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Genetic insights and phylogenetic relationships of Momordica balsamina using DNA barcoding: An accurate tool for species identification

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GENE REPORTS 2025年 39卷

作者： Aman, Iqra Maqsood, Quratulain Mehmood, Tahir Sumrin, Aleena Shahzad, Rehman Univ Punjab Ctr Appl Mol Biol CAMB Lahore 53700 Punjab Pakistan Univ Punjab Inst Microbiol & Mol Genet IMMG Lahore 54590 Punjab Pakistan Hitech Biotech SMC Pvt Ltd Lahore Pakistan

The Momordica balsamina, a member of Cucurbitaceae family is a plant species of immense botanical, phytochemicals, pharmacological, and medicinal value. Accurate identification is crucial in order to extract valuable bioactive compounds which is important for ensuring the quality and safety of herbal medicines. While traditional taxonomic methods aid in species identification but an innovative and authentic method for identifying species is DNA barcoding, which is based on the nucleotide diversity of conserved sequences. Plant DNA barcoding systems are more difficult to build and refine because of the enormous genetic variation among species. To ensure a more reliable approach, plant scientists should focus on conserved nuclear-transcribed regions to analyze genetic diversity, species differentiation, and phylogeny. In this study, we amplified and sequenced the rbcL, matK, and ITS regions of Momordica balsamina. Furthermore, there phylogenetic tree analysis, bootstrap percentage, sequence homology and failed amplification of matK were discussed. M. balsamina species were authenticated by using ITS primer (internal transcribed spacer). Through this intended research, we aim to genetically identify and characterize this medicinally important species through barcoding.

关键词： Momordica balsamina DNA barcoding rbcL matK and ITS gene phylogenetic tree analysis

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Probiotic potentiality, safety profiling and broad-spectrum antibacterial activity of lactic acid bacteria isolated from sour curd (Malda, India)

Microbe (Netherlands)

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Microbe (Netherlands) 2025年 7卷

作者： Nandi, Suchhanda Mandal, Shyamapada Department of Zoology University of Gour Banga Malda 732103 India

Lactic acid bacteria (LAB) obtained from fermented foods are recognized as probiotics that offer multiple health advantages and disease-preventing characteristics, like antibacterial properties. The employment of effective probiotic bacteria as dietary supplements has attracted considerable research attention because of their beneficial impacts. This study demonstrates the probiotic characteristics, safety aspects, and antibacterial efficacy of lactobacilli sourced from sour curd samples collected from Malda (West Bengal, India) using in vitro methods. The identity of LAB was verified through 16S-rRNA gene sequencing and subsequent analysis of the phylogenetic tree. The antibacterial activity was evaluated using agar well and agar overlay methods against gram-positive (Staphylococcus aureus, Bacillus cereus) and gram-negative (Escherichia coli, Pseudomonas aeruginosa) bacterial pathogens. The study identified six lactobacilli strains that exhibited probiotic features and satisfactory safety profiles. Three of these demonstrated notable broad-spectrum antibacterial effects against gram-positive and gram-negative bacterial pathogens, specifically identified as Levilactobacillus brevis LMEM 401B, Lacticaseibacillus casei LMEM 402 A, and Limosilactobacillus fermentum LMEM 403 C. All isolated lactobacilli displayed cumulative probiotic potentials (CPP) between 78 % and 100 %. Additionally, the probiotic LAB enhanced the action of antibiotics against the indicator bacteria. This study highlights locally available sour curd as a good source of probiotics, which could be used alone or in combination with antibiotics as biotherapeutic agents. © 2025 The Authors

关键词： 16S rRNA gene sequencing Antibacterial activity Lactobacilli phylogenetic tree analysis Probiotics

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Insights into the antibacterial activity of Limosilactobacillus fermentum curd isolate against gram-positive and gram-negative pathogenic bacteria

Pharmacological Research - Reports

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Pharmacological Research - Reports 2025年 3卷

作者： Suchhanda Nandi Shyamapada Mandal Department of Zoology University of Gour Banga Malda 732103 India

It is widely known that lactic acid bacteria (LAB), commonly found in fermented products like curd, extends health benefits and antibacterial properties, particularly in traditional Chinese medicine (TCM). In spite of the growing demand for sweet curd contained in earthen pots in our region, no research has specifically explored its unique properties. This study aims to address this gap by examining the probiotic characteristics, antibacterial activity and consumer appeal of sweet curd in earthen pots. A sweet curd sample containing in an earthen pot was microbiologically cultured to isolate LAB applying conventional methods and 16S rRNA gene sequencing. The strain thus isolated was identified to be Limosilactobacillus fermentum 406B, showed broad-spectrum antibacterial activity against gram-positive: Staphylococcus aureus , Bacillus cereus , Streptococcus pneumoniae and Listeria monocytogenes and gram-negative: Escherichia coli , Pseudomonas aeruginosa , Acinetobacter baumannii and Proteus vulgaris bacterial pathogens. Antimicrobial property and minimum inhibitory concentration (MIC) were assessed by agar well diffusion and broth dilution method, respectively. Initial tests based on colony morphology, gram staining and biochemical assays confirmed it as a Lactobacillus species, with sequencing confirming its identity as Limosilactobacillus fermentum 406B. The strain demonstrated excellent physiological tolerance to NaCl, bile, low pH and varying temperatures, confirming its suitability as a probiotic. Additionally, safety tests showed that it was non-pathogenic, as it tested negative for gelatinase and DNase production and displayed an acceptable antibiotic susceptibility profile. With a cumulative probiotic potential (CPP) of 100 %, L. fermentum 406B holds significant promise as a probiotic strain. These results underscore the strain’s potential not just in TCM but also as a prospective biotherapeutic agent, considering its extensive antibacterial activity and rob

关键词： LAB TCM 16S rRNA gene sequencing phylogenetic tree analysis Antibacterial action CPP

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Molecular cloning and characterization of a taxadienol acetyl transferase cDNA from Taxus x media

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PLANT SCIENCE 2004年第4期167卷 759-764页

作者： Kai, GY Miao, ZQ Qiu, CX Zhang, L Zhao, LX Li, ZG Xu, TF Zhang, LD Gong, YF Zhao, DL Liu, DH Sun, XF Tang, KX Shanghai Jiao Tong Univ Sch Agr & Biol Fudan SJTU Nottingham Plant Biotechnol R&D Ctr Plant Biotechnol Res Ctr Shanghai 200030 Peoples R China Shanghai Jiao Tong Univ Sch Life Sci & Technol Shanghai 200030 Peoples R China Fudan Univ Fudan SJTU Nottingham Plant Biotechnol R&D Ctr Morgan Tan Int Ctr Life Sci Sch Life SciState Key Lab Genet Engn Shanghai 200433 Peoples R China

A full-length cDNA encoding taxadienol acetyl transferase (designated as TmTAT), that catalyzes the first acylation step in Taxol biosynthesis, was cloned from young leaves of Taxus x media by rapid amplification of cDNA ends (RACE). The full-length cDNA of TmTAT was 1,159 bp and contained a 1317-bp open reading frame (ORF) encoding a protein of 439 amino acids. The deduced protein had a calculated molecular weight of about 49 kDa and an isoelectric point (pI) of 5.3, similar to acetyl transferases from Taxus cuspidata and Taxus chinensis. Sequence comparison analysis revealed that the TmTAT had high similarity with other members of plant transferase family. phylogenetic tree analysis showed that TmTAT had close relationship with acetyl transferases from T chinenisis and T cuspidata. Tissue expression pattern analysis revealed that TmTAT expressed strongly in leaves, weak in stems and no expression could be detected in fruits, indicating that TmTAT was a tissue-specific gene. (C) 2004 Elsevier Ireland Ltd. All rights reserved.

关键词： RACE taxadienol acetyl transferase Taxus x media TmTAT phylogenetic tree analysis

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Chaperone-dependent gene expression of organic solvent-tolerant lipase from Pseudomonas aeruginosa strain S5

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PROCESS BIOCHEMISTRY 2010年第3期45卷 346-354页

作者： Baharum, Syarul Nataqain Rahman, Raja Noor Zaliha Raja Abdul Basri, Mahiran Salleh, Abu Bakar Univ Putra Malaysia Fac Biotechnol & Biomol Sci Upm Serdang 43400 Selangor Malaysia Univ Putra Malaysia Fac Sci Upm Serdang 43400 Selangor Malaysia

The gene coding for the intracellular organic solvent-tolerant lipase of Pseudomonas aeruginosa strain S5 was isolated from a genomic DNA library and cloned into pRSET. The cloned sequence included two open reading frames (ORF) of 1575 bp for the first ORF (ORF1), and 582 bp for the second ORF (ORF2). The ORF2, known as chaperone, plays an important role in the expression of the S5 gene. The ORF2 is located downstream of lipase gene, and functions as the act gene for ORFI. The conserved pentapeptide, Gly-X-Ser-X-Gly, is located in the ORFI. A sequence coding for a catalytic triad that resembles that of a serine protease, consisting of serine, histidine, and aspartic acid or glutamic acid residues, was present in the lipase gene. Expression of the S5 lipase gene in E. coli resulted in a 100-fold increase in enzyme activity 9 h after induction with 0.75 mM IPTG. The recombinant protein revealed a size of 60 kDa on SDS-PAGE. The Lip S5 gene was stable in the presence of 25% (v/v) n-dodecane and n-tetradecane after 2 h incubation at 37 degrees C. (C) 2009 Elsevier Ltd. All rights reserved.

关键词： Organic solvent-tolerant lipase Chaperone Pseudomonos aeruginosa Shot-gun cloning Expression phylogenetic tree analysis

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Identification of Zygosaccharomyces mellis strains in stored honey and their stress tolerance

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FOOD SCIENCE AND BIOTECHNOLOGY 2016年第6期25卷 1645-1650页

作者： Liu, Gongliang Tao, Changli Zhu, Baosheng Bai, Weidong Zhang, Liangliang Wang, Zengpeng Liang, Xingting Zhongkai Univ Agr & Engn Coll Light Ind & Food Sci Guangzhou 510225 Guangdong Peoples R China Key Lab Tradit Cantonese Food Proc & Safety Contr Guangzhou 510225 Guangdong Peoples R China Guangdong Pharmaceut Univ Sch Life Sci & Biopharmacol Guangzhou 510006 Guangdong Peoples R China

To screen yeast with high sugar tolerance and evaluate their stress tolerance, six yeast strains were selected from 17 stored honey samples. The species were identified through 26S rRNA sequencing. Their stress tolerance was determined via the Durham fermentation method and ethanol production ability was determined via flask fermentation. The results demonstrated that all the six strains were Zygosaccharomyces mellis. Their sugar, ethanol, and acid tolerance ranges were 500-700 g/L, 10-12% (v/v), and pH 2.5-4.5, respectively. The SO2 tolerance was 250 mg/L. Among the six strains, 6-7431 had the best stress tolerance with sugar tolerance of 700 g/L, ethanol tolerance of 12% (v/v), and acid tolerance of pH 2.5. Furthermore, the strain of 6-7431 had the highest percentage of ethanol production at the same initial sugar content as the other strains. Therefore, the selected six yeast strains would be promising fermentation yeasts for wine-making, ethanol production, or other fermentation purposes.

关键词： Zygosaccharomyces mellis single colony isolation phylogenetic tree analysis stress tolerance

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Purification, characterization, and molecular cloning of chitinases from the stomach of the threeline grunt Parapristipoma trilineatum

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PROCESS BIOCHEMISTRY 2013年第9期48卷 1324-1334页

作者： Ikeda, Mana Kondo, Yujiro Matsumiya, Masahiro Nihon Univ Coll Bioresource Sci Dept Marine Sci & Resources Fujisawa Kanagawa 2520880 Japan

Two chitinase isozymes, PtChiA and PtChiB, were purified from the stomach of the threeline grunt, Parapristipoma trilineatum. The molecular masses of PtChiA and PtChiB were estimated to be 50 and 60 kDa by SDS-PAGE, respectively. Both chitinases were stable at pH 3.0-6.0 (acidic) and showed the optimum pH toward both short and long substrates in the acidic region (pH 2.5-5.0). PtChiA and PtChiB preferentially degraded the second and third glycosidic bonds from the non-reducing end of N-acetylchitooligosaccharides, respectively. PtChiA and PtChiB exhibited wide substrate specificities toward crystalline chitin. Moreover, 2 cDNAs encoding PtChiA and PtChiB, PtChi-1 and PtChi-2, respectively, were cloned. The deduced amino acid sequences of both chitinase cDNAs comprised N-terminal signal peptides, glycoside hydrolase 18 catalytic domains, linker regions, and C-terminal chitin-binding domains. phylogenetic tree analysis of vertebrate chitinases revealed that fish stomach chitinases including PtChi-1 and PtChi-2 form unique chitinase groups, acidic fish chitinase-1 (AFCase-1) and acidic fish chitinase-2 (AFCase-2), which differ from the acidic mammalian chitinase (AMCase) group. The present results suggest that fish have a chitin-degrading enzymatic system in which 2 different chitinases, AFCase-1 and AFCase-2, with different degradation patterns are expressed in the stomach. (C) 2013 Elsevier Ltd. All rights reserved.

关键词： Chitinase fish stomach Parapristipoma trilineatum Substrate specificity Molecular cloning phylogenetic tree analysis

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New geographic distribution and molecular diversity of Citrus chlorotic dwarf-associated virus in China

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Journal of Integrative Agriculture 2022年第1期21卷 293-298页

作者： YANG Zhen ZHANG Lan ZHAO Jin-fa ZHANG Xing-kai WANG Ying LI Tai-sheng ZHANG Wei ZHOU Yan National Citrus Engineering and Technology Research Center Citrus Research InstituteSouthwest UniversityChongqing 400712P.R.China Horticulture Research Institute Guangxi Academy of Agricultural SciencesNanning 530000P.R.China Plant Protection Station of Sichuan Agriculture and Rural Bureau Chengdu 610041P.R.China

In 2009, an emerging citrus viral disease caused by Citrus chlorotic dwarf-associated virus(CCDaV) was discovered in Yunnan Province of China. However, the occurrence and spread of CCDaV in other citrus-growing provinces in China is unknown to date. To better understand the distribution and molecular diversity of CCDaV in China, a total of 1 772 citrus samples were collected from 11 major citrus-growing provinces and were tested for CCDaV by PCR. Among these, 134 citrus samples from Guangxi, Yunnan and Guangdong were tested positive for CCDaV, demonstrating that the occurrence and spread of CCDaV are increasing in China. The complete genome sequences of 17 CCDaV isolates from different provinces and hosts were sequenced. Comparisons of the whole-genome sequences of the 17 CCDaV isolates as well as the 15 isolates available in GenBank revealed that the sequence identity was about 99–100%, showing that the CCDaV isolates were highly conserved. phylogenetic studies showed that the 32 CCDaV isolates belonged to four different groups based on geographical origins and host species, and that CCDaV isolates from China and Turkey were clustered into different groups. The results provide important information for clarifying the distribution and genetic diversity of CCDaV in China.

关键词： Citrus chlorotic dwarf-associated virus detection sequence alignments phylogenetic tree analysis

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Molecular Prevalence Study of QX-like Infectious Bronchitis Virus in Japan from 2016 to 2023

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AVIAN DISEASES 2023年第3期67卷 254-260页

作者： Nakanishi, Makoto Kayaba, Akira Soma, Junichi Zen-noh, Ja Natl Federat Agr Cooperat Assoc Inst Anim Hlth 7 Ohja Machi Sakura Chiba 2850043 Japan

Infectious bronchitis is an acute and highly contagious disease in chickens caused by the infectious bronchitis virus (IBV), which has caused significant economic losses to the poultry industry worldwide. The antigenic variant, the QX-like genotype (GI-19 lineage), has been currently reported in epidemics in East Asia, Southeast Asia, the Middle East, Europe, and Africa. We first reported an epidemic of Japanese QX-like IBVs genetically related to Chinese and South Korean strains in the Kyushu region of Japan in 2020. However, because their nationwide prevalence was unknown, we performed a nationwide survey. The testing of 419 reverse transcription (RT)-PCR-positive samples (376 layers and 43 broilers) of IBV field strains between April 2016 and March 2022 detected two QX-like IBVs in 2019 and 2021 broiler samples from one region. A survey of fecal samples collected from 122-layer farms nationwide between November 2022 and January 2023 detected QX-like IBV genes from seven farms in various regions. phylogenetic tree analysis on the basis of the S1 gene showed that all QX-like IBVs detected in Japan were genetically related to recent Chinese and South Korean strains. A new RT-PCR assay was developed to distinguish between QXlike IBV and other IBV variants prevalent in Japan, whose results were consistent with those of previously reported identification methods. These results suggest that QX-like IBV is rapidly spreading in Japan and that countermeasures are necessary.

关键词： infectious bronchitis virus QX-like genotype Japan molecular prevalence phylogenetic tree analysis RT-PCR

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Natural co-infection with two virulent wild strains of Marek's disease virus in a commercial layer flock

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VETERINARY MICROBIOLOGY 2020年 240卷 108501页

作者： Yu, Zhenghao Zhang, Yanping Lan, Xingge Wang, Yanan Zhang, Feng Gao, Yulong Li, Kai Gao, Li Pan, Qing Qi, Xiaole Cui, Hongyu Zhou, Linyi Sun, Guorong Wang, Xiaomei Liu, Changjun Chinese Acad Agr Sci Harbin Vet Res Inst Div Avian Immunosuppress Dis State Key Lab Vet Biotechnol Harbin 150069 Peoples R China

Marek's disease (MD) is a highly contagious lymphoproliferative poultry disease caused by the oncogenic herpesvirus, Marek's disease virus (MDV). MDV strains have shown a continued evolution of virulence leading to immune failure, and MD cases continue to occur. Co-infection of virulent MDV strains is an important factor leading to viral evolution and host immune failure. This study conducted a laboratory diagnosis and analysis of a MDV infected flock. Testing showed that all samples were MDV positive. PCR detection identified a variable 132-base pair repeat (132-bpr) sequence copy number. This indicated that two virulent strains of MDV were co-infecting the flock. Therefore, we performed homology, sequence alignment, and phylogenetic tree analysis of MDV variant genes including meq, pp38, and RLORF4. Two MDV strains had co-infected the flock;one was the 132bpr two-copy characteristic strain (AH2C) and the other was a 132bpr three-copy characteristic strain (AH3C). Specific mutations in AH3C were found, suggesting that it is a new variant strain. Furthermore, the viral load of the two strains in vivo indicated that both strains had high and similar replication ability. There was no significant difference in the proportion of positive samples of the two strains causing disease. In the whole flock, neither strain displayed an obvious advantage. However, there was a dominant strain in individual chickens, with the exception of one sample. This study reported the co-infection regularity of two virulent MDV strains in the same flock, and even in the same chicken in field conditions. In the context of overall epidemiology, this study is a useful reference.

关键词： Marek's disease virus Viral co-infection phylogenetic tree analysis Viral mutation Viral evolution

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