarded as safe” (GRAS) (FDA, 2004), as a food additive and
with its low molecular weight it can potentially degrade to biogenic levels before treated
products reaches the market (Simpson, 2004). This presents a significant advantage over
traditional pesticides, particularly so when considering numerous regulatory and financial
challenges resulting from maximum residue levels required for exports. In addition to these
safety perspective than methyl bromide and other comparable fumigants (Ren et al., 2008).
Ethyl formate has its challenges, which includes its flammability and registration efforts
(Armstrong et al., 2014). The flammability aspect has been eliminated through the
VaporMate™ delivery system, which reduces the concentration of the product to below its
flammability level in air while still effective to kill pests (Wolmarans, 2017). Ethyl formate
is mixed with liquid carbon dioxide and sold under the trade name, VaporMate™ by BOC
in both Australia and New Zealand, where it is approved for use on fresh produce and stored
products (Jamieson et al., 2009).
Ethyl formate is flammable at a at 2.7-16.5 volume concentration of a solution (% v/v) in air
(TCI America, 2006) and explosive when mixed at high concentrations (Ryan et al., 2003).
Researchers have demonstrated that mixing ethyl formate with carbon dioxide can enable it
safe to use as a fumigant (Jamieson et al., 2009a, Ducom, 2006). According to Haritos et al.,
(2003), the flammability risk of ethyl formate is virtually eliminated by the presence of high
concentrations of carbon dioxide or high concentrations (>99%) of nitrogen (Ren 2018, pers.
comm.).
Ducom, (2006), stated that “ethyl formate, not alone, but as a mixture with some other
compounds or in a vacuum, may be the most promising fumigant for grains and all other
stored and fresh products” when compared with current and potential alternatives to methyl
bromide (Armstrong et al., 2014).
1.4.1 Ethyl formate as a fumigant
Ethyl formate has been widely used as a fumigant for pests associated with dried fruit
(Vincent et al., 1971, 1972, Desmarchelier et al., 1999) and in stored wheat (Neifert et al.,
1925, Cotton et al., 1928, Roark et al., 1929, Simmons et al., 1954). Ethyl formate is a
fumigant with attributes such as rapid action, strong sorbtion and rapid decay to ethanol and
formic acid (Muthu et al., 1984, Ren et al., 2001, Haritos et al., 2003) and is a relatively safe
grain fumigant as well as a grain protectant (Desmarchelier et al., 1998, Annis, 2000, Annis,
42
2002, Mahon et al., 2003, Ren et al., 2003, Ren et al., 2006). Ethyl formate could be a useful
rapid fumigant for grains and similar durable commodities (Simmons et al., 1945) and was
evaluated for grain protection in the 1980’s (Muthu et al., 1983).
Shepard et al., (1937), found ethyl formate more toxic than carbon disulphide to Tribolium
confusum and Sitophilus granarius and Vincent et al., (1972), found that ethyl formate
compared favourably to phosphine against insects infesting dried fruits, while Muthu et al.,
(1984), regarded ethyl formate as a safe general fumigant for stored food.
Ethyl formate was also reported as an effective fumigant for control of insects in cereals and
pulses (Pruthi et al., 1945, CFTRI, 1979), clothing (Busvine et al., 1966, David, 1943) and
fresh fruit, vegetables and flowers (Aharoni et al., 1980, Stewart et al., 1983, Stewart et al.,
1984, Wang, 1982). Raghunathan et al., (1974) and Deo et al., (1986), found that ethyl
formate has fungicidal properties in cereals.
1.4.2. Toxicity of ethyl formate to insect pests
The toxicity of many fumigants is still under investigation. The toxicity of ethyl formate
toward insects may be due to formate inhibiting energy production in cytochrome-c-oxidase
system in mitochondria of insects. When ethyl formate enters the body of insects, ethyl
formate is broken down into ethanol and formate (Haritos et al., 2003). The toxicity of a
fumigant depends on the respiration rate of the target organism (Haritos, 2005). Generally,
the lower the temperature, the lower the respiration rate of the organism which tends to make
the pest less susceptible. Fumigation at lower temperatures requires a higher dosage rate for
a longer exposure period than fumigation at higher temperatures (USDA, 2016).
The toxicity of ethyl formate to stored-product insects has been investigated by Muthu et al.,
(1984) and Banks et al., (1996). The egg stages of both Sitophilus oryzae and Tribolium
castaneum (Herbst) were found to be the most susceptible and pupae the most tolerant stages
for ethyl formate fumigation (Muthu et al., 1984). Damcevski et al., (2002), found that all S.
oryzae larvae were killed within 48 hours at application rates of 109, 130 and 155 mg/L of
ethyl formate in 2.7 L glass desiccators containing 500, 1000 and 1500 g of wheat,
43
respectively. Dojchinov et al., (2003), investigated the toxicity of ethanol and formic acid,
the two derivative chemicals from ethyl formate, against two stored-product insects and
found that alkyl formates and formic acid were similarly toxic and twice as potent as ethyl
propionate, methyl acetate or ethanol. They found that alkyl formates were metabolised in
vitro to formic acid when incubated with insect homogenates to produce very high
concentrations of formic acid in the bodies of the target insects. It is therefore the insect’s
metabolism that transform ethyl formate into formic acid and these high levels of formic
acid in the insect causes death (Dojchinov et al., 2003).
An ethyl formate application rate of 90 g/m3 for a 24-hour exposure in an 868 L capacity
fumigation chamber (without grain) is effective for control of all the life stages of S. oryzae,
T. castaneum and Rhyzopertha dominica (Damcevski et al., 2002). Results from bin trials
wheat and peas (50-55 t) showed that an ethyl formate application rate of 90 g/m3 for a 48-
hour exposure is effective for all the life stages of T. castaneum and R. dominica but is
marginal for immature S. oryzae (Desmarchelier et al., 1998, Mahon et al., 2003).
When carbon doixide is used with ethyl formate, the toxicity to adults of S. oryzae, T.
castaneum and R. dominica can be greatly increased (Damcevski et al., 2003). Ren et al.,
(2006), reported that carbon dioxide resulting from insect respiration at different exposure
concentrations due to sorption of ethyl formate on the culture medium, were not adequately
considered. Damcevski et al., (2006), reported that the larger the grain quantity, the higher
the required ethyl formate dose to achieve 99% mortality. They further reported that the
toxicity of ethyl formate was highly dependent on relative humidity with toxicity higher at
higher humidities.
Toxicity of ethyl formate to insects has also been reviewed by Adu et al., (1985), Muthu et
al., (1984) and Hilton et al., (1997). Mortality (LD95) was achieved at 10.00 mg/L for eggs,
21.13 mg/L for larvae, 47.32 mg/L for pupae and 9.42 mg/L for adult of Callobruchus
chinensis (L) (Adu et al., 1985). This result showed pupae was most tolerant following by
larvae, egg and adults. Muthu et al., (1984), reported that eggs of both S. oryzae and T.
castaneum were found to be most susceptible and pupae were most tolerant stages for ethyl
formate fumigation.
44
Research has shown ethyl formate had very rapid action against S. oryzae adult with 50%
fill rate of wheat and exposure time of 12 minutes and two hours achieved 100% mortality
for concentration of 340 and 210 mg/L, respectively. Also, exposure time of three hours
achieved 94% mortality for concentration of 130 mg/L. All S. oryzae larvae were killed when
exposure to application of 109, 130 and 155 mg/L of ethyl formate on 500, 1000 and 1500
g wheat for a 48-hour exposure period (Damcevski et al., 2001). Ethyl formate at 25 mg/L
showed significant progeny reduction of psocids at 15, 25 and 30°C with 12, 24 and 48-hour
exposure period (Riudavets et al., 2001).
Ethyl formate was investigated by Simpson et al., (2007) as an alternative option for methyl
bromide as a treatment option for quarantine security and reported that susceptibility to many
types of control measures varies greatly with the pest’s age. A range of life stages for each
target pest was tested with ethyl formate. Arthropod tests indicated that LC99 concentrations
of ethyl formate would be in a range well tolerated by table grapes and that longer exposure
times might be necessary to achieve complete mortality for omnivorous leafroller and certain
other pest life stages.
Fumigation trials with Planococcus citri adults and eggs showed that ethyl formate was at
least as effective as MB at the EF and MB concentrations recommended under the current
phytosanitary disinfestation guidelines (Park et al. 2020). Ethyl formate fumigation were
also efficacious in controlling mealybugs and scale insects on the surface of a variety of
fruits and vegetables (Agarwal et al., 2015, Simpson et al., 2007, Misumi et al., 2013,
Jamieson et al., 2014, Pupin et al., 2013, Lee et al., 2016, Yang et al., 2016).
1.4.3 Determination of concentration × time (Ct)
1.4.3.1. Well-sealed enclosures that have passed a pressure test
The efficacy of a fumigant can be affected by the concentration of the fumigant and
fumigation time (Bell, 2000) and therefore the effect of the fumigant on insects and the
commodity is expressed as the concentration x time (Ct product). The Ct product is obtained
by multiplying the concentration by exposure time and expressed in g h/L. The cumulative
the average concentration of sequential observation and multiplying by the time interval
between them. Fumigants are monitored in terms of concentration values at timed intervals
over exposure time by means of gas chromatograph (GC) analysis (Bong et al., 2020). To
produce the best approximation of true total Ct products is to obtain numerous concentration
observations during an exposure (Asimah et al., 2015).
The Ct value of fumigants is calculated by using the equation
Ct = ∑(Ci+Ci+1) (ti+1−ti)/2
Where: C is fumigant concentration (g/m3),
t is time of exposure (h),
i is the order of measurement,
Ct is concentration × time product (g h/L).
Ren et al., (2006), reported that the Ct products for S. oryzae were at least 2200 g h/L in 75-
145 t farm bins but the true Ct product experienced by the target insects is not well defined,
as continuous measurements of prevailing concentrations were not taken and that sorption
effects lead to rapid loss of the fumigant. Calculating the Ct product based on applied
concentration and exposure period thus leads to an overestimation of the true Ct product
needed for control. It also does not consider that the required Ct products are probably
strongly time dependent, with lower values at short exposures having the same effect as
higher values at longer exposures (Banks et al., 1996).
The concentration × time (Ct) products of ethyl formate for adult Sitophilus oryzae,
Tribolium castaneum and Rhyzopertha dominica at 25°C and 70% relative humidity for the
six-hour exposure was, respectively: (1) LD50 107.8, 108.8 and 72.8 mg h/L and (2) LD99.5
207.4, 167.1 and 122.2 mg h/L. Endpoint mortality was reached within 24 hours of initial
exposure (Xin et al., 2008).
46
Park et al., (2020), compared the efficacy of ethyl formate and methyl bromide on P. citri
adults and eggs and both were effective at controlling P. citri adults with the L(Ct)99.9 value
lower than the currently recommended dose of ethyl formate and methyl bromide for
phytosanitary disinfestation of mealybugs in banana in South Korea. Complete control of P.
citri eggs was not achieved. When ethyl formate was applied at 35 g/m3 for 4 hours, mean
Ct product of 101.7 ± 11.6 g h/L with mean % egg hatch of 2.49 ± 3.44% (% egg mortality
of 97.5 ± 3.44%, ranging from 88.2 to 100%) was achieved.
1.4.3.2 Under Gas proof sheeting where gas losses are high
When gas loss rates are very high, it is better to calculate the Ct product by using the
geometric mean. This is done by multiplying two observed gas concentrations together
(g/m3) taken one after the other. Then multiplying the square root of their number by the
time interval in hours between the readings.
The Ct value of fumigants is then calculated by using the equation
Ct n, n+1 = (Tn+1 + Tn+1) x √Cn x Cn+1 gh/m3
Where:
Ct n, n+1 is the calculated Ct product between Tn and Tn+1 (gh/m3)
Tn is the first time the reading was taken in hours
Tn+1 is the second time the reading was taken in hours
Cn is the concentration reading at Tn in g/m3
Cn+1 is the concentration reading at Tn+1 in g/m3
The Ct product obtained from a series of readings can then be added to calculate the
cumulative Ct product for the entire fumigation period This value indicates whether the
fumigation is a success or a failure (P.E.S.T, 2017)
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Ethyl formate reacts with water under neutral conditions (non-acid or non-alkaline) to form
formic acid and ethanol which is a hydrolysis reaction (Mata-Segreda, 1999, Morrison et al.,
1959, Haritos et al., 2003). This reaction proceeds to an unstated equilibrium ratio in excess
liquid water at 30°C in approximately 4-5 hours, at 50°C in one hour and this reaction is
temperature dependent (Mata-Segreda, 1999). Ethyl formate is water soluble (105 g/L at
20°C) and undergoes slow hydrolysis to ethanol and formic acid (Budavari, 1996) and this
hydrolysis reaction is reported to proceed much faster when catalysed by acidic or alkaline
conditions (Mai, 2006). Formic acid and ethanol are metabolised by many organisms to
be incorporated into cellular components or used as sources of energy (Haritos et al.,
2003).
Other researchers have claimed that ethyl formate is very stable in distilled water (Ghittori
et al., 1984). The reaction equilibrium constants (Kc) of liquid based hydrolysis of ethyl
formate was measured at 25°C for dilute concentrations of ethyl formate in water of Kc =
0.03 and 0.009 mol ethyl formate/mol H2O with 0.025 mol/L of HCl catalyst to speed up the
reaction rates to be 0.38 and 0.40 respectively and these values were confirmed by measuring
the reaction equilibrium constants of the reverse esterification reaction of ethanol with
formic acid with values of Kc=1/2.6=0.38. This reaction equilibrium indicates that the
reaction of ethyl formate in water will proceed so that less than half the ethyl formate will
react to form ethanol and formic acid, (roughly 30 mol % for equivalent molar concentrations
of reactants, ethyl formate and H2O) (Mai, 2006).
This reaction is “thought to occur for ethyl formate adsorbed by grain although more
rapidly” (Haritos, 2005), however, specific evidence demonstrating that grain-based
hydrolysis occurs, was not provided. Ethyl formate added to samples of wheat to create a
grain concentration of 100 mg/kg and left for 24 to 72 hours in sealed flasks, was completely
recovered using 24 hours liquid methanol extraction to give 98-102 mg/kg concentration
48
estimates (Desmarchelier et al., 1999), indicating that the reaction of ethyl formate with grain
constituents was little for periods of 24 to 72 hours.
Haritos et al., (1999), reported that “the levels of ethyl formate directly ‘dripped’ onto wheat
when entering a sealed 50 ton silo at 90g ethyl formate per ton of wheat, where not
distinguishable from wheat at levels above control wheat samples of 1-3 mg/kg when out-
turned, after a 4 week ‘withholding’ period in the unaerated silo”, whereas Reuss et al.,
(2003), reported the formation of ethanol during adsorption of ethyl formate by rice
fractions, but conclude that the formation of ethanol in rice products could not be
distinguished from that occurring in pure air.
Natural levels of ethyl formate have been measured in wheat (Desmarchelier et al., 1998) at
levels of <=1 mg/kg, wheat and barley at 0.02 to 1.0 mg/kg respectively (Desmarchelier et
al., 1999) and grains (wheat, barley, oats and canola) at levels of 1-3 mg/kg (Ren et al.,
2001). Natural levels of the ethyl formate hydrolysis reaction product formic acid were
measured at relatively high levels of 197 to 243±22 mg/kg for wheat and 237±48 mg/kg for
barley (Desmarchelier et al., 1999). The hydrolysis reaction product ethanol occurs naturally
at much lower levels of 0.006±0.009 mg/kg for wheat and <0.01 mg/kg for barley
(Desmarchelier et al., 1999).
1.5 Nitrogen as a fumigant and carrier gas
Apart from poisonous gases that are used for pest control, natural components of the
atmosphere, e.g. oxygen, nitrogen and carbon dioxide can be manipulated to preserve food
and is referred to as "controlled " or "modified " atmosphere storage which are techniques
that are used in the storage of perishable commodities such as fruit, vegetables, cut-flowers,
etc., to slow down the ripening of fruit and vegetables and also reduce spoilage from micro-
organisms (Aharoni et al., 1980).
Nitrogen is an inert gas that comprises 78% of the atmosphere and has been researched as
an alternative fumigant for stored grains. Nitrogen is the atmosphere’s most dominant
component, with oxygen (20.9%) with argon and carbon dioxide completing the mix (Ren
et al., 2012).
Apart from being safe, environmentally friendly and cheap, nitrogen-based controlled
atmosphere technology has several other potential advantages over other fumigants (Ren et
al., 2012). Nitrogen is not toxic, which greatly reduce occupational health and safety and
environmental risks. Nitrogen is also residue-free, has no known resistance problems and
does not react with construction materials (Ren et al., 2012). In this study nitrogen was used
to eliminate flammability (by reduces the concentration of the product to below its
flammability level in air), act as a carrier gas for vaporized ethyl formate and increase the
effectiveness of the ethyl formate/nitrogen mix (synergistic effect) (Ren 2017, pers. comm.).
Controlled atmosphere is also an effective substitute for the fumigation of some commodities
because the gases involved do not leave harmful residues and may increase the effectiveness
of the treatment as carbon dioxide increases the toxicity of a number of fumigants to insects
(Kashi et al., 1975, Bernard et al., 1993, Bond, 1984, Leesch, 1995, Haritos et al., 2006).
Ren et al., (2012), has re-evaluated nitrogen based atmosphere as cheap and reliable and that
highly efficient nitrogen generators such as Pressure Swing Absorption (PSA) and
Membrane Nitrogen Separator (MNS) are available for the generation of nitrogen.
All fumigants that are identified as a potential alternative for methyl bromide must be
subjected to a cost benefit analysis. Cheap fumigants can become costly if an insect life stage
shows tolerance to the fumigant, or more fumigant being required to achieve an efficacious
fumigation for whatever reason. Very little economic data are available on these fumigants
and cost benefit analyses will require substantial efforts to support the commercial
application of any methyl bromide alternative (Armstrong et al., 2014, Adam et al., 2010).
1.6 Fumigants and synergism
Synergists of fumigants have been used for a long time and have contributed significantly to
the efficacy of insecticides (Bernard et al., 1993, Kashi et al., 1975). Synergism occur when
two compounds (either one toxic compound and a non-toxic synergist or two toxic
compounds) are combined and that toxicity produces significantly greater mortality to the
target pest than can be obtained using each of the compounds or toxicants individually (Cakir
et al., 2008, Casida et al., 1995).
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Carbon dioxide is commonly used to obtain synergistic effects (Bond, 1984) and it enhances
penetration and distribution of some fumigants, e.g., phosphine, through the substrate being
fumigated (Leesch, 1995). Carbon dioxide by itself is toxic to insects given adequate
concentration and exposure time (Bond, 1984). It can therefore be beneficial to fumigations
by increasing penetration into the commodity, thereby increasing efficacy (Leesch, 1995).
According to Bond, (1984), carbon dioxide may stimulate the respiratory movements and
opening of spiracles in insects. As early as 1930, Cotton, (1930, 1932), reported that the
addition of carbon dioxide increased the efficacy of fumigants i