scholarly journals Morphological and Molecular Characterization of Paragonimus skrjabini Complex from Yunnan, China: A Brief Report

2021 ◽  
Author(s):  
Qiu-Hong Shu ◽  
Shu-De Li ◽  
Ming Tian ◽  
Yong Meng ◽  
Shu-Mei-Qi He ◽  
...  
Keyword(s):  
Phylogenetic Analysis ◽  
Molecular Identification ◽  
Yunnan Province ◽  
Morphological Identification ◽  
Sprague Dawley ◽  
Parsimony Method ◽  
Maximum Parsimony Method ◽  
Freshwater Crabs ◽  
Sd Rats ◽  
Paragonimus Skrjabini

Abstract Purpose To perform environmental sampling and molecular identification of Paragonimus in endemic regions, which may help in minimizing transmission among humans. Methods Mountain crabs from the genus Potamiscus were collected and the encysted metacercariae were extracted and subjected to morphological identification, followed by animal inoculation in Sprague–Dawley (SD) rats. After 112 days of infection, animals were killed and adult worms were extracted from lungs and muscles. The morphology of adult worms was characterized by microscopy and molecular identification was done by polymerase chain reaction, followed by sequencing of cox1 and ITS2 genes. Phylogenetic analysis was done by maximum parsimony method. Results A total of 447 crabs were captured from the streams of Tongchang Town, Jinping County, Yunnan Province, China. The infection rate was found to be 41% (186 out of 447 crabs). The metacercariae of Paragonimus skrjabini was identified by the characteristics round or spherical encysted form measuring 410 to 460 × 400 to 460 µm. After animal infection in SD rats, adults were presumptively confirmed to be P. skrjabini, which was also confirmed by gene amplification and sequence analysis of cox1 and ITS2 regions. Paragonimus skrjabini clustered with previously reported P. skrjabini from Yunnan and Vietnam. The confidence values of their branches were > 95%. Phylogenetic analysis of the ITS2 region revealed two distinct clusters with distinct geographical grouping. Phylogenetic analysis with the combined data sets reiterated the geographical grouping with P. skrjabini from Yunnan clustering with strains from Vietnam. Conclusion Metacercariae of P. skrjabini was discovered in freshwater crabs in Yunnan province, China, and the strains were phylogenetically related to P. skrjabini from Vietnam.

scholarly journals Morphological and Molecular Characterization of Paragonimus Species Isolated from Freshwater Crabs in Southern Yunnan, China

2021 ◽  
Vol 2021 ◽  
pp. 1-11
Author(s):  
Qiu-Hong Shu ◽  
Yang Yang ◽  
Miao-Miao Wang ◽  
Shu-De Li ◽  
Ming Tian ◽  
...  
Keyword(s):  
Phylogenetic Analysis ◽  
Phylogenetic Tree ◽  
Molecular Characterization ◽  
Yunnan Province ◽  
Sprague Dawley ◽  
Natural Habitats ◽  
Freshwater Crabs ◽  
Multiple Organs ◽  
Locus Method ◽  
Paragonimus Skrjabini

Paragonimus species are highly prevalent in various regions of China. The study’s objective is to isolate and identify Paragonimus from natural habitats and compare the phylogenetic diversity of Paragonimus in southern Yunnan province, China. Metacercariae of Paragonimus was isolated from crabs, and morphologic identification was performed by microscopy. Metacercariae were injected into experimental Paragonimus free Sprague Dawley rats. After 114 days, adult worms and eggs were isolated from multiple organs. Morphologic identification confirmed the initial identification. DNA was extracted from 5 adult worms, and molecular characterization was performed by amplification and sequencing of CO1 and ITS2 regions, followed by phylogenetic analysis. Out of 447 crabs captured, 186 crabs were found to be infected. A total of 4 species of Paragonimus was observed from naturally infected crabs. Paragonimus microrchis (2), Paragonimus heterotremus (1), Paragonimus proliferus (1), and Paragonimus skrjabini (1) were isolated and identified. A total of 32 sequences were downloaded from the National Center for Biotechnology Information, and 5 sequences generated in the study were used for phylogenetic analysis. In the phylogenetic tree of the CO1 gene, Paragonimus proliferus, Paragonimus heterotremus, and Paragonimus skrjabini were clustered with the same species, and the confidence values of their branches were >95%. A congruent phylogenetic relationship was observed with the ITS2 phylogenetic tree. In the phylogenetic tree constructed with the combined dataset of CO1 and ITS2 datasets, Paragonimus proliferus, Paragonimus heterotremus, and Paragonimus skrjabini clustered with the same species, and their branch confidence values were >94%. Paragonimus microrchis clustered with Paragonimus bangkokensis in both datasets. Phylogenetic analysis revealed robustness of the double loci method as against the single-locus method with either CO1 or ITS2 alone. Paragonimus species isolated from the southern Yunnan province, China, was phylogenetically diverse, and the analysis revealed the clustering of multiple species of Paragonimus isolated from different geographic locations.

scholarly journals Morphological and molecular characterization of Paragonimus species isolated from freshwater crabs in Yunnan, China.

2020 ◽  
Author(s):  
Qiu-Hong Shu ◽  
Shu-De Li ◽  
Ming Tian ◽  
Wen-Wei Bai ◽  
Yong Meng ◽  
...  
Keyword(s):  
Phylogenetic Analysis ◽  
Molecular Characterization ◽  
Phylogenetic Diversity ◽  
Yunnan Province ◽  
Natural Habitat ◽  
Geographic Location ◽  
Sprague Dawley ◽  
Freshwater Crabs ◽  
Multiple Organs ◽  
Locus Method

Abstract Background Paragonimus species are highly prevalent in ethnic minority areas of Yunnan, China. The objective of the study is to isolate and identify Paragonimus from natural habitat and compare the phylogenetic diversity of Paragonimus in Yunnan province, China. Methods Metacercariae of Paragonimus was isolated from crabs and morphologic identification was performed by microscopy. Metacercariae were injected into experimental Paragonimus free Sprague Dawley rats. After 112 days, adult worms and eggs were isolated from multiple organs. Morphologic identification confirmed the initial identification. DNA was extracted from the adult worms and molecular characterization was performed by amplification and sequencing of CO1 and ITS2 regions followed by phylogenetic analysis. Results Out of 447 crabs captured, 186 crabs were found to be infected. A total of 5 occurrences of Paragonimus was observed from naturally infected crabs. Paragonimus microrchis (2), P,heterotremus (1), P.proliferus (1) and P.skrjabini (1) were isolated and identified. A total of 32 sequences downloaded from national center for biotechnology information and 5 sequences generated in the study was used for phylogenetic analysis. Phylogenetic analysis revealed robustness of the double loci method as against the single locus method with either CO1 or ITS2 alone. Considerable phylogenetic diversity depending on the geographic location was observed. Conclusion Paragonimus species isolated from Yunnan province, China were phylogenetically diverse and the analysis revealed the clustering of multiple species of Paragonimus isolated from different geographic locations.

scholarly journals Molecular record for the first authentication of Isaria cicadae from Vietnam

2021 ◽  
Vol 16 (1) ◽  
pp. 711-718
Author(s):  
Thuan Duc Lao ◽  
Hanh Van Trinh ◽  
Loi Vuong ◽  
Luyen Tien Vu ◽  
Thuy Ai Huyen Le ◽  
...  
Keyword(s):  
Phylogenetic Analysis ◽  
Genomic Dna ◽  
Morphological Analysis ◽  
Phylogenetic Analyses ◽  
Neighbor Joining ◽  
Pcr Assay ◽  
Parsimony Method ◽  
Maximum Parsimony Method ◽  
Cordyceps Cicadae ◽  
High Bootstrap

Abstract The entomopathogenic fungus T011, parasitizing on nymph of Cicada, collected in the coffee garden in Dak Lak Province, Vietnam, was preliminarily morphologically identified as Isaria cicadae, belonged to order Hypocreales and family Clavicipitaceae. To ensure the authenticity of T011, phylogenetic analysis of the concatenated set of multiple genes including ITS, nrLSU, nrSSU, Rpb1, and Tef1 was applied to support the identification. Genomic DNA was isolated from dried sample T011. The PCR assay sequencing was applied to amplify ITS, nrLSU, nrSSU, Rpb1, and Tef1 gene. For phylogenetic analysis, the concatenated data of both target gens were constructed with MEGAX with a 1,000 replicate bootstrap based on the neighbor-joining, maximum likelihood, maximum parsimony method. As the result, the concatenated data containing 62 sequences belonged to order Hypocreales, families Clavicipitaceae, and 2 outgroup sequences belonged to order Hypocreales, genus Verticillium. The phylogenetic analysis results indicated that T011 was accepted at subclade Cordyceps and significantly formed the monophyletic group with referent Cordyceps cicadae (Telemorph of Isaria cicadae) with high bootstrap value. The phylogenetically analyzed result was strongly supported by our morphological analysis described as the Isaria cicadae. In summary, phylogenetic analyses based on the concatenated dataset were successfully applied to strengthen the identification of T011 as Isaria cicadae.

Phylogenetic Analysis of Eight Malaysian Pineapple Cultivars using a Chloroplastic Marker (rbcL gene)

2013 ◽  
Vol 64 (2) ◽  
Author(s):  
Norfadilah Hamdan ◽  
Azman Abd Samad ◽  
Topik Hidayat ◽  
Faezah Mohd Salleh
Keyword(s):  
Phylogenetic Analysis ◽  
Sequence Similarity ◽  
Accurate Method ◽  
Ananas Comosus ◽  
Rbcl Gene ◽  
Ribulose Bisphosphate Carboxylase ◽  
Ribulose Bisphosphate ◽  
Parsimony Method ◽  
Bisphosphate Carboxylase ◽  
Maximum Parsimony Method

To date, Malaysian pineapple cultivars has only been characterized morphologically. A more consistent and accurate method such as biomarker is highly crucial to distinguish and establish the genetic relationship between different cultivars. In this work, we conducted a phylogenetic analysis of eight Malaysian pineapple cultivars using a chloroplastic DNA biomarker, ribulose-bisphosphate carboxylase (rbcL) gene. The rbcL gene was isolated from genomic DNA, amplified and sequenced. The rbcL gene of Ananas comosus is approximately 1100 bp. From the multialignment of eight cultivars, the percentage of sequence similarity ranged from 71.1% to 94.98% and is highly conserved throughout the sequences. Phylogenetic analysis which is carried out using maximum parsimony method revealed that the eight Malaysian pineapple cultivars can be classified into two groups. The first group consist of Yankee and Gandul cultivars while Moris, Moris Bentanggur, Moris Gajah, N36, Josaphine and Sarawak falls under the second group. Bootstrap values in some branches are low which reflect the small number of informative characters (981 are conserved, 12 are potentially informative). Formation of several group or subclades is due to its similar genetic pattern, thus supporting this classification. This study confirmed that rbcL gene is a good indicator to determine the phylogenetic relationship distinguishing the Malaysian pineapple cultivars.

scholarly journals Molecular identification and phylogenetic analysis of Chlorella isolates from Indonesia using rbcL gene

2021 ◽  
Vol 89 (1) ◽  
Author(s):  
Fauziatul FITRIYAH ◽  
Yora FARAMITHA ◽  
Dini Astika SARI ◽  
Irma KRESNAWATY ◽  
Tri PANJI ◽  
...  
Keyword(s):  
Phylogenetic Analysis ◽  
Molecular Identification ◽  
Morphological Identification ◽  
Rbcl Gene ◽  
Green Microalgae ◽  
Bisphosphate Carboxylase ◽  
Nannochloropsis Gaditana ◽  
Pcr Product ◽  
Ribulose 1 ◽  
Commercial Applications

Identifying the newly isolated species is crucial to establishing a reliable algal database with successful commercial applications for different biotechnological applications. Morphological identification does not give sufficient description, especially for tiny unicellular microalgae. The rbcL gene encodes the large unit of ribulose-1, 5-bisphosphate carboxylase /oxygenase (Rubisco) has been widely known for barcoding in plants and developed for microalgae molecular identification. In this study, we examined the local strains of green microalgae from Indonesia using the rbcL partial gene sequence to identify the strains. Green microalgae isolates originated from Yogyakarta, Serayu, Gondol, Ancol, Cilegon, and Teluk Jakarta were cultured in f/2 media and harvested for DNA extraction. The DNA extracted was proceeded to PCR using 1AB_rbcL primer pair to amplify the sequences of rbcL gene with target band located at 582 bp, followed by the sequencing of the PCR product was conducted. Molecular identification of local green microalgae isolates was successfully carried out using primers 1AB_rbcL with a genetic similarity of 99% toward identified species in the NCBI database. Among six isolates, TJ, G, S, C, and A isolates were identified as C. pyrenoidosa. Only CP isolate from Yogyakarta identified as C. sorokiniana. Nannochloropsis gaditana rbcL sequence was selected as an outgroup. The phylogenetic analysis indicated that the five isolates of Chlorella belong to one clade and clearly distinguished from C. sorokiniana isolate from Yogyakarta.

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