Microhaematuria and Proteinuria Performance as a Measured by Urine Reagent Strips in Estimating Intensity and Prevalence of Schistosomahaematobium Infection in Nigeria

Microhaematuria and proteinuria as measured by urine reagent strips are widely used to screen population at high risk o f urinary schistosomiasis. This investigation was conducted to assess, if microhaematuria and proteinuria as measured by reagent strips could estimate intensity of Schistosomahaematobium infection in endemic areas and evaluate their screening performance among children in Benue State, Nigeria. A total of 1,124 urine samples were collected, screened for microhaematuria and proteinuria using reagent strips (Combi 9) and compared to filtration technique the gold standard method. A significant correlation was observed between microhaematuria (rho= 0.66, p<0.01), proteinuria (rho = 0.71, p<0.01) and intensity of Schistosomahaematobium eggs. Proteinuria had sensitivity of 95.7% and specificity of 67.7%, while microhaematuria had sensitivity of 64.8% and specificity of 89.6%. The proportion of false positive diagnoses was higher in proteinuria (19.6%) than microhaematuria (6.0%). The find ings suggest that use of urine reagent strips could potentially estimate intensity of Schistosomahaematobium infection and their performance to screen urinary schistosomiasis agreed with previous observations.


Introduction
Urinary schistosomiasis is a major debilitating disease caused by Schistosomahaematobium and characterized by the presence of blood in urine. Other sympto ms are proteinuria, dysuria, b ladder carcino ma, b ladder stones, calcificat ion of bladder wall and somet imes renal failure.
The distribution of schistosomiasis has changed over time with some countries in South America, Asia, the Caribbean and the Middle East bringing down the prevalences of the disease through a concerted public health effort [1].In sub-Saharan Africa, prevalence levels have increased and vary fro m one country to another; this is mostly because of water resources development, roads and dams projects, irrigationof land for agricu ltural purposes, inactive control programme and mostly neglect from the part of governments to implement control programmes in endemic areas.
Nigeria is one of the highly endemic countries where the disease has been unsystematically reported and large areas remain whose disease status is unknown [2]. Screening using rapid, ind irect tests has been proposed as a procedure to simplify mapping surveys [3]. Haematuria (blood in urine) has been proposed as a valid indication of current infection in Schistosomahaematobium endemic populations [2,4,5,6].
Testing urine with reagent strips for microhaematuria and proteinuria is such a simp le and indirect diagnostic technique that could estimate the prevalence of urinary schistosomiasis in school children of endemic co mmun ities. Operat ional research studies in Africa showed that screening using reagent strips is an effective method to identify school children requiring treat ment and subsequently monitor control [7,8,9]. Several research studies reported high sensitivity and specificity of reagent strips compared to urine filtrationconsidered the most conclusive diagnostic for urinaryschistosomiasis and this procedure is expensive, cumbersome and too technical fo r lay use [2,7]. However, this study was undertaken to assess if microhaematuria and proteinuria as detected with reagent strips could estimate intensity of Schistosomahaematobiuminfection in endemic areas and evaluate their diagnostic performance in screening urinary schistosomiasis among children in Benue State, Nigeria.

Study Area
The study was conducted from November 2008 to September 2009 in Buru ku and Katsina-Ala local government areas (LGAs) of Benue State, Nigeria which are known for their endemicity for urinary schistosomiasis [10,11]. Before the co mmencement of the study, permission was sought fro m Directors of health and local government education authorities of both areas. The climate of the areas is tropical with two seasons, the dry season which starts from October to March and the rainy fro m April to October. Agricultural activ ities like crop farming and rearing of animals are the mainstay of the inhabitants.
Immediately after testing with the reagent strips, 1ml of ordinary household bleach was added to each collected urine sample to preserve any ova present and then taken to the laboratory within 2hrs for parasitological examination. 10 ml of urine was taken and filtered through a 12µm polycarbonate membrane in a filter holder. W ith the help of a forceps, the filter was removed fro m the filter holder and placed on a slide. A drop of Lugol's iodine was added and the slide examined under microscope using x10 and x40 objective lenses. The number of eggs was counted per 10 ml of urine and intensities of infection were classified as 1-10 eggs, 11-49 eggs and > 50eggs for light, moderate and heavy infections respectively.

Statistical Analysis
Collated data were double entered in Microsoft excel and analysed in PASW (Predict ive Analysis software) version 18.0. Associations between variables were tested using Spearman correlation (rho) at p < 0.01 significance level.
The diagnostic performance of microhaematuria and proteinuria was assessed by calculating sensitivity, specificity, positive pred ictive value and negative predictive value using the following formu lae.
The comparison betweenmicrohaematuria as indicator of urinary schistosomiasis and the true disease status as determined by filtrat ion technique shows that microhaematuria was detected in 370 (32.9 %), among these302 (26.9 %) had both microhaematuria and presence of eggs (true positive) and 68(6.0 %) had microhaematuria with no p resence of eggs (false negative). Of the 754 (67.1%) screened not havingmicrohaematuria in their urine, 164 (14.6 %) had Schistosomahaematobiumeggs (false positive) and 590 (52.5 %) were devoid of eggs (true negative). Table 2shows the relationship between proteinuria and intensity of Schistosomahaematobium eggs among children in Katsina-Ala and Buru ku LGAs o f Benue State. It was observed that of the 317 screened having proteinuria at Ca.30 (+), light infect ion recorded the highest rate with 118 (37.2 %), wh ile moderate in fection recorded the highest rate with 84 (37.3 %) out of the 225 screenedfor proteinuria at Ca.100(++). Heavy and moderate infect ion intensities recorded 52(43.3 %) and 54 (45.0%) respectively out of the 120 screened for proteinuria at Ca.500 (+++). It was observed that various degree of proteinuria concentrations, +, ++ and +++ co rresponded to highest rateof light, moderate and heavy intensity of eggs respectively although moderate infection was found having an edge over heavy infection at Ca.500. A significant spearman correlation (rho= 0.71, p < 0.01) was found between different degrees of proteinuria Estimating Intensity and Prevalence of Schistosomahaematobium Infection in Nigeria concentrations and intensity of S. haematobium eggs.
The comparison between proteinuria as indicator of urinary schistosomiasis and the true disease status as determined by filtrat ion technique shows that proteinuria was observed in 662 (58.9%) children, 446(39.7%) had both proteinuria and Schistosomahaematobiumeggs (true positive), while 216(19.2 %) had proteinuria with absence ofS. haematobium eggs (false positive). Of the 442 screened having no proteinuria in their urine, 20(1.8 %) had S.haematobium eggs, while 442(39.3%) were devoid of S. haematobium eggs (true negative).
The ability of microhaematuria and p roteinuria to accurately identify all those with the disease (sensitivity) was 64.8% and 95.7% respectively, wh ile their ab ility to correctly sort out all those without the disease (specificity) was 89.7 % and 67.2 % respectively.Microhaematuria had higher positive predictive value (PPV) (81.6 %) than proteinuria (67.7%), but had lower Negative Pred ictive Value (NPV) 78.2% against 95.6%.

Discussion
The present study demonstrates that the use of microhaematuria and proteinuria to estimate the intensity of urinary schistosomiasis has potential utility in discriminating intensity of infection among infected individuals in endemic areas.
The significant relat ionships observed between microhae maturia, p roteinuria and filtration technique clearly demonst rate that the presence or absence of microhaematuria or proteinuria in urine is function of Schistosomahaematobi um eggs excret ion in urine. However, the false positive results of microhaematuria and proteinuria observed entails the daily variat ion of S. haematobium eggs excretion in infected indiv iduals. The absence of microhaematuria and proteinuria in infected indiv iduals (false negative) could be the result of new infection in wh ich tissues of the urinary bladder and kidney have not been damaged yet.
The evaluation of only microhaematuria as indicator of urinary schistosomiasis shows sensitivity of 64.8 % and specificity of 89.6 %. This is closely related to findings of Ugbomoikoet al. [9] who reported sensivity of 68.3 % and specificity of 83.2 % among school child ren of two endemic areas in southwestern, Nigeria. Ho wever, the sensitivity of microhaematuria in this study is higher than that of Anosikeet al. [2] who obtained sensitivity of 41.0 % but with a similar specificity (82.0%) in a study conducted in Bende LGA of Abia State, Nigeria. Sensitiv ity and specificity of microhaematuria in this study are lower than that reported among zan zibari school children in Tan zania with sensitivity of 77.0%) and specificity of 97.0 % [8].Ho wever, variation in sensitivity and specificity of microhaematuria during Schistosomahaematobiuminfection has been reported in several studies conducted in different African settings. They have been reported to vary fro m 41.0 % to 93 % and fro m 67 % to 99 % fo r sensitivity and specificity respectively [2,4,6,8,9,12].  The diagnostic performance of proteinuria as indicator of urinary schistosomiasis showed sensitivity rate of 95.7 % and specificity of 67.2% wh ich are higher than findings of Brou weret al. [13 who obtained 65.0% and 60.0%respective ly among Zimbabwean school children. Ugbomo ikoet al. [9] reported lower sensitivity (67.7 %) and higher specificity (79.6 %) among school child ren in southwestern, Nigeria than the present study. Proteinuria is seen as a sign of b ladder damage occurring principally in severe infections. The high sensitivity (95.7%) of proteinuria observed in this study cannot be conclusive about S. haematobiuminfection; this is because of the relatively h igh rate o f false positive result (19.2%) than that of microhaematuria (6.0 %). However, this false positive result could be due to other urinary tract infections.
The use of urine reagent strips has been proposed as an indirect method in identify ing S.haematobium infected children, hence a useful tool to rapidly map the prevalence of urinary schistosomiasis in endemic areas.
The present study also shows that using microhaematuria and proteinuria as detected by reagent strips easily estimate intensity of infection; this is because they showed ability to discriminate between children at different level of infection intensity. Moreover, it was found that microhaematuria and proteinuria with higher positivity level detected infection of higher intensity (moderate and heavy).
Microhaematuria performed better in detecting moderate and heavy infection in the ch ildren than proteinuria. However, this corroboratesfindings of Anosikeet al. [2] and Ahmed [14]who found similar results among Nigerian and Yemeni children.
The positive predictive value (probability of infected children with S.haematobiumeggs among those having microhaematuria or proteinuria) was higher in children having microhaematuria(81.6 %) than proteinuria (67.7 %). This indicates that almost all children with microhaematuria were indeed infected with S.haematobium eggs.
In conclusion, using microhaematuria and proteinuria as detected by reagent strips is practical-cheap, fast and easy to use in estimat ing intensity and prevalence of S.haematobium infection. This can be used in Primary Health Care (PHC) setting with limited resources to screen and monitor S. haematobium infection in endemic areas.