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EN 1656

Evaluation of Bactericidal activity of Chemical Disinfectant

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BS EN 1656 : 2019 – Quantitative suspension test for evaluating the bactericidal activity of chemical disinfectants and antiseptics used in the veterinary area (phase 2, step 1)

EN 1656 is a phase 2, step 1 quantitative suspension test, which lays down the practical conditions and testing procedures for evaluating the efficacy of disinfectants and antiseptic products intended to be used in veterinary settings. The guideline specifies EN 1656 test to be conducted against different kinds of bacteria.

EN 1656 Test Conditions & Requirements

EN 1656 (phase 2, step 1 suspension test)

General disinfection products

Teat disinfection products

Mandatory test microorganisms

  • Enterococcus hirae (ATCC 10541)
  • Proteus hauseri (ATCC 13315)
  • Pseudomonas aeruginosa (ATCC 15442)
  • Staphylococcus aureus (ATCC 6538P)
  • Escherichia coli (ATCC 8739)
  • Staphylococcus aureus (ATCC 6538P)
  • Streptococcus uberis (ATCC 19436)

Test temperature

Between 5 °C to 40 °C

Between 20 °C to 30 °C

Contact time

30 sec to 5 min (interval of 30 sec) and

5 min to 120 min (interval of 5 min) 

For pre-milking teat disinfectants – Between 30 seconds and 3 minutes 

For post-milking teat disinfectants Between 1 minute to 30 minutes

Interfering substance 

Low level (3.0g/l bovine serum albumin) 

High level (10g/l bovine serum albumin + 10g/l yeast extract)

Pre milking disinfection (3.0g/l Bovine serum albumin)

Post milking disinfection (10g/l skimmed milk powder)

Log reduction



Scope of products that can be tested with EN 1656 guideline

EN 1656 test method is applicable for different category of disinfectants and antiseptics that are meant to be used in different veterinary field such as –

  • Animal breeding, husbandry, and production, 
  • Animal care facilities,
  • transport and disposal of all animals except when in the food chain following death and entry into the processing industry,

EN 1656 standard is also employed to test products used for teat disinfection of the milch animals.

EN 1656 Test Process

EN 1656 Test Method

  • Test product is added to the suspension mixture of test microorganisms and interfering substance. As per standard, the maximum concentration of the product tested is 80%, dilution of the test product is due to the interfering substances and test microorganism in the reaction mixture.
  • The test mixture (disinfectant + microorganism + interfering substance) is kept at a particular temperature and contact time as per the recommendation by the client and EN 1656 guideline.
  • After contact time, the test mixture is neutralized using a proper neutralizing solution to stop the reaction.
  • The recovered bacteria in the neutralizing medium is then enumerated by serial dilution and plating on a proper growth medium. After neutralization, mixture is further incubated for 2 days to check if any viable microorganisms are present.
  • After 2 days of incubation of the culture plates, bacterial colonies are counted and antimicrobial potency of the test product is determined.

Importance of EN 1656

Veterinary sites if not kept clean can potentially turn into breeding ground for highly pathogenic microbes. These microbes have the potential to cause diseases to both humans and animals, which can result in a huge financial loss. Hence, it is crucial to monitor cleanliness and implement appropriate hygiene practices in veterinary settings. 

EN 1656 is one of the widely accepted European standards that evaluates the efficacy of disinfectant and antiseptics intended to be used in the Veterinary settings. This standard thus enables checking of a proposed product to meet the necessary efficacy and safety criteria required to control different critical pathogens found in the veterinary area.

At MIS, we conduct EN 1656 test customized to meet the specific product requirements of our clients. In an effort to help clients with development of high performance antimicrobial products, we have implemented a  robust system that tests for quality and precision.

In addition to efficacy testing of veterinary disinfectants, we offer EN 1276 testing for disinfectants intended for use in food, industrial, domestic and institutional areas.

To request a quote for EN 1656 testing, contact us now to get consultation from our microbiology experts.


EN 1656 test is carried out to measure the bactericidal activity of chemical disinfectants and antiseptics utilized in veterinary areas. This test assesses the effectiveness of the sample product in eliminating mandatory pathogens described as critical in EN 1656 guideline.

EN 1656 is carried out for chemical disinfectants and antiseptic products used for general disinfection in veterinary areas. This test is also applicable for teat disinfectant products.

EN 1656 test usually takes 2 to 3 weeks to complete.

At Microbe Investigations, we test for EN 1656 using the following microbial strains: Enterococcus hirae (ATCC 10541), Proteus hauseri (ATCC 13315), Pseudomonas aeruginosa (ATCC 15442), Staphylococcus aureus (ATCC 6538P), Escherichia coli (ATCC 8739) and Streptococcus uberis (ATCC 19436).

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