Prevalence and Risk Factors of Intestinal Helminthiasis in Remote Mountainous Villages of Northern Lao PDR: A Cross-Sectional Study.

Helminth infections are prevalent in Lao People's Democratic Republic (Lao PDR). This study aimed at determining the prevalence and risk factors of intestinal helminthiasis in remote mountainous villages of northern Lao PDR. During the dry season in January 2017, a cross-sectional survey was conducted in 3 remote mountainous villages in Oudomxay province, Lao PDR. Villagers older than 18 years of age who agreed to submit stool samples or undergo an interview, were recruited. Stool samples from 198 individuals were examined by the Kato-Katz method, and a questionnaire surveyed 161 individuals among them. Univariable and multivariable logistic regression analyses were used to identify risk factors associated with the intestinal helminthiasis. An overall prevalence of intestinal helminthiasis was 75.8%. Hookworm infection was the most common (63.1%), followed by Opisthorchis viverrini/minute intestinal flukes (17.7%), Taenia spp. (15.2%), Trichuris trichiura (2.0%), Ascaris lumbricoides (1.5%), and Enterobius vermicularis (1.0%). Questionnaire analysis revealed sex (male) and absence of latrine to be significant risk factors for hookworm infection and consumption of raw meat for taeniasis. These results suggest that the mountainous area in northern Lao PDR has a different composition of helminth infections from other studies conducted in Lao PDR; a high prevalence of hookworm infection and taeniasis and low prevalence of T. trichiura and A. lumbricoides infections were observed. Also, liver flukes or intestinal flukes were similarly prevalent in the mountainous area.

Patients with severe schistosomiasis mekongi morbidity demonstrating ongoing transmission in Southern Lao People's Democratic Republic.

Chronic infection with Schistosoma mekongi may result in severe hepatosplenic morbidity. We report on eight patients with severe morbidity due to S. mekongi infection. The patients were diagnosed, treated and followed-up between 2007 and 2010 in Khong district, Southern Lao People's Democratic Republic (Lao PDR), eight years after the end of a control intervention. S. mekongi control programmes aimed to prevent morbidity and mortality associated with infection. The patients were visited and interviewed annually. In addition, clinical and abdominal ultrasound examinations were performed and faecal and blood samples were examined. The patients' ages ranged from 6 to 66 years. Of the eight patients, three were children and five were adults. The four youngest patients (aged 6-27 years) significantly improved after praziquantel treatment. One patient (age 46 years) worsened between 2007 and 2010. Two patients died due to bleeding of the oesophageal varices. One patient was lost to follow-up. The leading clinical signs were ascites, splenomegaly, collateral veins on the abdomen and a poor general nutrition status. Ultrasonography disclosed advanced liver fibrosis patterns in all patients; in seven patients, fibrosis pattern E or F was revealed, as per the Niamey protocol (pattern A normal, pattern B to F pathological with increasing severity). Stool microscopy revealed that five patients were co-infected with hookworm and Opisthorchis viverrini. The youngest patient (aged 6 years) was born after the schistosomiasis control program had ended. From her severe morbidity, we can conclude that S. mekongi transmission was on-going in Khong district, and that even in areas with low S. mekongi transmission intensities, severe morbidity from schistosomiasis can develop quickly. Early diagnosis and treatment are imperative, and close monitoring is required.

Efficacy and safety of tribendimidine versus praziquantel against Opisthorchis viverrini in Laos: an open-label, randomised, non-inferiority, phase 2 trial.

BACKGROUND: Praziquantel is the only option for treatment of the liver fluke infection Opisthorchis viverrini. Tribendimidine could be an alternative drug. We aimed to assess the efficacy and safety of a single, oral dose of tribendimidine, compared with praziquantel administered in two doses, in participants with O viverrini infection. METHOD: We did an open-label, randomised, non-inferiority, phase 2 trial in children (8-14 years) and adolescents and adults (≥15 years) in Champasack province, southern Laos. Participants infected with O viverrini were randomly assigned (1:1), via a computer-generated block-randomisation procedure (block sizes of two, four, and six), to receive a single, oral dose of tribendimidine (200 mg for children, 400 mg for adolescents and adults) or two oral doses of praziquantel (50 mg/kg bodyweight and 25 mg/kg bodyweight, 6 h apart). Physicians assessing adverse events and laboratory personnel were masked to treatment allocation, but the investigators administering treatment and the participants could have recognised the treatment group based on differences in the number, appearance, and odour of the tablets. The primary outcomes were cure rate, defined as no parasite eggs in stool at 3 weeks' follow-up, and egg reduction rate. We did available-case analysis of all participants with primary endpoint data. The non-inferiority margin for the difference in cure rates between the groups was pre-specified as -3 percentage points. Adverse events were monitored at 3 h and 24 h after treatment. This trial is registered, number ISRCTN96948551. FINDINGS: Between Feb 1, and April 30, 2014, we assigned 607 participants with confirmed O viverrini infection to receive tribendimidine (n=300) or praziquantel (n=307). 11 participants (five in the tribendimidine group and six in the praziquantel group) did not provide stool samples at 3 weeks' follow-up and were excluded from the available-case analysis. 276 (93·6%) of 295 participants in the tribendimidine group were cured compared with 293 (97·3%) of 301 participants in the praziquantel group. The difference in cure rates between the two groups was -3·8 percentage points (95% CI -7·1 to -0·4), thus the lower limit of the confidence interval exceeded the non-inferiority margin. In both treatment groups, egg reduction rates were 99·9%. Adverse events were of mild and moderate intensity and were more frequent in the praziquantel group than in the tribendimidine group (odds ratio 4·5, 95% CI 3·2-6·3; p<0·0001). The most frequent adverse events were headache, vertigo, nausea, and fatigue. INTERPRETATION: Tribendimidine has a slightly lower cure rate than praziquantel and non-inferiority was not shown. However, tribendimidine has a similar egg reduction rate to praziquantel and leads to fewer adverse events and thus might complement praziquantel in O viverrini control programmes, particularly in settings co-endemic for hookworm. FUNDING: Joint Global Health Trials scheme from the Wellcome Trust, Department for International Development, and Medical Research Council.

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

BACKGROUND: 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. METHODS: 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. RESULTS: 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. CONCLUSIONS: 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. TRAIL REGISTRATION NUMBER: 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.

BACKGROUND: 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. METHODS: 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. RESULTS: 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. CONCLUSIONS: 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.

Efficacy and Safety of Praziquantel Against Light Infections of Opisthorchis viverrini: A Randomized Parallel Single-Blind Dose-Ranging Trial.

Background: The liver fluke Opisthorchis viverrini, highly prevalent in Southeast Asia, is an important public health burden, including a risk factor for developing an aggressive bile duct cancer, cholangiocarcinoma, in chronically infected patients. Praziquantel, administered at a single 40 mg/kg dose in preventive chemotherapy programs and 3 × 25 mg/kg for individual treatment, is the drug of choice, yet information on the nature of the dose-response relationship is lacking. Methods: We performed a randomized, parallel, single-blind dose-ranging phase 2 trial in the Lao People's Democratic Republic in O. viverrini­infected adults. Patients were randomly assigned to 30 mg/kg, 40 mg/kg, 50 mg/kg, or 3 × 25 mg/kg praziquantel or placebo. Adverse events were recorded at baseline, 3 hours, and 24 hours posttreatment. Cure rates (CRs) and egg reduction rates (ERRs) were estimated 3 weeks after drug administration using available case analysis. Dose-response curves were predicted using Emax models. Results: Two-hundred seventeen O. viverrini­infected patients were assigned to the 5 treatment arms. The majority (94.3%) of patients harbored light infections. The Emax model predicted a high efficacy among the observed dose range. We observed CRs ranging from 92.7% to 95.5% and ERRs >99.5% for all praziquantel treatment groups. Adverse events were mild but higher in the standard treatment group (3 × 25 mg/kg) than in the single-dose treatment arms. Conclusions: Single-dose praziquantel appears to be as efficacious as the standard 3 × 25 mg/kg regimen for the treatment of O. viverrini infections, while presenting fewer adverse events. Further studies are necessary in moderate and heavy O. viverrini infections. Clinical Trials Registration: Randomized Controlled Trials (ISRCTN77186750).

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.

Efficacy and safety of tribendimidine against Opisthorchis viverrini: two randomised, parallel-group, single-blind, dose-ranging, phase 2 trials.

BACKGROUND: Treatment of the liver fluke infection Opisthorchis viverrini relies exclusively on praziquantel. Tribendimidine could be an alternative treatment option. We aimed to assess the efficacy and safety of ascending single, oral doses of tribendimidine in patients with O viverrini infection. METHODS: We did two randomised, parallel-group, single-blind, dose-ranging, phase 2 trials in children (aged 8-14 years) and adults and adolescents (≥15 years) in three O viverrini endemic villages in Champasack province, southern Laos. Patients with O viverrini infection were randomly assigned, via a computer-generated central block-randomisation procedure, with block sizes of three (study 1) and four, eight, and 12 (study 2), to receive oral tribendimidine at doses of 200 mg, 400 mg, or 600 mg in a 1:1:1 ratio (adults and adolescents in study 1); 25 mg, 50 mg, 100 mg, or 200 mg (four 50 mg tablets) in a 1:1:1:1 ratio (adults and adolescents in study 2); or 100 mg, 200 mg, or 400 mg in a 1:1:1 ratio (children in study 1). One non-randomised group of children received tribendimidine 50 mg (study 2). Participants, investigators, and laboratory technicians doing the diagnostic assessments were masked to group assignment, but the investigator administering treatment could have recognised the treatment group based on the number of tablets. The primary objective was to estimate the dose-response relation in terms of cure rate and egg reduction rate. We did available-case analysis of all patients with primary endpoint data. We predicted dose-response associations with Emax models. This trial is registered as an International Standard Randomised Controlled Trial, number ISRCTN96948551. FINDINGS: Between Oct 25, 2012, and Nov 5, 2013, 318 adolescents and adults were randomly assigned to seven tribendimidine dose groups: 200 mg (n=51), 400 mg (n=49), or 600 mg (n=47) in study 1, and 25 mg (n=39), 50 mg (n=47), 100 mg (n=44), or 200 mg (four 50 mg tablets; n=41) in study 2. 128 children were randomly assigned to receive tribendimidine 100 mg (n=44), 200 mg (n=40), or 400 mg (n=44) in study 1; 39 children were enrolled and received tribendimidine 50 mg in study 2. In adolescents and adults, the number of patients cured increased with increasing tribendimidine doses up to 100 mg: ten of 39 patients (25·6%, 95% CI 13·0-42·1) were cured in the 25 mg group, 20 of 47 patients (42·6%, 28·3-57·8) were cured in the 50 mg group, and 34 of 44 patients (77·3%, 62·2-88·5) were cured in the 100 mg group; geometric mean egg reduction rates were 86·9% (95% CI 74·8-93·4), 95·9% (92·7-97·7), and 99·1% (98·2-99·7), respectively. The 200 mg dose resulted in cure in 40 of 47 (83·0%, 69·2-92·5) adolescents and adults given the 200 mg tablet and 25 of 41 (61·0%, 95% CI 44·5-75·8) of those given four 50 mg tablets; the 400 mg dose resulted in cure in 43 of 47 patients (91·5%, 79·6-97·6) and the 600 mg dose resulted in cure in 36 of 45 patients (80·0%, 65·4-90·4). Corresponding egg reduction rates were 99·8% (95% CI 99·7-100·0) with one 200 mg tablet, 97·9% (95·9-99·2) with four 50 mg tablets, 99·9% (99·8-100·0) with 400 mg, and 99·8% (99·6-99·9) with 600 mg. The Emax model predicted an egg reduction rate of 99·0% (95% CI 95·7-99·8) at 111 mg in adolescents and adults. 50 mg tribendimidine had moderate efficacy in children, with cure recorded in 16 of 39 patients (41·0%, 95% CI 25·6-57·9). The 100 mg dose resulted in cure in 40 of 44 children (98·9%, 95% CI 78·3-97·5) and an egg reduction rate of 99·7% (95% CI 99·0-100·0), with no increased efficacy at higher doses. The Emax model predicted an egg reduction rate of 99·0% (95% CI 92·2-99·9) at 215 mg. Few adverse events were reported and were mostly mild, with few moderate and no serious events. The most common adverse events 3 h after treatment in adolescents and adults were vertigo (n=35 [11%]), headache (n=9 [3%]), nausea (n=6 [2%]), and fatigue (n=4 [1%]), and in children were headache (n=3 [2%]), vertigo (n=2 [1%]), and fatigue (n=2 [1%]). INTERPRETATION: Tribendimidine has excellent efficacy and tolerability at doses of 100 mg and above. Our study included mainly adults and children with low-intensity O viverrini infection; future studies should assess the efficacy of tribendimidine in patients with infections of moderate and high intensity. FUNDING: Department for International Development, Medical Research Council, Wellcome Trust Joint Global Health Trials Scheme.

Diagnosis of Opisthorchis viverrini Infection with Handheld Microscopy in Lao People's Democratic Republic.

Opisthorchiasis is a neglected tropical disease, yet it is of considerable public health importance in Southeast Asia given the predilection for chronically infected persons to develop cholangiocarcinoma. We evaluated a handheld microscope for the diagnosis of Opisthorchis viverrini in a community-based setting in Lao People's Democratic Republic in comparison with conventional light microscopy. In stool samples collected from 104 individuals, handheld microscopy revealed a sensitivity of 70.6% and a specificity of 89.5% for O. viverrini infection. Pearson's correlation for quantitative fecal egg counts between the two devices was 0.98 (95% confidence interval: 0.98-0.99). With small adjustments to further increase diagnostic sensitivity, a handheld microscope may become a helpful tool to screen for O. viverrini and other helminth infections in public health settings.

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

BACKGROUND: 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. METHODOLOGY: 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. PRINCIPAL FINDINGS: 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. CONCLUSIONS/SIGNIFICANCE: 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.

Subtle to severe hepatobiliary morbidity in Opisthorchis viverrini endemic settings in southern Laos.

Evidence of severe hepatobiliary morbidity associated with Opisthorchis viverrini liver fluke infection including cholangiocarcinoma (CCA) is scarce in Laos although O. viverrini infection is highly prevalent. We assessed hepatobiliary morbidity using abdominal ultrasonography (US) in O. viverrini adult patients in Saravan province, Southern Laos. A random sample of 431 O. viverrini patients from 10 villages underwent abdominal US. Mild, moderate and markedly advanced periductal fibrosis was diagnosed in 7.0%, 66.5%, and 17.0% of patients, respectively. Normal liver parenchyma was seen in only 9.5% of patients. Presence of gall stones (13.2%), sludge (1.4%), gall wall thickening (1.2%), bile duct dilatation (1.6%), fatty liver (12.0%), kidney stones (8.6%) and cysts (7.9%) were diagnosed in considerable frequencies. In five patients (1.2%) hepatobiliary lesions suggesting CCA were diagnosed. Tumour markers, i.e. Interleukin-6, plasminogen activator inhibitor and carbohydrate antigen 19-9 were within normal range. The number of CCA suspected liver masses and hepatobiliary morbidity diagnosed among clinically asymptomatic adult patients in O. viverrini endemic area presents a major public health concern in Laos. However, definitive diagnosis of Opisthorchis-related severe sequelae including CCA is urgently needed to gauge the burden of this deadly disease in Laos.

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.

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.

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 infections and associated risk factors. Only a sound awareness of worm infection and the underlying risk factors may prevent infection and re-infection after treatment.

Efficacy of praziquantel against Schistosoma mekongi and Opisthorchis viverrini: a randomized, single-blinded dose-comparison trial.

BACKGROUND: Schistosomiasis and opisthorchiasis are of public health importance in Southeast Asia. Praziquantel (PZQ) is the drug of choice for morbidity control but few dose comparisons have been made. METHODOLOGY: Ninety-three schoolchildren were enrolled in an area of Lao PDR where Schistosoma mekongi and Opisthorchis viverrini coexist for a PZQ dose-comparison trial. Prevalence and intensity of infections were determined by a rigorous diagnostic effort (3 stool specimens, each examined with triplicate Kato-Katz) before and 28-30 days after treatment. Ninety children with full baseline data were randomized to receive PZQ: the 40 mg/kg standard single dose (n = 45) or a 75 mg/kg total dose (50 mg/kg+25 mg/kg, 4 hours apart; n = 45). Adverse events were assessed at 3 and 24 hours posttreatment. PRINCIPAL FINDINGS: Baseline infection prevalence of S. mekongi and O. viverrini were 87.8% and 98.9%, respectively. S. mekongi cure rates were 75.0% (95% confidence interval (CI): 56.6-88.5%) and 80.8% (95% CI: 60.6-93.4%) for 40 mg/kg and 75 mg/kg PZQ, respectively (P = 0.60). O. viverrini cure rates were significantly different at 71.4% (95% CI: 53.4-84.4%) and 96.6% (95% CI: not defined), respectively (P = 0.009). Egg reduction rates (ERRs) against O. viverrini were very high for both doses (>99%), but slightly lower for S. mekongi at 40 mg/kg (96.4% vs. 98.1%) and not influenced by increasing diagnostic effort. O. viverrini cure rates would have been overestimated and no statistical difference between doses found if efficacy was based on a minimum sampling effort (single Kato-Katz before and after treatment). Adverse events were common (96%), mainly mild with no significant differences between the two treatment groups. CONCLUSIONS/SIGNIFICANCE: Cure rate from the 75 mg/kg PZQ dose was more efficacious than 40 mg/kg against O. viverrini but not against S. mekongi infections, while ERRs were similar for both doses. TRIAL REGISTRATION: Controlled-Trials.com ISRCTN57714676.

Spatial distribution of, and risk factors for, Opisthorchis viverrini infection in southern Lao PDR.

BACKGROUND: Opisthorchis viverrini is a food-borne trematode species that might give rise to biliary diseases and the fatal cholangiocarcinoma. In Lao PDR, an estimated 2.5 million individuals are infected with O. viverrini, but epidemiological studies are scarce and the spatial distribution of infection remains to be determined. Our aim was to map the distribution of O. viverrini in southern Lao PDR, identify underlying risk factors, and predict the prevalence of O. viverrini at non-surveyed locations. METHODOLOGY: A cross-sectional parasitological and questionnaire survey was carried out in 51 villages in Champasack province in the first half of 2007. Data on demography, socioeconomic status, water supply, sanitation, and behavior were combined with remotely sensed environmental data and fed into a geographical information system. Bayesian geostatistical models were employed to identify risk factors and to investigate the spatial pattern of O. viverrini infection. Bayesian kriging was utilized to predict infection risk at non-surveyed locations. PRINCIPAL FINDINGS: The prevalence of O. viverrini among 3,371 study participants with complete data records was 61.1%. Geostatistical models identified age, Lao Loum ethnic group, educational attainment, occupation (i.e., rice farmer, fisherman, and animal breeder), and unsafe drinking water source as risk factors for infection. History of praziquantel treatment, access to sanitation, and distance to freshwater bodies were found to be protective factors. Spatial patterns of O. viverrini were mainly governed by environmental factors with predictive modeling identifying two different risk profiles: low risk of O. viverrini in the mountains and high risk in the Mekong corridor. CONCLUSIONS/SIGNIFICANCE: We present the first risk map of O. viverrini infection in Champasack province, which is important for spatial targeting of control efforts. Infection with O. viverrini appears to be strongly associated with exposure to the second intermediate host fish, human behavior and culture, whereas high transmission is sustained by the lack of sanitation.

Discovery of Opisthorchis lobatus (Trematoda: Opisthorchiidae): a new record of small liver flukes in the Greater Mekong Sub-region.

Metacercariae, morphologically similar to those of small liver flukes, were found to parasitize red-tailed snakehead fish, Channa limbata, collected from the city of Vientiane, Lao People's Democratic Republic. Adult worms that were recovered from experimentally-infected hamsters showed characteristics distinctly different from Opisthorchis viverrini, but closely similar to Opisthorchis lobatus, which was first reported in poultry (Anas sp.) from Pakistan. The present study aimed to redescribe O. lobatus based on the adult worms recovered from experimentally-infected hamsters. Additionally, it aimed to document the genetic relationships among O. lobatus and other opisthorchiid liver flukes using the mitochondrial cytochrome c oxidase subunit 1 (COI) gene and the internal transcribed spacer 2 (ITS2) region. DNA alignment of the O. lobatus and O. viverrini COI partial sequences (330 bp) showed 3.03% fixed differences (2.72% of amino acids changed) while the ITS2 region (350 bp) indicated a 0.86% difference for nucleotides. Species boundaries between the 2 parasites were determined by neighbor-joining analysis using the molecular sequence data. The phenogram confirmed that O. lobatus was distinctly different from O. viverrini, representing the first reported instance of O. lobatus in the Greater Mekong Sub-region (GMS) and the first record of C. limbata as the second intermediate host of a small liver fluke. Questions regarding human infection and the extent of the geographic distribution of these species should be investigated further.

Efficacy and safety of mefloquine, artesunate, mefloquine-artesunate, tribendimidine, and praziquantel in patients with Opisthorchis viverrini: a randomised, exploratory, open-label, phase 2 trial.

BACKGROUND: Praziquantel is the only drug available for treatment of Opisthorchis viverrini, although in-vivo studies point to activity of mefloquine, artesunate, and tribendimidine against this liver fluke. We aimed to assess the efficacy and safety of these drugs compared with that of praziquantel in patients with O viverrini infection. METHODS: We did a randomised open-label trial between February and April, 2010, in the Saysetha district, Attapeu Province, Laos. Eligible patients were school children aged 10-15 years who had O viverrini infections. Patients were randomly assigned to one of five different treatment groups by use of a computer-generated randomisation code. We assessed efficacy as cure rate and egg reduction rate in intention-to-treat and per-protocol analyses. The trial was registered with Current Controlled Trials, ISRCTN23425032. RESULTS: 125 children were randomly assigned: 25 received mefloquine, 24 artesunate, 24 mefloquine-artesunate, 27 tribendimidine, and 25 praziquantel. 19 patients were lost to follow-up. In the intention to treat analysis, 14 patients receiving praziquantel were cured compared with none with mefloquine, one with artesunate (odds ratio 0·03, 95% CI 0·004-0·29), one with mefloquine-artesunate (0·03, 0·004-0·29), and 19 with tribendimidine (1·87, 0·60-5·85). Egg reduction rate was 98·4% for praziquantel, 30·2% for mefloquine (egg reduction-rate ratio 1·61, 95% CI 0·21-0·72), 31·5% for artesunate (0·43, 0·23-0·80), 41·3% for mefloquine-artesunate (0·60, 0·31-1·10), and 99·3% for tribendimidine (1·00, 0·44-2·30). Most adverse events were mild or moderate and affected all treatment groups; serious adverse events--vertigo, nausea, vomiting, and anxiety--were reported only by patients taking mefloquine or mefloquine-artesunate. INTERPRETATION: Tribendimidine seems to be at least as efficacious as the drug of choice, praziquantel, for the treatment of O viverrini infections; both drugs were well tolerated. Mefloquine, artesunate, and mefloquine-artesunate did not show an effect. Tribendimidine should be further investigated with large clinical trials. FUNDING: Swiss National Science Foundation, University of Basel.

Sero-epidemiological survey of gnathostomiasis in Lao PDR.

While human gnathostomiasis cases have been reported sporadically in Lao PDR since 1975, little is known about the disease in this country. We aimed to investigate sero-prevalence of gnathostomiasis and Gnathostoma species in Lao PDR. One village each in the north, central and south regions of Lao PDR was selected as the study sites. Overall, 125 (29.8%) of 420 sera from the randomly selected participants were sero-positive by immunoblot technique, with anti-Gnathostoma IgG antibody against the 24 kDa fraction. The sero-prevalence was high in the central (47.1%) and south (38.6%), but low (3.6%) in the north. Risk factor analyses revealed that the consumption of raw/undercooked fish was significantly associated with Gnathostoma sero-positivity (95% CI 1.05-17.05, P=0.042). The sero-positivity significantly increased with the age of the participants. Several fish, swamp eels, and frogs collected from central and southern Lao were infected with G. spinigerum advanced 3rd-stage larvae. Channa limbata (red-tailed snakehead fish) was identified as a natural second intermediate host of G. spinigerum. Eggs of G. spinigerum were found in dog feces collected in the south. Gnathostomiasis is endemic in central and southern Laos, so that preventive measures should be introduced for people living in these regions.

Monophyly of Opisthorchis viverrini populations in the lower Mekong Basin, using mitochondrial DNA nad1 gene as the marker.

The liver fluke, Opisthorchis viverrini, causes serious public-health problems in the Lower Mekong Basin. This study aimed to clarify whether O. viverrini populations may be genetically divided into sub-specific taxa. We collected 6 populations of O. viverrini from different places in Cambodia, Lao PDR, and Thailand, along both sides of the Mekong River, and analyzed the population structure of these using the mitochondrial nad1 gene as a marker. The results of the DNA polymorphism measurements, by theta-w (thetaw) and -pi (thetapi) values, neutrality tests, and mismatch distribution, suggested that the population of O. viverrini has expanded under the influence of purifying selection and selective sweep. The analysis of molecular variance (AMOVA) test revealed no significant genetic differences among the O. viverrini populations on opposite sides of the Mekong River. O. viverrini haplotypes occurred in multiple populations, and no distinct geographical clade. The star-like haplotype network confirmed a demographic expansion of the O. viverrini population. Overall, the genetic data from these populations suggested that the postulated existence of an O. viverrini species complex should be rejected. The bio-geographical diversity of O. viverrini populations should be explored further, using other appropriate markers and a wider range of samples from geographically different areas.