Emergence of Novel Type C Botulism Strain in Household Outbreak, Japan

In 2021, an outbreak of food poisoning caused by Clostridium botulinum type C occurred in Kumamoto, Japan. Analysis of the isolated strain revealed that it possessed the bont/C gene and was slightly different from the reference bont/C gene. The risk for human infection with this new toxin type may be low.


Emergence of Novel Type C Botulism Strain in Household Outbreak, Japan
Rika Maeda, Misato Mori, Seiya Harada, Ichiro Izu, Takaaki Hirano, Yukie Inoue, Shunsuke Yahiro, Hiromi Koyama to date (1), and in Japan, only 1 infant botulism case caused by type C has been reported (2).C. botulinum group III is primarily known as an animal infection, and many of its toxin types have been reported as mosaic types (primarily in birds with toxin type CD and cattle with toxin type DC).
In 2021, foodborne botulism occurred in Kumamoto, Japan.A meal eaten in a domestic residence was the assumed cause, and 4 patients were affected.Botulinum toxin and C. botulinum were detected in 3 of the 4 specimens.A commercially prepared chicken dish was suspected to be the cause, but because no food was remaining, we were unable to conduct tests on it.We neutralized the toxin present in the specimens with type C botulinum antitoxin serum, and the isolated strain was found to carry the bont/C gene using PCR targeting the bont genes (3).Next-generation sequencing data revealed full-length coding regions of the bont In 2021, an outbreak of food poisoning caused by Clostridium botulinum type C occurred in Kumamoto, Japan.Analysis of the isolated strain revealed that it possessed the bont/C gene and was slightly different from the reference bont/C gene.The risk for human infection with this new toxin type may be low.
gene of the isolated strains (GenBank accession no.LC759602).The next-generation sequencing method was as follows: after treating C. botulinum with 20 mg/mL lysozyme in 20 mM Tris-HCl, 2 mM EDTA, and 1% Triton X-100 (pH 8.0), we extracted DNA using a QIAamp DNA Mini Kit (QIAGEN, https://www.qiagen.com).We prepared genomesequencing libraries using the QIAseq FX DNA Library Kit (QIAGEN) and sequenced the samples on the Illumina iSeq 100 (https://www.illumina.com).We analyzed sequencing data using CLC Genomics Workbench 22.0.2(QIAGEN).The obtained contig was assembled from reads of 59× coverage and 29 kbp in size.In comparison with other known bont genes, the bont gene of the strain sequenced in this study had the highest amino acid sequence similarity with the bont/C gene (90%) but was partially different from the reference bont/C gene (Table).Detailed analysis revealed that the bont gene (LC759602) had the bont/C gene or the bont/CD gene in the protease domain (LC) and the translocation domain (H N ), as well as the bont/DC gene in the receptor-binding domain (H C ) (Table ;  The H C domain is involved in neurotoxin binding to specific receptors in peripheral nerve terminals.The bont gene (LC759602) possesses the bont/ DC gene in the H C domain, suggesting that human susceptibility to this gene might differ from that of the reference BoNT/C toxin.Unlike other BoNTs, BoNT/C interacts only with gangliosides, and no protein receptor for this toxin has been identified (4).However, BoNT/DC has been reported to interact with gangliosides and protein receptors (synaptotagmin I and II) (5).
The bont gene (LC759602) was determined to be BoNT/C using PCR, which can easily distinguish between types C, D, CD, and DC of C. botulinum group III (6).It should be noted that, because not all type C strains were subjected to sequencing, the presence of the bont gene (LC759602) as type C, as determined by typing PCR, might already exist in other samples.Further investigation is needed to determine the proportion of C. botulinum carrying the bont gene reported in this study.The risk for human infection with this new toxin type should also be investigated in I n February 2023, a 55-year-old man sought care at the emergency department of Vancouver General Hospital, Vancouver, British Columbia, Canada, for daily fevers, headache, and abdominal pain 5 days after returning from a 3-week trip to the Philippines.He stayed mostly in Manila but spent 4 days on Palawan Island in the western Philippines 4 days before his return to Canada; he had not taken malaria chemoprophylaxis.Bloodwork was notable for platelet nadir of 48 × 10 9 /L (reference range 150-450 × 10 9 /L), alanine transaminase of 329 U/L (reference range 10-55 U/L), and alkaline phosphatase of 177 U/L (reference range 30-135 U/L).Results of abdominal computed tomography were unremarkable and of a single-target Plasmodium falciparum histidine-rich protein 2 rapid diagnostic test were negative.Peripheral blood thin smear demonstrated variable intraerythrocytic parasite morphology, including band-like forms suggestive of P. malariae (<0.1% parasitemia) (Figure , panels A-C).Loopmediated isothermal amplification testing was Author affiliations: Kumamoto Prefectural Institute of Public Health and Environmental Science, Uto, Japan (R. Maeda, M. Mori, S. Harada, I. Izu, T. Hirano, Y. Inoue, S. Yahiro); Northern Kumamoto Administrative Headquarters Kamoto Area Development Bureau, Yamaga, Japan (H.Koyama) DOI: https://doi.org/10.3201/eid2910.230433B otulism is a neuroparalytic disease caused by the botulinum toxin, which is produced by Clostridium botulinum.C. botulinum is physiologically divided into groups I-IV, and botulinum neurotoxins (BoNT) are classified into 7 types, BoNT/A-G.Human botulism is caused primarily by toxin types A, B, and E, and cases of human infection with C. botulinum group III, which produces toxin types C and D, are rare.Only 5 foodborne botulism outbreaks caused by C. botulinum group III (4 outbreaks caused by type C and 1 outbreak caused by type D) have been reported Figure).The bont gene (LC759602) has not been previously reported, and we propose its designation as a new subtype of C. botulinum toxin.
research.However, given that human infections with a similar toxin type, C. botulinum group III, have rarely occurred, this new toxin type might pose little threat to human health.
2176 Emerging Infectious Diseases • www.cdc.gov/eid•Vol.29,No.10,October 2023 RESEARCH LETTERS *BoNT, botulism neurotoxin.future Emerging Infectious Diseases • www.cdc.gov/eid• Vol. 29, No. 10, October 2023 2177 RESEARCH LETTERS A 55-year-old man sought treatment for an uncomplicated febrile illness after returning to Canada from the Philippines.A suspected diagnosis of Plasmodium knowlesi infection was confirmed by PCR, and treatment with atovaquone/proguanil brought successful recovery.We review the evolving epidemiology of P. knowlesi malaria in the Philippines, specifically within Palawan Island.