Control approaches for Opisthorchis viverrini and co-infections in Lao PDR

Background: In Lao People’s Democratic Republic (Lao PDR) helminth infections are highly prevalent. All groups of helminths, including nematodes, trematode and cestodes can be found throughout the country. Besides high rates and intensity of infections, most individuals are infected with several different species of intestinal parasitic infections. As in many other countries, a main underlying factor for high worm infestations is the scarce economic resources which lead to a lack of appropriate sanitary facilities, and hygiene related education and adequate behaviour. Therefore, the parasite infection is common among the rural population. However in Laos, there is an additional, particular risk factor responsible for the populations, high worm-load; namely the consumption of raw or insufficiently cooked foodstuff such as raw meat, fish and vegetables. This habit is deeply culturally rooted, and widespread in the Lao population. In this PhD thesis, we tested and determined the appropriate control approaches for Opisthorchis viverrini (O. viverrini) and other important helminth infections such as Strongyloides stercoralis (S. stercoralis) and Schistosoma mekongi (S. mekongi). The tested approaches compose of latrine promotion programme, eco-health intervention study and novel urine test for S. mekongi diagnosis in Khong District, Champasack Province, Lao PDR. Goal and specific objectives: The present PhD study aimed to develop the appropriate control approaches for O. viverrini infection and co-infections in Lao PDR. Four specific objectives were pursued: i). To assess S. stercoralis infection and the risk of infection among the populations on three islands in Khong district, Champasack province, Southern Laos; ii). To define O. viverrini, S. mekongi and STH infections in humans in the ecological environment of Khong district, Champasack province where their potential animal reservoir, and intermediate hosts are living in close connectivity; iii).To compare the diagnostic tools for detection of S. mekongi infection in Lao People’s Democratic Republic and neighbouring country Cambodia; and iv). To assess the impact of improved sanitation and its use on the transmission of intestinal helminth infections in highly endemic areas, three islands in Khong district, Champasack province, Southern Laos. Methodology: Both cross-sectional and cohort studies were used in this PhD thesis’s research. All data of this PhD thesis were obtained from community-based studies. First study of this PhD research was latrine intervention which was called as latrine study, conducted in March 2011 to January 2013 on three islands, i.e. Donlong, Donthan and Donlieng island located in the Mekong River in Khong district, Champasack province, southern Laos. Given an experimental pre-test and post-test with one control group was used to assess the effects of latrine in the study villages on preventing of helminth infections particularly O. viverrini and S. mekongi. Household-based promoting latrine construction was conducted. Two stool samples were collected per study participants within a five day period. Each sample was examined by using Kato-Katz (KK) thick smears and Baermann technique. There were three different phases of the study which described elsewhere in material and methods section of Chapter 7. A baseline study of ecohealth intervention for O. viverrini, S. mekongi and STH was conducted in October 2011 and August 2012 on Done Khon and Done Som islands. Household members aged two years and older and potential animal reservoir hosts, i.e., dogs, cats, pigs and buffaloes, from selected households were enrolled and examined for helminth infections. For O. viverrini, snails of the genus Bithynia spp. were collected with a scoop from water bodies near the study villages (e.g., ponds, canals, and rice fields). Cyprinoid freshwater fish were captured from the same selected water bodies as well as from the Mekong using fishing net. For S. mekongi, N. aperta snails were hand-picked from the rocky area of the Mekong River. Two KK thick smears, formalin ether concentration technique (FECT), the shedding test and the pepsin enzyme digestion technique were performed for human, animal reservoir hosts, Bithynia spp. and N. aperta snail hosts and Cyprinoid fish, respectively to assess their infections. The evaluation of the new diagnostic techniques for S. mekongi was conducted between February and April 2016 in S. mekongi-endemic villages in Lao PDR and Cambodia. Urine and serum samples were obtained from each study participant to be tested for Schistosoma infection by POC-CCA, UCP-LF CAA and ELISA assays. We collected three stool samples from each participant during five consecutive days. Stool samples were subjected to examination by duplicate KK thick smears examined under a light microscope. Principle findings/results: Our first report was from the first phase of latrine study which emphasized on epidemiology of S. stercoralis infection on Mekong islands, showed 41.0% overall prevalence rate. The infection rate did not differ between the islands (Donlong 44.1% vs. Donthan/Donlieng 38.2%, p=0.107). The highest infection rate was observed with O. viverrini (72.2%), followed by hookworm (56.1%) and S. mekongi (12.8%). Trichuris trichiura (3.3%), Ascaris lumbricoides (0.3%) and Taenia spp. (0.3%). The most important risk factor was sex. Male study participants had a significantly higher risk for a S. stercoralis infection than female participants taking into account the age of the study participants (adjusted OR 1.97, 95% CI 1.45–2.67). The baseline result of our ecohealth intervention revealed that human infection rates with O. viverrini, hookworm, S. mekongi, T. trichiura, A. lumbricoides and Taenia spp. were 60.7%, 44.1%, 22.2%, 4.1%, 0.6% and 0.1%, respectively. Heavy intensity infections were 4.2%, 3.6% and 1.8% for O. viverrini, S. mekongi and hookworm, respectively. . O. viverrini infection rate among dogs and cats were 25.0% and 53.1%, respectively. S. mekongi infection rates among dogs were 14.7%.. Prevalence of O. viverrini and S. mekongi in snails was 0.3% and 0.01%, respectively. Overall prevalence of O. viverrini infection in fresh water fish was 26.9%, with the highest infection rates occurring in Hampala dispa (87.1%), Cyclocheilichthys apogon (85.7%) and Puntius brevis (40.0%). Illiteracy and lower socioeconomic status increased the risk of O. viverrini infection, while those aged 10–16 years and possessing latrines at home were less likely to be infected. Household dogs and cats that consumed raw fish were significantly and positively associated with O. viverrini infection of the household members. For S. mekongi, children under 9 years old were exposed significantly to this infection, compared to older age groups. Study on comparison of novel and standard diagnostic tools for the detection of S. mekongi infection in Lao PDR and Cambodia which was carried out between February and April 2016. Stool microscopy by KK thick smear revealed an overall prevalence of S. mekongi of 6.4% (one case in Cambodia and 23 cases in Lao PDR), while that of O. viverrini, hookworm, T. trichiura, A. lumbricoides and Taenia spp. were 50.4%, 28.1%, 3.5%, 0.3% and 1.9%, respectively. In total, 377 urine and serum samples were tested for S. mekongi infection. In the urine samples, the tests for CCA and CAA detected S. mekongi infections in 21.0% and 38.7% of the study participants, respectively. In the serum samples, the CAA assay revealed a prevalence of 32.4%, while a combination of the CAA assay in serum and in urine revealed a prevalence of 43.2%. There was a difference between the two study locations with a higher prevalence reached in the samples from Lao PDR. Study on assessment the effects of latrine in the study villages on preventing of helminth infections particularly O. viverrini and S. mekongi which was performed by using an experimental pre-test and post-test with one control group. In the intervention villages, the helminth infection rates at baseline for S. mekongi, O. viverrini, hookworm, A. lumbricoides, T. trichiura, Enterobius vermicularis, S. stercoralis and Taenia spp. were 28.6%, 79.5%, 48.8%, 0.3%, 3.5%, 0%, 43.1% and 0%, respectively. At follow-up they were reduced to 22.6%, 68.2%, 26.2%, 2.5%, 1.1%, 0.4%, 31.1% and 0.4%, respectively. Reduction in prevalence of four important helminthiasis were significantly observed (baseline vs follow-up, P-value), S. mekongi (28.6% vs 22.6%, P-value <0.001), O. viverrini (79.5% vs 68.2%, P-value <0.001), Hookworm (48.8% vs 26.2%, P-value <0.001) and S. stercoralis (43.1% vs 31.1%, P-value <0.001) in the intervention villages. While S. mekongi infection in the control villages was increased (1.8% vs 2.6%, P-value =0.74) and other helminths were decreased O. viverrini (71.8% vs 59.9%, P-value =0.027), Hookworm (65.6% vs 38.3%, P-value <0.001) and S. stercoralis (38.3% vs 34.8%, P-value =0.001). Conclusion/Significance: O. viverrini, S. mekongi, and STH particularly S. stercoralis were still high prevalence in Mekong islands as well as the multiparasitism was observed in all studies of this PhD thesis works. There is a pressing need to design and implement an integrated helminth control intervention on the Mekong Islands in southern Lao PDR. An appropriate integrated control approach involving interventions targeting human behaviour, animal reservoirs, and environmental modification, health education and improved access to clean water and adequate sanitation might improve the effectiveness of interventions and lead to the elimination of infections. Furthermore, the new diagnostic tool CCA, CAA and ELISA were evaluated and showed a substantially higher prevalence estimates for S. mekongi compared to Kato-Katz thick smears. Active schistosomiasis mekongi in Lao PDR and Cambodia might thus have been considerably underestimated previously. Hence, sustained control efforts are still needed to break transmission of S. mekongi. The pivotal role of highly sensitive diagnostic assays in areas targeting elimination cannot be overemphasised

Risk profiling of hookworm infection and intensity in southern Lao People's Democratic Republic using bayesian models

Among the common soil-transmitted helminth infections, hookworm causes the highest burden. Previous research in the southern part of Lao People's Democratic Republic (Lao PDR) revealed high prevalence rates of hookworm infection. The purpose of this study was to predict the spatial distribution of hookworm infection and intensity, and to investigate risk factors in the Champasack province, southern Lao PDR.; A cross-sectional parasitological and questionnaire survey was conducted in 51 villages. Data on demography, socioeconomic status, water, sanitation, and behavior were combined with remotely sensed environmental data. Bayesian mixed effects logistic and negative binomial models were utilized to investigate risk factors and spatial distribution of hookworm infection and intensity, and to make predictions for non-surveyed locations.; A total of 3,371 individuals were examined with duplicate Kato-Katz thick smears and revealed a hookworm prevalence of 48.8%. Most infections (91.7%) were of light intensity (1-1,999 eggs/g of stool). Lower hookworm infection levels were associated with higher socioeconomic status. The lowest infection levels were found in preschool-aged children. Overall, females were at lower risk of infection, but women aged 50 years and above harbored the heaviest hookworm infection intensities. Hookworm was widespread in Champasack province with little evidence for spatial clustering. Infection risk was somewhat lower in the lowlands, mostly along the western bank of the Mekong River, while infection intensity was homogeneous across the Champasack province.; Hookworm transmission seems to occur within, rather than between villages in Champasack province. We present spatial risk maps of hookworm infection and intensity, which suggest that control efforts should be intensified in the Champasack province, particularly in mountainous areas

Transmission of Opisthorchis viverrini, Schistosoma mekongi and soil-transmitted helminthes on the Mekong Islands, Southern Lao PDR

Prevalence of Opisthorchis viverrini, Schistosoma mekongi and soil-transmitted helminths (STH) remains high in Lao People's Democratic Republic (Lao PDR), despite control efforts including mass-drug administration, education and communication campaigns. New approaches are required to advance helminth control.; An ecohealth study was conducted on two Mekong islands in Southern Laos. Demographic and behavioural data were collected by questionnaire. Human and animal reservoir stools were examined. Bithynia spp. and Neotricula aperta snails were examined using shedding. Fresh water fish were examined using digestion technique. Multivariate random-effects analysis was used to find risk factors associated with helminth infections.; Human infection rates with O. viverrini, hookworm, S. mekongi, Trichuris trichiura, Ascaris lumbricoides and Taenia spp. were 60.7%, 44.1%, 22.2%, 4.1%, 0.6% and 0.1%, respectively. Heavy intensity infections were 4.2%, 3.6% and 1.8% for O. viverrini, S. mekongi and hookworm, respectively. O. viverrini and S. mekongi infection rates among dogs and cats were 25.0% and 14.7%, respectively. Of the cats tested, 53.1% were infected with O. viverrini. Prevalence of O. viverrini and S. mekongi in snails was 0.3% and 0.01%, respectively. Overall prevalence of O. viverrini infection in fresh water fish was 26.9%, with the highest infection rates occurring in Hampala dispa (87.1%), Cyclocheilichthys apogon (85.7%) and Puntius brevis (40.0%). Illiteracy and lower socioeconomic status increased the risk of O. viverrini infection, while those aged 10-16 years and possessing latrines at home were less likely to be infected. Household dogs and cats that consumed raw fish were significantly and positively associated with O. viverrini infection of the household members. For S. mekongi, children under 9 years old were exposed significantly to this infection, compared to older age groups.; There is a pressing need to design and implement an integrated helminth control intervention on the Mekong Islands in southern Lao PDR. Given the highly dynamic transmission of O. viverrini, S. mekongi, STH and extended multiparasitism, annual mass-drug administration is warranted along with environmental modifications, health education and improved access to clean water and adequate sanitation to consolidate morbidity control and move towards elimination.; Our findings presented here are from a cross-sectional study, therefore, it has not been registered

Comparison of novel and standard diagnostic tools for the detection of Schistosoma mekongi infection in Lao People's Democratic Republic and Cambodia

Given the restricted distribution of Schistosoma mekongi in one province in Lao People's Democratic Republic (Lao PDR) and two provinces in Cambodia, together with progress of the national control programmes aimed at reducing morbidity and infection prevalence, the elimination of schistosomiasis mekongi seems feasible. However, sensitive diagnostic tools will be required to determine whether elimination has been achieved. We compared several standard and novel diagnostic tools in S. mekongi-endemic areas.; The prevalence and infection intensity of S. mekongi were evaluated in 377 study participants from four villages in the endemic areas in Lao PDR and Cambodia using Kato-Katz stool examination, antibody detection based on an enzyme-linked immunosorbent assay (ELISA) and schistosome circulating antigen detection by lateral-flow tests. Two highly sensitive test systems for the detection of cathodic and anodic circulating antigens (CCA, CAA) in urine and serum were utilized.; Stool microscopy revealed an overall prevalence of S. mekongi of 6.4% (one case in Cambodia and 23 cases in Lao PDR), while that of Opisthorchis viverrini, hookworm, Trichuris trichiura, Ascaris lumbricoides and Taenia spp. were 50.4%, 28.1%, 3.5%, 0.3% and 1.9%, respectively. In the urine samples, the tests for CCA and CAA detected S. mekongi infections in 21.0% and 38.7% of the study participants, respectively. In the serum samples, the CAA assay revealed a prevalence of 32.4%, while a combination of the CAA assay in serum and in urine revealed a prevalence of 43.2%. There was a difference between the two study locations with a higher prevalence reached in the samples from Lao PDR.; The CCA, CAA and ELISA results showed substantially higher prevalence estimates for S. mekongi compared to Kato-Katz thick smears. Active schistosomiasis mekongi in Lao PDR and Cambodia might thus have been considerably underestimated previously. Hence, sustained control efforts are still needed to break transmission of S. mekongi. The pivotal role of highly sensitive diagnostic assays in areas targeting elimination cannot be overemphasised

Spatial Distribution of, and Risk Factors for, Opisthorchis viverrini Infection in Southern Lao PDR

The liver fluke Opisthorchis viverrini mainly occurs in Lao PDR and Thailand. Humans become infected through the consumption of raw or insufficiently cooked freshwater fish. Chronic infections may lead to severe liver (bile duct) diseases that eventually develop into a bile duct cancer with extremely poor prognosis. Current control efforts aim at preventing heavy morbidity and mortality. In recent years, spatial modeling, using data from well designed surveys, has been employed to better understand the distribution and determinants of parasitic diseases for guiding subsequent control. However, a spatial modeling approach has not been used for O. viverrini before. The purpose of the current study was to map the distribution of O. viverrini infection in Champasack province in southern Lao PDR, to identify risk factors of infection, and to predict the distribution at non-surveyed locations. We found that the risk of O. viverrini infection is higher for people living in close proximity to freshwater bodies, whereas the lack of sanitation sustained environmental contamination and transmission. High risk zones in Champasack province are concentrated in the Mekong River corridor, and hence control efforts should be targeted along the Mekong River

Helminth infection in southern Laos: high prevalence and low awareness

BACKGROUND: Helminthiasis is a public health concern in Lao People’s Democratic Republic (Lao PDR, Laos). This study aimed to understand helminth infection and associated risk factors in relation to existing local knowledge, attitudes and practices regarding worm infections in endemic communities. METHODS: A cross-sectional survey was conducted in 10 randomly selected villages in Saravane district, Southern Laos. Two stool samples obtained from 574 members (aged ≥2 years) of selected households were examined using the Kato Katz method. Household heads (n = 130) were interviewed. Eight focus group discussions (FGDs) and direct observations were performed. Uni- and multivariate logistic regression analyses were conducted to predict infection. Content analysis was conducted for qualitative data. RESULTS: The prevalence of Opisthorchis viverrini, hookworm, Trichuris trichiura, Ascaris lumbricoides and Taenia sp. was 88.7%, 86.6%, 32.9%, 9.8% and 11.5%, respectively. Most individuals were co-infected with O. viverrini and hookworm. More men had multiple helminth infections than did women. Only one-third of household heads had heard about liver fluke before, of which 59.2% associated it with eating raw fish dish. Among the soil-transmitted helminths, roundworm was the most well known (70.8%) but was attributed to raw food consumption (91.3%). Eating raw fish was a common practice (75.4%); few households possessed a latrine (16.1%); less than half of the study participants mentioned health benefits from latrine use and personal hygiene. Focus group discussion participants had a low level of awareness of worm infections; more men liked eating raw fish than did women; some disliked using latrines because they were not used to it and because of their bad smell. Poor personal and village hygiene practices were observed. CONCLUSIONS: This study highlights a high helminth infection rate and poor community awareness of worm and associated risk factors. Only a sound awareness of worm infection and the underlying risk factors may prevent infection and re-infection after treatment

Epidemiology of Strongyloides stercoralis on Mekong islands in southern Laos

Strongyloides stercoralis is a neglected helminth infection potentially that can lead to systemic infection in immunocompromised individuals. In Lao People's Democratic Republic (Lao PDR, Laos), information on S. stercoralis infection is scarce. We assessed S. stercoralis infection and associated risk factors and symptoms on the Mekong islands in Southern Laos. Baermann and Kato-Katz techniques were performed on two stool samples from each individual to detect S. stercoralis larvae and concomitant helminth infections. Among 729 individuals, 41.0% were infected with S. stercoralis. Men were at higher risk than women (OR 1.97, 95% CI 1.45-2.67). Urticaria and body itching was associated with S. stercoralis infection (OR 2.4, 95% CI 1.42-4.05). Infection with Opisthorchis viverrini (72.2%), Schistosoma mekongi (12.8%), and hookworm (56.1%) were very common. Few infections with Trichuris trichiura (3.3%), Ascaris lumbricoides (0.3%) and Taenia spp. (0.3%) were detected. The majority of helminth infections were of light intensity, with prevalences of 80.4%, 92.9%, 64.5%, 100% and 100%, for O. viverrini, hookworm, S. mekongi, T. trichiura and A. lumbricoides, respectively. Nevertheless, heavy infection intensities were observed for O. viverrini (1.0%), S. mekongi (14.0%) and hookworm (2.9%). S. stercoralis is highly endemic on the islands of Khong district, Champasack province, Southern Laos. The national helminth control programme should take action to control this helminth infection

Suspected cases of cholangiocarcinoma seen in reference hospitals in Lao People's Democratic Republic

Cholangiocarcinoma (CCA) poses a significant public health problem in Southeast Asia, with chronic Opisthorchis viverrini infection being the main risk factor. Epidemiological data on CCA is not available for Lao People Democratic Republic (Lao PDR), though there is a high prevalence of O. viverrini infection in the country. A hospital record-based retrospective study was carried out in six referral hospitals (four in the capital city, one in Savannakhet, one in Pakse) to assess the number of suspected CCA cases admitted between January 2006 and December 2010. In total, we identified 274 patients with liver lesions suggesting CCA, as diagnosed by ultrasound and CT scan. Male gender and age 50years and above were main characteristics of those identified. The majority were admitted to hospitals in the central (58.0%) and southern (29.6%) parts of Lao PDR. Abdominal pain (48.5%) and jaundice (14.2%) were the main symptoms leading to hospitalisation. Some 267 (97.4%) patients had dilated bile ducts; 40 (74.1%) had gallstone or intrahepatic stone, and 33 (12.0%) had a parasitologically confirmed O. viverrini infection. This study provides the first estimated numbers of suspected CCA cases admitted to selected hospitals in Lao PDR. Definitive diagnosis of CCA was not possible. Therefore, case diagnosis and registration of suspected CCA cases are warranted to gather evidence for a national burden estimate of CCA, to develop large-scale studies and adequate preventive and curative health services. A cancer registry would be a most useful tool

Schistosoma mansoni antigen detects Schistosoma mekongi infection

Northern Cambodia and Southern Laos are highly endemic for Schistosoma mekongi. However, there is currently no immunological assay available that is specific for this form of schistosomiasis. We have validated Schistosoma mansoni antigens to detect S. mekongi-directed antibodies in human sera collected from a highly S. mekongi endemic region in Laos. On two consecutive days stool samples of 234 individuals were analyzed by Kato-Katz for presence of S. mekongi eggs and the results were correlated with serology. A sensitivity of 94.5% was calculated for a combination of ELISA and indirect fluorescence assay (IFA) as compared to the detection of S. mekongi eggs in stool samples as gold standard. The results demonstrate that S. mansoni antigens can be used for the diagnosis of S. mekongi infections