Identification of optimal epitopes for Plasmodium falciparum rapid diagnostic tests that target histidine-rich proteins 2 and 3

Rapid diagnostic tests (RDTs) represent important tools to diagnose malaria infection. To improve understanding of the variable performance of RDTs that detect the major target in Plasmodium falciparum, namely, histidine-rich protein 2 (HRP2), and to inform the design of better tests, we undertook detailed mapping of the epitopes recognized by eight HRP-specific monoclonal antibodies (MAbs). To investigate the geographic skewing of this polymorphic protein, we analyzed the distribution of these epitopes in parasites from geographically diverse areas. To identify an ideal amino acid motif for a MAb to target in HRP2 and in the related protein HRP3, we used a purpose-designed script to perform bioinformatic analysis of 448 distinct gene sequences from pfhrp2 and from 99 sequences from the closely related gene pfhrp3. The frequency and distribution of these motifs were also compared to the MAb epitopes. Heat stability testing of MAbs immobilized on nitrocellulose membranes was also performed. Results of these experiments enabled the identification of MAbs with the most desirable characteristics for inclusion in RDTs, including copy number and coverage of target epitopes, geographic skewing, heat stability, and match with the most abundant amino acid motifs identified. This study therefore informs the selection of MAbs to include in malaria RDTs as well as in the generation of improved MAbs that should improve the performance of HRP-detecting malaria RDTs.

Blood transfer devices for malaria rapid diagnostic tests : evaluation of accuracy, safety and ease of use

Background: Malaria rapid diagnostic tests (RDTs) are increasingly used by remote health personnel with minimal training in laboratory techniques. RDTs must, therefore, be as simple, safe and reliable as possible. Transfer of blood from the patient to the RDT is critical to safety and accuracy, and poses a significant challenge to many users. Blood transfer devices were evaluated for accuracy and precision of volume transferred, safety and ease of use, to identify the most appropriate devices for use with RDTs in routine clinical care. Methods: Five devices, a loop, straw-pipette, calibrated pipette, glass capillary tube, and a new inverted cup device, were evaluated in Nigeria, the Philippines and Uganda. The 227 participating health workers used each device to transfer blood from a simulated finger-prick site to filter paper. For each transfer, the number of attempts required to collect and deposit blood and any spilling of blood during transfer were recorded. Perceptions of ease of use and safety of each device were recorded for each participant. Blood volume transferred was calculated from the area of blood spots deposited on filter paper. Results: The overall mean volumes transferred by devices differed significantly from the target volume of 5 microliters (p < 0.001). The inverted cup (4.6 microliters) most closely approximated the target volume. The glass capillary was excluded from volume analysis as the estimation method used is not compatible with this device. The calibrated pipette accounted for the largest proportion of blood exposures (23/225, 10%); exposures ranged from 2% to 6% for the other four devices. The inverted cup was considered easiest to use in blood collection (206/ 226, 91%); the straw-pipette and calibrated pipette were rated lowest (143/225 [64%] and 135/225 [60%] respectively). Overall, the inverted cup was the most preferred device (72%, 163/227), followed by the loop (61%, 138/227). Conclusions: The performance of blood transfer devices varied in this evaluation of accuracy, blood safety, ease of use, and user preference. The inverted cup design achieved the highest overall performance, while the loop also performed well. These findings have relevance for any point-of-care diagnostics that require blood sampling.

Global sequence variation in the histidine-rich proteins 2 and 3 of Plasmodium falciparum: implications for the performance of malaria rapid diagnostic tests

Background Accurate diagnosis is essential for prompt and appropriate treatment of malaria. While rapid diagnostic tests (RDTs) offer great potential to improve malaria diagnosis, the sensitivity of RDTs has been reported to be highly variable. One possible factor contributing to variable test performance is the diversity of parasite antigens. This is of particular concern for Plasmodium falciparum histidine-rich protein 2 (PfHRP2)-detecting RDTs since PfHRP2 has been reported to be highly variable in isolates of the Asia-Pacific region. Methods The pfhrp2 exon 2 fragment from 458 isolates of P. falciparum collected from 38 countries was amplified and sequenced. For a subset of 80 isolates, the exon 2 fragment of histidine-rich protein 3 (pfhrp3) was also amplified and sequenced. DNA sequence and statistical analysis of the variation observed in these genes was conducted. The potential impact of the pfhrp2 variation on RDT detection rates was examined by analysing the relationship between sequence characteristics of this gene and the results of the WHO product testing of malaria RDTs: Round 1 (2008), for 34 PfHRP2-detecting RDTs. Results Sequence analysis revealed extensive variations in the number and arrangement of various repeats encoded by the genes in parasite populations world-wide. However, no statistically robust correlation between gene structure and RDT detection rate for P. falciparum parasites at 200 parasites per microlitre was identified. Conclusions The results suggest that despite extreme sequence variation, diversity of PfHRP2 does not appear to be a major cause of RDT sensitivity variation.

Tests on price linkage between the U.S. and Japanese gold and silver futures markets

We tested the price linkage, the law of one price (LOP) condition, and the causality of the price linkage between the U.S. and Japanese gold and silver futures markets with consideration of structural breaks in the price series. The LOP condition did not hold for both the gold and silver markets when structural breaks were not considered but it sustained in some periods when it was tested for the break periods. We found from the causality test that the price linkage between the U.S. and Japanese gold and silver futures markets were led by the U.S. market.

Buckling tests of fire exposed cold-formed steel columns

Cold-formed steel sections are commonly used in low-rise commercial and residential buildings. During fire events, cold-formed steel structural elements in these buildings will be exposed to elevated temperatures. Hence after such events there is a need to evaluate the residual strength of these structural elements. However, only limited information is available in relation to the residual strength of fire exposed cold-formed steel sections. This means conservative decisions are often made in relation to fire exposed building structures. This research is aimed at investigating the buckling capacities of fire exposed cold-formed lipped channel steel sections. A series of compression tests of fire exposed, short lipped channel columns made of varying steel grades and thicknesses was undertaken in this research. Test columns were first exposed to different elevated temperatures up to 800 oC. They were then allowed to cool down at ambient temperatures before they were tested to failure. Similarly tensile coupon tests were also undertaken after being exposed to various elevated temperatures, from which the residual mechanical properties (yield stress and Young’s modulus) of the steels used in this study were derived. Using these mechanical properties, the residual compression capacities of tested short columns were predicted using the currently used design rules in AS/NZS 4600 and AISI cold-formed steel standards. This comparison showed that ambient temperature design rules for compression members can be used to predict the residual compression capacities of fire exposed short or laterally restrained cold-formed steel columns provided the maximum temperature experienced by the columns can be estimated after a fire event. Such residual capacity assessments will allow structural and fire engineers to make an accurate prediction of the safety of fire exposed buildings. This paper presents the details of this experimental study and the results.

Circulating antibodies against Plasmodium falciparum histidine-rich proteins 2 interfere with antigen detection by rapid diagnostic tests

Background Rapid diagnostic tests (RDTs) for detection of Plasmodium falciparum infection that target P. falciparum histidine-rich protein 2 (PfHRP2), a protein that circulates in the blood of patients infected with this species of malaria, are widely used to guide case management. Understanding determinants of PfHRP2 availability in circulation is therefore essential to understanding the performance of PfHRP2-detecting RDTs. Methods The possibility that pre-formed host anti-PfHRP2 antibodies may block target antigen detection, thereby causing false negative test results was investigated in this study. Results Anti-PfHRP2 antibodies were detected in 19/75 (25%) of plasma samples collected from patients with acute malaria from Cambodia, Nigeria and the Philippines, as well as in 3/28 (10.7%) asymptomatic Solomon Islands residents. Pre-incubation of plasma samples from subjects with high-titre anti-PfHRP2 antibodies with soluble PfHRP2 blocked the detection of the target antigen on two of the three brands of RDTs tested, leading to false negative results. Pre-incubation of the plasma with intact parasitized erythrocytes resulted in a reduction of band intensity at the highest parasite density, and a reduction of lower detection threshold by ten-fold on all three brands of RDTs tested. Conclusions These observations indicate possible reduced sensitivity for diagnosis of P. falciparum malaria using PfHRP2-detecting RDTs among people with high levels of specific antibodies and low density infection, as well as possible interference with tests configured to detect soluble PfHRP2 in saliva or urine samples. Further investigations are required to assess the impact of pre-formed anti-PfHRP2 antibodies on RDT performance in different transmission settings.

Pan-Plasmodium band sensitivity for Plasmodium falciparum detection in combination malaria rapid diagnostic tests and implications for clinical management

Background: Malaria rapid diagnostic tests (RDTs) are appropriate for case management, but persistent antigenaemia is a concern for HRP2-detecting RDTs in endemic areas. It has been suggested that pan-pLDH test bands on combination RDTs could be used to distinguish persistent antigenaemia from active Plasmodium falciparum infection, however this assumes all active infections produce positive results on both bands of RDTs, an assertion that has not been demonstrated. Methods: In this study, data generated during the WHO-FIND product testing programme for malaria RDTs was reviewed to investigate the reactivity of individual test bands against P. falciparum in 18 combination RDTs. Each product was tested against multiple wild-type P. falciparum only samples. Antigen levels were measured by quantitative ELISA for HRP2, pLDH and aldolase. Results: When tested against P. falciparum samples at 200 parasites/μL, 92% of RDTs were positive; 57% of these on both the P. falciparum and pan bands, while 43% were positive on the P. falciparum band only. There was a relationship between antigen concentration and band positivity; ≥4 ng/mL of HRP2 produced positive results in more than 95% of P. falciparum bands, while ≥45 ng/mL of pLDH was required for at least 90% of pan bands to be positive. Conclusions: In active P. falciparum infections it is common for combination RDTs to return a positive HRP2 band combined with a negative pan-pLDH band, and when both bands are positive, often the pan band is faint. Thus active infections could be missed if the presence of a HRP2 band in the absence of a pan band is interpreted as being caused solely by persistent antigenaemia.

Web crippling tests of cold-formed steel channels under two flange load cases

Cold-formed steel members have many advantages over hot-rolled steel members. However, they are susceptible to various buckling modes at stresses below the yield stress of the member because of their relatively high width-to-thickness ratio. Web crippling is a form of localized failure mode that can occur when the members are subjected to transverse high concentrated loadings and/or reactions. The four common loading conditions are the end-one-flange (EOF), interior-one-flange (IOF), end-two-flange (ETF) and interior-two-flange (ITF) loadings. Recently a test method has been proposed by AISI to obtain the web crippling capacities under these four loading conditions. Using this test method 42 tests were conducted in this research to investigate the web crippling behaviour and strengths of unlipped channels with stocky webs under ETF and ITF cases. DuraGal sections having a nominal yield stress of 450 MPa were tested with different web slenderness and bearing lengths. The flanges of these channel sections were not fastened to the supports. In this research the suitability of the currently available design rules for unlipped channels subject to web crippling was investigated, and suitable modifications were proposed where necessary. In addition to this, a new design rule was proposed based on the direct strength method to predict the web crippling capacities of tested beams. This paper presents the details of this experimental study and the results.

Pull-through failure tests of thin steel roof battens under wind uplift load

Thin profiled steel roof sheeting and battens are increasingly used in the construction of roofing systems of residential, commercial, industrial and farm buildings in Australia. The critical load combination of external wind suction and internal wind pressures that occur during high wind events such as thunderstorms and tropical cylcones often dislocate the roofing systems partially or even completely due to premature roof connection failures. Past wind damage investigations have shown that roof sheeting failures occured at their screw connections to battens. In most of these cases, the screw fastener head pulled through the thin roof sheeting whilst the screw fasteners also pulled out from the battens. Research studis undertaken on the roof sheeting to batten connection failures have improved this situation. However, the batten to rafter or truss connections have not been investigated adequately. Failure of these connections can cause the failure of the entire roof structure as observed during the recent high wind events. Therefore a detailed experimental study consisting of both small scale and full scale tests has been undertaken to investigate the steel roof batten pull-through failures in relation to many critical parameters such as steel batten geometry, thickness and grade, screw fastener head sizes and screw tightening. This paper presents the details of this experimental study and the pull-through failure load results obtained from them. Finally it discusses the development of suitable design rules that can be used to determine the pull-through connection capacities of thin steel roof battens under wind uplift loads.

Relationship between Barthel index with physical tests in adults with intellectual disabilities

We usually find low levels of fitness condition affect other aspects of living for people with ID like dependency in carrying out activivities of daily living. Therefore we find high levels of dependency in activities of daily living due to poor fitness condition. The aim of the study is to explore the criterion validity of the Barthel index with a physical fitness test. An observational cross-sectional study was conducted. Data from the Barthel index and a physical fitness test were measured in 122 adults with intellectual disability. The data were analysed to find out the relationship between four categories of the physical fitness test and the Barthel index. It needs to be stressed that the correlations between the Barthel index and leg, abdominal and arm strength can confirm that these physical test are predictive of the Barthel index. The correlations between the balance variables as functional reach and single-leg stance with eyes open shown relationships with Barthel Index. We found important correlations between the physical fitness test and the Barthel index, so we can affirm that some physical fitness features are predictor variables of the Barthel index.

Web crippling tests of rivet fastened rectangular hollow flange channel beams under two flange load cases

Recent research on hollow flange beams has led to the development of an innovative rectangular hollow flange channel beam (RHFCB) for use in floor systems. The new RHFCB is a mono-symmetric structural section made by intermittently rivet fastening two torsionally rigid closed rectangular hollow flanges to a web plate element, which allows section optimisation by selecting appropriate combinations of web and flange widths and thicknesses. However, the current design rules for cold-formed steel sections are not directly applicable to rivet fastened RHFCBs. To date, no investigation has been conducted on their web crippling behaviour and strengths. Hence an experimental study was conducted to investigate the web crippling behaviour and capacities of rivet fastened RHFCBs under End Two Flange (ETF) and Interior Two Flange (ITF) load cases. It showed that RHFCBs failed by web crippling, flange crushing and their combinations. Comparison of ultimate web crippling capacities with the predictions from the design equations in AS/NZS 4600 and AISI S100 showed that the current design equations are unconservative for rivet fastened RHFCB sections under ETF and ITF load cases. Hence new equations were proposed to determine the web crippling capacities of rivet fastened RHFCBs. These equations can also be used to predict the capacities of RHFCBs subject to combined web crippling and flange crushing conservatively. However, new capacity equations were proposed in the case of flange crushing failures that occurred in thinner flanges with smaller bearing lengths. This paper presents the details of this web crippling experimental study of RHFCB sections and the results.

Shear tests of hollow flange channel beams with real support conditions

This paper presents the details of experimental studies on the effect of real support conditions on the shear strength of hollow flange channel beams, known as LiteSteel beams (LSB). The LSB has a unique shape of a channel beam with two rectangular hollow flanges, made using a unique manufacturing process. In many applications in the building industry LSBs are used with only one web side plate (WSP) at their supports. The WSPs are also often not full height plates. Past research studies showed that these real support connections did not provide the required simply supported conditions. Many studies have been carried out to evaluate the behaviour and design of LSBs with simply supported conditions subject to pure bending and predominant shear actions. To date, however, no investigation has been conducted into the effect of real support conditions on the shear strength of LSBs. Hence a detailed experimental study based on 25 shear tests was undertaken to investigate the shear behaviour and strength of LSBs with real support conditions. Simply supported test specimens of LSBs with aspect ratios of 1.0 and 1.5 were loaded at mid-span until failure. It was found that the effect of using one WSP on the shear behaviour of LSB is significant and there is about 25% shear capacity reduction due to the lateral movement of the bottom flange at the supports. Shear capacity of LSB was also found to decrease when full height WSPs were not used. Suitably improved support connections were developed to improve the shear capacity of LSBs based on test results. Details of the recommended support connections and shear capacity results are given in this paper.

Use of objective psychomotor tests in health professionals

Evaluation of psychomotor skills is undertaken in a number of broad contexts. This includes testing of health professional populations as a measure of innate ability, to evaluate skill acquisition, or to compare professions. However, the use of psychomotor tests is frequently confounded by a lack of understanding of a particular tool's psychometric properties, strengths, and weaknesses. To identify and appraise the most commonly used tests on health professional populations, 86 articles were reviewed and the top nine tests identified. Few tests have had sufficient validity or reliability testing on health professionals. Based on the evidence available, use of the Grooved Pegboard Test, the Purdue Pegboard Test, or the Finger Tapping Test is recommended for the evaluation of dexterity in a health professional population; however, this choice may be dependent on the task(s) to which findings are generalised. More rigorous evaluation of validity and other psychometric properties is required.