Multiple parasitic infections are a common phenomenon noted in populations living in socio-economically disadvantaged settings. They represent a common fact in the daily realities of clinical health services (Petney & Andrews 1998; Drake & Bundy 2001). Despite these realities, relatively little attention have been given to systematically document this phenomenon and determine underlying risk factors. To close these knowledge gaps, a number of studies have been carried out in Africa (Keiser et al. 2002;
Raso et al. 2004; Matthys et al. 2007), China (Steinmann et al. 2008) and Southeast Asian countries (Ezeamama et al. 2005).
Discussion
In Lao PDR, only concurrent infections with O. viverrini and small intestinal trematode have been described (Chai et al. 2005, 2007). To our knowledge, no data on co-infections with different intestinal helminths as well as intestinal protozoa were available prior to out studies for Lao PDR. However, several parasites have been reported, starting from food-borne (e.g. O. viverrini, MIF, Paragonimus, and F. buski), to soil-transmitted helminths, S. mekongi, and intestinal protozoa, together with the absence of sanitary facility, low socio-economic status and raw food consumption behaviour. A nationwide survey among schoolchildren has shown a high geographical overlap of different helminth prevalence, hence co-infection must occur (Rim et al. 2003).
Our studies showed that people living in the rural areas of the country were currently suffering from multiparasitism rather than single-species infection. For example, in Saravane province (Sayasone et al. 2007) almost two-third of study participants harboured two or more parasite species. In Champasack province we could confirm rampant multiparasitism (Sayasone et al. 2009a). Fifteen different intestinal parasite species were discovered belonging to the family of Trematodiae, Nematodiae and Cestodiae and to intestinal protozoa, and 4 out of 5 study participants harboured two or more intestinal parasite species. A strong positive association was observed for co-infection between ‘O. viverrini’-like and S. mekongi, hookworm and A. lumbricoides, hookworm and T. trichiura and E. coli and A. lumbricoides. This is the first study pertaining to co-infection of different intestinal parasites starting from trematodiasis to intestinal protozoa infections in the country, and assessing the underlying risk factors.
Furthermore, a positive association between ‘O. viverrini’-like and S. mekongi was described, to our knowledge for the first time ever. The positive association between soil-transmitted helminths and intestinal protozoa have been investigated before for Côte d’Ivoire and China (Raso et al. 2004; Steinmann et al. 2008), and have now been extended to Lao PDR.
The presence of extensive multiparasitism is challenging the epidemiological study in the country; until today there is no a single standardized tool capable to detect the multiple intestinal parasitic infections with the high sensitivity (Bergquist et al. 2009).
Discussion
survey. However, those techniques have some limitations on their performance (Bergquist et al. 2009). The KK technique has been commended by WHO for epidemiological survey related to STH (WHO 1993). It equips with basic materials, allows screening a large sample size of population and is applicable in the field work.
However, it does not allow screening the infection with S. stercoralis and intestinal protozoa. Most importantly, it could not differentiate the O viverrini eggs from those of MIF. Moreover, the KK-technique is also known as having the low sensitivity detecting the parasite’s eggs in the stool if only one sample is analyzed (de Vlas & Gryseels 1992;
Kongs et al. 2001; Utzinger et al. 2001; Booth et al. 2003). Due to the relatively small amount of faeces (41.7 mg), light infections may be missed, consequently, underestimating the ‘true’ community prevalence and it has additionally noted as having a low sensitivity for hookworm diagnosis (Booth et al. 2003; Raso et al. 2004; Steinmann et al. 2008). A prolongation time from stool sample collection and smear slide preparation and reading results in a substantial reduction of eggs visualized under a light microscopy.
The FECT method is usually a standard parasitological tool of a laboratory. It is equipped with the basic materials e.g. tubes, formalin, ether, pipettes, normal saline and centrifuge (WHO 1998). This technique is able to screen a large sample size of conserved-stool samples. It is capable to detect the helminth eggs, cyst of intestinal protozoa and larvae of S. stercoralis. However, FECT is not the recommended technique for S. stercoralis diagnosis and hence the more reliable techniques are commended such as Koga agar plate and Baermann technique (Koga et al. 1991; Stothard et al. 2008;
Steinmann et al. 2008; Knopp et al. 2008). Utility of FECT method for distinguishing the eggs of O. viverrini from eggs of those MIF has been described in the literature (Elkins et al. 1991; Sithithaworn et al. 1991; Tesana et al. 1991). The O. viverrini diagnosis demonstrates the presence of rough eggshells, knob and prominent shoulder, which are more or less prominent in eggs of MIF. With a 400 times magnification these characters are clearly visible (Tesana et al. 1991).
With these limitations, the quantitative assessment of multiple parasitic infections in the population is only possible by the combination of different diagnostic techniques.
Discussion
abundant food-borne parasitic infections. The particular quantitative assessment of different species of FBT, including species assessments of liver and intestinal flukes makes this endeavour challenging. In our present study, we introduced the purgation of study participants after treatment as an additional technique to the routine stool examination, i.e. the examination of the entire stool collected after treatment. The adult flukes were collected, identified and counted. The results showed that 83% of the examined person harboured at least 2 parasites species (Sayasone et al. 2008b).
In addition to the stool sample analysis, six different species of small intestinal trematodes and one of Echinostomatidae were identified only in the purging study. These findings further document that multiparasitism is indeed the norm rather than the exception in the developing world and underscores the methodological challenges which need to be tackled when addressing quantitative research questions related to multiple parasitic infections in a specific setting (Raso et al. 2004; Sachs & Hotez 2006; Dung et al. 2007; Steinmann et al. 2008).
Furthermore, the evaluation of FECT performance in considering the purging results as the diagnostic ‘gold’ standard showed a correct O. viverrini egg diagnosis (e.g.
sensitivity 96.8% and specificity 100.0%). A weaker performance (e.g. sensitivity 85.0%
and specificity 70.6%) of later test was addressed for the MIF egg diagnosis. In fact, in the dissection of O. viverrini worms, we observed that a small proportion of O. viverrini eggs (5%) is very similar to MIF eggs, i.e. without prominent shoulders and knobs (Tesana et al. 1991), therefore a rigorous examination with FECT technique is necessary in order to correctly classify these eggs. In addition, the purgation underestimate the prevalence of MIF, since the collection of tiny heterophyid flukes sized 0.3-0.7 mm wide and 1.0-2.5 mm long (Radomyos et al. 2004; Garcia 2007) in the purging stool samples under a stereo-microscope is challenging, particularly in the cases with light infections.
Most recently a promising polymerase chain reaction (PCR) method has been described (Stensvold et al. 2006; Duenngai et al. 2008; Sato et al. 2009; Lovis et al. 2009) for accurate differentiation of O. viverrini and MIF infection. These techniques need
Discussion
are limited FECT method remains the method of choice for distinguishing O. viverrini infection from those of MIF.
The FLoTAC technique has recently been developed (Cringoli 2004, 2006;
Utzinger et al. 2008; Knopp et al. 2009). This method is capable to detect the concomitant infection of helminths and intestinal protozoa in SAF-conserved stool samples with high sensitivity. The screening of stool sample with highest microscopic magnification (100x) results in an accurate diagnosis of helminth species (Cringoli 2006;
Utzinger et al. 2008; Knopp et al. 2009). If this is also a case for food-borne trematode infections, in particular O. viverrini and those MIF, then it will offer a new means for assessment the diversity of FBT and in turn, it contributes to provide the comprehensive data on multiparasitism in places, where several food-borne trematode species are endemic.