SOME COMMENTS TO CONSIDER IN THE TRAINING OF NEW USERS OF ROTH BIOSCIENCE LLC R-CARD MEDIA FOR “WATER WATCHERS”
by Jonathan N. Roth Ph.D. & C.E.O. , Roth Bioscience, LLC
There are three main types of procedures that may be used, depending on the properties of the water being tested. Generally speaking, natural water sources may contain Coliform bacteria from fecal sources (human, four legged animals (farmer’s livestock, native four legged non-domesticated animals, and birds.), Coliform (non-fecal) from soil and other natural sources, and other types (many disease causing) e.g.-Salmonella, Pseudomonas, Listeria, etc.}
Potable (safe to drink)—no fecal bacteria (USEPA general government standard specifies Escherichia coli)
Low levels of bacteria (e.g.—less than 2 bacteria/mL -- = 50-100/100 mL sample)
Medium levels—2-5 bacteria/mL (200-500/100mL sample)
High levels (TNTC= too numerous to count)
The official categories: Each of the above is determined and regulated by the federal agency (USEPA) and lesser agencies (local –state). Most areas have set limits for whole body contact (swimming. Water skiing) and/or other water activities.) A whole body contact level is commonly 235 E. coli/100 mL of freshwater. Most public beaches test their bacteria levels every day and if levels exceed set standards, the beach is closed for the day. (It is a great advantage to have Rapid Methods that can provide results in less than 24 hr. from the time of sampling to accurate counts. ROTH BIOSCIENCE R-CARD® METHODS CAN GIVE FECAL COLIFORM (E. COLI FOR FRESH WATER AS EARLY AS 15-16 HRS WITH THEIR R-CARD ECC MEDIA, AND AS EARLY AS 8 HRS WITH THEIR USEPA approved KWIK COUNT® MF METHOD.
Enterococcus faecalis is favored as the fecal coliform of test choice by some who are testing saltwater beaches as it is thought that they live longer in an active state than E. coli. This may be true, but they generally require a significantly longer incubation period to give readable, accurate results than does E. coli.
Another continuing debate pertains to the method of using the different test methods and the handling of the water to be collected and tested for the accuracy and survivability of the bacteria that it holds.
- Until the recent advent of methods, such as the R-CARD®, water samples were collected in clean containers in the field packed on ice in insulated containers and shipped to labs where the counts were made. This may involve days of time and some have questioned whether the reported results are similar to the original samples.
- With the advent of the R-CARD®, it became easy to collect a fresh sample at its source, put it on ice and within a very short time have it in a protected lab and have an inoculated CARD with virtually no time for any significant bacterial population changes. However this obvious improvement on sample handling did not satisfy some persons, who were certain that the results could not match the collection and plating the samples “on site” by transferring the test water directly from the lake or river onto the R-CARD® itself in the field. (Despite the fact of potential rain, wind, dust and other factors being potentially present.) It also overlooked whether there were multiple samples taken from one single container or if each was collected separately for each CARD.
A should be noted that recent rains with their runoff will change a water population rapidly and make a big change in the type of bacterial population.
It should also be noted that the sending of iced samples has been the accepted standard protocol since the advent of commercial test laboratories without much rigorous testing to verify the method. The author of this note is not aware of any such comparative studies in the literature. It would appear that there was no need for such studies as there were no alternatives as the new rapid methods did not exist to provide any alternatives.
- In general, it is desirable to have a population of no less than 5-6 target organisms/CARD. There will likely be more accuracy in results if a 47 mm, 0.45µm micropore filter is used to filter 50-100 mL of the test inoculum and then placed right side up on the Card base and 1mL of sterile water added on its top prior to dropping the top piece onto it, and incubating. This procedure may be less convenient, but when very clean aqueous inoculum is used, the results will tend to be the most accurate.
Following are the results of a recent study, with results that compare “on site”, one hour post collection, 4 hour post collection, and 8 hour post collection results, with the same water sample being kept “on ice”, in a refrigerator (10 degrees Celsius) and at room temperature (about 25 degrees Celsius.)
River Test on R-CARD® ECC-A