Standardization of a method for the detection of helminth eggs and larvae in lettuce

Abstract

Despite reports that food-borne parasitic infections have been increasing worldwide, the methodologies employed to detect food contamination by helminths are still largely based on methodologies used to detect these pathogens in feces and water. This study sought to improve the diagnosis of parasitic contaminants in lettuce by standardizing a method for detecting helminth eggs and larvae and estimating their percentage of recovery. Sanitized lettuces were artificially contaminated with different amounts of Ascaris suum and hookworm eggs and larvae. To standardize the method, we tested liquid extractors, vegetable washing steps, and spontaneous sedimentation times. Higher percentages of egg and larvae recovery were obtained using 1 M glycine as the liquid extractor, manual shaking for 3 min and 2 h of sedimentation. Five different levels of artificial contamination (ten replicates each; n = 50) were tested using these standardized conditions, yielding an average recovery of 62.6 % (±20.2), 51.9 % (±20.0), and 50.0 % (±27.3) for A. suum eggs, hookworm eggs, and larvae, respectively. Tests were performed with a different matrix to evaluate the performance of the method. Furthermore, collaborative analytical studies performed by different laboratories produced satisfactory results. The method for the identification of helminth eggs and larvae proposed in this study proved to be simpler and more efficient than previously published procedures, thereby demonstrating its potential contribution to health surveillance and epidemiological studies.

Potentials and challenges in the isolation and detection of ascarid eggs in complex environmental matrices

“…The resulting pellet was centrifuged and screened for helminth eggs under a light microscope. The inter-laboratory confirmation of the procedure resulted in an average recovery of 52.1% (±37.9) with the detection of at least one egg in 96.3% of samples spiked with A. suum eggs ( Matosinhos et al, 2016 ). The utilisation of the protocol by Pineda et al (2021) yielded in a mean recovery efficiency of 57.1% (±37.6) for the lettuce samples and 50.7% (±29.0) for the arugula samples with a proposed detection limit of eleven eggs per gram of vegetable leaves.…”

“…The United States Food and Drug Administration (USFDA) published a protocol ( USFDA, 2021 ), initially designed to isolate protozoans from contaminated water ( Bier, 1991 ), that was adapted for isolating helminth eggs from food. Nevertheless, recovery of Ascaris eggs was rather low ( Matosinhos et al, 2016 ).…”

“…Generally, different techniques have been used for the isolation of ascarid ova from contaminated vegetables, whereas most of these methods are modifications of techniques established for the analysis of faecal material or water. However, only few studies analysed the effectivity of methods utilised for the recovery of ascarid eggs from vegetables, fruits or herbs ( Matosinhos et al, 2016 ; Pineda et al, 2021 ). Matosinhos et al (2016) established a technique to recover helminth eggs from leafy vegetables like lettuce and arugula.…”

“…However, only few studies analysed the effectivity of methods utilised for the recovery of ascarid eggs from vegetables, fruits or herbs ( Matosinhos et al, 2016 ; Pineda et al, 2021 ). Matosinhos et al (2016) established a technique to recover helminth eggs from leafy vegetables like lettuce and arugula. Within the scope of this study, the method was standardised in an inter-laboratory approach, which was later on pursued and extended by Pineda et al (2021) .…”

Development and Effectiveness Evaluation of a Propolis Hydroalcoholic Solution With Glycine in Reducing Microorganisms and Removing Helminth Eggs in Leafy Greens

“…In households, where there is a large number of food contaminating reports and minimal sanitary surveillance, the development of an ideal sanitizer, besides having a chemical composition harmless to humans, must have an antimicrobial effect to inactivate bacteria and fungi, and active mechanisms that facilitate the removal of parasite eggs from fresh food (Brazil 2007; Matosinhos et al 2016).…”