[Status of malaria epidemic and feasibility of malaria elimination in Guangxi Zhuang Autonomous Region].

OBJECTIVE: To analyze the status of malaria epidemic and discuss the feasibility of malaria elimination in Guangxi Zhuang Autonomous Region. METHODS: The data of blood smear examinations of febrile patients among local residents, focus residents and mobile population in Guangxi Zhuang Autonomous Region were collected, described and analyzed statistically from 2001 to 2011. RESULTS: A total of 4 916 343 blood smear slides of local residents who had fever, 195 967 slides of focus residents, 282 461 slides of returned emigrants, and 228 341 slides of immigrants were examined, and the average positive rates of blood examinations were 0.004 8%, 0.007 7%, 0.480% and 0.127%, respectively. The indigenous malaria cases reduced from 51 in 2001 to 1 in 2009. There were no indigenous malaria cases during the period of 2010-2011. The malaria incidence has been under 1/100 000 for 11 years. The imported malaria cases were dominated in the whole region. CONCLUSIONS: Malaria situation in this region has entranced the consolidating phase of elimination according to the WHO criteria of malaria elimination. The goal of malaria elimination in Guangxi would be achieved in 2018 as long as the government and other departments pay enough attention to the imported malaria control, and the technical measures are further strengthened.

[Surveillance of malaria in Global Fund Malaria Project counties of Guangxi Zhuang Autonomous Region in 2011].

OBJECTIVE: To analyze the monitoring results and epidemic trend of malaria in the Global Fund Malaria Project counties of Guangxi Zhuang Autonomous Region in 2011, so as to provide the evidence for improving the preventive measures. METHODS: The data about malaria surveillance were collected and statistically analyzed in 7 Global Fund Malaria Project counties of Guangxi in 2011. RESULTS: A total of 8 cases of malaria were reported in the Global Fund Malaria Project counties of Guangxi in 2011, the average annual incidence rate was 0.04 per 10000 which increased by 100% compared with that in 2010, accounting for 7.14% (8/112) of the total number of cases in Guangxi. Totally 42 064 residents with fever were examined with blood tests and no case was found; 3 867 floating people were examined with blood tests and 8 cases of malaria were found (0.21%), including 5 cases of vivax malaria and 3 cases of falciparum malaria. The 8 malaria cases distributed in Longlin, Tiane, Nandan, Youjiang counties (district), and they all had ever worked in Africa or Southeast Asia. There were no input secondary cases or deaths throughout the year. CONCLUSIONS: The preventive measures of malaria are effective in the Global Fund Malaria Project counties of Guangxi, and the malaria epidemic situation is stable. It is the key to strengthen the malaria surveillance of the floating population who returned from Africa or Southeast Asia for consolidating the achievement of malaria prevention and control.

[Epidemiological investigation of imported falciparum malaria in Longlin County, Guangxi Zhuang Autonomous Region].

Among 41 people returned from Equatorial Guinea, 16 persons were infected with Plasmodium falciparum (39.02%) in Longlin County, Guangxi Zhuang Autonomous Region from October 2011 to February 2012. Among the 16 cases, Plasmodiumfalciparum was found in their blood smears of 15 cases, and 1 patient was diagnosed clinically as falciparum malaria. All the 41 people were once suffered from malaria and they received antimalarial drugs when they were in Equatorial Guinea. These imported malaria cases received the standard treatment and the foci were sprayed with insecticides. No secondary malaria cases were found. In conclusion, the measures of imported malaria control are effective. The persons returned from Equatorial Guinea have a high risk of malaria; therefore, it is necessary to strengthen the monitoring of malaria for the floating population from high malaria prevalent areas.

[Impact of pesticide use on the density of Anopheles anthropophagus and malaria incidence].

OBJECTIVE: To investigate the impact of the application of chemical pesticides on the distribution of Anopheles anthropophagus in rice fields and the malaria incidence. METHOD: Twenty-four villages from 16 counties in the provinces of Zhejiang, Sichuan and Guangxi were chosen for the surveys in the period of 1983-1987. For the survey of An. anthropophagus, indoor human bait trapping until midnight and catching the mosquitoes in all the nets in early morning were carried out to get the density and population ratio of the mosquitoes. Historical data on Anopheles spp., malaria incidence, acreage of the single or double season cropping of rice and on the quantity of chemical pesticide used in rice fields were collected from the study areas. RESULTS: In Hang-jia-hu region of Zhejiang Province, double season rice cropping was performed at that time, the quantity of pesticides used in 1973 was 45 kg/hm2, which was as high as 50 times than that in the 1950s. The density of An. anthropophagus decreased yearly, no An. anthropophagus could be found at 11 survey points in late 1980s. The malaria incidence dropped to less than 1 per 10000. In Leshan and Yibin areas of Sichuan Province, the major cultivation was single cropping, pesticides were applied in paddy fields since 1960s, and the average quantity of pesticides used was 8.6 kg/hm2 during 1970s-1980s. No significant difference on the density of An. anthropophagus was revealed between 1980s (86.2%) and 1960s (82.2%) (chi2=0.63, P>0.05). After mid-1980s, pesticide use gradually increased, and reached to 18.18 kg/hm2 in average in the years after 2000. The density of An. anthropophagus decreased, no An. anthropophagus was found in 2010 in the area surveyed and no malaria cases were reported as well. With double season cropping in Huanjiang County of Guangxi, the pesticide amount consumed was 1.79kg/hm2, 25.13 kg/hm2 and 7.68 kg/hm2 in paddy fields in 1960s, 1970s, and 1980s, respectively. The proportion of An. anthropophagus in anophelines was 52%(1 747/3 392) in the beginning of the 1980s. After the year 2000, the average pesticide use increased to 20.38 kg/hm2 in paddy fields. It was difficult to find An. anthropophagus in human dwellings after 2008. The average annual malaria incidence dropped to 0.14 per 10 000. CONCLUSION: Change of farming activities and especially use of chemical pesticides in high quantity at the rice fields undermine the breeding environments of An. anthropophagus, greatly reduce the mosquito population and therefore the malaria incidence.

[Primary evaluation on the application of nested/multiplex PCR in malaria diagnosis and surveillance].

OBJECTIVE: To compare the usefulness of Tag-primer nested/multiplex PCR (UT-PCR) method with microscopy in malaria diagnosis and surveillance. METHODS: 400 blood smears and blood samples on filter paper were taken from febrile patients which were initially diagnosed as malaria or suspected malaria during surveillance in mixed endemic areas of Plasmodium falciparum (Pf) and P. vivax (Pv) in Hainan and Yunnan provinces and a malaria-controlled area in Guangxi Zhuang Autonomous Region. The initial test results of both UT-PCR and microscopy for the 400 samples were compared under double blind condition. Blood smears with discrepant results between the two methods were retested by an experienced microscopist, and also repeated by UT-PCR for 2-3 times. The sensitivity and specificity of the two methods were evaluated following the Tjitra's method. RESULTS: Among the 400 blood samples, 234 were found plasmodium-positive by microscopy with 125 Pf and 109 Pv; 235 were positive by UT-PCR including 124 Pf, 109 Pv and 2 mixed infection. Altogether, the coincidence between the two methods stood for 92.5 % (370/400), including 154 negatives and 216 positives (Pv 117, Pf 99). 25 samples with discrepancy from the initial detections were retested, which covered 11 microscopy negative and PCR positive, 10 microscopy positive and PCR negative, 3 microscopy Pf positive and PCR Pv positive, 1 microscopy Pv positive but PCR mixed infection. 15 of the 25 samples showed same UT-PCR results, 7 "false positives" and 3 "false negatives" . Therefore, the sensitivity and specificity of UT-PCR was 99.6% and 98.8% respectively. CONCLUSION: As a diagnosis method, UT-PCR is useful for confirmation of malaria diagnosis and differentiation of Plasmodium species, also for improving the effectiveness and quality of malaria surveillance.

[Sensitivity, specificity and stability of the Tag-primer nested/multiplex PCR for malaria diagnosis].

OBJECTIVE: To improve the sensitivity, specificity and stability of the Tag-primer nested/multiplex PCR for malaria diagnosis. METHODS: Filter paper blood samples were collected from 30 non-malaria fever patients and 20 infectious disease patients (common cold, influenza, typhoid, hepatitis, etc.). Four ml blood each taken from one falciparum malaria patient and one vivax malaria patient was serially diluted. Healthy blood sample was used as negative control. Improved direct heating method was used to prepare DNA template. The cytochrome oxidase gene (coxI) located in mitochondrion was selected as target gene. Relevant web resources and software (PUBMED, NCBI-BLAST, Mfold server and Primer Premier 5.0) were employed to design and optimize Tag-primer nested/multiplex PCR (UT-PCR) which was used to test all blood samples. RESULTS: A 611 bp band and a 255 bp band were seen in serially diluted infected blood samples (1,000, 100, 10 and 1 parasite/microl) from P.f and P.v patient tested by UT-PCR. The detection limit of either P. falciparum or P. vivax reached 1 parasite/microl, and the tested blood samples of non-malaria fever patients, patients with other infectious diseases and healthy persons were all negative. Consistent results of each sample in more than 3 duplicated tests were obtained. CONCLUSION: The optimized Tag-primer nested/multiplex PCR shows high sensitivity, specificity and stability in malaria diagnosis.

[Tag primer-nested/ multiplex PCR for detection of Plasmodium falciparum and Plasmodium vivax].

OBJECTIVE: To establish a sensitive, simple to use and low noise nested/multiplex PCR for simultaneously detection of Plasmodium falciparum (Pf) and Plasmodium vivax (P.v). METHODS: The tag primer amplification technique, software Primer Premier 5.0, NCBI-BLAST web resources and the matrix test were used to optimize the nested/multiplex PCR for detection of P.f and P.v with filter paper blood samples taken from malaria patients diagnosed by microscopy, and the results of the optimized nested/multiplex PCR and microscopy were evaluated. RESULTS: The sensitivity of the optimized PCR, determined by the examination of imitative filter paper blood samples, was about 1-2 parasites / microl for P.f and 5-10 parasites / microl for P.v. Primer-dimer and other PCR noise were removed. When 71 field filter paper blood samples taken from microscopically diagnosed patients (24 P.f, 47 P.v) were examined, the concordance between the optimized PCR and microscopy was 875% for Pf and 100% for P.v. CONCLUSION: The nested/multiplex PCR optimized by tag primer amplification technique is simple, with low noise and being able to detect Pf and P.v simultaneously. It is more sensitive in detecting cases with low parasitaemia and more accurate in identifying Plasmodium species than microscopy.

[A discussion on the CSP genotyping of Plasmodium vivax and malaria control in five southern provinces of China].

OBJECTIVE: To explore circumsporozoite protein (CSP) genotype structure of Plasmodium vivax in southern China and evaluate its epidemiological significance. METHODS: Filter paper blood samples were collected from 346 vivax malaria patients in 5 provinces (Autonomous Region) including Hainan, Yunnan, Guangxi, Guangdong and Guizhou for identifying CSP genotypes, by using the method of single-tube nested/multiplex PCR. The findings combined with relevant data were statistically analyzed. RESULTS: In Guangdong, Guangxi and Guizhou Provinces (Autonomous Region), the temperate zone family strains accounted for more than 90%, with only a few tropical zone family strains and no PV-type II each strain. In Yunnan Province, temperate strains and tropical strains accounted for 71.4% and 28.6% respectively, with occasional PV-type II strain. In Hainan Province, strains of temperate zone, tropical zone and PV-type II accounted for about one-third. CONCLUSIONS: The temperate zone family strains were the predominant ones in the Provinces (Autonomous Region) of Guangdong, Guangxi and Guizhou where malaria control was carried out effectively; while in Hainan and Yunnan Provinces the difficulties in malaria control may probably be related to the complex structure of P. vivax population and multiple infections of different genotypes. The findings indicate that the complexity of the P. vivax genotype structure might be an indicative epidemiological feature for malaria control and surveillance.