Rev. sci. tech. Off. int. Epiz., 1993,12 (4), 1135-1146

Risk assessment on the importation of milk and milk products (excluding cheese) from

countries not free from foot and mouth disease N.H. HENG and D.W. WILSON *

Summary: The authors discuss the risk assessment conducted by the Australian Quarantine and Inspection Service (AQIS) on the importation of milk and milk products (excluding cheese) from countries not free from foot and mouth disease (FMD). This assessment was undertaken in response to requests from countries wishing to export dairy products for sale on the Australian market.

AQIS conducted a public consultation on the proposal, in line with Australian Government policy on transparency and accountability in the quarantine decision-making process.

The authors examine the procedures involved in the investigation of the likely presence of FMD virus in milk of vaccinated and non-vaccinated cows, and of the heat treatment parameters effective in the inactivation of the virus. The data provide a useful aid in the assessment of the risk factors associated with the importation of milk and milk products, and in the development of quarantine conditions for importation.

KEYWORDS: Foot and mouth disease virus - Heat treatment - Milk - Milk products - Public consultation - Risk assessment - Risk factors.

BACKGROUND

Foot and mouth disease (FMD) is of significant quarantine concern to Australia, as this is a major exotic disease which has the potent ia l to devastate the Aust ra l ian livestock industry. The disease has not occurred in Australia since 1872.

Current Australian quarantine legislation permits the importation of dairy products other than cheese and lactose only from countries considered to be officially free from FMD. These countr ies include Canada , Denmark , Finland, Japan, New Zealand, Norway, the Republic of Ireland, Sweden, the United Kingdom and the United States of America.

In response to requests to export dairy products to the Australian market from the governments of several countries which are not considered to be free from F M D , the Australian Government decided to review the relevant quarantine conditions. The Australian Quarantine and Inspection Service (AQIS) conducted a risk assessment on the importation of dairy products (excluding cheese) from countries not free from FMD.

* The Australian Quarantine and Inspection Service, G.P.O. Box 858, Canberra, ACT 2601, Australia.

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A literature search was conducted on the epidemiology of the disease in vaccinated and non-vaccinated herds, particularly in relation to virus secretion in milk. As the role of milk and milk products in the spread of the disease is well established, information on the heat inactivation of FMD virus (FMDV) was sought, in order to establish conditions for the safe importation of dairy products.

Although FMDV was the main object of the risk assessment, other important viruses and bacter ia excreted in the milk of cows - including bovine virus d iar rhoea virus (BVDV), enzootic bovine leukosis virus (EBLV), Mycobacterium tuberculosis (TB), Listeria spp. and Brucella abortus - were also considered in the assessment. While a review of the scientific l i terature revealed that F M D V is not inactivated in the conventional pasteurisation of milk, other viruses and bacteria were found to be of negligible significance, as heat treatment in pasteurisation regimes has been shown to be effective in the inactivation of BVDV (2), EBLV (13), TB (3), Listeria spp. (9, 26) and B. abortus (21).

AQIS prepared and circulated the risk assessment discussion paper - for evaluation and comment - to the Austral ian States, industry, scientific organisations and all interested parties, including the governments of the countries seeking market access in Austral ia for their milk and milk products. The public consultative process is in conformity with Australian Government policy on transparency and accountability in the quarantine decision-making procedure. Relevant comments from respondents to the risk assessment process were incorporated in a position paper which was circulated, together with draft protocols for possible importation, for a further round of consultation.

ASSESSMENT OF DISEASE RISK

The four principal factors influencing the disease risk are as follows:

- zone/country factors (i.e. diseases present in the animal populat ion of the exporting zone/country)

- commodity factors (i.e. diseases present in the animal or product)

- risk reduction factors (i.e. methods of reducing risk by product treatment)

- probability of domestic exposure (i.e. risk of pathogens from the imported product reaching susceptible animal populations in the importing country).

The first three of these factors are discussed below.

FOOT AND MOUTH DISEASE CARRIER STATUS AND VIRUS EXCRETION

The FMD carrier status of cattle represents a major problem for disease control in situations where the disease is enzootic or where a vaccination policy is adopted. Studies have shown that both vaccinated and non-vaccinated cattle are likely to become carriers after exposure to F M D V infection. The virus was isolated from recovered, non-immunised cattle for several months after clinical infection (32,18). Vaccinated cattle exposed to the virus became carriers without showing clinical signs of the disease (32, 30 ,10 ,18 , 31,19) . Virus multiplication occurred primarily in the oropharyngeal

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mucosae (10). Observations conducted by Hedger (19) in a group of vaccinated cattle exposed to severe natural field challenge with FMDV, revealed that 85% became carriers without showing clinical symptoms. Virus-negative animals in the group were found to possess very high antibody titres, and it was suggested that vaccinated animals may possess sufficient immunity to resist clinical disease, as well as being carriers of infection. Partly-immunised cattle may exhibit clinical signs which could be overlooked in field conditions, as the symptoms may not be as severe as those seen in susceptible animals (27).

Several workers were unable to detect viraemia in immunised cattle exposed to FMDV infection (31,19, 27). McVicar and Sutmoller (27) also reported the absence of viraemia in partly-immunised cattle which exhibited clinical signs after challenge with the virus. Doubts were expressed over the possible excretion of FMDV in the milk of vaccinated cows, as Leeuw et al. (23) were unable to isolate F M D V from the milk of experimentally-infected, vaccinated cows. There is a high probability that the virus may be absent from milk in a well-immunised herd in which no outbreak of the disease has been reported for some years.

Some workers have also repor ted that spread of virus did not occur from carrier animals to susceptible in-contact animals (32,10,19). However, Hedger (18) reported evidence of transmission of the virus from a carrier animal to another animal within a herd in the field. Studies by Sellers et al. (29) on airborne dispersal of FMDV revealed that vaccinated and non-vaccinated cattle excreted virus in two phases - between 30 min and 22 h, and 2 and 7 days after exposure. The authors suggested that the first phase probably resulted from the mechanical shedding of the virus from the respiratory tract and the coat of animals, or from the clothing of people exposed in the experiment; shedding in the second phase was thought to result from the multiplication of the virus in the respiratory tract. Virus excretion was not detected after the first day in animals which had been vaccinated more than once. This finding suggested that higher immunity levels resulted in reduced or zero virus production. However, studies were not conducted on the infectivity of the virus for susceptible animals. Donaldson and Kitching (17) found that cattle vaccinated three weeks before challenge with FMDV did not transmit virus to susceptible in-contact animals. In cattle vaccinated two weeks before challenge, transmission of the virus to susceptible in-contact animals occurred but the infection was subclinical. Transmission and subsequent clinical disease occurred in susceptible in-contact animals which had been exposed to cattle challenged four or seven days after vaccination.

The above findings per ta ining to the spread of F M D V from carrier animals to susceptible in-contact animals support the conclusion by Anderson et al. (1) that in a well-immunised herd where reintroduction of the virus is prevented through livestock movement controls and other animal health measures, FMDV is unlikely to persist for a lengthy period in the population and, therefore, that elimination of the virus from the herd can be achieved.

FOOT AND MOUTH DISEASE VIRUS EXCRETION IN MILK

Li tera ture repor ts confirm the impor tan t role of milk in the spread of FMDV. Dawson (16) made references to early l i tera ture in which laboratory exper iments supported repor ts on the role of milk in the spread of F M D . Dawson p resen ted

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evidence on the involvement of milk and milk movement in the natural spread of the disease during the 1967-1968 outbreak in the Uni ted Kingdom. Studies by Burrows et al. (12) and Blackwell et al. (8) confirmed that F M D V replicated in the mammary gland.

The fact that F M D V is excreted in milk before the onset of clinical disease in infected cattle is of major epidemiological significance in the control of the disease. Burrows (11) had demonstrated virus excretion before the onset of clinical symptoms in experimental studies. Hedger and Dawson (20) demonstrated high titres of infective virus in milk, at least 33 h before the detection of clinical disease.

It is important to note that Leeuw et al. (23) were unable to demonstrate FMDV in milk from vaccinated cows infected by the intranasal route. Milk samples obtained from cows exposed to the virus between two and nine months after their last vaccination were found to be negative for FMDV when inoculated onto cell cultures, fed to susceptible pigs and calves or injected intramuscularly and/or intradermolingually into susceptible steers. The absence of viraemia in immunised cattle exposed to FMDV infection and in partly-immunised cattle which had exhibited clinical signs after challenge with the virus, support the findings of Leeuw et al.

CATEGORISATION OF COUNTRIES

As scientific evidence supports the contention that the probability of FMDV being present in milk from well-vaccinated herds would be low, the feasibility of importing milk products from countries with well-protected herds was assessed. Accordingly, countries were categorised according to disease status, disease control programmes and the ability to implement such programmes effectively.

AQIS categorised potential exporting countries as follows:

Category 1: FMD vaccination is prohibited, and no outbreaks of the disease have been reported in the past twelve months.

Category 2: Compulsory official control programmes are implemented and no outbreaks of FMD have been reported in the past two years. The veterinary authorities in these countries have demonst ra ted the ability to implement disease control programmes effectively and could be relied upon to diagnose and notify any disease occurrence rapidly.

Category 3: FMD outbreaks continue to occur, or the disease status is unclear. These countries have difficulty in the effective implementation of disease control programmes and in the early detection, diagnosis and notification of the disease.

Draft quarantine conditions were then developed to facilitate the safe importation of milk products from these countries, subject to a full public consultation.

FOOT AND MOUTH DISEASE VIRUS SURVIVAL IN MILK AND MILK PRODUCTS

The investigation of F M D V inactivation in milk has been conducted largely on samples obtained from susceptible cows inoculated by the in t ramammary and

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intravenous routes, although Blackwell et al. (7) reported similar results for susceptible cows infected by oral exposure.

Several workers have shown that current t empera tu re / t ime regimes used for pasteurisation (either 60-65.5°C for at least 30 min, or high-temperature, short-time [HTST] treatment at 71-72°C for at least 15 sec) are inadequate to inactivate F M D V reliably in milk.

Earlier work, such as that by Sellers (28), used relatively insensitive cell-culture techniques for residual virus detection, and added virus to milk instead of using milk from infected animals (Table I). Hyde et al. (22) and Leeuw et al. (25) demonstrated that virus in milk from infected cows survived longer than virus added to milk from healthy cows, due to the protection afforded by cells, proteins and fat in the milk. These authors also showed that susceptible cattle provided a more sensitive means for the detection of residual viral activity than cell cultures.

TABLE I

Results of inactivation of foot and mouth disease virus (FMDV) (28)

Time taken to inactivate FMDV to a Temperature survival of 0.00001%

at pH 6.7 at pH 7.6

56°C 6 min 30 min 63°C 1 min 2 min 72°C 17 sec 55 sec 80°C <5 sec <5 sec 85°C <5 sec <5 sec

pH (at 4°C) Time taken for inactivation

2 1 min 4 2 min 5.5 30 min 5.8 18 h

11 2 h 12 2.5 min 13 2.5 min

Note: traces of virus were detected after the times recorded

Hyde et al. (22) infected susceptible cows by the intravenous and int ramammary routes. Infected milk samples, heated at 72°C and 80°C for 15-17 sec, were found to be positive for F M D V by cell-culture techniques. Milk treated at the above temperatures and times and then heated at 65°C for 64.5 min to reduce the volume by 50% tested negative for F M D V in cell cultures. However, susceptible steers inoculated with the evaporated milk samples developed clinical disease (Table II).

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TABLE II

Results of inactivation of foot and mouth disease virus (FMDV) in milk (22)

Sample Treatment Virus survival

Milk obtained 24 h 72°C for 15-17 sec Detected in cell cultures post-inoculation 80°C for 15-17 sec Detected in cell cultures

Milk pasteurised at 65°C for 64.5 min Not detected in cell cultures 72°C and 80°C (evaporated to 50% Detected in cattle inoculation for 15-17 sec of original volume) tests

Similar results (Table III) were obtained by Blackwell and Hyde (6), who found that FMDV survived pasteurisation at 72°C for 15 sec in skimmed milk, cream and pelleted cellular components. FMDV survived in whole milk heated at 72°C for 5 min and 85°C for 15 sec, skimmed milk heated at 72°C for 2 min and 85°C for 15 sec, and in cream

TABLE III

Results of heat treatment on components of milk from cows infected with foot and mouth disease virus (FMDV)

(6)

Cattle inoculation Sample Heat treatment for FMDV

Whole milk

Skimmed milk

Cream

Pelleted cellular debris

Whole milk (heated at 72°C for 5 min)

Skimmed milk (heated at 72°C for 15 sec)

Skimmed milk (heated at 72°C for 30 sec)

72°C for 5 min 85°C for 15 sec

72°C for 2 min 85°C for 15 sec

93°C for 15 sec

72°C for 15 sec

65°C for 1 h maximum (evaporated to 50% of original volume)

as above

as above

+ positive - negative

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heated at 93°C for 15 sec. FMDV survived in whole milk which was heated at 72°C for 5 min and then evaporated. The virus survived in skimmed milk after this was heated at 72°C for 15 sec, but not when it was heated at 72°C for 30 sec and then evaporated.

Most of the heated samples were found to be negative for F M D V by cell-culture assay but were positive in cattle inoculation tests. These findings reinforced the report by Hyde et al. (22) that inoculation into susceptible cattle was a more sensitive method for the detection of residual virus.

The results support the findings of Hyde et al. (22) that conventional pasteurisation is insufficient to inactivate FMDV in milk and milk components.

Blackwell (4) showed that pasteurisation of cream at 93°C for 16 sec, followed by processing to butter and storage for up to 45 days at 4°C at a pH which fell from 5.9 to 5.4, did not completely inactivate the virus. FMDV was detected in cattle inoculation tests.

Leeuw et al. (25) also found that cell-culture assay was less sensitive than cattle inoculation in the detection of residual virus. Using cell-culture assay, F M D V was not detected in skimmed milk heated at 72°C for 30 sec. However, the same sample heated at 80°C for 30 sec was found to be positive for virus when inoculated into cattle.

Cunliffe and Blackwell (14) found that F M D V survived in casein prepared from skimmed milk pasteurised at 72°C for 15 sec. Casein had been maintained at p H 4.5-4.7 for 2-3 h during processing. F M D V was detec ted by the in t radermol ingual and intramuscular inoculation of cattle.

FMDV was not detected by either cell-culture assay or cattle inoculation in whey constituents manufactured from virus-positive milk (5).

The effectiveness of ultra-high temperature (UHT) processes in virus inactivation was examined by Cunliffe et al. (15), who found that heating milk at 138°C for 2-3 sec did not inactivate the virus completely, while heating at 148°C for 2-3 sec was effective (Table IV). Intradermolingual and intramuscular inoculation of susceptible steers was conducted to detect residual virus in the treated milk samples.

Walker et al. (33) confirmed that heating infected milk at 148°C for 2.5 sec could effectively inactivate F M D V and also demons t ra ted that more than 20 min were required to inactivate virus at 100°C. The data presented by Walker et al. were obtained from a collaborative project conducted at the Plum Island Animal Disease Centre (PIADC) in the United States of America and the Central Veterinary Institute (CVI) in the Nether lands . Da ta on the exper iments conducted at the CVI have also been reported separately by Leeuw and Van Bekkum (24). Milk was obtained from susceptible cows infected by the intramammary and intravenous routes. The CVI used whole and skimmed milk, whereas the P I A D C used only whole milk. However, no differences were detected in the thermal inactivation of F M D V in ei ther whole or skimmed milk. All assays for infectivity following heat t reatment were conducted in susceptible cattle which were inoculated by the intradermolingual and intramuscular routes. The results are presented in Table V.

DISCUSSION

The li terature review confirms that conventional pasteurisation of milk and milk products is inadequate for the reliable inactivation of FMDV.

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TABLE IV

Results of ultra-high temperature treatments on milk from cows infected with foot and mouth disease virus (FMDV)

(15)

Temperature Time Cattle inoculation

for FMDV

102°C 2-3 sec + 123°C 2-3 sec + 130°C 2-3 sec + 138°C 2-3 sec + 148°C 2-3 sec -

+ positive - negative

TABLE V

Results of inactivation of foot and mouth disease virus in milk heated at various temperatures and times

(33) Results of virus assay Temperature Time in susceptible cattle

80°C 30 sec + 85°C 30 sec + 90°C 3 min +

100°C 9 min + 100°C 27 min -110°C 30 sec + 110°C 3 min -120°C 30 sec -130°C 10 sec + 135°C 17 sec -138°C 2.5 sec + 138°C 5 sec -148°C 3 sec —

+ positive - negative

Most of the work reported was conducted in milk obtained from susceptible cows infected by intravenous and intramammary inoculation of the virus, although reports were also made on milk from cows infected by contact exposure (7) . Reports on virus excretion in milk (16 ,19) in the 1967-1968 outbreak in the United Kingdom pertained to susceptible cattle.

Leeuw et al. (23) failed to detect FMDV in milk from vaccinated cows up to 19 days after intranasal challenge, although virus was detected in oropharyngeal fluids during

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this period. F M D V was not detected when the milk samples were inoculated into cell cultures, fed to pigs and calves, or injected intramuscularly and/or intradermolingually into susceptible steers. The contention of Leeuw et al. (23) is that infected, vaccinated cattle are unlikely to excrete virus in milk and that , even if excretion did occur, the concentration of virus would not approach that recorded for susceptible cows infected via the intramammary route. These authors further suggested that standard commercial heat t reatments , coupled with the activity of neutralising antibodies in pooled milk, should be adequate to inactivate the virus. Repor t s by some workers that viraemia could not be detected in either well-immunised or partly-immunised cattle exposed to FMDV, and the suggestion made by Anderson et al. (1) that total disease eradication in a well-immunised herd is possible, would support this argument.

The above argument provides a sound basis for the development of quaran t ine conditions to facilitate the safe importation of milk and milk products from Category 2 countries.

In the case of Category 3 countries, there is a high risk of FMDV contamination in milk and milk products. As commercial pasteurisation procedures have been shown to be inadequate to inactivate the virus, only milk products which have been subject to commercial sterilisation treatments in cans or retort pouches (or the equivalent) would be considered to present an acceptable risk.

*

ÉVALUATION DES RISQUES LIÉS À L'IMPORTATION DE LAIT ET DE PRODUITS LAITIERS (À L'EXCEPTION DES FROMAGES) EN PROVENANCE DE PAYS NON INDEMNES DE FIÈVRE APHTEUSE. - N.H. Heng et D.W. Wilson.

Résumé : Les auteurs étudient la méthode utilisée par les Services australiens de quarantaine et d'inspection (Australian Quarantine and Inspection Service : AQIS) pour évaluer les risques liés à l'importation de lait et de produits laitiers (à l'exclusion des fromages) en provenance de pays non indemnes de fièvre aphteuse. Cette évaluation a été entreprise pour répondre aux demandes de pays désireux d'exporter des produits laitiers en Australie.

L'AQIS a effectué une enquête publique en la matière, conformément à la réglementation australienne sur la transparence et la responsabilité dans le processus de prise de décision relatif à la quarantaine.

Les auteurs examinent les procédures appliquées à la recherche du virus de la fièvre aphteuse dans le lait de vaches vaccinées et non vaccinées, ainsi qu'à l'étude des paramètres de traitement thermique efficaces pour l'inactivation du virus. Les données ainsi obtenues constituent une aide précieuse pour l'évaluation des facteurs de risques liés à l'importation de lait et de produits laitiers, ainsi que pour l'élaboration d'une réglementation de la quarantaine applicable aux marchandises importées.

MOTS-CLÉS : Enquête publique - Evaluation des risques - Facteurs de risques - Lait - Produits laitiers - Traitement thermique - Virus de la fièvre aphteuse.

* * *

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EVALUACIÓN DE RIESGOS POR IMPORTACIÓN DE LECHE Y DE PRODUCTOS LECHEROS (EXCEPTO QUESOS) PROVENIENTES DE PAÍSES NO LIBRES DE FIEBRE AFTOSA. - N.H. Heng y D.W. Wilson.

Resumen: Los autores estudian el método utilizado por el Servicio australiano de cuarentena e inspección (Australian Quarantine and Inspection Service: AQIS) para evaluar los riesgos derivados de la importación de leche y productos lecheros (excepto quesos) provenientes de países no libres de fiebre aftosa. Este método fue establecido en respuesta a los pedidos de países deseosos de exportar sus productos lecheros a Australia.

El AQIS llevó a cabo una encuesta pública al respecto, conforme a la reglamentación australiana que se refiere a la transparencia y la responsabilidad en el proceso de tomar decisiones sobre cuarentenas.

Los autores examinan los procedimientos que se aplican en la investigación del virus de la fiebre aftosa en la leche de vacas vacunadas y no vacunadas y los que se aplican en el estudio de los parámetros de tratamiento térmico eficaces para la inactivación del virus. Los datos así obtenidos son de gran ayuda para la evaluación de los factores de riesgo en la importación de leche y de productos lecheros, así como también para la elaboración de una reglamentación de la cuarentena aplicable a las mercaderías importadas.

PALABRAS CLAVE: Encuesta pública - Evaluación de riesgos - Factores de riesgo - Leche - Productos lecheros - Tratamiento térmico - Virus de la fiebre aftosa.

* * *

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