METHODS: A total of 50 serum samples were collected from patients clinically suspected for acute leptospirosis on admission in the Hospital Serdang, from June 2016 to June 2017. All the samples were subjected to MAT, lipL32 PCR and the two rapid tests (Leptocheck-WB and ImmuneMed Leptospira IgM Duo Rapid test).
RESULTS: Out of the 50 clinically suspected patients sampled, 19 were confirmed positive for leptospirosis. Six (12%) were confirmed by MAT and 13 (26%) by PCR. Similarly, of the 50 clinically suspected cases, 17 (34%) showed positivity for Leptocheck-WB and 7 (14%) for ImmuneMed Leptospira IgM Duo Rapid test. The overall sensitivity and specificity was 47.37% and 80.65% for Leptocheck-WB, and 21.05% and 90.32% for ImmuneMed Leptospira IgM Duo Rapid test. In another set of previously confirmed MAT positive samples (1:400-1:3600) obtained from a reference laboratory, Leptocheck-WB showed higher sensitivity (90.72%) than ImmuneMed Leptospira IgM Duo Rapid test (40.21%), and comparable specificity for ImmuneMed Leptospira IgM Duo Rapid test (88.89%) and Leptocheck-WB (82.86%).
CONCLUSION: The sensitivity was higher for Leptocheck-WB and had a comparable specificity with ImmuneMed Leptospira IgM Duo Rapid test. Therefore, based on the present study, Leptocheck-WB is found to be a more sensitive rapid immunodiagnostic test for acute leptospirosis screening in hospital settings.
METHODS: The halogen lamp was replaced by a low-cost, blue light-emitting diode (LED) lamp. Using a reformulated AO solution, the staining protocol was revised to make use of a concentration gradient instead of uniform staining. To evaluate this new AO diagnostic system, a pilot field study was conducted in the Lake Victoria basin in Kenya.
RESULTS: Without staining failure, malaria infection status of about 100 samples was determined on-site per one microscopist per day, using the improved AO diagnostic system. The improved AO diagnosis had both higher overall sensitivity (46.1 vs 38.9%: p = 0.08) and specificity (99.0 vs 96.3%) than the Giemsa method (N = 1018), using PCR diagnosis as the standard.
CONCLUSIONS: Consistent AO staining of thin blood films and rapid evaluation of malaria parasitaemia with the revised protocol produced superior results relative to the Giemsa method. This AO diagnostic system can be set up easily at low cost using an ordinary light microscope. It may supplement rapid diagnostic tests currently used in clinical settings in malaria-endemic countries, and may be considered as an inexpensive tool for case surveillance in malaria-eliminating countries.
METHODS: In-depth individual interviews with thematic saturation were conducted between May and July 2018. The data was analyzed using thematic analysis.
RESULTS: Based on expert opinion, diagnosis of severe dengue is challenging as it depends on astute clinical interpretation of non-dengue-specific clinical and laboratory findings. A specific test that detects impending manifestation of severe dengue could 1) overcome failure in identifying severe disease for referral or admission, 2) facilitate timely and appropriate management of plasma leakage and bleeding, 3) overcome the lack of clinical expertise and laboratory diagnosis in rural health settings. The most important feature of any diagnostics for severe dengue is the point-of-care (POC) format where it can be performed at or near the bedside.
CONCLUSION: The development of diagnostics to detect impending severe dengue is warranted to reduce the morbidity and mortality rates of dengue infection and it should be prioritized.
METHODS: Ten RDTs were evaluated: nine to detect clinical P. knowlesi infections from Malaysia, and nine assessing limit of detection (LoD) for P. knowlesi (PkA1-H.1) and P. falciparum (Pf3D7) cultures. Targets included Plasmodium-genus parasite lactate dehydrogenase (pan-pLDH) and P. vivax (Pv)-pLDH.
RESULTS: Samples were collected prior to antimalarial treatment from 127 patients with microscopy-positive PCR-confirmed P. knowlesi mono-infections. Median parasitaemia was 788/µL (IQR 247-5,565/µL). Pan-pLDH sensitivities ranged from 50.6% (95% CI 39.6-61.5) (SD BIOLINE) to 87.0% (95% CI 75.1-94.6) (First Response® and CareStart™ PAN) compared to reference PCR. Pv-pLDH RDTs detected P. knowlesi with up to 92.0% (95% CI 84.3-96.7%) sensitivity (Biocredit™). For parasite counts ≥200/µL, pan-pLDH (Standard Q) and Pv-pLDH RDTs exceeded 95% sensitivity. Specificity of RDTs against 26 PCR-confirmed negative controls was 100%. Sensitivity of six highest performing RDTs were not significantly different when comparing samples taken before and after (median 3 hours) antimalarial treatment. Parasite ring stages were present in 30% of pre-treatment samples, with ring stage proportions (mean 1.9%) demonstrating inverse correlation with test positivity of Biocredit™ and two CareStart™ RDTs.For cultured P. knowlesi, CareStart™ PAN demonstrated the lowest LoD at 25 parasites/µL; LoDs of other pan-pLDH ranged from 98 to >2000 parasites/µL. Pv-pLDH LoD for P. knowlesi was 49 parasites/µL. No false-positive results were observed in either P. falciparum-pLDH or histidine-rich-protein-2 channels.
CONCLUSION: Selected RDTs demonstrate sufficient performance for detection of major human malaria species including P. knowlesi in co-endemic areas where microscopy is not available, particularly for higher parasite counts, although cannot reliably differentiate among non-falciparum malaria.
METHODS: In this study, new binders derived from shark VNAR domains library were investigated. Following immunization of a wobbegong shark (Orectolobus ornatus) with three recombinant malaria biomarker proteins (PfHRP2, PfpLDH and Pvaldolase), a single domain antibody (sdAb) library was constructed from splenocytes. Target-specific VNAR phage were isolated by panning. One specific clone was selected for expression in Escherichia coli expression system, and study of binding reactivity undertaken.
RESULTS: The primary VNAR domain library possessed a titre of 1.16 × 106 pfu/mL. DNA sequence analysis showed 82.5% of isolated fragments appearing to contain an in-frame sequence. After multiple rounds of biopanning, a highly dominant clone specific to PfHRP2 was identified and selected for protein production in an E. coli expression system. Biological characterization showed the recombinant protein expressed in periplasmic has better detection sensitivity than that of cytoplasmic proteins. Assays of binding activity indicated that its reactivity was inferior to the positive control mAb C1-13.
CONCLUSIONS: Target-specific bacteriophage VNARs were successfully isolated after a series of immunization, demonstrating that phage display technology is a useful tool for selection of antigen binders. Generation of new binding reagents such as VNAR antibodies that specifically recognize the malaria biomarkers represents an appealing approach to improve the performance of RDTs.