Microbiological Test Data - Assuring Data Integrity
微生物測試數據-確保數據的完整性
Edward Charles Tidswell and Tim Sandle
PDA Journal of Pharmaceutical Science and Technology 2017,
Access the most recent version at doi:10.5731/pdajpst.2017.008151
Microbiological Test Data – Assuring Data Integrity
微生物測試數據-確保數據的完整性
Abstract 摘要
Marketed drugs and devices possess specifications including critical microbiological quality attributes purposed to assure efficacy and patient safety.
已上市的藥物和器械具有包括旨在確保療效和患者安全的關鍵微生物質量屬性在內的標準。
These attributes are legislated requirements intended to protect the recipient patient.
這些屬性是立法規定的要求，旨在保護受體病人。
Sampling, microbiological testing, interpretation of data for final products, raw materials and intermediates all contribute to a cohesive assessment in the assurance of finished product quality.
抽樣、微生物檢測、最終產品、原材料和中間體數據的解釋都有助於對最終產品質量的一致性評估。
Traditional culture-based microbiological methods possess inherent and unavoidable variability, recognized by the compendia and which might lead to erroneous conclusion pertaining to product quality.
傳統的基於培養的微生物學方法具有固有的和不可避免的可變性，這一點已被藥典所承認，並可能導致與產品質量有關的錯誤結論。
Such variability has been associated and intrinsically linked with data integrity issues; manufacturers have subsequently been encouraged by regulatory authorities to introduce multiple microbiologists or checks to prevent such issues.
這種可變性與數據完整性問題相關聯，並在本質上聯繫在一起；隨後，監管部門鼓勵生產商引入多種微生物學家或檢查方法，以防止此類問題的發生。
Understanding microbiological variability is essential such that genuine data integrity issues are identified.
了解微生物的可變性是至關重要的，這樣才能確定真正的數據完整性問題。
Furthermore, a range of meaningful preventative strategies are feasible beyond increasing the capacity of the quality control microbiological laboratory.
此外，除了增加質量控制微生物實驗室的能力之外，一系列有意義的預防策略是可行的。
This short review describes the legislative requirements, inherent microbiological variability and realistic actions and activities that genuinely assure patient safety.
本篇簡短的綜述描述了立法要求、內在的微生物多樣性以及真正確保患者安全的現實行動和活動。
Key Words關鍵詞
Microbiological tests, data integrity, microbiological quality attributes, sterility test, bioburden test
微生物測試，數據完整性，微生物質量屬性，無菌測試，生物負載測試
Introduction 簡介
The assessment of the microbiological quality of pharmaceutical finished drug products and devices is mandated through legislated requirements (21 CFR Parts 211 and 800 in the US and Volume 4 of Eudralex in the EU).
通過立法規定，對藥品成品和器械的微生物質量進行評估。
These laws are purposed to help assure the safety of the recipient patient population from potential microbiological adulteration of drugs and devices.
這些法律的目的是幫助確保接受治療的患者群體的安全，避免潛在的微生物摻假藥物和器械。
Such adulteration may be regarded as adventitious contamination manifested as the non- sterility of items purported to be sterile or the possession of an item with a certain quantity of microorganisms (bioburden) or type of microorganism (objectionable) occurring within items marketed as non-sterile.
這種摻假可被視為不定之症汙染，表現為聲稱無菌的物品不無菌，或擁有在作為非無菌商品銷售的物品中含有一定數量的微生物(生物負載)或微生物類型(特定微生物)。
The actual technical, microbiological criteria for sterile and non-sterile products are regarded as microbiological quality attributes and defined in the pharmaceutical compendia recognized by the relevant Health Authority.
無菌和非無菌產品的實際技術、微生物學標準被視為微生物質量屬性，並在相關衛生部門認可的藥典中進行了定義。
It is against these standards by which legislated requirements governing a finished product or device are assessed and determined to be fit for market.
這是違反這些標準的立法要求，管理的成品或器械是評估和確定適合市場。
Similarly the pharmaceutical compendia detail the reference or referee method or analytical test by which a drug or device are sampled, tested and assessed against the microbiological criteria (or offer routes through which alternative methods can be adopted).
類似地，藥典詳細說明了參照或仲裁方法或分析測試，通過這種方法，藥物或器械根據微生物學標準進行取樣、測試和評估(或提供可採用其他方法的途徑)。
Assurance of the microbiological quality of an item is not solely based upon end product sampling, testing and assessment; rather a complete cohesive and coordinated plan of process inputs (raw materials, additives, processing aids), process intermediates (including process steps such as intermediate hold) and environmental sampling, testing and assessment.
確保一項產品的微生物質量並不僅僅基於最終產品的抽樣、測試和評估;而是一個完整的、連貫的、協調的過程投入(原材料、添加劑、加工助劑)、過程中間體(包括中間貨艙等過程步驟)和環境取樣、測試和評估的計劃。
It is the complete, integrated and scientifically principled management of sampling, testing and assessment that provides the entire microbiological data which permits the disposition of product meeting its licensed label claim.
是對抽樣、測試和評估的完整、綜合和科學原則的管理，提供了完整的微生物數據，使產品符合其許可標籤要求的處置。
This complete end-to-end cohort of data for each batch of finished drug product or device is also legislated through the cGMPs (for example 21 CFR Parts 211.110, and 211.113).
每批成品或器械的完整端到端隊列數據也通過cGMPs進行立法(例如21 CFR第211.110和211.113部分)。
It is the combination of legislated requirements, established microbiological attributes,
sampling, testing and analysis which are purposed to help assure patient safety.
它是立法要求、已確立的微生物屬性、為確保病人安全而進行的抽樣、測試和分析。
Patient safety is the fundamentally crucial guiding principle and ethic in the provision of pharmaceutical finished drug products and devices.
在提供藥物成品和器械方面，患者安全是根本的關鍵指導原則和倫理。
Industry and regulators are entirely aligned in this, and what could be more unethical and paradoxical than the provision of harmful therapies supplied to treat patients.
行業和監管機構在這一點上完全一致，而且可能比為病人提供有害療法更不道德、更自相矛盾。
Equally the provision of safe therapies and assurance of patient safety in terms of microbiological quality is benefited with recognition of the following:
同樣，由於認識到下列情況，為病人提供安全的治療和保證病人的微生物質量也大有裨益:
Microbiological data does not per se materially control or modulate the bioburden or the
sterility of non-sterile products or products purported to be sterile.
微生物學數據本身並不能在實質上控制或調節生物群落非無菌產品或聲稱無菌的產品的無菌性。
Historically legislation has primarily relied upon data acquired from an analytical test as the proof of safety for the patient.
從歷史上看，立法主要依靠從分析測試中獲得的數據作為患者安全的證明。
This is understandable given the evidence-based foundation of judicial systems supporting legislation.
鑑於司法系統支持立法的證據基礎，這是可以理解的。
In actuality the assurance of the microbiological quality of products is better served with an emphasis upon design and control (as set out in International Conference on Harmonization guidance Q8 Pharmaceutical development; Q9 Quality risk management; Q10 pharmaceutical quality system; Q11 Development and manufacture of drug substances).
事實上，設計和控制是保證產品微生物質量的關鍵（如ICHQ8藥物開發；Q9質量風險管理；Q10藥品質量體系；Q11藥物的開發和製造）
Microbiological data has value; however this is only one part of the overall design, controls, checks and balances, necessary for assuring microbiological product quality.
微生物數據有價值;然而，這只是整體設計、控制、檢查和平衡的一部分，是保證微生物產品質量所必需的。
Acquisition of microbiological data is reliant upon a sample upon which a particular test is
performed.
微生物數據的獲取依賴於特定測試所依賴的樣本執行。
Meaningful microbiological data providing supporting evidence that a well- designed process is in control and that the end product meets the required microbiological attribute can only be gained if a representative sample is obtained.
只有獲得具有代表性的樣品，才能獲得有意義的微生物數據，這些數據提供了有力的證據，證明精心設計的過程處於控制之中，最終產品滿足所需的微生物屬性。
Samples must not only be authentic of the material tested, that is free from contamination during the activity of sampling but removed at a location and time that serves the purpose of the test (process control or material meeting a microbiological attribute).
樣品不僅必須是所測試材料的真實樣品，在取樣過程中不受汙染，而且必須在符合測試目的的地點和時間(過程控制或符合微生物特性的材料)被移除。
Typically a 『worst case』 time for sampling is at the end of any process hold period.
通常，採樣的「最壞情況」是在任何過程保持期的末尾。
Additionally the sample must be representative of material and account for any lack of homogeneity.
此外，樣品必須具有代表性的材料和解釋任何缺乏同質性。
An end-to-end integrated sampling plan based upon a genuine understanding of microbiological risk and control is the means to achieve this (Tidswell, 2017).
基於真正理解微生物風險和控制的端到端集成採樣計劃是實現這一目標的手段(Tidswell, 2017)。
Each sample is subject to a particular test methodology that is reliant upon a suitable method
development, qualification, and accompanying controls.
每個樣本都要遵循特定的測試方法，該方法依賴於合適的方法開發、確認和伴隨的控制。
Execution of the test is highly dependent upon the adequacy of the associated standard operating procedures (SOP) and the acumen of the microbiologist performing the method.
測試的執行高度依賴於相關標準操作規程(SOP)的充分性和微生物學家執行方法的敏銳性。
Finally once a sample has been tested the test is inspected and microbiological data is interpreted in order to make the microbiological assessment.
最後，一旦樣品被測試，測試是檢查和微生物數據的解釋，以便作出微生物評估。
Traditional microbiological test methods invariably rely upon the visual inspection of a liquid or solid medium and either the enumeration of colony forming units (CFUs), an assessment of the presence/absence of CFUs or the presence of turbidity in liquid media.
傳統的微生物測試方法總是依賴於對液體或固體培養基的目視檢查，以及菌落形成單位(CFUs)的計數，對CFUs的存在/不存在或液體培養基中是否存在渾濁度的評估。
The CFU is an arbitrary estimate at best since the only cells able to form colonies are those that can grow under the conditions of the test.
CFU充其量是一個武斷的估計，因為唯一能夠形成菌落的細胞是那些在測試條件下能夠生長的細胞。
These test conditions include: incubation temperature; incubation time; type of culture media; oxygen conditions and so on.
這些測試條件包括:培養溫度;培養時間;培養基類型;氧氣條件等等。
A subsequent recorded number of colony forming units (CFU) are not raw data but rather the
CFU is the microbiologist’s interpretation of the number of colonies on the plate (Sutton, 2012).
隨後記錄的菌落形成單元數(CFU)不是原始數據，而是CFU是微生物學家對平板上菌落數量的解釋(Sutton, 2012)。
This interpretation is affected by the number of colonies present, where there is too high a number leads to counting errors as a result of confluence or overcrowding across the surface of the plate; and where the number is too low error arises because the CFU’s follow Poisson distribution.
這種解釋受到當前菌落數量的影響，菌落數量過高會導致由於板塊表面的融合或過度擁擠而導致計數錯誤;由於CFU遵循泊松分布，所以誤差很小。
Of late several competent regulatory authorities have made inspectional observations of firms that are applying the microbiological test in alignment to the legislated cGMPs.
最近，幾個主管管理當局對正在按照立法的cGMPs進行微生物檢驗的公司進行了檢查。
Those observations note that a single microbiologist is visually inspecting the test and making an interpretation of the test.
這些觀察結果表明，一個微生物學家正在視覺上檢查測試並對測試進行解釋。
For example, there have been regulatory comments where colony forming units have been miscounted.
例如，有監管部門的評論指出，菌落形成單位的數量被錯誤計算。
This can arise due to inappropriate light or magnification; a failure to use a suitable counting device; the incorrect multiplication of a dilution and so on.
這可能是由於不適當的光或放大；未能使用適當的計數裝置;稀釋倍數的錯誤乘法，等等。
These observations have been categorized as data integrity issues.
這些觀察結果被歸類為數據完整性問題。
It has thus become popular to suggest that deployment of multiple microbiologists is an effective and expedient preventative action of microbiological data integrity issues.
因此，建議部署多個微生物學家是微生物數據完整性問題的有效和方便的預防措施，這已成為流行的觀點。
At first glance this may appear the most appropriate rectification; however this in itself is not a ubiquitous solution, may not be necessary and may in actuality prevent realization of our common goal of reduced patient risk, and maximized patient safety.
乍一看，這似乎是最恰當的糾正，然而，這本身並不是一個普遍存在的解決方案，可能沒有必要，而且實際上可能會阻礙我們實現降低患者風險和最大化患者安全的共同目標。
Our necessary history of evidence-based judicial systems and legislation founded upon indisputable and uncontestable data (laws of evidence or the rule of evidence) has helped to serve our industry well, however it does not match well with microbiological data with its well- recognized inherent variability (Sutton, 2011a).我們建立在無可爭議的數據(證據法或證據規則)基礎上的以證據為基礎的司法系統和立法的必要歷史有助於我們的行業更好地服務，然而，由於其固有的可變性，它與微生物數據並不匹配(Sutton, 2011a)。
Variability can be divided in to two categories (Singer and Sutton, 2011):
可變性可分為兩類(Singer和Sutton, 2011):
a) 「Avoidable」 variability (variability due to poor practice (Sutton 2010))
「可避免的」變異性(由於實踐不當造成的變異性(Sutton 2010))
b) Inherently unavoidable variability (variability due to limitations of the methods and the vagaries of dealing with biological samples (Jarvis, 1989).
固有的不可避免的變異性(由於方法的限制和處理生物樣本的反覆無常而產生的變異性)(Jarvis, 1989)。
Assurance of microbiological quality is not primarily achieved through test data.
微生物質量的保證主要不是通過測試數據來實現的。
It frequently appears confoundlingly difficult to reconcile the constraints of microbiological tests, and interpretative assessment of those tests with the precise details of design control, and sampling.
要調和微生物試驗的限制條件，以及對這些試驗的解釋性評估與設計控制和抽樣的精確細節之間的矛盾，常常顯得非常困難。
There is a need to ensure the data collected and purposed to demonstrate process control and product achieving microbiological quality attributes is as accurate as feasible, however there are many alternative means to achieve this other than multiple microbiologists performing the inspection and assessment.
有必要確保所收集和用於演示過程控制和產品實現微生物質量屬性的數據儘可能準確可行，然而，除了多個微生物學家進行檢查和評估之外，還有許多其他方法可以實現這一目標。
This short review is purposed to illustrate the inherent variability of culture based microbiological test methods (which cannot be reduced with any significance) and provide the range of options to ensuring the avoidance of potential data integrity issues.
這篇簡短的綜述旨在說明基於培養的微生物測試方法的固有可變性(不能有任何意義地降低這種可變性)，並提供了確保避免潛在數據完整性問題的選項範圍。
Microbial Quality Attributes
微生物質量屬性
All finished drug products and medical devices must be fit for use; granted marketing licenses based upon appropriate specifications, in combination with well-designed and controlled manufacturing processes conforming to the legislated cGMPs.
所有成品藥品和醫療器械必須適合使用;根據適當的規範，結合符合法規cGMPs的精心設計和控制的生產過程，授予銷售許可證。
For finished drug products this means a therapy must have appropriate characteristics of safety, identity, strength, purity, and quality.
對於成品藥物，這意味著治療必須具有適當的安全性、同一性、強度、純度和質量特徵。
Microorganisms (viable and non-viable), their products, or their inherent biochemical and physiological activity on or within therapies may adversely impact safety, strength, purity and quality.
：微生物(活的和不活的)、它們的產品，或它們在治療上或治療內固有的生化和生理活性可能對安全性、強度、純度和質量產生不利影響。
Unlike an inanimate chemical or physical impurities or contaminants, the animate and dynamic nature of microorganisms posse a far greater risk to fitness for use and ultimately patient safety.
與無生命的化學或物理雜質或汙染物不同，微生物的生命和動態特性對適宜使用和最終患者的安全構成了更大的風險。
In a certain quantity certain species of microorganisms may cause infection when introduced through the vehicle of a therapy, metabolically active microorganisms may transform a drug products formulation resulting in chemical changes, or changes in efficacy.
在一定數量的情況下，某些種類的微生物在通過治療載體引入時可能引起感染，代謝活性微生物可以改變藥物的配方，導致化學變化，或功效的變化。
Lastly the vestiges of microorganisms or sloughed off cell well and cell membrane may cause pyrogenic or toxic effect.
最後，微生物的殘留或脫落的細胞和細胞膜良好，可能導致發熱或毒性作用。
The microbial requirements of finished product specifications are therefore arguably the most important and are often underappreciated in their potential to adversely affect the recipient patient population.
因此，成品規範中的微生物要求可以說是最重要的，但由於其潛在的對受體患者群體產生不利影響的可能性，這些要求常常被低估。
Concomitantly pharmacopoeial compendia are very clear in stating the necessary microbial specifications of finished drug products.
與此同時，藥典在說明成品的必要微生物規範方面也非常清楚。
Sterility is required for all parenteral products are stipulated in by the United States Pharmacopeia (USP), (<71> Sterility tests), Pharmacopoeia Europa (EP), (2.6.1 Sterility), and Japanese Pharmacopoeia (JP), (4.06 Sterility test).
美國藥典(USP)、(<71>無菌試驗)、歐洲藥典(EP)、(2.6.1無菌)、日本藥典(JP)、(4.06無菌試驗)均規定注射用藥物產品必須無菌。
Non-sterile finished drug product are permitted a level of bioburden which is based upon the dosage form and route of administration and stipulated in USP (<1111> Microbiological examination of non-sterile products: acceptance criteria for pharmaceutical preparations and substances), EP (2.6.12 Microbiological examination of non-sterile products: total; viable aerobic count) and JP (4.05 Microbial limit test).
在USP（<1111>），EP（2.6.12），JP（4.05）中，非無菌成品按規定的劑型和給藥途徑允許有一定的生物負載。
The potential impact of microorganisms on or within therapies is undisputable and clearly represents a source of risk to patient safety; however it is imperative that we recognize the precise specifications for bioburden as described by the industry standard pharmacopeia.
微生物對治療或治療內的潛在影響是毋庸置疑的，並清楚地代表了對患者安全的風險來源;然而，我們必須認識到的準確規範的生物負載所描述的行業標準藥典。
USP <1111>, EP 2.6.12 EP and JP 4.05 list out the maximum limits of bioburden defined as total aerobic microbial counts (TAMC) and total yeast and mold count (TYMC) within different dosage forms of drug products; categorized as 10 CFU, 100CFUCFU and 1000CFU.
USP <1111>， EP 2.6.12 EP和JP 4.05列出了不同劑型藥物中生物負載的最大限值，生物負載定義為總需氧微生物計數(TAMC)和總酵母和黴菌計數(TYMC);分為10 CFU, 100CFU和1000CFU。
Each pharmaceutical compendia subsequently qualifies the interpretation of these maximum bioburden limits; the three harmonized compendia (USP, EP, JP) all stating:
每個藥典隨後都有確認解釋這些最大生物負載限度;三個協調案(USP, EP, JP)均聲明:
「When an acceptance criterion for microbiological quality is prescribed, it is interpreted as follows:
「規定微生物質量驗收標準時，解釋為:
101cfu: maximum acceptable count = 20;
102cfu: maximum acceptable count = 200;
103 cfu: maximum acceptable count = 2000;
and so forth.」
All compendia provide due and appropriate recognition to the inherent variability of microbiology and microorganisms; the expectation is that the expert microbiologist appropriately interprets the data.
所有的藥典都對微生物學和微生物的固有變異性提供了適當的認識;期望微生物學家專家能恰當地解釋這些數據。
A 200% leeway is permissible which is essentially the acceptance of a two-fold difference or variability in the enumeration of CFUs.
允許有200%的偏差，這本質上是接受CFUs計數中的雙重差異或變異性。
This variability is necessary because of the inherent dynamic nature of microorganisms, microbial populations and the means by which samples are handled and tested by culture-based methods.
這種變化是必要的，因為微生物的固有動態性質、微生物種群以及用培養方法處理和測試樣品的方法。
The same is true for process inputs (raw materials, additives, processing aids), process intermediates (including process steps such as intermediate hold) and environmental testing; enumeration to established microbial quality attributes is an important component of quality assurance however there is recognized variability.
工藝輸入(原料、添加劑、助劑)、工藝中間體(包括中間貨艙等工藝步驟)和環境試驗也是如此;對已建立的微生物質量屬性的枚舉是質量保證的一個重要組成部分，但也存在公認的可變性。
In addition to microbial quality attributes the pharmacopoeial compendia detail the standardized methodology of testing a sample purposed to enumerate bioburden or establish a presence or absence.
除了微生物質量屬性外，藥典還詳細說明了用於列舉生物樣本或確定存在或不存在的樣本檢測的標準化方法。
These methods are recognized as the referee or reference method which a firm needs to optimize and qualify for each of their sample formulations assuring the recovery of any resident culturable (using the media and conditions stated) microorganisms; the microbiology quality control (QC) laboratory tests.
這些方法被認為是裁判或參考方法，公司需要對其每一種樣品配方進行優化和確認，以確保回收任何可培養的(使用所述的培養基和條件)微生物;微生物學質量控制(QC)實驗室檢測。
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