Laboratory validation of the automated diagnosis of intestinal parasites via fecal sample processing for the recovery of intestinal parasites through the dissolved air flotation technique.

BACKGROUND: Techniques for diagnosing intestinal parasites need technological advancements in the preanalytical (collection/processing) and analytical (detection) stages. The dissolved air flotation (DAF) technique effectively recovers parasites from processed feces for routine diagnosis. Artificial intelligence (AI) is a practical and affordable alternative to modernize the analysis stage of microscopy images and generates high efficiency in the parasitological examination of feces. METHODS: The objective of this study was to standardize a laboratory protocol for stool processing using the DAF technique in conjunction with an automated diagnosis of intestinal parasites (DAPI) system. A total of 400 samples were obtained to perform the tests with the use of DAF to verify the recovery of the parasites as a function of the chemical reagent (polymer and surfactant), the volume of the flotation tube, and standardization of smear assembly on a microscopy slide, with automated analysis by DAPI. The DAF protocol that obtained the most satisfactory results in terms of parasite recovery (P < 0.05) and slide positivity was compared with the Three Fecal Test (TF-Test) protocol with manual (microscopists) and automated (DAPI) evaluation. We compared the sensitivity with the modified TF-Test technical protocol and the diagnostic agreement with the gold standard (Kappa) result. RESULTS: There was no significant difference in the parasite recovery between the 10 ml and 50 ml tubes (P > 0.05). The surfactants showed a range of parasite recoveries between 41.9% and 91.2% in the float supernatant. We obtained a maximum positivity of 73% of the assembled slides when we applied DAF processing with 7% CTAB surfactant and 57% positivity with the modified TF-Test technique. Regarding diagnostic performance, the TF-Test-modified and DAF techniques used in fecal processing for subsequent computerized analysis by AI presented sensitivities of 86% and 94%, with kappa agreements of 0.62 and 0.80 (substantial), respectively. CONCLUSIONS: The DAF protocol defined in this study and the DAPI system are innovative processes for parasite recovery and fecal debris elimination that are favorable for effectively detecting pathogenic structures in laboratory diagnosis.

First record of natural infection by Angiostrongylus cantonensis (Nematoda: Metastrongyloidea) in Belocaulus willibaldoi and Rattus norvegicus in an urban area of São Paulo city, SP, Brazil.

Angiostrongylus cantonensis, a rat lungworm, is one of the leading causes of eosinophilic meningitis in humans. Infection in humans occurs by the ingestion of intermediate hosts, undercooked paratenic hosts or contaminated vegetables and fruits by mucus from infected molluscs. This zoonosis is widespread in tropical and subtropical areas of Southeast Asia, it has also been reported in the Pacific Islands as well as in other regions of Americas. In Brazil, human cases of angiostrongyliasis have been reported since 2007 in Southeast, Northeast, and South regions. In January 2011, we collected a batch of 30 Belocaulus willibaldoi slides in a neighborhood of São Paulo city (Parque Fernanda). Six of them were used for identifying species, and the others (24) were used in parasitological tests through digestion in peptic solution and then larvae isolation by the Rugai method. A total of 250 larvae were obtained and they had morphological traits of Angiostrongylus spp. Later, four Golden hamsters (Mesocricetus auratus) were infected with 38 larvae that allowed the recovery of young worms from the brain and lungs of rodents on the 21st and 30th day of infection. In this same neighborhood we captured rodents (Rattus norvegicus) that, after necropsy led us to recovery of 22 adult worms in the pulmonary arteries (14 males and 8 females) in May 2011. The larvae and worms obtained from natural infection were evaluated by morphological and morphometric parameters, as well as biological behavior patterns and molecular profile. All methodologies identified the parasite as Angiostrongylus cantonensis. In this way, we report for the first time, the natural infection by A. cantonensis in intermediate (B. willibaldoi) and definitive (R. norvegicus) hosts in a new urban region of Brazil.