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Background: In Corynebacterium diphtheriae, the dtxR gene plays a role in regulating diphtheria toxin synthesis. The dtxR gene is often used as a marker for identifying C. diphtheriae by the polymerase chain reaction (PCR) method because it is present in all strains of this bacterium. Mutations in the dtxR gene can cause the over-synthesis of diphtheria toxin and reduce PCR assays' sensitivity. Objectives: This study aimed to describe the polymorphisms in the dtxR gene of C. diphtheriae isolated from a diphtheria outbreak in Indonesia. Methods: Forty-eight isolates of C. diphtheriae were obtained from clinical samples (throat/nasopharyngeal swabs) of diphtheria cases and close contacts. The isolates were revived on a blood agar plate (BAP), bacterial colonies were harvested, and deoxyribonucleic acid (DNA) was extracted. The DNA sequencing was carried out using a Whole-genome Sequencing (WGS) approach. The data were converted and analyzed with U-gene software. The dtxR gene analysis was performed with C. diphtheriae PW8 as references. Results: There were 59-point mutation locations in 48 isolates examined. None of these single nucleotide polymorphisms (SNPs) coded for amino acid changes. Based on the mutation pattern, seven clades/groups of the dtxR gene of 48 C. diphtheriae isolates were examined. Conclusions: At least seven types of DNA sequences and more than 50 SNPs of the dtxR gene were identified in 48 C. diphtheriae isolates from a diphtheria outbreak in Indonesia. Although all of them are silent mutations, they must be considered in the design of PCR examination in diphtheria laboratories.

Background: Corynebacterium diphtheriae, C. ulcerans, and C. pseudotuberculosis are known as diphtheria-causing bacteria. Although diphtheria therapy is administered based on the clinical manifestations, some cases are mild and atypical. The immediate and accurate identification of diphtheria-causing bacteria is of paramount importance to prevent the spread of the disease and provide case management as early as possible. Unfortunately, conventional methods as the gold standard are time-consuming. Objectives: This study aimed to develop and implement a multiplex real-time PCR with the dtxR and tox genes as the target to identify three species of diphtheria-causing bacteria and screen their toxigenicity quickly and accurately. Methods: The research sample encompassed seven reference strains, one synthetic DNA, 30 archived isolates, and 924 clinical specimens isolated from 311 diphtheria cases and 613 close contacts. The conventional methods as the gold standard and the established PCR assay were used to verify the results of multiplex real-time PCR developed in this study. Results: The multiplex real-time PCR could identify seven reference strains, one synthetic DNA, and 30 archived isolates as accurately as the conventional methods and the established PCR. Similar to established PCR, the multiplex real-time PCR identified diphtheria-causing bacteria in 120 (38.6%) out of 311 and 12 (2%) out of 613 clinical specimens from diphtheria cases and close contacts, respectively. Meanwhile, the conventional methods identified diphtheria-causing bacteria in 79 (25.4%) out of 311 and three (0.5%) out of 613 clinical specimens. Conclusions: The multiplex real-time PCR developed in this study can be used to identify three species of diphtheria-causing bacteria and screen their toxigenicity quickly and accurately. However, in this study, no diphtheria-causing bacteria other than C. diphtheriae was found in the clinical samples using the PCR or conventional methods. PCR is more sensitive than the conventional methods and can be used as an additional test in diphtheria laboratories.

Pertussis cases have been reported most frequently in developed countries, but they are predicted to be the most prevalent in developing countries. Indonesia, a developing country, routinely conducts case-based surveillance for pertussis. We reviewed the data on pertussis cases and close contacts based on clinical sample documents examined in the National Reference Laboratory for pertussis, Indonesia (2016–2020). Our objective was to analyze the laboratory and epidemiological aspects of pertussis cases and close contacts, particularly to evaluate the implementation of a 5-year case-based surveillance of pertussis in Indonesia. Data were collected from sample documents and annual laboratory reports between January 2016 and December 2020. We analyzed the proportion of pertussis cases and close contacts by geographic region, year, age, and sex. We used the χ2 test to correlate the laboratory and epidemiological data. In total, 274 clinical cases of pertussis and 491 close contacts were recorded in 15 provinces. The peak number of cases occurred in 2019, with a positivity rate (percentage of laboratory-confirmed cases) of 41.23% (47/114). Clinical cases were dominated by infants aged p = 0.00). Clinical and confirmed cases of pertussis occurred mostly in the early age group and may be lower in those aged ≥10 years, especially in confirmed cases. New policies are needed for pertussis prevention at an early age, as well as the application of serology tests to increase laboratory-confirmed cases in children aged ≥10 years.