|Year : 2022 | Volume
| Issue : 1 | Page : 126-131
Association with oral microbial alteration and oral disease among patients attending ruhengeri referral hospital, Rwanda: A case–control study
Callixte Yadufashije1, Joseph Mucumbitsi1, Marie Jeanne Umugwaneza Uwimana1, Liliane Muhimpundu1, Lydia Nzisa Mwanzia2, Martin Ndayambaje3, Emmanuel Munyeshyaka1, Gratien Twagirumukiza1, Albert Onyango Mala4
1 Department of Biomedical Laboratory Sciences, INES Ruhengeri Institute of Applied Sciences, Musanze, Rwanda
2 Department of Midwifery and Gender, Moi University, Eldoret, Kenya
3 Department of Biology, Université Hassan II de Casablanca, Casablanca, Morocco
4 Department of Medical Laboratory Sciences, Jomo Kenyatta University of Agriculture and Technology, Juja, Kenya
|Date of Submission||11-Oct-2021|
|Date of Acceptance||09-Nov-2021|
|Date of Web Publication||11-Mar-2022|
Department of Biomedical Laboratory Sciences, INES Ruhengeri Institute of Applied Sciences, Musanze
Source of Support: None, Conflict of Interest: None
Background: Dental disease remains a public health concern of this era. In 2020, World Health Organization reported that 3.5 billion of oral disease occurs every year. About 2.3 billion case is attributed to dental caries while gum disease affects 10% of the global population. Methods: This was a case control study carried out from November 2020 to February 2021. About 120 participants were recruited, of them, 60 were oral diseased, while the remaining 60 were selected as a control group. Oral swab samples were collected with the swab stick, and put into plastic sterile container to avoid external microbial contamination. Samples were transported to INES clinical microbiology laboratory for microbial identification. chi square was performed to test for association, while odd ratios and relative risk were performed to test for pathogenic microbial fraction. Results: The common oral diseases were tooth decay (56.6%) and gum disease (43.4%). the most affected age range was 5-19years (53.33%), while females (63.3%) were the most affected sex. The most isolated microorganism was Lactobacillus spp (15.8%) for patients, while Staphylococcus aureus (11.2%) was the most isolate in the control group. The statistical significant association with oral microbial alteration and oral disease was observed on Streptococcus mutans (x2=8.9, P= 0.002852), Lactobacillus spp (x2=9.84, P=0.001708), Candida spp (x2=5.2, P=0.02258), Staphylococcus aureus (x2 = 15.6, P= 0.000078), and Providencia spp (x2 = 6, P=0.014306). The overall oral microbial alteration (x2=53, P< 0.00001) was statistically significant. The ratio of pathogenic microorganisms (OR=4, 95%CI:2.3786-7.062 and RR=1.477, 95%CI:1.2478-1.7153) was significantly associated with oral disease. Conclusion: Oral microbial alteration contributes to oral disease. Early detection of oral microbial alteration, and oral diseases are recommended.
Keywords: Gum disease, microorganism, oral disease, tooth decay
|How to cite this article:|
Yadufashije C, Mucumbitsi J, Uwimana MJ, Muhimpundu L, Mwanzia LN, Ndayambaje M, Munyeshyaka E, Twagirumukiza G, Mala AO. Association with oral microbial alteration and oral disease among patients attending ruhengeri referral hospital, Rwanda: A case–control study. Biomed Biotechnol Res J 2022;6:126-31
|How to cite this URL:|
Yadufashije C, Mucumbitsi J, Uwimana MJ, Muhimpundu L, Mwanzia LN, Ndayambaje M, Munyeshyaka E, Twagirumukiza G, Mala AO. Association with oral microbial alteration and oral disease among patients attending ruhengeri referral hospital, Rwanda: A case–control study. Biomed Biotechnol Res J [serial online] 2022 [cited 2022 May 22];6:126-31. Available from: https://www.bmbtrj.org/text.asp?2022/6/1/126/339377
| Introduction|| |
A good oral health is defined as the state of mouth free of any disease affecting the oral cavity and its surrounding areas. The mouth harbors a wide variety of oral bacteria, such as Streptococcus species, Lactobacillus species, and Porphyromonas gingivalis that are believed to cause dental caries and gum disease. The maintenance of good oral hygiene is one of the most recommended strategies to curb dental disorders and oral diseases. The healthy oral cavity does not only ensure to feel healthy but also maintain the oral functions. The most common disorders of the oral cavity including dental caries and gum disease. The reduction and distribution of common oral disorders are significantly associated with their knowledge about their etiologic factors, causative agent, and prevention strategies.
Dental caries is characterized by the demineralization, and can be observed on the hard dental tissues. The disease process is initiated within the bacterial biofilm that covers the tooth surface. Dental caries and gum disease remain the major threat of public health globally, and the prevention measures may vary from population to population. Dental caries is not considered a communicable disease, and is recognized as part of the noncommunicable chronic. Gum disease is also known as periodontal disease. It is an inflammatory condition that affects the tissues surrounding the teeth. Gingivitis is the first early clinical characteristic, which occurs among patients with gum disease. The gum seems to be red, and sometimes bleed. The severe periodontitis leads to the loss of the gums surrounding the teeth, the loss of the bone, and the complete loss of the teeth. Gum disease is generally caused by the pathogenic bacteria in the mouth invading the tissue surrounding the teeth. Most adults in the UK have gum disease, and it is much less common in children. Periodontal diseases are mainly the result of infections and inflammation of the gums and bone that support the teeth. Periodontal disease and tooth decay are the two biggest threats to dental health.
In 2017, some studies estimated about 3.5 billion oral diseases globally, and 2.3 billion people suffered from dental caries of permanent teeth, which was the most common condition affecting more than 530 million children. Severe gum disease was reported as a common dental disorder, which results in the tooth loss among patients. It affects about 10% of the global population.
Developing countries ranked to be the most contributor of the high prevalence of oral diseases in the world. This is primarily caused by the inadequate exposure to fluoride (in the water supply and oral hygiene products such as toothpaste), improper brushing, tobacco use, hormone changes, chemotherapy, and poor access to oral health-care services in the community. Many low- and middle-income countries are incapable of providing adequate preventative measures to curb the prevalence of oral diseases, and treatment options of oral diseases remain largely inaccessible to large portions of the populace. This study identified the common microorganisms associated with dental disorders at Ruhengeri Referral Hospital.
| Methods|| |
The study was carried out at Ruhengeri Referral Hospital located in Musanze District, Northern Province, in Rwanda.
This was a case–control study carried from October 2020 to February 2021 at Ruhengeri Referral Hospital.
The study targeted all dental disorders outpatients accepted to participate in the study.
Sampling and sample size
Simple Random Sampling was used to recruit study participants of this study. This sampling type was used because the population selected had the same characteristics. About 120 participants were recruited. Out of which, 60 were oral disease patients, and the remaining 60 were taken as a control group.
The oral swab (on dental plaque and gum surface) samples were collected using the sterile cotton swab stick, and immediately put into sterile container and mixed with peptone water to prevent contamination. The samples were transported to INES clinical microbiology laboratory for microbial identification. Gram staining, culture, and biochemical tests were performed to identify microorganisms associated with dental disorders among patients attending Ruhengeri Referral Hospital.
We included patients who consented to participate in the study. Patients without any other health conditions were only recruited in the study. In the control group, the study targeted oral healthy individuals free from any other oral health conditions.
Healthy individuals with toothache were excluded from the control group. In the case group (patients), any patient with any other oral disease like oral candidiasis and others was excluded from the study. Patients who did not accept to participate were also excluded. Pregnant women were excluded.
Peptone water was used to enrich the growth of bacteria after collection of the samples from the plaques of the teeth. Then, samples from peptone water were inoculated on blood agar, MacConkey agar, and Sabouraud dextrose agar and incubated at 37°C for 18–24 h. The growing colonies were morphological identified and followed by Gram stain for bacteria colonies. Distinct biochemical tests were also performed to differentiate and confirm the isolated bacteria species. The identification of Gram-negative bacteria was done by using different biochemical culture media to test biochemical parameters. Simmons citrate agar was used to test bacteria with ability to use citrate as its only source of carbon. Indole test was performed to distinguish indole positive and indole negative. Urease test was also performed to test for urea catabolism by the enzyme urease produced by some bacteria strains. Kligler iron agar was performed to differentiate Gram-negative bacilli. For Gram-positive bacteria, catalase and coagulase tests were done to differentiate Gram-positive cocci such as Streptococcus and Staphylococcus species. However, for the growing yeast colonies on Sabouraud dextrose agar, germ tube test was performed to differentiate Candida albicans from other Candida spp.
Data and statistical analysis
Data were summarized and presented in the form of figures and tables. The analysis was supported by SPSS version 22 (IBM Company located in New York, USA). We tested the association with oral microbial alteration and dental disease with Chi-square (x2), whereas odds ratio (OR) and relative risk (RR) were analyzed to test for pathogenic microorganism's ratio among dental disease patients.
We have received the ethical clearance (972/RRH/DG/2020) from Ruhengeri Referral Hospital to access our study population in the hospital environment. The written informed consent was addressed to our study participants to voluntarily accept the participation in the study. No names or any personal information was mentioned to respect ethical standards.
| Results|| |
Demographic characteristics of participants
[Table 1] indicates the demographic characteristics of people with and without dental caries and gum disease with their frequencies and percentages. The most dominant age of participants with dental caries and gum disease was between 11 and 16 years (26.7%), followed by 5–10 (16.7%), 17–22 (16.7%), 35–40 (13.3%), 23–28 (10%), and 29–34 (10%). The lowest age was between 41 and 46 years (6.7%). About 26.7% of participants were in the age range of 17-22 years. About 20% of participants was attributed to 11-16 years, while 16.7% was for 5-10, 23-28, and 29-34 years. The remaining 3.3% was for 42-46 years. The lowest age was between 35 and 40 years (0.0%). According to gender, the most dominant were females (56.7%) followed by males (43.3%) among people with dental caries and gum disease, whereas the most dominant were males (56.7%) followed by females (43.3%) among people without dental caries and gum disease.
Burden of tooth decay and gum disease at Ruhengeri Referral Hospital
[Figure 1] indicates the prevalence of dental decay and gum disease. Tooth decay (56.6%) was high, whereas gum disease stood at 43.4%.
Distribution of tooth oral disease among males and females
This study investigated oral disease among women and men. Females (63.3%) were more affected than men (36.6%) [Figure 2].
Distribution of oral disease in age groups
[Figure 3] indicates the burden of tooth decay and gum disease among age groups. Ages were affected as follows 5–19 (53.33%), 20–30 (23.33%), 31–40 (16.66%), and 41–50 (6.66%). The disease reduced from adults to young people.
Alteration of isolated microorganisms between patients and control group
[Figure 4] indicates the microorganisms isolated from both patients with dental disorders and the control group selected. The observed microorganisms were Lactobacillus spp. (15.8%, 4.6%), Streptococcus mutans (9.2%, 1.9%), Pseudomonas spp. (6.6%, 5.9%), Enterobacter spp. (3.9%, 2.6%), Candida spp. (3.8%, 6.38%), Staphylococcus aureus (4.6%, 11.2%), C. albicans (4.47%, 4.47%), Staphylococcus epidermidis (3.3%, 4.6%), Providentia spp. (2.6%, 0%), Serratia spp. (1.3%, 0%), and Citrobacter spp. (1.3%, 0.33%). The couple of percentages in the bracket after each microorganism indicates the percentages of each microorganism in patients with dental disorders and the control group, respectively.
|Figure 4: Isolated microorganisms from people with oral disease and people without oral disease|
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Association of oral microbiota imbalance with tooth decay and gum disease
The association with oral microbial alteration and tooth decay and gum disease was performed. The alteration of Lactobacillus spp. (x2 = 9.84, P = 0.001708), S. mutans (x2 = 8.9, P = 0.002852), Candida spp. (x2 = 5.2, P = 0.02258), S. aureus (x2 = 15.6, P = 0.000078), and Providentia spp. (x2 = 6, P = 0.014306) was statistically significant to contribute to tooth decay and gum disease. There was no statistically significant association with Enterobacter spp. (x2 = 0.39, P = 0.532299, Pseudomonas spp. (x2 = 0.39, P = 0.532299), S. epidermidis (x2 = 2.39, P = 0.122114), C. albicans (x2 = 1.05, P = 0.305507), Serratia spp., (x2 = 2.95, P = 0.085878), and Citrobacter spp. (x2 = 0.94, P = 0.332278) to contribute to tooth decay and gum disease. The overall oral microbial alteration (x2 = 53, P < 0.00001) was statistically significant [Table 2].
Ratio of pathogenic microorganisms between the diseased participants and the control group
The ratio of oral pathogenic microorganisms was studied. OR = 4 with 95% confidence interval (CI) (2.3786–7.062) and RR = 1.463 with 95% CI (1.2478–1.7153) were calculated. The OR and RR were both above 1, which shows the high growth of pathogenic microorganisms among participants with tooth decay and gum disease [Table 3].
| Discussions|| |
Oral microbial alteration remains the major cause of oral diseases, which is a major risk of tooth loss among patients. Some studies about dental caries and gum disease were carried out in Rwanda, and in the region, but none of them investigated the contribution of oral pathogen to the oral disease. This study analyzed the association with oral microbial alteration and oral diseases. The study found that the main oral diseases at Ruhengeri Referral Hospital are tooth decay (56.6%) and gum disease (43.4%). The females (63.3%) were more affected compared to males (43.4%). The most age range affected was 5–19 years [Figure 1],[Figure 2],[Figure 3]. The findings of the study on the association with dental caries and severe periodontitis in Chilean adults reported the high prevalence in the mean age of 21.5 years, whereas the second effect was found in the mean age of above 35 years. The reported overall dental caries, gum disease, and periapical pathoses were 54.8% among males and 45.2% females. The findings of this study reported the same situation as the current study about the increase of oral disease in young age than adults, despite the difference about the prevalence of this health condition among males and females, where our study reported that the high prevalence of oral disease was high among females, but this study reported it high in males. The difference in reports may base on various factors such as nutritional factors and behavioral factors in any region. The differences and similarities of the burden of dental caries were again reported by previous findings in Ethiopia where it was observed that at young age, boys were the most affected, The study found that dental caries affected men and women at the same rate, with high dental restorations among women. Gum disease was also reported to be associated with age where the young age was at high risk of this health condition. More than 61% of cases were reported, but also about 16.8% were in severe conditions of the disease. The interest of our study was to investigate whether the alteration in the oral microorganisms contributes to oral disease. Various microorganisms were identified. Lactobacillus which is the main contributor to tooth decay progression was the predominantly isolated microorganisms among the population oral disease. The alteration of this harmful bacterium in the oral cavity was statistically significant. This bacterium was found at low number among participants without oral disease (control group). S. aureus is the know normal flora of the oral cavity area. It was the most isolated microorganism among the control group in our findings. Its derangement among the cases and control group was statistically significant. The bacterium was observed higher among the control group while lower in the patients with oral diseases. We could say that its low growth in the oral cavity due to various conditions favored the high growth of pathogenic and opportunistic pathogens in the oral cavity, and exposed the host to oral disease such tooth decay and gum disease that we identified during our study. S. mutans is the main cause of dental caries and gum disease. It was isolated with the higher percentage among the patient, and its alteration was statistically significant to be the contributor of oral diseases at Ruhengeri Referral Hospital. The alteration of Candida spp. was statistically significant. There was a critical reduction of it among patients with oral disease. Most of Candida spp. are the normal flora of the oral cavity except C. albicans which is the main agent for oral candidiasis. This yeast was isolated during our study, but its alteration was not statistically significant. Providentia spp. are opportunistic pathogens, and they are isolated during this study. Its alteration was statistically significant to cause the oral diseases. Any alteration in oral microbiome may awake the opportunistic pathogens to cause some side effects. Other bacteria such as Pseudomonas spp., Serratia, Enterobacter, S. epidermidis, and Citrobacter were a part of our findings [Figure 4] and [Table 2] and [Table 3]. Previous findings reported S. mutans as the main oral cariogenic microorganism. However, it is not always dominant among dental disease patients. Lactobacillus is one of the causes of dental caries, but the study conducted on oral microbial derangement did not find it statistically significant to cause dental caries.
The similar findings isolated Pseudomonas spp. (14.7%), Candida sp. (9.2%), Citrobacter (19.3%), Lactobacillus (29.2%), and Streptococcus (17.5%) overall in deep dentinal lesions where S. mutans was observed at low level (10.1%) in intact enamel and white spot lesions of diseased people also stood at 10%–20% of people with severe caries. Lactobacillus sp. and S. mutans are more prevalent. The dominance of Lactobacillus among dental caries patients was reported, whereas Staphylococcus spp. was observed in healthy individuals. The comparison of pathogenic and nonpathogenic microorganisms was performed in previous studies. S. mutans (28.1%) and Lactobacillus sp. (25.2%) were the main oral pathogens isolated, whereas Candida sp. (14.3%) and S. aureus were the microflora observed. The mean difference between the group was statistically significant (t = 4.1, P = 0.0068). The similar results reported Enterobacter (17.4%), Pseudomonas sp. (11.5%), Serratia (3%), Micrococcus sp. (28%), Lactobacillus sp. (21%6%), Streptococcus sp. (19.3%), S. aureus (16.1%), and C. albicans (8.0%) as the main microorganisms in the oral cavity of dental caries patients. Another study showed the contrasted results where Lactobacillus spp., C. albicans, S. aureus, S. mutans, Serratia, Pseudomonas sp, and Enterobacter spp were not main contributor of oral disease. In our findings, the derangement of Lactobacillus spp., S. aureus, and S. mutans was statistically significant to contribute to oral diseases. This contradiction may be due to different factors such as health conditions, behaviors, and environmental conditions.
We were limited to the number of samples; we cannot generalize the results of this study to the population of the region where the study was conducted
| Conclusion|| |
Oral microbial alteration was associated with oral disease at Ruhengeri Referral Hospital. Tooth decay and gum diseases were the major oral diseases observed, with the high prevalence of tooth decay compared to gum disease. Females were the most affected compared to males, whereas children and adolescents were the most affected by oral diseases. The alteration of S. mutans, Lactobacillus spp., Candida spp., S. aureus, and Providentia spp. was statistically significant to contribute to oral diseases. The OR and RR calculated showed the high grow of pathogenic microorganisms among the patients compared to the control group. The early detection of oral microbial community is recommended to dental clinics, and patients suspecting to have oral diseases should immediately attend the dental clinic for early testing to avoid uncontrolled alteration of the oral microbial community which is a leading cause of oral disease complication.
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Conflicts of interest
There are no conflicts of interest.
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[Figure 1], [Figure 2], [Figure 3], [Figure 4]
[Table 1], [Table 2], [Table 3]