The use of a new PCR-based method for the diagnosis of malaria in the Spanish Malaria Reference Laboratory has promoted an increase in confirmed cases of malaria. From August 1997 to July 1998, a total of 192 whole-blood samples and 71 serum samples from 168 patients were received from the hospitals of the Spanish National Health System. Most of the patients came from west-central African countries (85%). This molecular method showed more sensitivity and specificity than microscopy, detecting 12.4% more positive samples than microscopy and 13% of mixed infections undetectable by Giemsa stain. Plasmodium falciparum was the main species detected, with 68% of the total positive malaria cases, followed by Plasmodium malariae(29%), Plasmodium vivax (14%), and Plasmodium ovale (7%), including mixed infections in all cases. This report consists of the first wide, centralized survey of malaria surveillance in Spain. The reference laboratory conducted the analysis of all imported cases in order to detect trends in acquisition. The use of a seminested multiplex PCR permitted confirmation of the origins of the infections and the Plasmodium species involved and confirmation of the effectiveness of drug treatments. This PCR also allowed the detection of the presence in Spain of primaquine-tolerantP. vivax strains from west-central Africa, as well as the detection of a P. falciparum infection induced by transfusion.

The rise of imported malaria cases and the high fatality rate in Europe make the search for new and easy diagnostic methods necessary. Rapid diagnosis tests (RDTs) are, in part, developed to cover the lack of diagnosis experience. Unfortunately, our data suggest that the accuracy of RDTs is insufficient and could increase the number of incorrect malaria diagnoses.In recent years, countries in which malaria is not endemic have reported high and increasing numbers of imported malaria cases, with fatalities up from 3.8 to 20% (2). Preventing fatal outcomes in malaria cases requires early recognition of infection, accurate laboratory diagnosis, and prompt therapy (2). Unfortunately, health-care personnel in countries where malaria is not endemic frequently lack experience in the microscopic diagnosis of malaria. In Italy, 80% of cases had less than a 1-week elapse between the onset of malaria symptoms and the microscopic diagnosis, but the average diagnosis took 8.5 days and the range was 1 to 28 days (3). This fact makes the search for new and easy diagnostic methods necessary. Rapid diagnosis tests (RDTs) for malaria might offer a valid alternative to microscopy (5).We studied 206 pre-and posttreatment samples from 169 patients in 1998 and 1999 by microscopic diagnosis and seminested multiplex PCR (4). These samples were also tested using three commercial RDT kits; 189 samples from 149 patients were tested with the ParaSight-F Kit (Becton Dickinson), 197 samples from 126 patients were tested with the OptiMal Kit (Flow Incorporated), and 54 samples from 41 patients were tested with the ICT Pf/Pv kit (Amrad). All patients (with ages of 16 months to 72 years) presented a history of fever and travel in the previous year to an area of malaria endemicity.RDTs were performed according to the manufacturers' instructions. All microscopy-positive samples were confirmed by PCR (4). Furthermore, PCR detected 6 samples with mixed infections (4 Plasmodium falciparum plus P. malariae and 2 P. falciparum plus P. ovale) from samples that were characterized as P. falciparum-only by microscopy and 24 more positive samples (12 P. falciparum, 4 P. ovale, 6 P. malariae, and 2 P. vivax). The three RDT methods showed a high rate of false positives and false negatives (Table 1). Moreover, 29.2% of positive non-P. falciparum samples rendered a positive P. falciparum result when the ParaSight-F test was used, which suggests a high number of cross-reactions, as this test according to the manufacturer detects only P. falciparum infections. In the same way, according to the manufacturers, the OptiMal and ICT Pf/Pv kits are able to detect P. falciparum specifically and the other Plasmodium spp. unspecifically. Our data, however, show that e Average time in which a sample is still positive by RDT (negative by microscopy and PCR) after treatment.

In an effort to improve the diagnosis of intestinal amoebiasis, a real-time PCR has been used for the detection and differentiation of Entamoeba histolytica and E. dispar infections in African or South American immigrants who live in Spain. Faecal samples from all of the 130 subjects had apparently been found to contain E. histolytica/E. dispar cysts by microscopical examination. Using the real-time PCR, E. histolytica DNA was detected in faecal samples from only 10 (7.7%) of the immigrants, with E. dispar DNA detected in the samples from another 117 (90.0%) of the subjects. The use of such PCR in the routine investigation of patients found positive for E. histolytica/E. dispar cysts (by microscopy) is recommended, especially in non-endemic areas.

Background:The diagnostic approach to eosinophilia is complex, given the numerous reported etiologies. Intestinal parasites (especially helminths) are a concern in children from high-burden settings. We describe the diagnostic approach and clinical management of eosinophilia in a cohort of migrant children. Methods: We conducted a retrospective observational study that included children diagnosed with eosinophilia at a reference center for pediatric tropical diseases from 2014 to 2018. All patients were screened according to a unified protocol, including direct microbiologic and serologic tests. Results: A total of 163 children presented with eosinophilia during the study period [median age, 7.7 years (4.1-12.2); 57.1% boys], mostly from Asia (27.6%) and South America (22.1%). Most were internationally adopted children (43.6%) or migrants (26.4%). Only 34.4% of the children were symptomatic, and the main etiology for eosinophilia was helminths (56.4%). After a sequential diagnostic approach, no etiology was found for 40.5% of the patients. The independent risk factors for an unexplained etiology were younger age (≤2 years: odds ratio, 3.6; 95% CI, 1.3-10.2; P = 0.015), absence of symptoms (odds ratio, 4.8; 95% CI, 1.8-12.5; P = 0.001) and mild eosinophilia (<1000/µL: odds ratio, 4.2; 95% CI, 4.5-11.7; P = 0.005). Only 6 children were treated empirically. In those children with an identified cause and in those treated empirically, the eosinophilia resolved in 52% in a median of 7 months (5-9). Conclusions: Helminths are the main cause of eosinophilia in migrant children and need to be hunted, especially in older children with eosinophil counts >1000 eosinophils/µL.

BackgroundSubmicroscopic malaria (SMM) can be defined as low-density infections of Plasmodium that are unlikely to be detected by conventional microscopy. Such submicroscopic infections only occasionally cause acute disease, but they are capable of infecting mosquitoes and contributing to transmission. This entity is frequent in endemic countries; however, little is known about imported SMM.The goals of this study were two-fold: a) to know the frequency of imported SMM, and b) to describe epidemiological, laboratorial and clinical features of imported SMM.MethodsA retrospective study based on review of medical records was performed. The study population consisted of patients older than 15 years attended at the Tropical Medicine Unit of Hospital Carlos III, between January 1, 2002 and December 31, 2007. Routinely detection techniques for Plasmodium included Field staining and microscopic examination through thick and thin blood smear. A semi-nested multiplex malaria PCR was used to diagnose or to confirm cases with low parasitaemia.ResultsSMM was diagnosed in 104 cases, representing 35.5% of all malaria cases. Mean age (IC95%) was 40.38 years (37.41-43.34), and sex distribution was similar. Most cases were in immigrants, but some cases were found in travellers. Equatorial Guinea was the main country where infection was acquired (81.7%). Symptoms were present only in 28.8% of all SMM cases, mainly asthenia (73.3% of symptomatic patients), fever (60%) and arthromialgias (53.3%). The associated laboratory abnormalities were anaemia (27.9%), leukopaenia (15.4%) and thrombopaenia (15.4%). Co-morbidity was described in 75 cases (72.1%).ConclusionsResults from this study suggest that imported SMM should be considered in some patients attended at Tropical Medicine Units. Although it is usually asymptomatic, it may be responsible of fever, or laboratory abnormalities in patients coming from endemic areas. The possibility of transmission in SMM has been previously described in endemic zones, and presence of vector in Europe has also been reported. Implementation of molecular tests in all asymptomatic individuals coming from endemic area is not economically feasible. So re-emergence of malaria (Plasmodium vivax) in Europe may be speculated.