NATIONAL SALMONELLA, SHIGELLA & LISTERIA REFERENCE
LABORATORY OF IRELAND
(HUMAN HEALTH)
ANNUAL REPORT FOR 2012
NATIONAL SALMONELLA, SHIGELLA & LISTERIA REFERENCE LABORATORY
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Introduction
The National Salmonella, Shigella & Listeria Reference Laboratory was established in 2000 with
support from the Department of Health and Children to provide reference services related to human
health. It is a public service laboratory which currently operates with 2 full time scientific staff
representing a reduction from 3 staff working in the service until 2011. The service operates from the
Department of Bacteriology at the Clinical Science Institute, NUIGalway. The NSSLRL website is at
http://www.nuigalway.ie/salmonella_lab/. The reference laboratory uses a number of phenotypic
methods (serotyping, antibiotic-resistance testing and phage typing) and molecular methods (pulsefield gel electrophoresis (PFGE) and multi-locus variable number tandem repeat analysis (MLVA) to
precisely characterise Salmonella isolates. This process can be considered as genetic fingerprinting of
Salmonella. The goal of this genetic fingerprinting is to assist relevant agencies in protecting public
health by identifying and interrupting chains of transmission of Salmonella infection. In addition the
NSSLRL provides typing services for Shigella species and Listeria monocytogenes.
One of the interesting features in this years report is that as with last year the proportion of human S.
Typhimurium isolates accounted for by the phage type DT104 is less than 5%. For many years now S.
Typhimurium DT104 has accounted for 10 to 20% of human S. Typhimurium isolates. The reason for
this apparent decline is not known but given that the pattern that emerged in 2011 has been sustained
in 2012, it is unlikely to be random variation.
One of the major challenges that NSSLRL will need to address in the near future is the transformation
of molecular typing methods. It appears increasingly likely that whole genome sequencing is rapidly
being accepted as a key method in molecular typing of pathogens. In 2013, NSSLRL is exploring ways
in which it can develop and apply these methods within a very resource constrained environment.
The Laboratory is committed to providing a high quality and timely service and has achieved
accreditation to the ISO15189 standard from the Irish National Accreditation Board (INAB).
The
continued success of the laboratory is entirely dependent on the support of the staff in the laboratories
that submit isolates for typing. My colleagues and I appreciate that the preparation, packing and
dispatch of isolates is a significant burden and would like to thank you for your support over the years.
I would also like to acknowledge the support of all those agencies with whom we work closely to ensure
that the service we provide works as information for action. In particular I would like to thank
2
Galway University Hospitals, NUI Galway, the Food Safety Authority of Ireland, the Health Protection
Surveillance Centre and colleagues in Public Health Departments and Environmental Health
Departments throughout the country and to acknowledge the work of colleagues in the National
Reference Laboratory Salmonella (Food, Feed and Animal Health)1.
If you have any comments or questions arising from the report please feel free to contact me at the
email address given below.
Martin Cormican
Director National Salmonella, Shigella & Listeria Reference Laboratory
martin.cormican@hse.ie
1.
The Annual Report of National Reference Laboratory Salmonella (Food, Feed and Animal
Health is available at
http://www.agriculture.gov.ie/media/migration/animalhealthwelfare/labservice/nrl/NRLSalmo
nellaAnnualReport2011.pdf
Salmonella
In 2012, 868 isolates were submitted to the National Salmonella, Shigella & Listeria Reference Laboratory.
When non-Salmonella, QC, contaminants and duplicate isolates were removed a total of 710 Salmonella
isolates were typed. This represents a 1% increase in the number of isolates received in 2011.
There were 319 human clinical isolates, including 282 faecal isolates, 25 from blood (including 6 S.Typhi, 2
S.Paratyphi A and 1 S.Paratyphi B), 4 other invasive isolates, and 8 urine isolates. S.Typhimurium (n = 71)
and its monophasic variant 4,5,12:i:- (n = 52) and S.Enteritidis (n = 56) predominated (Table 2). There was
marked seasonal variation with the highest number of isolates occurring in months July to October. This
coincides with the warmer months of the year and with the peak season for foreign travel (Fig.1) and may be
related in part to one or both of these factors.
In some cases more than one isolate was received from a patient.
For example we may have received an
invasive isolate (e.g. from a blood culture) and an isolate from faeces from the same patient. Where invasive
and faecal isolates come from the same patient, only the invasive isolate is recorded to avoid duplication. The
average turnaround time for human isolates was 6 days (range 2-20 days). The number of human Salmonella
isolates received is now just over half that observed when the laboratory was established in 2000 (Table 1)
and has been sustained at that level for some years. This reduction in number of isolates submitted is likely to
reflect a true reduction in the incidence of human salmonellosis and represents a significant public health
achievement by those agencies working in this area.
3
Table 1: Number of Salmonella isolates received in NSSLRL
Year
Human Non-human
2012
319
391
2011
320
381
2010
364
559
2009
364
368
2008
447
815
2007
457
653
2006
430
308
2005
357
494
2004
420
650
2003
486
634
2002
394
540
2001
508
574
2000
636
214
Table 2: Top 15 serotypes of Human Isolates (inc typhoidal)
Serotype
Typhimurium
Frequency
%
71
22.3
1
Monophasic Typhimurium 52
16.3
Enteritidis
56
17.6
Stanley
11
3.4
9
2.8
Typhi
Newport
8
2.5
Infantis
7
2.2
Braenderup
6
1.9
Dublin
6
1.9
Bredeney
5
1.6
Saintpaul
5
1.6
Java
5
1.6
Agona
4
1.3
Kentucky
4
1.3
Mikawasima
4
1.3
Others
66
20.7
Total
319
100
4
1
The antigenic formula 4,5,12:i:- is that of S. Typhimurium except that the phase 2 antigen is not expressed.
These isolates are generally referred to as monophasic S. Typhimurium and are so called in this report.
Figure 1
1.
Seasonal Variation in numbers of Salmonella isolates 1
The line “FT (Foreign Travel)” describes the number of cases of salmonellosis for which an association
with recent foreign travel was reported to NSSLRL.
Reporting of recent foreign travel is likely to be
incomplete.
It is important to note that there is always an interval gap between the time of onset of symptoms and date of
isolate receipt in the NSSLRL. This includes time taken for patient to access doctor, taking and transporting
the sample to the primary laboratory, isolation of Salmonella, and referral to NSSLRL. Fig 1 refers to date
of receipt in the NSSLRL.
Salmonellosis non-typhoidal
S.Typhimurium and its monophasic variant.
S.Typhimurium 4,5,12;i:2 and its monophasic variant (4,5,12;i:-) together accounted for 38.6% of all cases of
Salmonella. Phage type U323 was the most common (24%) phage type among human isolates in 2012. Phage
types DT193 accounted for 13.8%, phage type Untypable* 11.4%, DT104 11.4% and DT120 accounted for
8.9% of the total number of S.Typhimurium/4,5,12:i:- isolates. The proportion of isolates accounted for by
phage type DT104b was low (4.9%) compared to recent years (2011 (3.5%), 2010 (11%), 2009 (12%), 2008
(20%), 2007 (12%)).
* Does not react with typing phages.
5
S.Enteritidis
S.Enteritidis accounted for 17.6% of all cases of Salmonella. The predominant phage types were PT8
(26.8%), PT1 (17.9%), PT21 (14.3%), PT4 (10.7%) and PT14b (8.9%).
Salmonellosis Typhi and Paratyphi
Nine isolates of S.Typhi, 3 isolates of S.Paratyphi A and 1 S.Paratyphi B isolate were received. A history of
recent travel was recorded for 7 of the S.Typhi isolates; 6 to the Indian subcontinent and 1 to the Philippines.
All 3 of the S.Paratyphi A isolates were associated with reported travel to Indonesia. The S.Paratyphi B
isolate had a history of travel to Peru.
Antimicrobial resistance
Fifty-three percent of isolates (n = 168) were susceptible to all antimicrobial agents tested. Thirty-eight
percent of isolates (n = 120) were multi-drug resistant (three or more different classes of antibiotics). Of the
38% of isolates that were multi-drug resistant, 39% (n=47) had the profile of resistance to ampicillin,
streptomycin, sulphonamide and tetracycline (ASSuT) and were mainly monophasic S. Typhimurium,U323
or DT193.
Four extended spectrum beta-lactamase (ESBL) producing isolates were detected. An ESBL-producing
S.Concord isolate was received from a child adopted from Ethiopia. The association of ESBL producing S.
Concord with children adopted from Ethiopia has been noted in a number of countries and a report of an
international collaborative study on this phenomenon in which NSSLRL participated was published in 2009.
The phenomenon has continued to occur in Ireland in the years since that report. Given the multi drug
resistant nature of these isolates it may be appropriate to consider screening infants adopted from Ethiopia for
carriage and measures to ensure that parents of these children are aware of the potential for carriage of these
organisms. Other ESBL producing isolates included a patient with travel history to Indonesia (S.Anatum) and
two patients with unknown foreign travel histories (S.Newport and S.Typhimurium DT120). An AmpC
producing S. Heidelberg isolate was received from a patient with a history of travel to Bangladesh.
Four isolates of Salmonella resistant to ciprofloxacin were detected. Two of these were S.Kentucky, one with
travel history to Bangladesh and the other to Nepal. Resistance to ciprofloxacin (based on widely used
interpretive criteria up to and including 2012) is rare among Salmonella but a ciprofloxacin-resistant S.
Kentucky clonal group has arisen and spread from North Africa in the last decade. Two (S.Java and
S.Typhimurium DT120) isolates with resistance to ciprofloxacin were received from patients with unknown
foreign travel history.
In addition 39 isolates resistant to nalidixic acid and with reduced susceptibility to ciprofloxacin (minimum
inhibitory concentration of ≥ 0.12 mg/L) were detected, compared with 43 in 2011. It is worth noting that the
issue of interpretive criteria for resistance to ciprofloxacin for Salmonella has been controversial for a number
of years and some would regard all those isolates with reduced susceptibility as resistant. The NSSLRL
6
reports all such isolates with an interpretive comment pointing out that there is some uncertainty about how
such isolates respond to therapy with ciprofloxacin.
Travel related infection
A history of recent foreign travel was recorded in 44.8 % (n = 143) of human cases of infection (Table 3).
Ireland was noted as the country of infection in 46.4 % (n = 148) of cases while 8.8 % (n = 28) had no
country of infection recorded. Spain and Thailand were the most commonly recorded travel destinations.
S.Enteritidis accounted for the majority of isolates associated with travel to Spain (73.7%) and Turkey
(85.7%) while S.Stanley was strongly associated with travel to Asia (81.8%). Forty-four (78.6 %)
S.Enteritidis isolates were associated with foreign travel compared to 22.8 % for S.Typhimurium and its
monophasic variant combined. Although NSSLRL does not have access to data on the number of Irish
people who travel to each country it is likely that the number of cases associated with each country is at least
in part accounted for by the popularity of the country as a holiday destination.
Table 3: Foreign travel history for Salmonella isolates
Continent
Country
Number
Spain
19
Turkey
7
Poland
6
Italy
5
Croatia
4
Portugal
3
France
3
UK
3
Latvia
3
The Netherlands
2
Belgium
1
Bosnia
1
Malta
1
Czech Republic
1
Nigeria
3
Europe (n = 59)
Africa
(n = 14)
7
Ghana
2
Kenya
2
Tunisia
2
Morocco
2
Ethiopia
1
Africa*
1
Zambia
1
Australasia (n = 58)
Thailand
17
The Philippines
8
India
7
Indonesia
6
Bangladesh
5
China
3
Pakistan
3
Vietnam
3
Cambodia
1
Nepal
1
Singapore
1
Asia, S.America & Australia
Malaysia/Borneo
1
Malaysia/China
1
1
Americas (n = 10)
Foreign travel
Brazil
2
Cuba
2
USA
2
Uruguay
1
Colombia
1
Jamaica
1
Peru
1
Country unknown (n = 1)
* Country not stated
8
Clusters
Twelve clusters of cases involving 57 isolates were identified in 2012. S.Typhimurium/monophasic S.
Typhimurium was involved in 6 clusters (43 isolates) while S.Enteritidis was implicated in 2 clusters (4
isolates).
Four of the clusters were family outbreaks, i.e. all patients affected were from one family,
while 3 outbreaks were associated with foreign travel (all patients had travelled to same country).
The outbreak of S.Typhimurium phage type DT8 that began in August 2009 continued through 2010 and
2011. This phage type is associated with ducks and several of the cases were linked epidemiologically with
ducks or consumption of duck eggs. However the number of DT8 isolates from human cases typed in the
NSSLRL decreased from 28 in 2010 to 9 in 2011 and just 2 in 2012. This suggests that the control procedures
put in place are working, leading to reduced burden of human infection from this source.
The NSSLRL liaises with the European Centre for Disease Control (ECDC) in the investigation of outbreaks
that may have an international dimension, e.g.
S. Newport outbreak associated with consumption of
contaminated watermelons from a particular region of Brazil of which four cases were detected in Ireland.
Pulsed field gel electrophoresis and whole genome sequencing showed the isolates to be related.
Evidence of Links between S. Typhimurium in Humans, Food and Animals.
Multi locus variable number tandem repeat analysis (MLVA) is a relatively new technology that allows for
very fine discrimination between isolates that appear very closely related by other methods including PFGE.
This precise discrimination has proved a useful technique in showing persuasive evidence of links between
human S.Typhimurium clusters and individual cases and recent food and animals sources.
Table 4
illustrates evidence of links between human infection and food /food animal types. In some cases the link is
complete from animal, to animal related food product and to human cases.
The importance of using standardised and readily comparable methods for “fingerprinting” human, food and
animal isolates of Salmonella not just within Ireland but across Europe and the world is illustrated by this
body of work over the past year.
9
Table 4 MLVA profiles of human and food/animal isolates
Isolate no.
Source
Phage type
Resistance
MLVA profile
Typhimurium
(n = 2)
Human
DT8
none
2-9-NA-12-212
(n = 8)
Duck/Poultry
DT8
none
2-9-NA-12-212
* Or closely related pattern, i.e. 2-9-NA-11-212, 2-9-NA-13-212
(n = 2)
Human
Untypable
none
2-15-16-11-212
(n = 2)
Bovine
Untypable
none
2-15-16-11-212
(n = 2)
Human
Untypable
ACSSuTTm
3-13-16-19-311*
(n = 2)
Bovine
Untypable
ACSSuTTm
3-13-16-19-311
* Or closely related pattern, i.e. 3-13-16-16-311
(n = 1)
Human
DT104
ACSSuT
3-14-14-14-311
(n = 1)
Bovine
DT104
ACSSuT
3-14-14-14-311
(n = 5)
Human
DT104
ACSSuT
3-14-15-14-311
(n = 1)
Bovine
DT104
ACSSuT
3-14-15-14-311
(n = 1)
Canine
DT104
ACSSuT
3-14-15-14-311
(n = 1)
Human
DT104
ACSSuT
3-14-16-23-311
(n = 1)
Bovine
DT104
ACSSuT
3-14-16-23-311
(n = 1)
Human
DT104b
ASu
3-14-6-12-311
(n = 1)
Swine
DT104b
NT
3-14-6-12-311
(n = 1)
Human
DT104b
ACSSuT
3-15-6-12-311
(n = 1)
Bovine
DT104b
ACSSuT
3-15-6-12-311
(n = 1)
Human
DT12
T
3-17-11-8-211
(n = 1)
Bovine
DT12
NT
3-17-11-8-211
10
4,[5],12:i:(n = 1)
Human
U311
ASSuT
3-12-10-NA-211
(n = 5)
Swine
U311
NT
3-12-10-NA-211
(n = 1)
Equine
U311
NT
3-12-10-NA-211
(n = 3)
Human
DT193
ASSuT
3-13-10-NA-211
(n = 1)
Swine
DT193
NT
3-13-10-NA-211
(n = 1)
Bovine
DT193
NT
3-13-10-NA-211
(n = 2)
Human
DT193
ASSuT
3-13-11-NA-211
(n = 1)
Bovine
DT193
ASSuT
3-13-11-NA-211
(n = 1)
Human
DT193
ASSuT
3-14-11-NA-211
(n = 1)
Swine
DT193
NT
3-14-11-NA-211
(n = 1)
Human
DT193
ASSuT
4-11-9-NA-211
(n = 1)
Swine
DT193
NT
4-11-9-NA-211
(n = 4)
Human
DT120
SSuT
3-12-9-NA-211
(n = 3)
Swine
DT120
NT
3-12-9-NA-211
NT = Not tested
Some of the animal isolates have not been serotyped in the NSSLRL
Animal Contact
A history of animal contact was recorded for 91 patients with salmonellosis including contact with snakes,
lizards, turtles, birds, fish, horses, dogs and farm animals (Table 5). Dogs were the most common contact
animal (n = 48) while contact with cats was less common (n = 16).
In some cases typing of isolates from patients and individual animals they had contact with, e.g. S.Monschaui
isolate from a baby and a bearded dragon, had indistinguishable PFGE patterns.
Other strong links included single cases of S. subsp II 1,13,23:z29:e,n,x and turtle contact; contact with
lizards and cases of S. subsp IIIb 61:l,v:5,7, S. subsp IIIb 48:z:-, S.Godesberg, S.Fomeco and S.Muenchen.
There was one case of S.Typhimurium DT135 linked to pet birds. This strain was linked with pet canaries in
previous years. However a very similar strain has also been isolated from cattle.
11
Although public information on the risk (particularly to children) of contact with reptiles has been circulated
(http://www.fsai.ie/WorkArea/DownloadAsset.aspx?id=11207) it appears that this may not be reaching
relevant sections of the population or may not have resulted in modification of risk behaviour.
It may be
appropriate to consider if further steps to limit exposure of children to risk of salmonellosis from contact with
reptiles and other exotic pets is appropriate. In total 28 isolates of Salmonella (approx. 8.8 % of all human
cases) were associated with contact with exotic animals, predominately reptiles.
Among people that had a recorded history of living on farms or working with farm animals (n = 26),
S.Typhimurium/monophasic S. Typhimurium predominated (n = 18).
Many of the patients that had a history of animal contact also had other risk factors, e.g. recent history of
foreign travel, consumption of particular food products, etc. It is important to note that Salmonella is
primarily a foodborne disease and only a certain percentage of these animal contacts resulted in the patients’
infections.
12
Table 5: Animal contact history or strains associated with animal contact
Source
Sub1.
Strain
Animal Contact
MS120315
Human
I
Infantis
Snakes, cats
MS120351
Human
I
Java
Snake, terrapin, reptile farm
MS120455
Human
I
Enteritidis PT8
Snake, dog
MS120104
Human
IIIb
48:z:-
Lizards, llamas, elephant,
MS120153
Human
I
Braenderup
Lizard
MS120423
Human
IIIb
61:l,v:5,7
Lizard
MS120801 *
Human
I
Godesberg
Lizard
MS120035
Human
I
Monschaui
Bearded dragon
MS120094
Bearded dragon I
Monschaui
-
MS120095
Bearded dragon I
Monschaui
-
MS120124
Human
I
Fomeco
Bearded dragon
MS120561
Human
I
Muenchen
Bearded dragon
MS120606 *
Human
I
Cotham
Bearded dragon, dog
MS120358
Human
I
Bovismorbificans
Reptiles
MS120042
Human
II
1,13,23:z29:e,n,x
Turtle
MS120736 #
Human
I
4,[5],12:i:- U323
Tortoise, pony
MS120861 *
Human
I
4,[5],12:i:- Untypable
Turtle, dog, sheep
MS120340
Human
I
4,[5],12:i:- DT120
Canaries, dog
MS120759
Human
I
Typhimurium DT135
Canaries
MS120791
Human
I
4,[5],12:i:- DT120
Canaries
MS120572 *
Human
I
Typhimurium DT4var
Budgies
MS120490
Human
I
Enteritidis PT21
Goldfish
MS120626
Human
I
Bredeney
Goldfish, dog
MS120681 #
Human
I
4,[5],12:i:- U323
Goldfish
MS120656
Human
I
Braenderup
Pet fish
MS120376 *
Human
I
Enteritidis PT14c
Jungle farm
MS120563 *
Human
I
Java
Zoo, dogs
MS120569 *
Human
I
4,[5],12:i:- DT120
Elephant
MS120575 *
Human
I
Enteritidis PT4
Dolphins
MS120583 *
Human
I
Enteritidis PT63
Dehu (Chilean squirrel)
MS120584 *
Human
I
Enteritidis PT8
Guinea pig
NSSLRL no.
Exotic animals
dogs
13
Farm or Food animals (including horses)
MS120359
Human
I
4,[5],12:i:- DT193
Pig farm
MS120599
Human
I
Altona
Pig, dog
MS120267 *
Human
I
Enteritidis PT15a
Cattle
MS120339
Human
I
Typhimurium DT104
Cattle, horses
MS120352
Human
I
Typhimurium DT104
Cattle, sheep
MS120452 *
Human
I
Mgulani
Cattle
MS120570
Human
I
Typhimurium Untypable
Cattle
MS120580
Human
I
Typhimurium Untypable
Cattle, dog
MS120586
Human
I
Typhimurium Untypable
Cattle
MS120661
Human
I
Typhimurium DT104
Cattle, dog, cat
MS120740 *
Human
I
Oslo
Cattle
MS120752 #
Human
I
4,[5],12:i:- U323
Cattle, sheep
MS120258
Human
I
Typhimurium DT132
Farm, dogs
MS120492 *
Human
I
Typhimurium U311
Farm animals, cats
MS120794
Human
I
Typhimurium DT104b
Farm, dog, cat
MS120365 *
Human
I
Stanley
Hens
MS120592
Human
I
Typhimurium U288
Hens, dog
MS120651
Human
I
Typhimurium U310
Hens, dog, cat
MS120652 #
Human
I
4,[5],12:i:- U323
Ducks, hens
MS120674 *
Human
I
Saintpaul
Hens, cat
MS120001
Human
I
Senftenberg
Dog
MS120037
Human
I
Saintpaul
Dog
MS120078 *
Human
I
Typhimurium DT120
Dogs
MS120083
Human
I
Stanley
Dog
MS120097 *
Human
I
Stanley
Dog
MS120118
Human
I
Typhimurium DT56 var
Dogs
MS120357
Human
I
Stanleyville
Dog
MS120420
Human
I
Umbilo
Dog, sheep
MS120446
Human
I
Typhimurium DT104
Dogs
MS120493
Human
I
Typhimurium DT8
Dog
MS120579
Human
I
Cotham
Dog
MS120588
Human
I
Newport
Dog
MS120630
Human
I
Typhimurium Untypable
Dog
MS120631
Human
I
4,[5],12:i:- U323
Dogs
MS120668 #
Human
I
4,[5],12:i:- U323
Dog, cat
MS120669 #
Human
I
4,[5],12:i:- U323
Dog
MS120671 *
Human
I
Enteritidis RDNC
Dog, cat, pony
Domestic pets
14
MS120684 *
Human
I
Typhimurium DT193
Dog
MS120686 #
Human
I
4,[5],12:i:- U323
Dog, horses
MS120690 #
Human
I
4,[5],12:i:- U323
Dog, cat
MS120709 *
Human
I
Braenderup
Dog
MS120710 #
Human
I
4,[5],12:i:- U323
Dog
MS120712 *
Human
I
4,[5],12:i:- DT193
Dogs
MS120713 *
Human
I
Enteritidis PT1
Dog
MS120750
Human
I
Enteritidis PT21
Dog
MS120751
Human
I
Typhimurium DT104
Dog, cat, horses, rabbits
MS120787
Human
I
Typhimurium DT104
Dog
MS120800 *
Human
I
Panama
Dog
MS120803
Human
I
4,[5],12:i:- DT193
Greyhounds
MS120814
Human
I
Indiana
Dog
MS120815 *
Human
I
Mikawasima
Dog
MS120839
Human
I
Bredeney
Dog
MS120036
Human
I
Typhimurium DT104b
Cat
MS120102
Human
I
4,[5],12:i:- DT193
Cat
MS120441 *
Human
I
Typhimurium Untypable
Cat
MS120578
Human
I
Enteritidis PT14b
Cat
MS120685 * #
Human
I
4,[5],12:i:- U323
Cats
MS120804 *
Human
I
4,[5],12:i:- U311
Cat
MS120072
Human
I
Typhimurium DT104
Pet, not specified
1. Sub = Subspecies. There are over 2500 Salmonella serotypes of which approximately 60% belong to
subspecies I. These account for about 99% of human infections. Subspecies I is present in both warm and cold
blooded animals while the other Salmonella subspecies are generally associated with cold-blooded animals.
* Patient had history of recent foreign travel
# Strain part of large, international, probable food-borne outbreak
15
Non-Human isolates
In 2012, 391 isolates of Salmonella of non-human origin were submitted to the NSSLRL. This represents an
increase of 2.6 % in the number of non-human isolates received in 2011. The majority of isolates were from
swine (n = 275), bovine (n = 64) and poultry (n = 22) sources. S.Typhimurium/monophasic S. Typhimurium
(n = 357) were the most prevalent serovars (Table 6).
Table 6 Top ten serotypes among non-human isolates
Serotype
Frequency
%*
Typhimurium
32
8.2
Monophasic Typhimurium 8
Not serotyped
1
2.0
317
81.1
Enteritidis
4
1.0
Dublin
4
1.0
Java
4
1.0
Give
3
0.8
Senftenberg
3
0.8
Kentucky
2
0.5
Montevideo
2
0.5
Monschaui
2
0.5
Others
10
2.6
Total
391
100
* Approximate figures
1
These are Typhimurium/monophasic S.Typhimurium isolates received from CVRL for phage typing only.
Salmonella serotypes and correlation with Human Infection
S.Typhimurium
S.Typhimurium and its monophasic variant 4,5,12;i:- accounted for 91.3% of all non-human isolates and were
isolated from a variety of sources predominantly swine (n = 267) but also including poultry (n = 15) and
bovine (n = 55) sources. Phage types DT193 (n = 119), DT120/DT120 low (n = 33), DT104b (n = 29), U311
16
(n = 17), Untypable (n = 15), U302 (n = 10), DT104 (n = 9) and DT12 (n = 7) were the most common phage
types from swine. Phage types DT104 (n = 26), Untypable (n = 10) and DT193 (n = 8) and were the most
common phage type among bovine isolates. As illustrated in Table 4 (above) there is evidence from
molecular typing of a significant link between isolates from pigs and pig products and also with cattle and
human infection.
Salmonella Enteritidis.
Four S.Enteritidis isolates were received from non-human sources in the NSSRL in 2012, 2 from poultry and
2 from bovine sources. It is of interest that S. Enteritidis is rarely detected from poultry or poultry products in
Ireland but that S. Enteritidis is the first or second most common serovar isolated from human infections each
year. In this context it is worth noting that a history of travel outside of Ireland is reported in relation to 78.6
% of S. Enteritidis cases compared with only 22.8 % of S. Typhimurium/monophasic S.Typhimurium cases in
2012.
Sources of Isolates
Swine
A total of 6 serovars accounted for the 275 isolates received from swine with S.Typhimurium and its
monophasic variant 4,5,12:i:- (n = 267) accounting for the majority of isolates. Phage type DT193 was by far
the most frequent phage type (n = 119), while DT120/DT120 low (n = 33), DT104b (n = 29), U311 (n = 17)
and Untypable (n = 15) were also common in the swine isolates tested. These phage types were also typed
from human cases in 2012; DT193 (n = 17), DT120/DT120 low (n = 11), Untypable (n = 14), DT104b (n =
6), U311 (n = 3) and U302 (n = 2). MLVA typing provided further evidence of links between Salmonella in
swine and human infection (see Table 4).
Poultry
A total of 22 isolates comprising 5 serovars were received from poultry sources. As with swine
S.Typhimurium and its monophasic variant 4,5,12:i:- (n = 15) were the most common serovars received.
S.Typhimurium isolates were from a variety of poultry sources including duck and chicken while the S. Java
(n = 4) was isolated from chicken. Two isolates of S.Enteritidis were received, a PT4 isolate from duck and a
PT3 isolate from unspecified poultry while a single isolate of S. Mbandaka was received from a boot cover
from a poultry farm.
Isolates of all of these serovars were associated with human infections (S. Typhimurium/ monophasic
Typhimurium (n = 123), S. Enteritidis (n = 56), S. Java (n = 5), and S. Mbandaka (n = 1) in 2012).
Bovine
A total of 64 isolates were received from bovine sources and again S.Typhimurium and its monophasic
variant 4,5,12:i:- accounted for the majority of isolates (n = 55) with DT104 (n = 26), Untypable (n = 10) and
DT193 (n = 8) being the most common phage types. These phage types were also received from human cases
17
of salmonellosis in 2012. MLVA typing provided further evidence of links between Salmonella in cattle and
human infection (see Table 4). There were also 4 S. Dublin isolates (and 6 human cases).
Antimicrobial Resistance
Antimicrobial susceptibility testing was performed on 68 of the 391 non-human isolates (323 were referred
for phage typing only). Of the isolates tested 42.6% (n = 29) were susceptible to all antimicrobial agents
tested while 42.6% (n = 29) were multi-drug resistant (three or more different classes of antibiotics). Nineteen
percent of isolates tested had the profile of resistance to ampicillin, chloramphenicol, streptomycin,
sulphonamide and tetracycline (ACSSuT) and most of these typed as S.Typhimurium DT104 or closely
related phage types.
None of the isolates were resistant to ciprofloxacin; however 7 isolates exhibited resistance to nalidixic acid
and reduced susceptibility to ciprofloxacin as discussed earlier in this report.
Laboratory Contamination
False-positive Salmonella results due to laboratory cross-contamination are a serious problem for laboratories
and can be difficult to detect. Cross contamination in a laboratory can result in inappropriate diagnosis of
patient infection or in unfounded concerns regarding the safety of a food product. Detailed subtyping of
isolates by the NSSLRL helps in detection and confirmation of laboratory contamination incidents (Role of
Subtyping in Detecting Salmonella Cross Contamination in the Laboratory; BMC Microbiology: 9; 155).
One definite and 3 suspected contamination incidents occurred with human clinical samples where the
contaminating isolates arose from other patient’s samples.
We would like to reiterate our request that all laboratories involved in testing Salmonella from any source use
Salmonella Nottingham NCTC 7382 as their positive control.
18
Other Testing:
Listeria monocytogenes
Fig. 2
Pulse Field Gel Electrophoresis image of L.monocytogenes isolates digested with AscI
The NSSLRL received 33 Listeria monocytogenes isolates in 2012. These included 8 human clinical isolates;
6 from blood cultures, 1 from CSF and 1 from placental swab. Five of the isolates from humans typed as
serotype 4b, 1 typed as serotype 1/2a and 2 typed as 1/2b.
The majority of the food/animal/environmental isolates serotyped as 1/2a (n = 15), 6 isolates were serotype
4b, 3 were 1/2c and 1 typed as 1/2b.
The NSSLRL is working with colleagues in food and veterinary microbiology in Ireland and with colleagues
in Europe to build a library of typing data that may help to identify sources of human infection. A critical
limiting factor is the availability of human isolates for typing.
The total number of human clinical isolates in
Ireland per year is very small therefore it is critical that all such isolates are available for typing and we
appeal for all isolates to be forwarded for typing.
19
Shigella species
Fig. 3
Pulse Field Gel Electrophoresis image of S.sonnei digested with XbaI
100
95
PFGE-XbaI
90
85
PFGE-XbaI
MH120036
India
MH120045
Ireland
MH120034
MH120005
India
MH120037
India
MH120021
Spain
MH120033
Tanzania
MH120040
Ireland
MH120046
Ireland
A total of 36 isolates were referred to the NSSLRL in 2012 for Shigella typing. When non-Shigella, QC and
duplicate isolates were removed a total of 20 Shigella isolates were typed. These included 12 S.sonnei, 6
S.flexneri. and 2 S.dysenteriae. The S.flexneri isolates were further divided into 3 S.flexneri 6, 1 S.flexneri 2a,
1 S.flexneri 3a, and 1 S.flexneri 4a.
Seventy five percent of isolates (n = 15) were multi-drug resistant (three or more different classes of
antibiotics). One ESBL producing Shigella sonnei was received in the NSSLRL in 2012 while 6 isolates were
resistant to ciprofloxacin. The ciprofloxacin resistant isolates included 4 Shigella sonnei, 3 of whom had
travelled to India, and 2 S.flexneri isolates, one with history of travel to India.
Thirteen patients had a recorded history of recent foreign travel, including Europe (n = 1), Africa (n = 3),
Australasia (n = 7) and the Americas (n = 2). Six patients had Ireland recorded as their country of infection
while there were no details for 1 patient.
20
Table 7: Foreign travel history for Shigella isolates
Continent
Africa
Country
Number
Morocco
2
Tanzania
1
Spain
1
(n = 3)
Europe (n = 1)
Australasia (n = 7)
India
5
India & Nepal
1
Pakistan
1
Canada
1
Americas (n = 2)
Dominican Republic 1
There was 1 family outbreak (n = 2) of Shigella sonnei in 2012.
21
NSSLRL Publications and Presentations 2012
Papers
Multilocus sequence typing as a replacement for serotyping in Salmonella enterica. Achtman M et al.
PLoS Pathog. 2012;8(6):e1002776
PFGE analysis of Listeria monocytogenes isolates of clinical, animal, food and environmental origin from
Ireland. Fox EM et al. J Med Microbiol. 2012;61: 540-7.
22