Metagenomic analysis of isolation methods of a targeted microbe, Campylobacter jejuni, from chicken feces with high microbial contamination
Abstract Background Originating from poultry, particularly chickens, Campylobacter jejuni is the leading foodborne pathogen worldwide and a major cause of campylobacteriosis. Isolating C. jejuni is difficult due to its specific growth requirements, the presence of viable but non-culturable bacteria,...
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| Format: | Article |
| Language: | English |
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BMC
2019-04-01
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| Series: | Microbiome |
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| Online Access: | http://link.springer.com/article/10.1186/s40168-019-0680-z |
| _version_ | 1819198727006453760 |
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| author | Junhyung Kim Jae-Ho Guk Seung-Hyun Mun Jae-Uk An Hyokeun Song Jinshil Kim Sangryeol Ryu Byeonghwa Jeon Seongbeom Cho |
| author_facet | Junhyung Kim Jae-Ho Guk Seung-Hyun Mun Jae-Uk An Hyokeun Song Jinshil Kim Sangryeol Ryu Byeonghwa Jeon Seongbeom Cho |
| author_sort | Junhyung Kim |
| collection | DOAJ |
| description | Abstract Background Originating from poultry, particularly chickens, Campylobacter jejuni is the leading foodborne pathogen worldwide and a major cause of campylobacteriosis. Isolating C. jejuni is difficult due to its specific growth requirements, the presence of viable but non-culturable bacteria, and because it is often masked by competing flora. Currently, there is no optimized method for isolating C. jejuni from chicken feces. Here, we evaluated the method for isolating C. jejuni from chicken feces using culture-independent sequence-based metagenomics and culture-dependent tools. Further, we assessed changes in microbial communities during microbe isolation to determine how the process can be improved. Results Fourteen different variations of C. jejuni isolation procedures were applied to all 35 chicken fecal samples. These variations included using different enrichment broths (without enrichment or enrichment in Bolton or Preston broth), different ratios of sample-to-enrichment broth (1:101, 1:102, and 1:103), and different selective agars (modified charcoal-cefoperazone-deoxycholate agar (mCCDA) or Preston agar). Enrichment during isolation of C. jejuni was evaluated on the basis of microbial diversity and taxonomic composition using metagenomics tools. The effect of selective media was evaluated using a combination of metagenomics and culture-dependent tools. Microbial diversity significantly decreased during the enrichment process, regardless of the type of enrichment broth, with the most significant decrease observed at a feces-to-broth ratio of 1:103. Particularly, in 103-Preston broth, the relative abundance of Campylobacter increased, while extended-spectrum beta-lactamase-producing Escherichia coli, which interfere with Campylobacter isolation, decreased. Metagenomics results were validated by quantitative PCR and culture-dependent analysis. Additionally, selective media affected the isolation results, although microbes with high relative abundance during enrichment were also frequently isolated using culture-dependent methods. Significantly more C. jejuni was isolated from mCCDA than from Preston agar enriched in 103 Preston broth. Conclusions Enrichment in Preston broth at a ratio of 1:103 followed by spreading onto mCCDA was the most effective method for isolating C. jejuni. This is the first study to apply metagenomics to evaluate a method for isolating a targeted microbe, C. jejuni, from chicken feces, a source with high microbial contamination. Thus, metagenomics can be applied to improve methods for isolating bacteria that are difficult to separate. |
| first_indexed | 2024-12-23T03:05:02Z |
| format | Article |
| id | doaj.art-04af74d96cf0411ab7e9edb5b4a6bb81 |
| institution | Directory Open Access Journal |
| issn | 2049-2618 |
| language | English |
| last_indexed | 2024-12-23T03:05:02Z |
| publishDate | 2019-04-01 |
| publisher | BMC |
| record_format | Article |
| series | Microbiome |
| spelling | doaj.art-04af74d96cf0411ab7e9edb5b4a6bb812022-12-21T18:02:19ZengBMCMicrobiome2049-26182019-04-017111610.1186/s40168-019-0680-zMetagenomic analysis of isolation methods of a targeted microbe, Campylobacter jejuni, from chicken feces with high microbial contaminationJunhyung Kim0Jae-Ho Guk1Seung-Hyun Mun2Jae-Uk An3Hyokeun Song4Jinshil Kim5Sangryeol Ryu6Byeonghwa Jeon7Seongbeom Cho8Research Institute for Veterinary Science and College of Veterinary Medicine, Seoul National UniversityResearch Institute for Veterinary Science and College of Veterinary Medicine, Seoul National UniversityResearch Institute for Veterinary Science and College of Veterinary Medicine, Seoul National UniversityResearch Institute for Veterinary Science and College of Veterinary Medicine, Seoul National UniversityResearch Institute for Veterinary Science and College of Veterinary Medicine, Seoul National UniversityDepartment of Food and Animal Biotechnology, Department of Agricultural Biotechnology, Research Institute for Agriculture and Life Sciences, and Center for Food and Bioconvergence, Seoul National UniversityDepartment of Food and Animal Biotechnology, Department of Agricultural Biotechnology, Research Institute for Agriculture and Life Sciences, and Center for Food and Bioconvergence, Seoul National UniversityDepartment of Food and Animal Biotechnology, Department of Agricultural Biotechnology, Research Institute for Agriculture and Life Sciences, and Center for Food and Bioconvergence, Seoul National UniversityResearch Institute for Veterinary Science and College of Veterinary Medicine, Seoul National UniversityAbstract Background Originating from poultry, particularly chickens, Campylobacter jejuni is the leading foodborne pathogen worldwide and a major cause of campylobacteriosis. Isolating C. jejuni is difficult due to its specific growth requirements, the presence of viable but non-culturable bacteria, and because it is often masked by competing flora. Currently, there is no optimized method for isolating C. jejuni from chicken feces. Here, we evaluated the method for isolating C. jejuni from chicken feces using culture-independent sequence-based metagenomics and culture-dependent tools. Further, we assessed changes in microbial communities during microbe isolation to determine how the process can be improved. Results Fourteen different variations of C. jejuni isolation procedures were applied to all 35 chicken fecal samples. These variations included using different enrichment broths (without enrichment or enrichment in Bolton or Preston broth), different ratios of sample-to-enrichment broth (1:101, 1:102, and 1:103), and different selective agars (modified charcoal-cefoperazone-deoxycholate agar (mCCDA) or Preston agar). Enrichment during isolation of C. jejuni was evaluated on the basis of microbial diversity and taxonomic composition using metagenomics tools. The effect of selective media was evaluated using a combination of metagenomics and culture-dependent tools. Microbial diversity significantly decreased during the enrichment process, regardless of the type of enrichment broth, with the most significant decrease observed at a feces-to-broth ratio of 1:103. Particularly, in 103-Preston broth, the relative abundance of Campylobacter increased, while extended-spectrum beta-lactamase-producing Escherichia coli, which interfere with Campylobacter isolation, decreased. Metagenomics results were validated by quantitative PCR and culture-dependent analysis. Additionally, selective media affected the isolation results, although microbes with high relative abundance during enrichment were also frequently isolated using culture-dependent methods. Significantly more C. jejuni was isolated from mCCDA than from Preston agar enriched in 103 Preston broth. Conclusions Enrichment in Preston broth at a ratio of 1:103 followed by spreading onto mCCDA was the most effective method for isolating C. jejuni. This is the first study to apply metagenomics to evaluate a method for isolating a targeted microbe, C. jejuni, from chicken feces, a source with high microbial contamination. Thus, metagenomics can be applied to improve methods for isolating bacteria that are difficult to separate.http://link.springer.com/article/10.1186/s40168-019-0680-zMetagenomicsMicrobial community analysisIsolation methodCampylobacter jejuni |
| spellingShingle | Junhyung Kim Jae-Ho Guk Seung-Hyun Mun Jae-Uk An Hyokeun Song Jinshil Kim Sangryeol Ryu Byeonghwa Jeon Seongbeom Cho Metagenomic analysis of isolation methods of a targeted microbe, Campylobacter jejuni, from chicken feces with high microbial contamination Microbiome Metagenomics Microbial community analysis Isolation method Campylobacter jejuni |
| title | Metagenomic analysis of isolation methods of a targeted microbe, Campylobacter jejuni, from chicken feces with high microbial contamination |
| title_full | Metagenomic analysis of isolation methods of a targeted microbe, Campylobacter jejuni, from chicken feces with high microbial contamination |
| title_fullStr | Metagenomic analysis of isolation methods of a targeted microbe, Campylobacter jejuni, from chicken feces with high microbial contamination |
| title_full_unstemmed | Metagenomic analysis of isolation methods of a targeted microbe, Campylobacter jejuni, from chicken feces with high microbial contamination |
| title_short | Metagenomic analysis of isolation methods of a targeted microbe, Campylobacter jejuni, from chicken feces with high microbial contamination |
| title_sort | metagenomic analysis of isolation methods of a targeted microbe campylobacter jejuni from chicken feces with high microbial contamination |
| topic | Metagenomics Microbial community analysis Isolation method Campylobacter jejuni |
| url | http://link.springer.com/article/10.1186/s40168-019-0680-z |
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