Dataset Information

Automated image analysis with ImageJ of yeast colony forming units from cannabis flowers.

ABSTRACT: Currently, in the state of Colorado and all other states within the United States of America with legalized marijuana programs, testing is required for bacteria, yeast, and mold on marijuana products. The Code of Colorado Regulations, 1 CCR 212-1, considers a passing result when a 1?g sample contains <104 colony forming units (CFU) for the total yeast and mold count (TYMC). These measurements are usually obtained by manually counting colonies on petri-dishes or 3?M™ Petrifilms™, which is a time consuming and user subjective process. Therefore, an automated counting method utilizing ImageJ has been developed for CFU analysis of TYMC on Petrifilms. The performance of this colony counting method was demonstrated by comparing manual and automated counts from marijuana flower samples containing spikes of Candida albicans as well as samples that tested positive for the presence of yeast and mold. Fifteen images of Petrifilms showing various concentrations of colonies were studied by fifteen users at two institutions using both the automated and manual counting methods. All counts from the automated ImageJ procedure were within 12% of those obtained manually. In twelve out of fifteen Petrifilms, the average count of the automated method was statistically similar to the manual counts. The statistical differences of the other three samples were observed to be random and caused by user errors. The automated counting method could be used to quickly count numbers that are as high as 400 CFUs, reducing time of analysis with improved documentation because the images and the electronic colony counts can be saved on a computer or cloud for long term storage and data access.

SUBMITTER: Stolze N

PROVIDER: S-EPMC6746339 | biostudies-literature | 2019 Sep

REPOSITORIES: biostudies-literature

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Automated image analysis with ImageJ of yeast colony forming units from cannabis flowers.

Stolze Nicholas N Bader Carly C Henning Christopher C Mastin Jared J Holmes Andrea E AE Sutlief Arin L AL

Journal of microbiological methods 20190802

Currently, in the state of Colorado and all other states within the United States of America with legalized marijuana programs, testing is required for bacteria, yeast, and mold on marijuana products. The Code of Colorado Regulations, 1 CCR 212-1, considers a passing result when a 1 g sample contains <10<sup>4</sup> colony forming units (CFU) for the total yeast and mold count (TYMC). These measurements are usually obtained by manually counting colonies on petri-dishes or 3 M™ Petrifilms™, which ...[more]

PMID: 31381981

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Project description:Measuring the abundance of microbes in a sample is a common procedure with a long history, but best practices are not well-conserved across microbiological ﬁelds. Serial dilution methods are commonly used to dilute bacterial cultures to produce countable numbers of colonies, and from these counts, to infer bacterial concentrations measured in colony-forming units (CFUs). The most common methods to generate data for CFU point estimates involve plating bacteria on (or in) a solid growth medium and counting their resulting colonies or counting the number of tubes at a given dilution that have growth. Traditionally, these types of data have been analyzed separately using different analytic methods. Here, we build a direct correspondence between these approaches, which allows one to extend the use of the most probable number method from the liquid tubes experiments, for which it was developed, to the growth plates by viewing colony-sized patches of a plate as equivalent to individual tubes. We also discuss how to combine measurements taken at different dilutions, and we review several ways of analyzing colony counts, including the Poisson and truncated Poisson methods. We test all point estimate methods computationally using simulated data. For all methods, we discuss their relevant error bounds, assumptions, strengths, and weaknesses. We provide an online calculator for these estimators.Estimation of the number of microbes in a sample is an important problem with a long history. Yet common practices, such as combining results from different measurements, remain sub-optimal. We provide a comparison of methods for estimating abundance of microbes and detail a mapping between different methods, which allows to extend their range of applicability. This mapping enables higher precision estimates of colony-forming units (CFUs) using the same data already collected for traditional CFU estimation methods. Furthermore, we provide recommendations for how to combine measurements of colony counts taken across dilutions, correcting several misconceptions in the literature.

Project description:Medical cannabis (MC) production is a rapidly expanding industry. Over the past ten years, many additional phytocannabinoids have been discovered and used for different purposes. MC was reported beneficial for the treatment of a variety of clinical conditions such as analgesia, multiple sclerosis, spinal cord injuries, Tourette's syndrome, epilepsy, glaucoma, Parkinson disease and more. Yet, there is still a major lack of research and knowledge related to MC plant diseases, both at the pre- and postharvest stages. Many of the fungi that infect MC, such as Aspergillus and Penicillium spp., are capable of producing mycotoxins that are carcinogenic, or otherwise harmful when consumed, and especially by those patients who suffer from a weakened immune system, causing invasive contamination in humans. Therefore, there are strict limits regarding the permitted levels of fungal colony forming units (CFU) in commercial MC inflorescences. Furthermore, the strict regulation on pesticide appliance application in MC cultivation exacerbates the problem. In order to meet the permitted CFU limit levels, there is a need for pesticide-free postharvest treatments relying on natural non-chemical methods. Thus, a decontamination approach is required that will not damage or significantly alter the chemical composition of the plant product. In this research, a new method for sterilization of MC inflorescences for reduction of fungal contaminantstes was assessed, without affecting the composition of plant secondary metabolites. Inflorescences were exposed to short pulses of steam (10, 15 and 20 s exposure) and CFU levels and plant chemical compositions, pre- and post-treatment, were evaluated. Steam treatments were very effective in reducing fungal colonization to below detection limits. The effect of these treatments on terpene profiles was minor, resulting mainly in the detection of certain terpenes that were not present in the untreated control. Steaming decreased cannabinoid concentrations as the treatment prolonged, although insignificantly. These results indicate that the steam sterilization method at the tested exposure periods was very effective in reducing CFU levels while preserving the initial molecular biochemical composition of the treated inflorescences.

Dataset Information

Automated image analysis with ImageJ of yeast colony forming units from cannabis flowers.

Publications

Automated image analysis with ImageJ of yeast colony forming units from cannabis flowers.

Similar Datasets

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