Ann. occup. Hyg., Vol. 47, No. 3, pp. 227–233, 2003
© 2003 British Occupational Hygiene Society
Published by Oxford University Press
DOI: 10.1093/annhyg/meg034
An In-field Demonstration of the True Relationship
between Skin Infections and their Sources in
Occupational Diving Systems in the North Sea
C. AHLÉN1*, L. H. MANDAL1 and O. J. IVERSEN2
1Division
of Microbial Exposure and Indoor Air, SINTEF Unimed, N-7034 Trondheim; 2Department of
Laboratory Medicine, Norwegian University of Science and Technology, N-7006 Trondheim, Norway
Received 15 July 2002; in final form 10 January 2003
Introduction: Skin infections in saturation diving are caused by microbes that flourish in
saturation environments. Improvements in the prevention of infections must therefore be based
on environmental control and elimination. Furthermore, only a few genotypes seem to be
responsible for the majority of infections in the Norwegian sector of the North Sea, and these
have all been demonstrated in saturation systems for many years. Although reservoirs of infectious genotypes have been identified, their true sources have not been identified.
Objectives: The purpose of this field study was to log the contamination by Pseudomonas
aeruginosa of the saturation system throughout a diving operation.
Materials and methods: Daily water samples from the vessels drinking water system and
from the heated seawater systems to divers suits were taken throughout the diving period of
1 month in the summer of 2001. All P.aeruginosa isolates were genotyped by pulsed field gel
electrophoresis.
Results: A total of 17 P.aeruginosa genotypes were identified in the course of this field study.
None of the most common infectious genotypes previously observed in the Norwegian sector
were among these strains. Two genotypes were involved in skin infections during the period of
operation: TP2 and TP12. TP2 was shown to be an inhabitant of the diving systems throughout
the investigation period, while TP12 was introduced from seawater in the course of the operation and rapidly spread and established itself throughout the diving system.
Conclusions: The study has demonstrated seawater as a true source of an infectious P.aeruginosa genotype in occupational diving systems.
Keywords: Pseudomonas aeruginosa; saturation diving; skin infections; source identification
normally is between 50 and 200 m (0.6–2.1 MPa).
The ambient temperature is typically between 28 and
30°C and relative humidity may reach 80–90%
during periods of intense diving activity. In order to
maintain thermal balance while working in the sea,
the divers wear protective suits, through which
heated (∼36°C) seawater is continuously flushed onto
the skin. All in all, saturation diving involves several
unusual environmental factors, any or all of which
may be of significance for occupational health
(Alcock, 1977; Hope et al., 1994; Ahlén et al.,
1998a,b).
Superficial skin infections are a dominant health
problem during occupational diving in the Norwegian
offshore sector (Norwegian Petroleum Directorate,
1999). The microbial flora of the living and working
environment is very rich, in terms of both numbers
INTRODUCTION
Occupational saturation diving is extensively employed
in the installation, inspection and maintenance of
offshore sub-sea oil production systems in the North
Sea. The occupational living and working environment is unique (Freitag and Woods, 1983). During
saturation, teams of divers live for weeks at a time in
confined steel chambers, pressurized with helium to
the actual working depth of the diving operation. The
partial pressure of oxygen is kept between 40 and
60 kPa. The ambient absolute pressure is dependent
on the working depth, which in the North Sea
*Author to whom correspondence should be addressed.
Tel: +47-73-592356; fax: +47-73-591005; e-mail:
[email protected]
227
228
C. Ahlén, L. H. Mandal and O. J. Iversen
Fig. 1. Identification of P.aeruginosa isolates from the ‘Upgrade study’ using PFGE and the endonuclease SpeI. The PFGE patterns
are consecutively coded (capital letters). The sources of the isolates are shown in Table 2.
and genera. Nevertheless, the dominant microbe in
divers’ skin infections is Pseudomonas aeruginosa, a
very common bacterium in waters all around the
world, and known as a very common challenge to
divers’ health since the beginning of saturation
diving (Thalman, 1974).
In the course of a long-term process of infection
monitoring and environmental control in Norwegian
occupational offshore saturation diving systems, a
great deal of specific knowledge about the infectionrelated P.aeruginosa strains has been gained. This
knowledge includes both phenotypic and genotypic
characteristics and patterns (Ahlén et al., 1998b). The
most important epidemiological findings from this
surveillance process have been gained from retrospective genetic studies of our collection of more
than 1000 field-related P.aeruginosa strains. A total
of 250 genotypes have been identified from infections, of which a few have been identified as frequent
infectious genotypes (Ahlén et al., 2000). These have
been shown to occur frequently both within the saturation environment and in infections over many
years. The relationships and significance of these
environmental isolates on the spectrum of infections
in occupational diving have recently been discussed
(Ahlén et al., 2001).
Potential reservoirs of these strains, involving both
water and gas systems on board diving vessels, have
been identified. In spite of the long period during
which monitoring has continued, it has so far proved
impossible to identify the source of the specific
frequent genotypes in saturation systems. This paper
describes seawater as a candidate true source of an
infection-related genotype of P.aeruginosa.
MATERIALS AND METHODS
Field study
The field study was undertaken during a 1 month
diving operation in July 2001 in the western part of
the Norwegian offshore sector of the North Sea. The
diving vessel was British, relatively new and had
never been to the Norwegian sector before. The
specific diving operation was pipe laying of a new
gas pipeline, and involved three different fields and
locations at which the vessel was stationary for
periods. The respective periods were 3–14 July, 15–
25 July and, finally, 26 July–3 August. Mobilization
for the operation was carried out in harbour in Haugesund, Norway.
Diving vessels usually have two separate water
production systems on board; one for drinking water
Pseudomonas aeruginosa in diving systems
229
Table 1. Pseudomonas aeruginosa genotypes and isolation sites during the field study
Genotype
Date of first occurrence
Dates of subsequent occurrence Sites of occurrence
TP 5
29 June
4, 5, 8 July
Galley, Ch3
TP 6
29 June
15, 21, 24 July
Galley, Ch3
TP 7/TP 14
29 June
TP 8
29 June
3, 21, 24 July
Ch3
TP9
7 July
8 July
Ch1
TP10
7 July
TP11
14 July
TP12
15 July
16, 21, 24, 25, 27 July
Sw intake, Ch1, Ch2, Ch4, Ch5, Equip.
room, Sat ctr, Suit rinse, infection
TP13
15 July
16 July
Sw intake
TP15
18 July
Sw intake
TP16
18 July
Sw systems
TP17
21 July
Sw systems
Sw systems
Sw systems
Sw intake
Ch, saturation chamber; Equip. room, equipment room; Sat ctr, saturation control room; Sw, seawater.
and one for hot water (seawater) for the divers. In
addition to the seawater production system, this
vessel had two separate drinking water production
systems on board; one employing evaporation and
the other reverse osmosis. Potable water was
bunkered from an onshore supply during mobilization on 29 June–2 July. Except for a certain amount
of evaporation, bunkered water was the main source
of water during the first 2 weeks of operation. Full
water production on board was first introduced on the
morning of 15 July, upon arrival at the second
stationary location.
Table 2. Pseudomonas aeruginosa genotype patterns and site
of isolation: ‘Upgrade study’
Date
Sampling site
PFGE pattern
(TP)
Lambda ladder
20–21 March 2001
Galley
1
20–21 March 2001
EL1 shower
2
20–21 March 2001
EL1 shower
2
20–21 March 2001
EL1 shower
2
20–21 March 2001
EL3 shower
3
20–21 March 2001
EL3 shower
3
20–21 March 2001
EL3 shower
3
Microbiological sampling and analyses
The field study focused entirely on P.aeruginosa.
The total microbial flora observed in the samples is
therefore not reported here.
The quality of microbiological sampling was assured
by the presence of licensed medical personnel
(nurses) on board throughout the operation. Microbiological analyses of samples were performed at the
same clinical microbiological laboratory onshore as
has been employed since the start of monitoring in
1985. Details of microbiological sampling procedures,
analyses and identification have been described elsewhere (Ahlén et al., 1998b), and are only described
briefly here.
20–21 March 2001
EL3 shower
3
20–21 March 2001
EL4 shower
3
20–21 March 2001
EL4 shower
3
Sampling period. The sampling period included
mobilization, operation and demobilization, and
lasted from 29 June, the date of arrival of the diving
vessel at the quay in Norway, to 3 August, the date of
return to the same quay.
room and equipment maintenance room. Internal
saturation systems: taps and showers in saturation
chambers 1–5.
Daily samples from the hot water system for divers
at work were taken from the seawater intake.
Sampling protocols were distributed to the vessel
prior to project start.
Daily water samples. Daily samples of fresh water
(potable water from both the general system on board
the vessel and from the water supply to the saturation
system) were taken from the following sites. External
saturation systems: taps in galley, saturation control
Lambda ladder
20–21 March 2001
EL5 tap
3
20–21 March 2001
EL5 tap
3
20–21 March 2001
EL5 tap
3
20–21 March 2001
EL5 tap
3
20–21 March 2001
EL3 shower, swab
3
20–21 March 2001
EL4 shower, swab
3
20–21 March 2001
EL5 hose toilet, swab
4
Lambda ladder
EL, entry lock.
Transport of samples to laboratory. Daily delivery
of samples to the laboratory was not possible due to
infrequent helicopter transport schedules. The daily
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C. Ahlén, L. H. Mandal and O. J. Iversen
Table 3. Pseudomonas aeruginosa genotypes present during
the first part of the field study
Table 4. Pseudomonas aeruginosa genotypes present in the last
part of the field study
Date
Date
Sampling site
PFGE pattern
(TP)
Lambda ladder
Sampling site
PFGE pattern
(TP)
Lambda ladder
29 June 2001
Galley dish hw, UK baseline
5
24 July 2001
Sat ctr tap mix
12
29 June 2001
Galley dish hw, UK baseline
6
24 July 2001
Equip. room, tap mix
12
29 June 2001
Sw, U1 AFT, UK baseline
7
24 July 2001
Suit rinse, tap mix
12
29 June 2001
EL4 tap cw
3
24 July 2001
EL1 shower mix
12
3 July 2001
EL3 shower mix
8
24 July 2001
EL2 shower mix
2
4 July 2001
EL3 shower mix
5
24 July 2001
EL3 shower mix
8
7 July 2001
EL1 shower mix
9
24 July 2001
EL3 shower mix
6
7 July 2001
EL1 tap cw
9
24 July 2001
EL4 shower mix
12
7 July 2001
EL3 tap hw
5
24 July 2001
EL5 shower mix
12
Lambda ladder
Lambda ladder
7 July 2001
Sw, FWD bell U3
10
24 July 2001
Sw intake
12
8 July 2001
EL1 shower mix
9
25 July 2001
Sat ctr tap mix
12
8 July 2001
EL3 shower mix
5
25 July 2001
Equip. room tap mix
12
14 July 2001
Sw intake
11
25 July 2001
Suit rinse, tap mix
12
15 July 2001
Sw intake
12
25 July 2001
EL2 shower mix
12
15 July 2001
EL2 shower mix
2
25 July 2001
EL4 shower mix
12
15 July 2001
EL3 shower mix
6
25 July 2001
EL5 shower mix
15 July 2001
EL4 shower mix
3
27 July 2001
D.E., infection
12
15 July 2001
Sw intake
13
27 July 2001
D.E., infection
12
Lambda ladder
cw, cold water; EL, entry lock; hw, hot water; Sw, seawater.
water samples were therefore stored at 4°C until they
were sent to the laboratory ashore. Samples were
collected for five working days at most.
Microbiological analyses. A 500 ml aliquot of each
sample of water was filtered (Millipore 0.45 µm) and
cultured on Pseudomonas agar (Cetrimidine). Specific
microbial findings were biochemically typed (API
systems) to species level. All P.aeruginosa isolates
were further analysed by genotyping.
Genotyping was done by means of pulsed-field gel
electrophoresis (PFGE). The method has been
described in detail in earlier reports and papers
(Ahlén et al., 1998b). Briefly, DNA was digested by
means of a rare-cutter endonuclease SpeI (Boehringer
Mannheim Biochemica) and electrophoresis was
performed in a commercial Gene Navigator (Pharmacia Biotech). The DNA fragment patterns were
visually compared with each other and with previously identified genotypes in our material from saturation diving. All new genotypes were labelled TP
with a consecutive number.
‘Upgrade study’: fitness for diving operations in the
Norwegian sector
The regulations for manned diving operations are
set on a national basis, which results in differences in
4
Lambda ladder
Equip. room, equipment room; EL, entry lock; Sat ctr,
saturation control room; Sw, seawater.
the regulations applied by individual countries. For
this specific project, harmonization of the UK and the
Norwegian procedures for hygiene and infection
control was undertaken as a separate project: the
‘Upgrade study’. This study was performed towards
the end of April 2001 while the vessel was in harbour
in the UK, and included a thorough survey of
contamination status on board the vessel, particularly
in the saturation systems.
A very high level of contamination of P.aeruginosa was demonstrated in the saturation systems,
with massive contamination of the freshwater supply
to the saturation chambers. Cleaning and disinfection
regimes were implemented by the vessel management in accordance with previously described guidelines (Ahlén et al., 1991).
The genotypes found in the saturation system on
board the diving vessel during the ‘Upgrade study’
were four different genotypes, none of which were
related to the frequent infectious genotypes known
from Norwegian saturation systems. The genotypes
isolated in the ‘Upgrade’ study were labelled TP1,
TP2, TP3 and TP4. The sampling sites and PFGE
patterns of these genotypes are shown in Table 2 and
Fig. 1.
Pseudomonas aeruginosa in diving systems
231
Fig. 2. Pseudomonas aeruginosa genotypes, identified by PFGE, present during the first part of the field study. The sources of the
isolates are shown in Table 3.
RESULTS
None of the most common P.aeruginosa infectious
genotypes seen in other studies in the Norwegian
sector were identified in this study.
Two of the genotypes identified from the ‘Upgrade
study’ (TP2 and TP3) were also found in the field
study. TP2 was identified in the showers in one of the
chambers on 15 July, and skin infections caused by
the same genotype were seen on 18 July. TP3 was
regularly observed throughout the field study, but has
not been involved in infection.
A total of 13 P.aeruginosa genotypes not previously observed in the Norwegian sector were isolated
in the course of the field study. These were consecutively labelled TP5–TP17 on the basis of their first
occurrence in the diving systems (with the exception
of TP14, which was present during the mobilization
period but was analysed at a later stage). In Table 1,
the first occurrence, later occurrences throughout the
field study and sites of isolation of each genotype are
shown. Table 3 and Fig. 2 list the genotypes identified during the first part of the field study.
Genotypes TP5, TP6, TP7 and T14 had already
been seen in the mobilization period (29–30 June).
TP5 and TP6 were identified from the drinking water
system outside the chamber systems and TP7 and
TP14 were identified from the hot seawater system.
Genotypes TP8 and TP9 were identified early in
the diving operations period, and both of these were
found in the drinking water supplied to the chamber
system.
In Table 4 and Fig. 3, the genotypes present during
the last part of the field study are shown. Genotypes
TP10–TP17 were all initially identified from the
seawater intake throughout the diving operations
period. Genotypes TP10 and TP11 were introduced
into the system during operations at the first location
and the remaining genotypes (TP12–TP17) were all
introduced at the second diving location.
Although several genotypes were introduced into
the systems in the course of the period of operation,
only one genotype, TP12, was widely disseminated
throughout the systems, spreading to both the
external vessel systems and the saturation systems
(Table 1). TP12 was demonstrated as a pure culture
from skin infections on 21 July (Fig. 3).
DISCUSSION
We have previously reported that skin infections in
occupational saturation divers in the Norwegian
offshore sector are related to certain frequent infectious genotypes of P.aeruginosa (Ahlén et al.,
1998a,b). These genotypes have been shown to be
present both in the saturation environment and as
232
C. Ahlén, L. H. Mandal and O. J. Iversen
Fig. 3. Pseudomonas aeruginosa genotypes, identified by PFGE, present during the last part of the field study. The dominance of
TP12 can be seen both in the external systems and in the saturation systems. The sources of the isolates are shown in Table 4.
skin infection strains over a period of many years
(Ahlén et al., 2000, 2001).
While several potential reservoirs of the frequent
infectious genotypes have been determined, it has
hitherto been impossible to identify the respective
sources of these genotypes. Knowledge of sources is
of great importance for improving control and elimination, and thereby prevention, of occupational
infections.
The field study presented in this paper was
performed on a diving vessel which had not previously been employed in the Norwegian sector. None
of the common frequent infectious genotypes previously observed were seen during this field study. The
fact that this operation involved the laying of a new
pipeline in fields not earlier exposed to oil exploration might be of relevance for this result.
Including the genotypes from the ‘Upgrade study’
in the UK sector, a total of 17 genotypes were identified from the diving system investigated in the field
study. The absence of the earlier well-known genotypes together with the new patterns from the UK
sector may suggest that P.aeruginosa genotypes are
to a certain extent field-specific.
A total of 13 P.aeruginosa genotypes were identified throughout the field study period. As mentioned
above, four of these had already been seen during the
mobilization period and are therefore not relevant to
the field study. These may have been introduced in
the UK sector, during the trip to Norway or in the
Norwegian harbour during mobilization.
Although diving operations started on 3 July, it was
not until 15 July that the contamination reached
significant levels. From that date, contamination
increased in terms both of number of genotypes and
number of bacteria. More than 50% of the genotypes
were introduced into the systems during the mid
diving period between 15 and 25 July. The change in
location and the introduction of full on-board water
production may have been factors of significance for
the level of contamination.
Most of the genotypes described in this study were
introduced into the vessel system through the
seawater systems, particularly via the seawater
intake. This study has thus demonstrated that
seawater is a potential source of contamination by
P.aeruginosa. Nevertheless, only one genotype
(TP12) was demonstrated as being widespread in
both the general vessel water systems and the satura-
Pseudomonas aeruginosa in diving systems
tion water systems on board. After its first occurrence
on 15 July, a massive spread could be observed in the
course of the following days (Fig. 3). This might indicate that certain P.aeruginosa genotypes have greater
potential to survive and spread within such systems
than do others. The fact that this genotype also
caused infection demonstrates that P.aeruginosa
obtained from seawater represents an infection threat
in diving systems. The demonstration that TP2 could
survive for several months, as could the infection
caused by that genotype, are examples of the contamination and contagion mechanisms seen in this
specific niche. Finally, the results also strengthen the
probability that certain genotypes are more infectious
than others.
In conclusion, this field study has for the first time
demonstrated seawater as a true source of a P.aeruginosa infectious genotype in a saturation diving
system. It is therefore reasonable to consider seawater
as a potential source of the frequent infectious genotypes seen in Norwegian diving systems in the course
of the past 18 years.
Acknowledgements—We thank all participating divers and the
health personnel on board the diving vessels for their skilled
cooperation and Marianne Aas, Grethe Lysholm Iversen, Evy
Mellemsæther and Therese Ahlén for skilled technical assistance. We wish to thank Statoil, Norsk Hydro, Saga Petroleum
and the Norwegian Petroleum Directorate for financial support.
233
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