ASTM D8412-21
Standard Guide for Quantification of Microbial Contamination in Liquid Fuels and Fuel-Associated Water by Quantitative Polymerase Chain Reaction (qPCR)

Standard No.

ASTM D8412-21

Release Date

2021

Published By

American Society for Testing and Materials (ASTM)

Latest

ASTM D8412-21

Scope

1.1 This guide covers procedures for using quantitative polymerase chain reaction (qPCR), a genomic tool, to detect, characterize and quantify nucleic acids associated with microbial DNA present in liquid fuels and fuel-associated water samples. 1.1.1 Water samples that may be used in testing include, but are not limited to, water associated with crude oil or liquid fuels in storage tanks, fuel tanks, or pipelines. 1.1.2 While the intent of this guide is to focus on the analysis of fuel-associated samples, the procedures described here are also relevant to the analysis of water used in hydrotesting of pipes and equipment, water injected into geological formations to maintain pressure and/or facilitate the recovery of hydrocarbons in oil and gas recovery, water co-produced during the production of oil and gas, water in fire protection sprinkler systems, potable water, industrial process water, and wastewater. 1.1.3 To test a fuel sample, the live and recently dead microorganisms must be separated from the fuel phase which can include any DNA fragments by using one of various methods such as filtration or any other microbial capturing methods. 1.1.4 Some of the protocol steps are universally required and are indicated by the use of the word must. Other protocol steps are testing-objective dependent. At those process steps, options are offered and the basis for choosing among them are explained. 1.2 The guide describes the application of quantitative polymerase chain reaction (qPCR) technology to determine total bioburden or total microbial population present in fuelassociated samples using universal primers that allow for the quantification of 16S and 18S ribosomal RNA genes that are present in all prokaryotes (that is, bacteria and archaea) and eucaryotes (that is, mold and yeast collectively termed fungi), respectively. 1.3 This guide describes laboratory protocols. As described in Practice D7464, the qualitative and quantitative relationship between the laboratory results and actual microbial communities in the systems from which samples are collected is affected by the time delay and handling conditions between the time of sampling and time that testing is initiated. 1.4 The values stated in SI units are to be regarded as standard. No other units of measurement are included in this standard with the exception of the concept unit of gene copies/mL (that is, 16S or 18S gene copies/mL) to indicate the starting concentration of microbial DNA for the intended microbial targets (that is, bacteria, archaea, fungi). 1.5 This standard does not purport to address all of the safety concerns, if any, associated with its use. It is the responsibility of the user of this standard to establish appropriate safety, health, and environmental practices and determine the applicability of regulatory limitations prior to use. 1.6 This international standard was developed in accordance with internationally recognized principles on standardization established in the Decision on Principles for the Development of International Standards, Guides and Recommendations issued by the World Trade Organization Technical Barriers to Trade (TBT) Committee.

ASTM D8412-21 Referenced Document

• ASTM D1129 Standard Terminology Relating to Water
• ASTM D4175 Standard Terminology Relating to Petroleum Products, Liquid Fuels, and Lubricants*， 2023-07-01 Update
• ASTM D6469 Standard Guide for Microbial Contamination in Fuels and Fuel Systems
• ASTM D6974 Standard Practice for Enumeration of Viable Bacteria and Fungi in Liquid Fuels—Filtration and Culture Procedures
• ASTM D7464 Standard Practice for Manual Sampling of Liquid Fuels, Associated Materials and Fuel System Components for Microbiological Testing

ASTM D8412-21 history

• 2021 ASTM D8412-21 Standard Guide for Quantification of Microbial Contamination in Liquid Fuels and Fuel-Associated Water by Quantitative Polymerase Chain Reaction (qPCR)