Evaluating the effect of tuf and secA gene sequence length for discrimination of phytoplasmas

Abstract Background In the present study, phylogenetic relationships within Heterobranchia in particular to Pulmonata were evaluated by means of Histone-3 (H3) gene sequence information. H3 gene is a slow evolving marker and is useful in resolving the deep level relationships. This is the first study to report the phylogeny of Pulmonata with more number of representatives from the group on the basis of H3 gene. Results The major groups within Heterobranchia viz. Lower Heterobranchia, Opisthobranchia, and Pulmonata were non-monophyletic. A few of the pulmonate groups’ viz. Planorbidae, Lymnaeidae, Siphonariidae, Veronicellidae, and Stylommatophora were recovered as monophyletic. The concepts of Eupulmonata and Geophila were not observed in the present study. Conclusions The present study was undertaken with an objective to study the phylogeny of Pulmonata reconstructed on the basis of H3 gene and its ability to resolve the deeper divergences in Pulmonata. However, the resolution at the deeper nodes is limited. There is a good resolution at the level of genera. In the future, inclusion of more number of taxa with increased sequence length of H3 marker may yield resolved topologies that may shed more insights into the phylogeny of Pulmonata.

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First report of 16SrI-B subgroup related phytoplasma associated with witches’-broom symptoms in Pericampylus glaucus in China

Plant Disease ◽

10.1094/pdis-09-20-2026-pdn ◽

2020 ◽

Author(s):

Shao-shuai Yu ◽

Qinghua Tang ◽

Yuan Wu ◽

Rui-ling Zhao ◽

Wei wei Song ◽

...

Keyword(s):

16S Rrna ◽

Gene Sequence ◽

Hainan Island ◽

Rrna Gene ◽

Yellow Leaf ◽

Active Components ◽

Plant Resource ◽

Phytoplasma Strain ◽

Ctab Method ◽

Seca Gene

Pericampylus glaucus is an important medicinal plant resource containing active components with potential antitumor activity in China (Zhao & Cui, 2009). During July through August 2020, plants displayed disease symptoms including “witches’ broom”, leaf chlorosis, leaflet and internode shortening that impacted their growth (Fig. 1). These plants were first found in Dingan county of Hainan province, China. Total DNA from 12 plants were extracted using 0.10 g fresh plant leaves based on CTAB method. After amplification using primers specific for phytoplasma 16S rRNA, tuf and secA gene targets, R16mF2R16mR1 (Lee et al, 1993), fTuf1/rTuf1 (Schneider et al., 1997) and secAfor1/secArev3 (Hodgetts et al., 2008), the target bands of the three gene fragments of phytoplasma were detected in the disease sample DNA from six disease plants, and not in the healthy sample DNA from six healthy plants. Nucleotide sequences of the three genes were obtained from the PCR products sequencing and analyzed by DNAMAN 5.0 software. The three gene fragments of the DNA extracted from the disease samples were identical, with length of 1334 bp 16S rRNA (GenBank accession: MT872515), 989 bp tuf (MT755960) and 750 bp secA (MT755961) gene fragments, putatively encoding 329 (tuf) and 249 (secA) amino acids sequence separately. The phytoplasma strain was named as Pericampylus glaucus witches’-broom (PgWB) phytoplasma, PgWB-hnda strain, belonging to 16SrI-B subgroup by iPhyClassifier analysis. Homology and phylogenetic analysis indicated that based on 16S rRNA gene fragments, PgWB-hnda, pepper yellow crinkle phytoplasma PYC-hnhk (MT760793), chinaberry witches’-broom phytoplasma CWB-hnsy1 (KP662119) and CWB-hn (EF990733), periwinkle virescence phytoplasma PeV-hnhk (KP662136), with 100.0 % identity value, arecanut yellow leaf phytoplasma AYL-hnwn (FJ998269) and AYL-hn (FJ694685), with 99.8 % identity value, were clustered into one clade. Based on the analysis of tuf gene sequence fragments, PgWB was closely related to PYC-hnhk (MT755960), CWB-hnsy1 (KP662155), PeV-hnhk (KP662172) with 99.9 % identity value. Based on the analysis of secA gene sequence fragments, PgWB was closely related to CWB-hnsy1 (KP662173) with 99.7 % identity value, PYC-hnhk (MT755961), PeV-hnhk (KP662190) with 99.4 % identity value. To our knowledge, this is the first time that Pericampylus glaucus witches’-broom disease caused by 16SrI-B subgroup phytoplasma strain was found in China. Multilocus sequence analysis showed that PgWB was closely related to the phytoplasma strains causing pepper yellow crinkle, chinaberry witches’-broom, periwinkle virescence and areca palm yellow leaf diseases, all occurred in Hainan Island of China.

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Tephrosia purpurea Represents a New Host of 16SrII-V Subgroup Phytoplasma Associated with Witches’-Broom Disease in China

Plant Disease ◽

10.1094/pdis-02-21-0357-pdn ◽

2021 ◽

Author(s):

Shao-shuai Yu ◽

Rui-ling Zhao ◽

Ming-xing Lin ◽

Yuan Wu ◽

Wei wei Song ◽

...

Keyword(s):

16S Rrna ◽

16S Rrna Gene ◽

Gene Sequence ◽

16S Rrna Gene Sequence ◽

Rrna Gene ◽

Rrna Gene Sequence ◽

Sequence Fragment ◽

Tephrosia Purpurea ◽

Seca Gene ◽

The 16S Rrna Gene

Tephrosia purpurea is a medical plant with excellent insecticidal activity belonging to the family of Leguminosae distributed throughout southern of China (Pei et al., 2013). During January to February 2021, the plants showing abnormal symptoms including witches’-broom, internode shortening, leaf chlorosis and leaflet formation, as shown in Fig.1, were found in Ledong County of Hainan Province, a tropical island in China, with about 60 % incidence. The Tephrosia purpurea disease symptoms were suspected to be induced by phytoplasma, a phloem-limited prokaryotic pathogen which can not be cultured in vitro and which causes severe financial loss and ecological damage to the island. Total DNA from the symptomatic and asymptomatic samples of Tephrosia purpurea were extracted using 0.10 g fresh plant leaves and branches by CTAB method (Doyle and Doyle, 1990). 16S rRNA and secA gene sequence fragments of phytoplasma were detected through PCR amplification using primers R16mF2/R16mR1 (Gundersen and Lee, 1996) and secAfor1/secArev3 (Hodgetts et al., 2008). The two gene sequence fragments of phytoplasma were obtained from the DNA of six symptomatic plant samples whereas not from the DNA of six asymptomatic plant samples. These amplified products were sequenced and the data were deposited in GenBank. The two gene sequence fragments of the DNA obtained from the diseased plant samples were all identical, with a length of 1335 bp for the 16S rRNA (GenBank accession: MW616560) and 729 bp for the secA gene (MW603929). The secA gene fragment putatively encodes for 242 amino acids. The phytoplasma strain was named as Tephrosia purpurea witches’-broom (TpWB) phytoplasma, TpWB-hnld strain. 16S rRNA gene sequence fragment of TpWB-hnld was analyzed by online tool iPhyClassifier (Wei et al., 2007), indicating that the pathogen strain was a member of subgroup 16SrII-V and a ‘Candidatus Phytoplasma aurantifolia’-related strain. Blast analysis based on the 16S rRNA gene sequence fragment of TpWB-hnld showed 100 % sequence identity with that of peanut witches’-broom group members (16SrII group), such as Cassava witches'-broom phytoplasma (KM280679) and Cleome sp. phytoplasma (KM280677); Blast analysis based on the secA gene sequence fragment of TpWB-hnld showed 100 % sequence identity with that of peanut witches’-broom group members (16SrII group), such as sesame phyllody phytoplasma (JN977044). Homology and phylogeny were analyzed using the software of DNAMAN 5.0 and MEGA 7.0, indicating that TpWB-hnld and other subgroup 16SrII-V phytoplasma strains, including Cassava witches'-broom phytoplasma, Cleome sp. phytoplasma, Crotalaria witches'-broom phytoplasma (EU650181) and Desmodium ovalifolium witches'-broom phytoplasma (GU113152), were clustered into one clade with 98 % bootstrap value based on the 16S rRNA gene sequence fragments; TpWB-hnld and sesame phyllody phytoplasma were clustered into one clade based on the secA gene sequence fragments. Multiple alignment based on the 16S rRNA gene sequence fragment showed that the TpWB-hnld phytoplasma strain showed 98 % sequence identity with TpWB phytoplasma strain (HG792252) belonging to 16SrII-M subgroup reported in India (Yadav et al., 2014). To our knowledge, this was the first time that 16SrII-V subgroup phytoplasma associated with Tephrosia purpurea witches’-broom disease was identified in China. Molecular analysis based on the 16S rRNA and secA gene sequence fragments indicated that TpWB-hnld phytoplasma was a member of subgroup 16SrII-V and a ‘Candidatus Phytoplasma aurantifolia’-related strain.

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Hepatic Encephalopathy and Liver Failure in Pediatric Patient with hepatitis A

International Journal of Drug Delivery Technology ◽

10.25258/ijddt.v9i3.16 ◽

2019 ◽

Vol 9 (o3) ◽

Author(s):

Shaima’a Dakhel AbdulHassa

Keyword(s):

Hepatic Encephalopathy ◽

Liver Failure ◽

Hepatitis A ◽

Gene Sequence ◽

Stool Examination ◽

Age Group ◽

Significant Difference ◽

Medical City ◽

Sequence Method ◽

Stool Samples

Gairdia lamblia is one of parasites that cause intestinal problems within the human body, particularly private travelers and children. In this study a total of (100) diarrheal patients, 20 patients with Giardiasis were identified by fecal antigen. 9 out of 20(20%) of them were infected by fecal antigen, while 9(9%) of them were infected by using the screening general stool examination (GSE). The stool samples were collected from patient how vested the Medical City/ Baghdad and Tikrit teaching Hospital during the period from 1 st may 2018 to 1 February 2019. The results revealing a significant difference (p andlt; 0.05) between the two methods of detection for G. lamblia (Fecal antigen method and GSE). IT has been shown that out of 20 infected individuals 12(12%) were males and 8(8%) were females, indicating regarding no significant deference in the distribution of Giardiasis among genders. In regard the age, our results showed that highest infection rate 8(3.2%) was recorded in the age group (10-19) years, followed by the age group (20-2) years which was 692.4%). In this study five mutations were recorded at position (926, 1094, 1202and 1304), by using tpiA gene sequence method, and tpiB gene was on point mutation change (G254A), in the position (85) of triose phosphate isomease.

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