S.I. No. 112/1996 - European Communities (Trade in Bovine Breeding Animlas, Their Semen,Ova and Embryos) Regulations, 1996

• (b) a serum agglutination test for brucellosis, carried out in accordance with the procedure laid down in Annex C to Council Directive 64/432/EEC, giving a count lower than 30 IU of agglutination per mililitre, or a complement fixation test showing a brucella count lower than 20 EEC units per mililitre (20 ICFT units)

• (c) a screening test for enzootic bovine leucosis, carried out in accordance with the procedure described in Annex G to Council Directive 64/432/EEC, with a negative result;

• (d) for infectious bovine rhinotracheitis/infectious pustular vulvo-vaginitis, a serum neutralization test or an Elisa test with a negative result. However,

— it is not necessary to carry out these tests on bulls which have already been subjected to such tests and have given a positive result to the serological test carried out in accordance with Directive 88/407/EEC as amended,

— vaccination against these diseases may be practised on sero-negative bulls, either with one dose of a temperature-sensitive live vaccine administered intranasally or two doses of an inactivated vaccine separated by an interval of not less than three weeks and not more than 4 weeks; the vaccination must be repeated subsequently at intervals of not more than 6 months.

• (e) either an immunofluorescent antibody test or a culture test for campylobacter foetus infection on a sample of preputial material or artificial vaginal washings; in the case of female animals a vaginal mucus agglutination test shall be earned out.

However, bulls which are not used for the production of semen may be exempt from the antibody test or a culture test for campylobacter foetus infection, with the proviso that such bulls may not be re-admitted to semen production until they have been subjected to such a test or culture and given a negative result.

Semen collected from all other animals at the centre since the date when the positive test was carried out shall be held in separate storage and may not be the subject of intra-Community trade until the health status of the centre has been re-established.

These provisions shall not apply to sero-positive bulls which, prior to their first vaccination in accordance with Council Directive 88/407/EEC (as amended), at the insemination centre, gave a negative reaction to the serum neutralization test or the ELISA test for infectious bovine rhinotracheitis or infectious pustular vulvo-vaginitis;

Sero-positive bulls referred to in the second subparagraph of Article 4(1) of Council Directive 88/407/EEC must be isolated, since their semen may be the subject of intra-Community trade in accordance with the provisions for trade in semen from such bulls.

Regulation 12

SCHEDULE H

CONDITIONS FOR THE APPROVAL OF AN OVA/EMBRYO COLLECTION AND OVA/EMBRYO PRODUCTION TEAMS

In order to be granted approval each embryo collection team shall fulfil the following conditions:

• (a) The collection, processing and storage of embryos shall be carried out either by a team veterinarian or under his responsibility by one or more technicians who are competent and trained by the team veterinarian in methods and techniques of hygiene.

• (b) The embryo collection team shall be placed under the general supervision and authority of an official veterinarian.

• (c) The embryo collection team shall have at its disposal permanent or mobile laboratory facilities where embryos can be examined, processed and packed, consisting of at least a work surface, a microscope and cryogenic equipment.

• (d) In the case of a permanently sited laboratory, the embryo collection team shall have at its disposal—–

• (i) a room where embryos can be manipulated which is adjacent to but physically separate from the area used to handle the donor animals during collection, and

• (ii) a room or area equipped for cleansing and sterilizing instruments and equipment used in embryo collection and manipulation.

• (iii) Where micromanipulation of the embryo which involves penetration of the zona pellucida is to be carried out, this shall be done in suitable laminar-flow facilities which shall be properly cleaned and disinfected between batches.

• (e) The embryo collection team shall have at its disposal in the case of a mobile laboratory a specially equipped part of the vehicle consisting of two separate sections–

• (i) one for the examination and manipulation of embryos which shall be a clean section, and

• (ii) the other for accommodating equipment and materials used in contact with the donor animals.

A mobile laboratory shall always have contact with a permanently sited laboratory to ensure the sterilization of its equipment and the provision of fluids and other products necessary for the collection and manipulation of embryos.

• (f) To be approved as a team for the production and processing of embryos derived by in vitro fertilization and/or in-vitro culture, an embryo production team must fulfil the following additional requirements:

• (i) the personal shall be trained in appropriate disease control and laboratory techniques, particularly in procedures for working in sterile conditions;

• (ii) it shall have at its disposal a permanently-sited processing laboratory which must:

— have adequate equipment and facilities, including a separate room for recovering oocytes from ovaries and separate rooms or areas for processing oocytes and embryos, and storing embryos and

— have laminar-flow facilities under which all oocytes, semen and embryos must be processed; however, the centrifugation of semen may be carried out outside the laminar-flow facility, as long as full hygienic precautions are taken;

• (iii) where oocytes and other tissues are to be collected in an abattoir, it must have at its disposal suitable equipment for the collection and transport of the ovaries and other tissues to the processing laboratory in a hygienic and safe manner.

• (g) It is the responsibility of the team veterinarian to ensure the welfare of the breeding female animals concerned. In particular embryos shall only be collected from breeding females to which suitable epidural anaesthesia have been administered which will have effect during the entire process of embryo collection.

Regulation 12

SCHEDULE I

CONDITIONS RELATING TO THE COLLECTION OR PRODUCTION, PROCESSING, STORAGE AND TRANSPORT OF OVA/EMBRYOS BY AN APPROVED EMBRYO

COLLECTION OR PRODUCTION TEAM

• (a) Embryos shall be collected and processed by an approved collection team without coming into contact with any other consignment of embryos not meeting the requirements of these Regulations.

• (b) Embryos shall be collected in a place which is isolated from other parts of the premises or holding and which shall be in good repair and easy to cleanse and disinfect.

• (c) Embryos shall be processed (examined, washed, treated and placed in identified and sterile containers) in either a permanent laboratory facility or a mobile laboratory facility, which is not situated in a zone subject to prohibition or quarantine measures.

• (d) All implements which come into contact with the embryos or the donor animal during collection and processing shall be disposable or shall be properly disinfected or sterilised prior to use.

• (e) Products of animal origin used during collection of the embryos and in the transport medium shall be obtained from sources which present no animal health risk or are to be so treated prior to use so that such risk is prevented. All media and solutions shall be sterilised by approved methods according to the recommendations of the manual of the International Embryo Transfer Society (IETS). Antibiotics may be added to the media in accordance with the IETS manual.

• (f) Storage flasks and transport flasks shall be properly disinfected or sterilized before the commencement of each filling operation.

• (g) The cryogenic agent used shall not have been previously used for other products of animal origin.

• (h) Each embryo container and the containers in which they are stored and transported shall be clearly code-marked in such a way that the date of collection of the embryos and the breed and identification of the donor sire and donor dam, as well as the registration number of the team can be readily established. The characteristics and form of this marking shall be in accordance with that specified by the Minister.

• (i) Each embryo shall be washed at least 10 times in a special fluid for embryos which shall be changed each time and which, unless decided otherwise, shall contain trypsin, in accordance with internationally recognized procedures. Each wash shall be a 100 -fold dilution of the previous wash and a sterile micropipette shall be used to transfer the embryo on each occasion.

• (j) After the last wash each embryo shall be subjected to microscopic examination at a magnification of at least x 50 over its entire surface to determine that the 'zona pellucida' is intact and is free from any adherent material. Any micro-manipulation which involves penetration of the zona pellucida shall be carried out in the facilities approved for the purpose, and after the last wash and examination. Such micromanipulation may only be carried out on an embryo having an intact zona pellucida.

• (k) Each consignment of embryos that has successfully undergone the examination provided for in subparagraph (j) shall be placed in a sterile container marked in accordance with sub paragraph (h) and which shall be sealed immediately.

• (l) Each embryo shall, where appropriate, be frozen as soon as possible and stored in a place which is under the control of the team veterinarian and which is subject to regular inspection by an official veterinarian.

• (m) All products in the processing, washing, transportation media must be obtained from sources which present no animal health risk or are so treated prior to use that such risk is prevented.

• (n) Each collection team shall submit to an approved laboratory specified by the Minister routine samples of flushing fluids, washing fluids, disintegrated embryos, non-fertilized ova etc, resulting from its activities for official examination for bacterial and viral contamination. The procedure for collecting of samples, conducting such examinations, together with the standards to be achieved shall be in accordance with that specified by the Minister. If the standards laid down are now achieved the Minister shall withdraw approval.

• (o) Each collection team shall keep a record of its activities in respect of embryo collection during the 12 months before and 12 months after storage including:

• (i) the breed, age and identification of the donor animals concerned,

• (ii) the place of collection, processing and storage of embryos collected by the team,

• (iii) the identification of the embryos together with details of their destination if known, and

• (iv) details of micromanipulation techniques which involve penetration of the zona pellucida or other techniques such as in vitro fertilization and/or in vitro culture which have been performed on the embryos. In the case of embryos derived by an in-vitro fertilization, the identification may be done on the basis of a batch, but must contain details of the date and place of collection of ovaries and/or oocytes. It must also be possible to identify the herd of origin of the donor animals.

• (p) The conditions laid down in paragraphs (a) to (o) shall apply as appropriate to the collection, processing, storage and transport of ovaries, oocytes and other tissues for use in in vitro fertilization and/or in vitro culture. Furthermore, the following additional conditions shall apply:

• (i) when ovaries and other tissues are to be collected at an abattoir, the abattoir should be officially approved and under the control of an official veterinarian whose responsibility it is to carry out ante and post mortem inspection of donors;

• (ii) materials and equipment coming into direct contact with ovaries and other tissues shall be sterilized before use and after sterilization, used exclusively for those purposes. Separate equipment shall be used to handle oocytes and embryos from different batches of donor animals;

• (iii) ovaries and other tissues shall not be allowed to enter the processing laboratory until completion of the post mortem inspection of the batch. If relevant disease is found in the batch of donors, or in any animals slaughtered in that abattoir on that day, all tissues from that batch must be traced and discarded;

• (iv) the washing and examination procedure laid down in subparagraphs (i) and (j) shall be carried out after the culture procedure has been completed;

• (v) any micromanipulation which involves penetration of the zona pellucida shall be carried out in accordance with the provisions of subparagraph (i), after the procedures laid down in subparagraph (s) have been completed;

• (vi) only embryos from the same batch of donors should be stored in the same ampoule/straw.

Each embryo collection or production team shall ensure that the embryos are stored at suitable temperatures in premises approved for the purpose by the Minister.

In order to be approved these premises shall:

• (i) comprise at least one lockable room intended exclusively for embryo storage;

• (ii) be easy to cleanse and disinfect;

• (iii) have permanent records of all incoming and outgoing movements of embryos and the final destination of the embryos in particular shall be specified in such records;

• (iv) be subject to inspection by the official veterinarian.

The competent authority may authorise the storage of semen that fulfils the requirements of Directive 88/407/EEC in the approved storage premises.

Embryos for trade shall be transported in hygienic conditions in sealed containers from the approved storage premises until their arrival at their destination.

The containers shall be marked in such a way that the number coincides with the number on the animal health certificate.

Regulation 12

SCHEDULE J

CONDITIONS APPLYING TO DONOR ANIMALS

• (a) they shall have spent at least the previous 6 months within the territory of the European Union or in the third country of collection;

• (b) they shall have been present in the herd of origin for at least 30 days prior to collection;

• (c) they shall come from herds which are:

— officially tuberculosis free

— officially brucellosis free or brucellosis free

— enzootic bovine leucosis free

in derogation from the third indent, they may come from a herd which is not leucosis free but for which certification has been obtained that there has not been any clinical case of enzootic bovine leucosis during the past 3 years;

• (d) during the previous year, they shall not have been present in a herd which has shown any clinical sign of infectious bovine rhinotracheitis/infectious pustular vulvovaginitis.

• (a) shall be kept in a holding which is not subject to veterinary prohibition or quarantine measures;

• (b) shall show no clinical signs of disease.

Regulation 20

SCHEDULE K

ZOOTECHNICAL CERTIFICATE

For trade in pure-bred breeding animals of the bovine species

The results of performance tests and results with origin of the assessment of the genetic value, on the animal itself and its parents and grandparents.

Known hereditary defects in the donor animal or its ancestors shall be indicated. (State none if applicable).

SCHEDULE K (contd)

An alternative may be provided in documentation accompanying the pure-bred breeding animal. In this event the competent authorities of the exporting country shall certify that the particulars required under paragraph 1 above are indicated in those documents, by the following formula:

"The undersigned certifies that these documents contain the particulars mentioned in Article 1 of Commission Decision 86/404/EEC."

Regulation 20

SCHEDULE L

ZOOTECHNICAL CERTIFICATE

For trade in the semen of pure bred breeding animals

The updated results of performance tests and updated results with origin of the assessment of the genetic value, on the animal itself and its parents and grandparents.

Known hereditary defects in the donor animal or its ancestors shall be indicated. (State none if applicable).

A.

B.

Origin of semen:

SCHEDULE L (contd)

An alternative certificate may be provided in documentation accompanying the semen. In this event the competent authorities of the exporting country shall certify that the particulars required under Paragraph 1 are indicated in those documents, by the following formula:

"The undersigned certifies that these documents contain the particulars mentioned in Article 1 of Commission Decision 88/124/EEC".

Regulation 20

SCHEDULE M

ZOOTECHNICAL CERTIFICATE

For trade in embryos of pure bred breeding animals

A. Particulars on donor bull:

The updated results of performance tests and updated results with origin of the assessment of the genetic value, on the donor bull itself and its parents and grandparents.

SCHEDULE M (contd)

B. Particulars on donor cow: