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The presence and distribution of various species of canine hookworms in Africa are poorly known. The main objective of this study, therefore, was to identify the hookworm species present in canine faecal samples from Morogoro, Tanzania, using molecular techniques. Faecal samples from 160 local dogs were collected and hookworm positive samples processed to recover larvae for further molecular characterization. DNA was extracted from pools of larvae from individual samples (n = 66), which were analysed subsequently using two different molecular approaches, polymerase chain reaction-linked restriction fragment length polymorphism (PCR-RFLP) and species-specific PCR coupled with Sanger sequencing. The PCR-RFLP technique detected only the presence of the ubiquitous Ancylostoma caninum in the 66 samples. However, by species-specific PCR coupled with Sanger sequencing we identified ten samples with A. braziliense, two with Uncinaria stenocephala and five with A. ceylanicum. Thus, all four known species of canine hookworms were identified in Morogoro, Tanzania. To our knowledge this is the first report of the detection of the presence of U. stenocephala and A. ceylanicum in Africa using molecular techniques. In addition to their veterinary importance, canine hookworms have zoonotic potential and are of public health concern.

This study evaluated the effect of mass drug administration (MDA) with praziquantel administered to school-aged children (SAC) combined with 'track and treat' of taeniosis cases in the general population on the copro-antigen (Ag) prevalence of taeniosis. The study was conducted in 14 villages in Mbozi and Mbeya district, Tanzania. SAC made up 34% of the population and received MDA with praziquantel (40mg/kg) in 2012 (both districts) and in 2013 (Mbozi only). Three cross-sectional population-based surveys were performed in 2012 (R0), 2013 (R1), and 2014 (R2). In each survey approximately 3000 study subjects of all ages were tested for taeniosis using copro-Ag-ELISA. In total 9064 people were tested and copro-Ag-ELISA positive cases were offered treatment 6-8 months after sampling. The copro-Ag prevalence of taeniosis was significantly higher (Χ 2 -test, p=0.007) in Mbozi (3.0%) at R0 compared to Mbeya (1.5%). Twelve months after MDA in both districts (R1), the copro-Ag prevalence had dropped significantly in both Mbozi (2.0%, p=0.024) and in Mbeya (0.3%, p=0.004), but the significant difference between the districts persisted (Χ 2 -test, p<0.001). Ten months after the second round of MDA in Mbozi and 22 month after the first MDA (R2), the copro-Ag prevalence had dropped significantly again in Mbozi (0.8%, p<0.001), but had slightly increased in Mbeya (0.5%, p=0.051), with no difference between the two districts (Χ 2 -test, p=0.51). The taeniosis cases tracked and treated between round R0 and R2 represented 9% of the projected total number of taeniosis cases within the study area, based on the copro-Ag prevalence and village population data. Among SAC in Mbozi, infection significantly decreased at R1 (p=0.004, OR 0.12, CI: 0.02-0.41) and R2 (p=0.001, OR 0.24, CI: 0.09-0.53) when comparing to R0. In Mbeya infection significant decreased at R1 (p=0.013, OR 0.14, CI: 0.02-0.55), but no difference was found for R2 (p=0. 089), when comparing to R0 among SAC. This study showed that school-based MDA with praziquantel in combination with 'track and treat' of taeniosis cases significantly reduced the copro-Ag prevalence of taeniosis, and that annual MDA was significantly better than single MDA. The persistence of taeniosis cases illustrates that a One Health approach must be emphasized for effective control., (Copyright © 2015 Elsevier B.V. All rights reserved.)

Background: Fish-borne zoonotic trematodes (FZT) infections including liver- and minute intestinal flukes are common in Southeast Asia in both humans and domestic animals eating raw fish and since 2010, the liver flukes are recognised as neglected tropical diseases by WHO. Mass drug treatment with praziquantel is advised for humans, but no recommendations for control of the FZT in the reservoir hosts exist., Methodology/principal Findings: A study was conducted to assess the ability of praziquantel treatment for control of FZT in farm dogs in an endemic area in Northern Vietnam. Initially, 101 dogs from 73 households were examined for small trematode eggs in their faeces. Forty seven copro-positive dogs were included in the study. Thirty eight dogs received treatment with a single dose of 40 mg/kg of praziquantel. A group of nine dogs were left untreated. Coprological examination for small trematode eggs was performed on day 0, 3, 10, 30, 60, 90 and 120 post treatment. Farmers were questioned about dog keeping practises. All dogs were copro-negative for small trematode eggs on both day 3 and 10 post treatment. From day 30 onwards previously negative dogs became positive again. The reinfection rates were 26.3% (day 30), 45.5% (day 60), 53.1% (day 90), 61.3% (day 120).The nine untreated dogs remained positive throughout the study period. There was no difference in the intensity of infection at day 0 and 120 neither in the treated or untreated group., Conclusion: Dogs had easy access to raw fish and did not receive treatment against flukes by their owner. More than 50% of the dogs were reinfected 3 months post treatment. We do not recommend controlling FZT infections in dogs by anthelmintic treatment alone since reinfection occurs fast under the existing farm management systems.

Copro-antigen detection has been advocated as a promising method for diagnosis of opisthorchiasis, particularly in people that harbored light infection or have had recent drug treatment. This study aimed to improve performance of a monoclonal antibody-based enzyme-linked immunosorbent assay (Mab-ELISA) for detection of Opisthorchis viverrini copro-antigen and assess the correlation between copro-antigen and intensity of infection. Four different treatment methods of 71 samples from the Lawa endemic area, Khon Kaen province were assessed for copro-antigen detection, namely (1) phosphate buffer saline (PBS), (2) heating (70°C 30min), (3) alkaline (pH 9.6 in carbonate buffer), and (4) trichloroacetic acid (TCA) treatment. Comparison of these protocols showed that the TCA method gave the best performance in discriminating O. viverrini positive and negative samples with high sensitivity (97.9%) and moderate specificity (54.2%) compared with other methods. Application of TCA-based Mab-ELISA method for antigen detection in fecal samples obtained from an endemic area of opisthorchiasis revealed that 86 of 141 samples (61.0%) were positive compared with 68 (48.2%) by PBS-based protocol, while the formalin ethyl-acetate concentration technique (FECT) yielded a positive proportion of 71.6%. Among 40 egg-negative samples confirmed by a gold standard parasitological method (FECT) from the same endemic area, 19 (47.5%) were positive by the TCA-based while only 6 (15%) were positive by PBS-based Mab-ELISA protocol. In addition, levels of antigen detection significantly correlated with intensity of infection (egg per gram feces). The results show that the improved Mab-ELISA method has high sensitivity and also quantifiable diagnosis of opisthorchiasis., (Copyright © 2013 Elsevier B.V. All rights reserved.)

The study was conducted to evaluate copro-diagnostic techniques for detection of small trematode eggs in dogs. FLOTAC, a novel flotation technique, and DBL, a sieving and sedimentation technique developed at the former Danish Bilharziasis Laboratory (DBL), were compared using 53 subsamples from four copro-positive dogs. Moreover, a modified version of the DBL technique and the Kato-Katz (KK) thick smear were later compared using faecal samples from 21 dogs. The four techniques were pair-wise compared regarding sensitivity, infection intensity and practical applicability. For the former two techniques, egg recovery subsequent to storage and reproducibility were also compared. The DBL technique detected all 53 subsamples positive for small trematode eggs. Based on 17 subsamples, mean infection intensity of 47±49 eggs per gram of faeces (EPG) was detected by the technique. Due to large amount of sediment, examination of a single subsample required an average of 3 hours. The FLOTAC technique was found less sensitive (82%) than the DBL technique and recovered significantly less eggs (4±6 EPG). Both sensitivity and intensity were further reduced following storage. As the FLOTAC technique requires specialised equipment, safety disposal and personal protective equipment, it was found less suited than the DBL technique for a basic laboratory. Additionally, poor reproducibilities were found for both the DBL and FLOTAC techniques (30±15% and 38±33%, respectively). Based on the 21 faecal samples, the modified version of the DBL technique was found more sensitive (85%) than the KK technique (68%), whereas egg counts were significantly higher for the latter (23±26 EPG vs. 482±909 EPG). By modifying the DBL technique, it was possible to diminish the retained sediment and examination time to a maximum of an hour, which was also the time required by the KK technique, although the latter was faster and more easily processed. Based on the results obtained in this study, none of the techniques evaluated were found applicable in their current form for detection of small trematode eggs in faeces from dogs in Vietnam., (Copyright © 2013 Elsevier B.V. All rights reserved.)

Opisthorchis viverrini is an important food-borne trematode in Southeast Asia. The infection causes significant morbidity in terms of hepatobiliary diseases and cholangiocarcinoma. The aim of this study was to improve the sensitivity of the PCR-based diagnosis of O. viverrini infection. A new fecal DNA extraction protocol for the detection of O. viverrini DNA using cetyltrimethyl-ammoniumbromide to remove PCR inhibitor was used and compared with the commercial stool kit method. The sensitivity of the new test was 79.3%, compared with the 44.8% of the previous method (P < 0.01). PCR-positive tests identified several cases judged parasite negative by the parasitological method (28.6%), indicating the new test's advantage in the diagnosis of individuals with light infections.

The present study sought to elucidate whether Schistosoma japonicum eggs could survive a passage through dogs and chickens. Three dogs and three chickens were used for the experiment. The dogs were allowed to eat goat faeces containing 80 S. japonicum eggs per gram. The chickens were given 2 ml of water containing more than 100 S. japonicum eggs per ml. The hatchability of the eggs was 51.8% prior to administration. All faeces were collected for 3 days from the dogs and for 2 days from the chickens and egg excretion and hatchability were measured. The results showed that nearly 50% of the eggs administered to the dogs were recovered in the faeces during the first 2 days and during the first 25 h 39% of the recovered eggs were viable and able to hatch. In the chickens 9% of the administered eggs were recovered within 27 h and within the first 3.5 h after administration 19% of the recovered eggs were viable and able to hatch. The results suggest that both dogs and chickens may act as transport hosts for S. japonicum in endemic areas.

Chemotherapy has been used on a large scale in countries where the blood fluke Schistosoma japonicum is endemic. This has led to a lower intensity of infections and consequently lower diagnostic values of commonly used diagnostic tests like serology and Kato-Katz stool smear. We designed a novel real-time PCR method for detection of S. japonicum in stool samples. Further, we evaluated different versions of an inexpensive, non-commercial extraction method, ROSE, as well as the commercial QIAamp DNA Stool Mini Kit. PCR primer sequences were designed targeting the mitochondrial NADH dehydrogenase I gene. Bovine serum albumin was added to the DNA extracts and SYBR Green was used for detection. The PCR method was evaluated with non-infected stool samples spiked with S. japonicum eggs. It demonstrated high sensitivity, even in samples containing a single egg. The two extraction methods were equally effective. The PCR was specific for S. japonicum when tested against other Schistosoma species, Trichuris trichiura, hookworm and Taenia sp. We conclude that this novel real-time PCR, in combination with either ROSE or QIAamp DNA Stool Mini Kit extraction, is a sensitive and specific tool for diagnosing S. japonicum in human stool samples.

In this study, a recently developed PCR test for the detection of Opisthorchis viverrini in human faecal samples was evaluated using two parasitological methods as references. During a survey of foodborne trematodes (FBT) in the Vientiane Province, Lao PDR, 85 samples were collected and evaluated for FBT eggs by the Kato Katz (KK) technique, the formalin ethyl acetate concentration technique (FECT) and a PCR analysis for the distinction between O. viverrini and other FBT. The two parasitological methods did not differ in the ability of detecting FBT eggs, and a single KK reading was characterized by a sensitivity of 85% when compared to two FECT readings. The PCR tested positive only in cases where eggs had been demonstrated by parasitological examination. However, the PCR tested negative in some samples with very high egg counts. Demonstrating a PCR sensitivity of approximately 50% in samples with faecal egg counts>1000, the previously reported PCR sensitivity based on in vitro studies was not supported. It is believed that technical problems rather than diagnostic reference related issues were responsible for the relatively low PCR performance. Further studies should aim at optimizing DNA extraction and amplification, and future PCR evaluation should include specificity control such as the scanning electron microscopy of eggs in test samples or the expulsion of adult trematodes from PCR tested individuals.

An improved laboratory method was developed for counting Schistosoma japonicum eggs in pig feces, which involves filtration, sedimentation and centrifugation, but avoids toxic chemicals. It is sensitive, allows easy differentiation from similar-sized and shaped protozoan cysts, and permits evaluation of egg viability both by direct viewing of eggs and miracidial hatching. It was found to be significantly better at recovering eggs than the modified Bell filtration technic. The sensitivity, specificity and practicality of this technic make it our method of choice for studies on porcine schistosomiasis japonica.

Sixteen non-infected control pigs housed together with 16 pigs infected with Trichuris suis, excreted T. suis eggs in their faeces (range 20-4, 960 eggs per gram faeces (EPG)). When the control pigs were moved to clean pens, their egg counts dropped to zero EPG within five days. A significant correlation was found between T. suis egg counts of infected and control pigs penned together (r = 0.89, P < 0.001). These results suggest that false-positive faecal egg counts may be the result of coprophagia.