Emergency response of infectious snails detected after interruption of schistosomiasis transmission in Hannan District, Wuhan / 中国热带医学

@#Abstract: Objective　To analyze the emergency response and long-term intervention effects after the detection of infectious snails epidemic by loop-mediated isothermal amplification (LAMP) assays in Hannan District, Wuhan City, and to explore the application of LAMP in early surveillance and early-warning of schistosomiasis transmission. Methods Snails picked up by the risk monitoring system in Hannan District were examined by anatomical microscopy and LAMP technology to identify the schistosomiasis infection. Emergency response and intensive intervention were initiated in the environment where positive snails appeared, and the long-term effects were evaluated. Results In May 2018, the infectious snails were detected by LAMP technology in Hannan District, and the positive snails were located in Zhujiacha, Dongzhuang Village, Obstacles and weeds were removed and buried by machine in Zhujiacha. 12 700 m2 of snails were killed by drugs, and the mortality rate of snails was more than 80%; no new seropositive persons were found in the emergency examination within 500 m of the positive snail sites. 506 people were examined in Dong Zhuang Village at the end of the year, and 30 positive IHA cases were detected with a blood positive rate of 5.93%, no positive fecal test was found, and all positive blood test patients took preventive medication. The monitoring results of sentinel rats and wild feces were all negative. Health education was carried out, 7 warning signs were deployed and refreshed, and 500 publicity brochures were distributed. After nearly three years of intensified intervention and monitoring in the villages where the positive environment is located, and the density of snails on the stubborn snail has dropped from 0.094/frame to 0.027/frame, and the positive rate of blood test in Dongzhuang Village has steadily dropped from 5.93% to 3.74%. Conclusions The infected snails missed by microscopy were detected by LAMP in Hannan District, which created conditions for the rapid emergency treatment of environment and elimination of positive snail and improved the sensitivity of the surveillance and early warning system in transmission-interrupted areas.

[Mutations of Plasmodium falciparum Multidrug Resistance 1 Gene in Imported Plasmodium falciparum in Wuhan].

Objective: To identify Plasmodium falciparum multidrug resistance 1（Pfmdr1） point mutations in imported Plasmodium falciparum in Wuhan. Methods: Blood samples were collected from returnees infected with P. falciparum in endemic areas of Africa and Myanmar during 2010-2015 in Wuhan City. Nested PCR primers were specifically designed for Pfmdr1 gene loci 86, 1042 and 1246 of P. falciparum. The Pfmdr1 gene was then amplified by nested PCR, and the products were digested by restriction enzyme ApoⅠ, AseⅠ and EcoRⅤ, respectively. The mutation rate for loci 86, 1042 and 1246 was analyzed. Results A total of 187 patients with falciparum malaria were involved in the study. Pfmdr1 was amplified from all the blood samples. Restriction enzyme digestion revealed mutation rate of 19.3%（36/187）, 4.3% （8/187） and 9.6%（18/187） for loci 86, 1042 and 1246, respectively. In detail, the mutation rate for loci 86, 1042 and 1246 was 20.6%（36/175）, 2.9%（5/175） and 10.3%（18/175） respectively in the 175 samples from Africa, and only 3 cases with locus 1042 mutation were found in the 12 samples from Myanmar. Results: A total of 187 patients with falciparum malaria were involved in the study. Pfmdr1 was amplified from all the blood samples. Restriction enzyme digestion revealed mutation rate of 19.3%（36/187）, 4.3% （8/187） and 9.6%（18/187） for loci 86, 1042 and 1246, respectively. In detail, the mutation rate for loci 86, 1042 and 1246 was 20.6%（36/175）, 2.9%（5/175） and 10.3%（18/175） respectively in the 175 samples from Africa, and only 3 cases with locus 1042 mutation were found in the 12 samples from Myanmar. Conclusion: The loci 86, 1042 and 1246 mutations of Pfmdr1 have all been found in the samples from Africa, with only one point mutation （locus 1042） found in samples from Myanmar.

[Analysis of MSP1 Allelotypes in Imported Cases of Plasmodium falciparum Malaria from Africa].

OBJECTIVE: To understand the allelotype characteristics of the merozoite surface protein 1 (MSP1) in imported Plasmodium falciparum. METHODS: Blood samples were collected from P. falciparum-infected patients returning from African malaria endemic areas. Nested PCR was used to amplify gene fragments of MSP1 coding for block 2 and block 3 motifs of MSP1 of P. falciparum by using the MSP1-specific primers. Then the allelotype of MSPI was analyzed. RESULTS: The MSP1 allelotype was detected in 117 of 135 blood samples, with a detection rate of 86.7%. In the 117 cases with successful PCR amplification, the detection rates for MAD20, K1, RO33, MAD20+K1, MAD20+RO33, K1+RO33 and MAD20+K1+R033 were 6.0%(7/117), 36.8%(43/117), 20.5%(24/117), 6.8% (8/117), 3.4% (4/117), 17.1% (20/117) and 9.4% (11/117), respectively, wherein the mixed infection accounted for 36.8%(43/135). The mean multiplicity of infection(MOI) of MSP1 allelotype was 1.46. There was no significant difference in the proportion of patients with major severity of illness among the MAD20, K1 and RO33 genotypes. The proportions of patients with major severity of illness were 25.7%(19/74) and 32.6%(14/43) in 74 cases of singular infection and 43 cases of mixed infection, respectively. The two infection types of patients had 241 ± 176 days and 285 ± 216 days of stay abroad, with no significant difference between them. CONCLUSION: The three genotypes of MSP1 and their four types of combination exist in imported cases of P. falciparum malaria from Africa. K1 and RO33 are the dominant genotves.

[Epidemiological analysis of overseas imported malaria in Wuhan City].

OBJECTIVE: To analyze the epidemiologic feature of overseas imported malaria in order to provide the evidence for improving the surveillance and control of malaria in Wuhan City. METHODS: The epidemiological investigation data of overseas imported malaria cases were collected and analyzed with an epidemiological method including plasmodium species, epidemic focus distribution, domestic distribution, population characteristics, attack-diagnosis interval, clinical grading, etc. in Wuhan City from 2008 to 2013. RESULTS: A total of 240 overseas imported malaria cases were reported, including 180 cases from African countries and the proportion of Plasmodiumfalciparum infections was 82.22%, and 60 cases from Southeast Asian countries and the proportion of Plasmodium vivax infections was 76.67%, and the proportions of the parasite species were significantly different between the two regions (χ2 = 105.53, P <0.001). The male and young adults were dominant. there were no statistical significance between clinical grading and attack-diagnosis interval (Z=-0.99, P = 0.32) , vocational background (χ2 = 10.61, P=0.10), then clinical symptoms aggravated the situation was occurred more easily among the first attack cases (χ2 = 7.66, P <0.05) and the falciparum malaria cases (χ2 =24.27, P<0.001). CONCLUSIONS: The overseas imported malaria cases are increasing rapidly with years and the malignant malaria cases were more than other malaria cases in Wuhan City. Therefore, the surveillance and management of the returnees from malaria endemic areas should be strengthened continually.

[An analysis on Pf60.1 gene polymorphism of Plasmodium falciparum from imported cases].

One hundred and one imported falciparum malaria cases in Wuhan City were confirmed by microscopy and Nest-PCR, and the blood samples were collected. The Pf60.1 gene was amplified by PCR. Among 101 blood samples, three Pf60.1 fragments [313 bp (56.5%, 52/92), 340 bp (37.0%, 34/92), 313 bp+340 bp (6.5%, 6/92)] were amplified from 92 samples. Among 83 blood samples from patients returning from Africa, 313 bp fragment were found in 46 samples (55.4%, 46/83), 340 bp fragment were found in 31 samples (37.1%, 31/83), and 7.2% (6/83) was mixed-fragment (313 bp+340 bp). Among 9 samples from southeast Asia, 6 samples were with 313 bp fragment and 3 samples with 340 bp fragment. The results indicated that the most common genotype was 313 bp-genotype, and there would be polyclonal P. falciparum infections.

[Two cases of African imported Plasmodium malariae malaria].

The process of diagnosis, epidemiological survey and treatment of two cases of African imported quartan malaria is reported in this paper.

[Investigation of Capillaria hepatica infection in rodents from Wuhan section of Yangtze River marshland].

OBJECTIVE: To understand the prevalence of Capillaria hepatica in rodents from Wuhan section of the Yangtze River marshland. METHODS: Rodents were trapped in Jiang'an section of Wuhan marshland of the Yangtze River. The livers of the rodents were examined for pathological changes by unaided eyes and the liver tissues were examined for the eggs of C. hepatica by a microscope. RESULTS: According to the natural conditions, the investigation was carried out in 6 survey areas. Each survey area was placed with 60 mousetraps, and all 360 mousetraps were recovered. A total of 31 rodents (rodent density 8.61%) were captured and examined, including 24 Apodemus agrarius, 3 Rattus norvegicus, 4 Sorex caecutiens, and C. hepatica eggs were found in 1 R. norvegicus (1/3) and not found in A. agrarius, and S. caecutiens. CONCLUSION: This study has documented a prevalence of C. hepatica in rodents from Wuhan section of the Yangtze River marshland where is a natural epidemic focus of capillariasis hepatica.

[Analysis of K76T Pfcrt genetic polymorphism in imported Plasmodium falciparum].

OBJECTIVE: To identify the incidence of the K76T mutation in Pfcrt genes of imported Plasmodium falciparum. METHODS: The blood samples were collected from returnees infected with P. falciparum in endemic areas of Southeast Asia and Africa from 2008 to 2012. According to the Pfcrt gene sequence of P. falciparum, nested PCR primers were designed. Nested PCR-RFLP was applied with falciparum DNA in the blood samples as templates. RESULTS: Among 92 blood samples of P. falciparum, the mutant Pfcrt alleles were found in 50 samples (54.3%), and the wild type Pfcrt alleles were found in 42 samples (45.7%). There were 33 samples (47.1%) with mutant Pfcrt alleles and 37 samples (52.9%) with wild type, respectively, from Africa. There were 17 samples (77.3%) with mutant Pfcrt alleles and 5 samples (22.7%) with wild type, respectively, from Southeast Asia. There was a significant difference between the Africa group and the Southeast Asia group (Chi(2) = 6.12, P < 0.05). CONCLUSION: The incidence of Pfcrt gene mutation is different among P. falciparum isolates from different regions. Therefore, Pfcrt K76T has an application value in the surveillance of the imported falciparum chloroquine-resistance.

[Analysis of Pfcrt gene polymorphism in Plasmodium falciparum from imported cases].

OBJECTIVE: To identify the incidence of the K76T mutation in Pfcrt gene of imported Plasmodium falciparum and study the Pfcrt gene polymorphism in Plasmodium falciparum. METHODS: Seventy-two blood samples were collected from patients infected with P. falciparum returning from Africa (Nigeria, Equatorial Guinea, Congo, Liberia, Angora and Mali) and Southeast Asia (Myanmar and Indonesia) from 2008 to 2012. According to Pfcrt gene sequence of P. falciparum, nested PCR primers were designed, and the reaction was applied with P. falciparum DNA in the blood samples as templates. PCR products were identified by Apo I digestion. RESULTS: Among 72 blood samples of P. falciparum, mutant Pfcrt alleles were found in 41 samples (57.7%, 41/71) and wild type Pfcrt alleles were found in 30 samples (42.3%, 30/71). There were 25 samples (50%, 25/50) each with mutant Pfcrt alleles or wild type that were from Africa, while 16 samples (76.2%, 16/21) with mutant Pfcrt alleles and 5 samples (23.8%, 5/21) with wild type that were from Southeast Asia, respectively. CONCLUSION: The incidence of Pfcrt gene mutation is different in P. falciparum isolates from different regions.

[Surveillance and forecast of schistosomiasis based on sentinel mouse technique in key water regions of Wuhan City in 2011].

OBJECTIVE: To understand the water infectivity of schistosome in key water regions of Wuhan City and explore the role of a sentinel mouse technique on surveillance and forecast system for schistosomiasis. METHODS: Schistosome-endemic areas of the Yangtze River, the Dongjing-Tongshun River system, the Fu-Lun River system and the Jinshui River of Wuhan City were chosen as the surveillance and forecast sites. The Oncomelania snail distribution and infection were surveyed before the flood season. The water infectivity was detected by using the sentinel mouse technique during the flood season. The infection status of people in the villages around the surveillance sites and the activities of human beings and domestic animals were surveyed. The correlation between the infection rate of sentinel mice and snail status was tested by the rank correlation method. The emergency response mechanism was initiated when the areas with schistosomes were detected in water. RESULTS: Among the 18 surveillance sites, 15 sites with infected sentinel mice were found, accounting for 83.33%. A total of 554 sentinel mice were placed and 540 recovered, with a recovery rate of 97.47%. All the recovered mice were dissected and 75 infected, with a total infection rate of 13.89%. Totally 172 adult worms were collected, with mean worm burden of 2.29 +/- 0.71 worms per mouse. The infection rates of sentinel mice in 4 water systems were 8.33%, 24.53%, 10.85% and 6.56%, respectively, and the mean worm burdens of infected sentinel mice were 2.33 +/- 0.71, 2.28 +/- 0.76, 2.22 +/- 0.60 and 2.75 +/- 0.96 worms per mouse, respectively. The infection rates of sentinel mice in 4 water systems had a statistically significant difference (chi2 = 19.131, P = 0.000). The mean worm burdens of the infected sentinel mice in 4 water systems had no statistically significant difference (F = 0.638, P = 0.593). The correlation coefficient among the infection rate of sentinel mice, snail area, the average density of living snails and infected snail rate had no statistical significance. Among the 15 sites with infected sentinel mice, 8 sites with fisherman activities, 8 sites with anglers or planters, 10 sites with cattle keepings and 4 of which with infected cattle. All the 15 sites with cercariae-infected water bodies started the emergency response and no epidemic situation happened. CONCLUSIONS: The water infectivity of schistosome in key water regions of Wuhan City was relatively high. Detecting water infectivity based on a sentinel mouse technique is an important part of surveillance and forecast system for schistosomiasis.

[Application of nested PCR in diagnosis of imported malaria].

OBJECTIVE: To evaluate the usefulness of nested PCR method in the diagnosis of imported malaria. METHODS: A total of 210 blood smears and blood samples on filter paper were taken from persons returned from highly malaria endemic countries. The results of both nested PCR and microscopy for 210 samples were compared. RESULTS: Among the 210 persons, 43 were hospitalized due to malaria, and positive by nested PCR test Among the rest 157 people at high risk of getting malaria, 3 were found plasmodium-positive by microscope (1.91%), and 5 were positive by nested PCR (3.18%). In four samples with discrepancy between the two methods, 1 was microscopy positive and PCR negative, and 3 were microscopy negative and PCR positive. Positive and negative coincidence rate between the two tests was 66.7% and 98.1%, respectively. The coincidence between the two methods was 97.5%. CONCLUSION: Nested PCR is useful for monitoring, identification and diagnosis of imported malaria.