Commission Delegated Regulation (EU) 2020/689Show full title

PART II U.K. DISEASE- SPECIFIC REQUIREMENTS FOR DISEASE-FREE STATUS OF AQUATIC ANIMALS

Part II covers the disease-specific requirements for disease-free status as regards the following listed diseases:

Viral haemorrhagic septicaemia (VHS)	Chapter 1
Infectious haematopoietic necrosis (IHN)	Chapter 1
Infection with HPR-deleted infectious salmon anaemia virus	Chapter 2
Infection with Marteilia refringens	Chapter 3
Infection with Bonamia exitiosa	Chapter 4
Infection with Bonamia ostreae	Chapter 5
Infection with white spot syndrome virus (WSSV)	Chapter 6

CHAPTER 1 U.K. Eradication, disease-free status and diagnostic methods for viral haemorrhagic septicaemia (VHS) and infectious hematopoietic necrosis (IHN)

Section 1 U.K. General requirements for health visits and sampling

Health visits and sampling for the surveillance referred to in point (b)(ii) of Article 3(2) must comply with the following requirements:

(a)

health visits and, where appropriate sampling, must be carried out during the period of the year when the water temperature is below 14 °C or when temperatures below 14 °C are not reached, samples must be taken at the lowest annual temperatures;

(b)

when targeted surveillance in wild populations is required due to the small number of aquaculture establishments in an eradication programme, the number and geographical distribution of sampling points must be determined to obtain a reasonable coverage of the Member State, zone or compartment. The sampling points must be representative of the different ecosystems where wild populations of susceptible species are located;

(c)

when establishments or wild populations are to be subject to health visits or sampled more than once per year, in accordance with Sections 2 to 4, the intervals between the health visits and between the collection of samples must be at least 4 months, or as long as possible, taking into account the temperature requirements provided for in point (a);

(d)

all production units, such as ponds, tanks and net cages, must be examined for the presence of dead, weak or abnormally behaving fish. Particular attention must be paid to the water outlet area where weak fish tend to accumulate because of the water current;

(e)

fish of listed species to be collected as samples must be selected as follows:

(i)

if rainbow trout are present, only fish of that species must be selected for sampling, except where other susceptible species are present which show typical signs of VHS or IHN; if rainbow trout are not present, the sample must be representative of all other susceptible species which are present;

(ii)

if weak, abnormally behaving or freshly dead but not decomposed fish are present, such fish must be selected; if more than one water source is utilised for fish production, fish representing all water sources must be included in the sample;

(iii)

the fish selected must include fish collected in such a way that all production units, such as net cages, tanks and ponds, of the establishment, as well as all year classes, are proportionally represented in the sample.

Section 2 U.K. Granting of the status free from VHS or free from IHN in Member States, zones and compartments of unknown health status

The status free from VHS or free from IHN may only be granted to a Member State, a zone or a compartment with an unknown health status with regard to VHS or IHN if:

(a)

all establishments and, when required, sampling points in wild populations selected in accordance with point (b) of Section 1, have been subject to one of the following scheme:

(i)

model A — 2-year scheme

The establishments or sampling points must have been subject to health visits and sampled for a minimum period of 2 consecutive years as laid down in Table 1.A.

During that 2-year period, the testing of all samples using the diagnostic methods set out in point 2 of section 5 must have produced negative results for VHS or IHN, and any suspicion of VHS or IHN must have been ruled out in accordance with the sampling and diagnostic methods set out in point 3 of Section 5;

(ii)

model B — 4-year scheme with reduced sample size

The establishments or sampling points must have been subject to health visits and sampled for a minimum period of 4 consecutive years as laid down in Table 1.B. During that 4-year period, the testing of all samples using the diagnostic methods set out in point 2 of Section 5 must have produced negative results for VHS or IHN and any suspicion of VHS or IHN must have been ruled out in accordance with the sampling and diagnostic methods set out in point 3 of Section 5;

(b)

if VHS or IHN have been detected during the surveillance referred to in point (a); before starting a new 2-year or 4-year scheme, relevant establishments in the Member State, zone or compartment must:

(i)

be subject to the minimum disease control measures laid down in Articles 58 to 65;

(ii)

be repopulated with fish from an establishment in a Member State, zone or compartment with status free from VHS or status free from IHN or from an establishment in a Member State, zone or compartment covered by an eradication programme for VHS or IHN.

Table 1.A

Scheme for Member States, zones and compartments for the 2-year control period referred to in point (a)(i) which precedes the achievement of status free from VHS and status free from IHN

a In the case of coastal zones or coastal compartments, the samples must be collected no sooner than 3 weeks after the transfer of the fish from fresh to saltwater.
b Ovarian or seminal fluid of broodstock shall be collected at the time of maturation, in connection with stripping.
Type of establishment	Number of health visits per year to each establishment	Number of samplings per year in each establishment	Number of fish in the samplea
			Number of growing fish	Number of broodstock fishb
(a) Establishments with broodstock	2	2	50 (first visit) 75 (second visit)	30 (first or second visit)
(b) Establishments with broodstock only	2	1	0	75 (first or second visit)
(c) Establishments without broodstock	2	2	75 (first AND second visit)	0
Maximum number of fish per pool: 10

Table 1.B

Scheme for Member States, zones or compartments using a reduced sample size for the 4-year control period referred to in point (a)(ii) which precedes the achievement of status free from VHS and status free from IHN

a In the case of coastal zones or coastal compartments, the samples must be collected no sooner than 3 weeks after the transfer of the fish from fresh to saltwater.
b Ovarian or seminal fluid of broodstock shall be collected at the time of maturation, in connection with stripping.
Type of establishment	Number of health visits per year to each establishment	Number of samplings per year in each establishment	Number of fish in the samplea
			Number of growing fish	Number of broodstock fishb
First 2 years
(a) Establishments with broodstock	2	1	30 (second visit)	0
(b) Establishments with broodstock only	2	1	0	30 (first or second visit)
(c) Establishments without broodstock	2	1	30 (first or second visit)	0
Last 2 years
(a) Establishments with broodstock	2	2	30 (first visit)	30 (second visit)
(b) Establishments with broodstock only	2	2		30 (first AND second visit)
(c) Establishments without broodstock	2	2	30 (first AND second visit)
Maximum number of fish per pool: 10

Section 3 U.K. Granting of the status free from VHS or free from IHN in Member States, zones and compartments known to be infected with either VHS or IHN

1. The status free from VHS or free from IHN may only be granted to a Member State, a zone or a compartment known to be infected with VHS or IHN, if all establishments keeping listed species within that Member State, zone or compartment have been subject to an eradication programme that complies with the following requirements: U.K.

(a)

the minimum control measures laid down in Articles 55 to 65 must have been effectively applied and a restricted zone of an appropriate size as provided for in point (c) of Article 58(1), where appropriate, divided into a protection zone and surveillance zone; must have been established in the vicinity of the establishment(s) declared infected with VHS or IHN, taking into account the requirements set out in point 2;

(b)

all establishments keeping listed species within the protection zone, or where a protection zone has not been established, the restricted zone, not infected with VHS or IHN must be subject to an investigation comprising at least the following elements:

(i)

the collection of samples for testing of 10 fish, when clinical signs or post-mortem lesions consistent with infection with VHS or IHN are observed or minimum 30 fish, when clinical signs or post-mortem lesions are not observed;

(ii)

in those establishments where the tests referred to in (i) have produced negative results; health visits must continue once per month during the period when the water temperature is below 14 °C, except when fish ponds, tanks, raceways or net cages are covered with ice, until the protection zone is withdrawn in accordance with point (c);

(c)

relevant establishments must be emptied in accordance with Article 62, cleaned and disinfected in accordance with Article 63 and fallowed in accordance with Article 64.

The duration of the fallowing period referred to in point (a) of Article 64(2) must be at least 6 weeks. When all establishments infected within the same protection zone, or where a protection zone has not been established, the restricted zone, are emptied, at least 3 weeks of synchronised fallowing must be carried out.

When fallowing of the infected establishments is carried out, the restricted zone or the protection zone, when it has been established, must be converted into a surveillance zone until the scheme set out in Section 2 is completed;

(d)

repopulation may only take place when all infected establishments have been emptied, cleaned, disinfected and fallowed in accordance with point (c);

(e)

all establishments other than those referred to in point (f) which keep listed species within the Member State, zone or compartment covered by the eradication programme and when surveillance in wild populations is required, all sampling points selected in accordance with point (b) of Section 1, must subsequently be subject to the scheme laid down in Section 2;

(f)

an individual establishment which keeps listed species and which has a health status which is independent of the health status of the surrounding waters is not required to comply with the scheme laid down in Section 2 following a disease outbreak, provided the establishment complies with the requirements set out in paragraph 3 of Article 80 and is repopulated with fish sourced from Member States, zones or compartments with status free from VHS or status free from IHN.

2. The restricted zone must have been defined on a case-by-case basis and: U.K.

(a)

it must take into account factors influencing the risks for the spread of VHS or IHN to kept and wild fish, such as:

(i)

the number, rate and distribution of the mortalities of fish on the establishment infected with VHS or IHN, or in other aquaculture establishments;

(ii)

the distance to and density of neighbouring establishments;

(iii)

the proximity to slaughterhouses;

(iv)

contact establishments;

(v)

species present at the establishments;

(vi)

the farming practices applied in the infected establishments and the neighbouring establishments;

(vii)

the hydrodynamic conditions; and

(viii)

other factors of epidemiological significance identified;

(b)

the geographical demarcation in coastal areas must comply with the following minimum requirements:

(i)

the protection zone must consist of an area included in a circle with a radius of at least one tidal excursion or at least 5 km, whichever is larger, centred on the establishment infected with VHS or IHN, or an equivalent area determined according to appropriate hydrodynamic or epidemiological data; and

(ii)

the surveillance zone must consist of an area surrounding the protection zone, of overlapping tidal excursion zones; or an area surrounding the protection zone and included in a circle of radius 10km from the centre of the protection zone; or an equivalent area determined according to appropriate hydrodynamic or epidemiological data;

or

(iii)

where separate protection and surveillance zones are not established, the restricted zone must consist of an area comprising both the protection zone and the surveillance zone;

(c)

the geographical demarcation in inland areas must comprise the entire water catchment area in which the establishment infected with VHS or IHN is located. The competent authority may limit the extent of the restricted zone to parts of the water catchment area, provided this limitation does not compromise the disease control measures with respect to VHS or IHN.

Section 4 U.K. Maintenance of status free from VHS and status free from IHN

1. When targeted surveillance is required in order to maintain the status free from VHS or free from IHN of a Member State, a zone or a compartment, in accordance with Article 81, all establishments keeping listed species within the Member State, zone or compartment concerned must be subject to health visits and fish must be sampled in accordance with Table 1.C, taking into account the risk level of the establishment for the contraction of VHS or IHN. U.K.

2. When determining the frequency of health visits required to maintain the status free from VHS or the status free from IHN of compartments, where the health status regarding VHS or IHN is dependent on the health status of the aquatic animal populations in surrounding natural waters, the risk for the contraction of VHS or IHN must be regarded as high. U.K.

3. Disease-free status must only be maintained as long as all samples tested, using the diagnostic methods set out in point 2 of Section 5, have produced negative results for VHS or IHN and any suspicion of VHS or IHN has been ruled out in accordance with the diagnostic methods set out in point 3 of Section 5. U.K.

Table 1.C
Scheme for Member States, zones or compartments to maintain status free from VHS or status free from IHN
a Risk level assigned to the establishment by the competent authority as set out in Chapter 2 of Part I other than in the case of dependent compartments where all establishments are deemed to be high risk.
b One sample to be taken during every health visit.
c In the case of coastal zones or coastal compartments, the samples must be collected no sooner than 3 weeks after the transfer of the fish from fresh to saltwater.
Risk levela	Number of health visits per year to each establishment	Number of fish in the sampleb,c
High	1 every year	30
Medium	1 every 2 years	30
Low	1 every 3 years	30
Maximum number of fish per pool: 10

Section 5 U.K. Diagnostic and sampling methods

1. The organs or tissue material to be sampled and examined must be the spleen, the anterior kidney, and either heart or encephalon. When sampling broodstock, ovarian or seminal fluid may also be examined. U.K.

In case of small fry, whole fish may be sampled.

Samples from a maximum of 10 fish may be pooled.

2. The diagnostic method for the granting or the maintenance status free from VHS or status free from IHN in accordance with Sections 2 to 4 must be: U.K.

(a)

virus isolation in cell culture with subsequent identification of the virus using ELISA, indirect fluorescent antibody test (IFAT), virus neutralisation test or virus genome detection; or

(b)

Reverse Transcription quantitative PCR (RT-qPCR) detection.

The detailed procedures to carry out these diagnostic methods must be those approved by the EURL for fish diseases.

3. When a suspicion of VHS or IHN is required to be confirmed or ruled out in accordance with Article 55, the following health visit, sampling and testing procedures must comply with the following requirements: U.K.

(a)

the suspected establishment must be subject to at least one health visit and one sampling of 10 fish, when clinical signs or post-mortem lesions consistent with infection with VHS or IHN are observed or minimum 30 fish, when clinical signs or post-mortem lesions are not observed. Samples shall be tested using one or more of the diagnostic methods set out in points 2(a) and 2(b) in accordance with the detailed diagnostic methods and procedures approved by the EURL for fish diseases;

(b)

the presence of VHS must be considered as confirmed, if one or more of those diagnostic methods are positive for VHSV. The presence of IHN must be considered as confirmed, if one or more of those diagnostic methods are positive for IHNV. The confirmation of the first case of VHS or IHN in Member States, zones or compartments previously not infected must be based on conventional virus isolation in cell culture with subsequent immunochemical or molecular identification or with genome detection including confirmation by sequencing of the amplification (RT-PCR) product;

(c)

Suspicion of VHS or IHN may be ruled out, if cell cultivation or RT-qPCR tests reveal no further evidence of the presence of VHSV or IHNV.

CHAPTER 2 U.K. Eradication, disease-free status and diagnostic methods for infection with HPR-deleted infectious salmon anaemia virus (HPR-deleted ISAV)

Section 1 U.K. General requirements for health visits and sampling

Health visits and sampling for the surveillance referred to in point (b)(ii) of Article 3(2) must comply with the following requirements:

(a)

when health visits and sampling of establishments must be carried out more than once per year in accordance with Sections 2 to 4, the intervals between the health visits or collection of samples shall be as long as possible;

(b)

when targeted surveillance in wild populations is required due to the low number of aquaculture establishments in the eradication programme, the number and geographical distribution of sampling points must be determined to obtain a reasonable coverage of the Member State, zone or compartment;

(c)

the sampling points must be representative of the different ecosystems where the wild populations of susceptible species are located;

(d)

all production units, such as ponds, tanks and net cages, must be examined for the presence of dead, weak or abnormally behaving fish. Particular attention must be paid to the edge of cages or the water outlet area as relevant, where weak fish tend to accumulate because of the water current;

(e)

fish of listed species to be collected as samples must be selected as follows:

(i)

if Atlantic salmon are present, only fish of that species must be selected for sampling, except where other susceptible species are present which show typical signs of infection with HPR- deleted ISAV. If there are no Atlantic salmon in the establishment, the sample must be representative of all other susceptible species which are present;

(ii)

if moribund or freshly dead, but not decomposed fish are present, such fish must be selected, in particular fish demonstrating anaemia, haemorrhages or other clinical signs suggesting circulatory disturbances; if more than one water source is utilised for fish production, fish representing all water sources must be included in the sample;

(iii)

the fish selected must include fish collected in such a way that all production units, such as net cages, tanks and ponds, of the establishment as well as all year classes are proportionally represented in the sample.

Section 2 U.K. Granting of the status free from infection with HPR-deleted ISAV in Member States, zones and compartments of unknown health status

The status free from infection with HPR-deleted ISAV may only be granted to a Member State, a zone or a compartment with an unknown health status with regard to infection with HPR-deleted ISAV if all establishments and, when required, selected sampling points in wild populations selected in accordance with (b) of Section 1, have been subject to the following scheme:

(a)

the establishments or sampling points have been subject to health visits and sampled for a minimum period of 2 consecutive years as laid down in Table 2.A;

(b)

during that 2-year period, the testing of all samples using the diagnostic methods set out in point 2 of Section 5 must have produced negative results for HPR-deleted ISAV and any suspicion of infection must have been ruled out in accordance with the diagnostic methods set out in point 3 of Section 5;

(c)

If infection with HPR-deleted ISAV is detected during the surveillance referred to in point (a); before re-starting the scheme, relevant establishments within the Member State, zone or compartment must:

(i)

be subject to the minimum disease control measures laid down in Articles 58 to 65;

(ii)

be repopulated with fish from an establishment in a Member State, zone or compartment free from infection with HPR-deleted ISAV or from an establishment in a Member State, zone or compartment covered by an eradication programme for that disease.

Table 2.A

Scheme for Member States, zones and compartments for the 2-year control period which precedes the achievement of status free from infection with HPR-deleted ISAV

a Samples must be collected during spring and autumn each year.
Maximum number of fish per pool: 5.
Year of surveillance	Number of health visits per year to each establishment	Number of laboratory examinations per yeara	Number of fish in the sample
Year 1	6	2	75
Year 2	6	2	75

Section 3 U.K. Granting of the status free from infection with HPR-deleted ISAV in Member States, zones and compartments known to be infected with HPR-deleted ISAV

1. The status free from infection with HPR-deleted ISAV may only be granted to a Member State, a zone or a compartment known to be infected with HPR-deleted ISAV if all establishments keeping listed species within the Member State, zone or compartment have been subject to an eradication programme that complies with the following requirements: U.K.

(a)

the minimum control measures laid down in Articles 55 to Article 65 have been applied and a restricted zone of an appropriate size as provided for in point (c) of Article 58(1), where appropriate, divided into a protection zone and a surveillance zone, must have been established in the vicinity of the establishment(s) infected with HPR-deleted ISAV, taking into account the requirements set out in point 2;

(b)

all establishments keeping listed species within the protection zone, or where a protection zone has not been established, the restricted zone, not infected with HPR-deleted ISAV must be subject to an investigation comprising at least the following elements:

(i)

the collection of samples for testing of minimum 10 moribund fish, when clinical signs or post-mortem lesions consistent with infection with HPR-deleted ISAV are observed, or minimum 30 fish when clinical signs or post mortem lesions are not observed;

(ii)

in those establishments where the tests referred to in (i) have produced negative results, the health visits must continue once per month until the protection zone is withdrawn in accordance with point (c);

(c)

relevant establishments must be emptied in accordance with Article 62, cleaned and disinfected in accordance with Article 63 and fallowed in accordance with Article 64.

The duration of the fallowing period referred to in point (b) of Article 64(2) shall be at least 3 months. When all establishments infected within the same protection zone, or where a protection zone has not been established, the restricted zone, are emptied, at least 6 weeks of synchronised fallowing must be carried out.

When fallowing of the infected establishments is carried out, the restricted zone or the protection zone, when it has been established, must be converted into a surveillance zone until the scheme set out in Section 2 is completed;

(d)

repopulation may only take place when all infected establishments have been emptied, cleaned, disinfected and fallowed in accordance with point (c);

(e)

all establishments other than those referred to in point (f) which keep listed species within the Member State, zone or compartment covered by the eradication programme and when surveillance in wild populations is required, all sampling points selected in accordance with point (b) of Section 1, must subsequently be subject to the scheme set out in Section 2;

(f)

an individual establishment which keeps listed species and which has a health status which is independent of the health status of the surrounding waters is not required to comply with the scheme set out in Section 2 following a disease outbreak provided the establishment complies with the requirements set out in paragraph 3 of Article 80 and is re-populated with fish sourced from Member States, zones or compartments with status free from infection with HPR-deleted ISAV.

2. The restricted zone must have been defined on a case-by-case basis and: U.K.

(a)

it must take into account factors influencing the risks for the spread of infection with HPR-deleted ISAV to kept and wild fish, such as:

(i)

the number, rate and distribution of the mortalities on the establishment infected with HPR-deleted ISAV or in other aquaculture establishments;

(ii)

the distance to and density of neighbouring establishments;

(iii)

the proximity to slaughterhouses;

(iv)

contact establishments;

(v)

species present at the establishments;

(vi)

the farming practices applied in the infected establishments and in the neighbouring establishments to the infected establishment;

(vii)

the hydrodynamic conditions; and

(viii)

other factors of epidemiological significance identified;

(b)

the geographical demarcation in coastal areas must comply with the following minimum requirements:

(i)

the protection zone must consist of an area included in a circle with a radius of at least one tidal excursion or at least 5 km, whichever is larger, centred on the establishment infected with HPR-deleted ISAV, or an equivalent area determined according to appropriate hydrodynamic or epidemiological data; and

(ii)

the surveillance zone must consist of an area surrounding the protection zone, of overlapping tidal excursion zones; or an area surrounding the protection zone and included in a circle of radius 10km from the centre of the protection zone; or an equivalent area determined according to appropriate hydrodynamic or epidemiological data;

or

(iii)

where separate protection and surveillance zones are not established, the restricted zone must consist of an area comprising both the protection zone and the surveillance zone;

(c)

the geographical demarcation in inland areas must comprise the entire water catchment area in which the establishment infected with HPR-deleted ISAV is located. The competent authority may limit the extent of the restricted zone to parts of the water catchment area, provided this limitation does not compromise the disease control measures with respect to infection with HPR-deleted ISAV.

Section 4 U.K. Maintenance of status free from infection with HPR-deleted ISAV

1. When targeted surveillance is required in order to maintain the status free from infection with HPR-deleted ISAV of a Member State, a zone or a compartment, in accordance with Article 81, all establishments keeping listed species within the Member State, zone or compartment concerned must be subject to health visits and fish must be sampled in accordance with Table 2.B, taking into account the risk level of the establishment for the contraction of infection with HPR-deleted ISAV. U.K.

2. When determining the frequency of health visits required to maintain the status free from infection with HPR-deleted ISAV of compartments where the health status is dependent on the health status of the aquatic animal population in surrounding natural waters, the risk for the contraction of infection with HPR-deleted ISAV must be regarded as high. U.K.

3. Disease-free status must only be maintained as long as all samples tested, using the diagnostic methods set out in point 2 of Section 5, have produced negative results for HPR-deleted ISAV and any suspicion of infection with HPR-deleted ISAV has been ruled out in accordance with the diagnostic methods set out in point 3 of Section 5. U.K.

Table 2.B
Scheme for Member States, zones or compartments to maintain status free from infection with HPR-deleted ISAV a
a Shall not apply to establishments rearing only rainbow trout (Oncorhynchus mykiss) or brown trout (Salmo trutta) or both rainbow trout and brown trout, and where the water supply is exclusively based on fresh water sources which are not populated with Atlantic salmon (Salmo salar).
b Risk level assigned to the establishment by the competent authority as set out in paragraph 1 of Chapter 2 of Part I other than in the case of dependent compartments where all establishments are deemed to be high risk.
c Samples must be collected during spring and autumn when two samples are required each year
d Samples must be collected during spring or autumn when one sample per year is required.
Maximum number of fish per pool: 5
Risk levelb	Number of health visits per year	Number of laboratory examinations per yearc,d	Number of fish in the sample
High	2	2	30
Medium	1	1	30
Low	1 every 2 years	1 every 2 years	30

Section 5 U.K. Diagnostic and sampling methods

1. The organs or tissue material to be sampled and examined must be: U.K.

(a)

Histology: anterior-kidney, liver, heart, pancreas, intestine, spleen and gill;

(b)

Immunohistochemistry: mid-kidney and heart including valves and bulbus arteriosus;

(c)

RT-qPCR analysis: mid-kidney and heart;

(d)

Virus culture: mid-kidney, heart, liver and spleen.

Organ pieces from a maximum of five fish may be pooled.

2. The diagnostic method to be used to grant or to maintain status free from infection with HPR-deleted ISAV in accordance with Sections 2 to 4 must be RT-qPCR, followed by conventional RT-PCR and sequencing of the HE-gene of positive samples in accordance with the detailed methods and procedures which must be those approved by the EURL for fish diseases. U.K.

In the case of a positive RT-qPCR result, further samples must be tested before the implementation of the initial control measures provided for in Articles 55 to 65.

Those samples must be tested as follows in accordance with the detailed methods and procedures approved by the EURL for fish diseases:

(a)

screening of the samples by RT-qPCR, followed by conventional RT-PCR and sequencing of the HE-gene to verify HPR-deletion; and

(b)

detection of ISAV antigen in tissue preparations by means of specific antibodies against ISAV; or

(c)

isolation in cell culture and subsequent identification of HPR-deleted ISAV.

3. When a suspicion of infection with HPR-deleted ISAV must be confirmed or ruled out in accordance with Article 55, the following visit, sampling and testing procedure must comply with the following requirements: U.K.

(a)

the suspected establishment must be subject to at least one health visit and one sampling of 10 moribund fish, when clinical signs or post-mortem lesions consistent with infection with HPR-deleted ISAV are observed, or minimum 30 fish when clinical signs or post-mortem lesions are not observed. Samples shall be tested using one or more of the diagnostic methods set out in point 2 in accordance with the detailed diagnostic methods and procedures approved by the EURL for fish diseases;

(b)

in the case of a positive result of RT-qPCR for HPR-deleted ISAV, further samples shall be tested before the implementation of the initial control measures provided in Article 58. A suspected case of infection with HPR-deleted ISAV shall be confirmed in accordance with the following criteria using the detailed methods and procedures approved by the EURL for fish diseases:

(i)

Detection of ISAV by RT-qPCR, followed by sequencing of the HE-gene to verify HPR-deletion, and detection of ISAV in tissue preparations by means of specific antibodies against ISAV;

(ii)

detection of ISAV by RT-qPCR, including sequencing of the HE-gene to verify HPR-deletion; and isolation and identification of ISAV in cell culture from at least one sample from any fish from the establishment;

(c)

where the presence of clinical, gross pathological or histopathological findings consistent with infection are observed, the findings must be corroborated by virus detection by two diagnostic methods with independent principles of detection, such as RT-qPCR and IHC, in accordance with the procedures approved by the EURL for fish diseases.

The suspicion of HPR-deleted ISAV may be ruled out, if tests and health visits over a period of 12 months from the date of the suspicion are found to reveal no further evidence of the presence of the virus.

CHAPTER 3 U.K. Eradication, disease-free status and diagnostic methods for infection with Marteilia refringens

Section 1 U.K. General requirements for health visits and sampling

Health visits and sampling for the surveillance referred to in point (b)(ii) of Article 3(2) must comply with the following requirements:

(a)

health visits and, where appropriate, the sampling must be carried out in the period of the year when prevalence of the parasite in the Member State, zone or compartment is known to be maximal. When such data is not available, sampling must be carried out just after the water temperature has exceeded 17 °C;

(b)

when molluscs must be sampled in accordance with the requirements set out in Sections 2 to 4, the following selection criteria must apply:

(i)

if Ostrea spp. are present, only oysters of that species must be selected for sampling. If Ostrea spp. are not present, the sample must be representative of all other susceptible species present;

(ii)

if weak, gaping or freshly dead but not decomposed molluscs are present in the production units, such molluscs must primarily be selected. If such molluscs are not present, the molluscs selected must include the oldest healthy molluscs;

(iii)

when sampling in mollusc establishments which utilise more than one water source for mollusc production, molluscs representing all water sources must be included for sampling in such a way that all parts of the establishment are proportionally represented in the sample;

(iv)

when sampling in mollusc establishments or groups of establishments, molluscs from a sufficient number of sampling points, must be included in the sample in such a way that all parts of the establishment or group of establishments are proportionally represented in the sample. The main factors to be considered for the selection of these sampling points are previous sampling points where Marteilia refringens was detected, stocking density, water flows, presence of susceptible species, presence of vector species, bathymetry and management practices. Natural beds within or adjacent to the establishment or group of establishments must be included in the sampling.

Section 2 U.K. Granting of the status free from infection with Marteilia Refringens in Member States, zones and compartments of unknown health status

1. The status free from infection with Marteilia refringens may only be granted to a Member State, a zone or a compartment with an unknown health status with regard to infection with Marteilia refringens if all establishments or groups of establishments keeping listed species within the Member State, zone or compartment and where required, sampling points in wild populations, have been subject to the following 3-year scheme: U.K.

(a)

the establishments or groups of establishments keeping listed species have been subject to health visits and sampled for a minimum period of 3 consecutive years as laid down in Table 3.A;

(b)

during that 3-year period, the testing of all samples using the diagnostic methods set out in point 2 of Section 5 have produced negative results for Marteilia refringens and any suspicion of Marteilia refringens has been ruled out in accordance with the diagnostic methods set out in point 3 of Section 5;

(c)

when Ostrea edulis sourced from a Member State, zone or compartment of disease-free status are to be included in the sample, they must have been introduced into the establishment or group of establishments at least in the spring just preceding the period when the scheme is carried out.

2. If Marteilia refringens is detected during the 3-year scheme set out in point 1, before starting a new 3-year scheme, relevant establishments in the Member State, zone or compartment must: U.K.

(a)

be subject to the minimum disease control measures laid down in Articles 58 to 65;

(b)

be repopulated with molluscs from an establishment in a Member State, zone or compartment free from infection with Marteilia refringens or from an establishment in a Member State, zone or compartment covered by an eradication programme for that disease.

Table 3.A
Scheme for Member States, zones or compartments for the 3-year control period which precedes the achievement of status free from infection with Marteilia refringens
Year of surveillance	Number of health visits per year to each establishment/group of establishments	Number of laboratory examinations per year	Number of molluscs in the sample
Year 1	1	1	150
Year 2	1	1	150
Year 3	1	1	150

Section 3 U.K. Granting of the status free from infection with Marteilia refringens in Member States, zones and compartments known to be infected with Marteilia refringens

1. The status free from infection with Marteilia refringens may only be granted to a Member State, a zone or a compartment known to be infected with Marteilia refringens, where the competent authority judges that eradication of this disease to be feasible, if all establishments or groups of establishments keeping listed species within that Member State, zone or compartment have been subject to an eradication programme that complies with the following requirements: U.K.

(a)

the minimum control measures laid down in Articles 55 to 65 have effectively been applied and a restricted zone of an appropriate size as provided for in point (c) of Article 58(1), where appropriate divided into a protection zone and surveillance zone, must have been established in the vicinity of the establishment(s) or group of establishments infected with Marteilia refringens, taking into account the requirements set out in point 2;

(b)

all establishments and groups of establishments keeping listed species within the protection zone, or where a protection zone has not been established, the restricted zone, not infected with Marteilia refringens must be subject to an investigation comprising at least the collection of samples for the testing of 150 molluscs after the beginning of the transmission period of Marteilia refringens. When the transmission period is not known, the sampling must begin in the period after the temperature of the water exceeds 17 °C;

(c)

relevant establishments and groups of establishments must be emptied in accordance with Articles 62, and if possible cleaned and disinfected in accordance with Article 63.

Fallowing must be carried out in accordance with Article 64 and the duration of the fallowing period must be at least:

(i)

2 months in case of the establishments and groups of establishments which can be fully drained and thoroughly cleaned and disinfected such as hatcheries and nurseries;

(ii)

2 months in case of the establishments and groups of establishments which cannot be drained and thoroughly cleaned and disinfected provided that the infected molluscs of the listed species and those molluscs of the listed species with epidemiological links with the infected establishment or group of establishments have been harvested or removed before the period of the year when the prevalence of Marteilia refringens is known to be maximal, or when that period is not known, before the period when water temperature exceeds 17 °C;

(iii)

14 months in case of the establishments and groups of establishments which cannot be drained and thoroughly cleaned and disinfected if the infected molluscs of the listed species and those molluscs of the listed species with epidemiological links with the infected establishment or group of mollusc establishments have not been harvested or removed before the period of the year when the prevalence of Marteilia refringens is known to be maximal or when such data is not known, when molluscs of the susceptible species have not been harvested or removed before the period when water temperature exceeds 17 °C.

When all infected establishments and infected groups of establishments are emptied, at least 4 weeks of synchronised fallowing must be carried out;

(d)

repopulation may only take place when all infected establishments or infected groups of establishments have been emptied, cleaned, disinfected and fallowed in accordance with point (c);

(e)

all establishments and groups of establishments other than those referred to in point (f) which keep listed species within the Member State, zone or compartment covered by the eradication programme, must subsequently be subject to the scheme set out in Section 2;

(f)

an individual establishment which keeps listed species and which has a health status which is independent of the health status of the surrounding waters is not required to comply with the scheme set out in Section 2 following a disease outbreak provided the establishment complies with the requirements set out in paragraph 3 of Article 80 and is repopulated with molluscs sourced from Member States, zones or compartments with status free from infection with Marteilia refringens.

2. The restricted zone must have been defined on a case-by-case basis and: U.K.

(a)

it must take into account factors influencing the risks for the spread of infection with Marteilia refringens including other establishments and wild molluscs, such as:

(i)

the number, age, rate and distribution of the mortalities of molluscs on the establishment or group of establishments infected with Marteilia refringens;

(ii)

the distance to and density of neighbouring establishments or groups of establishments and wild molluscs;

(iii)

the proximity to processing establishments, contact establishments or groups of establishments;

(iv)

the species, especially susceptible species and vector species, present at the establishments or groups of establishments;

(v)

the farming practices applied in the affected and neighbouring establishments and groups of establishments;

(vi)

the hydrodynamic conditions; and

(vii)

other factors of epidemiological significance identified;

(b)

the geographical demarcation must comply with the following minimum requirements:

(i)

the protection zone must consist of an area included in a circle with a radius of at least one tidal excursion or at least 5 km, whichever is larger, centred on the establishment infected with Marteilia refringens, or an equivalent area determined according to appropriate hydrodynamic or epidemiological data; and

(ii)

the surveillance zone must consist of an area surrounding the protection zone, of overlapping tidal excursion zones; or an area surrounding the protection zone and included in a circle of radius 10km from the centre of the protection zone; or an equivalent area determined according to appropriate hydrodynamic or epidemiological data;

or

(iii)

where separate protection and surveillance zones are not established, the restricted zone must consist of an area comprising both the protection zone and the surveillance zone.

Section 4 U.K. Maintenance of status free from infection with Marteilia refringens

1. When targeted surveillance is required in order to maintain the status free from infection with Marteilia refringens of a Member State, a zone or a compartment, in accordance with Article 81, all establishments keeping listed species within the Member State, zone or compartment concerned must be subject to health visits and molluscs must be sampled in accordance with Table 3.B, taking into account the risk level of the establishment for the contraction of infection with Marteilia refringens. U.K.

2. When determining the frequency of health visits required to maintain the status free from infection with Marteilia refringens of compartments, where the health-status regarding that disease is dependent on the health-status of the aquatic animal populations in surrounding natural waters, the risk for the contraction of infection with Marteilia refringens must be regarded as high. U.K.

3. The status free from infection with Marteilia refringens may only be maintained as long as all samples tested, using the diagnostic methods set out in point 2 of Section 5 have produced negative results for Marteilia refringens and any suspicion of infection with Marteilia refringens has been ruled out in accordance with the diagnostic methods set out in point 3 of section 5. U.K.

Table 3.B
Scheme for Member States, zones or compartments to maintain disease-free status for Marteilia refringens
a Risk level assigned to the establishment by the competent authority as set out in paragraph 1 of Chapter 2 of Part I other than in the case of dependent compartments where all establishments are deemed to be high risk.
Risk levela	Number of health visits to each establishment/group of establishments	Number of laboratory examinations	Number of molluscs in the sample
High	1 every year	1 every 2 years	150
Medium	1 every 2 years	1 every 2 years	150
Low	1 every 3 years	1 every 3 years	150

Section 5 U.K. Diagnostic and sampling methods

1. The whole animal must be submitted to the laboratory for the performance of the diagnostic tests provided for in points 2 and 3. U.K.

2. The diagnostic methods to be used to grant or maintain status free from infection with Marteilia refringens in accordance with Sections 2 to 4 must follow the detailed diagnostic methods and procedures approved by the EURL for Mollusc Diseases and must be histopathology, tissue imprints or PCR. U.K.

3. When a suspicion of infection with Marteilia refringens is required to be confirmed or ruled out in accordance with Article 55 the following visit, sampling and testing procedure must be complied with: U.K.

(a)

the investigation must include at least one sampling of 30 molluscs of susceptible species if the suspicion is based on a mortality report or if not, 150 molluscs of susceptible species after the beginning of the transmission period of Marteilia refringens. When the transmission period is not known, the sampling must begin in the period after the temperature of the water exceeds 17 °C;

(b)

samples must be tested using the diagnostic methods set out in point (i) following the detailed diagnostic methods and procedures approved by the EURL for Mollusc Diseases:

(i)

the presence of Marteilia refringens must be considered as confirmed when a positive result by histopathology, tissue imprints or in situ hybridisation is combined with a positive PCR result completed by sequencing. If biological material is not available for histopathology, tissue imprints or in situ hybridisation, the presence of Marteilia refringens must be considered as confirmed when positive results are obtained using two PCR assays targeting different fragments of the parasite genome and completed by sequencing;

(ii)

the suspicion of infection with Marteilia refringens may be ruled out, if the tests referred to in (i) reveal no further evidence of the presence of Marteilia refringens.

CHAPTER 4 U.K. Eradication, disease-free status and diagnostic methods for infection with Bonamia exitiosa

Section 1 U.K. General requirements for health visits and sampling

Health visits and sampling for the surveillance referred to in point (b)(ii) of Article 3(2) must comply with the following requirements:

(a)

health visits and, where appropriate, the sampling must be carried out in the period of the year when prevalence of the parasite in the Member State, zone or compartment is known to be maximal. When such data is not available, sampling shall be carried out twice a year, in spring and autumn;

(b)

when molluscs are to be sampled in accordance with the requirements set out in Sections 2 to 4, the following criteria must apply:

(i)

if Ostrea spp. are present, only oysters of that species must be selected for sampling. If Ostrea spp. are not present, the sample must be representative of all other susceptible species present;

(ii)

if weak, gaping or freshly dead but not decomposed molluscs are present, such molluscs must primarily be selected. If such molluscs are not present, the molluscs selected must include the oldest healthy molluscs;

(iii)

when sampling in establishments or groups of establishments which utilise more than one water source for mollusc production, molluscs representing all water sources must be included for sampling in such a way that all parts of the establishment are proportionally represented in the sample;

(iv)

when sampling in mollusc establishments or groups of establishments, molluscs from a sufficient number of sampling points must be included in the sample in such a way that all parts of the establishment or group of establishments are proportionally represented in the sample. The main factors to be considered for the selection of those sampling points are previous points where Bonamia exitiosa was detected, stocking density, water flows, the presence of susceptible species, the presence of vector species (e.g.Crassostrea gigas), bathymetry and management practices. Natural beds within or adjacent to the establishment or group of establishments shall be included in the sampling.

Section 2 U.K. Granting of the status free from infection with Bonamia exitiosa in Member States, zones and compartments of unknown health status

1. The status free from infection with Bonamia exitiosa may only be granted to a Member State, a zone or a compartment with an unknown health status with regard to infection with Bonamia exitiosa if all establishments or groups of establishments keeping listed species within the Member State, zone or compartment and where required, sampling points in wild populations, have been subject to the following 3-year scheme: U.K.

(a)

the establishments and groups of establishments keeping listed species have been subject to health visits and sampled for a minimum period of 3 consecutive years as laid down in Table 4.A;

(b)

during that 3-year period, the testing of all samples using the diagnostic methods set out in point 2 of Section 5 have produced negative results for Bonamia exitiosa and any suspicion of Bonamia exitiosa has been ruled out in accordance with the diagnostic methods set out in point 3 of Section 5;

(c)

when Ostrea edulis sourced from a Member State, zone or compartment of disease-free status are to be included in the sample, they must have been introduced into the establishment or group of establishments at least one year before the scheme is carried out.

2. If infection with Bonamia exitiosa is detected during the 3-year scheme referred to in point 1; before starting a new 3-year scheme, relevant establishments in the Member State, zone or compartment must: U.K.

(a)

be subject to the minimum disease control measures laid down in Articles 58 to 65;

(b)

be repopulated with molluscs from an establishment in a Member State, zone or compartment free from infection with Bonamia exitiosa or from an establishment in a Member State, zone or compartment covered by an eradication programme for that disease.

Table 4.A
Scheme for Member States, zones or compartments for the 3-year control period which precedes the achievement of status free from infection with Bonamia exitiosa
Year of surveillance	Number of health visits per year to each establishment or group of establishments	Number of laboratory examinations per year	Number of molluscs in the sample
Year 1	2	2	150
Year 2	2	2	150
Year 3	2	2	150

Section 3 U.K. Granting of the status free from infection with Bonamia exitiosa in Member States, zones and compartments known to be infected with Bonamia exitiosa

1. The status free from infection with Bonamia exitiosa may only be granted to a Member State, a zone or a compartment known to be infected with Bonamia exitiosa, where the competent authority judges that eradication of this disease to be feasible, if all establishments or groups of establishments keeping listed species within that Member State, zone or compartment have been subject to an eradication programme that complies with the following requirements: U.K.

(a)

the minimum control measures laid down in Articles 55 to 65 must have been effectively applied, and a restricted zone of an appropriate size as provided for in point (c) of Article 58(1), where appropriate, divided into a protection zone and surveillance zone; must have been established in the vicinity of the establishment or group of establishments declared infected with Bonamia exitiosa taking into account the requirements set out in point 2;

(b)

all establishments and groups of establishments keeping listed species within the protection zone or where a protection zone has not been established, within the restricted zone, not infected with Bonamia exitiosa must be subject to an investigation comprising at least the collection of samples for testing of 150 molluscs of susceptible species after the beginning of the transmission period of Bonamia exitiosa. When the transmission period is not known, the sampling must be done on oysters which have spent at least one year within the protection zone;

(c)

relevant establishments and groups of establishments must be emptied in accordance with Article 62, and if possible, cleaned and disinfected in accordance with Article 63.

Fallowing must be carried out in compliance with Article 64 and the duration of the fallowing period must be at least 6 months.

When all infected establishments or infected groups of establishments are emptied, at least 4 weeks of synchronised fallowing must be carried out;

(d)

repopulation may only take place when all infected establishments or infected groups of establishments have been emptied, cleaned, disinfected and fallowed in accordance with point (c);

(e)

all establishments and groups of establishments other than those referred to in point (f) which keep listed species within the Member State, zone or compartment covered by the eradication programme, must subsequently be subject to the scheme set out in Section 2;

(f)

an individual establishment which keeps listed species and which has a health-status which is independent of the health-status of the surrounding waters is not required to comply with the scheme set out in Section 2 following a disease outbreak provided the establishment complies with the requirements set out in paragraph 3 of Article 80 and is repopulated with molluscs sourced from Member States, zones or compartments with status free from infection with Bonamia exitiosa.

2. The restricted zone must have been defined on a case-by-case basis and: U.K.

(a)

it must take into account factors influencing the risks for the spread of infection with Bonamia exitiosa including other establishments and wild molluscs, such as:

(i)

the number, age, rate and distribution of the mortalities of molluscs on the establishment or group of establishments infected with Bonamia exitiosa;

(ii)

the distance to and density of neighbouring establishments or groups of establishments and wild molluscs;

(iii)

the proximity to processing establishments, contact establishments or groups of establishments;

(iv)

the species, especially susceptible species and vector species, present at the establishments or groups of establishments;

(v)

the farming practices applied in the affected and neighbouring establishments and groups of establishments;

(vi)

the hydrodynamic conditions; and

(vii)

other factors of epidemiological significance identified;

(b)

the geographical demarcation must comply with the following minimum requirements:

(i)

the protection zone must consist of an area included in a circle with a radius of at least one tidal excursion or at least 5 km, whichever is larger, centred on the establishment infected with Bonamia exitiosa, or an equivalent area determined according to appropriate hydrodynamic or epidemiological data; and

(ii)

the surveillance zone must consist of an area surrounding the protection zone, of overlapping tidal excursion zones; or an area surrounding the protection zone and included in a circle of radius 10km from the centre of the protection zone; or an equivalent area determined according to appropriate hydrodynamic or epidemiological data;

or

(iii)

where separate protection and surveillance zones are not established, the restricted zone must consist of an area comprising both the protection zone and the surveillance zone.

Section 4 U.K. Maintenance of status free from infection with Bonamia exitiosa

1. When targeted surveillance is required in order to maintain the status free from infection with Bonamia exitiosa of a Member State, a zone or a compartment, in accordance with Article 81, all establishments keeping listed species within the Member State, zone or compartment concerned must be subject to health visits and molluscs must be sampled in accordance with Table 4.B, taking into account the risk level of the establishment for the contraction of infection with Bonamia exitiosa U.K.

2. When determining the frequency of health visits required to maintain the status free from infection with Bonamia exitiosa of compartments where the health status regarding that disease is dependent on the health status of the aquatic animal populations in surrounding natural waters, the risk for the contraction of infection with Bonamia exitiosa must be regarded as high. U.K.

3. The status free from infection with Bonamia exitiosa may only be maintained as long as all samples, using the diagnostic methods set out in point 2 of Section 5 have produced negative results for Bonamia exitiosa and any suspicion of infection with Bonamia exitiosa has been ruled out in accordance with the diagnostic methods set out in point 3 of Section 5. U.K.

Table 4.B
Scheme for Member States, zones or compartments to maintain status free from infection with Bonamia exitiosa
a Risk level assigned to the establishment by the competent authority as set out in paragraph 1 of Chapter 2 of Part I other than in the case of dependent compartments where all establishments are deemed to be high risk.
Risk levela	Number of health visits to each establishment/group of establishments	Number of laboratory examinations	Number of molluscs in the sample
High	1 every year	1 every 2 years	150
Medium	1 every 2 years	1 every 2 years	150
Low	1 every 3 years	1 every 3 years	150

Section 5 U.K. Diagnostic and sampling methods

1. The whole animal must be submitted to the laboratory for the performance of the diagnostic tests provided for in points 2 and 3. U.K.

2. The diagnostic methods to be used to grant or maintain status free from infection with Bonamia exitiosa, in accordance with Sections 2 to 4 must follow the detailed diagnostic methods and procedures approved by the EURL for Mollusc Diseases and must be histopathology, tissue imprints or PCR. U.K.

3. When a suspicion of infection with Bonamia exitiosa is required to be confirmed or ruled out in accordance with Article 58, the following visit, sampling and testing procedure must be complied with: U.K.

(a)

the investigation must include at least one sampling of 30 molluscs of susceptible species if the suspicion is based on a mortality report, or if not, 150 molluscs of susceptible species after the beginning of the transmission period of Bonamia exitiosa. When the transmission period is not known, the sampling shall be carried out twice a year, in spring and autumn;

(b)

the samples must be tested using the diagnostic methods set out in point (i) following the detailed diagnostic methods and procedures which have been approved by the EURL for Mollusc Diseases:

(i)

the presence of Bonamia exitiosa must be considered as confirmed when a positive result by histopathology, tissue imprints or in situ hybridisation is combined with a positive result by PCR followed by sequencing. If biological material is not available for histopathology, tissue imprints or in situ hybridisation, the presence of Bonamia exitiosa must be considered as confirmed when positive results are obtained using two PCR assays targeting different fragments of the parasite genome and completed by sequencing;

(ii)

the suspicion of the presence of infection with Bonamia exitiosa must be ruled out, if those tests reveal no further evidence of the presence of Bonamia exitiosa.

CHAPTER 5 U.K. Eradication, disease-free status and diagnostic methods for infection with Bonamia ostreae

Section 1 U.K. General requirements for health visits and sampling

Health visits and sampling for the surveillance referred to in point (b)(ii) of Article 3(2) must comply with the following requirements:

(a)

health visits and, where appropriate, the sampling must be carried out in the period of the year when prevalence of the parasite in the Member State, zone or compartment is known to be maximal. When such data is not available, sampling must be carried out in winter or at the beginning of spring;

(b)

when molluscs are to be sampled in accordance with the requirements set out in Sections 2 to 4, the following criteria must apply:

(i)

if Ostrea edulis are present, only oysters of that species must be selected for sampling. If Ostrea edulis are not present, the sample must be representative of all other susceptible species present;

(ii)

if weak, gaping or freshly dead but not decomposed molluscs are present, such molluscs must primarily be selected. If such molluscs are not present, the molluscs selected must include the oldest healthy molluscs;

(iii)

when sampling in establishments or groups of establishments which utilise more than one water source for mollusc production, molluscs representing all water sources must be included for sampling in such a way that all parts of the establishment are proportionally represented in the sample;

(iv)

when sampling in mollusc establishments or groups of establishments, molluscs from a sufficient number of sampling points must be included in the sample in such a way that all parts of the establishment or group of establishments are proportionally represented in the sample. The main factors to be considered for the selection of those sampling points are previous points where Bonamia ostreae was detected, stocking density, water flows, the presence of susceptible species, the presence of vector species, bathymetry and management practices. Natural beds within or adjacent to the establishment or group of establishments shall be included in the sampling.

Section 2 U.K. Granting of the status free from infection with Bonamia ostreae in Member States, zones and compartments of unknown health status

1. The status free from infection with Bonamia ostreae may only be granted to a Member State, a zone or a compartment with an unknown health status with regard to infection with Bonamia ostreae if all establishments or groups of establishments keeping listed species within the Member State, zone or compartment and where required, sampling points in wild populations, have been subject to the following 3-year scheme: U.K.

(a)

the establishments and groups of establishments keeping listed species have been subject to health visits and sampled for a minimum period of 3 consecutive years as laid down in Table 5.A;

(b)

during that 3-year period, the testing of all samples using the diagnostic methods set out in point 2 of Section 5 have produced negative results for Bonamia ostreae and any suspicion of Bonamia ostreae has been ruled out in accordance with the diagnostic methods set out in point 3 of Section 5;

(c)

when Ostrea edulis sourced from a Member State, zone or compartment of disease-free status are to be included in the sample, they must have been introduced into the establishment or group of establishments at least one year before the scheme is carried out.

2. If infection with Bonamia ostreae is detected during the 3-year scheme referred to in point 1; before starting a new 3-year scheme, relevant establishments in the Member State, zone or compartment must: U.K.

(a)

be subject to the minimum disease control measures laid down in Articles 58 to 65;

(b)

be repopulated with molluscs from an establishment in a Member State, zone or compartment free from infection with Bonamia ostreae or from an establishment in a Member State, zone or compartment covered by an eradication programme for that disease.

Section 3 U.K. Granting of the status free from infection with Bonamia ostreae in Member States, zones and compartments known to be infected with Bonamia ostreae

1. The status free from infection with Bonamia ostreae may only be granted to a Member State, a zone or a compartment known to be infected with Bonamia ostreae, where the competent authority judges that eradication of this disease to be feasible, if all establishments or groups of establishments keeping listed species within that Member State, zone or compartment have been subject to an eradication programme that complies with the following requirements: U.K.

(a)

the minimum control measures laid down in Articles 55 to 65 must have been effectively applied, and a restricted zone of an appropriate size as provided for in point (c) of Article 58(1), where appropriate, divided into a protection zone and surveillance zone; must have been established in the vicinity of the establishment or group of establishments declared infected with Bonamia ostreae taking into account the requirements set out in point 2;

(b)

all establishments and groups of establishments keeping listed species within the protection zone or where a protection zone has not been established, within the restricted zone, not infected with Bonamia ostreae must be subject to an investigation comprising at least the collection of samples for testing of 150 molluscs of susceptible species after the beginning of the transmission period of Bonamia ostreae. When the transmission period is not known, the sampling must begin in winter or at the beginning of spring;

(c)

relevant establishments and groups of establishments must be emptied in accordance with Article 62, and if possible, cleaned and disinfected in accordance with Article 63.

Fallowing must be carried out in compliance with Article 64 and the duration of the fallowing period must be at least 6 months.

When all infected establishments or infected groups of establishments are emptied, at least 4 weeks of synchronised fallowing must be carried out;

(d)

repopulation may only take place when all infected establishments or infected groups of establishments have been emptied, cleaned, disinfected and fallowed in accordance with point (c);

(e)

all establishments and groups of establishments other than those referred to in point (f) which keep listed species within the Member State, zone or compartment covered by the eradication programme, must subsequently be subject to the scheme set out in Section 2;

(f)

an individual establishment which keeps listed species and which has a health status which is independent of the health status of the surrounding waters is not required to comply with the surveillance scheme set out in Section 2 following a disease outbreak provided the establishment complies with the requirements set out in paragraph 3 of Article 80 and is repopulated with molluscs sourced from Member States, zones or compartments with status free from infection with Bonamia ostreae.

2. The restricted zone must have been defined on a case-by-case basis and: U.K.

(a)

it must take into account factors influencing the risks for the spread of infection with Bonamia ostreae including other establishments and wild molluscs, such as:

(i)

the number, age, rate and distribution of the mortalities of molluscs on the establishment or group of establishments infected with Bonamia ostreae;

(ii)

the distance to and density of neighbouring establishments or groups of establishments and wild molluscs;

(iii)

the proximity to processing establishments, contact establishments or groups of establishments;

(iv)

the species, especially susceptible species and vector species, present at the establishments or groups of establishments;

(v)

the farming practices applied in the affected and neighbouring establishments and groups of establishments;

(vi)

the hydrodynamic conditions; and

(vii)

other factors of epidemiological significance identified;

(b)

the geographical demarcation must comply with the following minimum requirements:

(i)

the protection zone must consist of an area included in a circle with a radius of at least one tidal excursion or at least 5 km, whichever is larger, centred on the establishment infected with Bonamia ostreae, or an equivalent area determined according to appropriate hydrodynamic or epidemiological data; and

(ii)

the surveillance zone must consist of an area surrounding the protection zone, of overlapping tidal excursion zones; or an area surrounding the protection zone and included in a circle of radius 10km from the centre of the protection zone; or an equivalent area determined according to appropriate hydrodynamic or epidemiological data;

or

(iii)

where separate protection and surveillance zones are not established, the restricted zone must consist of an area comprising both the protection zone and the surveillance zone.

Table 5.A
Scheme for Member States, zones or compartments for the 3-year control period which precedes the achievement of status free from infection with Bonamia ostreae
Year of surveillance	Number of health visits per year to each establishment or group of establishments	Number of laboratory examinations per year	Number of molluscs in the sample
Year 1	1	1	150
Year 2	1	1	150
Year 3	1	1	150

Section 4 U.K. Maintenance of status free from infection with Bonamia ostreae

1. When targeted surveillance is required in order to maintain the status free from infection with Bonamia ostreae of a Member State, a zone or a compartment, in accordance with Article 81, all establishments keeping listed species within the Member State, zone or compartment concerned must be subject to health visits and molluscs must be sampled in accordance with Table 5.B, taking into account the risk level of the establishment for the contraction of infection with Bonamia ostreae. U.K.

2. When determining the frequency of health visits required to maintain the status free from infection with Bonamia ostreae of compartments where the health status regarding that disease is dependent on the health status of the aquatic animal populations in surrounding natural waters, the risk for the contraction of infection with Bonamia ostreae must be regarded as high. U.K.

3. The status free from infection with Bonamia ostreae may only be maintained as long as all samples, using the diagnostic methods set out in point 2 of Section 5 have produced negative results for Bonamia ostreae and any suspicion of infection with Bonamia ostreae has been ruled out in accordance with the diagnostic methods set out in point 3 of Section 5. U.K.

Table 5.B
Scheme for Member States, zones or compartments to maintain status free from infection with Bonamia ostreae
a Risk level assigned to the establishment by the competent authority as set out in paragraph 1 of Chapter 2 of Part I other than in the case of dependent compartments where all establishments are deemed to be high risk.
Risk levela	Number of health visits to each establishment/group of establishments	Number of laboratory examinations	Number of molluscs in the sample
High	1 every year	1 every 2 years	150
Medium	1 every 2 years	1 every 2 years	150
Low	1 every 3 years	1 every 3 years	150

Section 5 U.K. Diagnostic and sampling methods

1. The whole animal must be submitted to the laboratory for the performance of the diagnostic tests provided for in points 2 and 3. U.K.

2. The diagnostic methods to be used to grant or maintain status free from infection with Bonamia ostreae, in accordance with Sections 2 to 4 must follow the detailed diagnostic methods and procedures approved by the EURL for Mollusc Diseases and must be histopathology, tissue imprints or PCR. U.K.

3. When a suspicion of infection with Bonamia ostreae is required to be confirmed or ruled out in accordance with Article 58, the following visit, sampling and testing procedure must be complied with: U.K.

(a)

the investigation must include at least one sampling of 30 molluscs of susceptible species if the suspicion is based on a mortality report, or if not, 150 molluscs of susceptible species after the beginning of the transmission period of Bonamia ostreae. When the transmission period is not known, the sampling shall begin in the winter or at the beginning of spring;

(b)

the samples must be tested using the diagnostic methods set out in point (i) following the detailed diagnostic methods and procedures which have been approved by the EURL for Mollusc Diseases:

(i)

the presence of Bonamia ostreae must be considered as confirmed when a positive result by histopathology, tissue imprints or in situ hybridisation is combined with a positive result by PCR followed by sequencing. If biological material is not available for histopathology, tissue imprints or in situ hybridisation, the presence of Bonamia ostreae must be considered as confirmed when positive results are obtained using two PCR assays targeting different fragments of the parasite genome and completed by sequencing;

(ii)

the suspicion of the presence of infection with Bonamia ostreae must be ruled out, if those tests reveal no further evidence of the presence of Bonamia ostreae.

CHAPTER 6 U.K. Eradication, disease-free status and diagnostic methods for infection with white spot syndrome virus (WSSV)

Section 1 U.K. General requirements for health visits and sampling

Health visits and sampling for the surveillance referred to in point (b)(ii) of Article 3(2) must comply with the following requirements:

(a)

the sampling of crustaceans for laboratory examination must be carried out whenever the water temperature is likely to reach its highest annual point. That requirement concerning water temperature must also apply to health visits where these are feasible;

(b)

when farmed crustaceans must be sampled in accordance with the requirements set out in Sections 2 to 4, the following criteria must apply:

(i)

if weak or moribund crustaceans are present in the production units, such crustaceans must primarily be selected. If such crustaceans are not present, those selected must include crustaceans of different size cohorts namely juveniles and adults of the selected susceptible species, proportionally represented in the sample;

(ii)

if more than one water source is utilised for crustacean production, susceptible crustaceans representing all water sources must be included for sampling;

(c)

when targeted surveillance in wild populations is required due to the small number of establishments covered by the eradication programme, the number and geographical distribution of the sampling points must be determined to obtain a reasonable coverage of the Member State, zone or compartment. The sampling points must also be representative of the different ecosystems where the wild populations of susceptible species are located namely marine, estuary, river and lake systems. In such situations, the crustaceans to be sampled must be selected as follows:

(i)

in marine and estuary systems areas, one or more of the following species must be selected: Carcinus maenas, Cancer pagurus, Eriocheir sinensis, Liocarcinus depurator, Liocarcinus puber, Crangon crangon, Homarus gammarus, Palaemon adspersus or penaeid shrimp species namely Penaeus japonicus, Penaeus kerathurus, Penaeus semisulcatus. If those species are not present, the sample must be representative of other susceptible decapod species present;

(ii)

in river and lake systems, one or more of the following species must be selected: Pacifastacus leniusculus, Astacus leptodactylus, Austropotamobius pallipes or Orconectes limosus. If those species are not present, the sample must be representative of other susceptible decapod species present;

(iii)

if weak or moribund crustaceans are present, such crustaceans must primarily be selected. If such crustaceans are not present, those selected must include crustaceans of different size cohorts namely juveniles and adults of the selected susceptible species, proportionally represented in the sample.

Section 2 U.K. Granting of the status free from infection with WSSV in Member States, zones and compartments of unknown health status

1. The status free from infection with WSSV may only be granted to a Member State, a zone or a compartment with an unknown health status with regard to infection with WSSV if all establishments or groups of establishments keeping listed species within the Member State, zone or compartment and where required, sampling points in wild populations, have been subject to the following 2-year scheme: U.K.

(a)

the establishments or groups of establishments have been subject to health visits and sampled for a minimum period of 2 consecutive years as laid down in Table 6.A;

(b)

during that 2-year period, the testing of all samples using the diagnostic methods set out in point 2 of Section 5 have produced negative results for infection with WSSV and any suspicion of infection with WSSV has been ruled out in accordance with the diagnostic methods set out in point 3 of Section 5;

2. If infection with WSSV is detected during the 2-year scheme referred to in point 1, before starting a new 2-year scheme, relevant establishments in the Member State, zone or compartment must: U.K.

(a)

be subject to the minimum disease control measures laid down in Articles 58 to 65;

(b)

be repopulated with crustaceans from an establishment in a Member State, zone or compartment free from infection with WSSV or from an establishment in a Member State, zone or compartment covered by an eradication programme for that disease.

Section 3 U.K. Granting of the status free from infection with WSSV in Member States, zones and compartments known to be infected with WSSV

1. The status free from infection with WSSV may only be granted to a Member State, a zone or a compartment known to be infected with WSSV if all establishments keeping listed species within that Member State, zone or compartment have been subject to an eradication programme that complies with the following requirements: U.K.

(a)

the minimum control measures laid down in Articles 55 to 65 must have been effectively applied, and a restricted zone of an appropriate size as provided for in point (c) of Article 58(1), where appropriate, divided into a protection zone and surveillance zone; must have been established in the vicinity of the establishment(s) declared infected with WSSV taking into account the requirements set out in point 2;

(b)

all establishments keeping listed species within the protection zone, or where a protection zone has not been established, the restricted zone, not infected with WSSV must be subject to an investigation comprising at least the following:

(i)

the collection of samples for testing of 10 crustaceans, when clinical signs or post-mortem lesions consistent with infection WSSV are observed, or 150 crustaceans, when clinical signs or post-mortem lesions are not observed; and

(ii)

health visits; in those establishments where the tests referred to in (i) have produced negative results, health visits must continue once per month during the season when the water temperature is likely to reach its highest annual points, until the protection zone has been withdrawn in accordance with point (c);

(c)

relevant establishments must be emptied in accordance with Articles 62, cleaned disinfected in accordance with Article 63 and fallowed in accordance with Article 64. The duration of the fallowing period must be at least 6 weeks. When all infected establishments are emptied, at least 3 weeks of synchronous fallowing shall be carried out.

When fallowing of the officially declared infected establishments is carried out, the protection zones shall be converted into surveillance zones;

(d)

repopulation may only take place when all infected establishments have been emptied, cleaned, disinfected and fallowed in accordance with point (c);

(e)

all establishments other than those referred to in point (f) which keep listed species within the Member State, zone or compartment covered by the eradication programme and, when surveillance in wild populations is required, all sampling points selected to provide the greatest coverage of the geographical area included in the eradication programme must be subject at least to the scheme set out in Section 2;

(f)

an individual establishment which keeps listed species and which has a health status which is independent of the health status of the surrounding waters is not required to comply with the scheme set out in Section 2 following a disease outbreak provided the establishment complies with the requirements set out in paragraph 3 of Article 80 and is repopulated with crustaceans sourced from Member States, zones or compartments with status free from infection with WSSV.

2. The restricted zone must have been defined on a case-by-case basis taking into account factors influencing the risks for the spread of WSSV to farmed and wild crustaceans, such as: U.K.

(i)

the number, age, rate and distribution of the mortalities of crustaceans on the establishment or group of establishments infected with WSSV including other establishments and wild crustaceans;

(ii)

the distance to and density of neighbouring establishments or groups of establishments including wild crustaceans;

(iii)

the proximity to processing establishments, contact establishments or groups of establishments;

(iv)

the species, especially susceptible species and vector species, present at the establishments or groups of establishments;

(v)

the farming practices applied in the affected and neighbouring establishments and groups of establishments;

(vi)

the hydrodynamic conditions; and

(vii)

other factors of epidemiological significance identified.

Table 6. A
Scheme for Member States, zones and compartments for the 2-year control period which precedes the achievement of status free from infection with WSSV
Year of surveillance	Number of health visits per year to each establishment or group of establishments	Number of laboratory examinations per year	Number of crustaceans in the sample
Year 1	1	1	150
Year 2	1	1	150

Section 4 U.K. Maintenance of status free from infection with WSSV

1. When targeted surveillance is required in order to maintain the status free from infection with WSSV of a Member State, a zone or a compartment, in accordance with Article 81, all establishments keeping listed species within the Member State, zone or compartment concerned must be subject to health visits and crustaceans must be sampled in accordance with Table 6.B, taking into account the risk level of the establishment for the contraction of infection with WSSV. U.K.

2. In Member States, zones or compartments where the number of establishments is limited and targeted surveillance in those establishments does not provide sufficient epidemiological data, the surveillance to maintain disease-free status must include sampling points selected in accordance with the requirements laid down in point (b) of Section 1. U.K.

3. When determining the frequency of health visits required to maintain the status free from infection with WSSV of compartments where the health status regarding that disease is dependent on the health status of the aquatic animal populations in surrounding natural waters, the risk for the contraction of infection with WSSV must be regarded as high. U.K.

4. The status free from infection with WSSV may only be maintained as long as all samples, using the diagnostic methods set out in point 2 of Section 5 have produced negative results for WSSV and any suspicion of infection with WSSV has been ruled out in accordance with the diagnostic methods set out in point 3 of Section 5. U.K.

Table 6. B
Scheme for Member States, zones or compartments to maintain status free from infection WSSV
a Risk level assigned to the establishment by the competent authority as set out in paragraph 1 of Chapter 2 of Part I other than in the case of dependent compartments where all establishments are deemed to be high risk.
Risk levela	Number of health visits to each establishment/group of establishments	Number of laboratory examinations	Number of crustaceans in the sample
High	1 every year	1 every 2 years	150
Medium	1 every 2 years	1 every 2 years	150
Low	1 every 2 years	1 every 4 years	150

Section 5 U.K. Diagnostic and sampling methods

1. Samples of integumental epidermis, either dissected or contained within walking legs, pleopods, mouthparts or gills of the test animal must be fixed in 95 % ethanol prior to the preparation of samples for PCR. U.K.

Other samples, fixed for histology and transmission electron microscopy may be collected to support diagnostic data arising from PCR.

2. The diagnostic method and procedures to be used to grant or to maintain disease-free status with regard to infection with WSSV must be PCR followed by sequencing. When applying these diagnostic methods, the corresponding detailed methods and procedures which have been approved by the EURL for Crustacean Diseases must be followed. U.K.

In the case of a positive result from the PCR test, the result must be followed by sequencing of the amplicon before the initial control measures provided for in Article 63 of Regulation (EU) 2016/429 are implemented.

3. When a suspicion of infection with WSSV is required to be confirmed or ruled out in accordance with Article 58, the following visit, sampling and testing procedure must be complied with: U.K.

(a)

the investigation must include at least one health visit and one sampling of 10 crustaceans when clinical signs or post-mortem lesions consistent with infection with WSSV are observed or 150 crustaceans when clinical signs or post-mortem lesions are not observed. The samples must be tested using the diagnostic method set out in point 2;

(b)

the presence of WSSV must be considered as confirmed when PCR followed by sequencing, carried out in accordance with the detailed methods and procedures which have been approved by the EURL for Crustacean Diseases test positive for WSSV.

The suspicion of infection with WSSV may be ruled out, if those tests reveal no further evidence of the presence of the virus.