Brucella ovis – an update on control

Brucella ovis – an update on control

Anne Ridler
Massey University, Private Bag 11-222, Palmerston North 4442

Introduction and history

Brucella ovis was first recognised in NZ in the early 1950’s and breeding soundness examination of rams and
diagnosis of this condition have historically been, and still are, important reasons for veterinary involvement
on sheep farms. The concept of developing a NZ voluntary B. ovis accreditation scheme was first raised by Neil
Bruere in 1973 and in the late 1970’s such a scheme was trialled in stud and commercial flocks by Neil Bruere
and Dave West. In 1986 a nationwide voluntary accreditation scheme was launched.

Current status of B. ovis in NZ

No official national database is kept of the number of flocks that test for B. ovis but Gribbles Veterinary have
provided data on the number of infected flocks over the past three years. This shows cases are relatively
uncommon but have occurred throughout NZ including areas traditionally thought to be largely free of B. ovis
such as Otago and Southland (Figure 1). The actual number of flocks tested is not known but it is probable the
majority of commercial farmers do not get their rams tested to accredited-free standards.

Figure 1. Number of ram flocks known to be infected with B. ovis in New Zealand from 2009-2011

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 Brucella ovis – an update on control

Diagnosis of B. ovis

Scrotal palpation and epididymitis

Palpation of the scrotal contents is a quick, cost-effective procedure that is of primary importance for evaluating
ram breeding soundness but is also useful as a flock screening test for B. ovis. However it is important to
recognise that not all infected rams develop epididymitis. Anecdotally the percentage of seropositive rams with
epididymitis appears to vary markedly between flocks, possibly related to the chronicity of the infection and/or
the proportion of rams that are persistently vs. transiently infected. In one published study 30 out of 88 (34%)
of rams that were shedding B. ovis in semen had detectable epididymitis (Hughes and Claxton 1968). Absence
of rams with epididymitis in a flock does not necessarily mean it is free from B. ovis.

There is a common perception that B. ovis causes lesions in the tail of the epididymis while Gram-negative
pleomorphs cause lesions in the head of the epididymis. In one published study 90% of lesions due to B. ovis
were in the tail of the epididymis (Kennedy et al. 1956) but it is important to recognise that B. ovis can also cause
lesions in the epididymal head, just as Gram-negative pleomorphs can cause lesions in the epididymal tail. Hence
the location of epididymal lesions (tail vs. head) cannot be used to differentiate between the two infections.
Similarly another perception is that B. ovis only affects older rams whereas Gram-negative pleomorph infections
usually affect young rams. While B. ovis is probably more common in older rams it has been demonstrated in
rams as young as 4-6 months of age (Bulgin 1990). Gram-negative pleomorphic infections usually develop in
rams around puberty but the lesions persist indefinitely so in rarely inspected flocks older rams may be found
with epididymitis due to Gram-negative pleomorphs.

Serology
Serology is the most straightforward and cost-effective method of testing for B. ovis. In NZ 3 serological test
options are available – a Complement Fixation Test (CFT), an Enzyme-Linked Immunosorbent Assay (ELISA)
and a Gel Diffusion test (GD). There is occasionally debate about whether the CFT or ELISA is the best option
for use as a screening test as both have their own advantages and disadvantages but in NZ the CFT is routinely
used as the screening test of choice. Inevitably none of these tests are perfect so false-positive and false-negative
results will occur from time to time.

Semen culture
Semen culture can be useful in situations when doubt exists as to whether a ram(s) is truly infected or is a
serological false-positive, or in high-stakes situations where you want complete confidence that infection is truly
present (e.g. a potential breakdown in an accredited-free pedigree flock). Semen culture is often perceived as
difficult and unreliable but in reality B. ovis is easy to grow. However it is slow-growing (4-5 days for decent
growth) so it cannot compete in the face of contamination with other more rapidly growing bacteria. Best
results can be expected if the sample is collected with minimal contamination and dispatched to the lab as soon
as possible. Thus:
• Warn the lab ahead of time that you are going to send in a sample(s) for B. ovis culture – this will
give them time to order selective media (Thayer-Martin).
• Manually extrude the penis during semen collection.
• Collect into a sterile sample pot (we use sterile milk sample pots).
• Collect the semen in the morning early in the week and get it off to the lab a.s.a.p. (e.g. don’t
collect it on a Friday afternoon!). If it can’t be sent to the lab that day, refrigerate overnight.

Histopathology and/or culture of the genital organs

On rare occasions histopathology and/or culture of the genital organs may be used to confirm or rule-out B.
ovis infection. Brucella ovis most commonly resides in the epididymides and seminal vesicles so both these
organs should be submitted. If culture is required then the considerations outlined above regarding minimal
contamination and rapid dispatch to the lab should be adhered to. However a carefully collected semen sample
may be a better option.

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 Brucella ovis – an update on control

B. ovis investigation

Non-accredited flocks
The approach to investigation of B. ovis in non-accredited flocks should be tailored to individual farms based on
a risk assessment for that farm. For flocks that are considered low-risk then scrotal palpation alone (as part of a
breeding soundness examination), and blood sampling any rams with epididymitis, is likely to be sufficient. In
these flocks routine annual blood testing for B. ovis is likely to be an unnecessary extra expense. In contrast, for
flocks considered medium or high-risk then blood testing the flock in addition to palpation could be considered,
either as a routine annual or bi-annual event or in response to a specific risky incident(s).

Risk and risky practices

The following are not exhaustive lists but give an indication of factors which might be associated with increased
or decreased risk of a flock being or becoming infected with B. ovis.

Medium and high-risk flocks – ANY of the following:

• Unknown B. ovis status.
• Neighbouring flocks have B. ovis or have unknown B. ovis status and/or sheep are able to stray
between properties.
• Presence of feral sheep in the area.
• Buy/lease/borrow/share/lend rams from or to non-accredited-free sources.
• Buy winter ram lambs (assume that any ram at or after puberty can potentially be infected).

Low-risk flocks – ALL of the following

• Known to be free of B. ovis.
• Regularly inspected.
• Neighbours free from B. ovis and regularly inspected.
• Only ever buy/lease/borrow/share/loan rams from or to accredited-free sources, or quarantine
rams and have them tested after they come onto the property.
• Rams contained in a well-fenced central area of the farm when not in use.
• Excellent boundary fences and general biosecurity.

Hence in commercial flocks vets should decide on a farm-by-farm basis, based on risk assessment, whether
additional testing for B. ovis (over and above scrotal palpation) is warranted.

Accredited-free flocks
For accredited-free flocks risk assessment is also important. It should be recognised that the B. ovis scheme
guidelines are just that – guidelines. Veterinarians are advised to follow the guidelines as a minimum but in high-
risk situations they can, at their discretion, be more stringent. For example the scheme guidelines state that for
commercial sire rams only 20 rams (or the whole flock, whichever is the lesser) need be blood sampled. However
for high-risk flocks (e.g. if neighbours have uncertain B. ovis status and sheep occasionally stray between the
properties) it might be prudent to consider increasing the number of commercial rams that are blood sampled
or even testing all of them.

Dealing with B. ovis cases or breakdowns

Step 1: Determine if infection is actually present

Assuming infection is suspected on the basis of serological reactors, confirmation of infection is based on one
or a combination of:
• flock history and risk of introduction.
• number of rams that are serologically positive.
• number of rams with epididymitis and how this relates to serology.
• use of the GD test – see below.
• potentially semen culture and/or histopathology – see above.

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Assuming infection is confirmed:

Step 2: Identify the likely source(s) of infection

This will help determine how long the infection is likely to have been in the flock and which control policy may
be most appropriate. It also means steps can be put in place to minimise the chances of re-infection.

Step 3: Decide upon a control strategy in conjunction with your client

This will usually be based on the client’s motivation, within-flock prevalence, value of the rams, farm set-up and
management factors. The options are:
• Do nothing, or merely palpate the rams each year and cull those with epididymitis. This option
should only be considered if the client refuses to control the disease. The main cost of B. ovis
is ram wastage but a reduction in flock fertility may occur if the infection gets well entrenched.
Often this results in the farmer using a greater ram:ewe ratio which increases the replacement
cost of rams. Long-term, eradication of the disease is highly likely to be cost-effective. Infected
flocks also pose a risk to neighbours.
• 2-flock system – the infected rams are kept separate from newly purchased non-infected rams
and each group is mated to a completely separate mob of ewes. Over time the infected group
will be culled as they age. This option requires long-term excellent management and attention to
detail – any mixing of rams or ewes from the two mobs during mating will compromise it.
• Cull all rams and start again – in chronically infected flocks with a high prevalence of infection
this may be the most straightforward option
• Test and cull – depending on the chronicity and prevalence of infection, this is generally the
preferred option as it should result in a disease-free flock whilst still salvaging some rams. After
the initial diagnosis a minimum of two flock tests and more usually 3-4 will be required. On rare
occasions five or more rounds of testing may be required. As a generalisation test and cull in
chronically infected flocks is likely to be more challenging than in recently infected flocks.

Test and cull

Which test(s) to use?

Chronically infected flocks (likely infected for a year or more)

In these cases it is recommended to use the CFT and ELISA tests in parallel right from the initial test. This
is because some rams that have been infected for a long period of time (probably years) seem to have low
fluctuating antibody titres which vary between negative, suspicious or sometimes positive in either test (Webb
et al. 1980). Using the CFT and ELISA in parallel increases the likelihood of detecting these rams (West 2000).
The most cost-effective way of doing this is to use the CFT for all submitted samples and then request the ELISA
for all CFT-negative samples (or vice-versa). In some cases semen culture may also be of benefit to identify all
infected rams.

Recently infected flocks (likely infected for less than a year)

In these flocks it is unclear whether using the CFT and ELISA in parallel is advantageous. We would generally
expect recently-infected rams to have high antibody levels, provided they have had enough time since contracting
the disease to develop antibodies.

How frequently to re-test?

From a B. ovis infection study currently being undertaken at Massey it appears that infected rams start to develop
antibodies detectable by ELISA from 3-4 weeks after exposure. Detection of antibodies by CFT seems to take
slightly longer (4-5 weeks or more after exposure). However by four weeks after exposure one of the 15 rams
in this study was shedding B. ovis in semen, and by five weeks after exposure five of the 15 rams were shedding.
Thus having a re-test interval longer than four weeks will increase the risk that recently infected rams will have
started shedding B. ovis in semen and be infectious to others.

Further results from this infection study will follow at a later date but these preliminary results suggest that

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the optimal re-test interval is 3-4 weeks using the ELISA with or without the CFT. However interpretation of
suspicious ELISA results requires care as these sometimes occur in non-infected rams.

If re-testing after 3-4 weeks using the ELISA:

• If a ram tests positive then it should probably be regarded as infected and culled.
• If a ram tests suspicious then isolating that ram and re-testing in a few weeks time might be the
best option to avoid unnecessary culling.

Two clear flock tests are required before the flock can be considered free of B. ovis. Based on the B. ovis
accreditation guidelines these tests should be at least 60 days apart to ensure all rams that may seroconvert have
had time to do so.

Other considerations
• During a test and cull programme, splitting the ram flock into small mobs will reduce the extent
of transmission. Similarly if the test and cull process spans the mating period then splitting the
ewe flock into smaller groups for mating (ensuring rams are not mixed between mobs) will also
limit the extent of transmission. Rams mated to each mob need to be kept separate after mating
until they have been tested.
• Transmission of infection between rams is likely to be at its most rapid during mating.
• It is preferable if one vet takes responsibility for coordinating the test and cull programme.

Some challenges associated with B. ovis control

Differentiation between ‘true’ and ‘false’ positive serological reactors

In flocks where infection is not expected, a single or small number of positive serological reactors mean
differentiation is required as to whether these are truly infected or whether they are ‘false positives’. In these
cases it is recommended to request the GD test on positive reactors. The GD is considered to be highly specific
so if the sera test positive then they are highly likely to be truly infected. If they test negative in the GD then
they are most likely not infected, but because the sensitivity of the GD is only 91.7% a small number of truly
infected rams will test negative in the GD. In cases where doubt exists, semen culture can be undertaken and/
or euthanasia of the ram followed by histopathology and/or culture. It is essential that the ram(s) in question
are not culled until their true infection status is determined. Careful investigation into the risk of introduction
onto the property is also very important.

Protracted test and cull programmes

On occasion test and cull programmes, particularly in chronically-infected flocks, can become protracted and
frustrating. The most common reasons for this are likely to be:
• Chronically infected rams with low fluctuating antibody titres. Such rams sometimes go
undetected for some time and remain a source of infection within the flock. As described above
use of the ELISA and CFT in parallel, and potentially the use of semen culture, will help in
identifying these rams.
• Failure of the farmer to achieve a clean muster of rams at each testing period, or rogue infected
rams straying into the flock and re-infecting it. It is important that all rams are permanently
identified and a list is kept to ensure all rams are present at each test, that extra rams do not
randomly appear, and that the infected rams from the previous testing period have indeed been
culled. For a test and cull programme to be worthwhile rogue rams and straying sheep must be
prevented.
• The time period between re-testing is too long (see above).

Summary
• Brucella ovis is still an important disease throughout New Zealand.
• Scrotal palpation for detection of epididymitis is a useful and cheap flock-screening test but
absence of rams with epididymitis does not guarantee a flock is free from B. ovis.
• In non-accredited flocks vets should decide on a farm-by-farm basis, based on risk assessment,

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 Brucella ovis – an update on control

whether additional testing for B. ovis is warranted.

• In accredited free flocks the accreditation scheme guidelines should be applied but where a flock
is considered medium to high risk then more stringent testing may be appropriate.
• When dealing with a potential B. ovis case firstly confirm the infection is present, identify the
source of infection and decide upon an appropriate control strategy in conjunction with the
client and possibly neighbouring flocks
• The approach to test and cull may vary depending on whether the flock is chronically or recently
infected.
• During test and cull the optimal re-test interval is likely to be about 3-4 weeks using the ELISA
with or without the CFT, although ‘suspicious’ ELISA results should be interpreted with caution.

Help & support

Dealing with B. ovis cases can sometimes be difficult and frustrating. I am happy to provide advice and support
– email a.l.ridler@massey.ac.nz or phone 06 3569099.

Acknowledgements
Thanks to Gail Ross, Gribbles Veterinary, for compiling and sharing the data on B. ovis cases, and to Dave West
for helpful comments on this manuscript.

References
Bulgin MS. Brucella ovis epizootic in virgin ram lambs.
Journal of the American Veterinary Medical Association
196, 1120-1122, 1990
Hughes KL, Claxton PK. Brucella ovis infection. 1. An
evaluation of microbiological, serological and clinical
methods of diagnosis in the ram. Australian Veterinary
Journal 44, 41-47, 1968
Webb RF, Quinn CA, Cockram FA, Husband AJ.
Evaluation of procedures for the diagnosis of Brucella
ovis infection in rams. Australian Veterinary Journal 56,
172-175, 1980
West DM. Brucella ovis control: Dealing with problem
flocks. Proceedings of the 30th Seminar of the Society of
Sheep and Beef Cattle Veterinarians NZVA, 51-60, 2000

Kennedy PC, Frazier LM, McGowan B. Epididymitis in

rams: pathology and bacteriology. Cornell Veterinarian
46, 303-319, 1956

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