Egg viability in urinary schistosomiasis. III. Repeatability and reproducibility of new methods.

Two new field methods for the detection and quantification of egg viability/hatchability in urinary schistosomiasis due to Schistosoma haematobium have been described recently, and compared with reference methods; the new methods are cheap and simple to use, and may be used singly or together. Before recommending their widespread use in control programmes, repeatability (r) and reproducibility (R) were studied using the guide for determination prepared by the British Standards Institution. Both parameters were found to be low in relation to the mean proportions of eggs viable/hatchable except at very low egg counts, and the new methods can safely be used in PHC-based control programmes.

Egg viability in urinary schistosomiasis. II. Simplifying modifications and standardization of new methods.

Two new methods were recently introduced for the determination and quantification of egg viability in Schistosoma haematobium infections by staining eggs filtered from urine on Nytrel filters with mercurochrome and iodine. Both methods showed reasonable agreement with the best currently available method, centrifugation hatching, but it appeared that improved agreement could be obtained by modifications of timing and technique. The present paper describes the standardization of staining times and microscopic counting techniques; the modifications made gave substantially improved agreement with the reference method and achieved the most important objective, not to have zero estimates of viability when the centrifugation method detected hatching.

Egg viability in urinary schistosomiasis. I. New methods compared with available methods.

A new method of quantifying viability and hatchability of Schistosoma haematobium eggs in urine has been developed. It consists of filtering urine samples through Nytrel filters followed by mercurochrome staining to assist in the detection of viable (motile) miracidia in egg shells, and iodine staining to show hatched miracidia. The new method is compared with two existing techniques for viability/hatchability determination, centrifugation hatching and Nuclepore filtration trypan blue staining. Seventy-three egg positive urine samples are compared; the overall proportions of viable eggs are very close for centrifugation hatching and the new method, but 33% of eggs are lost in centrifugation and 19% of eggs are unclassifiable by Nuclepore filtration trypan blue staining. The main advantages and disadvantages of the new and existing methods are described and discussed; the cheapness, simplicity and ease of performance of the new method make it suitable for large-scale field use. Suggestions for improvements and standardization of the new method are made.

Repeatability and reproducibility of egg counts of Schistosoma haematobium in urine.

Egg output in urinary schistosomiasis can be influenced by several factors, such as time of collection of urine, day to day variations, seasonal variations and environmental conditions. Additionally there are unavoidable random errors inherent in every test procedure. To assess the precision of Schistosoma haematobium egg counts, carried out after paper filtration, two measures are determined, namely repeatability (r) and reproducibility (R). 10 aliquots of 10 urine samples are examined by 5 microscopists and values for r and R are calculated following the methods of the British Standards Institution (BSI, 1987). Results show unexpectedly high values for the 2 measures in relation to the overall means in all 10 urine samples, and analyses of variance confirm that there are large variations between aliquots of equal volume drawn from the same urine specimen, highly significant in all 10 urine specimens; variations between microscopists are considerably lower and only highly significant in one urine specimen. The Poisson heterogeneity test for all urine samples refutes in 9 out of 10 cases the hypothesis that aliquots of equal volume taken from a well mixed urine sample could reasonably be drawn from a Poisson distribution. It is therefore concluded that egg counts are very valuable for community studies and should always be performed, but results must be interpreted with more caution than before and great care taken in analyzing counts (especially in longitudinal studies), in classifying them and in relating them to morbidity.(ABSTRACT TRUNCATED AT 250 WORDS)

Quality control in schistosomiasis programmes.

The evaluation of a schistosomiasis control programme requires regular monitoring of the performance of technicians responsible for measuring prevalence rates and intensities of infection; simple quality control measures are the most effective form of monitoring. Quality control of microscopy has usually been carried out immediately after primary examination of fresh microscope specimens or by transport of preserved urine or stool samples to a central laboratory; the first method was impossible in Madagascar, and the second involved severe loss and breakage of specimen containers. A new method involving the transport of dried filters fixed on plastic sheets with adhesive tape was therefore used. Essential requirements of the quality control methods were simplicity, effectiveness, acceptability to field technicians, and the production of results valuable to the control programme. Acceptable limits of error (ALE) were determined by a combination of trial-and-error assessment of technicians' capabilities, and the application of a rule-of-thumb derived from quality control studies in clinical chemistry. Results were prepared in the form of simple 2 x 2 tables and easily understood charts for visual assessment of results. Technicians' work varied from excellent to poor; false negative rates (FNR) were usually higher than false positive rates (FPR) and technicians' field counts of eggs were most often lower than control counts. Control proved very useful in motivating technicians, in maintaining high work standards, and in identifying technicians needing further training.

False epidemiological results from the bulk transport of dried filter papers in urinary schistosomiasis.

In field studies in Schistosoma haematobium endemic areas, dried filters must be transported in bulk to central laboratories, either for primary screening and counting, or for quality control of field workers. We examined filters before and after bulk transport, from three populations, of high, medium and low prevalence rate. A total of 312 filters were examined before and after transport; 28.6% of all eggs on filters were lost during transport. Negative filters became positive during transport, and hence false prevalence rates resulted from after transport studies. Egg loss did not seem to be related to initial egg density. The operational results of these findings could be serious, in terms of distribution of treatment and evaluation of results and costs.

Quantitative egg counts in schistosomiasis surveys.

Methods for urine filtration and quantitative egg counts play a key role in schistosomiasis surveys, but recent WHO recommendations may be misleading. In this article, Reny Braun-Munzinger working in Madagascar discusses technical problems in the use of nytrel filters for schistosome egg counts.

Potential false positive egg-counts through the reuse of polyamide filters in the diagnosis of urinary schistosomiasis.

Nytrel monofilament polyamide filters have recently been strongly advocated as a cheap reusable method for filtering urine in field studies of Schistosoma haematobium endemic areas. We examined 1230 persons in a highly endemic area, filtering 10 ml urine through Nytrel filters and counting the eggs retained. It was found that staining Nytrel filters speeded up the counting process and made the detection of light infections much easier. After very careful washing and rinsing 12.0% of unstained filters and 4.8% of stained filters retained eggs. In view of the implication of these findings for epidemiological studies and control programmes, it is suggested that field workers consider the use of disposable paper filters.

Evaluation of the ELISA test as an epidemiological tool in schistosomiasis.

The ELISA test was studied to assess its applicability in epidemiological surveys of schistosomiasis. Specimens of serum, stool and urine from persons attending an outpatient clinic in an endemic area for S. mansoni and S. haematobium in Egypt were tested in the laboratories of the Theodor Bilharz Research Institute, Cairo. The ELISA test results were compared with parasitological findings and with the results of indirect immunofluorescence and indirect hemagglutination tests as reference data. The ELISA test showed an excellent sensitivity and specificity for both infections; the reading and recording of the extinction rates can be automated by using in time and money. The test can be strongly recommended for epidemiological surveys.

Preliminary studies on the histochemical differentiation of strains of Onchocerca volvulus microfilariae in Togo.

Skin snips were taken from 75 people living in four villages of northern Togo. The 7824 microfilariae that emerged were examined by staining for the presence of acid phosphatase. Four distinct patterns of enzyme staining were observed, and descriptions of the stained microfilariae are given. The study confirms the view that a number of biological strains or variants of Onchocerca volvulus coexist in West Africa, and suggestions are made for further research that could result in the practical application of these observations in onchocerciasis control programmes.

Bancroftian Filariasis in Togo 1. a comparative field study of the membrane filtration concentration technique and conventional blood films.

The membrane concentration filtration technique which has proved very successful in use in areas of subperiodic filariasis in the Pacific, has been used to study an area of nocturnally periodic in Togo. The results obtained were similar to those from areas of low endemicity in the Pacific. The method was easy to apply under field conditions in Africa and the taking of venous blood samples proved competely acceptable to a rural population in south-eastern Togo.

Epidemiological studies on onchocerciasis by means of a new field technique.

A new membrane filter concentration technique for the detection and quantification of Onchocerca volvulus microfilariae in skin snips was compared for sensitivity and efficiency with a widely used "standard" technique. A field study was carried out in five villages in an onchocerciasis focus north-east of the town of Sokodé, Mô river valley, Togo. Use of the new technique resulted in a substantial rise in the observed prevalence and density of microfilariae.

A new technique for the determination of microfilarial densities in onchocerciasis.

Precise measurement of the parasite load in helminthic infections makes it possible to correlate morbidity, the individual response to treatment, and the best control methods. A membrane filtration technique was recently developed for quantifying Wuchereria bancrofti microfilaraemia. The present paper describes its application to onchocerciasis in a study on 107 patients in northern Togo. The technique is cheap and simple to apply under field conditions, and is extremely efficient at recovering microfilariae from skin snips.