• Industrial Dipslide Price Document.

Microorganisms are responsible for enormous economic losses to industry. Microorganisms and their metabolic products accumulate in water, transportation fuels, petroleum products and industrial fluids resulting in alteration of their physical and chemical properties. This can cause obstruction of fluid delivery lines or chemical deterioration in machinery resulting in dangerous situations as well as poor performance and costly repairs. This is particularly true in metal-working fluids and stored hydrocarbon fuels, but the problem also exists in many other industrial processes requiring cooling waters. Bacterial and fungal growth in these systems can be responsible for corrosion, slime formation, foul odors, and decreases in fluid function.

A large number of opportunistic microorganisms are engaged in these destructive and costly processes. Many types of water based coolants and lubricants, for example, are used in aluminum processing and other metal processing operations. These fluids are quite susceptible to bacterial and fungal contamination/degradation which can result in the production of poor quality rolled material. Genera found in the fluids included Pseudomonas, Bacillus, Corynebacterium, Rhodococcus and Staphylococcus. Growth of microorganisms in fuel tanks is another source of concern to industry. The biological content of the sludge found in naval fuel tanks in Halifax, for example, included fungal material (Hormoconois resinae formerly known as Cladosporium resinae), "white fungi", penicillium ssp., bacteria, sporulating fungal bodies, Candida, yellow fungi, small marine invertebrates, algae and diatoms.

Particularly in companies following good manufacturing practices, it is standard practice to monitor microbial load in order to determine the status of industrial fluids and the efficacy of biocides and bacteriostatic components. Although alternative methods exist e.g. plate counts, direct microscopic counts, ATP measurement, and enzyme activity (catalase); the method of choice is the dipslide method. Solar-Cult industrial dipslides were developed to provide an inexpensive, fast, simple method for detecting microbial contamination in these problem liquids. To meet the challenge, Solar Biologicals offers dipslides in several configurations. Some of the many advantages include the Following:

• Specific dipslide are coated with agar based media which have been developed for optimal bacterial and/or fungal growth
• Dipslides are provided in kits that are complete and ready to use. No special equipment or training is necessary.
• Dipslides can often be used to effectively measure microorganisms in specimens not miscible with water. Separation into oily and aqueous layers and even extraction of medium components can occur with aqueous presence/absence broths making interpretation difficult.
• Only a few microliters of the specimen are taken up by the dipslide. Antibacterial agents and biocides, present in industrial fluids and on surfaces, are diluted in the gel allowing growth of the bacteria with less interference. Presence/absence broth commonly requires vials of additives such as sodium thiosulfate to neutralize halogen disinfectants such as chlorine, bromine and iodine.
• Test systems are available for simultaneous counting of both bacteria and fungi.
• The dipslide handle facilitates sampling and reduces the opportunity for contamination.
• The plastic vial serves as a lightweight, convenient incubator and transport container.
• The same area of culture medium is exposed each time to liquid specimens or solid surfaces ensuring highly consistent results.
• The transparent container allows colonies to be viewed and counted safely.
• Simple color changes enable colonies to be recognized and counted with ease in selective media.

Since the mix of offending organisms causing problems can be quite different from one industrial situation to another, a choice of Solar-CultÒ industrial dipslides is provided. Our dipslides are double sided – enabling simultaneous examination of test specimens with two different selective media e.g. a selective medium for yeast and fungi on one side and bacteria on the other. Contact us about the possibility of ordering tailor-made dipslides to more specifically suit your requirements.

At least one side of all of our industrial dipslides is coated with Nutrient Agar to which a small quantity of the dye 2,3,5- triphenyltetrazolium (TTC) is added. Aerobic bacteria species grow on this medium and they can be detected by their ability to reduce TTC to a red colored formozan dye. Bacterial colonies appear as red dots on an otherwise clear colorless medium, rendering them easily recognizable by persons untrained in microbiology. Tetrazolium salts are heterocyclic organic compounds that form colored formazan crystals when reduced. In bacterial cells, tetrazolium salts behave as electron acceptors (oxidizing agents), in conjunction with dehydrogenase systems in the metabolic pathway. Tetrazolium salts are toxic to bacteria at high concentration or incubation at high temperature.

MacConkey Agar is coated on the alternative side of one of our dipslides. In this configuration a broad spectrum of bacteria will grow on the Nutrient Agar - TTC side whereas the growth of gram-negative bacteria is selectively enhanced on the MacConkey Agar side.

Two of our dipslides have one side coated with Media specific for the growth of fungi, e.g. Rose Bengal Agar and Malt Extract Agar. These media are adjusted to the acid side of neutrality (pH approx. 5.5) which not only favors the growth of fungi but also suppresses growth of most bacterial species. Bacterial growth is detected on the alternate sides of these dipslides that are coated with Nutrient Agar - TTC.

For assessing the microbial content of fluids, the slide can either be dipped into the liquid or a little poured on each side of the slide, ensuring the entire surface is wet. For pipes and semisolid materials, it might be found that swabbing the media surfaces with specimen collected on sterile cotton swabs is the best method of inoculation. Excess fluid should be allowed to drain for a few seconds before the slide is returned to the container. A typical procedure might entail the following:

Collect a homogeneous sample of the industrial fluid. A specimen from a fuel tank, for example, should show no evidence of layering indicating a second non miscible liquid, e.g. water. It may be necessary to dilute highly viscous specimens such as paint with water. Remember to multiply results by a dilution factor.

1. The slide is removed from its container taking care to not touch the agar surfaces. It is then dipped into the fluid until both agar surfaces become completely wet (Procedures range from 2-3 seconds to as much as 30 seconds).
2. Withdraw slide. Allow excess fluid to drip off.
3. Wipe off the remaining drops of fluid by touching the end of the slide with a tissue. Do not touch the agar surfaces with the tissue.
4. Screw the slide back into the container, label and incubate. The incubation temperature is commonly that of the ambient temperature of the industrial fluid being sampled. Lower than expected numbers of colonies might be observed for media containing tetrazolium salts when incubated at temperatures greater than 50^oC.

In order to interpret the result, the user compares the density of colonies appearing on the slide after inoculation and incubation to a colony density chart. Each replicating bacterium collected on the dipslide produces a single red colored colony that appears on the slide as a red dot. The charts were derived by photographing dipslides inoculated by dipping into bacterial suspensions for which accurate counts of cells present were determined by a dilution plate count method. Optimum times for reading the bacterial side of the slides usually range from 12 to 48 hours.The time depends on incubation temperature and the growth characteristics of the particular bacteria in the specimen. After 2 to 4 days, confluent bacterial growth or interference by slow growing fungi and yeasts might make interpretation difficult. Yeasts generally appear as smooth circular colonies with the consistency of thick paste. They may be pigmented or colorless but do not generally become red if found on the bacterial side.

Because of growth inhibition of other bacteria, the number of colonies growing on MacConkey medium, which is a selective medium specific for gram-negative bacteria, may not be comparable to the number growing on the non-selective Nutrient Agar - TTC side.
Recognizing that most users are not microbiologists, the media for detecting fungal growth (Rose Bengal and Malt Extract Agar) have been modified to improve selectivity by inhibiting bacteria. Fungal growth (fungi, moulds and yeasts) is generally slower than that of bacteria. If no growth appears after 48 hours, incubate for an additional 48 hours.
Yeasts grow as smooth colonies whereas moulds tend to produce cottony, filamentous structures. Fungal levels are generally reported as nil, low, moderate or heavy contamination. Since fungi tend to adhere to surfaces making capture on dipslides difficult, counts are usually not meaningful as a measure of fungal contamination.

For quality assurance purposes (HACCP, cGMP, ISO9000, etc.); the procedures, results and interpretation need to be documented and reviewed for microbiological testing as with any other quality control activity. Charting results and establishing cause effect relations is often necessary at the outset to establish benchmarks and sampling frequencies. For example, the presence of moderate amounts of fungi or bacteria levels greater than 10⁵ might require remedial activity. After a baseline of contamination is established, testing on a less frequent basis is generally required to monitor changes from the baseline. It may be found useful to consider the microbial assay results with other factors such as specimen odor, color, effectiveness, etc. in decisions regarding remedial activity.

Kirko, H.T. and R.E. Burrell. 1991 The effects of tetrazolium salts on bacterial enumeration in aluminum hot rolling fluids. Lubrication Engineering. Journal of the Society of the Tribiologists and Lubrication Engineers. 47 (6): 505-508.