Competency of malaria laboratory diagnosis at national and provincial levels at the beginning of malaria post-elimination phase, China.

BACKGROUND: Qualified malaria diagnosis competency has contributed to the great achievement of malaria elimination in China. After eliminating malaria, it is still critical to the prevention of re-establishment of malaria transmission in China. This study was aimed to assess the malaria detection competency at national and provincial levels in China at the beginning of malaria post-elimination phase. METHODS: In the present study, different competency assessment activities on the laboratory malaria diagnosis were carried out for national and provincial malaria diagnostic laboratories based on the WHO scoring schedules, including malaria microscopy or nucleic acid amplification tests (NAAT), at the beginning of malaria post-elimination phase (2021-2022) in China. RESULTS: A total of 60 slides for malaria microscopy and 10 specimen for NAAT were included into the WHO External Quality Assessments of malaria parasite qualitative detection and species identification, and the scoring rate was 96.6% (microscopy: 171/177) and 85.0% (NAAT: 17/20), respectively. Moreover, 124 samples were included into the national NAAT quality assessment, and an accuracy of 87.9% (109/124) was found without significance among reference laboratories and non-reference laboratories. CONCLUSIONS: The findings suggest that there is still a need for sustained strengthening of malaria detection competency, particularly in the areas of parasite counting and detection of low-density parasitemia, to ensure prompt detection of the sources of infection and accurate identification of Plasmodium species, and contribute to case management and focus disposal, thereby effectively preventing the malaria re-establishment.

Epidemiological characteristics of Plasmodium malariae malaria in China: a malaria that should not be neglected post elimination.

BACKGROUND: Plasmodium malariae was always neglected compared with P. falciparum and P. vivax. In the present study, we aimed to describe the epidemiology of reported cases infected with P. malariae in the past decade to raise awareness of the potential threat of this malaria parasite in China. METHODS: Individual data of malaria cases infected with P. malariae reported in China in the past decade were collected via the China Information System for Disease Control and Prevention and Parasitic Diseases Information Reporting Management System, to explore their epidemiological characteristics. Pearson Chi-square tests or Fisher's Exact Test was used in the statistical analysis. RESULTS: From 2013 to 2022, a total of 581 P. malariae cases were reported in China, and mainly concentrated in 20-59 years old group (P < 0.001), and there was no significant trend in the number of cases reported per month. Moreover, four kinds of P. malariae cases were classified, including 567 imported cases from 41 countries in 8 regions and distributed in 27 provinces (autonomous regions, municipalities) in China, six indigenous cases in a small outbreak in Hainan, seven recurrent cases in Guangdong and Shanghai, and one induced case in Shanghai, respectively. In addition, only 379 cases (65.2%) were diagnosed as malaria on the first visit (P < 0.001), and 413 cases (71.1%) were further confirmed as P. malariae cases (P = 0.002). Meanwhile, most cases sought healthcare first in the health facilities at the county and prefectural levels, but only 76.7% (161/210) and 73.7% (146/198) cases were diagnosed as malaria, and the accuracy of confirmed diagnosis as malaria cases infected with P. malariae was only 77.2% (156/202) and 69.9% (167/239) in these health facilities respectively. CONCLUSIONS: Even though malaria cases infected with P. malariae didn't account for a high proportion of reported malaria cases nationwide, the threat posed by widely distributed imported cases, a small number of indigenous cases, recurrent cases and induced case cannot be ignored in China. Therefore, it is necessary to raise awareness and improve the surveillance and response to the non-falciparum species such as P. malariae, and prevent the reestablishment of malaria transmission after elimination.

Evaluation of PEN2-ATP6AP1 axis as an antiparasitic target for metformin based on phylogeny analysis and molecular docking.

BACKGROUND: Metformin (Met), the first-line drug used in the treatment for type 2 diabetes mellitus, is effective against a variety of parasites. However, the molecular target of Met at clinical dose against various parasites remains unclear. Recently, low-dose Met (clinical dose) has been reported to directly bind PEN2 (presenilin enhancer protein 2) and initiate the lysosomal glucose-sensing pathway for AMPK activation via ATP6AP1 (V-type proton ATPase subunit S1), rather than perturbing AMP/ATP levels. METHODS: To explore the possibility of PEN2-ATP6AP1 axis as a drug target of Met for the treatment of parasitic diseases, we identified and characterized orthologs of PEN2 and ATP6AP1 genes in parasites, by constructing phylogenetic trees, analyzing protein sequences and predicting interactions between Met and parasite PEN2. RESULTS: The results showed that PEN2 and ATP6AP1 genes are only found together in a few of parasite species in the cestoda and nematoda groups. Indicated by molecular simulation, Met might function by interacting with PEN2 on V37/W38/E5 (Trichinella spiralis) with similar binding energy, and on F35/S39 (Caenorhabditis elegans) with higher binding energy, comparing to human PEN2. Hence, these results indicated that only the T. spiralis PEN2-ATP6AP1 axis has the potential to be the direct target of low-concentration Met. Together with contribution of host cells including immune cells in vivo, T. spiralis PEN2-ATP6AP1 axis might play roles in reducing parasite load at low-concentration Met. However, the mechanisms of low-concentration Met on other parasitic infections might be mainly achieved by regulating host cells, rather than directly targeting PEN2-ATP6AP1 axis. CONCLUSIONS: These findings revealed the potential mechanisms by which Met treats various parasitic diseases, and shed new light on the development of antiparasitic drugs.

Blastocystis infection among diarrhea outpatients in Ningbo, Southeast China: A potential zoonotic health threat.

BACKGROUND: Blastocystis is one of the important zoonotic parasites which can infect humans and various animals worldwide and has become a growing global public health concern. The study aims to obtain the data of Blastocystis infection and the information of the genetic characteristic. METHODS: In the present study, 489 fecal samples were collected from diarrhea outpatients in Ningbo, Zhejiang province, and were examined the presence of Blastocystis by polymerase chain reaction combined with sequencing. RESULTS: A total of 10 samples (2.04%, 10/489) were positive for Blastocystis with no significant difference among sex and age groups, respectively. Eight samples were successfully sequenced, and five zoonotic ST3 and three zoonotic ST1 with two new sequences were identified. CONCLUSIONS: We first demonstrated the occurrence of Blastocystis infection in diarrhea outpatients in Ningbo, with two zoonotic subtypes (ST1 and ST3) and two new sequences being characterized. Meanwhile, mixed infection of Blastocystis and E. bieneusi was found which indicates the importance of investigation of multiple parasites. Finally, more extensive studies will be needed to better understand the transmission of Blastocystis at human-animal-environment interface and provide evidence for the development of one health strategies for the prevention and control of such diseases.

A review of malaria molecular markers for drug resistance in Plasmodium falciparum and Plasmodium vivax in China.

China has now achieved the elimination of malaria, but it still faces severe challenges in the post-elimination stage. China continues to be plagued by imported malaria cases, and preventing re-transmission of imported malaria is critical. The effectiveness of antimalarial drugs for malaria control largely depends on the study of drug resistance markers in vitro. Monitoring molecular markers of parasite-associated drug resistance can help predict and manage drug resistance. There is currently a lack of systematic reviews of molecular markers for indigenous and imported malaria in China. Therefore, this review summarizes the published articles related to molecular marker polymorphism of indigenous and imported malaria cases in China in the past two decades, to study the mutation frequency and distribution of crt, mdr1, dhps, dhfr and K13 gene resistance-related loci. This can provide a whole picture of molecular markers and the resistance mutations of imported cases in China, which has certain significance for drug resistance surveillance planning, safe and effective treatment, and preventing the recurrence of local transmission by imported malaria in China in the future.

1-3-7 surveillance and response approach in malaria elimination: China's practice and global adaptions.

There has been a significant reduction in malaria morbidity and mortality worldwide from 2000 to 2019. However, the incidence and mortality increased again in 2020 due to the disruption to services during the COVID-19 pandemic. Surveillance to reduce the burden of malaria, eliminate the disease and prevent its retransmission is, therefore, crucial. The 1-3-7 approach proposed by China has played an important role in eliminating malaria, which has been internationally popularized and adopted in some countries to help eliminate malaria. This review summarizes the experience and lessons of 1-3-7 approach in China and its application in other malaria-endemic countries, so as to provide references for its role in eliminating malaria and preventing retransmission. This approach needs to be tailored and adapted according to the region condition, considering the completion, timeliness and limitation of case-based reactive surveillance and response. It is very important to popularize malaria knowledge, train staff, improve the capacity of health centres and monitor high-risk groups to improve the performance in eliminating settings. After all, remaining vigilance in detecting malaria cases and optimizing surveillance and response systems are critical to achieving and sustaining malaria elimination.

Extracellular vesicles secreted by Echinococcus multilocularis: important players in angiogenesis promotion.

The involvement of Echinococcus multilocularis, and other parasitic helminths, in regulating host physiology is well recognized, but molecular mechanisms remain unclear. Extracellular vesicles (EVs) released by helminths play important roles in regulating parasite-host interactions by transferring materials to the host. Analysis of protein cargo of EVs from E. multilocularis protoscoleces in the present study revealed a unique composition exclusively associated with vesicle biogenesis. Common proteins in various Echinococcus species were identified, including the classical EVs markers tetraspanins, TSG101 and Alix. Further, unique tegumental antigens were identified which could be exploited as Echinococcus EV markers. Parasite- and host-derived proteins within these EVs are predicted to support important roles in parasite-parasite and parasite-host communication. In addition, the enriched host-derived protein payloads identified in parasite EVs in the present study suggested that they can be involved in focal adhesion and potentially promote angiogenesis. Further, increased angiogenesis was observed in livers of mice infected with E. multilocularis and the expression of several angiogenesis-regulated molecules, including VEGF, MMP9, MCP-1, SDF-1 and serpin E1 were increased. Significantly, EVs released by the E. multilocularis protoscolex promoted proliferation and tube formation by human umbilical vein endothelial cells (HUVECs) in vitro. Taken together, we present the first evidence that tapeworm-secreted EVs may promote angiogenesis in Echinococcus-infections, identifying central mechanisms of Echinococcus-host interactions.

Imported malaria from land bordering countries in China: A challenge in preventing the reestablishment of malaria transmission.

OBJECTIVE: No indigenous malaria cases have been reported since 2017 in China, but a large number of imported cases are still reported every year, including those from the land bordering countries. To characterize their epidemiological profiles will provide evidence for the development of appropriate strategies to effectively address the challenges of border malaria in the post-elimination phase. METHODS: Individual-level data of imported malaria cases from the land bordering countries were collected from 2017 to 2021 in China via the web-based surveillance systems, and analyzed by SPSS, ArcGIS and WPS software, to explore their epidemiological profiles. RESULTS: A total of 1170 malaria cases imported into China from six of the fourteen land bordering countries were reported between 2017 and 2021 with a decline trend. Overall, cases were widely distributed in 31-97 counties from 11 to 21 provinces but mainly in Yunnan. Moreover, these imported cases were mainly infected with P. vivax (94.8%), and a total of 68 recurrent cases were reported in 6-14 counties from 4 to 8 provinces. In addition, nearly 57.1% of the total reported cases could seek healthcare within 2 days of getting sick, and 71.3% of the reported cases could be confirmed as malaria on the day they sought medical care. CONCLUSIONS: China still needs to attach great importance to the risk and challenge of the imported malaria from bordering countries particularly from Myanmar in preventing reestablishment of malaria transmission in the post-elimination phase. It is necessary not only to strengthen collaboration and cooperation with the bordering countries, but also coordinate multiple departments at home to improve malaria surveillance and response system and prevent the reestablishment of malaria transmission in China.

Evolution of anti-malaria policies and measures in P.R. China for achieving and sustaining malaria-free.

Malaria is a major public health threat worldwide, and it was also widely prevalent in the history in China, seriously endangering people's health and affecting socioeconomic development. China was certified malaria elimination in 2021 with unremitting efforts since the founding of the People's Republic of China in 1949. This great achievement has been another milestone in the fight against major infectious diseases following the elimination of smallpox, poliomyelitis, leprosy, filariasis, neonatal tetanus and blinding trachoma in China. This paper briefly introduces the malaria burden dynamics and the corresponding malaria transmission risk stratificantions, as well as systematically reviews the evolution of anti-malaria policies and measures from severe epidemic to elimination in China. Meanwhile, five key lessons in malaria control and elimination in China are also briefly summarized. All of the above provide evidences for promoting global malaria eradication and preventing reestablishment of malaria transmission, finally benefit all individuals still suffering from the scourge of malaria.

First report on occurrence and genotypes of Enterocytozoon bieneusi, Cryptosporidium spp. and Cyclospora cayetanensis from diarrheal outpatients in Ningbo, Southeast China.

Enterocytozoon bieneusi, Cryptosporidium spp. and Cyclospora cayetanensis are three important zoonotic pathogens which were a major cause of foodborne or waterborne intestinal diseases in humans and animals. However, very little data about occurrence and genotypes of the three parasites in Ningbo in the south wing of the Yangtze River Delta, China, which is important for a tourist city. In the present study, molecular characterization of E. bieneusi, C. cayetanensis and Cryptosporidium spp. in fecal samples from 489 diarrheal outpatients were carried out. As a result, a total of 35 (7.16%, 35/489) and three (0.61%, 3/489) samples were positive for E. bieneusi and C. cayetanensis respectively. No Cryptosporidium-positive sample or mixed-infections were detected. Four known E. bieneusi genotypes (Type IV, D, I and CHN4) and 8 novel genotypes (NBH1-NBH8) were identified with type IV was the dominant genotype (n = 14), followed by genotypes D (n = 5), NBH8 (n = 5) and NBH7 (n = 3). The remaining genotypes were found in one sample each, and these genotypes were belonged to the previously described high-potential zoonotic group 1. One novel sequence named NBC315, and the other two sequences (NBC30 and NBC370) identical with the reported sequence were detected. Therefore, the existence and importance of zoonotic potential of E. bieneusi and C. cayetanensis in diarrheal outpatients in Ningbo indicates the public health threats, and more investigations should be carried out in human populations, animals and other environmental sources from the One Health perspective.

Risks of and response to cluster outbreak of imported malaria during malaria post-elimination era in China / 中国热带医学

@#Abstract: Objective　To analyze the recent cluster outbreaks of imported malaria and explore the risks, challenges and countermeasures for dealing with such events during malaria post-elimination era of malaria, and to provide reference for effectively addressing the risks and consolidating the achievements of malaria elimination. Methods　The individual malaria case data from "The Information System for Infectious Disease Surveillance" and "The Information System For Parasitic Diseases Prevention And Control" were collected，and the diagnosis classification, infection source, time and space distribution of cases were analyzed. Results　From January 1 to August 11, 2022, a total of 429 malaria cases were reported nationwide, an 18.9% decrease compared to the same period last year (529 cases), all of which were imported cases. The overall weekly trend of the outbreak remained stable, but since Week 31 (July 25-31), there has been a significant increase in the number of cases, with a peak on August 5. From July 25 to August 11, 2022, a total of 162 malaria cases were reported nationwide, up 315.4% from 39 cases in the same period last year, accounting for 37.8% of the total cases up to August 11, 2022. The main source of imported infections was Guinea (95 cases, 58.6%), with most cases reported in Longgang District, Shenzhen City, Guangdong Province (30 cases), Shilin County, Kunming City, Yunnan Province (21 cases), Chaoyang District, Beijing (11 cases), and Xiaoshan District, Hangzhou City, Zhejiang Province (7 cases). Conclusions　Due to the concentration of returnees to China, several entry port cities simultaneously experienced cluster outbreaks of imported malaria, which brought immense pressure and challenges to local medical and health institutions. Health facilities at all levels need to maintain high vigilance and sensitivity, be well prepared, and avoid death and secondary transmission caused by imported cases.

Malaria Microscopy Competency in the Subnational Verification, China: Implications for Malaria Elimination and the Prevention of Malaria Reestablishment.

Introduction: Qualified microscopy competency is a key indicator for certification of malaria elimination. To better prepare the country certification and identify the priorities that need improvement to prevent malaria reestablishment, microscopy competency at different levels were assessed in subnational verification of malaria elimination in China. Methodology. Microscopist representatives from centers for disease control and prevention (CDC)/institutes of parasitic diseases (IPD) and medical institutes for malaria diagnosis at the provincial and county levels in the subnational verification were analyzed. Specifically, five provincial microscopist representatives and ten county-level representatives were assessed in each of previously endemic provinces on qualitative identification (Plasmodium positive or negative) and Plasmodium species identification using standard slides from the National Malaria Diagnosis Reference Laboratory. Results: A total of 100 provincial-level representatives (60 from 42 CDCs/IPDs and 40 from 34 medical institutes) and 200 county-level representatives (61 from 41 CDCs and 139 from 118 medical institutes) were included. The qualitative accuracy was higher than 90% each (P = 0.137), but slides with low parasite density were easy to be misdiagnosed as negative. Furthermore, the accuracy of species identification was 80.0% and 83.6% in medical institutes and centers for disease control and prevention (CDCs) at the provincial level (P = 0.407) with relatively high misdiagnosis of P. vivax as P. ovale in the latter (16.2%) and 82.0% and 85.0% in medical institutes and CDCs at the county level (P = 0.330) for the identification of P. falciparum and non-P. falciparum with higher false-negative in medical institutions (P < 0.001). Conclusions: In conclusion, competent microscopy in subnational verification supported the quality in eliminating malaria in China, while the accurate identification of malaria parasites, especially slides with low parasite density still need to be improved through continuous diagnostic platform construction, continuous technological innovation, and targeted training to prevent reestablishment of malaria transmission.

Surveillance and Response to Imported Malaria During the COVID-19 Epidemic - Anhui Province, China, 2019-2021.

What is already known about this topic?: China was certified malaria-free on June 30, 2021. However, imported malaria continuously threatens the effort to prevent re-establishment of malaria in China. What is added by this report?: Measures such as international travel restrictions, entry quarantine, and screening in fever clinics during the coronavirus disease 2019 (COVID-19) period were associated with a significant decrease of imported malaria cases in Anhui Province, a higher proportion of non-Plasmodium falciparum (non-P. falciparum) malaria reported infections, and a higher proportion of cases requiring medical attention at their initial visit. What are the implications for public health practices?: It is necessary to be vigilant about imported malaria during the COVID-19 epidemic, especially for non-P. falciparum infections which are more difficult to detect, and to promote research, development, and introduction of more sensitive and specific point-of-care detection methods for non-P. falciparum species.

Genotyping and subtyping of Cryptosporidium spp. and Giardia duodenalis isolates from two wild rodent species in Gansu Province, China.

Cryptosporidium spp. and Giardia duodenalis are commonly detected intestinal protozoa species in humans and animals, contributing to global gastroenteritis spread. The present study examined the prevalence and zoonotic potential of Cryptosporidium spp. and G. duodenalis in Himalayan marmots and Alashan ground squirrels in China's Qinghai-Tibetan Plateau area (QTPA) for the first time. Four hundred ninety-eight intestinal content samples were collected from five counties of QTPA of Gansu province, China. All samples were examined for Cryptosporidium spp. and G. duodenalis by PCR amplification. The resultant data were statistically analyzed by chi-square, Fisher's test and Bonferroni correction using SPSS software 25. 0. Cryptosporidium positive samples were further subtyped through analysis of the 60-kDa glycoprotein (gp60) gene sequence. A total of 11 and 8 samples were positive for Cryptosporidium spp. and G. duodenalis, respectively. Prevalence of Cryptosporidium spp. and G. duodenalis were 2.5% (10/399) and 1.5% (6/399) in Himalayan marmots, 1.0% (1/99) and 2.0% (2/99) in Alashan ground squirrels, respectively. Sequence analysis confirmed the presence of C. rubeyi (n = 2), ground squirrel genotype II (n = 7), chipmunk genotype V (n = 1) and horse genotype (n = 1). The horse genotype was further subtyped as novel subtype VIbA10. G. duodenalis zoonotic assemblages A (n = 1), B (n = 6), E (n = 1) were identified in the present study. This is the first study to identify Cryptosporidium spp. and G. duodenalis in Himalayan marmots and Alashan ground squirrels, suggesting the potential zoonotic transmission of the two pathogens in QTPA.

A malaria-free China: global importance and key experience.

Malaria has a worldwide distribution and is the world's deadliest mosquito-borne disease. The goal of malaria elimination is also reflected in the United Nations Sustainable Development Goals and the Global Technical Strategy for Malaria 2016-2030 issued by the World Health Organization (WHO). China succeeded in its malaria elimination programme after being certified as malaria-free by the WHO on 30 June 2021. Therefore, we document some of the key lessons learnt in the course of the malaria elimination effort in China in this special volume, showing how different strategies made elimination feasible in different subregions of China with different epidemiological and socioeconomic characteristics, in order to present strong signals to other malaria-endemic countries that malaria elimination is feasible within one generation.

Profiling of Transcriptome-Wide N6-Methyladenosine (m6A) Modifications and Identifying m6A Associated Regulation in Sperm Tail Formation in Anopheles sinensis.

Recent discoveries of reversible N6-methyladenosine (m6A) methylation on messenger RNA (mRNA) and mapping of m6A methylomes in many species have revealed potential regulatory functions of this RNA modification by m6A players-writers, readers, and erasers. Here, we first profile transcriptome-wide m6A in female and male Anopheles sinensis and reveal that m6A is also a highly conserved modification of mRNA in mosquitoes. Distinct from mammals and yeast but similar to Arabidopsis thaliana, m6A in An. sinensis is enriched not only around the stop codon and within 3'-untranslated regions but also around the start codon and 5'-UTR. Gene ontology analysis indicates the unique distribution pattern of m6A in An. sinensis is associated with mosquito sex-specific pathways such as tRNA wobble uridine modification and phospholipid-binding in females, and peptidoglycan catabolic process, exosome and signal recognition particle, endoplasmic reticulum targeting, and RNA helicase activity in males. The positive correlation between m6A deposition and mRNA abundance indicates that m6A can play a role in regulating gene expression in mosquitoes. Furthermore, many spermatogenesis-associated genes, especially those related to mature sperm flagellum formation, are positively modulated by m6A methylation. A transcriptional regulatory network of m6A in An. sinensis is first profiled in the present study, especially in spermatogenesis, which may provide a new clue for the control of this disease-transmitting vector.

Key takeaways from China's success in eliminating malaria: leveraging existing evidence for a malaria-free world.

Although the total number of malaria cases and fatalities have declined globally since 2010, there were still 241 million malaria cases identified across 85 countries and territories in 2020. As the global malaria eradication process accelerates, more countries have launched their own initiatives of elimination. Notably, China achieved this goal by 2021, ending thousands of years of endemic. Undoubtedly, tremendous experience and vital lessons have been accrued en route to the malaria-free goal in malaria-eliminated countries including China. To enhance prospects of a malaria-free world by bridging the key evidence from a malaria-eliminated country to the contexts of affected, this personal view highlights concerted commitments and universal investment in healthcare, improved surveillance and response system, constant capacity building, demand-oriented scientific research, and multiway cooperation, which have helped China to eliminate this ancient scourge. We discuss how these key takeaways could be leveraged to different contexts. We also argue the long-term challenges and barriers on the pathway to malaria elimination and underline the needs for consistent efforts to maintain zero indigenous cases and prevent re-introduction of malaria. Through concerted efforts from global collaboration, a malaria-free world can become a reality.