Microbial Monitoring in the Pharmacy

Clinical Microbiology Newsletter Vol. 31, No. 4

www.cmnewsletter.com

$88 February 15, 2009

Microbial Monitoring in the Pharmacy Alice S. Weissfeld, Ph.D., D(ABMM), F(AAM) and Paula H. Vance, B.A., SM(ASCP), SM(NRM), M(CLSp), CIE, Microbiology Specialists Incorporated, Houston, Texas

Abstract In 2007, the United States Pharmacopeia finalized a program to ensure that compounded sterile products prepared for individual patients by physician order were indeed prepared aseptically. The program name became the chapter number in the national formulary, i.e., 具797典. Accredited clinical or environmental laboratories are required to support microbial testing of (i) the air in the i.v. area and hoods, (ii) the sterility of the compounded product itself, (iii) the reliability of surface decontamination, and (iv) the determination of whether the pharmacist or pharmacy technician can compound a surrogate product aseptically. This article reviews those quality metrics for pharmacies that involve microbial testing.

Background The July 15, 2008 (Vol.30, No.14) issue of the Clinical Microbiology Newsletter contained an article authored by Eric S Kastango, RPh (1) regarding the evolution of the environmental monitoring program now required by the United States Pharmacopeial Convention (USP). The USP has been responsible for maintenance of the U.S. National Formulary since the early 20th century. The National Formulary is published yearly with supplements as needed. The book is printed on over a thousand pages of tissue paper and also includes instructions for the pharmaceutical industry listing best practices. Embedded within more than 1,000 general information chapters are ones dealing with microbial assays in support of the industry. Chapter 具797典 was first published in 2004 (2) and revised after an extensive comment period in 2007 (3). The chapter entitled “Pharmaceutical Compounding — Sterile Preparations” is to the phar-

Mailing Address: Alice S. Weissfeld, Ph.D., D(ABMM), F(AAM), Microbiology Specialists Incorporated, 8911 Interchange Dr., Houston, TX 77054. Tel.: 713-6636888. Fax 713-663-7722. E-mail: [email protected]

Clinical Microbiology Newsletter 31:4,2009

macist what the Clinical Laboratory Improvement Act (CLIA) and subsequent Amendments is to the clinical microbiologist. It has taken almost 4 years for some individual state boards of pharmacy to adopt the regulations in chapter 具797典. Although not all 50 states have currently adopted all of the requirements in this chapter, most have it under serious consideration. The Joint Commission recognizes the chapter as a best-practice guideline. The Food and Drug Administration (FDA) provides federal oversight. There have been multiple incidences of morbidity and mortality following contamination of sterile preparations (4-12), so it is probably reasonable to assume that helping pharmacies with their 具797典 program will be a giant step toward fulfilling patient safety goals. When clinical microbiologists are asked to support microbial investigations in areas of the hospital outside of the laboratory, it may be the first time that they have ventured into a non-laboratory venue. Therefore, the first part of this article will be devoted to “learning the lingo” so that everyone is on the same page. To understand how the pharmacy works, it is important to understand the terminology. Pharmacy© 2009 Elsevier

specific terminology is in boldface below. Compounded sterile preparations (CSPs) are made by pharmacists or pharmacy technicians in response to a specific physician prescription. This is quite different than manufactured sterile products that are prepared by pharmaceutical companies. Manufactured sterile products are produced under strict FDA supervision according to specific product labeling that must be approved by that agency (13). CSPs have the potential to be among the most hazardous drugs given to a patient, as they are administered intrathecally, intravascularly, intraocularly, or into joints. They are also used as baths for live organs

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or tissues that are used in transplant patients. In order to comply with 具797典, the pharmacy is required to have performed a gap analysis. The pharmacist doing the gap analysis must analyze each CSP prepared by the pharmacy with an eye toward classifying each preparation as low risk, medium risk, high risk, or immediate use. The pharmacy’s ultimate risk level is determined by the riskiest preparation it produces. By definition, immediate-use products will be used within 12 hours of compounding. It will be important for the microbiologist to know their pharmacy’s risk level, as the microbial testing performed will depend on this assessment.

Risk Levels High-risk products are defined as those that start out using a nonsterile ingredient that must be sterilized during the course of preparation. An example of a high-risk compound is morphine, which is used for pain management when injected into the spinal column. Medium-risk CSPs are those in which there are multiple complex transfers of individual sterile products. An example of a medium-risk CSP is a total parenteral nutrition solution in which carbohydrates, fats, amino acids, and vitamins are individually added together. Low-risk CSPs involve a single volume transfer of a sterile ingredient. An example would be the preparation of an antibiotic. Immediate-use CSPs are defined as those prepared in emergency situations (such as in the emergency room), where the preparation will be used immediately. Only low-risk CSPs can be prepared for immediate use, and in some hospitals, they are prepared by physicians or nurses. As a general rule, most hospital pharmacies are medium risk.

Engineering Controls The microbiologist must also know what the primary and secondary engi-

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Table 1. Components of USP 具797典 Microbial Monitoring Program Category

Testing required

CSP sterility

Sterility Endotoxin

Personnel competency

Media fill testing Gloved fingertip testing

Environmental sampling

Count particles and culture air in primary and secondary engineering controlsa

Nonviable

Measure number of particles ≥ 0.5 mm

Viable

Culture air

a

Primary engineering controls: laminar airflow workbench, biological safety cabinet compounding aseptic isolator, compounding aseptic containment isolator. Secondary engineering controls: buffer area and anteroom.

neering controls are in their pharmacy to know how to set up the environmental monitoring program. A pharmacy’s primary engineering controls include a laminar airflow workbench (LAFW), a biological safety cabinet (BSC), a compounding aseptic isolator (CAI), and a compounding aseptic containment isolator (CACI). These are different types of hoods in which sterile compounding takes place. The CAI and the CACI look like our anaerobe chambers. The LAFW is an open workbench with unidirectional air flow, and the BSC is the same as those in microbiology laboratories. BSCs and CACIs are specifically used to compound chemotherapy preparations and must be exhausted directly to the outside. Primary engineering controls must maintain ISO class 5 air quality. The LAFW and BSC must be located within a buffer area. The buffer area should have HEPAfiltered air introduced at the ceiling so that it meets ISO class 7 air quality. The CAI and CACI may be situated outside of a buffer area as long as they can be certified to ISO class 5 standards during active compounding activities. Finally, an anteroom must be located adjacent to the buffer area. The anteroom must maintain ISO class 8 air quality.

© 2009 Elsevier

The anteroom is essentially a staging area for the pharmacist or pharmacy technician preparing to compound. It is also where the compounder dons his/her personal protective equipment. Compounders are required to wear booties, hair bonnets, beard covers, full-length disposable coats, and sterile gloves. In addition, compounders may not wear jewelry (including watches). Female compounders may not wear any makeup or have long or artificial nails. The pharmacy engineering controls, with the exception of the BSC or CACI used for compounding chemotherapy drugs, are all under positive pressure. The buffer zone is, by definition, at higher pressure than the anteroom so that net airflow is out of the buffer zone. The LAFW is at positive pressure to the buffer area. This is the exact opposite situation of what happens in the microbiology laboratory, where, for example, the acid-fast laboratory is at lower pressure than the adjacent rooms so that the net flow of air is into the laboratory. Similarly, the microbiology BSC is a negative-pressure cabinet. It pulls air from the room into the cabinet, and most of the air is then released from the BSC after HEPA filtration.

Clinical Microbiology Newsletter 31:4,2009

The Microbial Sampling Program Table 1 shows the components of a microbial sampling program. The program includes testing the environment in all of the i.v. preparation areas as well as the testing of personnel and the CSPs themselves. Frequencies of testing are listed in Table 2.

Sterility Testing CSPs should be randomly tested by either direct inoculation or membrane filtration of a product as defined in USP Chapter 具71典 (14). Tryptic soy broth (TSB) and thioglycollate broth (thio) are used for this purpose. The decision to use either direct inoculation or membrane filtration is made based on the volume of the CSP. Any CSP with a volume of <10 ml is tested by direct inoculation. Any CSP with a volume of >10 ml is tested by membrane filtration. For membrane filtration, the product is filtered and the membrane is aseptically cut so that one half is placed in TSB and the other is placed in thio. Thio is incubated at 32.5ºC ± 2.5ºC and TSB at 22.5ºC ± 2.5ºC for 14 days. The USP currently does not recognize the injection of CSPs into blood culture bottles as an acceptable method of testing them for sterility.

Endotoxin Testing Endotoxin testing is only required for high-risk CSPs. Therefore, it is unlikely that most clinical microbiologists will be approached by high-risk pharmacies to perform endotoxin testing, since most hospital pharmacies are medium risk. However, the single-tube pyrogen test at 1 EU (endotoxin unit) used for dialysis water is NOT appropriate or acceptable for CSPs. The reason is that most drugs in the national formulary contain specific bacterial endotoxin limits as stated in the official monographs for each drug. In addition, if the official monograph does not contain a limit, then the amount of endotoxin in the CSP should not exceed the amount of endotoxin units for the appropriate route of administration. The calculation per hour per kilogram of body weight can be found in USP chapter 具85典 on endotoxin testing (15). It is important to perform the testing using a method that allows plotting a standard curve so that the exact amount of endotoxin present can be determined. Clinical Microbiology Newsletter 31:4,2009

Table 2. Frequencies of environmental, personnel, and CSP testing Category

Frequency of testing

CSP sterility testing

Only required for high-risk CSPs; however, CSP sterility testing is a good way to randomly assess personnel competency by using CSPs that would otherwise be discarded.

CSP endotoxin testing

Only required for high-risk CSPs

Personnel competency Media fill testing Low and medium risk High risk

On employment and annually thereafter On employment and semi-annually thereafter

Gloved fingertip testing

3 times upon employment and annually thereafter

Environmental sampling Non-viable and viable air sampling

Surface sampling

Minimum of every 6 months, as well as: • On commissioning and certification of new facilities and equipment • Following any servicing of primary or secondary engineering controls • In response to identified problems with CSPs or any patient-related infection where the CSP may be implicated • Following renovations to the pharmacy Randomly on all primary and secondary surfaces

Media Fill Testing Media fill testing is designed to check the aseptic technique of individuals performing sterile compounding. Each risk level has a separate protocol that takes into account the number and types of manipulations performed by the compounder in each situation. For example, since medium-risk involves multiple transfers of sterile products, these types of manipulations are emphasized in the medium-risk media fill testing. Likewise, since high-risk testing starts with weighing a non-sterile powder, TSB powder, used as a surrogate for a pharmacopeial product, is weighed out and reconstituted. The microbiologist and the pharmacist can design their own test or the products necessary for each type of kit may be purchased. Hardy Diagnostics (Santa Maria, CA) has USP media fill kits that may be viewed on its website (http://www.hardydiagnostics. com). Their media fill kits are sold under the HardyVAL trademark.

Gloved Fingertip Testing The USP requires that compounders wear sterile gloves when performing their job, as the Expert Sterile Compounding Committee believes that the © 2009 Elsevier

easiest way to contaminate CSPs is by direct contact with hands. At the end of their 8-hour shift, the compounder should place the five fingers of each hand on a separate tryptic soy agar plate with polysorbate and lecithin (TSApl). The polysorbate and lecithin are necessary to neutralize the 70% alcohol that compounders use to sanitize their gloves between preparations. The TSApl is incubated at 35ºC ± 2ºC for 48 to 72 h in ambient air, and the colonies on each plate (one for the left hand and one for the right hand) are counted. Greater than 3 colony-forming units (CFU) for both hands is unacceptable. Corrective action for those individuals over the limit is usually retraining and retesting.

Environmental Sampling: Non-Viable Non-viable sampling is performed by the same individuals who certify hoods in our microbiology laboratories. They use particle counters to determine the number of particles in the air. These instruments count particles from <0.5 μm to ≥ 10 μm, and the range and sizes should be reflected in total counts. The number of particles corresponds to the ISO class, as shown in Table 3. 0196-4399/00 (see frontmatter)

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Environmental Sampling: Viable The thought of having to perform viable in addition to non-viable air sampling was probably the hardest thing for the pharmacy community to accept. The USP chapter calls for air sampling to be performed using a volumetric impaction sampler. Pharmacists who were doing air sampling were using settling plates. However, settling of particles by gravity is not a reliable method to follow trends. The settling of particles depends on particle size and may be influenced by air movement. Consequently, the number of CFUs on a settling plate may not always relate to concentrations of viable particles in the sampled environment. Therefore, volumetric air samplers that pull in a known amount of air must be used. A comparison of commonly used samplers is shown in Table 4. Pharmacies may either buy their own samplers or contract with a hood certifier, industrial hygienist, or microbiologist who may have one. The first thing that the pharmacist must do is write a sampling plan. The sampling plan should include a floor plan of the i.v. compounding area and the hoods. There should be an X on the plan at each location to be tested so that samples are taken at the same

Table 3. Non-viable air samplinga Engineering control

ISO particle count per m³ of air

5

LAFW, BSC, CAI, CACIb

3,520

7

Buffer area

352,000

8

Anteroom

3,520,000

ISO class

a

Adapted from reference 3. b LAFW, laminar airflow workbench; BSC, biological safety cabinet; CAI, compounding aseptic isolator; CACI, compounding aseptic containment isolator.

exact area each time. The plan should also include the method of collection, the frequency of sampling (at least every 6 months for low- or mediumrisk and weekly for high risk pharmacies), the volume of air sampled (400 to 1,000 L is suggested), and whether the sampling was active or passive. Active sampling is recommended in chapter 具797典. This means that air samples are collected while compounding is actually taking place. Passive sampling means that no one was working under the hood at the time of sampling. USP Chapter 具797典 also states that the environmental sampling results should be trended. In order to trend the results, many pharmacies are doing more than the minimum sampling, i.e., they are

sampling monthly or quarterly. Two media are required for air sampling, one for bacteria and one for fungi. The recommended bacterial medium is TSApl; the recommended fungal medium is MEA (malt extract agar). TSApl is incubated at 30 to 35ºC for 48 to 72 h, and MEA is incubated at 26 to 30ºC for 5 to 7 days. At the end of the incubation period, colonies are counted and the number of cfu/m³ is reported. Most air samplers have a conversion table based upon the amount of air sampled that converts the number of colonies on the plate to the final number. Recommended action levels are shown in Table 5. However, regardless of the number of CFU/m³ of air, the presence of certain organisms (gram-negative

Table 4. Volumetric air samplers Sample Air

Air Ideal

SAS Super 180

RCS High Flow

CI-90 CI-95

M and T

Quick-Take 30 + Biostage

Andersen N-6

AES Laboratoire

bioMérieux

Bioscience International

Biotest Diagnostics

CLiMET

Millipore

SKC

Thermo Fisher

AES Laboratoire media or standard prepared media from any medium manufacturer

Standard prepared media from any medium manufacturer

Standard prepared media from any medium manufacturer

Proprietary strips

Standard prepared media from any medium manufacturer

Must use their ready-to-use cassettes

Standard prepared media from any medium manufacturer

Standard prepared media from any medium manufacturer

Highvolume sampler

Yes, 9999 L in 99.99 h

Yes, 1,000 L in 10 min

Yes, 1,000 L in ∼5.5 min

Yes, 1000 L in 10 min

Yes, 1,000 L in 10 min

Yes, 1,000 L in <7 min

Yes, 1,000 L in ∼34 min

Yes, 1,000 L in ∼37 min

Flow rate

100 L/min

100 L/min

180 L/min

100 L/min

100 L/min

140 L/min for first 500 L, then 180 L/min for second 500 L

30 L/min

28.3 L/min

List price

$3,600

$5,100

$4,950

$4,695

Call factory for pricing

$4,300

$1,959

$1,200

Field calibration

Yes, and option to buy the calibration bench, including training

No

Yes

Yes, with anemometer

Unit contains a mass flow control, so it is not necessary to calibrate with each use

Kit available

Yes, comes with kit including rotameter, adapters, and tubing

No longer recommended; send back to recalibrate every 3 months

Direct impaction

Direct impaction

Direct impaction

Centrifugal impaction

Direct impaction

Direct impaction

Direct impaction

Direct impaction

Instrument Manufacturer Media

Type of sampler

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© 2009 Elsevier

Clinical Microbiology Newsletter 31:4,2009

rods, coagulase-positive staphylococci, molds, yeast, or other organisms which could be potentially fatal to the patient receiving the CSP) means that the compounding area must be thoroughly cleaned. One important criterion for determing who evaluates the plates is that the laboratory be either an accredited clinical or environmental laboratory able to identify both bacteria and fungi. Corrective action should include a re-evaluation of (i) personnel work practices, (ii) cleaning procedures, and (iii) a check of the efficiency of primary and secondary engineering controls. An investigation must be conducted to locate the source of the contamination, and the area must be re-sampled before compounding continues. Recently, one of our client pharmacies that is tested monthly had a spike in the number of CFU/m³ in one of their LAFW. The hood certifiers subsequently found a tear in the HEPA filter of the hood, although it had been re-certified just a month prior to the incident. Apparently some drilling had been done near the IV area to install a magnahelic gauge, and several high-speed particles had punctured the HEPA filter. If the monthly monitoring program had not been in place, it would have been another 5 months before the filter was recertified.

Surface Sampling Although the Centers for Disease Control and Prevention has long advised against surface sampling in hospitals, the USP has made surface sampling of primary and secondary engineering controls a requirement. The Sterile Expert Compounding Committee sees this as a necessary check on the decontamination of work surfaces and the success of a pharmacy’s chosen cleaning protocols. Surface sampling may be performed using contact (Rodac) plates or swabs. The medium should be TSApl, and results should be expressed as CFU/ contact plate surface area sampled (either cm² or in2). Recommended action levels for surface samples are shown in Table 6. Corrective action should include observing the cleaning of the IV room and the rotation of disinfectants.

Summary Although we have been told by a pharmacy director that he could comClinical Microbiology Newsletter 31:4,2009

Figure 1. rep PCR dendrogram and virtual gel image of Brevibacillus brevis strains 3031450 (lane 1) and 3031414 (lane 2). The dendrogram demonstrates 99.6 similarity.

pound a sterile preparation over “a bed of manure,” in fact, without scrupulous attention to aseptic technique, there is great danger of causing harm to patients. In spite of all of the primary and secondary engineering controls and in spite of all of the quality metrics that are in place, we have been able to find the same organism from a “sterile” CSP at the same time as it was recovered from the pharmacy’s buffer area. Figure 1 shows the results of the fingerprinting of both isolates using the automated Diversilab System (bioMérieux Clinical Diagnostics, Marcy L’Étoile, France). Fingerprinting was performed by Audrey Wanger and Mark Kruzel in the Department of Pathology, University of Texas Medical School, Houston, TX. The organism in this case was Brevibacillus brevis. It is tempting to speculate that sporadic outbreaks of Bacillus bacteremia may be caused by the introduction of Bacillus spores during infusion of supposedly sterile CSPs. It is our hope that by increasing understanding and advocating the implementation of best pharmacy practice guidelines as advocated in this article, clinical microbiologists can contribute to the reduction of both patient morbidity and mortality. References 1. Kastengo, E.S. 2008. USP chapter 具797典 and environmental monitoring. Clin. Microbiol. Newsl. 30:105-108. 2. U.S. Pharmacopeial Convention. 2004. Pharmaceutical compounding: sterile preparations, p. 2461-2477. In U.S. Pharmacopeia 27 – national formulary 22. U.S. Pharmacopeial Convention, Rockville, MD. 3. U.S. Pharmacopeial Convention. 2007. Revision bulletin, December 2007. Chapter 具797典 pharmaceutical com-

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Table 5. Recommended action levels of microbial contamination of aira Action limitb

ISO class 5

>1

7

>10

8 or worse

>100

a

Adapted from reference 3. b CFU per m³ (1,000 liters) of air per plate.

Table 6. Recommended action levels of microbial contamination on surfacesa Surface sampleb

ISO class 5

>3

7

>5

8 or worse

>100

a

Adapted from reference 3. b CFU/contact plate.

4.

5.

6.

7.

8.

pounding – sterile preparations. http://www.usp.org/pdf/EN/USPNF/ generalchapter797.pdf. Solomon, S.L. et. al. 1984. An outbreak of Candida parapsilosis bloodstream infection in patients receiving parenteral nutrition. J. Infect. Dis. 149:98-102. Hughes, C.F. et. al. 1986. Cardioplegia solution: a contamination crisis. J. Thorac. Cardiovasc. Surg. 91:296-302. Tresoldi, A.T. 2000. Enterobacter cloacae sepsis outbreak in a newborn unit caused by contaminated total parenteral nutrition solution. Am. J. Infect. Contr. 28:258-261. Centers for Disease Control and Prevention. 2002. Exophiala infection from contaminated injectable steroids prepared by a compounding pharmacy. MMWR Morb. Mortal. Wkly. Rep. 51:1109-1112. Selenic, D. et al. 2003. Enterobacter cloaecae bloodstream infections in pediatric patients traced to a hospital

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pharmacy. Am. J. Health Syst. Pharm. 60:1440-1446. 9. Centers for Disease Control and Prevention. 2005. Pseudomonas bloodstream infections associated with a heparin/saline flush – Missouri, New York, Texas, and Michigan. 2004-2005. MMWR Morb. Mortal. Wkly. Rep. 54:269-272. 10. Givens, R. et al. 2005. Outbreak of Serratia marsescens infection following injections of betamethasone compounded at a community pharmacy.

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Clin. Infect. Dis. 43:831-837. 11. Patel, R. et al. 2006. Hepatitis C virus infections from a contaminated radiopharmaceutical used in myocardial perfusion studies. JAMA 296:2005-2011. 12. Gresham, M.D. et al. 2008. Multistate outbreak of Pseudomonas fluorescens bloodstream infection after exposure to contaminated heparinized saline flush prepared by a compounding pharmacy. Clin. Infect. Dis. 47:1372-1379. 13. U.S. Food and Drug Administration. 2004. Guideline on sterile drug products

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produced by aseptic processing. FDA, Rockville, MD. http://www.fda.gov/ Cder/guidance/5882fnl.pdf. 14. U.S. Pharmacopeial Convention. 2004. Sterility tests, p. 2157-2162. In U.S. Pharmacopeia 27 – National formulary 22. U.S. Pharmacopeial Convention, Rockville, MD. 15. U.S. Pharmacopeial Convention. 2004. Bacterial endotoxins test, p. 2169-2173. In U.S. Pharmcopeia 27 — National Formulary 22. U.S. Pharmacopeial Convention, Rockville, MD.

Clinical Microbiology Newsletter 31:4,2009