Posted: June 20th, 2015

Nerita albicialla and Nerita Melanotragus Genes and Genomes

Introduction

The main goal in studying biology is to understand how animal species respond to temporal and spatial changes in their habitats/living environments. The species of snails varies depending on the environmental place of living. There are many species of snails, some of them are; Nerita albicilla and Nerita melanotragus. These two species are the widely spread species intertidal snailsand are distributed across a number of temporally and spatially fluctuating environmental gradients, including latitudinal clines in water temperature and abrupt changes in water temperature over a tidal cycle (Robinson, Didelot, Hood, & Crook, 2013).

The two species has variation in ecology and geographical ranges. N. albicilla is a low littorial species that are found in tropical and subtropical regions. On the other hand, N. melanotragus is a mid-littoral species that are found in areas from temperate to subtropical climes. The distribution of mid littorals species is more likely to retain long periods of temperature changes and the distribution of the low littoral species is mostly subjected by common flooding and is more shielded from temperature changes. When these two species are paced in different thermal environments, they show a pattern of changes in genetic transformations. In this report we shall discuss the genetic variations of the two snail species when exposed in two different temperature treatments i.e. 14 vs 31 degrees, and three replicates for each treatment condition (Robinson, Didelot, Hood, & Crook, 2013).

Aims of the Experiments

1. To study the genetic variations of the Nerita albicilla and Nerita melanotragus snails species when exposed to two different temperatures 14⁰C versus 31⁰
2. To determine whether the genes that are differentially expressed within each snail species under the two temperature treatments.
3. Check if certain genetic classes are more represented than other classes.
4. To determine if the differentially expressed genes have a similarity between species.

Materials and Methods

Snails sample collection and Pre-treatment

The snail’s species; Nerita albicilla and Nerita melanotragus were collected from the marine environment. To ensure compatibility, the snails of the same sizes were selected. These include the snails of 6 to 8 mm in length. It was preserved for one month and after which were washed and placed individually in water for 3 hours in a villa exposed to a light source at a temperature of 25 degrees. This helps in stimulating the emergence of cercarie for the identification of the possible infection caused by these organisms (Mathur, 2005).

The naturally infected snails were then identified and the snails were maintained on a sterile wet filter paper in petri dish. The petri dish was covered in a plastic mesh and placed in an incubator at 25 degrees previously supplied with some fresh humidified air with the filter paper being changed after every three days. When two weeks elapsed, the snails were then placed on a sterile, wet filter paper having a volume of 200U/ml penicillin and 200μg/ml streptomycin at 25 degrees for a week. Then the snails were further washed I a chemical solution of diethyl carbonate (DEPC) treated water containing 100μg/ml ampicillin. Dissection of the soft bodies of the snails was done individually out on a glass slides and examined microscopically to verify whether or not they were actually infected. All the snails were confirmed to have no infection. The whole soft tissue of each individual snail was carefully washed for three times in ice cold DEPC treated water having 0.3% sodium chloride solution. Dissection of the head-foot region was done individually under a microscope with infected and uninfected snails separated. This was prior to the storage in liquid nitrogen for future use (Mathur, 2005)

The overall snail’s structure of genetic environment was observed for any changes when kept in different temperature environment conditions of 14-31 degrees.

Results

Evaluation of Genetic composition

Ribonucleic acid (RNA) extraction, cDNA synthesis and construction of the suppression subtractive hybrization library.

The total RNA were extracted from head-foot of each of the species and treated separately using TROzol reagent according to manufacturer’s instructions. The traces of genomic DNA was removed through treatment of RNA samples and with DNase I (Promoga) prior to the cDNA synthesis. Evaluation of the RNA quality was done using gel electrophoreses to confirm the integrity of the RNA preparations. The cDNA synthesis and subsequent amplification were performed using 1μg of the total RNA and the super SMART PCR cDNA synthesis kit by following the manufacturer’s protocols (Wang, Zhao, Nie, Jiang & Song, 2012).

The putative biological function sequence was evaluated using gene ontology (GO) based on BLAST search technique analysis. The GO terms about a certain biological process; the molecular functions, and cellular components were taken from a sequence similarities using the corresponding application parameter filters. The resulting annotations were deduced to yield the putative gene functions (Wang, Zhao, Nie, Jiang & Song, 2012).

Discussion

As the climate change affects the marine environments, there is a shift in biotic and abiotic factors within the marine ecosystems. The way species respond to these climatic changes are important in determining the resulting behavior change in marine organisms. Understanding of the physiology of any marines’ species of animals especially the division Mollusca is listed and makes it hard to predict the expectations of the climatic changes. In this research the aim was developing transcriptomic data sets for the two snail species, selection of gene candidates that were involved in temperature, saline and oxygen environments, and calculation of the synonymous and non-synonymous substitution rates for the candidate genes and the comparison of the gene sequences with both closely and distantly related species (Robinson, Didelot, Hood, & Crook, 2013).

In the analysis using illumine sequencing, of Nerita melanotragus and Nerita
albicilla, the total number of reads produced a total of 68,678,334 and 61,367,256 respectively. From these two values, the de novo method of gene assembly assembled 77098 and 78577 contigs reads respectively. the candidates for the genes was identified from the datasets, whichinclude a suite of extensively heat shock proteins that were duplicated and having carbonic anyhrases and globin genes. These large number of contigs were annotated and their functionality characterized (Robinson, Didelot, Hood, & Crook, 2013)

Discussion Summary questions

(i)What genes are differentially expressed between the treatments the within species?

All the genes in the two species Nerita melanotragus and Nerita albicilla were differentially expressed between the treatments of the temperature changes.
(ii) Are some categories of genes over represented in the differentially expressed genes within species?

In the same sample proportion as shown above some genes were overrepresented than others i.e. the total number of reads produced a total of 68,678,334 and 61,367,256 for Nerita melanotragus and Nerita albicilla respectively.
(iii) Are similar genes differentially expressed between the two species?

The similar genes for the species were differentially expressed between the species because of the same impacting medium of temperature change.
(iv)If the genes are different between the species try and explain why?

By virtue of the slight variation of genetic composition between the species, the genes for the species were different in the response to the environmental changes.

References

1. Mathur, S. (2005). A Robust Statistical Method for Detecting Differentially Expressed Genes. Applied Bioinformatics, 4(4), 247-251.Doi – 10.2165/00822942-200504040-00004
2. MA, X., MA, X., WU, X., CHEN, B., & WANG, D. (2009). Differentially expressed genes in diabetes-induced embryopathy. Hereditas (Beijing), 31(3), 280-284. doi:10.3724/sp.j.1005.2009.00280
3. Wille, A., Gruissem, W., Bühlmann, P., & Hennig, L. (2007). TECHNICAL ADVANCE – EVE (external variance estimation) increases statistical power for detecting differentially expressed genes. The Plant Journal, 52(3), 561-569. Doi – 10.1111/j.1365-313x.2007.03227.x
4. Robinson, E., Didelot, X., Hood, D., & Crook, D. (2013). The omparative phylogenetics of ICEHin1056 family reveals deep evolutionary associations of mobile genetic elements responsible for transfer of antibiotic resistance genes. The Lancet, 381, S93. Doi-10.1016/s0140-6736(13)60533-4
5. Wang, H., Zhao, Q., Nie, P., Jiang, M., & Song, J. (2012). Identification of differentially expressed genes in Oncomelania hupensis chronically infected with Schistosoma japonicum. Experimental Parasitology, 130(4), 374-383. doi:10.1016/j.exppara.2012.02.004