Biotechnological Applications In Preserving Endangered Species

Dariyus KabrajiMSC 1 Batch 2 Roll No 18

INTRODUCTION

• The International Union for Conservation of Nature (IUCN) maintains a list of species whose existence is currently under threat from any natural or artificial malpractice, known as the Red list, or the Red Data Book

• Over 6800 species on that list face extinction in the near future unless certain steps are taken to preserve them

• With reserved status and efforts of protective organisations and breeding programs, these species are still under threat, thus advanced biological sciences seem to be the last resort.

REQUIREMENT

• Since the list of endangered species is so large, a singular effort or program to conserve all species would be astronomical

• Small scale projects are conducted to save endangered species every day by NGOs and Research facilities

• Such projects require certain information after choosing a single species to protect and focusing all efforts on protecting it

REQUIREMENT

The bare necessities of any such project include:• Knowledge of the species’ characteristics & habits• The optimal parameters for its habitat• The various natural and artificial threats to its

survival• Adequate funding and resources• Awareness generated amongst the locals who

might have personal experience with the species & might even depend on it for their own needs

Striped dolphins

• One example of a threatened species include Striped dolphins (Stenella coeruleoalba) which lives mainly near the Valencian community coast near the Mediterranean Sea

• The continuous cases of dying dolphins led to an investigation by a nearby vet school whose students performed autopsies on the bodies

• The procedures performed included necropsy, sample collection, histopathology, immunohistochemistry, conventional reverse transcription polymerase chain reaction (RT-PCR) and Universal Probe Library (UPL) RT-PCR assays

Striped dolphins

AUTOPSY PROCEDURE

• Fresh tissue samples (brain, lung, kidney, liver, lymph node, tonsil, thymus, spleen and skin) were fixed in 10% neutral buffered formalin for histopathology, refrigerated, and tissue samples were frozen for molecular diagnosis.

• Immunohistochemical staining with a Canine Distemper Virus monoclonal antibody specific for nucleoprotein, IgG2B isotype, was carried out on selected samples of brain, lung, kidney, urinary bladder, stomach and intestine.

• Frozen tissues were homogenized using a Bullet BlenderTM, and total nucleic acid was extracted using the NucleoSpin RNA II Kit and the High Pure PCR Template Preparation Kit.

• For the molecular CeMV diagnosis, real-time RT-PCR assays that target a sequence within the fusion protein gene was carried out.

RESULTS OF INFECTION

• In brain tissue, non- supportive encephalitis (inflammation) and acidophilic intra-nuclear viral inclusions detected

• Other symptoms include pneumonia, enteritis in large intestine with parasitic granulomas, tonsilitis, tongue pustules and many other effects

RESULTS OF INFECTION

DOLPHIN MORBILLIVIRUS

• Marine Viral pathogen that belongs to the same virus family as Canine Distemper Virus (CDV) and Porpoise Morbillivirus (PMV)

• Cetacean Morbillivirus (CeMV) is that family & has been responsible for many fatalities in marine mammals worldwide

• The US Navy Marine Mammal program collaborated with the University of Georgia to map out the DMV gene and prepare a DNA vaccine for the same

DOLPHIN MORBILLIVIRUS FAMILY

THE VACCINE

• In 2007, a DNA vaccine was created from the DMV virus and was the first of its kind to be successfully used on a mammal species

• The virus gene was mapped and the full length fusion and hemagglutinin genes were sub-cloned into the Sal 1 and Not 1 sites of expression vector pvR-1055, forming pvR-DMV-F and pvR-DMV-H

• The plasmid DNA was grown in E. coli and selected by observing their growth in kanamycin, then evaluated by Western Blot and DMV – Specific ELISA

• All dolphins received 1.5mg plasmid DNA in 4.5ml saline solution with booster injections

• By ELISA technique, their IgG was isolated and compared to the pre-inoculation samples

OTHER SPECIES UNDER THREAT

• Hector’s Dolphin (Cephalorhynchus hectori), one of New Zealand’s most famous dolphins

OTHER SPECIES UNDER THREAT

• Australian Snubfin Dolphin (Orcaella heinsohni)

FUTURE PROSPECTS

• The various Oceanographic institutes in Coastal US states and Europe are attempting to find the root cause of infection and how these viruses are incubated in the dolphins

• Research into why the Dolphin is the only family infected by this particular DMV is warranted

REFERENCES

• K. Vaughn, J. D. Crew, G. Hermanson, M. K. Wloch, R. H. Riffenburgh, C. R. Smith, W. G. V. Bonn; Immunogenic DNA vaccines against Dolphin Morbillivirus; Vetenary immunology and Immunopathology 120 (2007)

• Consuelo Rubio-Guerri, Mar Melero, Jose Manuel; Unusual Striped Dolphin Mass Mortality Episode related to Cetacean Morbillivirus (CeMV) in the Spanish Mediterranean Sea; BMC Vetenary Research 2013

• The IUCN Red List Of Threatened And Endangered Species