Pub Date : 2024-10-02DOI: 10.1007/s11686-024-00923-x
Kié Solange Millogo, Bérenger Kaboré, Paul Sondo, Eulalie W Compaoré, Amélé Fifi Chantal Kouevi, Sié A Elisée Kambou, Toussaint Rouamba, Adama Kazienga, Hamidou Ilboudo, Marc Christian Tahita, Ismaila Bouda, Karim Derra, Sanata Bamba, Halidou Tinto
Background: Seasonal malaria chemoprevention (SMC) is an effective malaria preventive intervention in sub-Sahara Africa. However, as with any other drug-based intervention, the large-scale deployment of this strategy could lead to Amodiaquine plus Sulfadoxine-Pyrimethamine (AQSP) drug pressure on the circulating parasites population with selection for specific alleles that could compromise the impact of the intervention in the near future. This study aimed to assess the distribution of the Pfmdr1 mutation involved in resistance to AQ before and after the annual campaign of SMC in the health district of Nanoro.
Methods: Randomly selected dried blood spots collected prior (n = 100) and after (n = 100) the 2021 SMC campaign were used for the detection of mutation in codons 86 and 184 of the Pfmdr1 gene using a nested PCR with restriction fragment length polymorphism approach.
Results: No significant change in the prevalence of Pfmdr1 N86Y mutation was observed before and after the SMC campaign (p = 0.28). The mutant allele 86Y was observed at low prevalences, representing only 2.17% and 6.12%, respectively, before and after the SMC campaign. Patients harboring the mutant Pfmdr1 86Y allele exhibited higher parasite densities compared to patients with the wild-type Pfmdr1 N86 allele (p = 0.04). A significant increase in the prevalence of the mutant allele 184 F was observed in the period before and after the SMC campaign (p = 0.03).
Conclusion: This selective pressure needs to be closely monitored in order to preserve the efficacy of this intervention for a long-term period in Burkina Faso.
{"title":"Trend of N86Y and Y184F Mutations in Pfmdr1 Gene in Children Under Seasonal Malaria Chemoprevention Coverage in Nanoro, Burkina Faso.","authors":"Kié Solange Millogo, Bérenger Kaboré, Paul Sondo, Eulalie W Compaoré, Amélé Fifi Chantal Kouevi, Sié A Elisée Kambou, Toussaint Rouamba, Adama Kazienga, Hamidou Ilboudo, Marc Christian Tahita, Ismaila Bouda, Karim Derra, Sanata Bamba, Halidou Tinto","doi":"10.1007/s11686-024-00923-x","DOIUrl":"https://doi.org/10.1007/s11686-024-00923-x","url":null,"abstract":"<p><strong>Background: </strong>Seasonal malaria chemoprevention (SMC) is an effective malaria preventive intervention in sub-Sahara Africa. However, as with any other drug-based intervention, the large-scale deployment of this strategy could lead to Amodiaquine plus Sulfadoxine-Pyrimethamine (AQSP) drug pressure on the circulating parasites population with selection for specific alleles that could compromise the impact of the intervention in the near future. This study aimed to assess the distribution of the Pfmdr1 mutation involved in resistance to AQ before and after the annual campaign of SMC in the health district of Nanoro.</p><p><strong>Methods: </strong>Randomly selected dried blood spots collected prior (n = 100) and after (n = 100) the 2021 SMC campaign were used for the detection of mutation in codons 86 and 184 of the Pfmdr1 gene using a nested PCR with restriction fragment length polymorphism approach.</p><p><strong>Results: </strong>No significant change in the prevalence of Pfmdr1 N86Y mutation was observed before and after the SMC campaign (p = 0.28). The mutant allele 86Y was observed at low prevalences, representing only 2.17% and 6.12%, respectively, before and after the SMC campaign. Patients harboring the mutant Pfmdr1 86Y allele exhibited higher parasite densities compared to patients with the wild-type Pfmdr1 N86 allele (p = 0.04). A significant increase in the prevalence of the mutant allele 184 F was observed in the period before and after the SMC campaign (p = 0.03).</p><p><strong>Conclusion: </strong>This selective pressure needs to be closely monitored in order to preserve the efficacy of this intervention for a long-term period in Burkina Faso.</p>","PeriodicalId":6932,"journal":{"name":"Acta Parasitologica","volume":null,"pages":null},"PeriodicalIF":1.2,"publicationDate":"2024-10-02","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142360919","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"农林科学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2024-10-02DOI: 10.1007/s11686-024-00920-0
Eva Bártová, Jiřina Marková, Alena Žákovská, Zuzana Čadková, Marie Budíková
Purpose: Parasites of genus Encephalitozoon are well known pathogens of domestic animals however less attention was paid to its spread among wildlife that can play an important role of reservoir of infection. The aim of the study was to conduct molecular detection and genotype characterization of Encephalitozoon spp. in wild small mammals trapped in localities both near to and at a large distance from residential areas.
Methods: In total, 300 wild small mammals (274 Rodentia and 26 Eulipotyphla) were trapped in 41 localities of the Czech Republic and tested by nested PCR for Encephalitozoon spp.
Results: The DNA of Encephalitozoon spp. was proved in tissues (brain or liver) of 11% (32/300) of animals. There was a statistically significant difference (p < 0.001) in positivity among animal species with the most infected species Micromys minutus (50%, 4/8) and Myodes glareolus (17%, 9/53). There was also statistically significant difference (p < 0.001) between localities with the higher positivity (29%, 12/42) in localities near to residential areas, compared to localities with a large distance from residential areas (8%, 20/258). Sex and age of wild small mammals did not have effect on their positivity. Genotyping analysis revealed E. cuniculi genotype II in 22 samples and E. hellem genotype 1 A in one sample.
Conclusion: This study brings new information on the molecular characterization of Encephalitozoon spp. isolated from wild small mammals trapped in two different areas (localities in near to residential areas and localities with a large distance from residential areas).
{"title":"Wildlife as Reservoirs of Encephalitozoon Cuniculi and Encephalitozoon Hellem and Molecular Genotyping of Encephalitozoon spp. in Small Mammals in the Czech Republic.","authors":"Eva Bártová, Jiřina Marková, Alena Žákovská, Zuzana Čadková, Marie Budíková","doi":"10.1007/s11686-024-00920-0","DOIUrl":"https://doi.org/10.1007/s11686-024-00920-0","url":null,"abstract":"<p><strong>Purpose: </strong>Parasites of genus Encephalitozoon are well known pathogens of domestic animals however less attention was paid to its spread among wildlife that can play an important role of reservoir of infection. The aim of the study was to conduct molecular detection and genotype characterization of Encephalitozoon spp. in wild small mammals trapped in localities both near to and at a large distance from residential areas.</p><p><strong>Methods: </strong>In total, 300 wild small mammals (274 Rodentia and 26 Eulipotyphla) were trapped in 41 localities of the Czech Republic and tested by nested PCR for Encephalitozoon spp.</p><p><strong>Results: </strong>The DNA of Encephalitozoon spp. was proved in tissues (brain or liver) of 11% (32/300) of animals. There was a statistically significant difference (p < 0.001) in positivity among animal species with the most infected species Micromys minutus (50%, 4/8) and Myodes glareolus (17%, 9/53). There was also statistically significant difference (p < 0.001) between localities with the higher positivity (29%, 12/42) in localities near to residential areas, compared to localities with a large distance from residential areas (8%, 20/258). Sex and age of wild small mammals did not have effect on their positivity. Genotyping analysis revealed E. cuniculi genotype II in 22 samples and E. hellem genotype 1 A in one sample.</p><p><strong>Conclusion: </strong>This study brings new information on the molecular characterization of Encephalitozoon spp. isolated from wild small mammals trapped in two different areas (localities in near to residential areas and localities with a large distance from residential areas).</p>","PeriodicalId":6932,"journal":{"name":"Acta Parasitologica","volume":null,"pages":null},"PeriodicalIF":1.2,"publicationDate":"2024-10-02","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142360920","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"农林科学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Background: For years, the Kato-Katz (KK) technique has been considered the gold standard for diagnosing schistosomiasis. The aim of this study was to compare the effectiveness of our previously developed gold nanoparticle-based lateral flow test strip (AuNPs-LFTS) for diagnosing active Schistosoma mansoni with that of the commercially available point-of-care Circulating Cathodic Antigen detection (POC-CCA) kit.
Methods: In this study, we collected sixty positive and twenty negative urine samples from patients in endemic hot spots in the Nile Delta, as well as from patients visiting the internal medicine clinic at Theodor Bilharz Research Institute (TBRI). We produced monoclonal antibodies (MAbs) against S. mansoni soluble egg antigen (SEA) from cloned hybridoma cells (4D/1D). These MAbs were conjugated with gold and mesoporous silica nanoparticles, and used to develop the LFTS.
Results: The LFTS demonstrated a limit of detection (LoD) of 3 ng/ml. The sensitivity and specificity of the developed LFTS were found to be 96.7% and 95%, respectively, compared to 85% and 90% for the POC-CCA detection kit. The cases were divided into groups based on egg count in the stool, categorized as light, moderate, and heavy infections. The sensitivity of the LFTS in the group with light infection was higher than that of the POC-CCA. When using the KK technique (eggs per gram of stool sample [EPG]) as the reference test, the kappa value for the nano-based strips was 0.902, compared to 0.672 for the CCA strips, indicating an almost perfect agreement between KK and our developed LFTS.
Conclusion: These results confirm the reliability and effectiveness of the LFTS compared to commercially available kits for rapid, sensitive, and early diagnosis of schistosomiasis. However, it is recommended to conduct further assessments of the developed strip on a larger scale with a broader range of cases before considering its introduction to local or international markets.
{"title":"Home-Made Lateral Flow Test Strip Versus POC-CCA Assay for Detection of Active Schistosomiasis in Egypt.","authors":"Manal Kamel, Faten Salah, Zeinab Demerdash, Sara Maher, Hanan El-Baz, Nahla Zakaria, Rania Abu-Shady, Amany Saad, Salwa Hassan, Doaa Abdel Aziz","doi":"10.1007/s11686-024-00917-9","DOIUrl":"https://doi.org/10.1007/s11686-024-00917-9","url":null,"abstract":"<p><strong>Background: </strong>For years, the Kato-Katz (KK) technique has been considered the gold standard for diagnosing schistosomiasis. The aim of this study was to compare the effectiveness of our previously developed gold nanoparticle-based lateral flow test strip (AuNPs-LFTS) for diagnosing active Schistosoma mansoni with that of the commercially available point-of-care Circulating Cathodic Antigen detection (POC-CCA) kit.</p><p><strong>Methods: </strong>In this study, we collected sixty positive and twenty negative urine samples from patients in endemic hot spots in the Nile Delta, as well as from patients visiting the internal medicine clinic at Theodor Bilharz Research Institute (TBRI). We produced monoclonal antibodies (MAbs) against S. mansoni soluble egg antigen (SEA) from cloned hybridoma cells (4D/1D). These MAbs were conjugated with gold and mesoporous silica nanoparticles, and used to develop the LFTS.</p><p><strong>Results: </strong>The LFTS demonstrated a limit of detection (LoD) of 3 ng/ml. The sensitivity and specificity of the developed LFTS were found to be 96.7% and 95%, respectively, compared to 85% and 90% for the POC-CCA detection kit. The cases were divided into groups based on egg count in the stool, categorized as light, moderate, and heavy infections. The sensitivity of the LFTS in the group with light infection was higher than that of the POC-CCA. When using the KK technique (eggs per gram of stool sample [EPG]) as the reference test, the kappa value for the nano-based strips was 0.902, compared to 0.672 for the CCA strips, indicating an almost perfect agreement between KK and our developed LFTS.</p><p><strong>Conclusion: </strong>These results confirm the reliability and effectiveness of the LFTS compared to commercially available kits for rapid, sensitive, and early diagnosis of schistosomiasis. However, it is recommended to conduct further assessments of the developed strip on a larger scale with a broader range of cases before considering its introduction to local or international markets.</p>","PeriodicalId":6932,"journal":{"name":"Acta Parasitologica","volume":null,"pages":null},"PeriodicalIF":1.2,"publicationDate":"2024-10-02","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142360917","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"农林科学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2024-10-02DOI: 10.1007/s11686-024-00924-w
Amira Meriem Rebah, Delphine Gey, Zouhour El Mouna Ayadi
Purpose: In this study we describe a new species Microcotyle tazeroutii n. sp. (Monogenea: Microcotylidae) found on the gills and operculum of the boarfish Capros aper (Caproidae) off the Algerian coast of the Western Mediterranean.
Methods: Monogeneans were observed alive or recently dead on the operculum and gills using a dissecting microscope, measured and drawn for morphological study. Furthermore, a molecular analysis was conducted using a partial fragment of the mitochondrial cytochrome c oxidase subunit 1 (cox1) of two specimens of monogeneans and a tissue sample of the fish's gills in which the parasites were found to confirm the identity of fish.
Results: The new species Microcotyle tazeroutii n. sp., exhibits a combination of morphological features that differentiate it from all other known species within the genus, such as the shape and the size of body, the haptor length, the number and the size of clamps and testes, the number of spines of the genital atrium and the size of eggs. Additionally, a molecular analysis of the mitochondrial cytochrome c oxidase subunit 1 (cox1 gene) revealed significant interspecific differences between Microcotyle tazeroutii n. sp. and previously published sequences of other Microcotyle species.
Conclusion: The morphological and molecular analyses revealed that Microcotyle tazeroutii n. sp. has unique characteristics that distinguish it from all previously identified species and confirmed the presence of Microcotyle within the Caproidae family for the first time.
目的:在本研究中,我们描述了在地中海西部阿尔及利亚海岸的野猪鱼 Capros aper(Caproidae)的鳃和厣上发现的新物种 Microcotyle tazeroutii n. sp.(Monogenea:Microcotylidae):方法:使用解剖显微镜观察厣和鳃上活着或刚死不久的单齿藻,测量并绘制形态研究图。此外,还利用两个单膜虫标本的线粒体细胞色素 c 氧化酶亚单位 1(cox1)的部分片段和发现寄生虫的鱼鳃组织样本进行了分子分析,以确认鱼类的身份:结果:新物种Microcotyle tazeroutii n. sp.具有不同于该属所有其他已知物种的形态特征,如身体的形状和大小、触角长度、夹子和睾丸的数量和大小、生殖器心房的棘数量以及卵的大小。此外,对线粒体细胞色素 c 氧化酶亚基 1(cox1 基因)的分子分析表明,Microcotyle tazeroutii n. sp.与之前发表的其他 Microcotyle 种类的序列存在显著的种间差异:形态学和分子分析表明,Microcotyle tazeroutii n. sp.
{"title":"Microcotyle Tazeroutii n. sp. (Monogenea: Microcotylidae) from the Gills of the Boarfish Capros aper Linnaeus, 1758 (Teleostei: Caproidae) off the Algerian Coast, Western Mediterranean.","authors":"Amira Meriem Rebah, Delphine Gey, Zouhour El Mouna Ayadi","doi":"10.1007/s11686-024-00924-w","DOIUrl":"https://doi.org/10.1007/s11686-024-00924-w","url":null,"abstract":"<p><strong>Purpose: </strong>In this study we describe a new species Microcotyle tazeroutii n. sp. (Monogenea: Microcotylidae) found on the gills and operculum of the boarfish Capros aper (Caproidae) off the Algerian coast of the Western Mediterranean.</p><p><strong>Methods: </strong>Monogeneans were observed alive or recently dead on the operculum and gills using a dissecting microscope, measured and drawn for morphological study. Furthermore, a molecular analysis was conducted using a partial fragment of the mitochondrial cytochrome c oxidase subunit 1 (cox1) of two specimens of monogeneans and a tissue sample of the fish's gills in which the parasites were found to confirm the identity of fish.</p><p><strong>Results: </strong>The new species Microcotyle tazeroutii n. sp., exhibits a combination of morphological features that differentiate it from all other known species within the genus, such as the shape and the size of body, the haptor length, the number and the size of clamps and testes, the number of spines of the genital atrium and the size of eggs. Additionally, a molecular analysis of the mitochondrial cytochrome c oxidase subunit 1 (cox1 gene) revealed significant interspecific differences between Microcotyle tazeroutii n. sp. and previously published sequences of other Microcotyle species.</p><p><strong>Conclusion: </strong>The morphological and molecular analyses revealed that Microcotyle tazeroutii n. sp. has unique characteristics that distinguish it from all previously identified species and confirmed the presence of Microcotyle within the Caproidae family for the first time.</p>","PeriodicalId":6932,"journal":{"name":"Acta Parasitologica","volume":null,"pages":null},"PeriodicalIF":1.2,"publicationDate":"2024-10-02","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142360918","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"农林科学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2024-10-02DOI: 10.1007/s11686-024-00925-9
Bankole Emmanuel Ofeniforo, David Chukwuma Nwikwe, Samuel Eleojo Agada, Elizabeth Abidemi Balogun
Purpose: Infections by Plasmodium parasite actuate oxidative stress. As malaria parasite actions overwhelm host antioxidant defense by producing excessive reactive species during haemoglobin degradation. This study aimed to evaluate the oxidative status by considering the antioxidant level of ethyl-acetate sub-fractions of Spilanthes filicaulis (ESSF) extract on Plasmodium berghei NK-65 parasitized mice.
Methods: Sixty female mice weighing 20.0 ± 3.0 g were inoculated intraperitoneally with 0.2 mL of parasitized erythrocytes randomly selected into five groups of 12 mice, Groups I and II were orally administered with normal saline (10 mL/kg) and chloroquine (10 mg/kg) while, Groups III, IV and V were administered 250,500 and 750 mg/kg per day respectively of ESSF. Mice in each group were sacrificed on days 4 and 8 post treatment, thereafter blood and liver samples were collected and prepared using standard methods to obtain erythrocytes and liver homogenates. Malondialdehyde (MDA), a measure of lipid peroxidation, superoxide dismutase (SOD) and catalase (CAT) level was assessed in the erythrocyte and liver.
Results: Administration of ESSF produced a significant (p < 0.05) decrease in the MDA concentration of the parasitized treated group when compared to parasitized untreated group on day 4. Concomitantly, a significant (p < 0.05) increase in SOD and CAT activity in the treated groups with a corresponding decrease in the untreated group on day 4. However, effects of this action were more pronounced on day 8 in both tissues.
Conclusion: These findings suggest that ESSF might contribute to the protection of malaria infected mice against oxidative disruptions by improving antioxidant status.
目的:疟原虫感染会产生氧化应激。疟原虫在血红蛋白降解过程中会产生过多的活性物质,使宿主的抗氧化防御能力不堪重负。本研究的目的是通过考虑丝兰(Spilanthes filicaulis)提取物乙酸乙酯子馏分(ESSF)的抗氧化水平,评估寄生在伯格希疟原虫(Plasmodium berghei)NK-65 上的小鼠的氧化状态:将 60 只体重为 20.0 ± 3.0 g 的雌性小鼠腹腔接种 0.2 mL 寄生红细胞,随机分为 5 组,每组 12 只,I 组和 II 组口服生理盐水(10 mL/kg)和氯喹(10 mg/kg),III 组、IV 组和 V 组每天分别服用 250、500 和 750 mg/kg 的 ESSF。各组小鼠在治疗后第 4 天和第 8 天被处死,然后收集血液和肝脏样本,并采用标准方法制备红细胞和肝脏匀浆。对红细胞和肝脏中的丙二醛(MDA)(一种衡量脂质过氧化的指标)、超氧化物歧化酶(SOD)和过氧化氢酶(CAT)水平进行评估:结果:服用 ESSF 后,红细胞和肝脏中的脂质过氧化反应、超氧化物歧化酶(SOD)和过氧化氢酶(CAT)水平均有明显改善:这些研究结果表明,ESSF 可通过改善抗氧化状态来保护受疟疾感染的小鼠免受氧化破坏。
{"title":"Assessing the Oxidative Stress Reducing Potential of Spilanthes filicaulis (Schumach & Thonn) Ethyl-Acetate Sub-fractions on Plasmodium berghei Infected Female Mice.","authors":"Bankole Emmanuel Ofeniforo, David Chukwuma Nwikwe, Samuel Eleojo Agada, Elizabeth Abidemi Balogun","doi":"10.1007/s11686-024-00925-9","DOIUrl":"https://doi.org/10.1007/s11686-024-00925-9","url":null,"abstract":"<p><strong>Purpose: </strong>Infections by Plasmodium parasite actuate oxidative stress. As malaria parasite actions overwhelm host antioxidant defense by producing excessive reactive species during haemoglobin degradation. This study aimed to evaluate the oxidative status by considering the antioxidant level of ethyl-acetate sub-fractions of Spilanthes filicaulis (ESSF) extract on Plasmodium berghei NK-65 parasitized mice.</p><p><strong>Methods: </strong>Sixty female mice weighing 20.0 ± 3.0 g were inoculated intraperitoneally with 0.2 mL of parasitized erythrocytes randomly selected into five groups of 12 mice, Groups I and II were orally administered with normal saline (10 mL/kg) and chloroquine (10 mg/kg) while, Groups III, IV and V were administered 250,500 and 750 mg/kg per day respectively of ESSF. Mice in each group were sacrificed on days 4 and 8 post treatment, thereafter blood and liver samples were collected and prepared using standard methods to obtain erythrocytes and liver homogenates. Malondialdehyde (MDA), a measure of lipid peroxidation, superoxide dismutase (SOD) and catalase (CAT) level was assessed in the erythrocyte and liver.</p><p><strong>Results: </strong>Administration of ESSF produced a significant (p < 0.05) decrease in the MDA concentration of the parasitized treated group when compared to parasitized untreated group on day 4. Concomitantly, a significant (p < 0.05) increase in SOD and CAT activity in the treated groups with a corresponding decrease in the untreated group on day 4. However, effects of this action were more pronounced on day 8 in both tissues.</p><p><strong>Conclusion: </strong>These findings suggest that ESSF might contribute to the protection of malaria infected mice against oxidative disruptions by improving antioxidant status.</p>","PeriodicalId":6932,"journal":{"name":"Acta Parasitologica","volume":null,"pages":null},"PeriodicalIF":1.2,"publicationDate":"2024-10-02","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142360916","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"农林科学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2024-09-27DOI: 10.1007/s11686-024-00929-5
Jose Ma M Angeles, Joseph Romeo O Paner, Elena A Villacorte, Pilarita T Rivera, Shin-Ichiro Kawazu
Purpose: Animal reservoirs are important targets for controlling and eliminating schistosomiasis. Prevalence studies showed that dogs (Canis familiaris) and water buffaloes (Bubalus bubalis) are important reservoirs of S. japonicum. Previous studies have validated the use of the recombinant proteins Sj1TR, Sj7TR, and SjTPx-1 in ELISA as diagnostics for dogs and water buffaloes from schistosomiasis-endemic areas. In this study, we aimed to determine the seroprevalence of S. japonicum among dogs and water buffaloes in New Corella, Davao del Norte, Philippines, using the recombinant proteins Sj1TR, Sj7TR, and SjTPx-1 in ELISA format.
Methods: Fecal and serum samples were collected from dogs (n = 63) and water buffaloes (n = 57). Schistosome-positive samples were detected using formalin-ether concentration technique (FECT), stool polymerase chain reaction, and enzyme-linked immunosorbent assay (ELISA) using soluble egg antigen (SEA), rSj1TR, rSj7TR, and rSjTPx-1. Positivity rates, sensitivity, specificity, predictive values, accuracy, and kappa values were calculated.
Results: Mean positivity rates for schistosome infection were high for both dogs (x = 15.40%) and water buffaloes (x = 6.32%). On dogs, the sensitivity and specificity of the tests are as follows: 66.67% and 100% for rSj7TR-ELISA, while 100% and 93.33% for rSjTPx-1-ELISA, respectively. rSjTPx-1-ELISA showed the highest agreement with stool PCR among all diagnostic tests, with an overall kappa value of 0.824. On water buffaloes, the sensitivity and specificity of both rSj1TR-ELISA and rSjTPx-1-ELISA are 100.0% and 98.15%. Both tests also had an overall kappa value of 0.84.
Conclusion: To ensure elimination and prevention of schistosomiasis in humans, the use of validated serological diagnostics such as recombinant antigen ELISA is preferable for field detection in animals, especially in resource-limited areas.
{"title":"Seroprevalence of Schistosoma japonicum Infection Among Dogs and Water Buffaloes Using Recombinant Antigen ELISA in New Corella, Davao del Norte, Philippines.","authors":"Jose Ma M Angeles, Joseph Romeo O Paner, Elena A Villacorte, Pilarita T Rivera, Shin-Ichiro Kawazu","doi":"10.1007/s11686-024-00929-5","DOIUrl":"https://doi.org/10.1007/s11686-024-00929-5","url":null,"abstract":"<p><strong>Purpose: </strong>Animal reservoirs are important targets for controlling and eliminating schistosomiasis. Prevalence studies showed that dogs (Canis familiaris) and water buffaloes (Bubalus bubalis) are important reservoirs of S. japonicum. Previous studies have validated the use of the recombinant proteins Sj1TR, Sj7TR, and SjTPx-1 in ELISA as diagnostics for dogs and water buffaloes from schistosomiasis-endemic areas. In this study, we aimed to determine the seroprevalence of S. japonicum among dogs and water buffaloes in New Corella, Davao del Norte, Philippines, using the recombinant proteins Sj1TR, Sj7TR, and SjTPx-1 in ELISA format.</p><p><strong>Methods: </strong>Fecal and serum samples were collected from dogs (n = 63) and water buffaloes (n = 57). Schistosome-positive samples were detected using formalin-ether concentration technique (FECT), stool polymerase chain reaction, and enzyme-linked immunosorbent assay (ELISA) using soluble egg antigen (SEA), rSj1TR, rSj7TR, and rSjTPx-1. Positivity rates, sensitivity, specificity, predictive values, accuracy, and kappa values were calculated.</p><p><strong>Results: </strong>Mean positivity rates for schistosome infection were high for both dogs (x = 15.40%) and water buffaloes (x = 6.32%). On dogs, the sensitivity and specificity of the tests are as follows: 66.67% and 100% for rSj7TR-ELISA, while 100% and 93.33% for rSjTPx-1-ELISA, respectively. rSjTPx-1-ELISA showed the highest agreement with stool PCR among all diagnostic tests, with an overall kappa value of 0.824. On water buffaloes, the sensitivity and specificity of both rSj1TR-ELISA and rSjTPx-1-ELISA are 100.0% and 98.15%. Both tests also had an overall kappa value of 0.84.</p><p><strong>Conclusion: </strong>To ensure elimination and prevention of schistosomiasis in humans, the use of validated serological diagnostics such as recombinant antigen ELISA is preferable for field detection in animals, especially in resource-limited areas.</p>","PeriodicalId":6932,"journal":{"name":"Acta Parasitologica","volume":null,"pages":null},"PeriodicalIF":1.2,"publicationDate":"2024-09-27","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142339031","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"农林科学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2024-09-23DOI: 10.1007/s11686-024-00897-w
Sowmya R Prabhu, Akshay Pramod Ware, Kapaettu Satyamoorthy, Abdul Vahab Saadi
Background: The rise in Plasmodium resistant strains, decreasing susceptibility to first-line combination therapies, and inadequate efficacy shown by vaccines developed to date necessitate innovative approaches to combat malaria. Drug repurposing refers to finding newer indications for existing medications that provide significant advantages over de novo drug discovery, leading to rapid treatment options. Growing evidence suggests that drugs could regulate the expression of disease-associated microRNAs (miRNAs), implying the potential of miRNAs as attractive targets of therapy for several diseases.
Methods: We aimed to computationally predict drug-disease relationships through miRNAs for the potential repurposing of the drugs as antimalarials. To achieve this, we created a model that combines experimentally validated miRNA-drug interactions and miRNA-disease correlations, assuming that drugs will be linked to disease if they share significant miRNAs. The first step involved constructing a network of drug-drug interactions using curated drug-miRNA relations from the Pharmaco-miR and SM2miR databases. Additionally, the drug-disease relations were acquired from the comparative toxicogenomics database (CTD), and the random walk with restart (RWR) algorithm was applied to the interaction network to anticipate newer drug indications. Further, experimentally verified miRNA-disease associations were procured from the human microRNA disease database (HMDD), followed by an evaluation of the model's performance by examining case studies retrieved from the literature.
Results: Topological network analysis revealed that beta-adrenergic drugs in the network that are closely linked may have a tendency to be used as antimalarials. Case studies retrieved from the literature demonstrated acceptable model performance. A few of the predicted drugs, namely, propranolol, metoprolol, epinephrine, and atenolol, have been evaluated for their association with malaria, thereby indicating the adequacy of our model and offering experimental leads for alternative drugs.
Conclusion: The study puts forth a computational model for forecasting potential connections between beta-adrenergic receptor targeting drugs and malaria to suggest potential for future drug repurposing. This takes into account the concept of commonly associated miRNA partners and providing a mechanistic basis for targeting diseases, elucidating the implication of miRNAs in novel drug-disease relations.
{"title":"MicroRNA Guided In Silico Drug Repositioning for Malaria.","authors":"Sowmya R Prabhu, Akshay Pramod Ware, Kapaettu Satyamoorthy, Abdul Vahab Saadi","doi":"10.1007/s11686-024-00897-w","DOIUrl":"https://doi.org/10.1007/s11686-024-00897-w","url":null,"abstract":"<p><strong>Background: </strong>The rise in Plasmodium resistant strains, decreasing susceptibility to first-line combination therapies, and inadequate efficacy shown by vaccines developed to date necessitate innovative approaches to combat malaria. Drug repurposing refers to finding newer indications for existing medications that provide significant advantages over de novo drug discovery, leading to rapid treatment options. Growing evidence suggests that drugs could regulate the expression of disease-associated microRNAs (miRNAs), implying the potential of miRNAs as attractive targets of therapy for several diseases.</p><p><strong>Methods: </strong>We aimed to computationally predict drug-disease relationships through miRNAs for the potential repurposing of the drugs as antimalarials. To achieve this, we created a model that combines experimentally validated miRNA-drug interactions and miRNA-disease correlations, assuming that drugs will be linked to disease if they share significant miRNAs. The first step involved constructing a network of drug-drug interactions using curated drug-miRNA relations from the Pharmaco-miR and SM2miR databases. Additionally, the drug-disease relations were acquired from the comparative toxicogenomics database (CTD), and the random walk with restart (RWR) algorithm was applied to the interaction network to anticipate newer drug indications. Further, experimentally verified miRNA-disease associations were procured from the human microRNA disease database (HMDD), followed by an evaluation of the model's performance by examining case studies retrieved from the literature.</p><p><strong>Results: </strong>Topological network analysis revealed that beta-adrenergic drugs in the network that are closely linked may have a tendency to be used as antimalarials. Case studies retrieved from the literature demonstrated acceptable model performance. A few of the predicted drugs, namely, propranolol, metoprolol, epinephrine, and atenolol, have been evaluated for their association with malaria, thereby indicating the adequacy of our model and offering experimental leads for alternative drugs.</p><p><strong>Conclusion: </strong>The study puts forth a computational model for forecasting potential connections between beta-adrenergic receptor targeting drugs and malaria to suggest potential for future drug repurposing. This takes into account the concept of commonly associated miRNA partners and providing a mechanistic basis for targeting diseases, elucidating the implication of miRNAs in novel drug-disease relations.</p>","PeriodicalId":6932,"journal":{"name":"Acta Parasitologica","volume":null,"pages":null},"PeriodicalIF":1.2,"publicationDate":"2024-09-23","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142278613","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"农林科学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2024-09-17DOI: 10.1007/s11686-024-00918-8
Jack Ingelbrecht, Karissa O. Lear, Alan J. Lymbery, Rebecca L. Bateman, Bradley M. Norman, Storm B. Martin, Travis Fazeldean, David L. Morgan
Purpose
This study examined the metazoan ectoparasites of the Critically Endangered giant shovelnose ray, Glaucostegus typus, in the eastern Indian Ocean.
Methods
We screened 186 G. typus for ectoparasites in four coastal regions of Western Australia between 2020 and 2022: the Pilbara Region, Exmouth Gulf, Ningaloo Coast and Shark Bay.
Results
Five parasite taxa were encountered on 186 G. typus: Caligus furcisetifer (Copepoda: Caligidae), Dermopristis cairae (Monopisthocotyla: Microbothriidae), Branchellion plicobranchus and Stibarobdella macrothela (Hirudinida: Piscicolidae), and praniza larvae of unidentified gnathiid isopod/s (Isopoda: Gnathiidae). Two of these species, B. plicobranchus and S. macrothela, are reported for the first time on G. typus. Only C. furcisetifer and S. macrothela were relatively common, encountered on 31% and 40% of G. typus, respectively. Gnathiids were observed infrequently, encountered on 13% of G. typus, and D. cairae and B. plicobranchus were scarce, encountered on 1% and 2% of G. typus, respectively. Intensity of infection for C. furcisetifer and gnathiids increased with host length. Likelihood of infection varied seasonally for C. furcisetifer, being considerably lower in summer, and regionally for gnathiids, being greatest at Shark Bay. Intensity and likelihood of infection for S. macrothela increased with host length and varied regionally, being greatest at Shark Bay.
Conclusion
These findings improve our understanding of the downstream impacts for dependent parasites that might arise should populations of G. typus continue to decline.
本研究考察了东印度洋极度濒危巨型锹鼻魟(Glaucostegus typus)的元虫类体外寄生虫:结果在 186 个典型鳕鱼身上发现了五种寄生类群:Caligus furcisetifer(桡足纲:Caligidae)、Dermopristis cairae(单鳃纲:Microbothriidae)、Branchellion plicobranchus 和 Stibarobdella macrothela(蛭形目:Piscicolidae),以及未确定的鮨科异足目(等足目:鮨科)的 praniza 幼虫。其中两个物种,B. plicobranchus 和 S. macrothela,是首次在 G. typus 上发现。只有 C. furcisetifer 和 S. macrothela 比较常见,分别在 31% 和 40% 的 G. typus 上发现。栉水母很少被观察到,仅在 13% 的 G. typus 上发现过,而 D. cairae 和 B. plicobranchus 则很少见,分别在 1% 和 2% 的 G. typus 上发现过。C.furcisetifer和gnathiids的感染强度随宿主长度而增加。糠虾的感染几率因季节而异,夏季低得多,贻贝的感染几率也因地区而异,鲨鱼湾的感染几率最大。这些发现加深了我们的理解,即如果典型海龟的数量继续减少,可能会对依附寄生虫造成的下游影响。
{"title":"Ectoparasites of the Critically Endangered Giant Shovelnose Ray Glaucostegus typus in the Eastern Indian Ocean, with a Summary of the Known Metazoan Parasites","authors":"Jack Ingelbrecht, Karissa O. Lear, Alan J. Lymbery, Rebecca L. Bateman, Bradley M. Norman, Storm B. Martin, Travis Fazeldean, David L. Morgan","doi":"10.1007/s11686-024-00918-8","DOIUrl":"https://doi.org/10.1007/s11686-024-00918-8","url":null,"abstract":"<h3 data-test=\"abstract-sub-heading\">Purpose</h3><p>This study examined the metazoan ectoparasites of the Critically Endangered giant shovelnose ray, <i>Glaucostegus typus</i>, in the eastern Indian Ocean.</p><h3 data-test=\"abstract-sub-heading\">Methods</h3><p>We screened 186 <i>G</i>. <i>typus</i> for ectoparasites in four coastal regions of Western Australia between 2020 and 2022: the Pilbara Region, Exmouth Gulf, Ningaloo Coast and Shark Bay.</p><h3 data-test=\"abstract-sub-heading\">Results</h3><p>Five parasite taxa were encountered on 186 <i>G</i>. <i>typus</i>: <i>Caligus furcisetifer</i> (Copepoda: Caligidae), <i>Dermopristis cairae</i> (Monopisthocotyla: Microbothriidae), <i>Branchellion plicobranchus</i> and <i>Stibarobdella macrothela</i> (Hirudinida: Piscicolidae), and praniza larvae of unidentified gnathiid isopod/s (Isopoda: Gnathiidae). Two of these species, <i>B. plicobranchus</i> and <i>S. macrothela</i>, are reported for the first time on <i>G. typus</i>. Only <i>C</i>. <i>furcisetifer</i> and <i>S</i>. <i>macrothela</i> were relatively common, encountered on 31% and 40% of <i>G</i>. <i>typus</i>, respectively. Gnathiids were observed infrequently, encountered on 13% of <i>G</i>. <i>typus</i>, and <i>D</i>. <i>cairae</i> and <i>B. plicobranchus</i> were scarce, encountered on 1% and 2% of <i>G</i>. <i>typus</i>, respectively. Intensity of infection for <i>C</i>. <i>furcisetifer</i> and gnathiids increased with host length. Likelihood of infection varied seasonally for <i>C. furcisetifer</i>, being considerably lower in summer, and regionally for gnathiids, being greatest at Shark Bay. Intensity and likelihood of infection for<i> S</i>. <i>macrothela</i> increased with host length and varied regionally, being greatest at Shark Bay.</p><h3 data-test=\"abstract-sub-heading\">Conclusion</h3><p>These findings improve our understanding of the downstream impacts for dependent parasites that might arise should populations of <i>G</i>. <i>typus</i> continue to decline.</p>","PeriodicalId":6932,"journal":{"name":"Acta Parasitologica","volume":null,"pages":null},"PeriodicalIF":1.5,"publicationDate":"2024-09-17","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142258280","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"农林科学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2024-09-15DOI: 10.1007/s11686-024-00922-y
Lórena Maciel Santos Silva, Camenas Vieira Barata, Weslania Souza Inacio da Silva, Manuel Benicio Oliveira Neto, Matheus Resende Oliveira, Allan Costa Gomes, André Mota Alves, Janilene de Oliveira Nascimento, Victor Fernando Santana Lima
Purpose
This study aimed to investigate the occurrence of leishmaniasis in domestic rabbits (Oryctolagus cuniculus) in the state of Sergipe, Brazil, and to evaluate the associated clinical signs.
Methods
A total of 31 rabbits from urban and rural areas were clinically examined using cytological, immunological, and serological tests. Blood and cytological samples were collected and analysed for the presence of Leishmania parasites and antibodies. Immunochromatographic tests were used to screen for anti-Leishmania antibodies, and cytological analysis of skin lesions was performed to detect the presence of Leishmania amastigotes.
Results
Of the rabbits tested, 19.35% were reactive in the anti-Leishmania antibody screening, and 3.33% tested positive for Leishmania amastigotes in skin lesion cytology. Clinical signs included cachexia, lymphadenomegaly, dehydration, apathy, dermatitis, ophthalmopathy, and alopecia. Cytological analysis revealed pyogranulomatous inflammation with Leishmania amastigotes present. The findings suggest that leishmaniasis is present among domestic rabbits in this region
Conclusion
This study demonstrates the presence of leishmaniasis in domestic rabbits in the Northeast of Brazil. The findings underline the importance of early diagnosis and intervention in preventing the spread of the disease, and highlight the need for further research into the role of rabbits as potential reservoirs of Leishmania.
{"title":"New Insights into Leishmaniasis in Rabbits Raised in the Northeast Region of Brazil","authors":"Lórena Maciel Santos Silva, Camenas Vieira Barata, Weslania Souza Inacio da Silva, Manuel Benicio Oliveira Neto, Matheus Resende Oliveira, Allan Costa Gomes, André Mota Alves, Janilene de Oliveira Nascimento, Victor Fernando Santana Lima","doi":"10.1007/s11686-024-00922-y","DOIUrl":"https://doi.org/10.1007/s11686-024-00922-y","url":null,"abstract":"<h3 data-test=\"abstract-sub-heading\">Purpose</h3><p>This study aimed to investigate the occurrence of leishmaniasis in domestic rabbits (<i>Oryctolagus cuniculus</i>) in the state of Sergipe, Brazil, and to evaluate the associated clinical signs.</p><h3 data-test=\"abstract-sub-heading\">Methods</h3><p>A total of 31 rabbits from urban and rural areas were clinically examined using cytological, immunological, and serological tests. Blood and cytological samples were collected and analysed for the presence of Leishmania parasites and antibodies. Immunochromatographic tests were used to screen for anti-Leishmania antibodies, and cytological analysis of skin lesions was performed to detect the presence of Leishmania amastigotes.</p><h3 data-test=\"abstract-sub-heading\">Results</h3><p>Of the rabbits tested, 19.35% were reactive in the anti-Leishmania antibody screening, and 3.33% tested positive for Leishmania amastigotes in skin lesion cytology. Clinical signs included cachexia, lymphadenomegaly, dehydration, apathy, dermatitis, ophthalmopathy, and alopecia. Cytological analysis revealed pyogranulomatous inflammation with Leishmania amastigotes present. The findings suggest that leishmaniasis is present among domestic rabbits in this region</p><h3 data-test=\"abstract-sub-heading\">Conclusion</h3><p>This study demonstrates the presence of leishmaniasis in domestic rabbits in the Northeast of Brazil. The findings underline the importance of early diagnosis and intervention in preventing the spread of the disease, and highlight the need for further research into the role of rabbits as potential reservoirs of Leishmania.</p>","PeriodicalId":6932,"journal":{"name":"Acta Parasitologica","volume":null,"pages":null},"PeriodicalIF":1.5,"publicationDate":"2024-09-15","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142258281","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"农林科学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2024-09-06DOI: 10.1007/s11686-024-00869-0
Nicol Bernardová, Jan Novák, Petr Horák, Chia-Kwung Fan, Libuše Kolářová
Toxocara canis larvae are one of the most overlooked agents of nervous system infection in paratenic hosts. Previous studies in mouse models have shown that infection with various (mainly high) numbers of larvae leads to neurobehavioral disturbances and pathological changes. Our study investigated whether the infection with low and moderate numbers of larvae could affect the physical condition, motor skills, and pathogenesis in the brains of experimentally infected mice.Two groups of BALB/c mice were orally infected with 10 and 100 T. canis larvae per animal and examined regularly until the 97th week after infection. General appearance, specific antibody responses, and motor/balance skills were assessed. The number and viability of larvae in the liver, spleen, lungs, and brain were assessed by quantitative compressed biopsy technique, while the pathological changes of the brain infection were studied histologically.As a result, changes were observed in overall appearance, activity, as well as motor and balance ability. The infections were associated with an increased IgG antibody response to the specific anti-T. canis excretory/secretory antigen and tissue damage in the brain characterized by necrosis, cell infiltrations, including foamy cells, and hemorrhages.The study demonstrated the effects of low and moderate T. canis infection in a paratenic host during the chronic phase of infection, which lasted up to 97 weeks for the first time.
{"title":"Neurobehavioral Disorders and Pathological Changes in the Brain of Mice Are Caused by Chronic Toxocara canis Larval Invasion with Low to Moderate Inoculum.","authors":"Nicol Bernardová, Jan Novák, Petr Horák, Chia-Kwung Fan, Libuše Kolářová","doi":"10.1007/s11686-024-00869-0","DOIUrl":"https://doi.org/10.1007/s11686-024-00869-0","url":null,"abstract":"<p><p>Toxocara canis larvae are one of the most overlooked agents of nervous system infection in paratenic hosts. Previous studies in mouse models have shown that infection with various (mainly high) numbers of larvae leads to neurobehavioral disturbances and pathological changes. Our study investigated whether the infection with low and moderate numbers of larvae could affect the physical condition, motor skills, and pathogenesis in the brains of experimentally infected mice.Two groups of BALB/c mice were orally infected with 10 and 100 T. canis larvae per animal and examined regularly until the 97th week after infection. General appearance, specific antibody responses, and motor/balance skills were assessed. The number and viability of larvae in the liver, spleen, lungs, and brain were assessed by quantitative compressed biopsy technique, while the pathological changes of the brain infection were studied histologically.As a result, changes were observed in overall appearance, activity, as well as motor and balance ability. The infections were associated with an increased IgG antibody response to the specific anti-T. canis excretory/secretory antigen and tissue damage in the brain characterized by necrosis, cell infiltrations, including foamy cells, and hemorrhages.The study demonstrated the effects of low and moderate T. canis infection in a paratenic host during the chronic phase of infection, which lasted up to 97 weeks for the first time.</p>","PeriodicalId":6932,"journal":{"name":"Acta Parasitologica","volume":null,"pages":null},"PeriodicalIF":1.2,"publicationDate":"2024-09-06","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142138872","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"农林科学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}