The first stage of control is carried out directly on the farms using rapid screening tests carried out by the farmer or by the driver of the tanker truck before milk is pumped to the tanker and transported to the dairy. However, not all farmers or tanker truck drivers check freshly milked milk directly on the farm. From the dairy's point of view, this level of control directly at the farmer is interesting because it eliminates the problem of contamination of milk from other farmers in the truck, and also it eliminates the transfer of risk and potential problems with the presence of veterinary drug residues to the next stage of the food chain. Food business operators carry out this control for their own money to eliminate the risk of supplying milk containing antibiotic residues. The purchase of rapid screening tests is provided directly by farmers or by dairies and distributed to farmers or milk tankers drivers.

The second stage of control takes place at the entrance to the dairy as a self-verification check. Following EC Regulation No 852/2004 (EC, 2004a), the established HACCP system and monitoring for the presence of inhibitory substances are performed routinely during milk intake. Each dairy has a customized monitoring system. In most cases, the laboratory worker will firstly take samples from individual farms sampled by the truck driver or autosampler. Also, it is necessary to take the “tanker sample” which contains milk from all farmers. Usually, this sample is sampled automatically on farms if the autosampler is used. Milk from all farmers is collected in one big bottle. In the case of manual sampling, laboratory workers need to take the sample from the tanker cistern or each tanker cistern chamber. This can be made following ISO 707 (ISO, 2008). Sometimes this tanker sample is sampled by dripping milk from the pipe valve leading to the receiving tank. Milk is waiting in the truck tanker or the designated intake tanker and can not be processed until is analyzed for drug residues presence. The sample is examined by a rapid 5 – 10 minute test (e.g. BetaStar, BetaStar Combo, ROSA Charm 3 SL3, ROSA Charm SLB, Twinsensor BT, SNAP BT ST test, etc.).

The sensitivity of rapid tests is either at the level of maximum residue limits or even higher (Zajác et al., 2006, 2007a, 2007b) and is suitable for milk testing by dairies.

If the residues of inhibitory substances are detected, then all samples from individual farms are tested. This is how the farm that caused the contamination is found. These positive samples are tested by the dairy only with a long 3-hour tests Delvotest^® SP-NT (MARD SR, 2006), Eclipse 50 (MARD SR, 2009a), or Kalidos TB MP (MARD SR, 2009b). The dairy reports this situation to the appropriate Regional Veterinary and Food Administration, but as this is a selfverification check, these results are not statistically evaluated by this institution.

Some dairies do not perform a long 3-hours test but they send positive samples directly to the accredited laboratory for confirmation, or they automatically consider the milk to be contaminated and therefore intended only for disposal.

The damage must be paid by the farmer who caused such contamination. Usually, the incurred damage is resolved as an insured event.

In Slovakia or European Union, the legislation does not specify which laboratory should test samples (EC, 2006). Even the legislation does not require accreditation of this laboratory! In practice, this situation can be handled directly by the legislation of the country or by the supplier-customer contract. In Slovakia, it is usually tested in one of the two accredited central testing laboratories specifically intended for milk analysis or in the National Reference Laboratory for Milk and Milk Products in Lužianky or by the National Reference Laboratory for Antibacterial Substances and Other Pharmacologically Active Substances Dyes and Some Prohibited Substances in Dolný Kubín (MARD, 2020).

In practice, dairies usually use the services of accredited laboratories for confirmation of positive samples only rarely. They usually immediately consider the sample to be positive. This is how it is solved in most supplier-customer contracts. Confirmation of positive samples can be performed in a contracted accredited central testing laboratory or one of the above-mentioned national reference laboratories. When the accredited laboratory confirms the positivity of the sample, then milk is legally disposed of by the authorized company under a decision of the Regional Veterinary and Food Administration.

Some dairies in Slovakia have recently started sending milk samples for analysis of milk composition, total bacterial count, somatic cells count, and drug residues to laboratories in Hungary, Austria, Germany, or the Czech Republic. The results from these laboratories are then not statistically processed by National Reference Laboratory for Milk and Milk Products. Only the results from central testing laboratories in Slovakia are processed by NRL.

The dairy reports the presence of a positive case of residues of inhibitory substances to the Regional Veterinary and Food Administration, which will carry out an inspection at the farmer who caused the contamination.

After the correction, the farmer will request a Regional Veterinary and Food Administration for sampling the milk, which will then be examined in an accredited testing laboratory. Only after this analysis can the farmer deliver the milk to the dairy again.

The third stage of control is performed in central testing laboratories or any other laboratory. Dairies in Slovakia are usually sent milk samples for the inhibitory drug residues substances test to the contracted one central testing laboratory. The frequency of testing is not prescribed in European legislation (EC, 2006). Usually, two samples per month are collected from each farm. This testing is performed as a part of the verification process based on the HACCP plans. Also, the frequency of testing is mentioned in the supplier-purchaser contract. The testing is generally carried out in conjunction with the examination of the freezing point of the milk because it is recommended to analyze not preserved milk samples. On the other hand, for some tests, preserved samples may be used.

Persons taking milk samples for payment purposes, whether drivers or directly the purchaser's staff, must be trained by the Veterinary and Food Institute, National Reference Laboratory for Milk and Milk Products. These persons periodically undergo an accredited educational activity every five years and hold a “milk sampler's certificate”, which entitles them to take milk samples. In principle, such samples are taken in duplicate. One sample remains on the farm and the other is used by a dairy company. The sampling plan created by the dairy for payment must be secret. Farmers must not know the date of this sampling. Sampler notifies the farmer immediately when this sample is collected. All samples are sent to a contracted laboratory (usually a central testing laboratory).

If residues of inhibitory substances are detected in the second or third stage of control, the dairy shall trace the source of the contamination to identify the farm which caused the contamination.

The farm that caused the contamination will then provide financial compensation for milk and costs associated with the legal disposal of the contaminated milk to all farmers affected by this contamination. From a technical point of view, single-chamber or multi-chamber tanks can be used to transport milk. In most cases, the milk is contaminated in the relevant chamber of the entire milk tank into which the milk is pumped is contaminated. In some cases, contamination of several chambers in the milk tank may occur if the capacity of the currently filled chamber is insufficient or by cross-contamination from residues of contaminated milk left in the hoses and distribution system of the milk tank truck. In the case of using a multi-chamber tank, it is, therefore, possible to save part of the milk supply, since the positive milk is usually only found in one of the chambers of the tank.

In the case of confirmation of a "positive" result for the presence of antibiotic residues, the dairy plant shall carry out the following:

According to the Decree of the Ministry of Industry and Trade of the Slovak Republic No. 148/2012 (MIT SR, 2012), such disposal must be carried out only under the official supervision of a veterinary inspector with his consent. According to EC Regulation no. 142/2011 (EC, 2011), the removal of milk by the wastewater stream is prohibited.

Depending on how the buyer has stated in the supplier-customer contract with the farmer, he can ensure that another confirmatory test is performed, which identifies and quantifies the antibiotic and proves whether or not the maximum residue limit for this particular antibiotic is exceeded. These confirmation tests are usually performed in accredited testing laboratory: Veterinary and Food Institute Dolný Kubín, National Reference Laboratory for Antibacterial Substances and Other Pharmacologically Active Substances, Dyes and Certain Prohibited Substances.

If the maximum residue limit is exceeded, the milk and milk products are disposed of legally. If the maximum residue limit is not exceeded, milk and products made from it are considered safe.

According to Kološta (2007), the biggest problem is mainly residues of antibiotics and sulfonamides, as these are mostly used in the treatment of dairy cows. The reference method for the detection of antibiotics and sulphonamides in raw milk and heat-treated milk is the CD 91/180 / EEC method on the principle of microbial inhibition. The mentioned method was also taken over into the system of Slovak technical standards STN 57 0531 Determination of antibiotics and sulfonamides in raw and heat-treated milk (STN, 2001).

The journals of the Ministry of Agriculture and Rural Development of the Slovak Republic state which laboratory methods can be used for the official control of the presence of inhibitory substances in milk. These are methods based on the principle of agar diffusion tests with a change in the color of the indicator due to the active metabolism of the test microorganism (Bacillus stearothermophilus var. Calidolactis) in the absence of an inhibitor. The determination of residues of inhibitory substances in milk and milk products can therefore be carried out in central testing laboratories by the following methods approved by the Ministry of Agriculture and Rural Development of the Slovak Republic:

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When testing raw cow's milk for payment, central testing laboratories in the Slovak Republic (MILEX Progres, as Bratislava and EXAMINALA – Dairy Research Institute in Žilina) use a diffusion test to determine residues of inhibitory substances, which lasts 3 hours: Delvotest^® SP-NT. This test is used also by the National Reference Laboratory for Milk and Milk Products in Nitra (NRLM) (Zajác and Čapla, 2010).

However, it is not possible to determine the exact amount of antibiotic residues or a specific species using these microbiological or antigen methods.

Veterinary and Food Institute Dolny Kubin, National Reference Laboratory for Antibacterial Agents and Other Pharmacologically Active Substances, Dyes and Certain Prohibited Substances quantify veterinary drug residues by HPLC/MC method (SVFI, 2021).

The State Veterinary and Food Administration of the Slovak Republic annually compiles the National Plan for the Control of Residues in Live Animals and Products of Animal Origin following the applicable legislation of the EU (EC, 2017; EC, 2019). The aim of the control through this plan is to obtain an overview of the current state of contamination of individual components of the food chain, to implement effective measures to eliminate the identified deficiencies and prevent their further occurrence. Longterm monitoring of the presence of inhibitory substances in milk leads to the finding that almost 95% of cases of secondary contamination with inhibitory substances are antibiotic contamination, approximately 2.5% of contamination is due to other drugs, especially sulfonamides and residual detergent and disinfectant residues, 1% may cause mycotoxins from feed (nuclear, bulk), 1% may be caused by agrochemical residues or some preservatives and then only about 0.5% remains on natural inhibitors in milk (Kološta, 2007).

Commission Regulation (EC) No. 853/2004 (EC, 2004b) contains specific hygiene rules for food of animal origin.

Commission Regulation (EC) No 1662/2006 (EC, 2006) defines health requirements for the production of raw milk. Raw milk must come from animals to which no unauthorized substances have been administered and the products (medicines or substances) administered have complied with the withdrawal periods prescribed for those products. This Regulation also defines the requirements for milking, transport, and transport hygiene. According to the Regulation, it is necessary to identify animals that have undergone treatment which could cause the excretion of drug residues in milk and colostrum, and that milk and colostrum obtained from such animals should not be used for human consumption before the expiry of the prescribed withdrawal period. Dips or sprays pods will be used only after authorization or registration. Based on these requirements, primary milk producers must put in place systems to control the treatment of dairy cows.

In the past, the State Veterinary and Food Administration published a guideline for control of raw milk in farms for the production of raw milk (SVFA, 2009), today the control is based on the requirements of the current European legislation (EC, 2017; EC, 2019), Title IV, Article 49 Control of milk and colostrum production holdings and Article 50 Control of milk and colostrum.

The Regional Veterinary and Food Administration checks the food business operator whether, according to EC Regulation no. 1662/2006 (EC, 2006) has procedures in place to ensure that raw milk is not placed on the market if it contains antibiotic residues in quantities which, for any of the substances listed in Annexes I and III to Regulation (EEC) No. 2377/90 (EC, 1990) exceeds the value permitted under that Regulation or if the combined total antibiotic residue level exceeds any maximum permitted value.

During the official control, the inspector fills in the prescribed official control record, in which he states the number of reports of positive cases of residues of inhibitory substances in raw milk. It also checks whether the supplier and the relevant regional veterinary and food administration of the Slovak Republic have been informed about unsatisfactory results of raw milk. In case of unsatisfactory results of raw milk, the inspector checks the documentation, the date of detection (notification from the Central testing laboratory, the State Veterinary, and Food Institute, the own laboratory, or any laboratory), the date of written notification to the relevant Regional Veterinary and Food Administration and the raw milk supplier.

The food business operator must therefore take measures to remedy this situation and should not place milk containing residues of inhibitory substances on the market. In the case of residues of inhibitory substances, the legislation does not further specify what measures the food business operator has to take. The legislation does not specify how the control body should proceed, nor does it specify whether samples are to be taken to prove remediation and where (what laboratory) these are to be tested. However, according to the methodological guideline (SVFA SR, 2009), inspectors of Regional Veterinary and Food Administrations are instructed as follows: the producer of raw milk must start procedures to ensure that raw milk is not placed on the market, i.e. unless corrective measures have been taken and it should be proved that raw milk does not exceed the maximum permitted level of the combined total antibiotic residues. Food business operators must not place such milk on the market. In practice, this means that after the situation has been remedied, the farmer must have a sample of milk examined at his own expense in the laboratory (it is not defined what kind of laboratory, but usually it is agreed with the inspector what laboratory should be used for this testing. Almost it is an accredited central testing laboratory or any accredited laboratory of the Veterinary and Food Institute – NRL for milk and milk products or NRL for Antibacterial Substances). If the result of the test shows that the milk does not contain antibiotic residues, it may place it on the market.

Control of antibiotic residues is necessary to protect the reputation of milk as safe food and to prevent exposure of the consumers to drug residues. Governmental authorities which are obliged to organize supervision and control systems by state supervision agencies must provide these requirements (Popelka, et al., 2003).

EU countries must monitor food of animal origin for the presence of residues. The legislation governing the design and implementation of residue monitoring plans is as follows (EC, 2021):

Specific EU legislation outlines the laboratory analysis and correct interpretation of results:

Commission Decision 97/747/EC lays down that the annual number of samples taken should be one per 15,000 tonnes of annual milk production, with a minimum of 300 samples (EFSA, 2019).

The overview of the non-compliant results for prohibited substances is present in Table 2. Chloramphenicol is the most dominant prohibited substance detected in milk in European Union.

The specific species of residues detected are listed in Table 3. There are several specific groups of residues according to the EU legislation: A (1-6), B (1-3). European Union member states report all positive samples from the monitoring of veterinary medicinal product residues and other substances in live animals and animal products including milk (EFSA, 2021).

The Slovak Republic has performed the monitoring and the targeted sampling according to the EU legislation. The number of positive samples was:

Positive samples present in Table 1 are not included in these reports (Table 2), because they were detected in the self-control by dairies and were not sampled as a part of the targeted monitoring performed by the Veterinary and Food Administration of the Slovak Republic.

The monitoring system in Slovakia is functional and will detect possible contamination of milk with residues of veterinary drugs. Dairies in Slovakia process only milk that does not contain residues of veterinary drugs.