Giardia duodenalis Contamination of Fresh Vegetables in a Wholesale Market

Authors

  • Jariya Sripanompong Department of Parasitology and Entomology, Faculty of Public Health, Mahidol University, THAILAND
  • Mathuros Tipayamongkholgul Department of Epidemiology, Faculty of Public Health, Mahidol University, THAILAND
  • Aongart Mahittikorn Department of Protozoology, Faculty of Tropical Medicine, Mahidol University, THAILAND
  • varakorn Kosaisavee Department of Parasitology and Entomology, Faculty of Public Health, Mahidol University

Keywords:

Giardia duodenalis, Giardiasis, fresh vegetables, contaminate, PCR, Giardia duodenalis, Giardiasis, Fresh vegetables, Contamination, Nested PCR

Abstract

Thai menus usually contain raw vegetables as the main or side dish. Currently, many outbreaks of intestinal parasitic infections that are related to eating raw vegetables have been revealed worldwide. Fresh vegetables can be contaminated by protozoan cysts, helminth eggs and larvae. The detection of helminth eggs and larvae by microscopy is easier than the detection of protozoan cysts which depends on expertise and techniques. The protozoan contamination in vegetables, which has been reported in the U.S. and other western countries, and Thailand, etc., is mostly from food- and water-borne protozoa. However, little is known about the prevalence or distribution of these waterborne protozoa due to difficulty related to detection methods. Giardia duodenalis is one of the waterborne protozoa that causes diarrhea in humans and many animals. Most infected adults are asymptomatic. They are less concerned about getting diagnosed, which results in a low estimated prevalence. However, infection has an adverse impact on children’s linear growth and psychomotor development. The conventional method of G. duodenalis detection is microscopy which has low sensitivity and specificity.

To estimate the possible occurrence of G. duodenalis in lettuce, white cabbage and cabbage from one large wholesale market, a sensitive molecular tool was used to detect protozoan DNA. These three vegetables are mostly eaten raw in many Thai food dishes. A total of 96 unwashed vegetables, of which 35 lettuces, 30 nappa cabbages and 31 cabbages were collected from September 2019 to June 2020. Samples were prepared as follows: 1) 200g of vegetable were chopped and washed with 0.85% saline, 2) sediment was concentrated by leaving it overnight, 3) DNA was extracted from the sediment. Nested polymerase chain reaction (Nested PCR) was used to detect G. duodenalis contamination targeting the glutamate dehydrogenase gene (gdh).

The results of this study showed 6 positives for G. duodenalis out of 96 (6.25%) samples. The protozoa were found in nappa cabbage (10.0%), cabbage (6.45%) and lettuce (2.86%). Unwashed vegetables from the largest wholesale vegetable market were contaminated with G. duodenalis. The Giardia cysts may be in the inner layers of these vegetable leaves due to the physical characteristics of the vegetables.  

 Although a small number of G. duodenalis contaminated vegetables were found, consistent and prolonged consumption of these vegetables may lead to Giardiasis. At present, many intestinal parasitic infections occur without knowing the sources of infection. One contaminated food is raw vegetables. Therefore, it is important to wash vegetables several times with running water or a washing solution, to reduce protozoal contamination from each leaf layer. The washing step is important to remove any pathogens from vegetables, before consuming them in their raw form.

The results of this study can be used as baseline data for G. duodenalis contamination in fresh vegetables, especially vegetables that are eaten raw. The benefit of a molecular detection method is that it can detect the minimum number of cysts, as one cyst can be detected by PCR. The development of easier, sensitive and faster detection methods could be valuable in the field of food sanitation, particularly pathogen surveillance systems for fresh vegetables and fruits.

References

Travaillé E, La Carbona S, Gargala G, Aubert D, Guyot K, Dumètre A, et al. Development of a qRT-PCR method to assess the viability of Giardia intestinalis cysts, Cryptosporidium spp. and Toxoplasma gondii oocysts. Food Control 2016; 59: 359-65.

Chonsawat P, Wongphan B. Prevalence of parasitic infections in patients at hospital for tropical diseases, Mahidol University. J Med Assoc Thai 2017; 45(2): 6073-84. (In Thai)

Korkes F, Kumagai FU, Belfort RN, Szejnfeld D, Abud TG, Kleinman A, et al. Relationship between intestinal parasitic infection in children and soil contamination in an urban slum. J Trop Pediatr 2009; 55(1): 42-5.

Aghaindum AG, Atud AQ, Nadège OAT. Implications of soils around domestic water points in the spread of intestinal parasites in the city of Yaounde (Cameroon). J Water Health 2019; 17(2): 318-28.

Hamilton KA, Waso M, Reyneke B, Saeidi N, Levine A, Lalancette C, et al. Cryptosporidium and Giardia in wastewater and surface water environments. J Environ Qual 2018; 47(5): 1006-23.

Rivero MR, Feliziani C, De Angelo C, Tiranti K, Salomon OD, Touz MC. Giardia spp., the most ubiquitous protozoan parasite in Argentina: Human, animal and environmental surveys reported in the last 40 years. Parasitol Res 2020; 119(10): 3181-201.

Fantinatti M, Bello AR, Fernandes O, Da-Cruz AM. Identification of Giardia lamblia assemblage E in humans points to a new anthropozoonotic cycle. J Infect Dis 2016; 214(8): 1256-9.

Leahy JG, Rubin AJ, Sproul OJ. Inactivation of Giardia muris cysts by free chlorine. Appl Environ Microbiol 1987; 53(7): 1448-53.

Sakkas H, Economou V, Bozidis P, Gousia P, Papadopoulou C, Karanis P. Detection of Cryptosporidium and Giardia in foods of plant origin in North-Western Greece. J Water Health 2020; 18(4): 574-8.

Chaidez C, Soto M, Gortares P, Mena K. Occurrence of Cryptosporidium and Giardia in irrigation water and its impact on the fresh produce industry. Int J Environ Health Res 2005; 15(5): 339-45.

Amorós I, Alonso JL, Cuesta G. Cryptosporidium oocysts and giardia cysts on salad products irrigated with contaminated water. J Food Prot 2010; 73(6): 1138-40.

Tantiamornkul K, Mataradchakul T. Detecting insecticide contamination and determining prevalence of protozoa in fresh vegetables from fresh markets, Maung Phayao, Phayao Province. Journal of Public Health 2019; 49(1): 118-29. (In Thai)

Caradonna T, Marangi M, Chierico FD, Ferrari N, Reddel S, Bracaglia G, et al. Detection and prevalence of protozoan parasites in ready-to-eat packaged salads on sale in Italy. Food Microbiol 2017; 67: 67-75.

Eraky MA, Rashed SM, Nasr ME-S, El-Hamshary AMS, Salah El-Ghannam A. Parasitic contamination of commonly consumed fresh leafy vegetables in Benha, Egypt. J Parasitol Res 2014; 2014: 613960.

Mohamed MA, Siddig EE, Elaagip AH, Edris AMM, Nasr AA. Parasitic contamination of fresh vegetables sold at central markets in Khartoum state, Sudan. Ann Clin Microbiol Antimicrob 2016; 15: 17.

Elsafi SH, Al-Maqati TN, Hussein MI, Adam AA, Hassan MM, Al Zahrani EM. Comparison of microscopy, rapid immunoassay, and molecular techniques for the detection of Giardia lamblia and Cryptosporidium parvum. Parasitol Res 2013; 112(4): 1641-6.

Miller KM, Sterling CR. Sensitivity of nested PCR in the detection of low numbers of Giardia lamblia cysts. Appl Environ Microbiol 2007; 73(18): 5949-50.

Suwatchinjaroen S, Songthamwa D. The development of Giardia lamblia detection in lettuce by using the real-time PCR method. J Med Assoc Thai 2016; 44(2): 5700-16. (In Thai)

Parker P. Sample size requirements for confidence intervals on distribution parameters. In: Meeker WQ, Hahn GJ, Escobar LA, eds. Statistical intervals: A guide for practitioners and researchers. 2nd ed. New Jersey: John Wiley & Sons. Inc.; 2017. p.284-307.

Read CM, Monis PT, Andrew TRC. Discrimination of all genotypes of Giardia duodenalis at the glutamate dehydrogenase locus using PCR-RFLP. Infect Genet Evol 2004; 4(2): 125-30.

Utaaker KS, Kumar A, Joshi H, Chaudhary S, Robertson LJ. Checking the detail in retail: Occurrence of Cryptosporidium and Giardia on vegetables sold across different counters in Chandigarh, India. Int J Food Microbiol 2017; 263: 1-8.

Ismail Y. Prevalence of parasitic contamination in salad vegetables collected from supermarkets and street vendors in Amman and Baqa'a - Jordan. Pol J Microbiol 2016; 65(2): 201-7.

Jongkolnee N, Songthamwat D. Detection of parasitic contamination in vegetables from Phra Nakhon Si Ayutthaya district, Phra Nakhon Si Ayutthaya. J Med Assoc Thai 2015; 43(1): 5141-50. (In Thai)

Punsawad C, Phasuk N, Thongtup K, Nagavirochana S, Viriyavejakul P. Prevalence of parasitic contamination of raw vegetables in Nakhon Si Thammarat Province, southern Thailand. BMC Public Health 2019; 19(1): 34.

Alharbi A, Toulah FH, Wakid MH, Azhar E, Farraj S, Mirza AA. Detection of Giardia lamblia by microscopic examination, rapid chromatographic immunoassay test, and molecular technique. Cureus 2020; 12(9): e10287.

Green MR, Sambrook J. Nested Polymerase Chain Reaction (PCR). Cold Spring Harb Protoc 2019; 2019(2): doi: 10.1101/pdb.prot095182.

Tram NT, Dalsgaard A. Water used to moisten vegetables is a source of Escherichia coli and protozoan parasite contamination at markets in Hanoi, Vietnam. J Water Health 2014; 12(4): 896-900.

Satjapala T, Toonsakool K, Pednog K. Contamination and reducing of parasite in fresh vegetables by washing. Bulletin of the Department of Medical Science 2014; 56(4): 205-12. (In Thai)

Kalani H, Daryani A, Sharif M, Ahmadpour E, Alizadeh A, Nasrolahei M, et al. Comparison of eight cell-free media for maintenance of Toxoplasma gondii tachyzoites. Iran J Parasitol 2016; 11(1): 104-9.

Climatological Center, Meteorological Department. Monthly and Annual Mean (Dry-Bulb) Temperature of Thailand 2020. Available from https://www.tmd.go.th/climate/climate.php?FileID=5, accessed 3 June 2021.

Downloads

Published

2021-12-30

Issue

Section

Original Articles