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YERSINIAVAX® TECHNICAL MANUAL

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C0NTENTS

1. Introduction pg 3

2. Yersiniosis – The Disease pg 3

3. Yersiniavax – product information pg 6

4. Efficacy and Field trials pg 9

5. Application and Timing pg 13

6. Investigation of Suspected Lack of Efficacy pg 16

7. References pg 18

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1. INTRODUCTION The first recognised outbreak of Yersiniosis (Yersinia pseudotuberculosis)in New Zealand was in Timaru in 1978. This rapidly became the most common bacterial disease of young deer on farms. In the early 1980’s, a collaboration between scientists at Invermay, Wallaceville and the University of Otago led to a better understanding of the epidemiology of the disease, and the development of a killed Yersinia pseudotuberculosis vaccine. After extensive field testing (see “Efficacy and Field Trial Data”) the vaccine Yersiniavax was made available commercially in 1992. Yersiniavax was the first vaccine developed for deer, and remains the only commercially available vaccine specifically for farmed deer.

2. YERSINIOSIS – THE DISEASE Yersiniosis (Y. pseudotuberculosis) is the leading cause of death in weaned deer. A 20011 survey demonstrated the most common causes of mortality in deer herds and included wasting in hinds, malignant catarrhal fever in stags and Yersiniosis in weaners (Table 1: Average Mortality by season for different classes of deer, Audige et. al.). Mortality is highest in weaners, with an average of 5 weaners per 100 dying, during autumn and winter.

Season Class of Deer

Autumn Winter Spring Summer Annual

3-15 months

2.41% 2.62% 0.42% 0.14% 5.87%

Females >15 months

0.23% 0.67% 0.58% 0.32% 1.77%

Males >15 months

0.38% 0.83% 0.82% 0.57% 2.60%

Table 1: Average Mortality by season for different classes of deer, Audige et. al.1

Most fawns are exposed to Y. pseudotuberculosis. Bacteria are shed in the faeces of carrier

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animals (including birds, rodents, rabbits, cattle, sheep, pigs, goats and deer). The bacteria survive well in soil, water and pasture, especially during the winter, and infect fawns when they suckle, drink or graze faecally contaminated feeds. Stress and exposure to large numbers of bacteria cause disease. Stressors can include (but not limited too):

• Weaning • Poor nutrition • Sudden changes in feed • Mixing of mobs • Cold wet weather • Yarding or transport • Heavy parasite burdens.

The digestive system of stressed fawns slows down, allowing Y. pseudotuberculosis to multiply enormously in their gut, leading to massive bacterial shedding and toxaemia. The first sign of disease is blood-tinged, watery, smelly diarrhoea or sudden death. Bacterial toxins damage the intestines, leading to rapid fluid loss, bleeding, dehydration and frequently death. Diseased animals are normally 4-8 months old, separate from the mob, and initially have green watery distinctive malodorous diarrhoea that turns dark or bloody. As diseased animals stand back and diarrhoea may not be obvious, yarding and inspection for subtle signs, such as dirty watery staining of the hocks, is necessary to detect all cases. If treated early (e.g. antibiotics and fluids) deer often recover. Mortality rates in non-vaccinated deer can be as high as 30% of a mob2. Yersinia bacteria have what are known as Yersinia Outer Proteins, or YOP’s. These resist phagocytosis by injection of YOP effectors into macrophages. This causes paralysis of the actin cytoskeleton of neutrophils and macrophages and allows for further replication of bacteria. This is illustrated in Figure 1, Inhibition of phagocytosis through YOP's.

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Figure 1: Inhibition of phagocytosis through YOP's

(adapted from Nature Immunology 8, 2007, Nature Publishing Group)

There are three main groups of Yersinia bacteria:

1. Y. pestis which was responsible for the Bubonic plague (which killed 1/3 of the population of Europe between 1347 and 1353).

2. Y. enterocolitica which is more often a cause of enteritis in humans and cattle. 3. Y. pseudotuberculosis, the focus of this document.

Y. pseudotuberculosis serotypes are based on somatic and flagellar antigens and classified as Ia, Ib, IIa, IIb, III, IVa, IVb, and V. Only I, II and III are pathogenic and cause disease in animals and humans in New Zealand. The most recent survey on serotypes was carried out in 1979-823 and the results are shown in Table 2, Typing of Yersinia, 1979-82. Interestingly the majority of cases in deer in Otago/Southland were Y. pseudotuberculosis Type I during this survey, whereas nationwide it was Y. pseudotuberculosis Type III. Recently, in 2012, a large outbreak in North Canterbury was also confirmed as Y. pseudotuberculosis Type III4.

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Species Number examined

Y. pstb I Y. pstb II Y. pstb III

Cattle 56 7% 1% 92% Sheep 8 13% - 87% Deer 117 28% 8% 64% Goats 13 15% - 85%

Table 2: Typing of Yersinia pseudotuberculosis, 1979-823.

3.YERSINIAVAX

3.1 PRESENTATION Liquid vaccine. Multi-strain (I, II, III) Yersinia pseudotuberculosis (‘Y. pstb’). Formalin inactivated, containing DEAE Dextran as adjuvant, and 0.01% w/v thiomersal as a preservative.

3.2 USES For the immunisation of susceptible deer against losses due to the effects of infection by Y. pseudotuberculosis, namely diarrhoea, dehydration and death.

3.3 AGE TO VACCINATE

Deer most at risk are young stock in their first winter. Hence, usually it is weaners that are targeted by a vaccination programme (see later). It is recommended vaccination does not start before 12 weeks of age.

3.4 DOSE Two doses, each of 2mLs, are required. The second should be administered 3-6 weeks after the first, this course should be completed before the risk period. A single dose does not provide adequate protection. See Section 5: “Application and timing” for further information.

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3.5 ADMINISTRATION • Shake vaccine well before use. • Inject subcutaneously in the anterior half of the neck, not intramuscular. • Do not administer on the same side of neck as Tb tests as a scar could confuse the

Tb reading. • Change needle frequently, at least every 20 animals. • Ideal needles to use are 18G 3/8” or 18G 1/4” to ensure good subcutaneous

technique.

3.6 CONTRA-INDICATIONS • Do not vaccinate wet animals. • Do not inject through dirty areas of the skin. • Administer only to healthy animals.

3.7 ADVERSE EFFECTS/SAFETY

An injection site reaction may occur following treatment with Yersiniavax vaccine. This can be a hard swelling, a small abscess or a plaque-like leathery thickening of the skin, but will usually disappear within 10-12 weeks. It is important that animals are injected under the skin, and not into the muscle, to avoid the potential for a site reaction. Animals should not be submitted for slaughter prior to lesions subsiding. In a very small number of cases, a systemic effect due to the effect of the lipopolysaccharides, cell components or the DEAE adjuvant may occur leading to pyrexia and inappetance. These effects are transient, but if severe the animals should be treated with Non-Steroidal Anti-inflammatory Drugs (NSAID’s). Trials have revealed that in cases of an overdose (5mL vaccine per animal) a slight temperature rise was seen and a reduced appetite in some animals for 24-48 hours. There were also slightly larger injection site reactions, but these receded over the next few weeks.

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3.8 WITHHOLDING PERIOD There is no withholding period, however deer should not be submitted for slaughter prior to injection-site lesions subsiding. Injection sites too far down the neck of the deer may lead to carcass damage and good technique is essential.

3.9 WARNINGS/PRECAUTIONS

• Accidental self-injection can cause serious local reactions. If this does occur, seek medical attention at once. Infection by transfer of bacteria via a dirty needle may occur, and some people may react to the DEAE Dextran adjuvant. Contact the National Information Centre for Poisons and Hazardous Chemicals on 0800 POISON (0800 764 766).

• Yersiniavax should be stored between +2⁰C and +8⁰C. • Do not freeze. • Protect from heat and sunlight. • Use before expiry date on the label.

3.10 PARTIALLY USED PACKS

As long as the sterility is maintained and storage instructions adhered to, partially used packs may be stored for up to six weeks.

3.11 PACK SIZE

Each pack of Yersiniavax is sufficient for 50 doses and contains: • One 100mL vaxipack containing prepared vaccine. • Draw off tube for connection of vaxipack to vaccinating gun. • Instruction leaflet.

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4. EFFICACY AND FIELD TRIAL DATA

Efficacy challenge trials

In 1989 and 1990 two experimental trials were conducted using weaner red deer fawns that were 3-6 months old5,6 in which two different adjuvants, STM (oil) and DEAE Dextran, were compared.

Doses

In 1989 one 2mL subcutaneous dose of vaccine was given, whereas in 1990 two vaccinations were given.

Stress/challenge

In both years, a group of deer were put through a combination of normal farm stressors (yarding, weighing, fasting for 24 hours, transport in a truck for 3 hours, and unloading at strange deer yards), prior to their being challenged with an oral dose of live virulent Y. pseudotuberculosis organisms: 8 x 109 in 1989 and 3 x 109 in 1990. A control group in 1990 was also challenged with, Y. pseudotuberculosis but without prior stress.

There were a small number of non-challenged deer (vaccinated and unvaccinated) run in contact with the challenged deer to test the degree of natural challenge from the experimentally infected deer. None of these ‘in contact’ deer developed Yersiniosis. This contrasts with the high rate of Yersiniosis in the experimentally infected deer, indicating that the experimental challenge was a severe challenge. The severity of the experimental challenge is further indicated by the fact that 30% of the unstressed deer in the 1990 trial were also affected by the challenge.

Results

Significantly (p>0.05) fewer vaccinated animals were affected than similarly stress/challenged unvaccinated deer under these conditions of a heavy challenge. However, of those that were affected, a similar number of vaccinated and unvaccinated deer died.

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In the 1990 trial, DEAE Dextran vaccine performed as well as STM, however because it resulted in fewer injection-site reactions it was considered more acceptable.

Treatments Number of animals Clinical Yersiniosis seen

Yersiniavax (STM) – stressed 70 22 (31%)

Unvaccinated control – stressed 69 37 (54%) Table 3: 1989 Trial.

Treatments Number of animals Clinical Yersiniosis seen

Yersiniavax (STM) – stressed 30 8 (27%)

Yersiniavax (DEAE) – stressed 30 10 (33%)

Unvaccinated control – stressed 30 18 (60%)

Unvaccinated control – no stress 30 9 (30%) Table 4: 1990 Trial.

Field trials

In 1991 a large scale field trial was set up involving almost 5,000 red deer weaners. In order to increase the chances of a successful trial, farms were chosen that had a high probability of a Yersiniosis outbreak based on history and a group size of >150 fawns. The monitoring of the trial concentrated on investigating outbreaks and assessing how the vaccine performed.

Trial protocol

The aim of this trial was to test Yersiniavax on 4,958 deer fawns on 17 high risk farms. Twelve veterinarians assisted during the 6 month period March-Sept 1991.

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Two doses of DEAE Dextran adjuvanted vaccine (2mL by subcutaneous injection in the neck) were given in March and April 1991, the second dose 3-6 weeks after the first, to half of the weaners chosen at random. The remaining half of each mob was not vaccinated. Randomisation was achieved by bringing through small groups of weaners and alternately vaccinating or not vaccinating the individuals in each groups and identifying them. Farmers were left with recording sheets on which to record the ear tag numbers of any fawns that became sick, developed diarrhoea or died.

Results

Outbreaks occurred in 3 of the 17 trial mobs, 2 in the North Island and 1 in Canterbury.

Outbreak A (Farm #1) involved a total of 43 clinical cases of Yersiniosis, all of which died, with 33 deaths (22%) in unvaccinated calves, and 10 deaths (6.7%) in vaccinated calves, from a total of 299 calves, (P<0.001).

Outbreak B (Farm #2) involved a total of 98 cases of Yersiniosis, 88 cases (34%, 55 of which died) in 260 unvaccinated calves, compared with 10 cases (4%, 3 died) in 260 vaccinated calves, (P<0.001).

Outbreak C (Farm #3) involved 22 cases of Yersiniosis (14%) and 3 deaths in the unvaccinated group, and 13 cases (8%) and 3 deaths in the vaccinated group. This difference was not statistically significant (P<0.2), but the outbreak started less than 2 weeks after the first dose of vaccine was given (i.e. before the second dose was administered and protection could be expected) and no new cases occurred after the second dose of vaccine.

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Table 5: Results of a large 1991 scale field trial.

*significant difference between vaccinated and unvaccinated, (P<0.001)

# An outbreak occurred before the second dose of vaccine was given.

Farm No.

No. Vaccinated

No. unvaccinated

Vaccinated Unvaccinated Comments

No. Clinical disease

No. died No. clinical disease

No. died

1 150 149 10 10* (6.7%)

33 33* (22%)

Y. pseudotuberculosis isolated from 5 unvacc, 3 vacc

2 260 260 10* (4%)

3* (1%)

88* (34%)

55* (21%)

Y. pseudotuberculosis confirmed 4 cases

3 152 152 13 (9%)

3 # 22 (15%)

3 Cases occurred after first vacc

4 100 96 2 2 1 1 None confirmed

5 186 185 1 1 2 2 None confirmed

6 127 124 6 6 # 2 2 Cases occurred after first vacc

7 159 160 4 4 3 2 None confirmed

8 241 261 4 3 3 3 None confirmed

9 111 111 0 0 0 0 No cases

10 135 135 1 1 2 0 None confirmed

11 121 143 0 0 0 0 No cases

12 127 127 3 3 1 1 None confirmed

13 97 140 1 1 3 2 None confirmed

14 146 146 0 0 0 0 No cases

15 100 97 1 1 1 1 Y. pseudotuberculosis from both groups

16 100 100 1 1 1 1 None confirmed

17 106 109 0 0 0 0 No cases

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Conclusions

• 2 doses of vaccine 3-6 weeks apart give significant protection against heavy experimental and natural Y. pseudotuberculosis challenge.

• The field trials are likely to have been different from what would happen when all of a mob are vaccinated because:

a. The degree of challenge experienced by unvaccinated animals was decreased when an outbreak did occur.

b. Each infected animal drastically increases the amount of contamination in the environment and challenges peers in this group.

5. APPLICATION AND TIMING

Vaccination with Yersiniavax will provide significant protection against Yersiniosis in deer. The aim of vaccination is to prevent a serious epidemic by reducing the spread of disease through the mob, however will not protect every individual. Yersiniavax therefore enhances, rather than is a substitute for, good herd management.

Young deer are most at risk of the disease being rare in adults, suggesting that exposure during their first winter leads to strong immunity. Farmers should therefore aim to vaccinate weaners prior to their first winter.

Vaccination with Yersiniavax is only part of what should be done by farmers to reduce the risk of an outbreak of Yersiniosis. Reducing the effects of stressors (see Section 2) and weaning before the rut, when it is warmer and more feed is available, will help with preventing outbreaks. All general farm practices should be carried out with this in mind.

Deer should have completed their full course of Yersiniavax vaccination at least 7-10 days prior to the risk period. In New Zealand the disease is much more prevalent in late autumn and winter, as that is the time when onset may be precipitated by stressors, as well as the addition to the unavoidable stress imposed by weaning.

Ideally, young deer should be vaccinated as early as possible, however two further

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considerations are important.

1. Maternally Derived Antibodies (MDA) may interfere with vaccination if given too early which may reduce the weaners’ immunological response to the vaccine, thereby compromising the vaccines effectiveness. This effect is unlikely to persist beyond 10-12 weeks of age; hence the label stated vaccination from 3 months of age. For the majority of farms where weaners are born in November or December, vaccination from March onwards is best.

2. Vaccination ideally needs to fit in with the normal farming calendar to avoid excessive yarding and disturbing late-fawning hinds. Although early vaccination is desirable, the benefit of this must be weighed up with the potential stress caused by multiple yardings or movements of mobs.

From a farm management perspective, the best time for vaccination is a balance between the inconveniences or disadvantages of vaccinating early, and the risk that later vaccination will leave deer exposed without protection at a time of the greatest challenge. The management practice with the largest bearing on this is whether a farmer weans before or after the rut. It is important to also consider the stress of the weaning process itself, which may reduce the weaners’ response to vaccinations.

Timing of Vaccination

1. Both vaccinations completed at least 10 days before weaning:

This provides maximum protection over weaning and through the high risk period. Especially beneficial if weaners are sold or transported soon after weaning. However, this may be difficult due to late fawning hinds and many fawns being less than 10-12 weeks of age at the time of first (or even second) vaccination. As the majority of hinds conceive in the first 2 cycles of mating, this may mean that the majority of the fawns will be old enough at the time of vaccination. If weaning after the rut, an option is to coincide the 2nd injection with mating, for example when mating groups are yarded to change the stags.

If management priorities do not allow this, alternative timing can be considered, however it should be noted that altered timing of vaccination may affect the level of protection

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afforded.

2. First injection 3-6 weeks before weaning, the second injection at weaning.

This is probably the most common programme currently used. Some protection against Yersiniosis triggered by weaning stressors, but the peak immune response is not complete until a minimum of 7-10 days after second injection, so doesn’t fully protect weaners at weaning. This programme reduces risk of disease associated with bad weather after weaning but does not protect weaners that are sold or transported at weaning time.

3. First injection at weaning, second injection 3-6 weeks later.

This is the least favourable option as this provides no protection against Yersiniosis triggered by weaning stress or poor weather until 7-10 days after the second injection. In some cases this means weaners are unprotected for a considerable time during the most risky period.

4. Use of a 3rd vaccine into the programme.

This involves giving all fawns (even those less than 12 weeks of age) their first vaccination 3-4 weeks before weaning, the second at weaning and the smallest (and youngest) weaners are given a third vaccination 3-4 weeks after weaning. This is to ensure that those who were too young to respond to the initial vaccination due to MDA (hence the weaning vaccination acted as a sensitiser not a booster) receive a full course, while still ensuring the majority of the mob are protected early. Preliminary results after three seasons show good results with little extra expense. These management options highlight the need for Yersiniavax vaccine to be used to augment, rather than substitute for, good management. It is too late to vaccinate a mob once an outbreak has started in that mob.

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6. INVESTIGATION OF SUSPECTED LACK OF EFFICACY

Every season we investigate a small number of instances in winter where Y. pseudotuberculosis is associated with deaths in deer vaccinated with Yersiniavax. These cases are individually investigated to determine the cause of death, and in each case a satisfactory reason for the deaths is usually determined.

When investigating cases, veterinarians should consider a number of factors:

Management factors

1. Did deer receive the full dose at least 7-10 days prior to the risk period? 2. Were 2 doses given, the second 3-6 weeks after the first? A single dose does not give

protection. 3. Was vaccination accompanied by bad weather, under feeding or high stress? Animal

response to vaccination is compromised by these conditions. 4. Was the vaccine stored and handled correctly? 5. Was the vaccine used before its expiry date? 6. Did the outbreak occur before or soon after the second dose? Maximally protective

antibody levels do not develop until 7-10 days after the second dose. 7. What age were the animals at first vaccinations, and could MDA have interfered with

the response to vaccination?

Animal related factors

1. Not every animal has the genetic capability to respond to every bacterial antigen. Some deer have a poor immune response.

2. Some deer appear to be genetically more susceptible to Y. pseudotuberculosis infection. In 1998 a farm in Pahiatua was investigated and there was some trends seen with 4 of the 100 progeny of one stag dying. This prompted an immunological study comparing the remaining progeny of this stag to others, and those results, albeit with small numbers, suggested an immunological basis for sire differences2.

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3. Some deer have their response compromised at the time of vaccination by stress (nutritional, weaning, transport, social etc.), or concurrent disease (lungworm, gastro-intestinal parasites, facial eczema etc.).

4. Some deer may have their overall immune response compromised at the time of disease challenge, by similar stressors and diseases as in (3) above.

5. Other diseases such as Malignant Catarrhal Fever (MCF) or infections with Salmonella, Y. enterocolitica and Campylobacter organisms are all also capable of causing death and are not protected against by Yersiniavax.

6. Deer may have been exposed to a new strain of Y. pseudotuberculosis. This is possible but unlikely and recent work to investigate this did show this was not the case4. Yersiniavax incorporates all three known serotypes in NZ (I,II and III)

7. The antibody levels produced in response to the vaccine may wane more quickly in some animals than others.

8. Often the size of the challenge dose of Y. pseudotuberculosis organisms is high enough to overwhelm the protective immune response in some animals.

No vaccine is 100% protective. The aim of mass vaccination in a group of animals is to produce ‘flock’ or ‘herd’ immunity. This will prevent a serious epidemic of disease from spreading through a herd, even if some individual animals are not protected. Use of the Yersiniavax vaccine does not remove the need to monitor the health of weaners daily over the winter, and to treat sick animals promptly.

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7. REFERENCES

1. Audi’ge et al; Disease and mortality on red deer farms in New Zealand. Vet Record 148: 334-340(2001).

2. Wilson, P.R; Mackintosh, C.G; Griffin, J.F.T; Yersiniosis: Resistance, susceptibility and vaccination. Proceedings of the Deer Branch of New Zealand Veterinary Association, 16:133-142 (1999).

3. Henderson, T.G; Yersiniosis in Deer from the Otago-Southland region of New Zealand. New Zealand Vet Journal 31: 221-4(1983).

4. MSD, Data on file.

5. Mackintosh, C.G; Buddle, B.M; Griffin, J.F.T; Cross, J.P; Yersiniosis vaccine update. Proceedings of the Deer Branch of New Zealand Veterinary Association, 8, (1991).

6. Mackintosh, C.G; Buddle, B.M; Griffin, J.F.T; Yersiniosis Vaccine Research. Proceedings of the Deer Branch of New Zealand Veterinary Association, 7, (1990).

Further reading of relevance:

Hodges, R.T al; Serotypes of Yersinia pseudotuberculosis recovered from domestic livestock. New Zealand Vet Journal 32:11-13(1984).

Mackintosh, C.G; Vaccine against yersiniosis in deer. Surveillance 20(2), 25(1993).

Obwolo, M.J; A review of yersiniosis. The Veterinary Bulletin 46 (3) 167-171(1976).

Mackintosh, C.G; Henderson, T.G; The epidemiology of yersiniosis in deer. Proceedings of the Deer Branch of New Zealand Veterinary Association, 1: 34-42 (1984).

Mackintosh, C.G; Buddle, B.M; Griffin, J.F.T; Yersiniavax® efficacy under field conditions in weaner red deer. Proceedings of the Deer Branch of New Zealand Veterinary Association, 9:107-110 (1992).

Mackintosh, C.G; Yersiniosis vaccination – timing it right. The Deer Farmer magazine, (1995).

AVAILABLE ONLY UNDER VETERINARY AUTHORISATION. ACVM No: A6151. ®Registered trademark. Schering-Plough Animal Health Ltd. Phone: 0800 800 543. www.msd-animal-health.co.nz . NZ/YSVX/0115/0002