EH-15 Potentially Hazardous Amoebae Found in Eyewash Stations
                         ENVIRONMENT, SAFETY & HEALTH

                                   BULLETIN
Assistant Secretary for                              U.S. Department of Energy
Environment, Safety, & Health                        Washington, D.C. 20858

DOE/EH-0010                      Issue No. 15                         May 1986

POTENTIALLY HAZARDOUS AMOEBAE FOUND IN EYEWASH STATIONS

Acanthamoebae, small amoebae capable of causing serious eye infections, were
found in numerous portable and stationary eyewash stations at a DOE facility.
Such infections can be particularly severe -- they have often caused the loss
of the infected eye.

Diagnosis has been difficult and often delayed: the organism cannot be
identified on the usual corneal scrapings and smears; special culture plates
are needed.  Often Acanthamoeba is considered only after treatment for more
common causes of infection has failed.

Treatment, too, has been difficult because most antibiotics are ineffective
against this amoeba.

Acanthamoebae are able to survive conventional water plant treatment regimens.
Therefore, extra precautions are needed to rid them from water used to flush
traumatized eye tissue.

Evaluation of the Problem

Both the presence of acanthamoebae in potable water and their potential
harmfulness has been documented at one DOE site.

Water samples were taken from eleven eyewash stations at four geographically
disparate locations throughout the site and acanthamoeba were found in
samples from all four areas.  To identify the amoebae, the water samples were
filtered through 1.2-micrometer cellulose membranes; the filters were
inverted and placed onto nonnutrient agar plates seeded with a lawn of live
Escherichia coli, then the plates were incubated at 37 degrees C for amoebic
growth.  Amoebic outgrowths from the filters were analyzed morphologically
for the presence of Acanthamoebae.

Many of the resultant acanthamoebae isolates were treated for pathogenicity by
intranasal inoculation into weanling ICR mice.  Moribund mice were sacrificed
14-90 days after inoculation and the brain and lung tissues plated on E. coli-
seeded agar plates.  The presence of acanthamoebae growing from lung tissue
weeks or months after inoculation proved that some of the samples from the
eyewash stations were capable of causing chronic lung infections in mice.

Evaluation of the Controls

To test the efficacy of chlorine treatment, the portable stations were
treated with calcium hypochlorite to effect a free chlorine residual of
approximately 25 ppm and then tested at intervals between 30-150 days later
for the presence of Acanthamoebae.  This treatment was effective in
destroying them.  After only one treatment, 6 of 7 portable eyewash stations
were free of acanthamoebae.  Corrosion was found in some of the stainless
steel eyewash stations, however, suggesting that this level of chlorine is

too harsh for the equipment.  One-minute and three-minute flushing regimens
were also tested: one-minute flushings did little to reduce the number of
acanthamoebae; three-minute flushings drastically reduced the number of
positive samples.  How long this diminution persists has not been delineated
but from one test it appears to be less than six weeks.

Present Status and Current Recommendation

There are still many important questions about their control remaining to be
investigated: the optimal level of chlorination -- one that destroys the
amoeba but not the equipment -- needs to be determined; the use of hydrogen
peroxide instead of chlorine needs to be investigated; we need to learn how
long the diminution of amoebae lasts after three-minute flushings and whether
there is an accumulative effect; and whether there is a cumulative effect
after one-minute flushings, which would make them more effective than we are
now led to believe.

Meanwhile, in light of the confirmation that eyewash stations filled with or
connected to potable water supplies may be contaminated with Acanthamoeba,
we recommend that eyewash stations be flushed weekly, preferably for three
minutes.

For more technical information, contact: Dr. R. L. Tyndall, Biology Division,
Oak Ridge National Laboratory, Y-12 Area, Oak Ridge, Tennessee 37831.
Telephone: FTS: 624-0686; Commercial: 615-574-0686.

MEDICAL TREATMENT OF ACANTHAMOEBA EYE INFECTION REPORTED

Eye infections caused by Acanthamoeba are especially challenging.  Until
recently, excisional keratoplasty was the most successful diagnostic and
therapeutic measure.  The first effective medical treatment -- a combination
of dibromopropamidine isethionate drops(1) and propamidine isethionate
ointment(2) and neomycin drops -- has been reported in the British Journal
of Ophthalmology(3).

These infections are difficult to recognize and treat, and the outcome of
all reported cases has been very poor.

Although they have characteristic features, such as a slow relapsing course
with development of ring abscess, recurrent opithelial breakdown, hypopyon
which waxes and wanes, and secondary glaucoma, diagnosis is difficult because
the organism cannot be identified on the usual corneal scrapings and smears
which are taken and air dried to fix cells and bacteria.  Such air drying
causes rupture of these delicate organisms, leaving no distinctive residuum,
but preserves cysts where present.  Special culture plates of plain agar
preseeded with aerobacter or E. coli to encourage growth of Acanthamoeba are
needed.  Too often, Acanthamoebae are considered only after conventional
treatment for more common causes of suppurative keratitis has failed.
Treatment is difficult because most antibiotics are ineffective.

This particular case differed from other reported cases in that it did not
involve direct ocular trauma or previous corneal ulcerations(4).  The patient
was a physically fit 44-year-old meat porter who was splashed in the right
eye with blood from a condemned carcass of meat but did not suffer any direct
trauma to the eye.  This incident occurred in April 1979.  The infection which
insued was treated with various topical medicaments by the patient's general
practitioner and local ophthalmologist until June, when the patient was
referred to Moorfields Eye Hospital, London.  Treatment continued but by

September the corneal epithelium had broken down, with increasing stromal
supporation and hypopyon, producing an appearance similar to that seen in
another patient known to have had acanthamoeba infections.  Smears continued
to show only pus cells and debris with no organisms.  Special cultures from
both cornea and conjuctiva grew Acanthamoeba, permitting their positive
identification.

Both propamidine and dibromopropamidine have long been available in Great
Britain for ocular therapy.  In addition to the amoebicidal effect, these
compounds show antibacterial and antifungal properties.

The optimum treatment regimen still needs to be determined.  In this case,
treatment began with propamidine isethionate drops every hour and
dibromopropamidine isethionate ointment every four hours, day and night.
After nine days, the eye became much redder with rapidly worsening lid
swelling despite improvement in corneal signs.  The doctors attributed the
local irritation to toxicity, seen in some patients after topical application
to wounds for more than 10 days.

Since drops were used again four times daily after a penetrating keratoplasty
22 months later and were well tolerated, the doctors suggested that intensive
initial treatment probably should not last longer than one week and,
thereafter, treatment four times daily should suffice.

After 9 days, the treatment was changed to neomycin drops every four hours
day and night.  By October the epithelium had healed and prednisolone drops
were added every four hours, with steady improvements in the corneal
inflammatory signs.  The treatment was slowly reduced until the patient was
using only neomycin drops four times daily.  These were continued for one
year, at which time limbal follicles developed with some increased palpebral
conjunctival hyperaemia and cellularity but no sign of any skin irritation.
All topical therapy was stopped and penetrating keratoplasty proposed.  It was
performed 22 months after initial presentation.  Postoperative treatment
included propamidine isethionate drops four times daily plus the usual
topical steroid and antibiotics.  After therapy was stopped, the graft
remained clear for nine months, when a rejection episode developed.  It was
easily controlled with topical steroids.

The doctors reported that at the time of writing the graft was clear and
vision was 6/24, being reduced by lens opacities.  The intraocular pressure is
being controlled with timolol maleate 0.25% drops twice daily.

1. & 2. Not approved by FDA.

3. Wright. P., Warhurst, D., and Jones, B.J.,"Acanthamoeba Keratitis
   Successfully Treated Medically." British Journal of Ophthalmology,
   1985, 69: 778-782.

4. A case associated with soft contact lens wear was reported recently in the
   United States.  Successful treatment included propamidine isethionate.
   Moore, M.B., McCulley, J.P., et al, "Acanthamoeba Keratitis Associated
   with Soft Contact Lenses," American Journal of Ophthalmology, September
   1985, 100: 396-403.

The complete article, "Acanthamoeba Keratitis Successfully Treated
Medically," British Journal of Ophthalmology, can be obtained from Nona F.
Shepard, Office of Occupational Safety, EH-134, Department of Energy,
Washington, D.C. 20545.  FTS: 233-2958; Commercial: (301) 353-2958.

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| SOME REMINDERS ABOUT EMERGENCY EYEWASH STATIONS                            |
|                                                                            |
| Any delay in treating chemical burns of the eye will generally aggravate   |
| the injury*.  The initial treatment of choice for chemical burns is        |
| active mechanical flushing of the eyes with copious amounts of water. For  |
| serious burns (i.e. strong alkalies or acids), authoritative sources       |
| indicate that active irrigation should generally continue for a period of  |
| 20 to 30 minutes.  All employees potentially exposed to corrosive          |
| materials should be instructed in the proper use of eyewash fountains and  |
| the length of time necessary to flush eyes.                                |
|                                                                            |
| Providing 20 to 30 minutes of eye irrigation requires a considerable       |
| volume of water.  American National Standard for emergency eyewash and     |
| shower equipment (ANSI) Z358.1-1981, requires that plumbed and             |
| self-contained eyewash equipment provide at least 0.4 gallons per minute   |
| (gpm) flow for at least 15 minutes.  Most commercial plumbed and portable  |
| equipment provide 1.5 to 2 gpm.  Therefore 20 to 30 minutes of irrigation  |
| may require 30 to 40 gallon of water.  Many portable eyewash stations      |
| have a capacity of 5 to 10 gallons for a maximum usefulness of 5           |
| minutes.  Squeeze bottles and other plastic container devices have even    |
| lower water capacities.                                                    |
|                                                                            |
| All eyewash stations require maintenance.  ANSI Z358.1 requires that       |
| plumbed eyewash units be activated weekly to flush the line and verify     |
| proper operation.  Portable units are generally sealed pressurized         |
| systems.  In addition to the need to change water, pressurized equipment   |
| requires frequent maintenance checks to recharge if necessary.  Squeeze    |
| bottles also require frequent inspection and maintenance since they lose   |
| water to evaporation, become contaminated, and are easily misplaced.       |
|                                                                            |
| Based on the above considerations, the following guidance is provided:     |
|                                                                            |
| 1. In general, squeeze bottles should not be used since they hold          |
|    inadequate amounts of water and might delay proper eye irrigation.      |
|    The exception would be where hazard severity or distance requires       |
|    personal equipment at the work station for immediate flushing prior to  |
|    prolonged flushing at a plumbed or self-contained unit.  Be sure that   |
|    the shelf-life expiration is marked on the squeeze bottle and that it   |
|    is not exceeded.  Each unit should be tested, refilled, and maintained  |
|    in accordance with the manufacturer's instructions.                     |
|                                                                            |
| 2. Self-contained eyewash stations should not be used in areas where a     |
|    continuous source of potable water is available.  They should be used   |
|    only in remote areas where installation of a potable water              |
|    distribution system is not economically feasible.  Self-contained       |
|    units should meet the 0.4 gpm flow rate and 15 minutes duration         |
|    requirements of ANSl Z358.1                                             |
|                                                                            |
| 3. Plumbed eyewash units should be activated weekly to flush the line and  |
|    to verify proper operation.  Self-contained units must be inspected in  |
|    accordance with the manufacturer's instructions.                        |
|                                                                            |
| 4. Weekly flushing for three minutes is recommended to reduce              |
|    Acanthamoebae in potable water (see accompanying article).              |
|                                                                            |

|                                                                            |
|                                                                            |
|                                                                            |
| *OSHA standard 29 CFR 1910.151, paragraph C, states "Where the eyes or     |
| body of any person may be exposed to injurious corrosive materials,        |
| suitable facilities for quick drenching or flushing of the eyes or body    |
| shall be provided within the work area for immediate emergency use."       |
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The Bulletin is published so that DOE program managers and contractors can
share information about potential health and safety problems relevant to DOE
operations.  For additional copies, contact Nona Gilbert Shepard, Editor,
Office of Operational Safety, Assistant Secretary for Environment, Safety &
Health, U.S. Department of Energy, Washington, D.C. 20545, telephone FTS
233-2958; Commercial (301) 353-2958.
.
.