Next Article in Journal
Needs Assessment for a VR-Based Adult Nursing Simulation Training Program for Korean Nursing Students: A Qualitative Study Using Focus Group Interviews
Previous Article in Journal
Does Improvement in Health-Related Lifestyle Habits Increase Purpose in Life among a Health Literate Cohort?
 
 
Search for Articles:
Title / Keyword
Author / Affiliation / Email
Journal
Article Type
 
 
Advanced Search
 
Section
Special Issue
Volume
Issue
Number
Page
 
Journals
IJERPH
Volume 17
Issue 23
10.3390/ijerph17238877
ijerph-logo
Submit to this Journal Review for this Journal Propose a Special Issue
► Article Menu

Article Menu

share Share announcement Help format_quote Cite question_answer Discuss in SciProfiles thumb_up
...
Endorse textsms
...
Comment
first_page
settings
Order Article Reprints
Font Type:
Arial Georgia Verdana
Font Size:
Aa Aa Aa
Line Spacing:
Column Width:
Background:
Article

Prevalence and Subtype Distribution of Blastocystis Infection in Patients with Diabetes Mellitus in Thailand

by
Noppon Popruk
1,
Satakamol Prasongwattana
2,
Aongart Mahittikorn
3,
Attakorn Palasuwan
4,
Supaluk Popruk
3,* and
Duangdao Palasuwan
4,*
1
Department of Transfusion Medicine and Clinical Microbiology, Faculty of Allied Health Sciences, Chulalongkorn University, Bangkok 10330, Thailand
2
Department of Nursing, Bang Pa-in Hospital, Bang Pa-in District, Phra Nakhon Si Ayutthaya 13160, Thailand
3
Department of Protozoology, Faculty of Tropical Medicine, Mahidol University, Bangkok 10400, Thailand
4
Oxidation in Red Cell Disorders Research Unit, Department of Clinical Microscopy, Faculty of Allied Health Sciences, Chulalongkorn University, Bangkok 10330, Thailand
*
Authors to whom correspondence should be addressed.
Int. J. Environ. Res. Public Health 2020, 17(23), 8877; https://doi.org/10.3390/ijerph17238877
Submission received: 20 October 2020 / Revised: 25 November 2020 / Accepted: 25 November 2020 / Published: 29 November 2020
(This article belongs to the Section Infectious Disease Epidemiology)
Browse Figure
Versions Notes

Abstract

:
Diabetes mellitus (DM) is a major global public health problem with an increasing prevalence. DM increases the risk of infections caused by bacteria, fungi, viruses, and parasites. We examined the prevalence, subtypes, and risk factors of Blastocystis infection in patients with and without DM in central Thailand. Stool samples and questionnaires were obtained from 130 people in the DM group and 100 people in the non-DM group. Blastocystis infection was identified via a nested polymerase chain reaction and subtyped via sequencing of the partial small-subunit ribosomal RNA (SSU rRNA) gene. Analysis of potential risk factors was conducted via binary logistic regression. The overall prevalence of Blastocystis infection was 10.8%, including rates of 9% and 12.3% in the non-DM and DM groups, respectively. The most prevalent subtype was ST3, followed by ST1, and ST4. Factors that potentially increased the risk of Blastocystis infection include patients being >65 years old, the presence of DM, a DM duration of ≥10 years, a low level of education, and animal ownership. In conclusion, this is the first study of Blastocystis infection in DM, and a high prevalence was found among this population. Therefore, health education promoting sanitation and hygiene is necessary to reduce and prevent infection in the community.

1. Introduction

Intestinal parasitic infection has a global distribution, and its prevalence is especially high in developing countries. Simultaneously, these countries have increasingly higher levels of public health problems related to non-communicable diseases, such as cardiovascular diseases, cancers, chronic respiratory diseases, and diabetes mellitus (DM) [1]. People with diabetes may be more susceptible to infectious disease than those without diabetes. Both innate immune response defects (including neutrophil and macrophage dysfunction) and dysfunction of the adaptive immune response (including T cells) are believed to be responsible for immune system weakness against invading pathogens in people with diabetes [2]. DM is associated with increased rates of infection, especially those caused by bacteria [3,4,5]. A few studies of parasitic infection among people with diabetes have been reported to date [6]. These studies suggest that DM is significantly associated with the prevalence of intestinal parasites or common intestinal parasites, such as Ascaris lumbricoides, Entamoeba histolytica, Giardia duodenalis, and Opisthorchis viverrini [7,8,9]. Thailand has a rapidly increasing prevalence of type 2 DM (T2DM) [10,11]. Several studies on the prevalence of intestinal parasites in healthy subjects in Thailand have been reported [12,13,14]. Surprisingly, there is a paucity of information on Blastocystis sp., one of the most frequent protozoa found in humans.
Blastocystis sp. is an enteric protozoan found in both humans and animals with a worldwide distribution [15]. Because of its genetic heterogeneity, the genetic variants have been grouped into subtypes based on sequence similarity [16]. At present, 17 subtypes of Blastocystis sp. have been reported. Subtypes (ST)1–ST8 have been detected in both humans and animals. Conversely, ST9 has only been found in humans, while other subtypes have been found in animals. ST1–ST4 are the most prevalent Blastocystis subtypes found in humans [17]. The role of Blastocystis sp. as a human pathogen is unclear. Gastrointestinal symptoms such as diarrhea, abdominal pain, bloating and constipation, and extraintestinal disorders, such as cutaneous lesions, may be associated with Blastocystis infection [18,19,20]. In Thailand, studies on Blastocystis infection have been conducted in various groups in different community settings [21,22,23]. The prevalence of Blastocystis sp. has been reported to be as high as 45% in Thailand [24]. To date, no studies have been conducted on the prevalence of Blastocystis infection in patients with DM in the country.
Therefore, the present study investigated the prevalence and subtype distribution of Blastocystis infection in patients with diabetes using PCR-based methods. In addition, we also examined the associations between risk factors and Blastocystis infection.

2. Materials and Methods

2.1. Study Area

A cross-sectional study was conducted at primary health care hospitals between November 2019 and February 2020. This study focused on participants living in the Bang Pa-in district in the Phra Nakhon Si Ayutthaya province, which is located in central Thailand. The Bang Pa-in district is located approximately 64 km north of Bangkok. It is a semi-urban community with the second highest population after the Phra Nakhon Si Ayutthaya district. According to population-based health information system data, the local population in the fiscal year of 2019 was approximately 90,000, including 4000 people with diabetes. The most important river of the Phra Nakhon Si Ayutthaya province, namely Chao Phraya River, flows along this study area. Villagers living along the river use the water for agriculture, farming, and transportation. Furthermore, Bang Pa-in Industrial Estate requires raw water from the river for production. As a result, these activities may produce and discharge waste into water resources, including canals. These characteristics make this area suitable for conducting the study.

2.2. Study Population and Study Design

To increase the efficiency of population recruitment in this study, we used a primary health care database. The primary health care system is the smallest and most effective infrastructure of the Thai health care system [25]. The study population consisted of 130 participants with DM and 100 participants without DM. The criteria for participant selection were as follows: older than 40 years of age, native resident of the district, and no use of anthelmintic/antiprotozoal drugs for at least 3 months prior to enrolment. All participants were asked to provide fresh stool samples for parasite detection. All participants gave written informed consent to participate in this study, and they were directly interviewed to obtain basic demographic information using questionnaires. The study protocol was reviewed and approved by the Research Ethics Review Committee for Research Involving Human Research Participants, Health Sciences Group, Chulalongkorn University (certificate of approval number: 231/2562).

2.3. Stool Collection and Processing

After providing informed consent, the participants were asked to complete a brief questionnaire. Then, the study participants were given a stool collection kit and standard instructions on proper and safe collection. The participants were asked to provide one stool sample. All stool samples were shipped under cool conditions to the laboratory of Protozoology Department, Faculty of Tropical Medicine, Mahidol University (Thailand) within 4–6 h after evacuation for processing. To detect Blastocystis sp., the stool samples were aliquoted and frozen in −20 °C until further DNA extraction.

2.4. DNA Extraction and Nested PCR Amplification

All stool samples were extracted using a QIAamp Fast DNA Stool Mini Kit (Qiagen, Hilden, Germany) following the manufacturer’s protocol. The extracted DNA was stored at −20 °C until use. To identify Blastocystis sp., the 1.1-kb SSU rRNA gene was detected using nested PCR. RD3 (5′-GGGATCCTGATCCTTCCGCAGGTTCACCTAC-3′) and RD5 (5′-GGAAGCTTATCTGGTTGATCCTGCCAGTA-3′) were the external primers used for primary PCR [26], and an internal set of forward (5′-GGAGGTAGTGAC AATAAATC-3′) and reverse primers (5′-ACTAGGAATTCCTCGTTCATG-3′) was used for secondary PCR [27]. Each 25-µL reaction mixture contained 1× PCR buffer, 1.5 mM MgCl2, 0.2 mM dNTPs, 1 µM each primer and 2.5 U of Taq DNA polymerase (Thermo Fisher Scientific, Waltham, MA, USA). PCR products were separated by electrophoresis in 1.5% agarose gel in the presence of ethidium bromide, visualized using ultraviolet transillumination, and photographed.

2.5. Sequencing and Phylogenetic Analysis

The positive PCR products of the 1100-bp fragment of the Blastocystis SSU rRNA gene were sequenced in two directions using appropriate internal primers on an ABI 3730xl automated DNA sequencer by Bio Basic Inc. (Bukit Batok, Singapore). Blastocystis subtypes were identified using a BLAST search of the National Center for Biotechnology Information database (https://blast.ncbi.nlm.nih.gov/Blast.cgi). The raw nucleotide sequences and 18 reference sequences were edited manually using BioEdit v.7.2.5 Software (Ibis Biosciences, Carlsbad, CA, USA), and a multiple alignment was performed using ClustalW (Table 1). Finally, MEGA version 6 software was used for phylogenetic analysis. The best model to account for the evolution of the DNA sequences was the Hasegawa–Kishino–Yano model with gamma distribution. A phylogenetic tree was constructed with the maximum likelihood method and tested with 1000 bootstrap replicates. The 25 nucleotide sequences generated in this study were deposited in GenBank under the following accession numbers: MT330258–MT330260, MT330263, MT330265–MT330267, MT330269–MT330277, and MT947108–MT947116.

2.6. Statistical Analysis

Descriptive analysis was used to describe the characteristics, prevalence, and subtype distribution. The chi-squared test was used to analyze the potential risk factors for Blastocystis infection. Odds ratios (ORs) and the corresponding 95% confidence intervals (CIs) were used to measure the degree of association between Blastocystis infection and potential risk factors. All statistical analyses were performed using IBM SPSS Statistics for Windows, version 22 (IBM Corp., Armonk, NY, USA), and p < 0.05 was considered statistically significant.

3. Results

3.1. Basic Characteristics of the Individuals

The mean age of the study participants was 66.7 years (range, 44–88), and 56.5% (130/230) of the participants had DM. Most study participants were female (66.5%). The participant characteristics of the DM and non-DM groups are presented in Table 2. There was a slightly significant difference in the level of education between the non-DM and DM groups.

3.2. The Prevalence and Subtype of Blastocystis sp.

The overall prevalence of Blastocystis infection was 10.8% (25/230). The prevalence rates of Blastocystis infection were 9% (9/100) and 12.3% (16/130) in the non-DM and DM groups, respectively (Table 3). There was no association between participant characteristics and Blastocystis infection. Male sex (OR = 1.373, 95% CI = 0.586–3.218), age ≥ 65 years (OR = 1.630, 95% CI = 0.673–3.949), presence of DM (OR = 1.419, 95% CI = 0.599–3.36), DM duration ≥ 10 years (OR = 1.439, 95% CI = 0.499–4.153), less than secondary school education (OR = 1.525, 95% CI = 0.338–6.877), and presence of animals in the household (OR = 1.337, 95% CI = 0.565–3.167) tended to increase the risk of Blastocystis infection, but none of the associations were significant.
The 25 nucleotide sequences of the partial SSU rRNA gene in the present study displayed an extremely high similarity (≥98%) to existing sequences of Blastocystis reported in GenBank (Table 4), and they were identified as three distinct subtypes: ST1, ST3, and ST4. Blastocystis ST3 was the most prevalent subtype found in both groups, followed by ST1 and ST4. Blastocystis ST4 was only found in one sample in the non-DM group (Table 5).

3.3. Phylogenetic Analysis

We performed a phylogenetic analysis of 25 nucleotide sequences of Blastocystis-positive samples compared with the 1–4 reference subtype sequences in GenBank. The sequence of the Blastocystis ST4-positive sample from the non-DM group was closely related to rat-derived sequences in GenBank, as presented in Figure 1.

4. Discussion

Blastocystis sp. has been reported in humans worldwide. In the present study, our data, obtained using nested PCR, revealed a high prevalence of Blastocystis infection among subjects with and without DM. However, the prevalence of Blastocystis infection in this study was lower than that in prior studies conducted in asymptomatic individuals in Thailand [22,41,42] but was similar to findings in other developing countries [43,44]. The highest prevalence of Blastocystis infection of 45.2% was reported in children in an orphanage in Pathum Thani province, Thailand [25]. Conversely, the prevalence was low in developed countries such as Japan (0.5–1%) [45] and Singapore (3.3%) [27]. Prevalence varies between countries and between regions within the same country. This variation could be related to the health status of the study population, geographic distribution, and detection method.
Our finding that ST3 was the predominant Blastocystis subtype was similar to the results of previously reported studies in Thailand [24,42,46,47] and other countries such as Japan, Bangladesh, Pakistan [38], and China [48]. Contrarily, other studies found that Blastocystis ST1 was the most predominant subtype [49,50]. Blastocystis ST3 is most commonly found in humans in Europe, Africa, Asia, and Australia, whereas the most common subtype in America is ST1 [51]. Nevertheless, Blastocystis ST1–ST4 have been identified as the most common subtypes in humans [18,21]. In this study, Blastocystis ST1 was the second-most common subtype, and it was found in almost all individuals with animals in their homes. It has been suggested that Blastocystis ST1 is associated with zoonotic transmission to humans [52]. Our finding indicates that Blastocystis ST1 detected in these people might have been caused by exposure to animal stools. Blastocystis ST4 was found in only one sample in the non-DM group. Our result is consistent with a previous study reporting a low prevalence of ST4 in Africa, America, and Asia [51].
In our study, 230 completed questionnaires were used to assess possible risk factors for Blastocystis infection. Blastocystis infection was not significantly related to any participant characteristics. This result is consistent with previous findings in patients with DM undergoing hemodialysis [53]. Meanwhile, another study found that Blastocystis infection was significantly more prevalent in patients with DM than in those without DM [54]. Interestingly, our results illustrated that most infected individuals consume safe drinking water. On the contrary, several studies indicated that Blastocystis infection was significantly associated with the quality of drinking water [55,56]. This discrepancy could be attributed to other potential factors associated with Blastocystis infection, such as sanitation and hygiene practices. However, the fecal–oral route is considered to be the major mode of transmission of this protozoan [19]. The prevalence of Blastocystis infection among the two groups was not statistically different in the present study. The findings of other studies indicate that type 2 diabetes in humans is associated with compositional changes in the intestinal microbiota that decrease the abundance of some universal butyrate-producing bacteria and increase the likelihood of various opportunistic pathogens [57,58]. Blastocystis can release proteases that affect the integrity of the epithelial tissue and possibly facilitate colonization by other intestinal pathogens and changes in the intestinal microbiota diversity and composition [59,60].
The phylogenetic tree indicated that the 25 nucleotide sequences in the same subtype clusters had good bootstrap support and belonged to three subtypes: ST1, ST3, and ST4. One nucleotide sequence with ST4 in this study was located on the same branch as the reference sequence of wild rats in Japan [39]. Unlike Blastocystis ST1 and ST3, which are commonly found in humans, rodents have been suggested to be the reservoir host of Blastocystis ST4 [61]. Our findings suggest that Blastocystis ST4 may have been transmitted from rodents to this participant.

5. Conclusions

This report is the first regarding the prevalence and subtype distribution of Blastocystis sp. in patients with DM in Thailand. This protozoan was more prevalent in the DM than in the non-DM group. Although, we observed no association between Blastocystis infection and potential risk factors (participant characteristics), the potential risk factors for Blastocystis infection, including DM and close contact with animals, should not be excluded. To better understand the association between potential risk factors and Blastocystis infection, it will be necessary to increase the sample size, examine a wide variety of populations, including immunocompromised people, and expand the survey area.

6. Limitations

First, we recruited study participants based on the presence of diabetes diagnosed by the Thai health care system. The DM and non-DM groups differed at baseline concerning the glycemic control profile. The difference in glycemic control between the groups may have affected the risk of infection in these individuals. Although the control group consisted of people without diabetes, the presence of other underlying diseases may have affected the study results. Second, we did not obtain other data about the participants such as medical history or confidential information because such data cannot be obtained without patient permission or legal authorization. Moreover, the sample size was a limitation in this study as well.

Author Contributions

Conceptualization, N.P., D.P., and S.P. (Supaluk Popruk); methodology, D.P., S.P. (Supaluk Popruk), and S.P. (Satakamol Prasongwattana); software, N.P. and S.P. (Supaluk Popruk); validation, N.P.; formal analysis, N.P. and S.P. (Supaluk Popruk); writing—original draft preparation, N.P.; writing—review and editing, N.P., D.P., A.P., A.M., S.P. (Supaluk Popruk), and S.P. (Satakamol Prasongwattana); funding acquisition, D.P.; supervision, S.P. (Supaluk Popruk) and D.P. All authors have read and agreed to the published version of the manuscript.

Funding

This research was funded by grant from the 90th Anniversary of Chulalongkorn University Fund, Ratchadaphiseksomphot Endowment Fund batch#46 (2/2020), Thailand.

Acknowledgments

The authors would like to thank all of those who participated in the study and staff at all the primary health care hospitals in Bang Pa-in, Phra Nakhon Si Ayutthaya province. We thank the staff of Protozoology Department, Faculty of Tropical Medicine, Mahidol University and the Pathology Laboratory, Veterinary Medicine Department, Armed Forces Research Institute of Medical Sciences (Bangkok, Thailand) for excellent laboratory assistance. We thank Kittisak Thawnashom for statistical advice.

Conflicts of Interest

The authors declare no conflict of interest.

References

  1. World Health Organization. Global Status Report on Noncommunicable Diseases 2014; WHO: Geneva, Switzerland, 2014; pp. 1–298. [Google Scholar]
  2. Berbudi, A.; Rahmadika, N.; Tjahjadi, A.; Ruslami, R. Type 2 Diabetes and its Impact on the Immune System. Curr Diabetes Rev. 2020, 16, 442–449. [Google Scholar] [CrossRef] [PubMed]
  3. Shah, B.R.; Hux, J.E. Quantifying the risk of infectious diseases for people with diabetes. Diabetes Care 2003, 26, 510–513. [Google Scholar] [CrossRef] [PubMed] [Green Version]
  4. Carey, I.M.; Critchley, J.A.; DeWilde, S.; Harris, T.; Hosking, F.J.; Cook, D.G. Risk of Infection in Type 1 and Type 2 Diabetes Compared with the General Population: A Matched Cohort Study. Diabetes Care 2018, 41, 513–521. [Google Scholar] [CrossRef] [PubMed] [Green Version]
  5. Casqueiro, J.; Casqueiro, J.; Alves, C. Infections in patients with diabetes mellitus: A review of pathogenesis. Indian J. Endrocrinol. Metab. 2012, 16, S27–S36. [Google Scholar]
  6. Akinbo, F.O.; Olujobi, S.O.; Omoregie, R.; Egbe, C. Intestinal parasitic infections among diabetes mellitus patients. Biomarkers Genomic Med. 2013, 5, 44–47. [Google Scholar] [CrossRef] [Green Version]
  7. Machado, E.R.; Matos, N.O.; Rezende, S.M.; Carlos, D.; Silva, T.C.; Rodrigues, L.; Almeida, M.J.R.; de Oliveira, M.R.F.; Muniz-Junqueira, M.I.; Gurgel-Gonçalves, R. Host-Parasite Interactions in Individuals with Type 1 and 2 Diabetes Result in Higher Frequency of Ascaris lumbricoides and Giardia lamblia in Type 2 Diabetic Individuals. J. Diabetes Res. 2018, 2018, 4238435. [Google Scholar] [CrossRef] [Green Version]
  8. Htun, N.S.N.; Odermatt, P.; Paboriboune, P.; Sayasone, S.; Vongsakid, M.; Phimolsarn-Nusith, V.; Tran, X.D.; Ounnavong, P.S.; Andriama-Hefasoa, N.; Senvanpan, N.D.; et al. Association between helminth infections and diabetes mellitus in adults from the Lao People’s Democratic Republic: A cross-sectional study. Infect. Dis. Poverty 2018, 7, 105. [Google Scholar] [CrossRef]
  9. Nuchprayoon, S.; Siriyasatien, P.; Kraivichian, K.; Porksakorn, C.; Nuchprayoon, I. Prevalence of parasitic infections among Thai patients at the King Chulalongkorn Memorial Hospital, Bangkok, Thailand. J. Med. Assoc. Thai. 2002, 85, S415–S423. [Google Scholar]
  10. Nanditha, A.; Ma, R.C.; Ramachandran, A.; Snehalatha, C.; Chan, J.C.; Chia, K.S.; Shaw, J.E.; Zimmet, P.Z. Diabetes in Asia and the Pacific: Implications for the Global Epidemic. Diabetes Care 2016, 3, 472–485. [Google Scholar] [CrossRef] [Green Version]
  11. Aekplakorn, A.; Chariyalertsak, S.; Kessomboon, P.; Assanangkornchai, S.; Taneepanichskul, S.; Putwatana, P. Prevalence of Diabetes and Relationship with Socioeconomic Status in the Thai Population: National Health Examination Survey, 2004–2014. J. Diabetes Res. 2018, 2018, 1654530. [Google Scholar] [CrossRef]
  12. Kitvatanachai, S.; Rhongbutsri, P. Intestinal parasitic infections in suburban government schools, Lak Hok subdistrict, Muang Pathum Thani, Thailand. Asian Pac. J. Trop. Med. 2013, 6, 699–702. [Google Scholar] [CrossRef] [Green Version]
  13. Wongsaroj, T.; Nithikathkul, C.; Rojkitikul, W.; Nakai, W.; Royal, L.; Rammasut, P. Brief communication (Original). National survey of helminthiasis in Thailand. Asian Biomed. 2014, 8, 779–783. [Google Scholar] [CrossRef] [Green Version]
  14. Punsawad, C.; Phasuk, N.; Bunratsami, S.; Thongtup, K.; Siripakonuaong, N.; Nongnaul, S. Prevalence of intestinal parasitic infection and associated risk factors among village health volunteers in rural communities of southern Thailand. BMC Public Health 2017, 17, 564. [Google Scholar] [CrossRef] [PubMed]
  15. Tan, K.S. Blastocystis in humans and animals: New insights using modern methodologies. Vet Parasitol. 2004, 126, 121–144. [Google Scholar] [CrossRef] [PubMed]
  16. Stensvold, C.R.; Suresh, G.K.; Tan, K.S.; Thompson, R.C.; Traub, R.J.; Viscogliosi, E.; Yoshikawa, H.; Clark, C.G. Terminology for Blastocystis subtypes—A consensus. Trends Parasitol. 2007, 23, 93–96. [Google Scholar] [CrossRef]
  17. Alfellani, M.A.; Taner-Mulla, D.; Jacob, A.S.; Imeede, C.A.; Yoshikawa, H.; Stensvold, C.R.; Clark, C.G. Genetic Diversity of Blastocystis in Livestock and Zoo Animals. Protist 2013, 164, 497–509. [Google Scholar] [CrossRef] [Green Version]
  18. Tan, K.S. New insights on classification, identification, and clinical relevance of Blastocystis spp. Clin. Microbiol. Rev. 2008, 21, 639–665. [Google Scholar] [CrossRef] [Green Version]
  19. Katsarou-Katsari, A.; Vassalos, C.M.; Tzanetou, K.; Spanakos, G.; Papadopoulou, C.; Vakalis, N. Acute urticaria associated with amoeboid forms of Blastocystis sp. subtype 3. Acta Derm. Venereol. 2008, 88, 80–81. [Google Scholar] [CrossRef]
  20. Stensvold, C.R.; Lewis, H.C.; Hammerum, A.M.; Porsbo, L.J.; Nielsen, S.S.; Olsen, K.E.; Arendrup, M.C.; Nielsen, H.V.; Mølbak, K. Blastocystis: Unravelling potential risk factors and clinical significance of a common but neglected parasite. Epidemiol. Infect. 2009, 137, 1655–1663. [Google Scholar] [CrossRef] [Green Version]
  21. Taamasri, P.; Leelayoova, S.; Rangsin, R.; Naaglor, T.; Ketupanya, A.; Mungthin, M. Prevalence of Blastocystis hominis carriage in Thai army personnel based in Chonburi, Thailand. Mil. Med. 2002, 167, 643–646. [Google Scholar] [CrossRef] [Green Version]
  22. Popruk, S.; Udonsom, R.; Koompapong, K.; Mahittikorn, A.; Kusolsuk, T.; Ruangsittichai, J.; Palasuwan, A. Subtype distribution of Blastocystis in Thai-Myanmar border, Thailand. Korean J. Parasitol. 2015, 53, 13–19. [Google Scholar] [CrossRef] [PubMed]
  23. Pipatsatitpong, D.; Leelayoova, S.; Mungthin, M.; Aunpad, R.; Naaglor, T.; Rangsin, R. Prevalence and Risk Factors for Blastocystis Infection among Children and Caregivers in a Child Care Center, Bangkok, Thailand. Am. J. Trop. Med. Hyg. 2015, 93, 310–315. [Google Scholar] [CrossRef] [Green Version]
  24. Saksirisampant, W.; Nuchprayoon, S.; Wiwanitkit, V.; Yenthakam, S.; Ampavasiri, A. Intestinal parasitic infestations among children in an orphanage in Pathum Thani province. J. Med. Assoc. Thai. 2003, 86, S263–S270. [Google Scholar] [PubMed]
  25. Jongudomsuk, P.; Srithamrongsawat, S.; Patcharanarumol, W.; Limwattananon, S.; Pannarunothai, S.; Vapatanavong, P.; Sawaengdee, K.; Fahamnuaypol, P. The Kingdom of Thailand Health System Review; World Health Organization, Regional Office for the Western Pacific: Manila, Philippines, 2015. [Google Scholar]
  26. Clark, C.G. Extensive genetic diversity in Blastocystis hominis. Mol. Biochem. Parasitol. 1997, 87, 79–83. [Google Scholar] [CrossRef]
  27. Wong, K.H.; Ng, G.C.; Lin, R.T.; Yoshikawa, H.; Taylor, M.B.; Tan, K.S. Predominance of subtype 3 among Blastocystis isolates from a major hospital in Singapore. Parasitol. Res. 2008, 102, 663–670. [Google Scholar] [CrossRef] [PubMed]
  28. Whipps, C.M.; Boorom, K.; Bermudez, L.E.; Kent, M.L. Molecular characterization of Blastocystis species in Oregon identifies multiple subtypes. Parasitol. Res. 2010, 106, 827–832. [Google Scholar] [CrossRef]
  29. Banaticla, J.E.; Rivera, W.L. Detection and subtype identification of Blastocystis isolates from wastewater samples in the Philippines. J. Water Health 2011, 9, 128–137. [Google Scholar] [CrossRef] [Green Version]
  30. Udonsom, R.; Prasertbun, R.; Mahittikorn, A.; Mori, H.; Changbunjong, T.; Komalamisra, C.; Pintong, A.R.; Sukthana, Y.; Popruk, S. Blastocystis infection and subtype distribution in humans, cattle, goats, and pigs in central and western Thailand. Infect. Genet. Evol. 2018, 65, 107–111. [Google Scholar] [CrossRef]
  31. Wylezich, C.; Belka, A.; Hanke, D.; Beer, M.; Blome, S.; Höper, D. Metagenomics for broad and improved parasite detection: A proof-of-concept study using swine faecal samples. Int. J. Parasitol. 2019, 49, 769–777. [Google Scholar] [CrossRef]
  32. Arisue, N.; Hashimoto, T.; Yoshikawa, H. Sequence heterogeneity of the small subunit ribosomal RNA genes among blastocystis isolates. Parasitology 2003, 126, 1–9. [Google Scholar] [CrossRef]
  33. Rivera, W.L. Phylogenetic analysis of Blastocystis isolates from animal and human hosts in the Philippines. Vet. Parasitol. 2008, 156, 178–182. [Google Scholar] [CrossRef] [PubMed]
  34. Pintong, A.R.; Sunyanusin, S.; Prasertbun, R.; Mahittikorn, A.; Mori, H.; Changbunjong, T.; Komalamisra, C.; Sukthana, Y.; Popruk, S. Blastocystis subtype 5: Predominant subtype on pig farms, Thailand. Parasitol. Int. 2018, 67, 824–828. [Google Scholar] [CrossRef] [PubMed]
  35. Palasuwan, A.; Palasuwan, D.; Mahittikorn, A.; Chiabchalard, R.; Combes, V.; Popruk, S. Subtype Distribution of Blastocystis in Communities along the Chao Phraya River, Thailand. Korean J. Parasitol. 2016, 54, 455–460. [Google Scholar] [CrossRef] [PubMed] [Green Version]
  36. Chavatte, J.M.; Jureen, L. Incidental Detection of Cyclospora Cayetanensis during General Health Screening: A Case Study from Singapore. J. Trop. Dis. 2016, 4, 1–5. [Google Scholar] [CrossRef]
  37. Han, J.Q.; Li, Z.; Zou, Y.; Pu, L.H.; Zhu, X.Q.; Zou, F.C.; Huang, C.Q. Prevalence, Molecular Characterization and Risk Factors of Blastocystis sp. from Farmed Pigs in Yunnan Province, Southwestern China. Acta Parasitol. 2020, 65, 1005–1010. [Google Scholar] [CrossRef]
  38. Yoshikawa, H.; Wu, Z.; Kimata, I.; Iseki, M.; Ali, I.K.; Hossain, M.B.; Zaman, V.; Haque, R.; Takahashi, Y. Polymerase chain reaction-based genotype classification among human Blastocystis hominis populations isolated from different countries. Parasitol. Res. 2004, 92, 22–29. [Google Scholar]
  39. Katsumata, M.; Yoshikawa, H.; Tokoro, M.; Mizuno, T.; Nagamoto, T.; Hendarto, J.; Asih, P.B.S.; Rozi, I.E.; Kimata, I.; Takami, K.; et al. Molecular phylogeny of Blastocystis isolates from wild rodents captured in Indonesia and Japan. Parasitol. Res. 2018, 117, 2841–2846. [Google Scholar] [CrossRef]
  40. Silberman, J.D.; Sogin, M.L.; Leipe, D.D.; Clark, C.G. Human parasite finds taxonomic home. Nature 1996, 380, 398. [Google Scholar] [CrossRef]
  41. Yaicharoen, R.; Sripochang, S.; Sermsart, B.; Pidetcha, P. Prevalence of Blastocystis hominis infection in asymptomatic individuals from Bangkok, Thailand. Southeast Asian J. Trop. Med. Public Health 2005, 36, 17–20. [Google Scholar]
  42. Yowang, A.; Tsaousis, A.D.; Chumphonsuk, T.; Thongsin, N.; Kullawong, N.; Popluechai, S.; Gentekaki, E. High diversity of Blastocystis subtypes isolated from asymptomatic adults living in Chiang Rai, Thailand. Infect. Genet. Evol. 2018, 65, 270–275. [Google Scholar] [CrossRef] [Green Version]
  43. Belleza, M.L.; Reyes, J.C.; Tongol-Rivera, P.N.; Rivera, W.L. Subtype analysis of Blastocystis sp. isolates from human and canine hosts in an urban community in the Philippines. Parasitol. Int. 2016, 65, 291–294. [Google Scholar] [CrossRef] [PubMed]
  44. Sanpool, O.; Laymanivong, S.; Thanchomnang, T.; Rodpai, R.; Sadaow, L.; Phosuk, I.; Maleewong, W.; Intapan, P.M. Subtype identification of human Blastocystis spp. isolated from Lao People’s Democratic Republic. Acta Trop. 2017, 168, 37–40. [Google Scholar] [CrossRef] [PubMed]
  45. Hirata, T.; Nakamura, H.; Kinjo, N.; Hokama, A.; Kinjo, F.; Yamane, N.; Fujita, J. Prevalence of Blastocystis hominis and Strongyloides stercoralis infection in Okinawa, Japan. Parasitol. Res. 2007, 101, 1717–1719. [Google Scholar] [CrossRef] [PubMed]
  46. Jantermtor, S.; Pinlaor, P.; Sawadpanich, K.; Pinlaor, S.; Sangka, A.; Wilailuckana, C.; Wongsena, W.; Yoshikawa, H. Subtype identification of Blastocystis spp. isolated from patients in a major hospital in northeastern Thailand. Parasitol. Res. 2013, 112, 1781–1786. [Google Scholar] [CrossRef] [PubMed]
  47. Srichaipon, N.; Nuchprayoon, S.; Charuchaibovorn, S.; Sukkapan, P.; Sanprasert, V. A Simple Genotyping Method for Rapid Differentiation of Blastocystis Subtypes and Subtype Distribution of Blastocystis spp. in Thailand. Pathogens 2019, 8, 38. [Google Scholar] [CrossRef] [PubMed] [Green Version]
  48. Deng, L.; Chai, Y.; Zhou, Z.; Liu, H.; Zhong, Z.; Hu, Y.; Fu, H.; Yue, C.; Peng, G. Epidemiology of Blastocystis sp. infection in China: A systematic review. Parasite 2019, 26, 41. [Google Scholar] [CrossRef] [Green Version]
  49. Leelayoova, S.; Siripattanapipong, S.; Thathaisong, U.; Naaglor, T.; Taamasri, P.; Piyaraj, P.; Mungthin, M. Drinking Water: A Possible Source of Blastocystis spp. Subtype 1 Infection in Schoolchildren of a Rural Community in Central Thailand. Am. J. Trop. Med. Hyg. 2008, 79, 401–406. [Google Scholar] [CrossRef] [Green Version]
  50. Thathaisong, U.; Siripattanapipong, S.; Mungthin, M.; Pipatsatitpong, D.; Tan-ariya, P.; Naaglor, T.; Leelayoova, S. Identification of Blastocystis subtype 1 variants in the Home for Girls, Bangkok, Thailand. Am. J. Trop. Med. Hyg. 2013, 88, 352–358. [Google Scholar] [CrossRef] [Green Version]
  51. Alfellani, M.A.; Stensvold, C.R.; Vidal-Lapiedra, A.; Onuoha, E.S.; Fagbenro-Beyioku, A.F.; Clark, C.G. Variable geographic distribution of Blastocystis subtypes and its potential implications. Acta Trop. 2013, 126, 11–18. [Google Scholar] [CrossRef] [Green Version]
  52. Noël, C.; Dufernez, F.; Gerbod, D.; Edgcomb, V.P.; Delgado-Viscogliosi, P.; Ho, L.C.; Singh, M.; Wintjens, R.; Sogin, M.L.; Capron, M.; et al. Molecular Phylogenies of Blastocystis Isolates from Different Hosts: Implications for Genetic Diversity, Identification of Species, and Zoonosis. J. Clin. Microbiol. 2005, 43, 348–355. [Google Scholar] [CrossRef] [Green Version]
  53. Gil, F.F.; Barros, M.J.; Macedo, N.A.; Júnior, C.G.; Redoan, R.; Busatti, H.; Gomes, M.A.; Santos, J.F. Prevalence of intestinal parasitism and associated symptomatology among hemodialysis patients. Rev. Inst. Med. Trop. Sao Paulo 2013, 55, 69–74. [Google Scholar] [CrossRef] [PubMed] [Green Version]
  54. Mohtashamipour, M.; Hoseini, S.G.; Pestehchian, N.; Yousefi, H.; Fallah, E.; Hazratian, T. Intestinal parasitic infections in patients with Diabetes Mellitus: A case-control study. J. Anal. Res. Clin. Med. 2015, 3, 157–163. [Google Scholar] [CrossRef]
  55. Taamasri, P.; Mungthin, M.; Rangsin, R.; Tongupprakarn, B.; Areekul, W.; Leelayoova, S. Transmission of intestinal blastocystosis related to the quality of drinking water. Southeast. Asian J. Trop. Med. Public Health 2000, 31, 112–117. [Google Scholar] [PubMed]
  56. Abdulsalam, A.M.; Ithoi, I.; Al-Mekhlafi, H.M.; Ahmed, A.; Surin, J.; Mak, J.W. Drinking water is a significant predictor of Blastocystis infection among rural Malaysian primary schoolchildren. Parasitology 2012, 139, 1014–1020. [Google Scholar] [CrossRef] [PubMed] [Green Version]
  57. Larsen, N.; Vogensen, F.K.; van den Berg, F.W.; Nielsen, D.S.; Andreasen, A.S.; Pedersen, B.K.; Al-Soud, W.A.; Sørensen, S.J.; Hansen, L.H.; Jakobsen, M. Gut microbiota in human adults with type 2 diabetes differs from non-diabetic adults. PLoS ONE 2010, 5, e9085. [Google Scholar] [CrossRef]
  58. Qin, J.; Li, Y.; Cai, Z.; Li, S.; Zhu, J.; Zhang, F.; Liang, S.; Zhang, W.; Guan, Y.; Shen, D. A metagenome-wide association study of gut microbiota in type 2 diabetes. Nature 2012, 490, 55–60. [Google Scholar] [CrossRef]
  59. Ajjampur, S.S.; Tan, K.S. Pathogenic mechanisms in Blastocystis spp.—Interpreting results from in vitro and in vivo studies. Parasitol. Int. 2016, 65, 772–779. [Google Scholar] [CrossRef]
  60. Alzate, J.F.; Toro-Londoño, M.; Cabarcas, F.; Garcia-Montoya, G.; Galvan-Diaz, A. Contrasting microbiota profiles observed in children carrying either Blastocystis spp. or the commensal amoebas Entamoeba coli or Endolimax nana. Sci. Rep. 2020, 10, 15354. [Google Scholar] [CrossRef]
  61. Stensvold, C.R.; Alfellani, M.A.; Nørskov-Lauritsen, S.; Prip, K.; Victory, E.L.; Maddox, C.; Nielsen, H.V.; Clark, C.G. Subtype distribution of Blastocystis isolates from synanthropic and zoo animals and identification of a new subtype. Int. J. Parasitol. 2009, 39, 473–479. [Google Scholar] [CrossRef] [Green Version]
Ijerph 17 08877 g001 550
Figure 1. Maximum likelihood analysis of the Blastocystis SSU rRNA (small subunit ribosomal RNA) gene based on the general time reversible model. The outgroup sequences were Proteromonas lacertae (U37108) and Karotomorpha sp. (DQ431242). Symbol ▲—Diabetes DM, ■—Non-DM and *—Presence of animals in the household.
Figure 1. Maximum likelihood analysis of the Blastocystis SSU rRNA (small subunit ribosomal RNA) gene based on the general time reversible model. The outgroup sequences were Proteromonas lacertae (U37108) and Karotomorpha sp. (DQ431242). Symbol ▲—Diabetes DM, ■—Non-DM and *—Presence of animals in the household.
Ijerph 17 08877 g001
Table
Table 1. GenBank references for the Blastocystis subtypes (ST)1–ST4 sequences used to construct a phylogenetic tree.
Table 1. GenBank references for the Blastocystis subtypes (ST)1–ST4 sequences used to construct a phylogenetic tree.
SubtypeAccession NumberHost
1EU679349Human [28]
GU992416Wastewater [29]
MH104999Human [30]
MK801358Pig [31]
2AB070987Human [32]
EU445487Pig [33]
3EU445494Human [33]
EU445496Human [33]
KT819620Human [34]
KT819622Human [34]
KU051536Human [35]
KX618192Human [36]
MG214877Human (unpublished)
MH104993Human [30]
MK375226Pig [37]
4AY244620Human [38]
MH127499Rat [39]
U51152Guinea pig [40]
Table
Table 2. Characteristics of study participants with diabetes mellitus (DM, n = 130) and without diabetes mellitus (Non-DM, n = 100).
Table 2. Characteristics of study participants with diabetes mellitus (DM, n = 130) and without diabetes mellitus (Non-DM, n = 100).
CharacteristicsDM
n = 130 (%)		Non-DM
n = 100 (%)		Total
n = 230 (%)		p
GenderMale43/130 (33.1%)34/100 (34%)77/230 (33.5%)0.883
Female87/130 (66.9%)66/100 (66%)153/230 (66.5%)
Age<65 years54/130 (41.5%)43/100 (43%)97/230 (42.2%)0.824
≥65 years76/130 (58.5%)57/100 (57%)133/230 (57.8%)
Level of educationLow
(≤primary school)		120/130 (92.3%)84/100 (84%)204/230 (88.7%)0.049 *
High
(≥secondary school)		10/130 (7.7%)16/100 (16%)26/230 (11.3%)
EmployedNo66/130 (50.8%)58/100 (58%)124/230 (53.9%)0.275
Yes64/130 (49.2%)42/100 (42%)106/230 (46.1%)
Source of drinking waterTreated water (bottled and tap water)118/130 (90.8%)96/100 (96%)214/230 (93%)0.122
Untreated water (surface and rainwater)12/130 (9.2%)4/100 (4%)16/230 (7%)
Animals in the householdNo53/130 (40.8%)44/100 (44%)97/230 (42.2%)0.623
Yes77/130 (59.2%)56/100 (56%)133/230 (57.8%)
* p < 0.05; DM, diabetes mellitus.
Table
Table 3. Association between participant characteristics and Blastocystis infection in present study.
Table 3. Association between participant characteristics and Blastocystis infection in present study.
CharacteristicsNumber of Examined% InfectedOR (95% CI) *
GenderMale7713 (10/77)1.373 (0.586–3.218)
Female1539.8 (15/153)1
Age<65 years978.2 (8/97)1
≥65 years13312.8 (17/133)1.63 (0.673–3.949)
DM statusNon-DM1009 (9/100)1
DM13012.3 (16/130)1.419 (0.599–3.36)
Duration of DM≤10 years8310.8 (9/83)1
>10 years4714.9 (7/47)1.439 (0.499–4.153)
Education statusLow
(≤primary school)		20411.3 (23/204)1.525 (0.338–6.877)
High
(≥secondary school)		267.7 (2/26)1
EmployedNo12412.1 (15/124)1.321 (0.567–3.078)
Yes1069.4 (10/106)1
Source of drinking waterTreated water (bottled and tap water)21411.2 (24/214)1.895 (0.239–14.99)
Untreated water (surface and rainwater)166.3 (1/16)1
Presence of animals in householdYes13312 (16/133)1.337 (0.565–3.167)
No979.3 (9/97)1
* Binary logistic regression, p > 0.05. DM, diabetes mellitus; OR, odds ratio; CI, confidence interval.
Table
Table 4. Accession numbers of positive samples used in the phylogenetic reconstruction in this study.
Table 4. Accession numbers of positive samples used in the phylogenetic reconstruction in this study.
GenBank Accession No.SubtypeGroup (DM/Non-DM)Query CoverSequence Similarity (%)Similar GenBank Reference Sequence
MT3302583DM97%99.53KT819620
MT3302591DM98%99.44MH104999
MT3302601DM97%99.44GU992416
MT3302631DM98%99.07EU679349
MT3302653Non-DM99%98.44KT819620
MT3302663DM99%98.52KT819620
MT3302671Non-DM98%98.98MK801358
MT3302693DM98%99.17KT819620
MT3302701Non-DM99%98.99MK801358
MT3302713DM98%99.44KT819620
MT3302723DM98%99.26KT819620
MT3302733DM97%99.81KX618192
MT3302743DM97%99.81KT819620
MT3302753DM97%99.81KT819620
MT3302763DM97%99.63KX618192
MT3302773DM97%99.54MN914073
MT9471081DM100%99.5MH104999
MT9471093DM100%99.9MH104993
MT9471103DM100%99.26KT819622
MT9471114Non-DM100%99.8MH127499
MT9471123Non-DM100%99.8MG214877
MT9471133Non-DM100%99.9MK375226
MT9471143Non-DM100%100MK375226
MT9471153Non-DM100%99.8MK375226
MT9471163Non-DM100%99.9MK375226
DM, diabetes mellitus.
Table
Table 5. Subtype distribution in the diabetes mellitus (DM) and non-DM groups.
Table 5. Subtype distribution in the diabetes mellitus (DM) and non-DM groups.
GroupSubtype Distribution Positive Case/Total No. (%)Total
ST1ST3ST4
DM4/130 (3%)12/130 (9.3%)0/130 (0%)16/130 (12.3%)
Non-DM2/100 (2%)6/100 (6%)1/100 (1%)9/100 (9%)
DM, diabetes mellitus.
Publisher’s Note: MDPI stays neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Share and Cite

MDPI and ACS Style

Popruk, N.; Prasongwattana, S.; Mahittikorn, A.; Palasuwan, A.; Popruk, S.; Palasuwan, D. Prevalence and Subtype Distribution of Blastocystis Infection in Patients with Diabetes Mellitus in Thailand. Int. J. Environ. Res. Public Health 2020, 17, 8877. https://doi.org/10.3390/ijerph17238877

AMA Style

Popruk N, Prasongwattana S, Mahittikorn A, Palasuwan A, Popruk S, Palasuwan D. Prevalence and Subtype Distribution of Blastocystis Infection in Patients with Diabetes Mellitus in Thailand. International Journal of Environmental Research and Public Health. 2020; 17(23):8877. https://doi.org/10.3390/ijerph17238877

Chicago/Turabian Style

Popruk, Noppon, Satakamol Prasongwattana, Aongart Mahittikorn, Attakorn Palasuwan, Supaluk Popruk, and Duangdao Palasuwan. 2020. "Prevalence and Subtype Distribution of Blastocystis Infection in Patients with Diabetes Mellitus in Thailand" International Journal of Environmental Research and Public Health 17, no. 23: 8877. https://doi.org/10.3390/ijerph17238877

Note that from the first issue of 2016, this journal uses article numbers instead of page numbers. See further details here.

Article Metrics

Back to TopTop