Efficacy of rapid multiplex polymerase chain reaction for early diagnosis and treatment of pertussis

Background Pertussis, a highly infectious respiratory disease caused by Bordetella pertussis, causes airway inflammation and severe, persistent (lasting 2 weeks or more) characteristic whooping cough. In severe cases, complications such as atelectasis and bronchopneumonia may occur. Recently, the prevalence of pertussis has increased in South Korea due to reduced effect of the DTaP vaccination in infants as their age increased. Although culture is the gold standard test for diagnosis, polymerase chain reaction (PCR) method is most commonly used for diagnosis of pertussis due to the low sensitivity and long turnaround time of the culture method. Recently, a rapid multiplex PCR test has been introduced for comprehensive detection of respiratory pathogens (17 viruses and 3 bacteria), including Bordetella pertussis, with a turnaround time of 1 hour. This study aimed to investigate the efficacy of multiplex PCR for early diagnosis and treatment of pertussis. Methods We performed a retrospective study on patients with pertussis diagnosed from May 2017 to June 2019 at Yeungnam University Hospital. Nasopharyngeal swab specimens were tested using multiplex PCR. Medical records collected included data on age, sex, symptoms at the time of diagnosis, admission, hospitalization, isolation, vaccination history, past medical history, and accompanying diseases. Results A total of 27 patients were diagnosed with pertussis, nine (33.3%) of whom were men, with a median age of 48.9 years (3.3–82.2). Eleven (40.7%) had fever, 12 (44.4%) had dyspnea, 3 (11.1%) had paroxysmal cough, and 9 (33.3%) had inspiratory whooping. Seventeen (62.9%) and 24 (88.8%) patients had coughing for <8 days and ≤14 days, respectively. Median time from first symptom to diagnosis

All but one patient was prescribed macrolide antibiotics; all patients were isolated, with 22 (81.5%) requiring hospitalization. Three patients (11.1%) received ICU care for ventilation. All patients survived.
Conclusion A rapid multiplex PCR test can ensure early diagnosis, isolation, and treatment of pertussis. Testing of patients with respiratory symptom with multiplex PCR can led to early diagnosis of pertussis, proper treatment, and may help in outbreak control.

Background
Pertussis is a highly contagious disease that causes a wide range of clinical symptoms ranging from trivial to serious illnesses, resulting in death in young infants [1]. Pertussis spreads through respiratory droplets and can be transmitted by coughing, sneezing, or sharing breathing space for extended periods of time [2].
The most common side effect of pertussis is apnea followed by pneumonia, seizures, and encephalopathy. In more severe cases, pertussis can be life-threatening with refractory pulmonary hypertension and pneumothorax [1].
In South Korea, the government introduced the DTaP (diphtheria, tetanus, and acellular pertussis) vaccine in the National Immunization Program (NIP) in 1989.
During 1955-2011, the secular trend in pertussis incidence was characterized by a gradual decrease in the reported number of cases from 1955 to the late 1990s, followed by a recent increase from the early 2000s [3]. The pertussis vaccination is administered as three doses of basic vaccination with DTaP administered at 2, 4 and 6 months. The pertussis booster vaccination is administered at 15-18 months, at 4-6 years with DTaP, and at 11-12 years with Tdap. The Korean DTaP vaccination 5 coverage for infants under 36 months of age was 96-97% in 2017 [4]. Furthermore, the national immunization program budget is approximately 250 million dollars [5].
Despite these efforts, pertussis outbreaks occur every 3 years in South Korea and has recently increased [6]. Therefore, it is evident that the pertussis outbreak is not properly controlled. The reason for this includes non-immunization in adolescents and adults aged ≥ 15 years in a timely manner and underdiagnosis due to atypical symptoms in adolescents and adults [3,7]. Thus, such patients are the major cause of transmission within their families.
The gold standard diagnosis for pertussis is culture. Nevertheless, recently, polymerase chain reaction (PCR) is widely used because of the rapid diagnosis rate and high sensitivity. Particlularly, multiplex PCR (mPCR) is faster because different pathogens can be detected in only one trial. Therefore, this test is used for the diagnosis of many infectious disease including respiratory infectious disease [8][9][10], infectious gastroenteritis [11], and infections of the central nervous system such as meningitis and encephalitis [12]. This study was conducted retrospectively to investigate the efficacy of mPCR for the early diagnosis and treatment of pertussis.

Statistical analyses
Statistical analyses were performed using the SPSS 25.0 (IBM Corp., Armonk, N.Y., USA). Pearson's Correlation test was used for analyzing the relationship between age and cough duration. P-value less than 0.05 were considered statistically significant. results

Demographics and clinical symptoms
Twenty-seven patients, with a mean age of 48.9 (range 3.3-82.2) years and a male to female ratio of 1:2, were diagnosed with pertussis using mPCR. (Table 1). There  In mPCR, pathogens coinfected with pertussis were found in 10 patients, and AdV and HRV/EV were the most common organisms with (5, 18.5% each), followed by
Median time from first symptom to diagnosis was 9.0 (1-31) days. Twenty-four patients (81.5%) were diagnosed within 2 weeks. (Table 2). No significant differences between cough duration and age (p = 0.630). Pertussis has an incubation period of 7 to 10 days [13]. There was no significant difference in cough duration between pediatric adolescents and adults (Figure 3; p=0.711); the cough duration for patients diagnosed with pertussis was 7 days on median, 8.48 days on average, which was less than that in previous studies [6,7,15]. In this study, we retrospectively analyzed the clinical symptoms of patients diagnosed with pertussis. Only 9 patients (33.3%) showed CSP, which seemed to be the result of early diagnosis.

Treatment & Outcome of patients
In South Korea, according to the National Vaccination Management Guidelines, all citizens received three basic vaccinations with DTaP at 2, 4 and 6 months of age, three booster vaccinations at 15-18 months, with DTaP at 4 to 6 years of age; with Tdap at 11 to 12 years of age, followed by a dose of Td vaccine every 10 years [16].
Despite this high immunization rate, reports on the incidence of pertussis are increasing, and nonetheless, the Korean surveillance system for pertussis is likely to underestimate the burden of pertussis [17,18].
In addition, most adolescents do not receive regular booster vaccines; it has been reported that the probability of pertussis as a cause of chronic cough has increased [7]. Furthermore, pertussis infection in adolescents and adults has been found to be responsible for household transmission of pertussis to susceptible infants [19].
In the US, reported pertussis incidence has increased since the 1980s, with peaks every 2-5 years. In 2013, there were 28,639 cases of reported pertussis in the US and 13 pertussis-related deaths [20]. In the US, a claims database analysis study showed that considerable underreporting of pertussis in people aged under 50 years exists, especially with increasing age. Thus, it is necessary to develop public health programs to reduce the pertussis burden [21]. In our study, adolescents and adults infected with pertussis accounted for 85.2% of the total burden. Therefore, if the mPCR test using FA-RP is applied to patients in this age group, the outbreak of pertussis may be more effectively controlled.

Conclusions
In conclusion, testing of patients with respiratory symptoms through mPCR containing pertussis can lead to early diagnosis. Early diagnosis and proper treatment, may help in outbreak control of pertussis.

Competing interests
The authors declare that they have no competing interests.