A Survey of Human Face Mites Demodex (Acari, Demodicidae) in Patients with Dermatological Symptoms in Baghdad, Iraq

Demodex spp. mites are external obligate parasites; they are transmitted between hosts through direct contact, and may induce several dermatological symptoms when found in large numbers. However, these symptoms may be similar to other commonly known diseases; this often leads dermatologists to neglect the pathogenic role of these mites. Therefore, a better diagnosis is recommended in order to avoid mistreatment. The aim of this study was to investigate the correlation between Demodex mites and dermatological diseases. Infestation rates in patients suffering from acne, rosacea, folliculitis, and psoriasis were compared with asymptomatic patients, along with the mites’ relation to gender, age, personal hygiene, time of year, and residency. These mites were mainly found in patients with rosacea (66.67%) followed by acne (64.41%), while they were only observed in 48.15% of asymptomatic patients, 40.00% in folliculitis, and 0.00% in psoriasis. The statistical analysis showed a significant relationship between infestation and the previously mentioned dermatological diseases (P<0.01). The infestation rates increased significantly with age, reaching 83.33% in patients above 60 years old, and especially during cold months with higher humidity rates. These mites were also more predominantly found in urban areas (56.10%), followed by rural areas (48.45%). However, there was no significant relationship according to gender or personal hygiene.


Introduction
Demodex spp. mites are obligate universal ectoparasites found within the human pilosebaceous units, they feed on nutrients and dead cells from hair roots, and may damage cell walls [1]. Demodex folliculorum (Simon), 1842, lives inside hair follicles, while Demodex brevis Akbulatova, 1963, lives within the secretory ducts of sebaceous glands, along with the Meibomian glands [2]. These mites are unable to survive without a host for extended periods of time, thus they are known to be transmitted between hosts via direct contact. These mites were detected in almost all races and ages with an estimated colony of 1000 to 2000 mites on each human [3,4]. Their infestation is believed to be in correlation with dermatological symptoms, but it is still unclear if their increased numbers are the cause or the result of the inflammation [5]. The mere presence of these mites is usually asymptomatic, however, symptoms may occur when the host suffers from immunosuppression [6], or when they are present in great densities within a small area of the skin [7]. Infestation of the skin (more than 5 mites/cm 2 ) may be associated with symptoms such as itching, pustules, dryness, nodulocystic lesions, erythema, and inflammation [8]. Their movement within the skin pores may result in irritation combined with increased oil production, which could trigger a vicious cycle of sensitive skin in patients suffering from rosacea [9]. The optimal method for collecting and counting these mites so far is the repeated extraction using Standardized Surface Skin Biopsy (SSSB) from their habitat inside the pilosebaceous units and then examining them under microscope [10].

Materials and Methods
During the period from November 2018 to June 2019, clinical specimens were collected from 220 patients suffering from dermatological diseases. The selected patients comprised 86 males, and 134 females, within different age groups ranging from 10 to 70 years old, with both symptomatic and asymptomatic cases. All studied cases were diagnosed and confirmed by specialized experienced dermatologists, and were collected from different hospitals in Baghdad, Iraq; Al-Imamain Al-Kadhimain Medical City, Al-Yarmuk Teaching Hospital, Al-Amirat Private Hospital, Al-Shahid Al-Hakim General Hospital, Al-Karama Teaching Hospital, and a private clinic in Al-Kadhimya district, in addition to The Department of Biology/College of Science/University of Baghdad. Written informed consent was obtained from all patients, and their information was recorded in a questionnaire sheet. The skin of patients was cleaned using a cotton pad soaked with alcohol. After it was allowed to dry, a drop of alpha-cyanoacrylate adhesive (super glue) was placed on a slide, spread into an area of 1 cm 2 , then pressed onto the skin. After 1-2 min, the glass slide was gently removed [11]. Figure 1 presents the standard procedure for the Standardized Surface Skin Biopsy (SSSB). An SSSB sample was collected from each patient, and the collected skin samples were then examined under microscope [11]. For each month, air temperature (T) and humidity (H) ranges in the city of Baghdad were recorded, as shown in Table 1.

Sample examination
All collected specimens were examined immediately under compound light microscope (10× and 40× magnifications). A drop of normal saline was added to enhance the visibility of the mites. Different life stages of Demodex spp. were observed and classified based on phenotypic characteristics. The classification was confirmed by specialists in The Iraqi Natural History Museum and Research Center in the University of Baghdad, Baghdad, Iraq.

Results
The prevalence of Demodex spp. mites was observed in 232 patients, 81 were asymptomatic, 118 suffered from acne, 22 suffered from psoriasis, 6 suffered from rosacea, and 5 suffered from folliculitis. The statistical analysis of these results, based on the infestation rates in each case, showed that these mites were predominantly found in patients with rosacea (66.67%), followed immediately by acne (64.41%). However, these mites were not observed in any of the patients suffering from psoriasis, and were only seen in 48.15% of asymptomatic patients and 40.00% of patients with folliculitis. Also, a significant relationship was found when compared statistically (P<0.01). According to gender, these mites were present in females (55.22%) at a higher percentage than in males (47.67%). However, there was no significant difference when compared statistically. In relation to age, these mites were consistently found in elderly patients in the age group of 61~70 (83.33%), followed by 51~60 (66.67%), 21~30 (61.47%), and 31~40 (59.46%). While it was rarely observed in young patients in the age groups of 10~20 (21.95%), followed by the age 41~50 (33.33%). Significant statistical relationship was found between infestation rates and age (P<0.01). Based on hygiene of patients, it was noticed that the group with < 4 washes/day showed a slightly higher percentage (46.67%), while the ≥ 4 washes/day group showed a slightly lower percentage (41.00%). However, both percentages were too close and no significant relationship was found when compared statistically. As for the mites' presence during the months of the year, these mites were mostly found in December (60.00%), followed by February (57.14%), June (57.14%), May (55.56%), November (51.92%), and January (50.00%). However, the lowest occurrence rates were recorded in March (44.12%), followed by April (44.44%), (P<0.05), (P<0.01). In total, it was found in (52.73%) of dermatological infestations in Baghdad, Iraq. However, not all hospitals and districts were studied. Statistical analysis revealed that there was a significant relationship between the prevalence of Demodex mites in dermatological diseases with the variations in temperature (P<0.05) and humidity (P<0.01) specified by the different seasons. Also, in relation to the type of residency, these mites were mostly found in urban areas (56.10%), followed by rural areas (48.45%). However, there was no significant relationship between infestation rates and residency when compared statistically. Figure 2 presents the prevalence percentages of Demodex mites in skin according to dermatological diseases, personal hygiene, gender, residency, age, and their presence during the months of year, in Baghdad, Iraq.

Discussion
Statistical analysis of the observed data revealed that there was a significant relationship between the presence of Demodex mites and the previously mentioned dermatological diseases (Rosacea, Acne and Folliculitis) (P<0.01). These result are in agreement with the previously published studies, where Demodex mites were significantly found in patients with rosacea [9], acne-like demodicosis [12], and folliculitis [8]. However, this study is in disagreement with the outbreak of mites reported in patients with psoriasis [13], which may be due to the patients being undergoing different treatment methods other than phototherapy. The relation between gender and Demodex infestation found in this study was also in agreement with previous studies where no significant correlation was found [9,14]. However, some studies found a higher incidence rate in females [15]. These differences may be related to the health state, skin type, geography, or heredity of the individual patient, causing a fluctuation in the prevalence rates [16,17], along with the difference in sebaceous gland morphology between genders and their sebum production rates [18]. Previous literature also confirmed the significant correlation between Demodex infestation and age [9,14,15]. This increase may be explained by the fact that elderly tissues contain mature follicles and glands that are fully developed, which in turn may provide the mites with the required nutrition [9], along with the fact that they are relatively immuno-compromised [5,19]. On the other hand, it is important to point out the noticeable low prevalence rates in the age group of 41~50 years, along with the low rates found in the 10~20 years group. This may indicate that Demodex prevalence could be affected by the hormonal changes. Patients in these groups are either undergoing puberty or menopause, meaning that estrogen and androgen levels are fluctuating, which are known to target the sebaceous glands and control their size and sebum secretion [20,21]. Personal hygiene was found to have no significant relationship with infestation rates. This agrees with previously published studies [9,14,22], despite the common belief where bad hygiene was considered suitable for the infestation of these mites. However, few studies suggested that Demodex prevalence was higher in patients with a low level of personal hygiene [23,24]. This disagreement of findings may be explained by the fact that the majority of the studied patients washed their faces with water only and did not use soap in all washes, which may not have cleaned the face efficiently from excess oils and mites.
The relationship between the mites' infestation rates and time of year may indicate that the mites are positively affected by low temperatures and high humidity, and may be predominantly found during winter in Iraq; this is in agreement with previous literature conducted on laboratory cultures [14,25]. However, it is recommended to study the changes of humidity and pH of the skin and their effects on the mites. Previous studies on the relationship between Demodex prevalence and location of residency are relatively scarce; however, the results of this study are in agreement with few previous studies where no significant difference was found between types of residency [14,22,26]; however, it is in disagreement with studies that reported higher rates in rural areas [15]. Perhaps this difference in findings is due to the fact that patients in urban areas are more likely to use artificial lipids and cosmetics, which may have had an effect on the prevalence rates of these mites when compared to rural areas. In addition, it could be affected by the differences in the environment, biotic, and abiotic factors within both areas [5].