滅菌/消毒過程中，例如，VHP消毒，指示劑出現(xiàn)陽性，一定是失敗了嗎？我們參考USP1221.11和PDA TR51，其中提及流氓 Bl，即BI出現(xiàn)出乎意料的陽性，可能會發(fā)生在經(jīng)過認證的生物指示劑批次中。如果檢測到陽性生物指示劑，并且已經(jīng)排除了所有可能的物理參數(shù)原因，則有可能是生物指示劑本身的可變性原因。使用重復樣品（指同一位置放置多個相同指示劑）可以減輕生物指示劑陽性的影響。

 

Addressing Anomalous Bl Results

 

處理異常的 Bl 結(jié)果

 

Any indicator system that relies on living organisms is liable to variation. The organisms themselves are inherently variable (genotypic factors) and are affected by external factors which lead to phenotypic Variation.

 

任何依賴于活體微生物的指示系統(tǒng)都容易發(fā)生變化。微生物本身本質(zhì)上是可變的（基因因素），并受到導致表型變異的外部因素的影響。

 

The Bl represents an integration of several biological, chemical, and physical factors, and it is inappropriate to make comparisons with simpler systems such as chemical indicating strips or temperature sensors, The Bl cannot be calibrated and is not a calibration device.

 

Bl 代表生物、化學和物理因素的綜合，不宜與化學指示劑或溫度傳感器等更簡單的系統(tǒng)進行比較，Bl 無法校準，也不是校準設(shè)備。

 

Rogue Bls (unexpected survivors) may occur in batches of certified Bls. In the event positive Bls are detected and all the cycle's physical parameters have been eliminated as possible causes, then it is necessary to investigate the basis of the variability of the Bls themselves. The impact of positive Bls may be mitigated by using duplicates (43).

 

流氓 Bl（出乎意料的陽性）可能會發(fā)生在經(jīng)過認證的生物指示劑批次中。如果檢測到陽性生物指示劑，并且已經(jīng)排除了所有可能的物理參數(shù)原因，則有必要調(diào)查生物指示劑本身的可變性原因。使用重復樣品（指同一位置放置多個相同指示劑）可以減輕生物指示劑陽性的影響 。

 

 

Investigation of Anomalous Results

 

異常結(jié)果的調(diào)查 

 

 

lf the process fails, an investigation should be conducted. This investigation may include, but is not limited to, the following:

 

如果過程失敗，則應(yīng)進行調(diào)查。這項調(diào)查可能包括但不限于以下內(nèi)容：

 

confirmation of cycle parameters

 

確認循環(huán)參數(shù)

 

review of the certificate of conformity supplied with the Bls and the results of in house test data

 

審查生物指示劑隨附的COA和內(nèi)部測試數(shù)據(jù)的結(jié)果

 

review of process records for compliance with relevant SOPs

 

審查程序記錄是否符合相關(guān) SOP

 

review of the certificate of conformity supplied with the sporicidal agent

 

審查殺孢子劑隨附的COA

 

review of equipment calibration data

 

審查設(shè)備校準數(shù)據(jù)

 

review of maintenance records

 

審查維護記錄

 

review of trends from previous cycles

 

審查前幾次滅菌的趨勢

 

review of temperature and chemical indicator data

 

審查溫度和化學指示劑數(shù)據(jù)

 

Then, if necessary, further modes of investigation should be pursued, including：

 

然后，如有必要，應(yīng)采取進一步的調(diào)查方式，包括：

 

re-running the cycle using a different lot of Bls

 

使用其他批次的Bl重新運行循環(huán)

 

usage of duplicate or triplicate Bls in the same location

 

在同一位置使用雙重或三重Bl

 

The investigation should also take into account previous cycles and a history of the site where the Bl failed as well as those of adjacent and similar sites. One investigational approach is to expose multiple Bls at the problem site(s) and determine the number of survivors. One hundred percent survival may indicate a change in cycle conditions; occasional or sporadic survival may indicate an issue involving variation in the Bls' resistance.

 

調(diào)查還應(yīng)考慮以前的周期和 Bl 失敗位置的歷史以及相鄰和相似位置的歷史。一種研究方法是在問題位置身上放置多個Bl 并確定陽性的數(shù)量。100% 陽性可能表明該循環(huán)條件發(fā)生了變化；偶爾或散發(fā)的陽性可能表明存在涉及 Bl 抗性變化的問題。

 

If the assessment method used to verify the specified kill level shows occasional random positive growth that is not repeated in the same test position, then it may be possible to argue that the results are due to rogue Bls. Ongoing random positives are an indication that the cycle is not robust and requires further investigation.

 

如果用于確認指定殺滅水平的評估方法顯示偶爾的隨機陽性生長，并且在同一測試位置沒有重復，那么有可能認為結(jié)果是由于流氓 Bl造成的。持續(xù)的隨機陽性表明該周期不穩(wěn)健，需要進一步調(diào)查。

 

Current industrial experience indicates that occasional positive Bls occur even in well defined cycles. Such rogue results may not be indicative of a failed cycle. Appropriate statistical methods may be used to support the acceptance of such rogue results in both primary validation and revalidation programs.

 

目前的工業(yè)經(jīng)驗表明，即使在明確的周期中，偶爾也會發(fā)生陽性 Bl。這種流氓結(jié)果可能并不表示循環(huán)失敗。可以使用適當?shù)慕y(tǒng)計方法來支持在主要驗證和重新驗證計劃中接受此類流氓結(jié)果。

 

Table-Factors That May Affect the Performance of Biological Indicators

 

可能影響生物指示劑性能的因素

 

Variation in the inoculum loaded on to the carriers (volume and spore count)

 

載體上的接種物的變化（體積和孢子數(shù)）

 

Purity of the inoculum on the carrier

 

載體上接種物的純度

 

Genotypic variability amongst the spore population

 

孢子種群中的基因型變異性

 

Aggregation of spores

 

孢子聚集

 

Presence of cel/media debris in the inoculum

 

接種物中細胞/介質(zhì)殘骸的存在

 

Orientation of carrier within primary package

 

內(nèi)包裝中載體的方向

 

Surface characteristics of the carrier

 

載體的表面特性

 

Handling of naked (i.e.., no packaging) Bls (aseptically, gloved hands, UAF Hood to avoid addition of oils or dirt that will occlude the inoculum)

 

BI（即拆開包裝后）的處理（無菌操作、戴手套、UAF層流罩以避免添加會影響接種物的油或污垢）

 

Composition of the primary packing material (when this is not removed during the cycle)

 

內(nèi)包裝材料（循環(huán)期間未去除）的成分

 

Storage conditions used for the Bis

 

BI的儲存條件

 

Equilibrium relative humidity of the spores on the carrier immediately prior to exposure to stress

 

載體上的孢子在暴露于壓力之前的平衡相對濕度

 

Method used to release spores from carriers when carrying out spore counts

 

進行孢子計數(shù)時用于從載體中釋放孢子的方法

 

Age of the Bis

 

BI的新鮮度

 

Retention of sterilant gases by the spore coat, carrier materials and primary packaging (and subsequent outgassing)

 

滅菌氣體在孢子涂層、載體材料和內(nèi)包裝上的殘留

 

Time between exposure and recovery of the Bls

 

從暴露到Bls回收之間的時間

 

Composition of the recovery medium, including the presence of substances that may stimulate/inhibit the germination and subsequent outgrowth of injured spores

 

回收培養(yǎng)基的組成，包括可能刺激/抑制受傷孢子發(fā)芽和隨后生長的物質(zhì)的存在

 

Table 2 Manufacturing Issues That May Affect Bacterial Spore Resistance to Physical and Chemical Stresses

 

表 2 可能影響細菌孢子物理和化學抵抗力的制造問題

 

Genetic complement-species/strain variation (Genotypic variability)

 

遺傳互補—物種/品系變異（基因型變異）

 

Composition of growth/sporulation medium (e.g., variation in concentration of divalent cations especially Mn++and Ca++)

 

生長/產(chǎn)孢培養(yǎng)基的組成（例如，二價陽離子，尤其是 Mn++ 和 Ca++ 濃度的變化）

 

Temperature of sporulation

 

產(chǎn)孢溫度

 

Growth limiting nutrient during sporulation

 

孢子形成過程中限制生長的營養(yǎng)物質(zhì)

 

Age of the spore

 

孢子的年齡

 

Temperature at which spore suspensions are stored and temperature fluctuations

 

孢子懸浮液的儲存溫度和溫度波動

 

pH of the suspending medium

 

懸浮介質(zhì)的 pH 值

 

Chemical composition of the suspending medium

 

懸浮介質(zhì)的化學成分