Working Regime Criteria for Microscale Electrohydrodynamic Conduction Pumps.

We investigated the microscale electrohydrodynamic (EHD) conduction pumps in a wide range of working regimes, from the saturation regime to the ohmic regime. We showed that the existing macro- and microscale theoretical models could not accurately predict the electric force of microscale EHD conduction pumps, especially for the cases of a strong diffusion effect. We clarified that the failure is caused by a rough estimate of the heterocharge layer thickness. We revised the expression of heterocharge layer thickness by considering the diffusion effect and developed a new theoretical model for the microscale EHD conduction pumps based on the revised expression of heterocharge layer thickness. The results showed that our model can accurately predict the dimensionless electric force of the microscale EHD conduction pumps even for the cases of a strong diffusion effect. Furthermore, we developed a working regime map of microscale EHD conduction pumps and found that the microscale EHD conduction pumps more easily fall into the saturation regime compared with the macroscale EHD conduction pumps due to the enhanced diffusion effect; in other words, the microscale EHD conduction pumps have a wider saturation regime. We showed that the conduction number C0 could not distinguish the working regime of the microscale EHD conduction pumps because it does not take the diffusion effect into account. By employing the revised expression of heterocharge layer thickness, we proposed a new dimensionless number, C0D to distinguish the working regimes of microscale EHD conduction pumps.

Small-scale spatial analysis of intermediate and definitive hosts of Angiostrongylus cantonensis.

BACKGROUND: Angiostrongyliasis is a food-borne parasitic zoonosis. Human infection is caused by infection with the third-stage larvae of Angiostrongylus cantonensis. The life cycle of A. cantonensis involves rodents as definitive hosts and molluscs as intermediate hosts. This study aims to investigate on the infection status and characteristics of spatial distribution of these hosts, which are key components in the strategy for the prevention and control of angiostrongyliasis. METHODS: Three villages from Nanao Island, Guangdong Province, China, were chosen as study area by stratified random sampling. The density and natural infection of Pomacea canaliculata and various rat species were surveyed every three months from December 2015 to September 2016, with spatial correlations of the positive P. canaliculata and the infection rates analysed by ArcGIS, scan statistics, ordinary least squares (OLS) and geographically weighted regression (GWR) models. RESULTS: A total of 2192 P. canaliculata specimens were collected from the field, of which 1190 were randomly chosen to be examined for third-stage larvae of A. cantonensis. Seventy-two Angiostrongylus-infected snails were found, which represents a larval infection rate of 6.1% (72/1190). In total, 110 rats including 85 Rattus norvegicus, 10 R. flavipectus, one R. losea and 14 Suncus murinus were captured, and 32 individuals were positive (for adult worms), representing an infection rate of 29.1% of the definitive hosts (32/110). Worms were only found in R. norvegicus and R. flavipectus, representing a prevalence of 36.5% (31/85) and 10% (1/10), respectively in these species, but none in R. losea and S. murinus, despite testing as many as 32 of the latter species. Statistically, spatial correlation and spatial clusters in the spatial distribution of positive P. canaliculata and positive rats existed. Most of the spatial variability of the host infection rates came from spatial autocorrelation. Nine spatial clusters with respect to positive P. canaliculata were identified, but only two correlated to infection rates. The results show that corrected Akaike information criterion, R2, R2 adjusted and σ2 in the GWR model were superior to those in the OLS model. CONCLUSIONS: P. canaliculata and rats were widely distributed in Nanao Island and positive infection has also been found in the hosts, demonstrating that there was a risk of angiostrongyliasis in this region of China. The distribution of positive P. canaliculata and rats exhibited spatial correlation, and the GWR model had advantage over the OLS model in the spatial analysis of hosts of A. cantonensis.

[Species composition and distribution of medical mollusca in Shanghai City].

OBJECTIVE: To investigate the species diversity and distribution of medical mollusca in Shanghai City. METHODS: From August 2012 to October 2013, all kinds of habitats in 8 districts and counties in Shanghai City, namely Jiading, Qingpu, Baoshan, Minhang, Songjiang, Jinshan, Chongming, Pudong, were selected for the field survey according to the distribution characteristics of the river system, and all the specimens of medical mollusca in the investigation sites were collected and classified by morphological identification. Meanwhile, the species composition, habitats as well as the fauna of the medical mollusca collected were analyzed. RESULTS: A total of 5,211 specimens were collected, which belonged to 2 classes, 14 families, 18 genera and 25 species, including Oncomelania hupensis hupensis, Pomacea canaliculata, Parafossarulus striatulus, Alocinma longicornis, Physa acuta, Galba pervia, Hippeutis cantori, etc. The species numbers of medical mollusca in Chongming, Jinshan, Pudong new area and Qingpu districts (counties) were 22, 22, 21 and 20, respectively, which were more than those of other areas. The habitat analysis suggested that the species numbers in the river and wetland were the most, both of which were 14 species. The main faunas of the medical mollusca in Shanghai were the cosmopolitan and oriental species. CONCLUSIONS: The freshwater gastropod species are paucity in Shanghai City, but almost of them can be served as the intermediate hosts of certain parasites to transmit snail-related parasitic diseases, so the surveillance of medical mollusca should be strengthened.

[Comparative analysis of mitochondrial genomes of Angiostrongylus cantonensis].

OBJECTIVE: To compare the diversity of mitochondrial genomes of Angiostrongylus cantonensis in the mainland of China. METHODS: According to the population genetic of A. cantonensis, seven female worms were selected to characterize the mitochondrial (MT) genomes. Twelve primer pairs based on known MT genome (GQ398121) were used for PCR. The target fragments were sequenced and aligned. The gene localization, genome structure, composition of nucleotide, distribution of variable sites, and phylogeny were analyzed by employing multiple softwares. RESULTS: Five distinct types were identified from seven complete MT genomes. They were similar in size and structure, i.e., ranging 13,491-13,502 bp, including 12 protein-coding genes, 2 ribosomal genes, 22 tRNA genes, and 2 major non-coding regions. All the genes were localized at the same strand and had the same transcription direction. A total of 745 variable sites were identified, accounting for 5.5%. Among the variable sites, 59 were deletion/insert mutations, 105 transversions, and 581 transitions. The variable sites distributed evenly at the complete genome. CONCLUSION: The study reveals the mutation profile in the whole MT genome of A. cantonensis and thus will facilitate the development of intraspecific differential diagnosis.

[Investigation on the species distribution and infection status of host snails of Angiostrongylus cantonensis in Shanghai].

OBJECTIVE: To investigate the species distribution and infection status of host snails of Angiostongylus cantonensis in Shanghai. METHODS: From August 2012 to October 2014, 10 districts or counties were selected for this investigation in Shanghai, including Jiading, Qingpu, Putuo, Baoshan, Yangpu, Minhang, Songjiang, Jinshan, Chongming, and Pudong. All of the snails were examined by lung-microscopy and tissue homogenate methods. RESULTS: Totally 1074 snail samples were identified, belonging to 8 families, 10 species, including Pomacea canaliculata (133), Achatina fulica (25), Cipangopaludina chinensis (183), Bellamya aeruginosa (78), Physa acuta (349), Radix swinhoei (224), Bradybaena similaris (45), B. ravida sieboldiana (32), Limax flavus (2), and Philomycus bilineatus (3). A. fulica was sold in market, and was not found in the field. The natural population of P. canaliculata was found in the field in Jinze Town of Qingpu District. A. cantonensis larvae were not detected. CONCLUSION: No snails were found to be infected with A. cantonensis in Shanghai. Attention needs to be paid to the monitoring of P. canaliculata.

The complete mitochondrial genome of the rodent intra-arterial nematodes Angiostrongylus cantonensis and Angiostrongylus costaricensis.

The two rodent intra-arterial nematodes, Angiostrongylus cantonensis and Angiostrongylus costaricensis, can cause human ill-health. The present study aimed to characterize and compare the mitochondrial (mt) genomes of these two species, and clarify their phylogenetic relationship and the position in the phylum Nematoda. The complete mt genomes of A. cantonensis and A. costaricensis are 13,497 and 13,585 bp in length, respectively. Hence, they are the smallest in the class of Chromadorea characterized thus far. Like many nematode species in the class of Chromadorea, they encode 12 proteins, 22 transfer RNAs, and two ribosomal RNAs. All genes are located on the same strand. Nucleotide identity of the two mt genomes is 81.6%, ranging from 77.7% to 87.1% in individual gene pairs. Our mt genome-wide analysis identified three major gene arrangement patterns (II-1, II-2, and II-3) from 48 nematode mt genomes. Both patterns II-1 and II-2 are distinct from pattern II-3, which covers the Spirurida, supporting a closer relationship between Ascaridida and Strongylida rather than Spirurida. Thymine (T) was highly concentrated on coding strands in Chromadorea, but balanced between the two strands in Enoplea, probably due to the gene arrangement pattern. Interestingly, the gene arrangement pattern of mt genomes and phylogenetic analysis based on concatenated amino acids indicated a closer relationship between the order Ascaridida and Rhabditida rather than Spirurida as indicated in previous studies. These discrepancies call for new research, reassessing the position of the order of Ascaridida in the phylogenetic tree. Once consolidated, the findings are important for population genetic studies and target identification.

[Population genetics of Pomacea spp. in mainland of China].

OBJECTIVE: To reveal the population structure of Pomacea spp. using genetic markers so as to provide the evidence for studying the invasion and expansion of it in the mainland of China. METHODS: The genetics of 581 specimens of Pomacea spp. from 60 sites was analyzed by sequencing CO I gene. The diversity of nucleotide and haplotypes were calculated in DnaSP 5.10.01. The haplotype network analysis was performed in Network 4.2.0.1. A phylogenetic tree was produced based on the haplotypes from the present study and those available from GenBank in order to understand the taxonomic status of Pomacea spp. in China. RESULTS: A total of 556 sequences were acquired in the present study and produced 25 unique haplotypes. Six haplotypes frequently occurred in the specimens and accounted for 96.0%. The phylogenetic analysis identified two Pomacea species in China, i. e. P. canaliculata and P. insularum. The usage of haplotypes of P. insularum in China reversed from the existing pattern in other countries. CONCLUSION: The complexity of population structure of Pomacea spp. in the mainland of China indicates multi-original introduction and complicated expansion patterns.

[Survey on the focus of Angiostrongylus cantonensis in Guangdong Province].

OBJECTIVE: To understand the distribution of Angiostrongylus cantonensis foci in Guangdong Province for making surveillance program. METHODS: Survey sites were chosen by strata sampling according to different geographic locations. Totally 22 survey sites were selected in four regions: East Guangdong, West Guangdong, North Guangdong and the Pearl River Delta. One or two administrative villages in each site were randomly selected for the investigation. Pomacea canaliculata and Achatina fulica collected from fields and other species of freshwater or terrestrial snails obtained in the restaurants and wet markets were examined for the third stage larvae by tissue grinding or lung examination. Rats were captured in the fields, and their hearts and lungs were dissected for adult worms. Rat feces were also collected for the detection of first stage larvae by water precipitation. RESULTS: Large number of P. canaliculata was found in all sites. A. fulica was found in most surveyed sites. Totally 2929 P. canaliculata and 1354 A. fulica were collected with a larva infection rate of 5.9% (172/2929) and 16.5% (223/1354), respectively (P<0.01). The average prevalence among the regions was different (P<0.01) with the highest prevalence in Pearl River Delta (15.6%, 152/975), especially in Dongguan City of the Delta (34.7%, 78/225). 114 Cipangopaludina sp. and 252 Bellamya sp. were bought from wet markets of 9. sites. Larvae were found only in Bellamya snails from Luoding and Kaiping cities with an infection rate of 1.4% (1/70) and 3.3% (3/91), respectively. Totally 491 rats were captured in 9 sites including Rattus norvegicus, R. flavipectus, Suncus murinus, Mus musculus, Bandicota indica, R. losea and R. rattus, with an average infection rate of 11.4% (56/491). Adult worms were found in R. norvegicus, R. flavipectus and B. indica with a prevalence of 19.8% (52/263), 2.5% (3/118) and 10.0% (1/10), respectively. Thirty-four rodent fecal samples were collected in 7 sites and examined with a larva positive rate of 44.1% (15/34). CONCLUSION: Foci of Angiostrongylus cantonensis are widely distributed in Guangdong Province as natural infection has been found in its intermediate and definitive hosts.

[Multiplex PCR assay for the detection of Angiostrongylus cantonensis larvae in Pomacea canaliculata].

OBJECTIVE: To establish a multiplex PCR assay for detecting Angiostrongylus cantonensis larvae in Pomacea canaliculata. METHODS: A pair of specific primers was designed based on the sequences of the small subunit rDNA of A. cantonensis (GenBank jAY295804), in combination with 16s rDNA specific primers of P. canaliculata, a multiplex PCR was developed. The PCR was performed on positive and negative snails, and the amplified products were analyzed by agarose gel electrophoresis and DNA sequencing. DNA template of 200 III stage larvae of A. cantonensis was diluted by negative snail DNA (1200 ng/microl, 120 ng/microl, 12 ng/microl, 1200 pg/microl, 120 pg/microl and 12 pg/microl), to find the minimum detectable level. Single blind method was used to evaluate the accuracy. After being detected by lung microscopy, 172 snails from field were tested by the multiplex PCR to assess the sensitivity and specificity. RESULTS: Agarose gel electrophoresis and DNA sequencing analysis indicated that the target sequences were efficiently amplified by the PCR assay (550 bp for P. canaliculata, 405 bp for A. cantonensis). The minimum detectable level was 120 pg/microl. The coincidence between the two methods stood for 84.3% (145/172), including 45 positives and 100 negatives. 24 snails were PCR positive and microscopy negative, 3 snails were PCR negative and microscopy positive. The sensitivity and specificity of multiplex PCR was 93.8% and 80.6%, respectively. Its positive rate (40.1%, 69/172) was significant higher than that of lung-microscopy (27.9%, 48/172)(chi2 = 14.8, P < 0.01). CONCLUSION: A multiplex PCR method has been developed for the detection of A. cantonensis larvae in P. canaliculata.

Invasive snails and an emerging infectious disease: results from the first national survey on Angiostrongylus cantonensis in China.

BACKGROUND: Eosinophilic meningitis (angiostrongyliasis) caused by Angiostrongylus cantonensis is emerging in mainland China. However, the distribution of A. cantonensis and its intermediate host snails, and the role of two invasive snail species in the emergence of angiostrongyliasis, are not well understood. METHODOLOGY/PRINCIPAL FINDINGS: A national survey pertaining to A. cantonensis was carried out using a grid sampling approach (spatial resolution: 40x40 km). One village per grid cell was randomly selected from a 5% random sample of grid cells located in areas where the presence of the intermediate host snail Pomacea canaliculata had been predicted based on a degree-day model. Potential intermediate hosts of A. cantonensis were collected in the field, restaurants, markets and snail farms, and examined for infection. The infection prevalence among intermediate host snails was estimated, and the prevalence of A. cantonensis within P. canaliculata was displayed on a map, and predicted for non-sampled locations. It was confirmed that P. canaliculata and Achatina fulica were the predominant intermediate hosts of A. cantonensis in China, and these snails were found to be well established in 11 and six provinces, respectively. Infected snails of either species were found in seven provinces, closely matching the endemic area of A. cantonensis. Infected snails were also found in markets and restaurants. Two clusters of A. cantonensis-infected P. canaliculata were predicted in Fujian and Guangxi provinces. CONCLUSIONS/SIGNIFICANCE: The first national survey in China revealed a wide distribution of A. cantonensis and two invasive snail species, indicating that a considerable number of people are at risk of angiostrongyliasis. Health education, rigorous food inspection and surveillance are all needed to prevent recurrent angiostrongyliasis outbreaks.

Angiostrongylus cantonensis: morphological and behavioral investigation within the freshwater snail Pomacea canaliculata.

An infection with Angiostrongylus cantonensis, the main causative agent for human eosinophilic encephalitis, can be acquired through the consumption of the freshwater snail Pomacea canaliculata. This snail also provides a suitable model to study the developmental morphology and behavior of A. cantonensis larvae, facilitated by the snail's distinct lung structure. We used microanatomy for studying the natural appearance and behavior of A. cantonensis larvae while developing within P. canaliculata. The distribution of refractile granules in the larval body and characteristic head structures changed during the developmental cycle. Two well-developed, rod-like structures with expanded knob-like tips at the anterior part were observed under the buccal cavity as early as the late second developmental stage. A "T"-shaped structure at the anterior end and its tenacity distinguished the outer sheath from that shed during the second molting. Early first-stage larvae obtained from fresh rat feces are free moving and characterized by a coiled tail, whereas a mellifluous "Q"-movement was the behavioral trait of third-stage A. cantonensis larvae outside the host tissue. In combination, the distribution of refractive granules, distinct head features, variations in sheaths, and behavioral characteristics can be utilized for differentiation of larval stages, and for distinguishing A. cantonensis larvae from those of other free-living nematodes.

[The first national survey on natural nidi of Angiostrongylus cantonensis in China].

OBJECTIVE: To reveal the natural distribution of Angiostrongylus cantonensis in the mainland of China. METHODS: The potential areas of A. cantonensis and its main intermediate host Pomacea canaliculata were predicted based on degree-day models using GIS technique. A grid sampling was performed on the prediction map and 5% grids were randomly sampled. A total of 55 sampled sites were selected for the survey on A. cantonensis and its hosts in September-October of 2006. RESULTS: Nineteen Provinces/Municipalities/Autonomous Regions were found as potential habitats for P. canaliculata in the mainland of China. It was then confirmed that the snails distributed in the provinces of Fujian, Jiangxi, Zhejiang, Hunan, Guangdong, Guangxi, Hainan, and Yunnan. Higher prevalence of A. cantonensis in P. canaliculata was detected in Jianou of Fujian (36.6%), Xingguo of Jiangxi (19.9%), Rui'an of Zhejiang (16.0%), Rucheng of Hunan (5.0%), Huazhou of Guangdong (6.3%), Shangsi of Guangxi (39.1%) and Wuzhishan of Hainan (25.0%). CONCLUSION: Natural nidi of A. cantonensis have been found in seven provinces where natural infection in P. canaliculata has been detected.

[Morphology of III stage larvae of Angiostrongylus cantonensis in Pomacea canaliculata].

OBJECTIVE: To observe the morphologic characteristics of III stage larvae of Angiostrongylus cantonensis from Pomacea canaliculata. METHODS: P. canaliculata, the intermediate host snail of A. cantonensis, was infected with I stage larvae of A. cantonensis in laboratory. After 61 days, III stage larvae of A. cantonensis were harvested from snail's lungs and muscle of head-foot, followed by HE stain to observe morphological characteristics. RESULTS: The whole body of III stage larva was curling with obtuse head. Its pharyngeal canal extends from the buccal hole on the top of the head to the intestines at the pharyngeal intestine joint place, with apex cauda and clear anal tube. The tegument of the III stage larva was eosin-stained, with a transparent sheath outside of tegument. Some of the larvae cauda showed in circular cylinder, and some larvae presented ventral gland with two very short uterine which used to be the feature only showed in early IV stage larva. CONCLUSION: Morphologically characteristics of the III stage larvae is helpful to better understand the life-cycle and the control of A. cantonensis.

[Enzyme histochemistry: the effect of META-Li on Oncomelania hupensis].

OBJECTIVE: To explore the killing mechanism of META-Li against Oncomelania hupensis by observing the change of enzyme activity in snail tissue. METHODS: Sixty snails were divided into 2 groups. Snails in experiment group were immersed in META-Li (100 mg/L) for 2d and soft tissue was separated for frozen sections. Histochemical staining for the enzymes CCO, LDH, SDH, AChE and NOS was done by routine method and the average grey density was measured under microscopy. Tissue sections of 10 snails were used to detect grey density for each enzyme. Snails without META-Li treatment served as control. RESULTS: The enzyme activity of CCO and AChE in the experiment group was significantly lower than that in the control (t=12.26, P<0.01), that of LDH and NOS in the experiment group was significantly higher than that in the control (t=3.41, P<0.05). There was no significant difference on the enzyme activity of SDH between the two groups (t=0.51, P>0.05). CONCLUSION: The snail-killing effect of META-Li may be relevant to the enzyme activity in energy metabolism and the blocking of the nerve transmission.

[A comparative study of three methods in detecting Angiostrongylus cantonensis larvae in lung tissue of Pomacea canaliculata].

OBJECTIVE: To compare the efficiency of three methods, lung-microscopy, tissue homogenate and enzyme digestion in the detection of Angiostrongylus cantonensis larvae from the lungs of snails. METHODS: 60 Pomacea canalicular infected by the first-stage larvae of A. cantonensis were divided into 2 groups and the lung of each snail from the two groups was separated from the soft body. All the lungs were examined under microscope and larval nodes were counted. Each lung from one group was ground and that from the other was artificially digested by enzyme, the number of larvae in each lung was recorded. The efficiency of three methods was compared. Enzyme digestion was also used to estimate number of larvae in lung and in other body parts. RESULTS: By using enzyme digestion as the standard method, the detection rate of lung-microscopy, tissue homogenate and enzyme digestion was 96.7%, 93.4% and 100% respectively (chi2=2.069, P>0.05), while the lung-microscopy was significantly faster (Z=4.782, P<0.01). The number of larvae in snail lung was positively correlated with that in other part (r=0.847, P<0.01). CONCLUSIONS: The lung-microscopy in larvae detection is similarly efficient to the other two methods but faster, which is therefore more suitable for snail screening in the field.

[Studies on the growth-development and infectivity of Angiostrongylus cantonensis in dormant Pomacea canaliculata].

OBJECTIVE: To study the impact of dormancy of Pomacea canaliculata on the growth-development and infectivity of Angiostrongylus cantonensis. METHODS: The intermediate host snails (P. canaliculata) were infected with the first stage larvae of A. cantonensis from the laboratory. One day after infection the snails were kept dormant under 25.0-25.5 degrees C, and a sample of the snails was selected and dissected to examine the larval growth-development at various interval. Twenty days after infection, they were placed in room with natural winter conditions. Every 10 days a sample of the snails was dissected for larval activity. The third-stage larval infectivity from each group was identified by infecting SD rats. Meanwhile the survival and weight change of snails in the two groups were recorded, and were compared with those snails cultured in water under the corresponding temperature conditions mentioned above. RESULTS: The time for first-stage larvae of A. cantonensis in dormant snails to develop to third stage was shorter than that in the snails in aquarium. All the third stage larvae at various degree of activity recovered from snails in winter room conditions, including dormant snails and active snails in water, infected rats successfully. The dormant snails in winter room conditions stopped growing with decreased weight, but the survival rate was significantly lower than that of the snails in aquarium with the same condition along with an extending time of dormancy. CONCLUSION: The development of A. cantonensis larvae has not been affected when snails are kept dormant under 25.0-25.5 degrees C. The third stage larvae from snails at natural winter room temperature or in aquarium were all infective. As of the overwintering ways, it is better to keep the infected snails dormant than in the aquarium.

[Experimental study on compatibility of three species of freshwater snails with Angiostrongylus cantonensis].

OBJECTIVE: To compare the compatibility of three species of freshwater snails, Pomacea canaliculata, Cipangopaludina chinensis, Bellamy a aeruginosa, with Angiostrongylus cantonensis. METHODS: The snails were infected by the first-stage larvae of A. cantonensis under the same conditions. Twenty snails of each species were randomly sampled after exposed to the larvae for 1, 3, 6, 12, 24 hours, respectively. Each group was placed into an aquarium. Each species with same number was established as control. All the aquaria were equipped with a filter, the water temperature was kept at (24 +/- 1) degrees C. In the first two weeks, the number of death was recorded. Later, the snails were successively examined to record the weight and worm burden of each snail. RESULTS: Some snails were dead which mainly happened in the first week postinfection. The death rate and infection rate were not associated with exposure time and snail species. Worm burden of P. canaliculata was significantly higher than the other two species, however, the worm density of P. canaliculata and B. aeruginosa was not significantly different but higher than that of C. chinensis. The worm burden and worm density of P. canaliculata and B. aeruginosa, respectively, were significantly different among five exposure-time groups, but that of C. chinensis was not. CONCLUSION: All the three snail species show a high compatibility with A. cantoneasis. In general, the compatibility of P. canaliculata is superior to the other two species.

[Study on the molluscicidal effect of META-Li against Oncomelania hupensis].

OBJECTIVE: To evaluate the molluscicidal effect of the META-Li (40% META liquid formulation) against Oncomelania snails in laboratory and field. METHODS: The experiment of META-Li against the snails by spraying, immersion and climbing-inhibition test was carried out in laboratory. Spray method was performed in the field to compare with the wettable powder of 50% niclosamide ethanolamine salt. RESULTS: In laboratory, the LC50 of META-Li by spraying for 1, 2, 3 days was 0.78, 0.44 and 0.46 g/m2 respectively; by immersion method for 1, 2, 3 days, it was 44.4, 27.4 and 24.8 mg/L respectively. When sprayed with active ingredient 2 g/mW of META-Li in the field, the snail death rate was above 90% after 7 days, similar to that of niclosamide. CONCLUSION: META-Li shows high molluscicidal and climbing-inhibition effect on Oncomelania snails.

The effect of temperature on the development of Angiostrongylus cantonensis (Chen 1935) in Pomacea canaliculata (Lamarck 1822).

Angiostrongyliasis cantonensis, clinically presented as eosinophilic meningitis, is a snail-borne parasitic disease. We studied the effects of different temperatures on the larval development of Angiostrongylus cantonensis in the freshwater snail Pomacea canaliculata. Six groups of snails were infected and each group was cultured under different temperature conditions. At predefined intervals, four snails from each group were dissected to examine the larval development. The development-time curve of each group was drawn according to the fraction of third-stage larvae present. The developmental time was defined as the time needed until 50% of the first-stage larvae developed into third-stage larvae. A linear regression model was established based on the time (D; in days) and the corresponding temperature (T; in degrees Celsius): DT = 15.04 x D + 262.53. The threshold temperature for larval development was 15.04 degrees C and the thermal constant was 262.53 degree-days. These parameters could be helpful in estimating the number of parasite generations in a year and the impact of climate change on the distribution of A. cantonensis.

[Development of PCR assay for detection of Angiostrongylus cantonensis in Pomacea canaliculata].

OBJECTIVE: To establish a PCR assay for detecting the third-stage larvae of Angiostrongylus cantonensis in Pomacea canaliculata. METHODS: Polymerase chain reaction primers were designed by the software Lasergene, based on the specific cDNA of the third-stage larvae of A.cantonensis in Genbank. The total RNA was prepared from the third-stage larvae of A.cantonensis and of the snails by TRIzol one-step protocol. Amplification by RT-PCR was carried out following the kit protocol. RESULTS: RT-PCR assay revealed a clear differentiation between infected and negative snails. When a mixture of the total RNA from the negative snails and the third-stage larvae of A.cantonensis was tested by the PCR assay, the detectable level was 128 pg RNA, a concentration close to one third-stage larva of A.cantonensis, minimum concentration that could be found by naked eyes. The minimum detected total RNA concentration of the third-stage larvae of A.cantonensis was 105 pg by PCR assay. CONCLUSION: A PCR assay has been developed for detecting A.cantonensis larva in Pomacea canaliculata.