灭菌/消毒过程中，例如，VHP消毒，指示剂出现阳性，一定是失败了吗？我们参考USP1221.11和PDA TR51，其中提及流氓 Bl，即BI出现出乎意料的阳性，可能会发生在经过认证的生物指示剂批次中。如果检测到阳性生物指示剂，并且已经排除了所有可能的物理参数原因，则有可能是生物指示剂本身的可变性原因。使用重复样品（指同一位置放置多个相同指示剂）可以减轻生物指示剂阳性的影响。

 

Addressing Anomalous Bl Results

 

处理异常的 Bl 结果

 

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.

 

任何依赖于活体微生物的指示系统都容易发生变化。微生物本身本质上是可变的（基因因素），并受到导致表型变异的外部因素的影响。

 

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、化学和物理因素的综合，不宜与化学指示剂或温度传感器等更简单的系统进行比较，Bl 无法校准，也不是校准设备。

 

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（出乎意料的阳性）可能会发生在经过认证的生物指示剂批次中。如果检测到阳性生物指示剂，并且已经排除了所有可能的物理参数原因，则有必要调查生物指示剂本身的可变性原因。使用重复样品（指同一位置放置多个相同指示剂）可以减轻生物指示剂阳性的影响 。

 

 

Investigation of Anomalous Results

 

异常结果的调查 

 

 

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

 

如果过程失败，则应进行调查。这项调查可能包括但不限于以下内容：

 

confirmation of cycle parameters

 

确认循环参数

 

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

 

审查生物指示剂随附的COA和内部测试数据的结果

 

review of process records for compliance with relevant SOPs

 

审查程序记录是否符合相关 SOP

 

review of the certificate of conformity supplied with the sporicidal agent

 

审查杀孢子剂随附的COA

 

review of equipment calibration data

 

审查设备校准数据

 

review of maintenance records

 

审查维护记录

 

review of trends from previous cycles

 

审查前几次灭菌的趋势

 

review of temperature and chemical indicator data

 

审查温度和化学指示剂数据

 

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

 

然后，如有必要，应采取进一步的调查方式，包括：

 

re-running the cycle using a different lot of Bls

 

使用其他批次的Bl重新运行循环

 

usage of duplicate or triplicate Bls in the same location

 

在同一位置使用双重或三重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.

 

调查还应考虑以前的周期和 Bl 失败位置的历史以及相邻和相似位置的历史。一种研究方法是在问题位置身上放置多个Bl 并确定阳性的数量。100% 阳性可能表明该循环条件发生了变化；偶尔或散发的阳性可能表明存在涉及 Bl 抗性变化的问题。

 

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.

 

如果用于确认指定杀灭水平的评估方法显示偶尔的随机阳性生长，并且在同一测试位置没有重复，那么有可能认为结果是由于流氓 Bl造成的。持续的随机阳性表明该周期不稳健，需要进一步调查。

 

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.

 

目前的工业经验表明，即使在明确的周期中，偶尔也会发生阳性 Bl。这种流氓结果可能并不表示循环失败。可以使用适当的统计方法来支持在主要验证和重新验证计划中接受此类流氓结果。

 

Table-Factors That May Affect the Performance of Biological Indicators

 

可能影响生物指示剂性能的因素

 

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

 

载体上的接种物的变化（体积和孢子数）

 

Purity of the inoculum on the carrier

 

载体上接种物的纯度

 

Genotypic variability amongst the spore population

 

孢子种群中的基因型变异性

 

Aggregation of spores

 

孢子聚集

 

Presence of cel/media debris in the inoculum

 

接种物中细胞/介质残骸的存在

 

Orientation of carrier within primary package

 

内包装中载体的方向

 

Surface characteristics of the carrier

 

载体的表面特性

 

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（即拆开包装后）的处理（无菌操作、戴手套、UAF层流罩以避免添加会影响接种物的油或污垢）

 

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

 

内包装材料（循环期间未去除）的成分

 

Storage conditions used for the Bis

 

BI的储存条件

 

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

 

载体上的孢子在暴露于压力之前的平衡相对湿度

 

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

 

进行孢子计数时用于从载体中释放孢子的方法

 

Age of the Bis

 

BI的新鲜度

 

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

 

灭菌气体在孢子涂层、载体材料和内包装上的残留

 

Time between exposure and recovery of the Bls

 

从暴露到Bls回收之间的时间

 

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

 

回收培养基的组成，包括可能刺激/抑制受伤孢子发芽和随后生长的物质的存在

 

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

 

表 2 可能影响细菌孢子物理和化学抵抗力的制造问题

 

Genetic complement-species/strain variation (Genotypic variability)

 

遗传互补—物种/品系变异（基因型变异）

 

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

 

生长/产孢培养基的组成（例如，二价阳离子，尤其是 Mn++ 和 Ca++ 浓度的变化）

 

Temperature of sporulation

 

产孢温度

 

Growth limiting nutrient during sporulation

 

孢子形成过程中限制生长的营养物质

 

Age of the spore

 

孢子的年龄

 

Temperature at which spore suspensions are stored and temperature fluctuations

 

孢子悬浮液的储存温度和温度波动

 

pH of the suspending medium

 

悬浮介质的 pH 值

 

Chemical composition of the suspending medium

 

悬浮介质的化学成分