Use of CO1 gene sequences for computing genetic diversity between Cirrhinus mrigala from two different habitats (Farm and River), 2018
I S S N : 2 4 5 6-6 2 6 8 A B S T R A C T DNA barcoding technique involving documentation of anim... more I S S N : 2 4 5 6-6 2 6 8 A B S T R A C T DNA barcoding technique involving documentation of animal species based on a partial sequence of cytochrome C Oxidase 1 gene from mitochondrial DNA attracted many scientists at the start of 21 st century. The flaws in morphometric methods for identification of larvae of fishes and other animals, their eggs, and damaged specimen with incomplete morphology upraised the question for an alternate method for identification and grouping of many animal species. Partial sequence of CO1 gene used as barcode has been attested as a valuable tool for identification of fish species as well as supportive in computing evolutionary history and genetic diversity. Current study was conducted to identify the fishes by using the DNA barcoding technique resulting partial sequences of CO1 gene, and then use of these sequences in evaluation of the evolutionary history and genetic diversity of Cirrhinus mrigala inhabiting different areas. Short sequences from 5 end of CO1 gene (650 base pair) were amplified, sequenced and analyzed using different soft wares. According to results haplotypes were found which showed genetic variations among fishes from two different habitats. The phylogenetic analysis established a neighboring relationship and a shared ancestor of fish from different habitats. Haplotype diversity of 0.7143 was found in partial sequences of CO1 gene of all experimental fish. From these results it is obvious that fish species sharing identical genus and family but with diverse habitats show genetic variations even though involving a common ancestor.
Study of Induce Histological and Biochemical Alterations in Hypophthalmicthyes molitrix after Acute Exposure to Deltamethrine, 2018
Abstract | Pesticides have brought us great economic benefits, at the same time they are also
roo... more Abstract | Pesticides have brought us great economic benefits, at the same time they are also root of serious ecological harms like non-target toxicity of organism. Synthetic pyrethroid (deltamethrine) is widely used due to its less toxicity and easy degradation in soil. In this study fresh water fish Hypophthalmicthyes molitrix were exposed to sub lethal concentrations of deltamethrine. Fish were divided into 4 groups and each group contains six fish. One of them is control and A, B and C were experimental groups. Experimental groups viz. A, B and C were exposed to 25, 50 and 75% of LC50, respectively for four days. After 96 h; sample of blood for biochemical analysis (blood glucose, cholesterol and total protein) and gills tissue for histology were taken. After acute exposure to pesticide, significant changes were observed in serum biochemistry and histology. Serum glucose and cholesterol were increased while total protein was decreased. Histopathological result reveled that gills tissue of experimental fish was damage severely resulting Necrosis, Damaged nuclei , rupturing of epithelial cells, Mucous cells and severe lamellar fusion .it was concluded that, deltamethrine is highly toxic for non-target aquatic organisms like silver carp (Hypophthalmicthyes molitrix). Artic
Taxonomy has always been a challenging job for the scientists. Traditional methods for identifica... more Taxonomy has always been a challenging job for the scientists. Traditional methods for identification of which sometimes create great confusion due to great morphological similarities in closed species. DNA barcoding is a modern species identification method in which a short standardized mitochondrial DNA region is u fragment near the 5 (CO1 capable enough for identification of processed, fishes as compared to traditional morphometric methods and is also appropriate for rapid identification of juveniles as well as adult sta fishes. Present study is Labeogonius of CO1 and same fishes which were identified morphologically at the b study with the help of identification key. The phylogenetic tree analysis confirmed the evolutionary relationships between from Pakistan and fishes from other parts of the world. From these findings, it is marker for identification of most of fish species at molecular phylogenetic analysis.
Deltamethrin is a broad spectrum synthetic pyrethroid pesticide. It is considered less toxic as c... more Deltamethrin is a broad spectrum synthetic pyrethroid pesticide. It is considered less toxic as compared to organo-chlorine and organo-phosphate pesticides. Deltamethrin was selected to study the biochemical and histological alterations in liver of silver carp. The LC50 1.6 µg/L was determined. Fish (n=6) were dividing into 4 groups A (control) B, C and D (experimental). Experimental groups B, C and D were exposed to 25%, 50% and 75% of LC50 respectively for acute exposure. The blood and liver tissue sampling was done after 96hrs for histology and biochemical analysis [alanine amino transferase (ALT), and aspartate amino-transferase (AST)]. Liver histology revealed that necrosis, nuclear pycnosis, hypertrophy of hepatocytes, vacuolization, nuclear atrophy and congestion of blood vessels were observed as compare to control group. This result was also supported by the significant increase in levels of hepatic enzymes AST and ALT in blood plasma of exposed fish as compared to control fish. These results suggested that deltamethrin is highly toxic for aquatic organism like silver carp causing malfunctioning of liver.
ABSTRACT
The Interferon Regulatory Factor (IRF) family of transcription factors which play differ... more ABSTRACT The Interferon Regulatory Factor (IRF) family of transcription factors which play different roles in response to microbial pathogen invasion and coordinate both innate and adaptive immune systems. This happens when pathogen-associated molecular patterns (PAMPS) bind to cell surface and recognize cytosolic receptors. Most IRFs studies focus on the functional role of IRFs in antiviral responses and their functional and developmental roles in macrophages, dendritic cells, B lymphocytes and T lymphocytes. So far IRF 1, 2, 3, 4. 7& 8 have been cloned in rainbow trout. During present study, a portion of IRF5 gene was cloned. RNA was extracted from cells and tissues of spleen, and head kidney. RNA was converted to cDNA. Six primers (four forward and two reverse) were used during amplification of IRF5 gene under different PCR conditions. These primers were designed from the data base of Salmon ESTs (Expressed Sequence Tags). PCR products were ligated, transformed and cloned. After Polymerase Chain Reaction all samples were run on agarose gel. Gel images showed many bands less than 1000 base pairs in length. Samples which resulted into brighter bands were sent for sequencing. Results were analyzed with BLAST2 which showed that all sequences were part of IRF5 gene and size of product was 678 base pair. After translation these nucleotides were converted into 226 amino acids. These amino acids showed complete alignment with the Salmon IRF5 gene. Key words: Interferon regulatory factors, IRF5, cloning, rainbow trout INTRODUCTION It is a natural phenomenon that all multicellular organisms have a built in defense system against any disease causing agents such as microbes. This distinctive feature of organisms is due to the presence of natural or innate immune system. Natural immune system is characterized by responding against microbes and it does not react against non-microbes. This system is also activated when host cells are damaged by microbes. The most important components of innate immunity include epithelia, phagocytes and natural killer (NK) cells. Cytokines like interferons and plasma proteins, including the proteins of the complement system are also part of innate immune system. This system provides the first defense against infections, and gives “second signals” along with antigens for the activation of B and T lymphocytes. B and T lymphocytes are part of adaptive immune system. In this way adaptive immunity is stimulated by innate immunity. Interferons are cytokines which hinder the activity of viruses in a cell, in other words they stop breeding of viruses in cells. Interferons are also produced from neighboring cells as a result of virus attack on cells (Vilcek, 2006). Interferons are capable of stopping virus growth in cells surrounding the infected cells. Interferons have many functions. They were
Abstract
DNA bar-coding is a taxonomic method that uses small genetic markers in organisms’
mitoc... more Abstract DNA bar-coding is a taxonomic method that uses small genetic markers in organisms’ mitochondrial DNA (mt DNA) for identification of particular species. It uses sequence diversity in a 658-base pair fragment near the 50 end of the mitochondrial cytochrome c oxidase subunit 1 (CO1) gene as a tool for species identification. DNA barcoding is more accurate and reliable method as compared with the morphological identification. It is equally useful in juveniles as well as adult stages of fishes. The present study was conducted to identify three farm fish species of Pakistan (Cyprinus carpio, Cirrhinus mrigala, and Ctenopharyngodon idella) genetically. All of them belonged to family cyprinidae. CO1 gene was amplified. PCR products were sequenced and analyzed by bioinformatic software. Conspecific, congenric, and confamilial k2P nucleotide divergence was estimated. From these findings, it was concluded that the gene sequence, CO1, may serve as milestone for the identification of related species at molecular level
Use of CO1 gene sequences for computing genetic diversity between Cirrhinus mrigala from two different habitats (Farm and River), 2018
I S S N : 2 4 5 6-6 2 6 8 A B S T R A C T DNA barcoding technique involving documentation of anim... more I S S N : 2 4 5 6-6 2 6 8 A B S T R A C T DNA barcoding technique involving documentation of animal species based on a partial sequence of cytochrome C Oxidase 1 gene from mitochondrial DNA attracted many scientists at the start of 21 st century. The flaws in morphometric methods for identification of larvae of fishes and other animals, their eggs, and damaged specimen with incomplete morphology upraised the question for an alternate method for identification and grouping of many animal species. Partial sequence of CO1 gene used as barcode has been attested as a valuable tool for identification of fish species as well as supportive in computing evolutionary history and genetic diversity. Current study was conducted to identify the fishes by using the DNA barcoding technique resulting partial sequences of CO1 gene, and then use of these sequences in evaluation of the evolutionary history and genetic diversity of Cirrhinus mrigala inhabiting different areas. Short sequences from 5 end of CO1 gene (650 base pair) were amplified, sequenced and analyzed using different soft wares. According to results haplotypes were found which showed genetic variations among fishes from two different habitats. The phylogenetic analysis established a neighboring relationship and a shared ancestor of fish from different habitats. Haplotype diversity of 0.7143 was found in partial sequences of CO1 gene of all experimental fish. From these results it is obvious that fish species sharing identical genus and family but with diverse habitats show genetic variations even though involving a common ancestor.
Study of Induce Histological and Biochemical Alterations in Hypophthalmicthyes molitrix after Acute Exposure to Deltamethrine, 2018
Abstract | Pesticides have brought us great economic benefits, at the same time they are also
roo... more Abstract | Pesticides have brought us great economic benefits, at the same time they are also root of serious ecological harms like non-target toxicity of organism. Synthetic pyrethroid (deltamethrine) is widely used due to its less toxicity and easy degradation in soil. In this study fresh water fish Hypophthalmicthyes molitrix were exposed to sub lethal concentrations of deltamethrine. Fish were divided into 4 groups and each group contains six fish. One of them is control and A, B and C were experimental groups. Experimental groups viz. A, B and C were exposed to 25, 50 and 75% of LC50, respectively for four days. After 96 h; sample of blood for biochemical analysis (blood glucose, cholesterol and total protein) and gills tissue for histology were taken. After acute exposure to pesticide, significant changes were observed in serum biochemistry and histology. Serum glucose and cholesterol were increased while total protein was decreased. Histopathological result reveled that gills tissue of experimental fish was damage severely resulting Necrosis, Damaged nuclei , rupturing of epithelial cells, Mucous cells and severe lamellar fusion .it was concluded that, deltamethrine is highly toxic for non-target aquatic organisms like silver carp (Hypophthalmicthyes molitrix). Artic
Taxonomy has always been a challenging job for the scientists. Traditional methods for identifica... more Taxonomy has always been a challenging job for the scientists. Traditional methods for identification of which sometimes create great confusion due to great morphological similarities in closed species. DNA barcoding is a modern species identification method in which a short standardized mitochondrial DNA region is u fragment near the 5 (CO1 capable enough for identification of processed, fishes as compared to traditional morphometric methods and is also appropriate for rapid identification of juveniles as well as adult sta fishes. Present study is Labeogonius of CO1 and same fishes which were identified morphologically at the b study with the help of identification key. The phylogenetic tree analysis confirmed the evolutionary relationships between from Pakistan and fishes from other parts of the world. From these findings, it is marker for identification of most of fish species at molecular phylogenetic analysis.
Deltamethrin is a broad spectrum synthetic pyrethroid pesticide. It is considered less toxic as c... more Deltamethrin is a broad spectrum synthetic pyrethroid pesticide. It is considered less toxic as compared to organo-chlorine and organo-phosphate pesticides. Deltamethrin was selected to study the biochemical and histological alterations in liver of silver carp. The LC50 1.6 µg/L was determined. Fish (n=6) were dividing into 4 groups A (control) B, C and D (experimental). Experimental groups B, C and D were exposed to 25%, 50% and 75% of LC50 respectively for acute exposure. The blood and liver tissue sampling was done after 96hrs for histology and biochemical analysis [alanine amino transferase (ALT), and aspartate amino-transferase (AST)]. Liver histology revealed that necrosis, nuclear pycnosis, hypertrophy of hepatocytes, vacuolization, nuclear atrophy and congestion of blood vessels were observed as compare to control group. This result was also supported by the significant increase in levels of hepatic enzymes AST and ALT in blood plasma of exposed fish as compared to control fish. These results suggested that deltamethrin is highly toxic for aquatic organism like silver carp causing malfunctioning of liver.
ABSTRACT
The Interferon Regulatory Factor (IRF) family of transcription factors which play differ... more ABSTRACT The Interferon Regulatory Factor (IRF) family of transcription factors which play different roles in response to microbial pathogen invasion and coordinate both innate and adaptive immune systems. This happens when pathogen-associated molecular patterns (PAMPS) bind to cell surface and recognize cytosolic receptors. Most IRFs studies focus on the functional role of IRFs in antiviral responses and their functional and developmental roles in macrophages, dendritic cells, B lymphocytes and T lymphocytes. So far IRF 1, 2, 3, 4. 7& 8 have been cloned in rainbow trout. During present study, a portion of IRF5 gene was cloned. RNA was extracted from cells and tissues of spleen, and head kidney. RNA was converted to cDNA. Six primers (four forward and two reverse) were used during amplification of IRF5 gene under different PCR conditions. These primers were designed from the data base of Salmon ESTs (Expressed Sequence Tags). PCR products were ligated, transformed and cloned. After Polymerase Chain Reaction all samples were run on agarose gel. Gel images showed many bands less than 1000 base pairs in length. Samples which resulted into brighter bands were sent for sequencing. Results were analyzed with BLAST2 which showed that all sequences were part of IRF5 gene and size of product was 678 base pair. After translation these nucleotides were converted into 226 amino acids. These amino acids showed complete alignment with the Salmon IRF5 gene. Key words: Interferon regulatory factors, IRF5, cloning, rainbow trout INTRODUCTION It is a natural phenomenon that all multicellular organisms have a built in defense system against any disease causing agents such as microbes. This distinctive feature of organisms is due to the presence of natural or innate immune system. Natural immune system is characterized by responding against microbes and it does not react against non-microbes. This system is also activated when host cells are damaged by microbes. The most important components of innate immunity include epithelia, phagocytes and natural killer (NK) cells. Cytokines like interferons and plasma proteins, including the proteins of the complement system are also part of innate immune system. This system provides the first defense against infections, and gives “second signals” along with antigens for the activation of B and T lymphocytes. B and T lymphocytes are part of adaptive immune system. In this way adaptive immunity is stimulated by innate immunity. Interferons are cytokines which hinder the activity of viruses in a cell, in other words they stop breeding of viruses in cells. Interferons are also produced from neighboring cells as a result of virus attack on cells (Vilcek, 2006). Interferons are capable of stopping virus growth in cells surrounding the infected cells. Interferons have many functions. They were
Abstract
DNA bar-coding is a taxonomic method that uses small genetic markers in organisms’
mitoc... more Abstract DNA bar-coding is a taxonomic method that uses small genetic markers in organisms’ mitochondrial DNA (mt DNA) for identification of particular species. It uses sequence diversity in a 658-base pair fragment near the 50 end of the mitochondrial cytochrome c oxidase subunit 1 (CO1) gene as a tool for species identification. DNA barcoding is more accurate and reliable method as compared with the morphological identification. It is equally useful in juveniles as well as adult stages of fishes. The present study was conducted to identify three farm fish species of Pakistan (Cyprinus carpio, Cirrhinus mrigala, and Ctenopharyngodon idella) genetically. All of them belonged to family cyprinidae. CO1 gene was amplified. PCR products were sequenced and analyzed by bioinformatic software. Conspecific, congenric, and confamilial k2P nucleotide divergence was estimated. From these findings, it was concluded that the gene sequence, CO1, may serve as milestone for the identification of related species at molecular level
Uploads
root of serious ecological harms like non-target toxicity of organism. Synthetic pyrethroid
(deltamethrine) is widely used due to its less toxicity and easy degradation in soil. In this
study fresh water fish Hypophthalmicthyes molitrix were exposed to sub lethal concentrations
of deltamethrine. Fish were divided into 4 groups and each group contains six fish. One of
them is control and A, B and C were experimental groups. Experimental groups viz. A, B and
C were exposed to 25, 50 and 75% of LC50, respectively for four days. After 96 h; sample of
blood for biochemical analysis (blood glucose, cholesterol and total protein) and gills tissue for
histology were taken. After acute exposure to pesticide, significant changes were observed in
serum biochemistry and histology. Serum glucose and cholesterol were increased while total
protein was decreased. Histopathological result reveled that gills tissue of experimental fish was
damage severely resulting Necrosis, Damaged nuclei , rupturing of epithelial cells, Mucous cells
and severe lamellar fusion .it was concluded that, deltamethrine is highly toxic for non-target
aquatic organisms like silver carp (Hypophthalmicthyes molitrix).
Artic
The Interferon Regulatory Factor (IRF) family of transcription factors which play different roles in response to microbial pathogen invasion and coordinate both innate and adaptive immune systems. This happens when pathogen-associated molecular patterns (PAMPS) bind to cell surface and recognize cytosolic receptors. Most IRFs studies focus on the functional role of IRFs in antiviral responses and their functional and developmental roles in macrophages, dendritic cells, B lymphocytes and T lymphocytes. So far IRF 1, 2, 3, 4. 7& 8 have been cloned in rainbow trout. During present study, a portion of IRF5 gene was cloned. RNA was extracted from cells and tissues of spleen, and head kidney. RNA was converted to cDNA. Six primers (four forward and two reverse) were used during amplification of IRF5 gene under different PCR conditions. These primers were designed from the data base of Salmon ESTs (Expressed Sequence Tags). PCR products were ligated, transformed and cloned. After Polymerase Chain Reaction all samples were run on agarose gel. Gel images showed many bands less than 1000 base pairs in length. Samples which resulted into brighter bands were sent for sequencing. Results were analyzed with BLAST2 which showed that all sequences were part of IRF5 gene and size of product was 678 base pair. After translation these nucleotides were converted into 226 amino acids. These amino acids showed complete alignment with the Salmon IRF5 gene.
Key words: Interferon regulatory factors, IRF5, cloning, rainbow trout
INTRODUCTION
It is a natural phenomenon that all multicellular organisms have a built in defense system against any disease causing agents such as microbes. This distinctive feature of organisms is due to the presence of natural or innate immune system. Natural immune system is characterized by responding against microbes and it does not react against non-microbes. This system is also activated when host cells are damaged by microbes. The most important components of innate immunity include epithelia, phagocytes and natural killer (NK) cells. Cytokines like interferons and plasma proteins, including the proteins of the complement system are also part of innate immune system. This system provides the first defense against infections, and gives “second signals” along with antigens for the activation of B and T lymphocytes. B and T lymphocytes are part of adaptive immune system. In this way adaptive immunity is stimulated by innate immunity.
Interferons are cytokines which hinder the activity of viruses in a cell, in other words they stop breeding of viruses in cells. Interferons are also produced from neighboring cells as a result of virus attack on cells (Vilcek, 2006). Interferons are capable of stopping virus growth in cells surrounding the infected cells. Interferons have many functions. They were
DNA bar-coding is a taxonomic method that uses small genetic markers in organisms’
mitochondrial DNA (mt DNA) for identification of particular species. It uses sequence diversity in
a 658-base pair fragment near the 50 end of the mitochondrial cytochrome c oxidase subunit 1
(CO1) gene as a tool for species identification. DNA barcoding is more accurate and reliable
method as compared with the morphological identification. It is equally useful in juveniles as
well as adult stages of fishes. The present study was conducted to identify three farm fish
species of Pakistan (Cyprinus carpio, Cirrhinus mrigala, and Ctenopharyngodon idella) genetically.
All of them belonged to family cyprinidae. CO1 gene was amplified. PCR products were
sequenced and analyzed by bioinformatic software. Conspecific, congenric, and confamilial
k2P nucleotide divergence was estimated. From these findings, it was concluded that the
gene sequence, CO1, may serve as milestone for the identification of related species at
molecular level
root of serious ecological harms like non-target toxicity of organism. Synthetic pyrethroid
(deltamethrine) is widely used due to its less toxicity and easy degradation in soil. In this
study fresh water fish Hypophthalmicthyes molitrix were exposed to sub lethal concentrations
of deltamethrine. Fish were divided into 4 groups and each group contains six fish. One of
them is control and A, B and C were experimental groups. Experimental groups viz. A, B and
C were exposed to 25, 50 and 75% of LC50, respectively for four days. After 96 h; sample of
blood for biochemical analysis (blood glucose, cholesterol and total protein) and gills tissue for
histology were taken. After acute exposure to pesticide, significant changes were observed in
serum biochemistry and histology. Serum glucose and cholesterol were increased while total
protein was decreased. Histopathological result reveled that gills tissue of experimental fish was
damage severely resulting Necrosis, Damaged nuclei , rupturing of epithelial cells, Mucous cells
and severe lamellar fusion .it was concluded that, deltamethrine is highly toxic for non-target
aquatic organisms like silver carp (Hypophthalmicthyes molitrix).
Artic
The Interferon Regulatory Factor (IRF) family of transcription factors which play different roles in response to microbial pathogen invasion and coordinate both innate and adaptive immune systems. This happens when pathogen-associated molecular patterns (PAMPS) bind to cell surface and recognize cytosolic receptors. Most IRFs studies focus on the functional role of IRFs in antiviral responses and their functional and developmental roles in macrophages, dendritic cells, B lymphocytes and T lymphocytes. So far IRF 1, 2, 3, 4. 7& 8 have been cloned in rainbow trout. During present study, a portion of IRF5 gene was cloned. RNA was extracted from cells and tissues of spleen, and head kidney. RNA was converted to cDNA. Six primers (four forward and two reverse) were used during amplification of IRF5 gene under different PCR conditions. These primers were designed from the data base of Salmon ESTs (Expressed Sequence Tags). PCR products were ligated, transformed and cloned. After Polymerase Chain Reaction all samples were run on agarose gel. Gel images showed many bands less than 1000 base pairs in length. Samples which resulted into brighter bands were sent for sequencing. Results were analyzed with BLAST2 which showed that all sequences were part of IRF5 gene and size of product was 678 base pair. After translation these nucleotides were converted into 226 amino acids. These amino acids showed complete alignment with the Salmon IRF5 gene.
Key words: Interferon regulatory factors, IRF5, cloning, rainbow trout
INTRODUCTION
It is a natural phenomenon that all multicellular organisms have a built in defense system against any disease causing agents such as microbes. This distinctive feature of organisms is due to the presence of natural or innate immune system. Natural immune system is characterized by responding against microbes and it does not react against non-microbes. This system is also activated when host cells are damaged by microbes. The most important components of innate immunity include epithelia, phagocytes and natural killer (NK) cells. Cytokines like interferons and plasma proteins, including the proteins of the complement system are also part of innate immune system. This system provides the first defense against infections, and gives “second signals” along with antigens for the activation of B and T lymphocytes. B and T lymphocytes are part of adaptive immune system. In this way adaptive immunity is stimulated by innate immunity.
Interferons are cytokines which hinder the activity of viruses in a cell, in other words they stop breeding of viruses in cells. Interferons are also produced from neighboring cells as a result of virus attack on cells (Vilcek, 2006). Interferons are capable of stopping virus growth in cells surrounding the infected cells. Interferons have many functions. They were
DNA bar-coding is a taxonomic method that uses small genetic markers in organisms’
mitochondrial DNA (mt DNA) for identification of particular species. It uses sequence diversity in
a 658-base pair fragment near the 50 end of the mitochondrial cytochrome c oxidase subunit 1
(CO1) gene as a tool for species identification. DNA barcoding is more accurate and reliable
method as compared with the morphological identification. It is equally useful in juveniles as
well as adult stages of fishes. The present study was conducted to identify three farm fish
species of Pakistan (Cyprinus carpio, Cirrhinus mrigala, and Ctenopharyngodon idella) genetically.
All of them belonged to family cyprinidae. CO1 gene was amplified. PCR products were
sequenced and analyzed by bioinformatic software. Conspecific, congenric, and confamilial
k2P nucleotide divergence was estimated. From these findings, it was concluded that the
gene sequence, CO1, may serve as milestone for the identification of related species at
molecular level