Olfactory disorders are caused by various factors, such as sinonasal disease, head trauma, upper respiratory tract infections, and neurodegenerative diseases. Viral infections, particularly those caused by human rhinovirus, coronavirus, parainfluenza virus, and Epstein-Barr virus, are considered the leading cause of post-infection olfactory dysfunction [1-5]. The COVID-19 pandemic caused by SARS-CoV-2 is associated with temporary olfactory loss in about 53% of patients [6-10]. Even with the development of vaccines and treatments, newer variants, such as Omicron, still induce olfactory dysfunction in adults [11-16]. While some patients experience temporary parosmia or phantosmia, some patients develop a persistent olfactory disorder [17,18]. Although there are many potential treatments, little evidence indicates their effectiveness. In this regard, corticosteroids are commonly used, and the effective role of olfactory rehabilitation is also highlighted [19,20].

Traditional acupuncture is one of the oldest treatments in China, which has been used to treat patients for over 2,000 years [21]. Recent studies in China have reported that acupuncture may regulate microcirculation under physiological and pathological conditions [22]. Olfactory dysfunction might be related to the impaired movement of energies in the body, especially in the meridians associated with the nose and the olfactory system. Some studies suggested that acupuncture might help in olfactory dysfunction via the regeneration of olfactory neurons and attenuation of the inflammatory response of olfactory mucosa by regulating neuropeptides [22]. Therefore, acupuncture treatment for anosmia could be a hopeful option, even found to be effective in post-COVID-19 olfactory dysfunction, although it has not been sufficiently explored [23].

Auricular acupuncture is a simple, painless, and cost-effective treatment for neurological dysfunctions and has beneficial effects on olfactory dysfunction [24]. In the frequency acupuncture method, some points in the body or ear can be more accurately diagnosed and treated by using lasers and bioresonance frequencies and strengthening energies in several sessions. Thus, the present study aimed to investigate the role of frequency-controlled ear acupuncture in persistent olfactory dysfunction in COVID-19 patients through a clinical trial for the first time.

1. Study design, participants, and randomization

This single-center, participant-blind, randomized clinical trial with a 1:1 allocation ratio included individuals who suddenly lost their sense of smell during the COVID-19 outbreak and did not improve or had a slight improvement after 10-20 weeks. Other inclusion criteria were adults aged ≥ 18 years with a positive COVID-19 test result or a history of close contact with a confirmed COVID-19 case, patients without previous history of olfactory dysfunction, and those being able to provide informed consent. In our clinical trial registration, we originally planned to enroll 90 patients. However, due to unforeseen challenges, we recruited and analyzed data from 40 participants. Despite the smaller sample size, the design and methodology remained consistent with the registered protocol. Forty patients who were referred to the otorhinolaryngology department of the Hazrat Rasoul Hospital (20 and 20 patients in experimental and control groups, respectively) because of olfactory dysfunction underwent the Smell Identification Test. This test is reliable and valid for olfactory assessment among different ethnicities [25]. Moreover, for this subjective score, the participants were asked to rank their ability to smell before and after the intervention on a 10-point visual analog scale by marking the appropriate point between two extremes of anosmia to normal status [26].

Exclusion criteria comprised olfactory test scores > 20 (based on Table 1), history of any head surgery or head trauma, chronic and severe inflammatory diseases, degenerative diseases, abnormal nasal anatomy, and any treatment for COVID-19-induced olfactory dysfunction, such as intranasal spray, omega 3, or vitamin A nasal drops, except for betamethasone drops, in last two months. Although some conditions, including nasal allergy, were initially exclusion criteria, we later recognized the importance of including a diverse range of participants to enhance the external validity of our findings. Importantly, our subsequent analysis revealed no statistically significant differences in the prevalence of these conditions between intervention and control groups. This lack of difference assured us that the inclusion of such participants did not introduce bias into our study. In this participant-blind study, patients were randomly assigned to experimental and control groups by randomly selecting from two pots, one with participant numbers and one with either experiment or control group status. Operator blinding was not feasible. The Iran University of Medical Sciences (IR.IUMS.FMD.REC.1399.841) approved the study protocol. The trial was registered at the Iranian Registry of Clinical Trials (IRCT20210311050671N1). All participants provided informed consent before the study. The study protocol conformed to the ethical guidelines of the 1975 Declaration of Helsinki, as reflected in a priori approval by the institutional Human Research Committee.

Table 1

Diagnostic score of olfactory dysfunctions determined by Smell Identification Test [25].

N	Score (0-24 essential odors)	Olfactory status
1	19-24	Normal
2	16-18	Mild hyposmia
3	14-15	Moderate hyposmia
4	11-13	Severe hyposmia
5	Less than 11	Anosmia

2. Procedures

The Smell Identification Test contains 24 essential odors. Diagnosing 19-24 essential odors out of 24 is considered normal. If a patient recognizes 16-18, 14-15, and 11-13 essential odors, they suffer from mild, moderate, and severe hyposmia, respectively. Moreover, recognizing < 11 essential odors is diagnostic of anosmia (Table 1) [25]. In our study, patients with olfactory dysfunction after COVID-19 were included if their olfactory test scores were ≤ 20, although the questionnaire categorized scores > 19 as normal. The rationale behind this decision was to ensure a more focused enrollment of participants who subjectively reported a loss of smell post-COVID-19. We chose this threshold as it aligns with our aim to investigate the impact of the intervention on individuals who have experienced a substantial decline in olfactory function. Excluding patients with scores > 20 was deemed appropriate to prioritize those with a discernible olfactory impairment, enhancing the relevance and sensitivity of our study to the targeted population. The patients were treated for two sessions at a one-week interval, and their test results were evaluated after two sessions. Additionally, we could extend the treatment if necessary. Both groups were asked to use the nasal betamethasone drops (3 drops for each nasal cavity) 3 times daily for a maximum of one month. The control group included patients who received two sessions of switched-off lasers. The switched-off laser, during which patients wear safety glasses to prevent eye damage, could serve as a valid placebo in laser acupuncture studies without energy output [27]. Auricular microsystem acupuncture is a combination of Western and Eastern medicine, which has been justified by anatomical, embryological, and neurophysiologic pathways, describing how each point stimulation could influence another part of the body. In this study, acupuncture points on the auricle were selected in each session according to a study by Dr. Nogier and Dr. Soliman. They suggested some points that could play an important role in olfactory dysfunction with energy blockage [28]. Accordingly, we mostly identified the stagnation of energy movement among patients. The virus also caused a significant drop in the patient’s energy. The two diagnoses were the reasons for choosing the protocol type to determine treatment points according to the patient’s condition and a specific treatment protocol. The procedure was performed with RJ laser (Physiolaser Olympic model) by a trained acupuncturist. We stimulated the treatment points with a completely painless laser. Hence, the anxiety aroused by the fear of needles, or the possibility of infection was eliminated.

Acupuncture was performed in 2 sessions with a 1-week interval. Each session lasted 20 minutes and used stimulation by Izar laser with a diode radiation source, power of 400 mW, wavelength of 810, and specific frequency. You et al. [29] put together light wavelengths that share similar tissue optical properties and found that 808 nm falls under the category of 800 nm+, which is suitable for tissue penetration and was used in previous studies for neurological diseases. Therapeutic frequencies also varied based on the patient’s problem in each session. The amount of energy applied to each point was determined using the patient’s vascular autonomic reflux (VAS). VAS (changing pulse strength in a few pulses) is a physical phenomenon introduced by Paul Nogier in 1966, who noticed a decrease or an increase in radial pulses when he exercised a slight pressure on specific ear acupuncture points [30,31].

The use of the VAS signal enabled the development of interference field acupuncture and the so-called therapeutic “controlled acupuncture” or “frequency acupuncture,” in which the laser with specific frequencies is positioned under VAS control and not, as in the classical Chinese tradition, by other measurement units or particular points with fix position. In this study, 1622/22 Hz and solfeggio frequencies were mainly used.

3. Outcomes and statistical analysis

Given no previous research on how laser acupuncture can impact COVID-19-related olfactory dysfunction, it was not possible to calculate a sample size beforehand due to the unknown effect size. The study planned to recruit 40 participants as this is a reasonable number based on previous recruitment rates and is sufficient for an exploratory clinical trial [32]. While exploratory, our primary focus was on evaluating treatment effects to inform future larger trials. This trial serves as an essential initial step in assessing the potential therapeutic impact of laser acupuncture.

In addition to the descriptive statistics (i.e., percentage and mean), statistical tests, such as two independent sample t-tests, were used for comparing general characteristics. One-way analysis of covariance, two independent sample t-tests, and Paired sample T-tests were used for pre- and post-test olfactory scores. SPSS software version 22 was used to perform the statistical analysis of the recorded data. A p < 0.05 indicated statistical significance.

1. Baseline characteristics of the study sample

Forty patients (20 and 20 patients in experimental and control groups, respectively) with hyposmia and olfactory test scores ≤ 20 were included in the study (Fig. 1). The recruitment period spanned from May 2021 to July 2021. All patients underwent the second treatment session after one week in experimental and control groups, without patient loss in both groups. None of the participants had anosmia, and the control group encompassed 20 patients with the same condition as those in the experimental group. There was no statistically significant difference between the groups in terms of age, gender, underlying diseases, body mass index (BMI), and tobacco smoking based on two independent sample T-tests (Table 2).

Figure 1. The CONSORT diagram showing the flow of participants through each stage of a randomized trial.

*Allergy (22.2%), sinusitis (64.3%), multiple sclerosis (7.4%), rhinoplasty (7.4%), and hypothyroidism (7.4%). †Sinusitis (50%), rhinoplasty (12.5%), allergy (12.5 %), and asthma and allergy (25%)..

&md=tbl&idx=2' data-target="#file-modal"">Table 2

Baseline characteristics of the study participants.

Variables	Cases (20)	Controls (20)	p-value
Age (mean ± sd) (years)	31.95 ± 8.37	30.90 ± 7.09	0.671
Sex (n)	11 (Females)	13 (Females)	0.530
Medical condition (%)	70%*	40%†	0.580
Smoking (n)	12	10	0.593
BMI	24.07 ± 2.80	24.61 ± 3.10	0.563

*Allergy (22.2%), sinusitis (64.3%), multiple sclerosis (7.4%), rhinoplasty (7.4%), and hypothyroidism (7.4%). †Sinusitis (50%), rhinoplasty (12.5%), allergy (12.5 %), and asthma and allergy (25%)..

2. Participants’ hyposmia status before and after two intervention sessions in experimental and control groups

There was no statistically significant difference between the groups in terms of olfactory dysfunction before the intervention sessions based on paired sample T-tests. The comparison of intervention sessions revealed statistically significant differences based on two independent sample T-tests.

According to the results of the one-way analysis of covariance, there was a statistically significant difference between the adjusted means in experimental and control groups after two intervention sessions (F [37, 1] = 37.463; p = 0.000, Eta² = 0.503). According to the eta-squared coefficient, the intervention caused about 50% variation between the groups. Fig. 2 demonstrates the mean scores of hyposmia in both groups before and after treatment.

Figure 2. Effects of frequency-controlled ear acupuncture on COVID-19 induced olfactory dysfunction. After 2 sessions of treatment, scores on the smell identification test were significantly increased in the intervention group. The scores in the control group did not show any significant changes. Error bars are standard errors of the mean. *Before the treatment, there was no significant difference between the scores of both groups (p-value = 0.593). ***After the treatment, there was a significant difference (p-value < 0.001).

Moreover, the subjective visual analog score regarding the sense of smell before and after treatment indicated an improvement. The mean scores of patients significantly improved from 2.80 ± 1.76 to 5.22 ± 3.40 after treatment in the study group, but the control group did not show significant improvement from 4.20 ± 1.43 to 5.00 ± 2.22 (p = 0.007, 0.184, respectively).

For side effects, assessed during patients’ telephone follow-ups, only a small number of patients in the experimental group (3 patients) reported short and transient side effects, including nausea, headache, and dizziness, representing the side effects of laser treatment in the first hours after treatment.

The present study aimed to evaluate the efficacy of frequency-controlled ear acupuncture in treating patients with persistent post-COVID-19 olfactory dysfunction. The results revealed a significant difference between experimental and control groups in terms of recovery progress after two intervention sessions. As treatment continued, this difference became more pronounced, and the intervention caused a 50% variation between the groups.

Traditional Chinese acupuncture showed promising results in treating olfactory dysfunction induced by viral infections [21]. Our findings are in line with previous observations, showing that acupuncture may regulate microcirculation under physiological or pathological conditions. For example, Brandt et al. [33] reported a significant improvement in smell and taste after acupuncture treatment. A case report documented the effectiveness of acupuncture in treating anosmia [34]. Moreover, Vent et al. [35] used traditional Chinese acupuncture to treat 15 patients with post-viral olfactory dysfunction, 8 of whom experienced improved olfactory function, compared with only 2 patients treated with vitamin B complex. In line with these observations, Dai et al. [36] claimed that approximately one-third (32%) of patients with post-viral olfactory dysfunction exhibited improvement during 1 year, and longer durations between baseline and post-treatment testing were correlated with a higher probability of olfactory function improvement. They also found that 44% of the traditional Chinese acupuncture group experienced recovery in olfactory function within 3 months. The recovery rate in this study was higher than the 32% reported in the literature. However, observation time in reported studies was no longer than a year. Regarding auricular olfactory points, Tanaka et al. [22] stimulated the lung point with acupuncture on the cavum conchae, which is supplied by the auricular branch of the vagus nerve, influencing the olfactory sensory recognition. With traditional acupuncture, there is a slight risk of pain, bleeding, and infection. However, using a laser provides no risk, except for eye damage, which could be prevented by safety glasses.

The recovery rate in our study was higher than the rate reported in the literature for post-viral olfactory dysfunction. Although the frequency of ear acupuncture intervention caused a significant change in the olfactory recovery rate, it revealed relatively low patient satisfaction because there was no expected sense of satisfaction in terms of quality.

Olfactory training is a low-cost and effective therapy used to treat patients with post-infectious olfactory dysfunction [37]. Oral and intranasal corticosteroids are used to exclude an inflammatory component in patients with post-infectious olfactory dysfunction. The initiation of corticosteroid treatment may benefit post-infectious olfactory dysfunction induced by COVID-19 [38-40]. However, to date, there is no evidence indicating that other promising medications, such as intranasal sodium citrate, intranasal vitamin A, or systemic omega-3, are effective in patients with olfactory dysfunction induced by COVID-19 [37-41].

One of the strengths of this study is the research design (i.e., a clinical trial) evaluating the role of a non-invasive technique, such as frequency-controlled ear acupuncture, in the treatment of COVID-19-induced olfactory dysfunction. In addition to the smell identification test, we also compared patients with olfactory dysfunction before and after treatment. All patients in the experimental group received higher scores after treatment compared to baseline data. However, only 8 out of 20 patients in the control group received higher scores. Finally, according to the statistical analysis of the standard quantitative olfactory test results, the improvement rate showed a significant difference between the groups. Although the sample size was small, the cause of relative dissatisfaction with olfactory function could be associated with patients’ expectations. Furthermore, most participants claimed to have a powerful sense of smell before experiencing olfactory dysfunction. Out of 20 patients in the experimental group, 8 patients had phantosmia, 2 patients were completely cured, 1 patient experienced no change, and 5 individuals partially recovered. In addition to the small sample size, short-term follow-up is a limitation of this study.

In conclusion, our findings suggest that frequency-controlled ear acupuncture may be a promising therapy for the treatment of COVID-19-induced olfactory dysfunction. Future clinical trials with longer follow-ups and larger sample sizes are needed to confirm the clinical significance of these findings. Additionally, further investigations are required to determine the optimal frequency and duration of treatment, as well as to compare the efficacy of frequency-controlled ear acupuncture with other therapies, in post-COVID-19 olfactory dysfunction.

No funding sources.

All authors contributed to the conception or design of the work, drafting the article, and critical revision. Maryam Arab additionally contributed to data analysis.

The authors declare no conflict of interest.