Seroprevalence of SARS-CoV-2 in blood donors from the Lodi Red Zone and adjacent Lodi metropolitan and suburban area.

OBJECTIVES: To define the seroprevalence of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) in blood donors (referred to the first lockdown area (Lodi Red-Zone) of the Lombardy region and in a contiguous area that was not included in the first lockdown); to define the agreement between a commercial serological assay and a reference microneutralization assay; and to evaluate the persistence of SARS-CoV-2 neutralizing antibodies in a cohort of blood donors. METHODS: Blood donors referred to the first lockdown area in Lombardy Region and the neighbouring area were analysed for SARS-CoV-2 IgG-specific antibodies during the period 18 March to 24 June 2020. Serum samples were analysed using both a chemiluminescent immunoassay (LIAISON® SARS-CoV-2 S1/S2 IgG, DiaSorin) for the quantitative characterization of SARS-CoV-2 anti-S1 and anti-S2 IgG antibodies and a neutralizing antibodies (NT-Abs) assay. RESULTS: In the period from 18 March to 24 June, 1922 blood donors were tested for the presence of SARS-CoV-2 IgG showing a prevalence of 378/1922 (19.7%). A subgroup of 1139 blood donors were tested in parallel with a SARS-CoV-2 IgG assay and a microneutralization assay showing a prevalence of 22.2% and 21.6%, respectively. SARS-CoV-2 IgG quantification was correlated with NT-Abs titres. In 78.2% of participants the NT-Abs titre was maintained, but in 15.8% it decreased by one four-fold dilution and in 6.0% it increased by one four-fold dilution. CONCLUSIONS: The duration of immunity of SARS-CoV-2 is crucial for the course of the pandemic and for this reason the monitoring of NT Abs is important. Despite a stable NT-Abs titre being observed in the majority of blood donors, our findings need to be validated in a long-term period of follow up.

Cerebral Toxocariasis as a Cause of Epilepsy: A Pediatric Case.

Toxocarosis is the consequence of human infection by Toxocara spp. larvae and is one of the most common ascarioses, not only in developing countries, but also in the European region, where its prevalence reaches 14%. Due to their particular behavior, children are at higher risk of this parasitic infection, whose clinical features depend on the localization of the Toxocara larvae. Neurotoxocariasis is very uncommon in children and may take different forms depending on the underlying physiopathologic process: immune reaction against the parasite antigens, vasculitis, treatment complications, or, very rarely, brain localization of Toxocara spp. larvae. The association between neurotoxocariasis and the onset of childhood epilepsy has been postulated but is still debated. Moreover, a Toxocara spp. abscess causing epileptic seizures in children has been rarely described, especially in western countries. Hereby we present a 9-year-old patient with a new diagnosis of epilepsy definitely secondary to brain abscess due to the localization of Toxocara canis larvae. Diagnosis was confirmed by neuroimaging and serological test. The successful treatment with albendazole and steroids was documented with a close and long-term clinical and neuroradiological follow-up. Our experience confirms that every case of cryptogenetic epilepsy in children deserves a neuroimaging study and, in case of cystic images, Toxocara serology is mandatory to avoid further unnecessary invasive diagnostic investigations and to set the specific drug therapy.

Prevalence of SARS-CoV-2 specific neutralising antibodies in blood donors from the Lodi Red Zone in Lombardy, Italy, as at 06 April 2020.

We evaluated SARS-CoV-2 RNA and neutralising antibodies in blood donors (BD) residing in the Lodi Red Zone, Italy. Of 390 BDs recruited after 20 February 2020 - when the first COVID-19 case in Lombardy was identified, 91 (23%) aged 19-70 years were antibody positive. Viral RNA was detected in an additional 17 (4.3%) BDs, yielding ca 28% (108/390) with evidence of virus exposure. Five stored samples collected as early as 12 February were seropositive.

Multilocus sequence typing of Dientamoeba fragilis identified a major clone with widespread geographical distribution.

The flagellated protozoan Dientamoeba fragilis is often detected in humans with gastrointestinal symptoms, but it is also commonly found in healthy subjects. As for other intestinal protozoa, the hypothesis that genetically dissimilar parasite isolates differ in their ability to cause symptoms has also been raised for D. fragilis. To date, only two D. fragilis genotypes (1 and 2) have been described, of which genotype 1 largely predominates worldwide. However, very few markers are available for genotyping studies and therefore the extent of genetic variation among isolates remains largely unknown. Here, we performed metagenomics experiments on two D. fragilis-positive stool samples, and identified a number of candidate markers based on sequence similarity to the phylogenetically related species Trichomonas vaginalis. Markers corresponding to structural genes and to genes encoding for proteases were selected for this study, and PCR experiments confirmed their belonging to the D. fragilis genome; two previously described markers (small subunit ribosomal DNA and large subunit of RNA polymerase II) were also included. Using this panel of markers, 111 isolates of human origin were genotyped, all of which, except one, belonged to genotype 1. These isolates had been collected at different times from symptomatic and asymptomatic persons of different age groups in Italy, Denmark, Brazil and Australia. By sequencing approximately 160kb from 500 PCR products, a very low level of polymorphism was observed across all the investigated loci, suggesting the existence of a major clone of D. fragilis with a widespread geographical distribution.

Failure of repeated treatment with praziquantel and arthemeter in four patients with acute schistosomiasis.

Schistosomiasis is on the rise but still difficult to treat in international travelers; it should be suspected in patients returning from endemic areas. Praziquantel (PZQ) is not effective and may aggravate symptoms. More recently, combination treatment with artemisinin derivatives have shown promising results. We report four cases of acute schistosomiasis (AS) in which several courses of combined therapy had been necessary to obtain negative serology.

Immunoblotting with human native antigen shows stage-related sensitivity in the serodiagnosis of hepatic cystic echinococcosis.

The diagnosis of hepatic cystic echinococcosis is based on ultrasonography and confirmed by serology. However, no biological marker of cyst viability is currently available implying years-long patient follow-up, which is not always feasible in endemic areas. We characterized the performance of an immunoblotting test based on human hydatid cyst fluid with particular regard to its ability to distinguish between cyst stages. Sera from patients with cysts in different stages showed distinctive band pattern recognition. Most importantly, the test discriminated in 80% of cases CE3a from CE3b transitional cysts, known to have different viability profiles. Interestingly, we observed a rapid change in band pattern recognition of sera from one patient at time points when his cyst passed from active to transitional to inactive stages. Further identification of different antigens expressed by different cyst stages will support the development of diagnostic tools that could early define cyst viability, to guide clinical decision making, and shorten patient follow-up.

Isolation and genotyping of Acanthamoeba strains from corneal infections in Italy.

Acanthamoeba keratitis (AK) is a corneal disease caused by members of a genus of free-living amoebae and is associated predominantly with contact lens (CL) use. This study reports 16 cases of culture-proven AK diagnosed in northern Italy. Genotype identification was carried out with a PCR assay based on sequence analysis of the 18S rRNA gene, and sensitivity and specificity were evaluated in comparison with traditional parasitological techniques. A 405 bp region of the 18S rRNA gene (ASA.S1) including diagnostic fragment 3 (DF3) was amplified using the genus-specific primers JDP1 and JDP2. Genotype assignment was based on phenetic analysis of the ASA.S1 subset of the nuclear small-subunit rRNA gene sequence excluding the highly variable DF3 region. Phylogenetic analysis was also performed on the sequences obtained. All patients complained of monolateral infection; 11 (68.75%) admitted improper CL disinfection. In 14/16 (87.5 %) subjects, corneal scrapings were stained with calcofluor white and haematoxylin and eosin and, in ten cases (62.5 %), microscopy was positive for Acanthamoeba cysts. In vitro culture on 3 % non-nutrient agar plates was obtained in all cases (100 %), whereas cloning and axenic growth were positive for 14 amoebic stocks (87.5 %). PCR analysis had 100 % sensitivity and specificity compared with in vitro axenic culture, showing positive amplification from 15 isolates. All Acanthamoeba strains belonged to the T4 genotype, the main AK-related genotype worldwide. These results confirmed the importance of a complete diagnostic protocol, including a PCR assay, for the clinical diagnosis of AK on biological samples. Genotyping allowed inclusion of all isolates in the T4 group, thus demonstrating the prevalence of this genotype in northern Italy.

Amebic infections due to the Entamoeba histolytica-Entamoeba dispar complex: a study of the incidence in a remote rural area of Ecuador.

An epidemiologic field study was conducted in the village of Borbòn in Esmeraldas province in northern Ecuador to compare different parasitologic methods in the diagnosis of infection with the Entamoeba histolytica/Entamoeba dispar complex. The results of two stool antigen detection assays (the Prospect Entamoeba histolytica microplate assay and the E. histolytica II assay) were compared with isoenzyme characterization of the amebic isolates. Nearly all (176 of 178, 98.9%) subjects were positive for intestinal parasites on direct microscopic examination, and cysts and/or vegetative forms morphologically consistent with the E. histolytica/E. dispar complex were recorded in 48 of 178 cases (27%). Culture in Robinson's medium was positive for amebic stocks in 89 (50%) of the 178 samples tested. Of the 37 isolates successfully stabilized, cloned, and characterized by zymodeme analysis, seven (18.9%) showed isoenzyme patterns of E. histolytica, whereas 26 (70.3%) showed patterns of E. dispar. The remaining four strains were identified as Entamoeba coli (three isolates; 8.1%) and Dientamoeba fragilis (one strain; 2.7%).The immunochromatographic tests showed different degrees of sensitivity and specificity when compared with isoenzyme characterization as the reference technique. The microplate assay, which does not discriminate between E. histolytica and E.dispar, showed a sensitivity of 54.5% and a specificity of 94% for both these amebic species. In contrast, the second-generation E. histolytica II test had a sensitivity of 14.3% and a specificity of 98.4% for E. histolytica sensu stricto. Our survey clearly demonstrated that more specific and sensitive diagnostic tests, such as stool antigen detection assays and isoenzyme analysis, are needed to establish the actual worldwide distribution of E. histolytica and E. dispar.