MPS DISINFECTION

Disinfection Efficacy of MPDS Against Pseudomonas

By Ruth A. Rosenthal, MS, Cindy L. McAnally, BS, Ralph P. Stone, PhD, and Barry A. Schlech, PhD
January 2001

This multi-purpose solution's disinfecting efficacy was tested against 25 strains of Pseudomonas aeruginosa.

Infections among contact lens wearers seem to have a different etiology than those among the non-contact lens population. Pseudomonas aeruginosa, which is only occasionally isolated from non-contact lens-related keratitis, is the most frequently recovered organism from contact lens wearers with corneal ulcers.

P. aeruginosa, a Gram-negative aerobic bacterium, is an opportunistic pathogen capable of causing chronic and acute infections in compromised hosts. A corneal Pseudomonas infection usually begins with minor trauma to the eye, causing interruption in the epithelial surface that allows invasion into the underlying stroma. P. aeruginosa can be an intracellular or extracellular pathogen to the eye, either initiating an inflammatory response or it can be cytotoxic and kill eucaryotic cells. A transmission electron micrograph of one of our P. aeruginosa isolates is shown in Figure 1.

Pseudomonas keratitis is usually more common in humid environments. It has been isolated from soil, sewage, water, plants and animals, and is sometimes part of man's normal flora. It has minimal nutritional requirements and can grow in distilled water. P. aeruginosa has been found in a variety of aqueous solutions, including contact lenses and solutions, cosmetics, disinfectants, ointments, soaps, irrigation fluids, eye drops and dialysis fluids. It can also frequently be found in aerators, traps of sinks, whirlpool and hydrotherapy baths, respiratory equipment and showerheads.

The prevalence of P. aeruginosa raises concern about non-compliance among contact lens wearers. Improper handling of the lens may add potential contaminants, and bacterial adhesion to contact lenses may lead to infection. Appropriate care of the lenses and lens cases, as well as use of a contact lens disinfecting solution which kills these organisms, can reduce the risk of contamination.

During development of new contact lens disinfecting solutions, products are tested according to FDA guidelines and draft ISO standards against representative bacteria, yeast and mold in order to demonstrate disinfecting efficacy. One of the required organisms includes a representative strain of P. aeruginosa. However, the single strain of the organism used during the basic testing process may not be totally representative of the entire species. The FDA guidelines and draft ISO 14729 standards provide guidance for the performance of products labeled as contact lens disinfectants. These procedures require that the disinfecting (soaking) step of products demonstrate greater than or equal to an average of a 3.0 log reduction of the representative bacteria and greater than or equal to an average of a 1.0 log reduction of the yeast and mold. The literature has shown that Opti-Free Express Multi-Purpose Disinfecting Solution meets the primary criteria of the FDA guidelines and draft ISO standards for disinfection of contact lenses against these regulatory organisms. There are no standards for other strains of organisms.

We carried out this study to confirm the performance of Opti-Free Express MPDS with a variety of P. aeruginosa isolates.

Materials

Opti-Free Express MPDS with Aldox is an enhanced disinfection formula preserved with 0.001% Polyquad (polyquaternium-1) and 0.0005% Aldox (myristamidopropyl dimethylamine). We obtained media, reagents and laboratory supplies used in this study from commercial sources.

The Pseudomonas aeruginosa strains used in this study included isolates from microbial keratitis, non-corneal sources and a contact lens solution and lens case. Some of the strains are identified as invasive and cytotoxic. Also, some of the serogroups are identified. Seven different serogroups were evaluated. A serogroup refers to those strains of organisms that are serotype specific. Serotyping is used for detecting bacterial antigens on the cell surface and is one of the classic tools for epidemiological studies. A complete description of the isolates is shown in Table 1.

Methods

Each isolate of Pseudomonas aeruginosa was grown on soybean-casein digest agar (Becton, Dickinson and Company; BBL) at 30 to 35 degrees C for 18 to 24 hours. Three lots of the product were challenged with separate batches of each isolate of P. aeruginosa using procedures based on FDA guidelines and draft ISO standards for contact lens disinfecting solutions. The product was sampled for survivors at two, four, six and 24 hours. We used serial dilutions in Dey Engley Neutralizing Broth (D/E Broth; DIFCO) and duplicate plating with soybean-casein digest agar containing 0.07% lecithin and
0.5% polysorbate 80 (BBL) for recovery of the survivors. The number of CFU/ml was determined following appropriate incubation. Appropriate recovery media and inoculum controls were prepared according to the specified procedure.

Results

Opti-Free Express MPDS reduced the populations of all isolates of Pseudomonas aeruginosa by 3.8 to 5.3 log units after two hours. Following four hours of exposure to the product, a 4.2 to 5.2 log reduction was achieved. At six hours, 84 percent of the P. aeruginosa isolates showed no detectable survivors. The PA17 isolate showed a slightly lower average log reduction, although two of the three lots tested showed a 4.7 log reduction against this isolate. At 24 hours, no survivors of any of the 25 P. aeruginosa isolates were detected.

A similar distribution of kill was obtained regardless of the source of the isolate, mechanism of infection or serogroup of the isolates tested. The two isolates from the lens case (PA08 and PA09) were reduced by approximately 5.0 log units ± 0.1 standard deviation (SD) by Opti-Free Express MPDS at six hours. The 12 keratitis isolates were reduced by about 4.8 log units ± 0.2 SD. The seven non-corneal isolates were reduced by about 5.0 log units ± 0.1 SD at the six-hour soak time. In this study, a log reduction of 4.7 to 5.2 log units indicated no detectable survivors, depending upon inoculum size. In general, the cytotoxic strains were reduced by about 4.8 log units ± 0.3 SD and the invasive strains by about 5.0 log units ± 0.2 SD at six hours. The seven serogroups tested were also reduced by about 4.8 log units ± 0.2 SD.

Discussion and Conclusions

Pseudomonas aeruginosa is one of the most frequent causes of corneal ulcers in contact lens wearers. Approximately 60 percent of the contact lens-related cases of microbial keratitis are attributable to this organism.

P. aeruginosa can cause rapid perforation of the corneal stroma because of its proteolytic enzyme production. It has a high incidence in patients with predisposing ocular conditions. Pseudomonas keratitis typically begins as a small central ulcer that spreads concentrically and may involve the entire cornea, sclera and deeper portions of the stroma. Since a Pseudomonas corneal ulcer may lead to rapid loss of ocular function, appropriate therapy is essential.

Many lens wearers are unaware of the potential risks of non-compliance. Among the causes of corneal infections, inadequate hygiene and overextended wear may play an important role. As many as 50 to 70 percent of lens wearers do not follow the recommended regimen. Lens cases are frequently cited as a source of various microbial contaminants. The degree of contamination may be related to compliance with patient instructions for use of the product.

Opti-Free Express MPDS demonstrated high performance against a large number of P. aeruginosa isolates, reducing the populations of all isolates of P. aeruginosa tested more than 3.8-log units by two hours. The P. aeruginosa isolates were reduced about 5.0 log units of a 5 to 6 log CFU/ml inoculum, or essentially no survivors after six hours exposure. The current FDA guidelines and draft ISO standards require an average 3.0-log reduction of designated bacteria at the disinfection time to meet the primary criteria. This high level of efficacy occurred regardless of whether the isolate was from a corneal infection or contact lens accessories, regardless if the strain was invasive or cytotoxic and regardless of the serogroup tested. By 24 hours, there were no survivors detected for any of the 25 different P. aeruginosa isolates. These results were similar to those reported by researchers at the Indiana State University for Opti-Free Express MPDS and provided reduction of two P. aeruginosa isolates of 5.0 and 5.3 log units, respectively.

The results showed that Opti-Free Express MPDS typically reduced the level of P. aeruginosa to no detectable survivors. It can be concluded that Opti-Free Express MPDS is very effective in killing a variety of P. aeruginosa isolates from contact lens solutions and cases, from microbial keratitis and other human non-corneal isolates.

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Ruth A. Rosenthal, MS, is the Director of Consumer Products Microbiology at Alcon Research, Ltd.

Cindy L. McAnally, BS, is a Scientist in R&D Microbiology in Consumer Products Microbiology at Alcon Research, Ltd.

Dr. Stone is the Vice President of Research and Development for Consumer Products at Alcon Research, Ltd.

Dr. Schlech is Vice President of R&D Pharmaceutical Technology and Microbiology at Alcon Research, Ltd.