| NMKL newsletter | |
| Nr. 43/00, March 2000 | |
New method
Methods withdrawn
Seminar concerning Genetically Modified
Foodstuffs
Questions and comments regarding the Listeria
monocytogenes method
Questions regarding the Salmonella
method
Rising of the prices
NMKL methods in Finnish recently published
New NMKL method:
NMKL-method No 144, Enterobacteriaceae Determination in foods and feeds, has been revised
.This method is intended for use in examinations of foods and
feeds. Known amounts of dilutions of foods are mixed with violet
red-bile-glucose agar. Bacteria belonging to the family Enterobacteriaceae
form pink to red colonies with or without a precipitation zone. After incubation
at 37oC for 22-26 hours a representative selection of colonies are
confirmed by the oxidase test.
In 1994 the National Food Administration, Sweden arranged a collaborative study
on this method (National Food Administration report No 6/94). The result was not
satisfactory due to several factors. In the method it was indicated that the
pink to red colonies with diameter >0.5 mm should be counted. It was
difficult to determine the size of the colony, especially when they were as
small as 0.5 mm. The readings therefore became subjective. In addition the
variations in incubation time and temperature influenced the size of the colony.
In the revised method the incubation time is specified to be 24±
2 hours, and the limit of the size of the colony is removed. All the pink to red
colonies are to be counted.
Per Norberg
NMKL gives thanks to Per Norberg and the National Food Administration, Sweden, for elaboration of this method.
The following NMKL methods are withdrawn, and therefore requested to be removed from your NMKL collection:
No. 1, 1950: Boric acid. Determination in foods.
No. 72, 1970: Biphenyl. Determination in citrus fruits, marmalade of citrus fruits, and in apples and pears.
No. 19, 1955: Fluorine. Determination in calcium compounds and cereals with added calcium compounds.
No. 78, 1970: Lactose. Determination in milk chocolate.
No. 73, 1970: o-Phenylphenol. Determination in citrus fruits, marmalade of citrus fruits, and in apples and pears.
Questions and comments regarding
the Listeria monocytogenes method
Questions and comments regarding method No 136, 2nd ed. 1999, are quoted below since it might be of general interest
.- In the method, the plates should be examined after 24± 3 hours and then after 48± 4 hours (as requested in NMKL method no. 66, 1999). In accordance with accreditation and prospective documentation requirements, it would be an advantage if time limits were only quoted in cases when requested for professional reasons. Could the examination of the plates after 24 hours be considered as a support to the examination after 48± 4 hours, and if so would ± 3 hours in respect to the 24 hours be considered as unnecessary?
The referee
Birgit Nørrung, Danish Veterinary and Food Administration answers: Your notion
is correct. In NMKL methods the time specifications are harmonised at different
temperature specifications, that is the reason for ±
3. However, this should not cause any problem.
Some laboratories have experienced that it is helpful to replace one of the sub-cultivations of well-isolated colonies with a streaking on blood agar from the most densely populated area on the selective plate. This will make it easier to detect a low number of haemolytic Listeria spp. among a high background of non-haemolytic Listeria spp. Other laboratories have found it helpful to inoculate a blood agar plate in addition to the selective plates at the first isolation step. In this case, it will also be easy to detect haemolytic colonies among a high number of non-haemolytic colonies
.The prices have not been raised since 1993. The estimated incomes of this rising will be used to stimulate the work within elaboration of methods.
From 1st of March NMKL will operate with the following prices (in NOK):
|
|
NOK |
| Subscription fee (annually) |
500 |
|
Compiled collection of NMKL methods |
1800 |
|
Method (single) |
150 |
|
Ringbinder (for each) |
40 |
| Report (except number 8 that costs NOK 250,-) |
150 |
| Report nr. 8 |
250 |
| Procedure |
200 |
Questions regarding the Salmonella method
NMKL has received questions about NMKL method No 71, 5th Ed., 1999. The questions, which are answered below by the referee, Christer Wiberg, National Food Administration, Sweden, are quoted here since others might have the same questions.
There are quote instances of support in the literature that 18±2 hours give the optimal result. After about 20 hours a dramatic pH decrease might occur in some samples, which may result in fewer positive samples. In my opinion the correct incubation temperature is 18±2 hours. 21±3 hours was prescribed due to a compromise between the national committees for making it possible for a laboratory to pre-incubate samples from one morning to the next. The final remark is that you should always choose 18±2 hours if possible. This time is also in accordance with several international methods, as ISO for instance.
This question can be answered similarly to the previous question above. Technically, the extension to 48 hours does not give a less accurate result in respect of the frequency of positive samples, some trials even show that you get more positive samples after 48 hours than you do after 24 hours. Practically, you want the results as quickly as possible. The main reason for the 48 hours alternative is due to making it possible to start the analysis on Friday and incubate the enrichment broth from Saturday until Monday morning, avoiding work on Sundays.
Actually, section 6.2 in the method gives the answer to this question. However, I strongly recommend use of a water-bath. If I did not have to compromise on this method the alternative with an incubator would not have been included. My notion is based on concrete examinations of temperature variations over time and the incubator environment
. I have also an extensive practical experience from own research and from our laboratory’s activities in arranging performance studies that shows that use of water-bath in comparison with incubator generally gives more positive results. The optimal enrichment temperature is around 42.0°C, and some serotypes are sensitive to enrichment temperatures about 42.5°C. The best incubators have a variation of temperature within 1°C while the temperature variation in water-baths (with stirring) can hardly be measured.In the pre-incubation step 18± 2 hours was used and 24±3 hours was used in
the pre enrichment with RVS-broth.
Follow the method, subculture on a suitable non-selective agar plate and perform all confirmations with polyvalent antiserum from this plate. We talk about polyvalent H-antiserum that contains a mixture of antiserum against the common excisting
H-anti genes. Nor polyvalent O- or H-antiserum gives a 100% confidence, but using both antisera increases the confidence. There exist serotypes that give O+, H- or O-, H+ when testing with these polyvalent antisera.
Seminar concerning Genetically Modified Foodstuffs
The seminar will be presented in English.
In connection with NMKL’s Annual Meeting 2000 at Bornholm, Denmark, a seminar concerning Genetically Modified Foodstuffs will be arranged. The seminar will be held at Hotel Fredensborg, Friday 25th of August 10am – 6pm.
The seminar will provide a wide overview of the topic, with attention to regulatory aspects and risk evaluation of genetically modified foodstuffs. The main topic during the seminar will be qualitative and quantitative methods for detection of Genetically Modified Organisms (GMO), with mutual updating on status and experiences within the Nordic countries. As a closure there will be a discussion of future perspectives and prospective needs for a Nordic co-operation on this matter.
The seminar will focus on the modified organisms and the extension of GMO used, the associated changes in plants, as well as on the ratio of GM- and non-GM acreage for each of the crops, growing conditions the genetically modified plants require in comparison to non-genetically modified plants and any unintended effects that may occur with gene splicing.
Registration for the seminar can be forwarded (also in connection with registration to NMKL’s Annual Meeting) no later that 1st of June to the secretary of the Danish National Committee, Inge Meyland, Danish Veterinary and Food Administration, Mørkhøj Bygade 19, DK-2860 Søborg. E-mail: ime@fdir.dk. The participation fee is not yet settled.
The program of the seminar will be available at the beginning of April. The program will be distributed to the national committees of NMKL and can also be requested from Jens Kirk Andersen, Danish Veterinary and Food Administration, Mørkhøj Bygade 19, DK-2860 Søborg. E-mail: jka@fdir.dk.
You will find the program here.
NMKL methods in Finnish recently published:
Nro. 136, 2. painos, 1999 Listeria monocytogenes. Osoittaminen elintarvikkeista.
Nro. 66, 3. painos, 1999 Koagulaasipositiiviset stafylokokit. Määrittäminen elintarvikkeista.