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The potential of various quantitative lateral flow (LF) based assays utilizing up-converting phosphor (UCP) reporters for the diagnosis of schistosomiasis is reviewed including recent developments. Active infections are demonstrated by screening for the presence of regurgitated worm antigens (genus specific polysaccharides), whereas anti-Schistosoma antibodies may indicate ongoing as well as past infections. The circulating anodic antigen (CAA) in serum or urine (and potentially also saliva) is identified as the marker that may allow detection of single-worm infections. Quantitation of antigen levels is a reliable method to study effects of drug administration, worm burden and anti-fecundity mechanisms. Moreover, the ratio of CAA and circulating cathodic antigen (CCA) is postulated to facilitate identification of either Schistosoma mansoni or Schistosoma haematobium infections. The UCP-LF assays allow simultaneous detection of multiple targets on a single strip, a valuable feature for antibody detection assays. Although antibody detection in endemic regions is not a useful tool to diagnose active infections, it gains potential when the ratio of different classes of antibody specific for the parasite/disease can be determined. The UCP-LF antibody assay format allows this type of multiplexing, including testing a linear array of up to 20 different targets. Multiple test spots would allow detection of specific antibodies, e.g. against different Schistosoma species or other pathogens as soil-transmitted helminths. Concluding, the different UCP-LF based assays for diagnosis of schistosomiasis provide a collection of tests with relatively low complexity and high sensitivity, covering the full range of diagnostics needed in control programmes for mapping, screening and monitoring.

Trichinellosis is a serious zoonositc parasitosis worldwide. Because its clinical manifestations aren't specific, the diagnosis of trichinellosis is not easy to be made. Trichinella spiralis muscle larva (ML) excretory-secretory (ES) antigens are the most widely applied diagnostic antigens for human trichinellosis, but the major drawback of the ES antigens for assaying anti-Trichinella antibodies is the false negative in the early Trichinella infection period. The aim of this study was to characterize the T. spiralis putative serine protease (TsSP) and to investigate its potential use for diagnosis of trichinellosis. The full-length TsSP sequence was cloned and expressed, and recombinant TsSP (rTsSP) was purified by Ni-NTA-Sefinose Column. On Western blotting analysis the rTsSP was recognized by T. spiralis-infected mouse serum, and the natural TsSP was identified in T. spiralis ML crude and ES antigens by using anti-rTsSP serum. Expression of TsSP was detected at various T. spiralis developmental stages (newborn larvae, muscle larvae, intestinal infective larvae and adult worms). Immunolocalization identified the TsSP principally in cuticles and stichosomes of the nematode. The sensitivity of rTsSP-ELISA and ES-ELISA was 98.11% (52/53) and 88.68% (47/53) respectively (P > 0.05) when the sera from trichinellosis patients were examined. However, while twenty-one serum samples of trichinellosis patients' sera at 19 days post-infection (dpi) were tested, the sensitivity (95.24%) of rTsSP-ELISA was distinctly higher than 71.43% of ES-ELISA (P < 0.05). The specificity (99.53%) of rTsSP-ELISA was remarkably higher than 91.98% of ES-ELISA (P < 0.01). Only one out of 20 serum samples of cysticercosis patients cross-reacted with the rTsSP. Specific anti-Trichinella IgG in infected mice was first detected by rTsSP-ELISA as soon as 7 dpi and antibody positive rate reached 100% on 10 dpi, whereas the ES-ELISA did not permit detection of 100% of infected mice before 16 dpi. The rTsSP is a potential early diagnostic antigen for human trichinellosis.

Schistosomiasis is a neglected infection affecting millions of people, mostly living in sub-Saharan Africa. Morbidity and mortality due to chronic infection are relevant, although schistosomiasis is often clinically silent. Different diagnostic tests have been implemented in order to improve screening and diagnosis, that traditionally rely on parasitological tests with low sensitivity. Aim of this study was to evaluate the accuracy of different tests for the screening of schistosomiasis in African migrants, in a non endemic setting. A retrospective study was conducted on 373 patients screened at the Centre for Tropical Diseases (CTD) in Negrar, Verona, Italy. Biological samples were tested with: stool/urine microscopy, Circulating Cathodic Antigen (CCA) dipstick test, ELISA, Western blot, immune-chromatographic test (ICT). Test accuracy and predictive values of the immunological tests were assessed primarily on the basis of the results of microscopy (primary reference standard): ICT and WB resulted the test with highest sensitivity (94% and 92%, respectively), with a high NPV (98%). CCA showed the highest specificity (93%), but low sensitivity (48%). The analysis was conducted also using a composite reference standard, CRS (patients classified as infected in case of positive microscopy and/or at least 2 concordant positive immunological tests) and Latent Class Analysis (LCA). The latter two models demonstrated excellent agreement (Cohen's kappa: 0.92) for the classification of the results. In fact, they both confirmed ICT as the test with the highest sensitivity (96%) and NPV (97%), moreover PPV was reasonably good (78% and 72% according to CRS and LCA, respectively). ELISA resulted the most specific immunological test (over 99%). The ICT appears to be a suitable screening test, even when used alone. The rapid test ICT was the most sensitive test, with the potential of being used as a single screening test for African migrants.

Most studies assessing praziquantel (PZQ) efficacy have used relatively insensitive diagnostic methods, thereby overestimating cure rate (CR) and intensity reduction rate (IRR). To determine accurately PZQ efficacy, we employed more sensitive DNA and circulating antigen detection methods. A sub-analysis was performed based on a previously published trial conducted in children from Côte d'Ivoire with a confirmed Schistosoma mansoni infection, who were randomly assigned to a standard (single dose of PZQ) or intense treatment group (4 repeated doses of PZQ at 2-week intervals). CR and IRR were estimated based on PCR detecting DNA in a single stool sample and the up-converting particle lateral flow (UCP-LF) test detecting circulating anodic antigen (CAA) in a single urine sample, and compared with traditional Kato-Katz (KK) and point-of-care circulating cathodic antigen (POC-CCA). Individuals positive by all diagnostic methods (i.e., KK, POC-CCA, PCR, and UCP-LF CAA) at baseline were included in the statistical analysis (n = 125). PCR showed a CR of 45% (95% confidence interval (CI) 32-59%) in the standard and 78% (95% CI 66-87%) in the intense treatment group, which is lower compared to the KK results (64%, 95% CI 52-75%) and 88%, 95% CI 78-93%). UCP-LF CAA showed a significantly lower CR in both groups, 16% (95% CI 11-24%) and 18% (95% CI 12-26%), even lower than observed by POC-CCA (31%, 95% CI 17-35% and 36%, 95% CI 26-47%). A substantial reduction in DNA and CAA-levels was observed after the first treatment, with no further decrease after additional treatment and no significant difference in IRR between treatment groups. The efficacy of (repeated) PZQ treatment was overestimated when using egg-based diagnostics (i.e. KK and PCR). Quantitative worm-based diagnostics (i.e. POC-CCA and UCP-LF CAA) revealed that active Schistosoma infections are still present despite multiple treatments. These results stress the need for using accurate diagnostic tools to monitor different PZQ treatment strategies, in particular when moving toward elimination of schistosomiasis. www.clinicaltrials.gov, NCT02868385.

Reliable diagnosis of human helminth infection(s) is essential for ongoing disease surveillance and disease elimination. Current WHO-recommended diagnostic assays are unreliable in low-endemic near-elimination settings and typically involve the invasive, onerous and potentially hazardous sampling of bodily fluids such as stool and blood, as well as tissue via biopsy. In contrast, diagnosis by use of non-invasive urine sampling is generally painless, more convenient and low risk. It negates the need for specialist staff, can usually be obtained immediately upon request and is better accepted by patients. In some instances, urine-based diagnostic assays have also been shown to provide a more reliable diagnosis of infection when compared to traditional methods that require alternative and more invasive bodily samples, particularly in low-endemicity settings. Given these relative benefits, we identify and review current research literature to evaluate whether non-invasive urine sampling is currently exploited to its full potential in the development of diagnostic tools for human helminthiases. Though further development, assessment and validation are needed before their routine use in control programmes, low-cost, rapid and reliable assays capable of detecting transrenal helminth-derived antigens and cell-free DNA show excellent promise for future use at the point-of-care in high-, medium- and even low-endemicity elimination settings.

The global elimination of lymphatic filariasis (LF) is a major focus of the World Health Organization. One key challenge is locating residual infections that can perpetuate the transmission cycle. We show how a targeted sampling strategy using predictions from a geospatial model, combining random forests and geostatistics, can improve the sampling efficiency for identifying locations with high infection prevalence. Predictions were made based on the household locations of infected persons identified from previous surveys, and environmental variables relevant to mosquito density. Results show that targeting sampling using model predictions would have allowed 52% of infections to be identified by sampling just 17.7% of households. The odds ratio for identifying an infected individual in a household at a predicted high risk compared to a predicted low risk location was 10.2 (95% CI 4.2–22.8). This study provides evidence that a ‘one size fits all’ approach is unlikely to yield optimal results when making programmatic decisions based on model predictions. Instead, model assumptions and definitions should be tailored to each situation based on the objective of the surveillance program. When predictions are used in the context of the program objectives, they can result in a dramatic improvement in the efficiency of locating infected individuals.

Parasite-released extracellular vesicles (EVs) deliver signals to the host immune system that are critical to maintaining the long-term relationship between parasite and host. In the present study, total EVs (FhEVs) released in vitro by adults of the helminth parasite Fasciola hepatica were isolated using a recently described gravity flow method that protects their structural integrity. The FhEVs molecular cargo was defined using proteomic analysis and their surface topology characterised by glycan microarrays. The proteomic analysis identified 618 proteins, 121 of which contained putative N-linked glycosylation sites while 132 proteins contained putative O-linked glycosylation sites. Glycan arrays revealed surface-exposed glycans with a high affinity for mannose-binding lectins indicating the predominance of oligo mannose-rich glycoproteins, as well as other glycans with a high affinity for complex-type N-glycans. When added to bone-marrow derived dendritic cells isolated FhEV induced a novel phenotype that was categorised by the secretion of low levels of TNF, enhanced expression of cell surface markers (CD80, CD86, CD40, OX40L, and SIGNR1) and elevation of intracellular markers (SOCS1 and SOCS3). When FhEV-stimulated BMDCs were introduced into OT-II mice by adoptive transfer, IL-2 secretion from skin draining lymph nodes and spleen cells was inhibited in response to both specific and non-specific antigen stimulation. Immunisation of mice with a suspension of FhEV did not elicit significant immune responses; however, in the presence of alum, FhEVs induced a mixed Th1/Th2 immune response with high antigen specific antibody titres. Thus, we have demonstrated that FhEVs induce a unique phentotype in DC capable of suppressing IL-2 secretion from T-cells. Our studies add to the growing immuno-proteomic database that will be an important source for the discovery of future parasite vaccines and immunotherapeutic biologicals.

Background For years, the Kato-Katz (KK) technique has been considered the gold standard for diagnosing schistosomiasis. The aim of this study was to compare the effectiveness of our previously developed gold nanoparticle-based lateral flow test strip (AuNPs-LFTS) for diagnosing active Schistosoma mansoni with that of the commercially available point-of-care Circulating Cathodic Antigen detection (POC-CCA) kit. Methods In this study, we collected sixty positive and twenty negative urine samples from patients in endemic hot spots in the Nile Delta, as well as from patients visiting the internal medicine clinic at Theodor Bilharz Research Institute (TBRI). We produced monoclonal antibodies (MAbs) against S. mansoni soluble egg antigen (SEA) from cloned hybridoma cells (4D/1D). These MAbs were conjugated with gold and mesoporous silica nanoparticles, and used to develop the LFTS. Results The LFTS demonstrated a limit of detection (LoD) of 3 ng/ml. The sensitivity and specificity of the developed LFTS were found to be 96.7% and 95%, respectively, compared to 85% and 90% for the POC-CCA detection kit. The cases were divided into groups based on egg count in the stool, categorized as light, moderate, and heavy infections. The sensitivity of the LFTS in the group with light infection was higher than that of the POC-CCA. When using the KK technique (eggs per gram of stool sample [EPG]) as the reference test, the kappa value for the nano-based strips was 0.902, compared to 0.672 for the CCA strips, indicating an almost perfect agreement between KK and our developed LFTS. Conclusion These results confirm the reliability and effectiveness of the LFTS compared to commercially available kits for rapid, sensitive, and early diagnosis of schistosomiasis. However, it is recommended to conduct further assessments of the developed strip on a larger scale with a broader range of cases before considering its introduction to local or international markets. Graphical Abstract

Despite the successful removal of traces, the elimination of potential false positivity resulted in no detection of true positives. [...]individuals with active schistosomiasis were misdiagnosed as negative by the new test version in low endemic areas. In the past, custom-tailored kits were tested, aiming to overcome the debatable accuracy and performance of POC-CCA in a low-prevalence population of school children [18]. Because only limited information was disclosed by the manufacturers, one assumes that changes on test sensitivity were also performed since the last commercially available assay. [...]predefinition of trace as positive or negative may end up misleading test interpretation. POC-CCA very low intensity reactions may hide low parasite–load-induced infections. [...]removal of trace might result in no detection of Schistosoma infection pre- and post-PZQ use and, as a result, improvement of the test must be encouraged.

The aim of this study was to determine if the protozoa that cause dysentery might have been present in Jerusalem, the capital of the Kingdom of Judah, during the Iron Age. Sediments from 2 latrines pertaining to this time period were obtained, 1 dating from the 7th century BCE and another from the 7th to early 6th century BCE. Microscopic investigations have previously shown that the users were infected by whipworm (Trichuris trichiura), roundworm (Ascaris lumbricoides), Taenia sp. tapeworm and pinworm (Enterobius vermicularis). However, the protozoa that cause dysentery are fragile and do not survive well in ancient samples in a form recognizable using light microscopy. Enzyme-linked immunosorbent assay kits designed to detect the antigens of Entamoeba histolytica, Cryptosporidium sp. and Giardia duodenalis were used. Results for Entamoeba and Cryptosporidium were negative, while Giardia was positive for both latrine sediments when the analysis was repeated three times. This provides our first microbiological evidence for infective diarrhoeal illnesses that would have affected the populations of the ancient near east. When we integrate descriptions from 2nd and 1st millennium BCE Mesopotamian medical texts, it seems likely that outbreaks of dysentery due to giardiasis may have caused ill health throughout early towns across the region.