Investigating Coxiella burnetii infection in a breeding cattery at the centre of a Q fever outbreak
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487360
2013
JFM0010.1177/1098612X13487360Journal of Feline Medicine and SurgeryKopecny et al
Original Article
Journal of Feline Medicine and Surgery
Investigating Coxiella burnetii 0(0) 1–9
© ISFM and AAFP 2013
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infection in a breeding cattery at the sagepub.co.uk/journalsPermissions.nav
DOI: 10.1177/1098612X13487360
centre of a Q fever outbreak jfms.com
Lucy Kopecny, Katrina L Bosward, Amanda Shapiro and
Jacqueline M Norris
Abstract
The potential role of cats in transmitting Coxiella burnetii to humans was highlighted in a Q fever outbreak, linked
to a caesarean section in a breeding queen, in an Australian small animal veterinary hospital. The objectives of
this study were to evaluate the C burnetii seroreactivity of the breeding queen and other cats residing at the same
breeding cattery (n = 27) and to evaluate C burnetii infection of the breeding queen by molecular and histological
methods. Three assays [complement fixation test (CFT), indirect immunofluorescence assay (IFA) and enzyme-
linked immunosorbent assay (ELISA)] were used for serological evaluation. Additionally, uterine and ovarian samples
collected from the breeding queen 11 weeks post-parturition were assessed by routine and specialised histological
methods and polymerase chain reaction. The breeding queen showed strong seropositivity using CFT (titre 1/32),
IFA (titre phase I 1/8192 and phase II 1/8192) and ELISA; however, the reproductive tract showed no evidence of
pathology or C burnetii infection. A number of cattery-confined cats were identified as seropositive to phase II and/
or phase I C burnetii. Serological detection of C burnetii in a breeding cattery linked to a Q fever outbreak indicates
likely infection by this bacterium in Australian feline populations, re-confirming the relevance of this zoonosis.
Accepted: 28 March 2013
Introduction
Coxiella burnetii is the aetiological agent of Q fever, a 1.9–42% of cats have been seropositive,4,5,8,18 suggesting
highly significant worldwide zoonosis with a large res- infection of cats may be common.
ervoir encompassing wild and domestic mammals, Several community-acquired Q fever outbreaks have
birds and arthropods.1 It is an obligately intracellular likely resulted from direct or indirect exposure to
Gram-negative bacterium with extreme environmental queens and/or their kittens during or shortly after par-
resilience.1–3 Coxiella burnetii produces several poten- turition,9–11,13,14,17 probably via the infected animal’s
tially debilitating forms of Q fever in humans, render- birth products, where up to 109 bacteria per gram of
ing it a significant public health concern.1 While placental tissue have been reported in other species.1,19
infection is most commonly asymptomatic, 40% of pri- Coxiella burnetii has been detected in the blood, urine
mary infections in humans are symptomatic, with seri- and genital tracts of infected cats.11,13,16,20 While it
ous acute or chronic illnesses possible.1 Death can appears cats may be infected by and shed C burnetii, it
occur in both acute and, more commonly, chronic is uncertain whether infection is associated with clini-
infections.1 cal disease in cats. Several reports have suggested
Q fever is traditionally associated with contact with
cattle, sheep and goats in the livestock and meat indus-
tries.1,3 Less commonly, cats have been suggested as Faculty of Veterinary Science, The University of Sydney, Sydney,
sources of C burnetii in southern Africa,4 Japan,5–8 Korea,5 Australia
Canada9–15 and the USA.16–18 In maritime Canada, expo-
sure to parturient cats and newborn kittens has been Corresponding author:
Jacqueline Norris BVSc, MVS, PhD, MASM, GradCertEd, Faculty
identified as a significant risk factor for Q fever,11 with of Veterinary Science, Building B14, The University of Sydney,
seroprevalence of C burnetii infection in cats in these Sydney, NSW 2006, Australia
regions varying from 6.2 to 24%.12,15 In other countries, Email: jacqui.norris@sydney.edu.au2 Journal of Feline Medicine and Surgery 0(0)
Table 1 Collection dates and sequence of serum samples taken from cats in the breeding cattery
Collection date Months since outbreak Cat/s sampled First collection Second collection
July 2010 1.1 Index cat
April 2011 11 Cats 2–22
September 2011 16 Cat 23
February 2012 22 Cats 2, 6, 9
April 2012 23 Cats 24–27
infection in cats, as in other species, may be linked to Materials and methods
reproductive disorders.1,6,11 Samples
Diagnosis of C burnetii infection in animals is com- Serum samples were collected from the index cat and 26
plex owing to the lack of known disease associations other cats from the same breeding cattery. A second
and few available sensitive, specific diagnostic tech- serum sample was acquired from three of these cats at
niques. In animals, diagnosis primarily relies on serol- different time points. Collection dates of all serum sam-
ogy.21 The World Organisation for Animal Health ples (n = 30) are given in Table 1. Whole blood (0.5–3 ml)
(OIE) reference test for serological diagnosis of C bur- from each cat was collected in serum separator tubes and
netii remains the complement fixation test (CFT),22 centrifuged at 12,000 g for 10 min. Serum was harvested,
despite various more sensitive tests, including the divided into 300 μl aliquots and stored at –20°C until
indirect immunofluorescence assay (IFA) and enzyme- tested. Samples from the ovaries, left and right uterine
linked immunosorbent assay (ELISA), being available horns, and uterine body were collected from the index
for use in humans, where the gold standard is IFA.1,21 cat at ovariohysterectomy, 11 weeks post-parturition,
While CFT is highly specific, IFA and ELISA have with half fixed in formalin and the other half frozen
higher sensitivity, detect seroconversion earlier and immediately at –20°C.
are more rapidly performed.1,21 Other techniques for
demonstrating C burnetii in biological samples include
polymerase chain reaction (PCR) and immunohisto- Serological testing
chemistry.1,21 Recently, fluorescent in situ hybridisa- CFT Serum samples (n = 30 from 27 cats) were submitted
tion (FISH) has also been evaluated for detecting to the Elizabeth Macarthur Agricultural Institute, Menan-
C burnetii in the placental tissues of naturally-infected gle, Australia, for testing by CFT (OIE reference test22).
ruminants, with comparable sensitivity and specific-
ity to immunohistochemistry.23 IFA A modification of a commercial human C burnetii
The potential role of cats in C burnetii transmission to IgG/IgM/IgA IFA (Vircell) standardised previously by
humans was highlighted by a Q fever outbreak in a the authors, was used to detect IgG antibodies to phase
Sydney small animal veterinary hospital following a cae- I and phase II C burnetii (Nine Mile strain). Briefly, feline
sarean section in a breeding queen in June 2010.24 Nine serum samples (n = 30 from 27 cats) were initially
veterinary personnel, in addition to the queen’s owner, screened at 1/256 dilution using 5% skim milk powder
showed evidence of recent C burnetii infection. Six of the (SMP) in phosphate buffered saline (PBS; Vircell) for
nine veterinary personnel were symptomatic with mild- C burnetii seropositivity. Two sets of phase I and phase II
to-severe ‘flu-like’ signs, with two requiring extended slide wells in each run contained positive and negative
hospitalisation. Those most severely affected had human control solutions from the kit. Diluted feline
assisted the caesarean section and performed mouth- serum was added to remaining wells and slides were
to-mouth resuscitation on newborn kittens, and there- then incubated in a humid chamber at 37°C for 30 min.
fore likely experienced the most significant exposure to Following rinsing in PBS and then water, slides were
the queen’s birth products.24 To date, investigations of allowed to air dry. Anti-human IgG fluorescein isothio-
cats involved in Q fever outbreaks have focused on CFT cyanate (FITC) conjugate solution from the kit was
and IFA to demonstrate C burnetii antibodies.9,10,13,14 This applied to positive and negative control wells and anti-
article uses three standardised serological assays (CFT, feline IgG FITC conjugate solution (VMRD) to the
IFA and ELISA) in combination with histological and remaining wells. Slides were incubated, washed and
molecular techniques (FISH and PCR) to evaluate C bur- dried as above then read under a fluorescent microscope
netii infection in the breeding queen identified as the (Olympus BX60F-3; Olympus) at 400× shortly thereafter
probable primary source in the Q fever outbreak (index by two of the authors independently. Samples read as
cat) and the seroreactivity of cats residing at the same positive on phase I and/or phase II at titre 1/256 were
breeding cattery. serially diluted to end titre using the same method.Kopecny et al 3
ELISA A modification of the commercially available approximate 95% confidence interval (CI).25 Minitab
Panbio Q fever IgG ELISA (Alere) (previously stan- Version 15 (Minitab) was used for analysis.
dardised by the authors) was used to detect IgG antibod-
ies to phase II C burnetii (Henzerling strain) in feline Histological examination and FISH
serum samples (n = 30 from 27 cats). With the exception Formalin-fixed, paraffin-embedded 5 μm sections of the
of microwells containing control and calibrator samples, index cat’s ovaries, left and right uterine horns, and uter-
microwells were incubated at room temperature for 1 h ine body were stained using haematoxylin and eosin,
with 5% SMP in PBS. Dilution of positive and negative Gram Twort and Giemsa. To assess for C burnetii within
control and calibrator samples to 1/100 was performed the index cat’s ovaries, left and right uterine horns, and
using Tris-buffered saline (pH 7.2–7.6) (Alere). To dilute uterine body, formalin-fixed, paraffin-embedded
feline serum samples to 1/100, 5% SMP in PBS was used. histological sections (4 µm) from each tissue were
Positive and negative control samples were run singly, mounted on Probe-On Plus slides (Fisher Scientific) and
the calibrator sample in triplicate and feline serum sam- evaluated by FISH using a eubacterial probe (EUB-338;
ples in duplicate. The plate was covered and incubated GCTGCCTCCCGTAGGAGT) by a previously described
for 30 min at 37°C after applying samples to microwells. technique.26 Slides were examined under a fluorescent
The microwells were washed six times using PBS con- microscope (Olympus BX60F-3) at 400×. Control samples
taining 0.05% Tween 20. To microwells containing con- were formalin-fixed, paraffin-embedded tissues pre-
trol and calibrator samples, horseradish peroxidase pared by forming a ‘bacterial sandwich’ using lung tis-
(HRP) conjugated anti-human IgG was applied. The sue.27 Control bacteria in these preparations were
same volume of HRP conjugated anti-feline IgG (Peroxi- Staphylococcus pseudintermedius and Pseudomonas aerogi-
dase-conjugated AffiniPure Goat Anti-Cat IgG; Jackson nosa. Coxiella species controls could not be prepared
ImmunoResearch Laboratories) was applied to microw- owing to the lack of physical containment 3 facilities
ells containing feline samples. The plate was covered, required for handling this bacterial species.
incubated and manually washed as above. Tetramethyl-
benzidine was applied to each microwell, the plate incu- PCR
bated for 10 min and then the reaction stopped by adding Frozen samples of the index cat’s ovaries, left and right
1 M phosphoric acid. Within 30 min plate microwells uterine horns, and uterine body were submitted to the
were read in a microtitre plate reader at wavelength 450 Australian Rickettsial Reference Laboratory, Geelong,
nm with reference filter 600 nm to determine optical Australia, for real-time PCR testing for C burnetii.
density (OD).
Two methods were used to assess OD results.
According to the manufacturer’s instructions, a sample
Results
absorbance/calibrator absorbance ratio (index value)
Summary of Q fever outbreak in a small animal
was calculated, with calibrator absorbance determined
veterinary hospital
by calculation of the average absorbance of the calibra-
Sydney South West Public Health Unit conducted an
tor in triplicate and multiplication of this by the calibra-
epidemiological investigation into the cluster of human
tion factor supplied with each kit batch. Positive
Q fever cases at a small animal veterinary hospital.24 All
samples were defined as having an index value >1.1.
animals that underwent caesarean section in the identi-
Those that were negative were4 Journal of Feline Medicine and Surgery 0(0)
Study population were also positive to anti-phase I antibodies. All cats
The index cat was a 4-year-old Burmese queen from an seropositive using IFA had antibody titres to phase II C
urban breeding cattery with a history prior to 2010 of burnetii greater than or equal to their antibody titres to
having produced three litters normally, although some phase I C burnetii. Calculating ELISA results by S/P%
were of small size (one kitten only). The breeding for resulted in 6/27 (22%) cats being positive to anti-phase II
each of these litters occurred within the cattery. She was C burnetii antibodies; these cats were also all detected as
bred to an external Burmese male stud cat in April 2010 seropositive to phase II C burnetii by IFA. Eleven of 27
– the only occasion she had been outside the cattery. At (41%) cats were seropositive when ELISA results were
this mating, she did not have contact with any animals calculated using index values. Of five additional cats
outside that cattery. She had no known prior illnesses, detected as seropositive based on index values com-
and, during the pregnancy in 2010, was in good health. pared to S/P% none were detected as seropositive to
The index cat and cat 23 were confined indoors, while phase II C burnetii by IFA.
remaining cattery-confined cats had free-roaming out- In three cats from which a second sample was collected,
side access. Most cats in the cattery were fed a diet com- there were no changes to seroreactivity using the serologi-
prised predominantly of raw beef supplemented by cal assays with the exception of ELISA results for cat 2. The
commercial pet food. The index cat and cat 23 refused anti-phase I and II C burnetii antibody titres of cat 2 using
raw beef and were therefore fed commercial pet food IFA also decreased between the two sampling events (from
and raw chicken necks. 1/1024 to 1/512 and 1/8192 to 1/512, respectively).
The index cat commenced parturition on 31 May 2010 A comparison between phase II IgG IFA and ELISA
with a live kitten born without assistance that evening. results is provided in Table 3. Results for IFA and
Prolonged labour without fetal delivery and failure to ELISA S/P% suggest good agreement and, for IFA and
respond to 2.5 IU oxytocin subcutaneously led to caesar- ELISA index values, moderate agreement between
ean section the following morning at which the veteri- these tests.25
narian assessed the uterus to be normal in view of the
duration of dystocia. The placenta was unremarkable. Histological and molecular analysis
One live and one dead kitten were extracted and received The ovarian cortical stroma contained moderate num-
mouth-to-mouth resuscitation; both appeared small for bers of growing follicles. The endometrium was lined by
gestational stage. Following caesarean section, the index simple cuboidal-columnar epithelium in longitudinal
cat was administered 75 mg amoxicillin subcutaneously folds, with moderate hyperplasia of endometrial glands
and was discharged immediately. Antibacterial therapy that were also tortuous and contained scant luminal
continued with 50 mg amoxicillin clavulanate orally secretions. There was moderate myometrial hypertro-
twice daily. After identification as the probable primary phy. Throughout the endometrium and myometrium
source of C burnetii in the Q fever outbreak, the index cat were moderate numbers of macrophages, some of which
was administered 25 mg doxycycline orally twice daily were multinucleated, containing brown granular pig-
for 14 days commencing 4 weeks after the initial caesar- ment (likely haemosiderin). There was marked conges-
ian. The index cat’s ovaries and uterus were grossly nor- tion of vessels in the stratum vasculare of the myometrium
mal at ovariohysterectomy 11 weeks after caesarean and perimetrium. These findings were consistent with a
section. The two live kittens from the gestation devel- post-partum uterus.28 Bacteria were not visualised in
oped normally. Giemsa or Gram Twort-stained sections. Staining using
Of the 27 cats from the breeding cattery, 11 (41%) were the EUB-338 probe did not reveal bacteria. Further stain-
female entire, 11 (41%) were female neutered, four were ing using a C burnetii-specific probe was therefore not
male entire (15%) and one male was neutered (3.7%). undertaken. PCR analysis of the index cat’s reproductive
Burmese comprised five (19%) and Cornish Rex 22 (81%) tissues for C burnetii was negative.
of 27 cats within the cattery population. The median age
at initial sampling was 6 years (range 3 months to 17 Discussion
years). From initial sample collection to May 2012, three This study was a feline-focused investigation of an
of 27 (11%) cats died. Signalment of individual cats and important Q fever outbreak in a small animal veterinary
available further information are provided in Table 2. hospital following a caesarean section on a breeding
queen, the index cat. Previous investigations of feline-
Serological testing associated outbreaks have demonstrated serological evi-
Coxiella burnetii serology results from CFT, IFA and dence of C burnetii infection in the cat at their centre, but
ELISA in individual cattery-confined cats are summa- focused on human perspectives.9–11,13,14 In this study, the
rised in Table 2. Of the 27 cats, two (7.4%) were positive index cat showed marked antibody responses to phase I
using CFT. Using IFA, 7/27 (26%) cats were positive to and phase II C burnetii using all serological assays.
anti-phase II C burnetii antibodies, of which six (22%) Extending the investigation to assess seroprevalence inTable 2 Signalment and Coxiella burnetii serological results for all cattery-confined cats. The reciprocal antibody titre is provided for positive complement fixation
test (CFT) and indirect immunofluorescence assay (IFA) results. Where a second sample was collected, results are given as first sample collection/second sample
collection
Kopecny et al
Cat Breed Sex Age at first sample Further information CFT IFA ELISA
collection (years)
Phase I Phase II (S/P%) Index value
Index cat Burmese FE 4 32 8192 8192 + +
2 Cornish Rex FN 8 –/– 1024/512 8192/512 + /– +/–
3 Cornish Rex FN 14 – – – – +
4 Cornish Rex FE 5 – – – – –
5 Cornish Rex MN 9 – – – – –
6 Cornish Rex FE 6 Now deceased — feline infectious peritonitis –/– –/– –/– –/– –/–
7 Cornish Rex FE 6 – 512 512 + +
8 Cornish Rex FN 8 – 256 256 + +
9 Cornish Rex FE 3 Neutered after two stillborn litters –/– –/– –/– –/– –/–
10 Cornish Rex FE 3 – – – – +
11 Cornish Rex FE 5 – – – – –
12 Cornish Rex FN 15 Now deceased — congestive heart failure – – 256 + +
13 Cornish Rex FN 14 – 256 512 – –
14 Cornish Rex FN 4 Daughter of cat 17 – – – – –
15 Cornish Rex ME 3 – – – – –
16 Cornish Rex FE 9 – – – – –
17 Cornish Rex FN 6 – – – – –
18 Cornish Rex FE 3 – – – – –
19 Cornish Rex FN 9 – – – – +
20 Burmese ME 7 Source of reproductive failure — small litter size – 512 1024 + +
21 Burmese FN 17 – – – – +
22 Burmese FN 14 Now deceased — chronic renal failure – – – – +
23 Burmese FE 3 Daughter of cat 1 – – – – –
24 Cornish Rex ME 0.3 Son of cat 6 8 – – – –
25 Cornish Rex ME 0.3 Son of cat 6 – – – – –
26 Cornish Rex FE 0.3 Daughter of cat 6 – – – – –
27 Cornish Rex FE 2 Daughter of cat 6 – – – – –
ELISA = enzyme linked immunosorbent assay; S/P% = sample/positive ratio; FE = female entire; FN = female neutered; MN = male neutered; ME = male entire
56 Journal of Feline Medicine and Surgery 0(0)
Table 3 Percent agreement and Cohen’s kappa coefficient for phase II IgG indirect immunofluorescence assay (IFA)
and enzyme-linked immunosorbent assay (ELISA) results
Tests compared Percent agreement 95% CI Cohen’s kappa coefficient 95% CI
IFA and ELISA S/P% 93 78, 99 0.81 0.46, 1.2
IFA and ELISA index value 77 58, 90 0.47 0.12, 0.82
CI = confidence interval; S/P% = sample/positive ratio
cats from the same cattery demonstrated past or current demonstrated, approximately 4 weeks post-outbreak,
C burnetii infection in 26% of these cats using IFA. This strong positivity to anti-C burnetii IgG antibodies, in
evaluation of seroprevalence was enabled by using three accordance with her probable role as the source of C bur-
standardised serological assays. netii in the Q fever outbreak. Previously reported anti-
Determining C burnetii infection in cats has been com- body titres using IFA in cats linked to community-acquired
plicated by the absence of sensitive, specific diagnostics, Q fever outbreaks have varied between 1/32 and 1/8192
particularly as serological tests cannot be translated to phase I and between 1/8 and 1/8192 to phase II anti-
directly from one species to another without standardisa- gens;10,11,13,14 the index cat’s high antibody titres therefore
tion. In humans, IFA and ELISA are more sensitive than appear consistent with those from earlier investigations.
CFT1,21 (the OIE reference technique)22 and therefore val- Variable anti-phase I and phase II antibody titres were
uable for serodiagnosis in felines. The IFA and ELISA detected in the remaining cattery-confined cats up to 22
methodologies used in this study were recently standard- months post-outbreak. This suggests such antibodies
ised in cats by the authors. Coxiella burnetii seropositivity may remain raised for some time after infection in cats,
in the index cat was detected by all three serological though it is uncertain whether exposure to C burnetii
assays. A challenge in assessing the remaining cattery- continued in the population. In humans, anti-phase II
confined cats seropositive by IFA and/or ELISA, but neg- IgG titres have been identifiable 12 years after a Q fever
ative by CFT, was the lack of a ‘gold standard’ against outbreak, though CFT antibodies generally decrease ear-
which to evaluate their validity. While inter-test agree- lier.35,36 Cell-mediated immunity, important in control-
ment suggested IFA and ELISA to be sensitive and spe- ling C burnetii infection in humans,1 was not assessed in
cific in the study population,25,29 definitively establishing the study, likely limiting detection of all infected cats
these as alternatives to CFT requires evaluation in large, Previously, seroprevalence studies have indicated
representative feline populations. Further evaluation is stray cats or dogs may have higher rates of C burnetii
also required to determine the preferred method of ELISA seropositivity than those that are client-owned, with
analysis as the small sample size in this study was rodents a possible reservoir of infection.5,18 In view of the
limiting. high seroprevalence in the breeding cattery studied, cat-
Cross reactions between C burnetii and other organ- tery-confined breeding cats may also represent a popula-
isms, including Legionella species and Bartonella species tion at increased risk of C burnetii infection, though this
have been described in human sera using certain sero- requires further investigation. Likely important risk fac-
logical techniques.1,29–31 Cross reactions involving IgA tors within breeding catteries include more frequent
and IgM have been described between C burnetii and exposure to parturient animals (C burnetii is abundant in
Bartonella henselae and Bartonella quintana in human sera the birth products of infected individuals1), as well as
using IFA, likely owing to similarity between protein high population densities. The primary source of C bur-
antigens.1,30 The extent of serological cross-reactions netii for cats in the breeding cattery at the centre of this
involving C burnetii is unknown in feline sera; however, outbreak was unable to be established. The exposure of
as cats are recognised as the primary reservoir hosts for the index cat to the male stud at mating could not be
several Bartonella species, including B henselae,32 poten- excluded as we were not able to acquire serum samples
tial cross reactions may warrant consideration. In our to assess this cat’s seroreactivity. Alternatively, reactiva-
study, the absence of cross-reactions using IFA between tion of previously latent C burnetii in the next cat may
C burnetii and B henselae was able to be confirmed in test- have occurred. The active predatory behaviours of some
ing by the manufacturer (Vircell). cats have been proposed to increase their likelihood of
In C burnetii-infected humans, elevated phase II IgM infection by C burnetii by close contact with reservoirs
antibodies are detected within 14 days of signs appear- such as rodents.37,38 Contact with rodents or other poten-
ing, followed later by IgM antibodies to phase I.1,33,34 tial animal sources of C burnetii was unlikely in the index
From 2–3 weeks, IgG antibodies to phase II reach high cat, but possible for cattery-confined cats with outside
levels, with phase I IgG responses developing more access. Other forms of dietary access, such as ingestion
slowly and at lower titres.1,33,34 The index cat of raw meat, are possible, with ingestion of high doses ofKopecny et al 7
C burnetii a recognised, albeit rare, source of infection in recommendations to wear appropriate personal protec-
humans.1 tive equipment (PPE) where potential for exposure to
Studies describing well-defined disease associations infective materials, particularly birth products, exists and
with feline C burnetii infection are lacking. Experimental avoid direct mouth-to-mouth or mouth-to-nose resuscita-
infection of cats has caused fever, inappetence and leth- tion of neonates.41–43 However, as well as needing to
argy.20 Reproductive abnormalities in periparturient cats improve and expand these guidelines, adopting such pro-
associated with Q fever outbreaks have also been cedures within hospitals will be dependent on adherence
described, yet such relationships are likely complex. The to and awareness of written infection control guidelines
reported history of some queens and their kittens was by veterinary staff, in particular the requirements for
unremarkable,9,10,17 while others have exhibited signs, PPE.44,45 In conjunction with infection control procedures,
including bleeding per vaginum for 3 weeks prior to vaccination of all small animal veterinary personnel
parturition,14 stillbirths and death of kittens shortly post- against Q fever may gain importance as the epidemiology
parturiently.13 In a study of Q fever cases connected to of C burnetii infection in companion animals unfolds.
exposure to felines, 7/10 queens had one or more still- Future seroprevalence studies will be valuable in
born kittens.11 In other species, manifestations of determining the importance of C burnetii in feline popu-
C burnetii infection during pregnancy encompass abor- lations and enabling enhanced understanding of the risks
tion, prematurity, stillbirth, low birth weight and neona- cats pose as sources of C burnetii in Australia. Together
tal weakness, with these connected to placentitis and/or with epidemiological data, elucidation of disease associa-
direct fetal injury.1,3,39 Signs in the index cat, including tions with C burnetii is required to advance our under-
stillbirth and small kittens relative to gestational stage, standing of the pathogenesis of feline infections.
may have been due to C burnetii infection, though other
contributing factors cannot be excluded. Of other C bur- Conclusions
netii seropositive cattery-confined cats with known dis- Serological detection of C burnetii in a breeding cattery
ease, it was difficult to determine the contributions of linked to a Q fever outbreak amongst veterinary hospital
their serological status. For several years, a breeding personnel indicates there is likely infection by this bacte-
male (cat 20) was considered a source of reproductive rium in Australian feline populations. However, many
failure that resolved when affected females were bred to questions remain regarding the epidemiology and disease
a different male. The reproductive failures associated outcomes following infection in this species, both in
with cat 20 have now resolved. It is questionable whether Australia and globally. This outbreak’s association with a
C burnetii was the underlying cause, particularly given caesarean section on a parturient queen, together with the
that this cat was C burnetii seropositive. However, inter- elevated C burnetii seroprevalence in the cattery from which
estingly, in humans, C burnetii infection has been associ- this queen arose, re-confirm the relevance of this zoonosis.
ated with orchitis, epididymitis and priapism.1,36
Duration of C burnetii shedding from the reproductive Acknowledgements We would like to thank Dr Susan
tract of naturally infected cats is unknown, though shed- Piripi for reviewing the histologic sections. We are especially
ding in animals is primarily recognised periparturiently.1 grateful to the practice owners and veterinary staff at the
Coxiella burnetii has been isolated from feline uteruses 3 and hospital at the centre of this outbreak, and also to the owner
8 weeks post-parturition, but could not be isolated from of the cattery who has been so incredibly cooperative in
another uterus 10 weeks post-parturition.11,13 In the present allowing cats to be tested and for providing thorough clini-
cal and breeding histories. We are grateful to the Australian
investigation, inability to detect C burnetii histologically
Rickettsial Laboratory in Geelong Melbourne for PCR test-
may represent true- or false-negative results. Several fac-
ing, free of charge. The corresponding author (Norris) would
tors, particularly the interval between caesarean section like to thank Professor Kenneth Simpson for teaching her to
and ovariohysterectomy, during which the index cat was perform FISH.
administered doxycycline—the antibacterial of choice in
empirical treatment of acute Q fever in humans36—were Funding This research was supported by a competitive grant
likely to limit findings in this case. Although false-negative from the Australian Companion Animal Health Foundation.
results can occur using FISH with metabolically inactive
bacteria due to low ribosomal RNA content,40 negative Conflict of interest The authors do not have any potential
PCR results on these tissues increased our suspicion that C conflicts of interests to declare.
burnetii was, indeed, no longer present.
As cats are recognised as C burnetii sources, develop- References
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