A Novel 16S rRNA PCR-Restriction Fragment Length Polymorphism Assay to Accurately Distinguish Zoonotic Capnocytophaga canimorsus and C. cynodegmi
ABSTRACT The zoonotic bacteria Capnocytophaga canimorsus and C. cynodegmi, the predominant Capnocytophaga species in the canine oral biota, can cause human local wound infections or lethal sepsis, usually transmitted through dog bites. Molecular surveying of these Capnocytophaga species using conven...
| Main Authors: | , , , , |
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| Format: | Article |
| Language: | English |
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American Society for Microbiology
2023-06-01
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| Series: | Microbiology Spectrum |
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| Online Access: | https://journals.asm.org/doi/10.1128/spectrum.02916-22 |
| _version_ | 1797803379214254080 |
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| author | Cheng-Hung Lai Yu-Sin Lin Chao-Min Wang Poa-Chun Chang Wei-Yau Shia |
| author_facet | Cheng-Hung Lai Yu-Sin Lin Chao-Min Wang Poa-Chun Chang Wei-Yau Shia |
| author_sort | Cheng-Hung Lai |
| collection | DOAJ |
| description | ABSTRACT The zoonotic bacteria Capnocytophaga canimorsus and C. cynodegmi, the predominant Capnocytophaga species in the canine oral biota, can cause human local wound infections or lethal sepsis, usually transmitted through dog bites. Molecular surveying of these Capnocytophaga species using conventional 16S rRNA-based PCR is not always accurate due to their high genetic homogeneity. In this study, we isolated Capnocytophaga spp. from the canine oral cavity and identified them using 16S rRNA and phylogenetic analysis. A novel 16S rRNA PCR-restriction fragment length polymorphism (RFLP) method was designed based on our isolates and validated using published C. canimorsus and C. cynodegmi 16S rRNA sequences. The results showed that 51% of dogs carried Capnocytophaga spp. Among these, C. cynodegmi (47/98, 48%) was the predominant isolated species along with one strain of C. canimorsus (1/98, 1%). Alignment analysis of 16S rRNA sequences revealed specific site nucleotide diversity in 23% (11/47) of the C. cynodegmi isolates, which were misidentified as C. canimorsus using previously reported species-specific PCR. Four RFLP types could be classified from all the isolated Capnocytophaga strains. The proposed method demonstrates superior resolution in distinguishing C. cynodegmi (with site-specific polymorphism) from C. canimorsus and especially in distinguishing C. canimorsus from other Capnocytophaga species. After in silico validation, this method was revealed to have an overall detection accuracy of 84%; notably, accuracy reached 100% in C. canimorsus strains isolated from human patients. Overall, the proposed method is a useful molecular tool for the epidemiological study of Capnocytophaga in small animals and for the rapid diagnosis of human C. canimorsus infections. IMPORTANCE With the increased number of small animal breeding populations, zoonotic infections associated with small animals need to be taken more seriously. Capnocytophaga canimorsus and C. cynodegmi are part of common biota in the mouths of small animals and can cause human infections through bites or scratches. In this study, C. cynodegmi with site-specific 16S rRNA sequence polymorphisms was erroneously identified as C. canimorsus during the investigation of canine Capnocytophaga by conventional PCR. Consequently, the prevalence of C. canimorsus is incorrectly overestimated in epidemiological studies in small animals. We designed a new 16S rRNA PCR-RFLP method to accurately distinguish zoonotic C. canimorsus from C. cynodegmi. After validation against published Capnocytophaga strains, this novel molecular method had high accuracy and could detect 100% of C. canimorsus-strain infections in humans. This novel method can be used for epidemiological studies and the diagnosis of human Capnocytophaga infection following exposure to small animals. |
| first_indexed | 2024-03-13T05:19:56Z |
| format | Article |
| id | doaj.art-3115a8bcdedc4dc9b417a456cbcd7521 |
| institution | Directory Open Access Journal |
| issn | 2165-0497 |
| language | English |
| last_indexed | 2024-03-13T05:19:56Z |
| publishDate | 2023-06-01 |
| publisher | American Society for Microbiology |
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| series | Microbiology Spectrum |
| spelling | doaj.art-3115a8bcdedc4dc9b417a456cbcd75212023-06-15T13:18:33ZengAmerican Society for MicrobiologyMicrobiology Spectrum2165-04972023-06-0111310.1128/spectrum.02916-22A Novel 16S rRNA PCR-Restriction Fragment Length Polymorphism Assay to Accurately Distinguish Zoonotic Capnocytophaga canimorsus and C. cynodegmiCheng-Hung Lai0Yu-Sin Lin1Chao-Min Wang2Poa-Chun Chang3Wei-Yau Shia4Department of Veterinary Medicine, College of Veterinary Medicine, National Chung Hsing University, Taichung, TaiwanDepartment of Veterinary Medicine, College of Veterinary Medicine, National Chung Hsing University, Taichung, TaiwanDepartment of Veterinary Medicine, National Chiayi University, Chiayi City, TaiwanGraduate Institute of Microbiology and Public Health, National Chung Hsing University, Taichung, TaiwanDepartment of Veterinary Medicine, College of Veterinary Medicine, National Chung Hsing University, Taichung, TaiwanABSTRACT The zoonotic bacteria Capnocytophaga canimorsus and C. cynodegmi, the predominant Capnocytophaga species in the canine oral biota, can cause human local wound infections or lethal sepsis, usually transmitted through dog bites. Molecular surveying of these Capnocytophaga species using conventional 16S rRNA-based PCR is not always accurate due to their high genetic homogeneity. In this study, we isolated Capnocytophaga spp. from the canine oral cavity and identified them using 16S rRNA and phylogenetic analysis. A novel 16S rRNA PCR-restriction fragment length polymorphism (RFLP) method was designed based on our isolates and validated using published C. canimorsus and C. cynodegmi 16S rRNA sequences. The results showed that 51% of dogs carried Capnocytophaga spp. Among these, C. cynodegmi (47/98, 48%) was the predominant isolated species along with one strain of C. canimorsus (1/98, 1%). Alignment analysis of 16S rRNA sequences revealed specific site nucleotide diversity in 23% (11/47) of the C. cynodegmi isolates, which were misidentified as C. canimorsus using previously reported species-specific PCR. Four RFLP types could be classified from all the isolated Capnocytophaga strains. The proposed method demonstrates superior resolution in distinguishing C. cynodegmi (with site-specific polymorphism) from C. canimorsus and especially in distinguishing C. canimorsus from other Capnocytophaga species. After in silico validation, this method was revealed to have an overall detection accuracy of 84%; notably, accuracy reached 100% in C. canimorsus strains isolated from human patients. Overall, the proposed method is a useful molecular tool for the epidemiological study of Capnocytophaga in small animals and for the rapid diagnosis of human C. canimorsus infections. IMPORTANCE With the increased number of small animal breeding populations, zoonotic infections associated with small animals need to be taken more seriously. Capnocytophaga canimorsus and C. cynodegmi are part of common biota in the mouths of small animals and can cause human infections through bites or scratches. In this study, C. cynodegmi with site-specific 16S rRNA sequence polymorphisms was erroneously identified as C. canimorsus during the investigation of canine Capnocytophaga by conventional PCR. Consequently, the prevalence of C. canimorsus is incorrectly overestimated in epidemiological studies in small animals. We designed a new 16S rRNA PCR-RFLP method to accurately distinguish zoonotic C. canimorsus from C. cynodegmi. After validation against published Capnocytophaga strains, this novel molecular method had high accuracy and could detect 100% of C. canimorsus-strain infections in humans. This novel method can be used for epidemiological studies and the diagnosis of human Capnocytophaga infection following exposure to small animals.https://journals.asm.org/doi/10.1128/spectrum.02916-22Capnocytophagaphylogenetic analysisdogsPCR-restriction fragment length polymorphismzoonosis |
| spellingShingle | Cheng-Hung Lai Yu-Sin Lin Chao-Min Wang Poa-Chun Chang Wei-Yau Shia A Novel 16S rRNA PCR-Restriction Fragment Length Polymorphism Assay to Accurately Distinguish Zoonotic Capnocytophaga canimorsus and C. cynodegmi Microbiology Spectrum Capnocytophaga phylogenetic analysis dogs PCR-restriction fragment length polymorphism zoonosis |
| title | A Novel 16S rRNA PCR-Restriction Fragment Length Polymorphism Assay to Accurately Distinguish Zoonotic Capnocytophaga canimorsus and C. cynodegmi |
| title_full | A Novel 16S rRNA PCR-Restriction Fragment Length Polymorphism Assay to Accurately Distinguish Zoonotic Capnocytophaga canimorsus and C. cynodegmi |
| title_fullStr | A Novel 16S rRNA PCR-Restriction Fragment Length Polymorphism Assay to Accurately Distinguish Zoonotic Capnocytophaga canimorsus and C. cynodegmi |
| title_full_unstemmed | A Novel 16S rRNA PCR-Restriction Fragment Length Polymorphism Assay to Accurately Distinguish Zoonotic Capnocytophaga canimorsus and C. cynodegmi |
| title_short | A Novel 16S rRNA PCR-Restriction Fragment Length Polymorphism Assay to Accurately Distinguish Zoonotic Capnocytophaga canimorsus and C. cynodegmi |
| title_sort | novel 16s rrna pcr restriction fragment length polymorphism assay to accurately distinguish zoonotic capnocytophaga canimorsus and c cynodegmi |
| topic | Capnocytophaga phylogenetic analysis dogs PCR-restriction fragment length polymorphism zoonosis |
| url | https://journals.asm.org/doi/10.1128/spectrum.02916-22 |
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