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| SYMPOSIUM |
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| Year : 2017 | Volume
: 7
| Issue : 2 | Page : 86-91 |
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Laboratory diagnosis of soil transmitted helminthiasis
Sumeeta Khurana, Shveta Sethi
Department of Medical Parasitology, Post Graduate Institute of Medical Education and Research, Chandigarh, India
| Date of Acceptance | 16-Jun-2017 |
| Date of Web Publication | 25-Sep-2017 |
Correspondence Address:
Sumeeta Khurana
Department of Medical Parasitology, Postgraduate Institute of Medical Education and Research, Chandigarh
India
 Source of Support: None, Conflict of Interest: None  | Check |
DOI: 10.4103/tp.TP_29_17
 Abstract | | |
Soil-transmitted helminths (STHs) include, i.e., hookworms (Ancylostoma duodenale, Necator americanus), roundworm (Ascaris lumbricoides), whipworm (Trichuris trichiura) and Strongyloides stercoralis. Globally, around 1.5 billion people are infected with STHs. STHs contribute to significant impairment of mental and physical growth, especially in developing countries. Unfortunately, these infections mostly remain undiagnosed due to lack of trained personnel and appropriate technologies. Intermittent shedding of eggs or larvae further makes the diagnosis difficult. Thus, there is a dire need of rapid and accurate tests for the diagnosis of STHs. The diagnostic methods include conventional and molecular methods. Conventional methods include microscopy, culture, and egg counting. Serology has a role, especially in case of S. stercoralis where conventional methods have very poor sensitivities. The rapid, highly sensitive molecular techniques, particularly quantitative polymerase-chain reaction make it suitable for diagnosing STH over insensitive as well as labor-intensive conventional methods. Until now, molecular detection of STH was mainly restricted to the research setting, but now, there is recommendation of adopting molecular tests in the World Health Organization STH elimination programs. Thus, STH infections are important public health problems and should be appropriately diagnosed and managed to reduce the mortality and morbidity significantly. Keywords: Diagnosis, helminths, soil
How to cite this article:
Khurana S, Sethi S. Laboratory diagnosis of soil transmitted helminthiasis. Trop Parasitol 2017;7:86-91 |
| Introduction | |  |
Soil-transmitted helminths (STHs) comprise a number of intestinal nematodes i.e. hookworms (Ancylostoma duodenale, Necator americanus), roundworm (Ascaris lumbricoides), whipworm (Trichuris trichiura), and Strongyloides stercoralis though World Health Organization (WHO) formally does not include S. stercoralis in STH list. STH have been included in the WHO's list of 17 neglected tropical diseases (NTDs) because of the associated poverty, significant morbidity, and DALY's lost.[1] Most infections occur in the tropical and subtropical countries including India. Globally, around 1.5 billion people are infected with STHs.[1] Ascariasis is reported in 771.7–891.6 million people, trichuriasis in 429.6–508 million people, and about 406.3–480.2 million are infected with hookworms.[2] STHs contribute to significant impairment of mental and physical growth and anemia, especially in children. STH infections are often asymptomatic but sometimes result in severe gastrointestinal (GI) symptoms. The WHO has set a goal of eliminating STHs as a public health problem in childhood by 2020.[3]
Most parasitic diseases cannot be diagnosed by physical examination alone, and laboratory investigations are necessary. Unfortunately, these infections mostly remain undiagnosed due to lack of trained personnel and appropriate technologies. Intermittent shedding of eggs or larvae further makes the diagnosis difficult. Thus, there is a dire need of rapid and accurate tests for the diagnosis of STHs. As there is no adequate gold standard for STH detection, this further makes the comparison and standardization of any new technique difficult.[4] The diagnostic methods include conventional and molecular methods. Moreover, quantitation of worm burden is also important to assess the intensity of infection and prognosis.
| Conventional Diagnostics | | |
Conventionally, the STHs are diagnosed by the examination of fecal or other GI specimens for the presence of helminthic eggs, larvae or sometimes adult worms or their segments. Due to intermittent shedding of eggs and/or larvae, several specimens (at least 3) collected over the period of 10 days is required to detect parasites.[5] Fresh fecal specimen of approximately large teaspoon amount or about 10 ml of liquid stool sample should reach the laboratory within 1 h of being collected.
Gross examination
The specimen should be examined for the color, consistency (formed/semiformed/unformed/watery), presence of worm or segments, presence of pus, or blood etc.
Microscopy-based techniques
Direct microscopy
Make a smooth, thin preparation (with saline and iodine) and cover with a cover glass. In case of dysenteric and unformed specimens, select a portion of the specimen which contains blood and mucus to make smear and without adding saline or stain, cover with a cover glass.[5] Examine the preparation under microscope to identify any larvae or helminthic eggs. They should be carefully observed for their shape, size, bile staining, etc., Eggs of A. lumbricoides are bile stained while those of other STHs are not. Fertilized eggs of A. lumbricoides are rounded, 45 to 75 μm long and have a thick shell with an external mammillated layer. In some cases, the outer layer is absent (decorticated eggs). Unfertilized eggs are elongated and larger than fertile eggs (~90 μm), their shell is thinner and mammillated layer is more variable, either with large protuberances or practically none.[6] The eggs of Ancylostoma and Necator cannot be differentiated microscopically. The eggs are thin-shelled, colorless, and measure 60–75 × 35–40 μm.[6]T. trichiura eggs are 50–55 × 20–25 μm, thick-shelled barrel-shaped, and have a pair of polar “plugs” at each end.[5] In case of S. stercoralis, unsheathed larvae measuring 200–250 μm × 16 μm are observed. They show a typical rhabditiform large bulbed esophagus.[5] If stool sample remains in warm and humid environment for >24 h, the larvae may hatch from the hookworm eggs. These must be differentiated from the larvae of S. stercoralis.[7] In T. trichiura infection, eosinophils and Charcot-Leyden crystals may be present in the stool.
Fecal concentration methods
If the number of organisms is less in feces, then wherever possible stool should be concentrated. Most recommended procedure is formal-ether concentration (FEC) method. Formalin-ether solution which has lower specific gravity than the parasitic organisms, thus concentrating the latter in the sediment.[8] Floatation techniques (zinc sulfate floatation, saturated sodium chloride floatation, etc.) are not widely used because infertile eggs of Ascaris and larvae of Strongyloides do not float and thus cannot be easily recovered by this method.[5]
Egg counting
Kato-Katz method
This is the most common method employed for quantitation of helminth eggs. The WHO recommends the examination of duplicate slides in Kato-Katz (K-K) method.[9] Slides must be examined after glycerol-clearing time of 40–60 min, else hookworm eggs tend to disintegrate and disappear. The helminthic eggs per gram of stool (EPG), as a standard infection intensity measure, is calculated as: No. of eggs in smear ×24.[10]
According to the WHO, 5000 and 50,000 EPG for A. lumbricoides, 2000 and 4000 EPG for hookworm, and 1000 and 10,000 EPG for T. trichiura, are the thresholds for moderate and heavy infections, respectively.[10] K-K is good for the detection of A. lumbricoides and T. trichiura infections while it has relatively poor sensitivity for the detection of hookworm eggs because hookworm eggs tend to disintegrate if there is a time delay in examination of samples.[11] Currently, this is the most widely employed technique in epidemiological field survey due to its low cost and relatively low level of skill required.[12]
McMaster technique
This is another egg counting technique which is comparatively easy to standardize than K-K, and performance is also comparable.[13] Here, a known volume of fecal suspension is examined microscopically using a counting chamber. Afterward the number of EPG is calculated as: Total number of eggs or oocysts ×50.[14] Here, the eggs are floated free of debris; hence, the advantage of relatively quick counting. Need of a special counting chamber is the only disadvantage here.
FLOTAC and mini-FLOTAC techniques
These are recently developed novel fecal egg counting methods. Here, FLOTAC® (University of Naples, Italy)apparatus is used to perform the flotation of sample in a centrifuge, followed by cutting off the apical portion of the floating suspension, and quantitation of eggs.[15] The major advantage is its multivalency so that this can be used for the detection of different helminths as well as intestinal protozoa simultaneously.[16] FLOTAC has many practical hindrances including high cost, requirement of a centrifuge, and long procedure time for sample preparation, decreasing its value as a sole diagnostic method.[16] Studies are underway for validation of this technique. Mini-FLOTAC method, a simplified form of FLOTAC, was developed which is optimal in the settings with limited facilities.[17] It has been suggested in the previous studies that sensitivity of mini-FLOTAC is comparable to the K-K but lower than FLOTAC. Additional advantage is that this can be used on the preserved fecal samples.[12]
| Comparison of Different Microscopy-Based Techniques | | |
A meta-analysis has compared different microscopic techniques in low intensity as well as in high intensity areas.[12] This comparison was based on Bayesian latent class model by assuming a probabilistic model for the relationship and results have been presented in [Table 1]. | Table 1: Comparison of sensitivites (%) of different Microscopy based techniques[12]
Click here to view |
FLOTAC method had overall highest sensitivity among all. Sensitivities were comparable for both Mini-FLOTAC and K-K methods. Direct microscopy had the lowest sensitivity. Test sensitivities strongly varied by intensity of infection (low and high intensity groups) for all tests, especially for the K-K method where sensitivity was lower in the low-intensity group compared with the high-intensity group (high sensitivity group, 74–95%). This variation needs to be taken into consideration while selecting any diagnostic test in a specific setting.[12]
The comparison of average egg counts by different techniques shows that K-K method estimates higher egg count compared to the FLOTAC method. FLOTAC method generally underestimates the average egg counts.[18] In McMaster technique also, higher egg counts were observed in comparison to K-K, especially in cases of T. trichiura and hookworm infections.[13],[19]
Culture
Culture is mainly used to recover S. stercoralis which is viviparous while hookworm may be artificially cultured in the laboratory to produce rhabditiform larvae. There are many methods, some of which are obsolete while others are used in research settings.
Harada-Mori filter paper strip culture
This was initially introduced by Harada and Mori in 1955.[7] This technique requires filter paper to which fecal material is added, and a test tube containing some water into which it is inserted. After incubation in suitable environment, hatching of ova and/or development of larvae occur. In addition to the low sensitivity of 28%, other drawbacks are that refrigerated or preserved specimen cannot be used for culture. In addition, filter paper containing infective larvae is biohazardous.[7]
Filter paper/slant culture technique ( Petri dish More Details)
This technique is quite similar to Harada-Mori technique, only difference is that this is a slide-based method where slide containing fresh fecal material is placed in a glass or plastic petri dish containing water. This technique allows the direct examination with dissecting microscope to look for larvae.[7]
Charcoal culture
This is another way to culture using granulated charcoal. Charcoal provides an environment for larval development that mimics conditions in the nature.[7]
Koga agar plate culture
This is mainly used for the culture of S. stercoralis. Stool sample is placed on the agar plate and incubated in a humid chamber at ambient temperature. Following incubation, the plates are examined for the presence of larvae of S. stercoralis under a light microscope. As larvae crawl over the agar, they carry bacteria with them. This creates visible tracks over the agar. The plates are rinsed with 10% acetyl-formalin solution. The eluent is centrifuged, and the sediment is examined under microscope.[20] These larvae should be differentiated from those of hookworms. This is more sensitive than direct smear and fecal concentration methods.
Baermann culture technique
It is mainly used for S. stercoralis larvae detection. A walnut-sized stool sample (10 g) is placed on gauze which is kept into a glass funnel, and covered with tap water. After 2 h, liquid at the bottom is collected, centrifuged, and supernatant is removed. The sediment is examined under a microscope at a ×400 magnification to confirm the larvae.[7],[10] Although this technique is less time-consuming, labor intensive and large quantity of stool are required which may not be available.[7]
Drawbacks of culture techniques include requirement of technical expertise, difficult to perform under field conditions, and take longer time. Moreover, old and refrigerated samples are not suitable.[7]
Serology-based assays
In situations where fecal samples are unavailable, serology can have a role in diagnosis. This falls into two categories: antigen-detection assays and antibody-detection assays. These include the enzyme-linked immunosorbent assay (ELISA), and its derived forms such as Falcon Assay Screening Test ELISA, Dot-ELISA, Luciferase Immunoprecipitation System (LIPS), indirect immunofluorescent antibody test (IFAT) or direct immunofluorescent antibody test, immunoblotting, and some rapid immunochromatographic tests (RDTs).[21]
Disadvantages associated with serodiagnosis are its more invasive nature, antibodies tend to remain persistent after treatment and thus may not be indicative of active infection, cross-reaction with other nematodes, particularly with filarial infections.[22] However, serology plays an important role in the diagnosis of S. stercoralis infection. It replicates within the host leading to chronic infection, with wide spectrum of clinical presentations, ranging from asymptomatic stage to life-threatening hyperinfection due to dissemination of larvae, particularly in immunocompromised patients. Conventional diagnostic tests have very low sensitivity in these patients. A single-direct fecal smear examinaxtion (DS) fails to detect almost 70% of cases but examination of multiple samples improves the sensitivity.[22] Here, serological techniques have higher sensitivity compared to other conventional parasitological tests (81–98%). Recently, developed indirect immunofluorescence microscopy (IFAT) has been widely used with 97% sensitivity and 98% specificity.[22] Requirement for whole live infective-larvae is the main disadvantage of IFAT and to overcome this drawback, gelatin particle agglutination test was developed with reported sensitivity of 81% and specificity of 74%.[22] Using the immunodominant antigen of S. stercoralis and S. ratti, immunoblot tests have been developed which showed sensitivity between 65–100%.[23] Several ELISAs have been developed which have demonstrated better sensitivity (94%) compared to IFAT. Recently, LIPS, has been developed and demonstrated very good sensitivity (97%) and specificity (100%).[22] This technique can be performed relatively fast (2.5 h), and an even faster version (<2 min) is under research.[24]
Coproantigen detection
Coproantigen assays have been developed since long for the diagnosis of a range of human and animal intestinal infections. For STH, these are mainly available for Strongyloides and hookworms. Polyclonal rabbit antiserum raised against Strongyloides ratti excretory/secretory (E/S) antigen and coated on microtiter plates to capture fecal antigen.[25]Strongyloides coproAg remains stable if frozen as formalin-extracted fecal supernatants stored at −20°C. There are many trials of conversion of the Strongyloides coproAg-based coproELISA into a rapid RDT, many of them are commercially available also.[26] The sensitivity for the detection of S. stercoralis antigen is reported to be 0.5 mg/ml.[26] IgG antibody raised against Ancylostoma ceylanicum ES is used to capture hookworm coproantigens. This is capable of detecting ES proteins from 10 ng/mL to 10 ug/mL.[27]
| Molecular-Based Approaches | | |
Advancements in molecular techniques have provided an advantage of rapid detection as well as accurate quantification of STHs eggs. The much better sensitivity of molecular techniques makes them especially useful to monitor effect of treatment or control strategies. The molecular targets used mainly are ITS-1, ITS-2, 18S, etc., Several different polymerase-chain reaction (PCR) based methods are available, i.e., conventional PCR, quantitative PCR (qPCR), multiplex PCR reaction, etc.[28],[29] There are many technical problems associated with the molecular methods. Stool samples contain high amounts of suspended solids, bile acids, etc., that may inhibit the molecular reactions.[30] This obstacle can be overcome using flotation or sedimentation step before the extraction process.[31] Many nucleic acid extraction kits have been developed with additional inhibitor removal steps. Another problem is that egg shells of STHs eggs are much tougher than the cell walls of bacteria, leading to low yield of nucleic acid. Destruction of the egg shells can be facilitated by using methods such as sonication or tissue homogenization with degradation resistant beads, etc.[32] Despite the limitations, molecular tests have been proved to be more sensitive than conventional methods in detecting very low numbers of eggs.[8] With advancement, it is possible to detect multiple parasites using multiplex PCR in one go but they tend to lag behind in sensitivity compared to the singleplex assays.[28]
| Comparison of Conventional Versus Molecular Techniques for Sth Diagnosis | | |
The rapid, highly sensitive properties of qPCR make it suitable for diagnosing STH over insensitive as well as labor-intensive age old conventional methods. Many studies have compared the sensitivities of both methods. According to these studies, for A. lumbricoides, microscopy had a sensitivity of 70–88% while molecular tests had a sensitivity of 85–100%; for hookworms, sensitivity of microscopy was reported 30–88% versus molecular techniques with sensitivity of 75–100%; for S. stercoralis, sensitivity of microscopy was 16–50% versus molecular techniques 76–93%; for T. trichiura, microscopy had the sensitivity of 88% while molecular techniques had the reported sensitivity of 100%.[28]
Until now, molecular detection of STH was mainly restricted to the research setting. However, now considering its accuracy and cost-effectiveness, there is recommendation of adopting molecular tests in the WHO STH elimination programs as well.
| Other Molecular Techniques | | |
Certain new tests have been introduced, for example:
- Loop-mediated isothermal amplification: Act as a cheaper alternative because water bath is used here for reaction in place of expensive thermocycler[8]
- Droplet digital PCR, a third-generation PCR technology, was introduced for absolute quantification of targeted genes applicable for pathogens.[8]
| Other Diagnostic Parameters | | |
- Hemogram: Eosinophilia (>600 μL or >6% of the thin-layer chromatography) may be used as a marker of STH infection. Many studies have shown a correlation between STH infection and eosinophilia that is usually observed in about 47% cases of ascariasis, 78% cases of trichuriasis and 82% cases of strongyloidiasis, up to 60% cases of hookworm infection.[33],[34],[35],[36] Low hemoglobin of ≤5 g/dl (60%) and low ferritin (33%) concentration (due to iron deficiency anemia) may be seen in patients with hookworm infestation[7]
- Occult blood test: Occult blood in stool in patients with severe anemia due to chronic hookworm infection has been used as a surrogate marker[37]
- Other samples: In case of ascariasis, hookworm or S. sterocoralis, during the larval migration phase of infection, diagnosis can be made by finding the larvae in the sputum or gastric washings. In ascariasis, charcot-Leyden crystals and eosinophils may be found in the sputum[7]
- Endoscopy: In the case of S. stercoralis, most common endoscopic findings, which are usually incidental, are ulceration, bleeding, duodenal spasm, mucosal edema, thickened duodenal folds. In case of hookworm infection also upper digestive endoscopy plays an important role.[22]
String test (entero-test capsule)
In case of S. stercoralis infection, due to irregular larval shedding in stool, especially in cases of chronic low infection, duodenal material may be examined for the presence of larvae. Briefly, a nylon yarn coiled inside a lined gelatin capsule is swallowed, and the capsule is delivered to the duodenum. Then, the line is pulled back with adhered bile-stained duodenal mucus.[22]
- Diagnostic radiology: In ascariasis, worm intestinal tracks may be visualized. This may be particularly obvious when 2 worms are lying parallel, like 'trolley car lines'. Appendicitis and cholecystitis can also be seen. In Hookworm infection, intestinal hypermotility, proximal jejunal dilatation, coarsening of the mucosal folds are seen. In S. stercoralis infection, findings ranging from normal appearance of GI tract or mild edema with thickened folds to significant dilatation of the small bowel mucosa with stricture in hyperinfected patients may be seen[22]
- Histological examinations: this can also confirm the diagnosis by showing larvae, eggs or occasionally adult forms, predominantly in the gastric, or duodenal crypts along with eosinophilic infiltration[22]
- Intradermal skin tests. The immediate hypersensitivity reaction to different somatic and excretory/secretory antigens has been reported to be a reliable skin test for the diagnosis of strongyloidiasis. Cross-reactions and persistence after treatment are the major drawbacks.[22]
Thus, STHs are important public health problems and should be appropriately diagnosed and managed to reduce the mortality and morbidity significantly. There is a need to develop and validate newer, rapid, sensitive, and specific tests for detection of STHs.
Financial support and sponsorship
Nil.
Conflicts of interest
There are no conflicts of interest.
| References | | |
| 1. | World Health Organization. Investing to Overcome the Global Impact of Neglected Tropical Diseases: Third WHO Report on Neglected Diseases. Geneva, Switzerland: WHO Press; 2015. p. 1-191.  |
| 2. | Pullan RL, Smith JL, Jasrasaria R, Brooker SJ. Global numbers of infection and disease burden of soil transmitted helminth infections in 2010. Parasit Vectors 2014;7:37. |
| 3. | World Health Organization. Eliminating Soil-Transmitted Helminthiases as a Public Health Problem in Children, Progress Report 2001-2010 and Strategic Plan 2011-2020. Geneva, Switzerland: WHO Press; 2012. p. 1-78. |
| 4. | Tarafder MR, Carabin H, Joseph L, Balolong E Jr., Olveda R, McGarvey ST. Estimating the sensitivity and specificity of Kato-Katz stool examination technique for detection of hookworms, Ascaris lumbricoides and Trichuris trichiura infections in humans in the absence of a 'gold standard'. Int J Parasitol 2010;40:399-404. |
| 5. | Cheesbrough M. Direct Laboratory Practice in Tropical Countries (Part-1). New York: Cambridge University Press; 2009. p. 29-35. |
| 6. | |
| 7. | Garcia LS. Diagnostic Medical Parasitology. 4 th ed. Washington, D.C: ASM Press; 2001. p. 786-801. |
| 8. | Amoah ID, Singh G, Stenström TA, Reddy P. Detection and quantification of soil-transmitted helminths in environmental samples: A review of current state-of-the-art and future perspectives. Acta Trop 2017;169:187-201. |
| 9. | WHO Expert Committee. Prevention and control of schistosomiasis and soil-transmitted helminthiasis. World Health Organ Tech Rep Ser 2002;912:i-vi, 1-57. |
| 10. | Knopp S, Mgeni AF, Khamis IS, Steinmann P, Stothard JR, Rollinson D, et al. Diagnosis of soil-transmitted helminths in the era of preventive chemotherapy: Effect of multiple stool sampling and use of different diagnostic techniques. PLoS Negl Trop Dis 2008;2:e331. |
| 11. | Tarafder MR, Carabin H, Joseph L, Balolong E Jr., Olveda R, McGarvey ST. Estimating the sensitivity and specificity of Kato-Katz stool examination technique for detection of hookworms, Ascaris lumbricoides and Trichuris trichiura infections in humans in the absence of a 'gold standard'. Int J Parasitol 2010;40:399-404. |
| 12. | Nikolay B, Brooker SJ, Pullan RL. Sensitivity of diagnostic tests for human soil-transmitted helminth infections: A meta-analysis in the absence of a true gold standard. Int J Parasitol 2014;44:765-74. |
| 13. | Albonico M, Rinaldi L, Sciascia S, Morgoglione ME, Piemonte M, Maurelli MP, et al. Comparison of three copromicroscopic methods to assess albendazole efficacy against soil-transmitted helminth infections in school-aged children on Pemba Island. Trans R Soc Trop Med Hyg 2013;107:493-501. |
| 14. | Vadlejch J, Petrtýl M, Zaichenko I, Cadková Z, Jankovská I, Langrová I, et al. Which McMaster egg counting technique is the most reliable? Parasitol Res 2011;109:1387-94. |
| 15. | Utzinger J, Rinaldi L, Lohourignon LK, Rohner F, Zimmermann MB, Tschannen AB, et al. FLOTAC: A new sensitive technique for the diagnosis of hookworm infections in humans. Trans R Soc Trop Med Hyg 2008;102:84-90. |
| 16. | Knopp S, Glinz D, Rinaldi L, Mohammed KA, N'Goran EK, Stothard JR, et al. FLOTAC: A promising technique for detecting helminth eggs in human faeces. Trans R Soc Trop Med Hyg 2009;103:1190-4. |
| 17. | Barda BD, Rinaldi L, Ianniello D, Zepherine H, Salvo F, Sadutshang T, et al. Mini-FLOTAC, an innovative direct diagnostic technique for intestinal parasitic infections: Experience from the field. PLoS Negl Trop Dis 2013;7:e2344. |
| 18. | Knopp S, Rinaldi L, Khamis IS, Stothard JR, Rollinson D, Maurelli MP, et al. A single FLOTAC is more sensitive than triplicate Kato-Katz for the diagnosis of low-intensity soil-transmitted helminth infections. Trans R Soc Trop Med Hyg 2009;103:347-54. |
| 19. | Albonico M, Ame SM, Vercruysse J, Levecke B. Comparison of the Kato-Katz thick smear and McMaster egg counting techniques for monitoring drug efficacy against soil-transmitted helminths in schoolchildren on Pemba Island, Tanzania. Trans R Soc Trop Med Hyg 2012;106:199-201. |
| 20. | Koga K, Kasuya S, Khamboonruang C, Sukhavat K, Ieda M, Takatsuka N, et al. A modified agar plate method for detection of Strongyloides stercoralis. Am J Trop Med Hyg 1991;45:518-21. |
| 21. | Ndao M. Diagnosis of parasitic diseases: Old and new approaches. Interdiscip Perspect Infect Dis 2009;2009:278246. |
| 22. | Requena-Méndez A, Chiodini P, Bisoffi Z, Buonfrate D, Gotuzzo E, Muñoz J. The laboratory diagnosis and follow up of strongyloidiasis: A systematic review. PLoS Negl Trop Dis 2013;7:e2002. |
| 23. | Buonfrate D, Formenti F, Perandin F, Bisoffi Z. Novel approaches to the diagnosis of Strongyloides stercoralis infection. Clinical Microbiology and Infection. 2015;21:543-52. |
| 24. | Ramanathan R, Burbelo PD, Groot S, Ladarola MJ, Neva FA, et al. A luciferase immunoprecipitation systems assay enhances the sensitivity and specificity of diagnosis of Strongyloides stercoralis infection. J Infect Dis 2008;198:444-51. |
| 25. | Johnson MJ, Behnke JM, Coles GC. Detection of gastrointestinal nematodes by a coproantigen capture ELISA. Res Vet Sci 1996;60:7-12. |
| 26. | Sykes AM, McCarthy JS. A coproantigen diagnostic test for Strongyloides infection. PLoS Negl Trop Dis 2011;5:e955. |
| 27. | Bungiro RD Jr., Cappello M. Detection of excretory/secretory coproantigens in experimental hookworm infection. Am J Trop Med Hyg 2005;73:915-20. |
| 28. | O'Connell EM, Nutman TB. Molecular diagnostics for soil-transmitted helminths. Am J Trop Med Hyg 2016;95:508-13. |
| 29. | Gordon CA, Gray DJ, Gobert GN, McManus DP. DNA amplification approaches for the diagnosis of key parasitic helminth infections of humans. Mol Cell Probes 2011;25,143-52. |
| 30. | Al-Soud WA, Ouis IS, Li DQ, Ljungh S, Wadstrom T. Characterization of the PCR inhibitory effect of bile to optimize real-time PCR detection of Helicobacter species. FEMS Immunol Med Microbiol 2005;44:177-82. |
| 31. | Bass D, Stentiford GD, Littlewood DT, Hartikainen H. Diverse applications of environmental DNA methods in parasitology. Trends Parasitol 2015;31:499-513. |
| 32. | Gyawali P, Ahmed W, Jagals P, Sidhu JP, Toze S. Comparison of concentration methods for rapid detection of hookworm ova in wastewater matrices using quantitative PCR. Exp Parasitol 2015;159:160-7. |
| 33. | Darlan DM, Tala ZZ, Amanta C, Warli SM, Arrasyid NK. Correlation between soil transmitted helminth infection and eosinophil levels among primary school children in medan. Open Access Maced J Med Sci 2017;5:142-146. |
| 34. | Ashrafi K, Tahbaz A, Rahmati B. Strongyloides stercoralis: The most prevalent parasitic cause of eosinophilia in gilan province, Northern Iran. Iran J Parasitol 2010;5:40-7. |
| 35. | Kucik CJ, Martin GL, Sortor BV. Common intestinal parasites. Am Fam Physician 2004;69:1161-8. |
| 36. | Jiero S, Ali M, Pasaribu S, Pasaribu AP. Correlation between eosinophil count and soil-transmitted helminth infection in children. Asian Pac J Trop Dis 2015;5:813-6. |
| 37. | Wakid MH. Fecal occult blood test and gastrointestinal parasitic infection. J Parasitol Res 2010;2010. pii: 434801. |
[Table 1]
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