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Mark-recapture techniques have been widely used and specialized to study organisms throughout the field of biology. To mark-recapture ticks (Ixodida), we have created a simple method to mark ticks using nail polish applied with an insect pin secured in a pencil that allows for a variety of questions to be answered. For measuring tick control efficacy, estimating population estimates, or measuring movement of ticks, this inexpensive mark-recapture method has been easily applied in the field and in the lab to provide useful data to answer a variety of questions about ticks., (© The Author(s) 2020. Published by Oxford University Press on behalf of Entomological Society of America.)

Objective: To establish an evaluation system for the field assessment of the Kato-Katz technique in detecting soiltransmitted nematodes, so as to provide insights into the field application of the Kato-Katz technique., Methods: The initial evaluation indicators were determined through literature search, brainstorming and expert consultation. The evaluation indicatorswere improved and the weight of each indicator was decided through three rounds of expert consultation by using the Delphimethod. In addition, the expert authority coefficient and the coordination coefficient of each indicator were calculated at eachround of expert consultations., Results: The recovery rates of the questionnaire were 100.00%, 100.00% and 89.29% at the firstto the third round of the expert consultations, respectively, and the expert authority coefficients were all more than 0.85 at eachround. The final evaluation system included 4 first-level indicators and 15 second-level indicators. In the first-level indicators, "detecting effect" and "funds investment" had the mean weighted value of 4.53 and 4.49, which were relatively higher than that of"person-time investment" and "operability" (both 4.34). Among the second-level indicators under each first-level indicator, thefour most significant indicators included "ability of personnel in egg discrimination", "cooperation of village cadres and doctors","Person-time on testing" and "organizational start-up cost", with the mean weighted values of 4.74, 4.43, 4.39 and 4.17, respectively. The coordination coefficients were 0.39 to 0.65, 0.28 to 0.58 and 0.45 to 0.65 at the first to the third round of the expertconsultations, respectively, and there were significant differences in the coordination coefficients at all three rounds of the consultations (all P < 0.05)., Conclusions: An evaluation system for the field assessment of the Kato-Katz technique in detecting soiltransmitted nematodes is successfully established, among which "ability of personnel in egg discrimination" and "cooperation ofvillage cadres and doctors" have the greatest mean weighted values of the significance.

Giardia cysts stained with hot carbolfuchsin were used as internal controls in a concentration method for surface water samples. The morphological integrity of stained cysts and the stain's stability and intensity were tested with each of the chemical reagents used in the aluminum sulfate flocculation method. No alterations in morphology or color were noted. The stained cyst preparation has a low cost, high stability, and suitability for both light and immunofluorescent microscopy, making it affordable to researchers in low- and middle-income countries., (Copyright © 2019 Elsevier B.V. All rights reserved.)

The purpose of the study was to substantiate the theoretical and methodical principles of pricing for laboratory services in the diagnosis of giardiasis, taking into account their iterability and peculiarities of parasitic research methods. The methods of laboratory study of gum disease of native smear, treated with Lyulol solution, and ether-formalin enrichment on the criteria of their quality and effectiveness are analyzed. On the basis of the study of the effectiveness of the first and repeated analyzes, the conclusion on the iterative nature of laboratory studies of giardiasis and the effectiveness of the use of an iterative approach to the determination of prices for laboratory services is substantiated. The approaches to pricing laboratories providing diagnostic services for giardiasis in Ukraine are analyzed. The necessity of applying the price trajectory for laboratory diagnosis of giardiasis on the basis of multiplicity of researches (interactive approach) and the determination of the minimum and maximum price levels (the minimax approach) is proved. The main factors of pricing for laboratory diagnostics of giardiasis are identified and characterized: iterative research, economic efficiency, social value, value for the patient, competitiveness and reputation.

Fifty years after anthelmintic resistance in livestock parasites was first reported, the prevalence of resistance has increased globally, and is of increasing significance in animal industries. It is now timely to reflect on what we have learnt, how research has unfolded, and what we hope to learn in the future. This Opinion paper examines ten important research events that were pivotal in resistance research. The moments include the discovery, description, and diagnosis of parasite resistance, as well as important physiological and genetic findings, and the development of online tools to help manage resistance. Despite our efforts, resistance remains the greatest challenge in parasite control. The future directions for research, including people and funding, are discussed., (Copyright © 2018 Elsevier Ltd. All rights reserved.)

Laboratory workers, in resource-poor countries, still consider PCR detection of Giardia lamblia more costly and more time-consuming than the classical parasitological techniques. Based on 2 published primers, an in-house one-round touchdown PCR-RFLP assay was developed. The assay was validated with an internal amplification control included in reactions. Performance of the assay was assessed with DNA samples of various purities, 91 control fecal samples with various parasite load, and 472 samples of unknown results. Two cysts per reaction were enough for PCR detection by the assay with exhibited specificity (Sp) and sensitivity (Se) of 100% and 93%, respectively. Taking a published small subunit rRNA reference PCR test results (6%; 29/472) as a nominated gold standard, G. lamblia was identified in 5.9% (28/472), 5.2%, (25/472), and 3.6% (17/472) by PCR assay, RIDA(®) Quick Giardia antigen detection test (R-Biopharm, Darmstadt, Germany), and iodine-stained smear microscopy, respectively. The percent agreements (kappa values) of 99.7% (0.745), 98.9% (0.900), and 97.7% (0.981) were exhibited between the assay results and that of the reference PCR, immunoassay, and microscopy, respectively. Restriction digestion of the 28 Giardia-positive samples revealed genotype A pattern in 12 and genotype B profile in 16 samples. The PCR assay with the described format and exhibited performance has a great potential to be adopted in basic clinical laboratories as a detection tool for G. lamblia especially in asymptomatic infections. This potential is increased more in particular situations where identification of the parasite genotype represents a major requirement as in epidemiological studies and infection outbreaks.

An easy and low-cost method to elaborate a container to dehydrate nematodes and other meiofauna in order to process them for scanning electron microscopy (SEM) is presented. Illustrations of its elaboration, step by step, are included. In addition, a brief methodology to process meiofauna, especially nematodes and kinorhynchs, and illustrations are provided. With this methodology it is possible to easily introduce the specimens, to lock them in a closed chamber allowing the infiltration of fluids and gases (ethanol, acetone, carbon dioxide) but avoiding losing the specimens. After using this meiofauna basket for SEM the results are efficient. Examples of nematode and kinorhynch SEM pictures obtained using this methodology are also included., (© 2015 Wiley Periodicals, Inc.)

Background: As efforts to contain artemisinin resistance and eliminate Plasmodium falciparum intensify, the accurate diagnosis and prompt effective treatment of malaria are increasingly needed in Myanmar and the Greater Mekong Sub-region (GMS). Rapid diagnostic tests (RDTs) have been shown to be safe, feasible, and effective at promoting appropriate treatment for suspected malaria, which are of particular importance to drug resistance containment. The informal private sector is often the first point of care for fever cases in malaria endemic areas across Myanmar and the GMS, but there is little published information about informal private provider practices, quality of service provision, or potential to contribute to malaria control and elimination efforts. This study tested different incentives to increase RDT use and improve the quality of care among informal private healthcare providers in Myanmar., Methods: The study randomized six townships in the Mon and Shan states of rural Myanmar into three intervention arms: 1) RDT price subsidies, 2) price subsidies with product-related financial incentives, and 3) price subsidies with intensified information, education and counselling (IEC). The study assessed the uptake of RDT use in the communities by cross-sectional surveys of 3,150 households at baseline and six months post-intervention (6,400 households total, 832 fever cases). The study also used mystery clients among 171 providers to assess quality of service provision across intervention arms., Results: The pilot intervention trained over 600 informal private healthcare providers. The study found a price subsidy with intensified IEC, resulted in the highest uptake of RDTs in the community, as compared to subsidies alone or merchandise-related financial incentives. Moreover, intensified IEC led to improvements in the quality of care, with mystery client surveys showing almost double the number of correct treatment following diagnostic test results as compared to a simple subsidy., Conclusions: Results show that training and quality supervision of informal private healthcare providers can result in improved demand for, and appropriate use of RDTs in drug resistance containment areas in eastern Myanmar. Future studies should assess the sustainability of such interventions and the scale and level of intensity required over time as public sector service provision expands.

Comparisons were undertaken to investigate cost-effective methods of implementing the enzyme-linked immunosorbent assay (ELISA) for sporozoite determination in anophelines when large numbers require processing. Comparisons between ELISA plate reader and visual assessments were performed with Anopheles funestus and Anopheles gambiae s.l. (Diptera: Culicidae), as were comparisons between whole-body mosquito samples, heads and thoraces, and abdomens alone. Rates obtained from pools of five or 10 mosquitoes were compared with those for individual mosquitoes, as were rates obtained using different sampling methods. A total of 41 792 An. funestus and 9431 An. gambiae s.l. collected in light traps, and 22 323 An. funestus and 6860 An. gambiae s.l. from exit collections were analysed. Visual assessments gave results similar to those of machine readings. Sporozoite rates were similar in both species, as were rates by collection method. The use of whole mosquitoes increased estimates of infection rate by 0.6%. Pool size did not affect infection rates of An. gambiae s.l., but rates were higher among individually tested An. funestus than among those tested in pools. For large-scale surveys, the use of whole mosquitoes in pools of 10 mosquitoes, with correction for overestimation, and the noting of results according to a simple three-stage visual assessment of positivity is the most cost-effective approach and is sufficient to obtain reliable data for comparative purposes., (© 2014 The Royal Entomological Society.)

Background: The emergence of artemisinin-resistant Plasmodium falciparum parasites in Southeast Asia threatens global malaria control efforts. One strategy to counter this problem is a subsidy of malaria rapid diagnostic tests (RDTs) and artemisinin-based combination therapy (ACT) within the informal private sector, where the majority of malaria care in Myanmar is provided. A study in Myanmar evaluated the effectiveness of financial incentives vs information, education and counselling (IEC) in driving the proper use of subsidized malaria RDTs among informal private providers. This cost-effectiveness analysis compares intervention options., Methods: A decision tree was constructed in a spreadsheet to estimate the incremental cost-effectiveness ratios (ICERs) among four strategies: no intervention, simple subsidy, subsidy with financial incentives, and subsidy with IEC. Model inputs included programmatic costs (in dollars), malaria epidemiology and observed study outcomes. Data sources included expenditure records, study data and scientific literature. Model outcomes included the proportion of properly and improperly treated individuals with and without P. falciparum malaria, and associated disability-adjusted life years (DALYs). Results are reported as ICERs in US dollars per DALY averted. One-way sensitivity analysis assessed how outcomes depend on uncertainty in inputs., Results: ICERs from the least to most expensive intervention are: $1,169/DALY averted for simple subsidy vs no intervention, $185/DALY averted for subsidy with financial incentives vs simple subsidy, and $200/DALY averted for a subsidy with IEC vs subsidy with financial incentives. Due to decreasing ICERs, each strategy was also compared to no intervention. The subsidy with IEC was the most favourable, costing $639/DALY averted compared with no intervention. One-way sensitivity analysis shows that ICERs are most affected by programme costs, RDT uptake, treatment-seeking behaviour, and the prevalence and virulence of non-malarial fevers. In conclusion, private provider subsidies with IEC or a combination of IEC and financial incentives may be a good investment for malaria control.

As large numbers of metacercariae of Fasciola hepatica are necessary for research, experimental infections of Galba truncatula and Pseudosuccinea columella with this digenean were carried out to determine the better intermediate host for metacercarial production and, consequently, the most profitable snail for decreasing the cost price of these larvae. Pre-adult snails (4 mm in shell height) originating from two populations per lymnaeid species were individually exposed to two or five miracidia, raised at 23 °C and followed for cercarial shedding up to their death. Compared to values noted in G. truncatula, the survival of P. columella on day 30 post-exposure was significantly greater, while the prevalence of F. hepatica infection was significantly lower. In the four P. columella groups, metacercarial production was significantly greater than that noted in the four groups of G. truncatula (347-453 per cercariae-shedding snail versus 163-275, respectively). Apart from one population of G. truncatula, the use of five miracidia per snail at exposure significantly increased the prevalence of F. hepatica in P. columella and the other population of G. truncatula, whereas it did not have any clear effect on the mean number of metacercariae. The use of P. columella for experimental infections with F. hepatica resulted in significantly higher metacercarial production than that noted with G. truncatula, in spite of a lower prevalence for the former lymnaeid. This finding allows for a significant decrease in the cost price of these larvae for commercial production., (© P. Vignoles et al., published by EDP Sciences, 2015.)

Echinococcus multilocularis is one of the most pathogenic zoonotic parasites in the temperate and arctic region of the Northern Hemisphere. For estimating the potential risk of human infection in endemic areas, reliable antemortem methods are needed to detect the parasite in carnivore definitive hosts. The sensitivity of routine flotation techniques for detection of E. multilocularis eggs was found to be low (3-33%) depending on the flotation solution used (specific gravities = 1.3-1.4). An improved faecal flotation followed by a species-specific PCR is described with a sensitivity of 74% (95% CI = 62-84%) and a specificity of 100% (95% CI = 94-100%). These parameters are similar to those of the intestinal scraping technique (sensitivity = 78%, specificity = 100%). The sensitivity of the improved flotation was significantly higher (P < 0.0001) than that of routine flotation techniques. The costs of the method are similar or lower than those of other antemortem diagnostic methods. Based on these data, the method is suitable for surveys of domesticated and wild carnivores.

Objectives: This study evaluates the diagnostic accuracy and cost-effectiveness of the Kato-Katz and Mini-FLOTAC methods for detection of soil-transmitted helminths (STH) in a post-treatment setting in western Kenya. A cost analysis also explores the cost implications of collecting samples during school surveys when compared to household surveys., Methods: Stool samples were collected from children (n = 652) attending 18 schools in Bungoma County and diagnosed by the Kato-Katz and Mini-FLOTAC coprological methods. Sensitivity and additional diagnostic performance measures were analyzed using Bayesian latent class modeling. Financial and economic costs were calculated for all survey and diagnostic activities, and cost per child tested, cost per case detected and cost per STH infection correctly classified were estimated. A sensitivity analysis was conducted to assess the impact of various survey parameters on cost estimates., Results: Both diagnostic methods exhibited comparable sensitivity for detection of any STH species over single and consecutive day sampling: 52.0% for single day Kato-Katz; 49.1% for single-day Mini-FLOTAC; 76.9% for consecutive day Kato-Katz; and 74.1% for consecutive day Mini-FLOTAC. Diagnostic performance did not differ significantly between methods for the different STH species. Use of Kato-Katz with school-based sampling was the lowest cost scenario for cost per child tested ($10.14) and cost per case correctly classified ($12.84). Cost per case detected was lowest for Kato-Katz used in community-based sampling ($128.24). Sensitivity analysis revealed the cost of case detection for any STH decreased non-linearly as prevalence rates increased and was influenced by the number of samples collected., Conclusions: The Kato-Katz method was comparable in diagnostic sensitivity to the Mini-FLOTAC method, but afforded greater cost-effectiveness. Future work is required to evaluate the cost-effectiveness of STH surveillance in different settings.

Background: Soil-transmitted helminths and intestinal protozoa infection are widespread in developing countries, yet an accurate diagnosis is rarely performed. The aim of this study was to evaluate the recently developed mini-FLOTAC method and to compare with currently more widely used techniques for the diagnosis of intestinal parasitic infections in different settings., Methodology/principal Findings: The study was carried out in Dharamsala, Himachal Pradesh, India, and in Bukumbi, Tanzania. A total of 180 pupils from two primary schools had their stool analyzed (n = 80 in Dharamsala and n = 100 in Bukumbi) for intestinal parasitic infections with three diagnostic methods: direct fecal smear, formol-ether concentration method (FECM) and mini-FLOTAC. Overall, 72% of the pupils were positive for any intestinal parasitic infection, 24% carried dual infections and 11% three infections or more. The most frequently encountered intestinal parasites were Entamoeba coli, Entamoeba histolytica/dispar, Giardia intestinalis, hookworm, (and Schistosoma mansoni, in Tanzania). Statistically significant differences were found in the detection of parasitic infections among the three methods: mini-FLOTAC was the most sensitive method for helminth infections (90% mini-FLOTAC, 60% FECM, and 30% direct fecal smear), whereas FECM was most sensitive for intestinal protozoa infections (88% FECM, 70% direct fecal smear, and 68% mini-FLOTAC)., Conclusion/significance: We present the first experiences with the mini-FLOTAC for the diagnosis of intestinal helminths and protozoa. Our results suggest that it is a valid, sensitive and potentially low-cost alternative technique that could be used in resource-limited settings--particularly for helminth diagnosis.

With the Global Program for the Elimination of Lymphatic Filariasis continuing to make strides towards disease eradication, many locations endemic for the causative parasites of lymphatic filariasis are realizing a substantial decrease in levels of infection and rates of disease transmission. However, with measures of disease continuing to decline, the need for time-saving and economical molecular diagnostic assays capable of detecting low levels of parasite presence is increasing. This need is greatest in locations co-endemic for both Wuchereria bancrofti and Brugia parasites because testing for both causative agents individually results in significant increases in labor and reagent costs. Here we describe a multiplex, TaqMan-based, real-time PCR assay capable of simultaneously detecting W. bancrofti and Brugia malayi DNA extracted from human bloodspots or vector mosquito pools. With comparable sensitivity to established singleplex assays, this assay provides significant cost and labor savings for disease monitoring efforts in co-endemic locations., (Copyright © 2013 Elsevier B.V. All rights reserved.)

Background: The most severe clinical form of neurocysticercosis (NC) occurs when cysticerci are located in the subarachnoid space at the base of the brain (SaB). The diagnosis, monitoring and treatment of NC-SaB, constitutes a severe clinical challenge. Herein we evaluate the potential of the HP10 antigen detection enzyme-linked immunosorbent assay (HP10 Ag-ELISA) in the long term follow-up of NC-SaB cases. Assay performance was compared with that of Magnetic Resonance Imaging (MRI). In addition, the robustness of the HP10 Ag-ELISA was evaluated independently at two different institutions., Methodology/principal Findings: A double-blind prospective cohort trial was conducted involving 38 NC-SaB cases and a total of 108 paired serum and cerebrospinal fluid (CSF) samples taken at intervals of 4 to 8 months for up to 43 months. At each medical visit, results of sera and CSF HP10 Ag-ELISA and MRI obtained at last visit were compared and their accuracy was evaluated retrospectively, considering radiological evolution between appointments. In the long-term follow-up study, HP10 Ag-ELISA had a better agreement than MRI with retrospective radiological evaluation. High reproducibility of HP10 Ag-ELISA between laboratories was also demonstrated., Conclusions: Results reported in this study establish for the first time the usefulness of the comparatively low cost HP10 Ag-ELISA for long term follow-up of NC-SaB patients.

The primers traditionally used to detect Plasmodium ovale infections are known for not binding all P. ovale parasites within the small-subunit rRNA gene when used alone. We describe a simple, cost- and time-efficient multiplex nested PCR and a nested PCR using a novel set of primers for the simultaneous detection of P. ovale curtisi and P. ovale wallikeri.

Background: Rapid diagnostic tests (RDT) for malaria have been demonstrated to be effective and they should replace microscopy in certain areas., Method: The cost-effectiveness of five RDT and thick smear microscopy was estimated and compared. Data were collected on Brazilian Extra-Amazon Region. Data sources included the National Malaria Control Programme of the Ministry of Health, the National Healthcare System reimbursement table, laboratory suppliers and scientific literature. The perspective was that of the Brazilian public health system, the analytical horizon was from the start of fever until the diagnostic results provided to patient and the temporal reference was that of year 2010. Two costing methods were produced, based on exclusive-use microscopy or shared-use microscopy. The results were expressed in costs per adequately diagnosed cases in 2010 U.S. dollars. One-way sensitivity analysis was performed considering key model parameters., Results: In the cost-effectiveness analysis with exclusive-use microscopy, the RDT CareStart™ was the most cost-effective diagnostic strategy. Microscopy was the most expensive and most effective, with an additional case adequately diagnosed by microscopy costing US$ 35,550.00 in relation to CareStart™. In opposite, in the cost-effectiveness analysis with shared-use microscopy, the thick smear was extremely cost-effective. Introducing into the analytic model with shared-use microscopy a probability for individual access to the diagnosis, assuming a probability of 100% of access for a public health system user to any RDT and, hypothetically, of 85% of access to microscopy, this test saw its effectiveness reduced and was dominated by the RDT CareStart™., Conclusion: The analysis of cost-effectiveness of malaria diagnosis technologies in the Brazilian Extra-Amazon Region depends on the exclusive or shared use of the microscopy. Following the assumptions of this study, shared-use microscopy would be the most cost-effective strategy of the six technologies evaluated. However, if used exclusively for diagnosing malaria, microscopy would be the worst use of resources. Microscopy would not be the most cost-effective strategy, even when structure is shared with other programmes, when the probability of a patient having access to it was reduced. Under these circumstances, the RDT CareStart™ would be the most cost-effective strategy.

The quantitative buffy coat (QBC) technique is a method of diagnosing malarial parasites based on micro-centrifugation, fluorescence, and density gradient of infected red blood cells. The aim of the present study was to modify the QBC technique in order to reduce the cost per test of malaria diagnosis. This was achieved by introducing some modifications to routine QBC wherein REMI centrifuge (cost Rs 19000/-) and ultra-violet microscope (Rs 115000) were used instead of parafuge (Rs 108000) and paralens (Rs 293625/-). With the above modification, the cost per test for laboratories dealing with high patient load was reduced by 13%, whereas for smaller laboratories with low patient load, the cost per test was reduced by 48%. This is a significant difference in cost. The results of the modified QBC method were compared with the current diagnostic methods: peripheral blood smear (PBS) and routine QBC. Blood samples collected from 96 patients were subjected to the above tests. Considering PBS as the gold standard, routine QBC showed 91% sensitivity and 96% specificity for Plasmodium vivax- and 91% sensitivity and 94% specificity for Plasmodium falciparum-infected patients. It was seen that the modified QBC technique had 91% sensitivity and 98% specificity for P. vivax and 91% sensitivity and 96% specificity for P. falciparum. It was concluded that modification of the QBC technique renders it cheaper without compromising the specificity and sensitivity of the method.

Diagnosis of Acanthamoeba by microscopic examination, culture, and polymerase chain reactions (PCRs) has several limitations (sensitivity, specificity, lack of detection of several strains, cost of testing for discrimination among strains). We developed a new high-resolution melting real-time PCR (HRM) to detect and characterize Acanthamoeba infections. HRM performances were evaluated with strains from the American Type Culture Collection (ATCC) and with 20 corneal scrapings. The DNA extracted from specimens were amplified, detected, and characterized in 1 run using 2 original primers diluted in a solution containing an intercalating dye. Detection and molecular characterization of Acanthamoeba infections could be achieved in less than 2.5 h with a dramatic reduction in cost of reactants (postamplification procedures and radioactive or fluorescent-labeled molecular probes were unnecessary). HRM detection limits were 0.1 cyst/μL or less (including genotypes T5 and T11), and its sensitivity and specificity were higher than other molecular tests. For the tested strains from the ATCC, the HRM drafted 4 different profiles: Type I (genotypes T2 and T4), Type II (T5 and T7), Type III (T8), and Type IV (T1, T3, T6, T9, T11, T12, and T13)., (Copyright © 2012 Elsevier Inc. All rights reserved.)

Background: Accurate sampling of sub-microscopic gametocytes is necessary for epidemiological studies to identify the infectious reservoir of Plasmodium falciparum. Detection of gametocyte mRNA achieves sensitive detection, but requires careful handling of samples. Filter papers can be used for collecting RNA samples, but rigorous testing of their capacity to withstand adverse storage conditions has not been fully explored., Methods: Three gametocyte dilutions: 10/μL, 1.0/μL and 0.1/μL were spotted onto Whatman™ 903 Protein Saver Cards, FTA Classic Cards and 3MM filter papers that were stored under frozen, cold chain or tropical conditions for up to 13 weeks . RNA was extracted, then detected by quantitative nucleic acid sequence-based amplification (QT-NASBA) and reverse-transcriptase PCR (RT-PCR)., Results: Successful gametocyte detection was more frequently observed from the Whatman 903 Protein Saver Card compared to the Whatman FTA Classic Card, by both techniques (p<0.0001). When papers were stored at higher temperatures, a loss in sensitivity was experienced for the FTA Classic Card but not the 903 Protein Saver Card or Whatman 3MM filter paper. The sensitivity of gametocyte detection was decreased when papers were stored at high humidity., Conclusions: This study indicates the Whatman 903 Protein Saver Card is better for Pfs25 mRNA sampling compared to the Whatman FTA Classic Card, and that the Whatman 3MM filter paper may prove to be a satisfactory cheaper option for Pfs25 mRNA sampling. When appropriately dried, filter papers provide a useful approach to Pfs25 mRNA sampling, especially in settings where storage in RNA-protecting buffer is not possible.

We have evaluated a new simple technique using whole blood from experimentally infected cattle for the isolation and cultivation of Theileria annulata. The study was carried out on 20 Holstein-Frisian bovines that had been experimentally infected with a virulent lethal dose of Theileria annulata. This technique has been compared to the classical peripheral blood monocyte isolation with Ficoll carried out on 22 experimentally infected Holstein-Friesian calves. The effectiveness of the reference technique was estimated to 86.4%, whilst the effectiveness of the new technique was 100%. Moreover, this new technique leads to time and money saving estimated to € 3.06 per sample. It decreases the contamination risks by reducing the steps of sample manipulation.

Cryptosporidiosis caused by Cryptosporidium spp. is an important diarrhoeal disease observed in farm animals and humans, especially in young or immunocompromised individuals. A novel cell culture assay for testing extracts and pure compounds against Cryptosporidium parvum in 96-well microplate format was established and evaluated. It is based on previously described indirect fluorescent antibody techniques and was optimised for higher sample throughput. Rapid assessment of minimal inhibitory concentrations (MICs) was done by checking each well microscopically for the presence or absence of parasite stages. As a novelty, parasite development was quantified by enumeration of clusters of secondary infection (CSI), which typically appeared upon infection with a distinct parasite inoculum after a defined incubation time. Host cell (HCT-8) viability was measured by an integrated non-destructive water-soluble tetrazolium salt assay (WST-1), which facilitated discrimination of antiparasitic activity from possible cytotoxic effects of a test compound against the host cells. Host cell viability was regarded unimpaired when cultures had 75% or more viability when compared to control cultures without test substance. In this study, a maximum density of distinguishable CSI was obtained when cultures were infected with 2.5 × 10(3) oocysts and incubated for 48 h. The applicable inoculum has to be optimised for each batch of oocysts and before each experimental series. Parasite development was inhibited completely by monensin at 134 nM and silymarin at 50 mg/mL. These concentrations were non-toxic to the host cells and comparable to literature data. The percentages of parasite inhibition were determined for monensin and a 50% inhibitory concentration (IC(50)) of 36.6 nM (27.4-45.5) and a 90% inhibitory concentration of 65.9 nM (54.8-90.2) were calculated. The introduced assay is economic because relatively low parasite numbers may be used. If MICs are determined, evaluation is fast, as each well is viewed only briefly under the fluorescence microscope for presence or absence of CSI. Furthermore it is highly critical because only full parasite inhibition is assessed. Counting of CSI is more laborious and time-consuming, but it allows calculation of parasite inhibition rates and parameters like the half maximal inhibitory concentration (IC(50)). This assay shall be used to assess anticryptosporidial activities of various plant waste materials and by-products from the food and the pharmaceutical industries in the course of the EU project SAFEWASTES. Comparison with in vivo models should be performed to further corroborate the results. Automated evaluation by flow cytometry might facilitate higher sample throughput and reduce operator bias.

Human helminthiases are of considerable public health importance in sub-Saharan Africa, Asia, and Latin America. The acknowledgement of the disease burden due to helminth infections, the availability of donated or affordable drugs that are mostly safe and moderately efficacious, and the implementation of viable mass drug administration (MDA) interventions have prompted the establishment of various large-scale control and elimination programmes. These programmes have benefited from improved epidemiological mapping of the infections, better understanding of the scope and limitations of currently available diagnostics and of the relationship between infection and morbidity, feasibility of community-directed or school-based interventions, and advances in the design of monitoring and evaluation (M&E) protocols. Considerable success has been achieved in reducing morbidity or suppressing transmission in a number of settings, whilst challenges remain in many others. Some of the obstacles include the lack of diagnostic tools appropriate to the changing requirements of ongoing interventions and elimination settings; the reliance on a handful of drugs about which not enough is known regarding modes of action, modes of resistance, and optimal dosage singly or in combination; the difficulties in sustaining adequate coverage and compliance in prolonged and/or integrated programmes; an incomplete understanding of the social, behavioural, and environmental determinants of infection; and last, but not least, very little investment in research and development (R&D). The Disease Reference Group on Helminth Infections (DRG4), established in 2009 by the Special Programme for Research and Training in Tropical Diseases (TDR), was given the mandate to undertake a comprehensive review of recent advances in helminthiases research, identify research gaps, and rank priorities for an R&D agenda for the control and elimination of these infections. This review presents the processes undertaken to identify and rank ten top research priorities; discusses the implications of realising these priorities in terms of their potential for improving global health and achieving the Millennium Development Goals (MDGs); outlines salient research funding needs; and introduces the series of reviews that follow in this PLoS Neglected Tropical Diseases collection, "A Research Agenda for Helminth Diseases of Humans."

Background: Malaria is a serious infectious disease. According to the World Health Organization, it is responsible for nearly one million deaths each year. There are various techniques to diagnose malaria of which manual microscopy is considered to be the gold standard. However due to the number of steps required in manual assessment, this diagnostic method is time consuming (leading to late diagnosis) and prone to human error (leading to erroneous diagnosis), even in experienced hands. The focus of this study is to develop a robust, unsupervised and sensitive malaria screening technique with low material cost and one that has an advantage over other techniques in that it minimizes human reliance and is, therefore, more consistent in applying diagnostic criteria., Method: A method based on digital image processing of Giemsa-stained thin smear image is developed to facilitate the diagnostic process. The diagnosis procedure is divided into two parts; enumeration and identification. The image-based method presented here is designed to automate the process of enumeration and identification; with the main advantage being its ability to carry out the diagnosis in an unsupervised manner and yet have high sensitivity and thus reducing cases of false negatives., Results: The image based method is tested over more than 500 images from two independent laboratories. The aim is to distinguish between positive and negative cases of malaria using thin smear blood slide images. Due to the unsupervised nature of method it requires minimal human intervention thus speeding up the whole process of diagnosis. Overall sensitivity to capture cases of malaria is 100% and specificity ranges from 50-88% for all species of malaria parasites., Conclusion: Image based screening method will speed up the whole process of diagnosis and is more advantageous over laboratory procedures that are prone to errors and where pathological expertise is minimal. Further this method provides a consistent and robust way of generating the parasite clearance curves.

Objective: To determine a cost-minimizing option for congenital toxoplasmosis in the United States., Methodology/principal Findings: A decision-analytic and cost-minimization model was constructed to compare monthly maternal serological screening, prenatal treatment, and post-natal follow-up and treatment according to the current French (Paris) protocol, versus no systematic screening or perinatal treatment. Costs are based on published estimates of lifetime societal costs of developmental disabilities and current diagnostic and treatment costs. Probabilities are based on published results and clinical practice in the United States and France. One- and two-way sensitivity analyses are used to evaluate robustness of results. Universal monthly maternal screening for congenital toxoplasmosis with follow-up and treatment, following the French protocol, is found to be cost-saving, with savings of $620 per child screened. Results are robust to changes in test costs, value of statistical life, seroprevalence in women of childbearing age, fetal loss due to amniocentesis, and to bivariate analysis of test costs and incidence of primary T. gondii infection in pregnancy. Given the parameters in this model and a maternal screening test cost of $12, screening is cost-saving for rates of congenital infection above 1 per 10,000 live births. If universal testing generates economies of scale in diagnostic tools-lowering test costs to about $2 per test-universal screening is cost-saving at rates of congenital infection well below the lowest reported rates in the United States of 1 per 10,000 live births., Conclusion/significance: Universal screening according to the French protocol is cost saving for the US population within broad parameters for costs and probabilities.

Background: Rapid and accurate diagnosis of malaria is central to clinical management and the prevention of drug-overuse, which may lead to resistance development, toxicity and economic losses. So far, light microscopy (LM) of Giemsa-stained thick blood smears is the gold standard. Under optimal conditions the procedure is fast and reliable; nevertheless a gain in speed would be a great advantage. Rapid diagnosis tests are an alternative, although they cost more and give qualitative instead of quantitative results. Light-emitting diode (LED) fluorescence microscopy (ledFM 400 ×, 1000 ×) may offer a reliable and cheap alternative, which can be used at the point of care., Methods: LedFM and conventional fluorescence microscopy (uvFM) were compared to LM in 210 samples from patients with history of fever in the last 24 hours admitted to the Albert Schweitzer Hospital in Lambaréné, Gabon., Results: Sensitivities were 99.1% for ledFM and 97.0% for uvFM, specificities 90.7% for ledFM 400 × and 92.6% for ledFM 1000 × and uvFM. High agreement was found in Bland-Altman-plot and Kappa coefficient (ledFM 1000 ×: 0.914, ledFM 400 × and uvFM: 0.895). The time to diagnosis for both FM methods was shorter compared to LM (LM: 43 min, uvFM: 16 min, ledFM 1000 ×: 14 min, ledFM 400 ×: 10 min)., Conclusion: ledFM is a reliable, accurate, fast and inexpensive tool for daily routine malaria diagnosis and may be used as a point of care diagnostic tool., (© 2011 Lenz et al; licensee BioMed Central Ltd.)

Background: Loop-mediated isothermal amplification (LAMP) is a high performance method for detecting DNA and holds promise for use in the molecular detection of infectious pathogens, including Plasmodium spp. However, in most malaria-endemic areas, which are often resource-limited, current LAMP methods are not feasible for diagnosis due to difficulties in accurately interpreting results with problems of sensitive visualization of amplified products, and the risk of contamination resulting from the high quantity of amplified DNA produced. In this study, we establish a novel visualized LAMP method in a closed-tube system, and validate it for the diagnosis of malaria under simulated field conditions., Methods: A visualized LAMP method was established by the addition of a microcrystalline wax-dye capsule containing the highly sensitive DNA fluorescence dye SYBR Green I to a normal LAMP reaction prior to the initiation of the reaction. A total of 89 blood samples were collected on filter paper and processed using a simple boiling method for DNA extraction, and then tested by the visualized LAMP method for Plasmodium vivax infection., Results: The wax capsule remained intact during isothermal amplification, and released the DNA dye to the reaction mixture only when the temperature was raised to the melting point following amplification. Soon after cooling down, the solidified wax sealed the reaction mix at the bottom of the tube, thus minimizing the risk of aerosol contamination. Compared to microscopy, the sensitivity and specificity of LAMP were 98.3% (95% confidence interval (CI): 91.1-99.7%) and 100% (95% CI: 88.3-100%), and were in close agreement with a nested polymerase chain reaction method., Conclusions: This novel, cheap and quick visualized LAMP method is feasible for malaria diagnosis in resource-limited field settings.

Migration is a channel through which Chagas disease is imported, and vertical transmission is a channel through which the disease is spread in non-endemic countries. This study presents the economic evaluation of Chagas disease screening in pregnant women from Latin America and in their newborns in a non endemic area such as Spain. The economic impact of Chagas disease screening is tested through two decision models, one for the newborn and one for the mother, against the alternative hypothesis of no screening for either the newborn or the mother. Results show that the option "no test" is dominated by the option "test". The cost effectiveness ratio in the "newborn model" was 22€/QALYs gained in the case of screening and 125€/QALYs gained in the case of no screening. The cost effectiveness ratio in the "mother model" was 96€/QALYs gained in the case of screening and 1675€/QALYs gained in the case of no screening. Probabilistic sensitivity analysis highlighted the reduction of uncertainty in the screening option. Threshold analysis assessed that even with a drop in Chagas prevalence from 3.4% to 0.9%, a drop in the probability of vertical transmission from 7.3% to 2.24% and with an increase of screening costs up to €37.5, "test" option would still be preferred to "no test". The current study proved Chagas screening of all Latin American women giving birth in Spain and of their infants to be the best strategy compared to the non-screening option and provides useful information for health policy makers in their decision making process., (Copyright © 2011 Elsevier B.V. All rights reserved.)

Parasitic gastrointestinal infections are a major cause of morbidity and mortality in the developing world, with stool microscopy being the mainstay of diagnostic practice. Both direct microscopy and concentration techniques can be utilized; direct microscopy may be time consuming and tedious; however clinical laboratories in developing countries lack trained staff who can effectively use concentration methods. In our practice we used the Parasep O and P filter concentrator tubes (manufactured by DiaSys Ltd, Berkshire, England. Product Code 146000) along with direct microscopic techniques and found that Parasep filters enhanced the ability to detect intestinal parasites that would have been missed on routine microscopy. We found the Parasep filter concentration method to be easy, cost-effective and reliable for routine stool examinations.

Background: In areas with limited structure in place for microscopy diagnosis, rapid diagnostic tests (RDT) have been demonstrated to be effective., Method: The cost-effectiveness of the Optimal® and thick smear microscopy was estimated and compared. Data were collected on remote areas of 12 municipalities in the Brazilian Amazon. Data sources included the National Malaria Control Programme of the Ministry of Health, the National Healthcare System reimbursement table, hospitalization records, primary data collected from the municipalities, and scientific literature. The perspective was that of the Brazilian public health system, the analytical horizon was from the start of fever until the diagnostic results provided to patient and the temporal reference was that of year 2006. The results were expressed in costs per adequately diagnosed cases in 2006 U.S. dollars. Sensitivity analysis was performed considering key model parameters., Results: In the case base scenario, considering 92% and 95% sensitivity for thick smear microscopy to Plasmodium falciparum and Plasmodium vivax, respectively, and 100% specificity for both species, thick smear microscopy is more costly and more effective, with an incremental cost estimated at US$549.9 per adequately diagnosed case. In sensitivity analysis, when sensitivity and specificity of microscopy for P. vivax were 0.90 and 0.98, respectively, and when its sensitivity for P. falciparum was 0.83, the RDT was more cost-effective than microscopy., Conclusion: Microscopy is more cost-effective than OptiMal® in these remote areas if high accuracy of microscopy is maintained in the field. Decision regarding use of rapid tests for diagnosis of malaria in these areas depends on current microscopy accuracy in the field.

Background: Increasing emphasis on integrated control of neglected tropical diseases (NTDs) requires identification of co-endemic areas. Integrated surveys for lymphatic filariasis (LF), schistosomiasis and soil-transmitted helminth (STH) infection have been recommended for this purpose. Integrated survey designs inevitably involve balancing the costs of surveys against accuracy of classifying areas for treatment, so-called implementation units (IUs). This requires an understanding of the main cost drivers and of how operating procedures may affect both cost and accuracy of surveys. Here we report a detailed cost analysis of the first round of integrated NTD surveys in Southern Sudan., Methods and Findings: Financial and economic costs were estimated from financial expenditure records and interviews with survey staff using an ingredients approach. The main outcome was cost per IU surveyed. Uncertain variables were subjected to univariate sensitivity analysis and the effects of modifying standard operating procedures were explored. The average economic cost per IU surveyed was USD 40,206 or USD 9,573, depending on the size of the IU. The major cost drivers were two key categories of recurrent costs: i) survey consumables, and ii) personnel., Conclusion: The cost of integrated surveys in Southern Sudan could be reduced by surveying larger administrative areas for LF. If this approach was taken, the estimated economic cost of completing LF, schistosomiasis and STH mapping in Southern Sudan would amount to USD 1.6 million. The methodological detail and costing template provided here could be used to generate cost estimates in other settings and readily compare these to the present study, and may help budget for integrated and single NTDs surveys elsewhere.

This study evaluates Cryptosporidium parvum oocyst production in dexamethasone suppressed CF-1 and C57BL/6 mice. Both models can yield 1 x 10(9) total oocysts over a 20-day production period; however, only 20 CF-1 mice are required to reliably achieve this goal compared to 40 C57BL/6 mice. Although oocyst yields per mouse are similar for both mouse strains, the survival rate for CF-1 mice is higher, resulting in reduced lost production time per study when compared to the C57BL/6 mice. This study presents a more efficient and cost effective dexamethasone suppressed murine model of propagating high concentrations of C. parvum oocysts.

The applications of highly specific and sensitive molecular techniques based on polymerase chain reaction (PCR) have constituted a valuable tool for the diagnosis of schistosomiasis and also for the detection of schistosome infections in the snail intermediate hosts. The common method of detecting PCR amplicons is gel electrophoresis in the presence of ethidium bromide, a carcinogen, which is followed by UV transillumination. Other methods, which are available for detecting PCR products, are real-time PCR, PCR-enzyme-linked immunosorbent assay (PCR-ELIZA) and mass spectrometry but they are cumbersome while they are sometimes complex and expensive. Therefore, a simple method of PCR product detection would be a welcome idea and a most valuable tool particularly in disease endemic countries with limited research facilities and resources. In this study, we applied a simple and rapid method for the detection of Schistosoma haematobium and Schistosoma mansoni PCR amplified DNA products using oligochromatographic (OC) dipstick. The amplicons are visualized by hybridization with a gold conjugated probe, while a control for the chromatographic migration is incorporated in the assay. The lower detection limit observed was 10fg of genomic DNA from each of the two species, while the dipstick was also specific for each of the species used in this study.

Host-parasitoid models including integrated pest management (IPM) interventions with impulsive effects at both fixed and unfixed times were analyzed with regard to host-eradication, host-parasitoid persistence and host-outbreak solutions. The host-eradication periodic solution with fixed moments is globally stable if the host's intrinsic growth rate is less than the summation of the mean host-killing rate and the mean parasitization rate during the impulsive period. Solutions for all three categories can coexist, with switch-like transitions among their attractors showing that varying dosages and frequencies of insecticide applications and the numbers of parasitoids released are crucial. Periodic solutions also exist for models with unfixed moments for which the maximum amplitude of the host is less than the economic threshold. The dosages and frequencies of IPM interventions for these solutions are much reduced in comparison with the pest-eradication periodic solution. Our results, which are robust to inclusion of stochastic effects and with a wide range of parameter values, confirm that IPM is more effective than any single control tactic.

Objective: Rapid diagnostic tests for malaria seem cost effective in standard analyses, but these do not take account of clinicians' response to test results. This study tested the impact of clinicians' response to rapid diagnostic test or microscopy results on the costs and benefits of testing at different levels of malaria transmission and in different age groups., Design: Cost-benefit analysis using a decision tree model and clinical data on the effectiveness of diagnostic tests for malaria, their costs, and clinicians' response to test results., Setting: Tanzania., Methods: Data were obtained from a clinical trial of 2425 patients carried out in three settings of varying transmission., Results: At moderate and low levels of malaria transmission, rapid diagnostic tests were more cost beneficial than microscopy, and both more so than presumptive treatment, but only where response was consistent with test results. At the levels of prescription of antimalarial drugs to patients with negative tests that have been found in observational studies and trials, neither test methodis likely to be cost beneficial, incurring costs 10-250% higher, depending on transmission rate, than would have been the case with fully consistent responses to all test results. Microscopy becomes more cost beneficial than rapid diagnostic tests when its sensitivity under operational conditions approaches that of rapid diagnostic tests., Conclusions: Improving diagnostic methods, including rapid diagnostic tests, can reduce costs and enhance the benefits of effective antimalarial drugs, but only if the consistency of response to test results is also improved. Investing in methods to improve rational response to tests is essential. Economic evaluations of diagnostic tests should take into account whether clinicians' response is consistent with test results.

A survey was conducted in a low-endemic and in a non-endemic area of Sudan to evaluate the specificity and efficiency of different serological antibody detection techniques for Trypanosoma brucei gambiense. Comparisons were made of the card agglutination test for trypanosomiasis (CATT) on diluted blood, on diluted plasma and on eluates from blood dried on filter paper, the LATEX test on diluted plasma and an ELISA on diluted plasma and filter paper. The specificities of all the serological tests were not significantly different from CATT on diluted blood (99.5%). The specificity of CATT on diluted blood was similar (99.3%). The highest sensitivities (100%) were observed with CATT on diluted blood and with CATT and LATEX on diluted plasma. CATT on diluted blood was more cost-efficient than the classic test, CATT on whole blood.

Cryptosporidium hominis and Cryptosporidium parvum are associated with massive disease outbreaks worldwide. Because these two species have different transmission cycles, identification of these parasites to the species level in clinical samples may provide laboratory data of crucial importance in epidemiologic investigations. To date, the most reliable way to differentiate C. hominis and C. parvum is based on DNA sequencing analysis of PCR amplicons. Although this approach is very effective for differentiation of Cryptosporidium species, it is labor-intensive and time-consuming compared with methods that do not require DNA sequencing analysis as an additional step and that have been successfully used for specific identification of a number of pathogens. In this study, we describe a novel Luminex-based assay that can differentiate C. hominis from C. parvum in a rapid and cost-effective manner. The assay was validated by testing a total of 143 DNA samples extracted from clinical specimens, environmental samples, or samples artificially spiked with Cryptosporidium oocysts. As few as 10 oocysts per 300 microl of stools could be detected with this assay. The assay format includes species-specific probes linked to carboxylated Luminex microspheres that hybridize to a Cryptosporidium microsatellite-2 region (ML-2) where C. hominis and C. parvum differ by one nucleotide substitution. The assay proved to be 100% specific when samples that had been characterized by direct fluorescent antibody test (DFA) and DNA sequencing analysis were tested. In addition, the assay was more sensitive than DFA and provided species identification, which is an advantage for epidemiologic studies.

The development of new diagnostic tools particularly for some parasitic "neglected diseases", is slowed or even hindered by limited resources assigned for basic and applied research in public institution and private sector. Even if the time-line and costs needed for developing a new In Vitro Diagnostic (IVD) test are generally lower compared to vaccines or new drugs, industry is poorly engaged in investing resources due to the perception of limited markets. To accelerate the development of diagnostics for the world's most deadly diseases, the World Health Organization's (WHO) Special Programme for Research and Training in Tropical Diseases (TDR), the United Nations Development Programme, the World Bank and the Gates Foundation, last year launched a new initiative, FIND (Foundation for Innovative New Diagnostics, www.finddiagnostics.org). The aim is to "apply the latest biotechnology innovations to develop and validate affordable diagnostic tests for diseases of the developing world". Ideally, a new diagnostic test should be accurately evaluated prior to use in medical practice. The first step would be a pre-clinical evaluation, an analytic study to determine its laboratory performance. A crucial point in this phase is the calibration of reagents (antigens, antibodies, DNA probes, etc.) against a standard reference preparation. WHO, through the WHO International Laboratories for Biological Standards, "provides International Biological Reference Preparations which serve as reference sources of defined biological activity expressed in an internationally agreed unit" (www.who.int/biologicals/IBRP/index.htm). Standardization allows "comparison of biological measurements worldwide" and ensures the reliability of diagnostic procedures. These preparations are generally intended for use in the characterization of the activity of secondary reference preparations (regional, national or in-house working standards). Unfortunately, international reference standards for parasitic diseases are not available at present, except for Toxoplasma antibodies. The first international standard reagent for Anti-Toxoplasma Serum was established in 1968 and at present, an international standard reference serum, Anti-toxoplasma serum, human TOXM is available at the National Institute for Biological Standards and Control (NIBSC) in UK. Several collaborative, multicenter studies were carried out to assess the performance of different methods and commercial tests for the diagnosis of toxoplasmosis, by providing to participating laboratories a panel of well-defined sera to be tested. A four-phase process following well-accepted methodological standards for the development of diagnostics, analogous to those internationally accepted for drugs and vaccines was recently proposed. The pre-clinical evaluation, the analytic study to assess sensitivity, specificity, predictive values in laboratory (phase I), should be followed by a proof of principle study to distinguish diseased from healthy persons in easily accessible populations (phase II). The evaluation of test performance in populations of intended use (phase III), and finally the delineation of cost-effectiveness and societal impact of new tests in comparison with existing tools (phase IV) should complete the validation procedure. In this context, national regulatory agencies play a major role in pre-market approval and post-market surveillance of IVDs. The European Community in 1998 approved a directive (Directive 98/79/EC) which rules the marketing of IVD medical devices, in order to harmonise the performance levels and standards in European countries. But, among IVDs for parasitic diseases, only those to detect congenital toxoplasmosis are submitted to defined procedures to provide the verification of products before their placing on the market and the surveillance after their marketing by a notified body, which perform appropriate examinations, tests and inspections to production facilities to verify if the device meets the requirements of the directive. In U.S.A., the Food and Drug Administration (FDA), through the Office of In Vitro Diagnostic Device Evaluation and Safety (OIVD), provides a comprehensive and regulatory activity for IVDs through pre-market evaluation and post-market surveillance. In developing countries, the scarcity of resources limits the procedures through which the national control authority can assure safety, quality and efficacy of products marketed, both imported and locally manufactured.

The way in which the huge Australian parasite fauna is described (discovery and naming) is the subject of this address. The approach to the task has never been well-organised so that a few groups of parasites are now relatively well-known because of the efforts of small groups of workers who have made sustained efforts in these groups, but equally some host-parasite systems have been almost completely ignored in that no worker has ever given them sustained attention. A high proportion of Australian parasites have been described by international workers. The sustaining of interest in a group of parasites over a long period is the key to real progress being made. The nature of the organisation of Australian science presently means that few positions are available for parasite taxonomists and funding for taxonomic research is scarce. Thus, parasite taxonomy (like the taxonomy of many groups of Australian plants and animals) can only be considered to be in crisis.

Introduction: To determine the cost and volume of work implied in providing a parasitological diagnosis when attending immigrants in a referral Tropical Medicine Unit., Methods: The total number and type of samples, requests processed and parasitology laboratory costs were quantified after attending 1258 immigrants during the period January 1989 to December 2001. Estimation of laboratory workload for the total was made on the basis of results from 748 of these patients. A reference to relative value units (RVU) was established for each of the tests used in the parasitological diagnosis. The evaluation included costs related to disposable material and time spent by technicians and medical staff, but did not include costs related to sample extraction or processing of the test request. Modified indicators from the Catalogue of Microbiology studies of the Valencian Community for the year 2000 were used, establishing the equivalence of 1 RVU as the value of one urine culture 5 2.39 3 (398 pesetas)., Results: The overall cost was 99,680.99 3 (16,585,522 pesetas) or 45,934.94 RVU, and the cost per patient was 79.24 3 (13.185 pesetas) or 36.51 URV. The volume of work dedicated to attending immigrants was 9.7% of the total of samples received in the laboratory., Conclusions: The cost and volume of work involved in attending this group of patients in the parasitology laboratory was not high.

The future of veterinary parasitology is discussed at a time when R&D funding from the pharmaceutical industry is declining, yet the opportunities for veterinary parasitologists to diversify their activities has never been greater. Emerging and re-emerging areas requiring input from veterinary parasitologists include: veterinary public health; conservation and wildlife diseases; emerging and exotic infectious diseases; surveillance strategies; economic effects of parasitic diseases; aquaculture; molecular epidemiology; dietary and biological control of parasitic diseases; animal welfare; organic agricultural systems; novel vaccination strategies; drug target characterisation and rational drug design. Without change, the survival of veterinary parasitology as a viable, distinct discipline is under threat. In this environment, veterinary parasitologists must be adaptable, imaginative and pro-active in terms of setting the agendas for establishing strategic alliances, promoting research needs and developing research programs.

Current evidence suggests research in veterinary parasitology is in decline despite its importance. This is particularly true in the UK where research funds have been diverted into BSE. Decline in interest in veterinary parasitology is at least in part due to the success of major pharmaceutical companies in producing a range of effective and safe anti-parasitic drugs. Research is needed because of the effects of parasites on animal welfare and the economic costs of parasites. However, there is little information on the actual costs of animal parasites. Another major reason for research is the development of drug resistance in protozoa, helminths and arthropods of veterinary importance. This is a serious problem particularly for sheep and goats in the southern hemisphere. A prioritised list of research requirements is suggested: (i) new drugs; (ii) resistance management; (iii) vaccines; (iv) breeding for resistance; (v) improved diagnostics; (vi) zoonoses; (vii) global warming and parasites. There is a major political challenge to raise the profile of veterinary parasitology and thus the funding essential for its advancement and the continued welfare and productivity of animals.