Horizontal Gene Transfer in Evolution of Nematode

Parasitism

Priyank H. MhatreRoll No - 5021

Division of Nematology

A gene is a unit of heredity in a living organism

It is a segment of DNA  that code for a specific  protein

Genes hold the information to build and maintain an organism's cells

Gene

What is Gene Transfer?

Vertical gene transfer Horizontal gene transfer

Types of Gene Transfer

Vertical Gene Transfer : - transmission of genes from an organism to it’s offspring

- Sexual reproduction in higher animals and plants is the way of VGT

Vertical Gene Transfer

Horizontal gene transfer (HGT) - implies the non-sexual exchange of genetic material between species, in some cases even across kingdoms

Horizontal gene transfer

Frederick Griffith (1879-1941) Joshua Lederberg (1925-2008)

History of HGT

J Lederberg in 1946 – 47 first time seen conjugation and in 1956 transduction in bacteria

M Syvanen (1985 ) proposed this mechanism as cross-species gene transfer in evolution

Hilario & Gogarten (1993 ) use the term horizontal gene transfer between organisms as an alternative explanation for those conflictive phylogenetic events

(Luis Boto., 2009)

Frederick Griffith in 1928 first time seen the transformation in bacteria

Cont…

HGT is the non-genealogical transmission of genetic material from one organism to another

It is a source of new genetic material to the recipient

It is a mechanism that permits the acquisition of evolutionary novelties

It is common mechanism of gene transfer in bacteria and archaea

HGT is the non-genealogical transmission of genetic material from one organism to another

It is a source of new genetic material to the recipient

It is a mechanism that permits the acquisition of evolutionary novelties

It is common mechanism of gene transfer in bacteria and archaea

(Luis boto., 2009)

Some facts…

HGT

Recent

Ancient

Types of HGT

Ancient :- genes transferred between organisms separated a long time ago

-The ancient HGT is difficult to detect through codon usage bias and

differential base composition

Foreign genes in

cellonce

they are part of the

same genome

support

same mutational bias with resident

and ameliorated after many generations

Recent :- genes transferred between organisms separated recently - Easy to detect based on Criteria of codon usage bias and differential base composition.

(Luis Boto., 2009)

Types of HGT

Transduction

HGTConjugation

Transduction

Transformation

How it happens?How it happens?

Magic of Horizontal Gene Transfer

The photosynthetic sea slug Elysia chlorotica appears like a dark green leaf as a result of retaining chloroplasts from its algal prey, Vaucheria litorea, in cells lining of its digestive tract.

The sea slug has acquired photosynthesis-supporting genes by horizontal gene transfer and can use the chloroplasts to carry out photosynthesis for several months

(Rumpho et al., 2008)

(Rumpho et al., 2008)

How It Grows?

Cont…  It is also called as “solar-powered” sea slug

Nuclear gene of oxygenic photosynthesis, psbO, have been acquired by the animal via HGT

Photosynthesis is by the plastids which provides E. chlorotica with energy for its entire lifespan of 10 months∼

The plastids are not transmitted vertically (i.e. are absent in eggs) and do not undergo division in the sea slug.

(Rumpho et al., 2008)

What about nematodes?

CENOZOIC

MESOZOICCretaceousJurassicTriassic

65 --- Flowering plants (PPN)145200

PALEOZOIC

PermianCarboniferousDevonianSilurianOrdovicianCambrian

253--- Insect (EPN)300336440484542 --- Nematodes origin (FLN)

Era Period Millions Year ago

Land Plants

Evolutionary History of Nematodes

Parasitism

Pre-adaptations-1. Dauer larvae2. Toxicity tolerance3. Low oxygen tolerance

Free-living

Phoresy

Necromeny

(Dieterich et al., 2009)

Evolution of Parasitism In Nematodes

This intermediate association considered as a step toward parasitism

Found in a necromenic association with scarab beetle representing an intermediate type of association between phoretic and parasites

In this nematode the number of detoxification enzymes are Increase, in comparison to phoretic nematodes

It has also acquired cellulase genes Ppa-cel-1, 2, & 3 via HGT from bacteria and archaea

This is the first report of cellulase found in non plant parasitic nematodes

(Mayer et al., 2011)

Pristionchus pacificus

Enzymes Activity

Cellulase, Xylanases

Degradation of cellulose and hemicelluloses

Polygalacturonases,Pectate lyases,Candidate arabinanases

Degradation of pectins

Expansin Soften the plant cell wall

(Danchin et al., 2010)

Nematode proteins with their activity

Plant cell wall modifiers with their closest relatives

(Danchin et al., 2010)

(Danchin et al., 2010)

Phylogenetic Analysis of Polygalacturonases, Pectate lyases, Candidate arabinases

Phylogenetic Analysis of Cellulase, Xylanases., and Expansin.

(Danchin et al., 2010)

Cellulases are enzymes that hydrolyse the β-1,4 linkages of cellulose

Cellulases are from three structurally and phylogenetically unrelated families found in nematodes :- 1. GHF5- genes common in PPN 2. GHF16- genes are known from B. xylophilus , B. mucronatus 3. GHF45- genes are known from B. xylophilus

These genes are acquired via HGT from bacteria, fungus, or other microbes

Cellulase in nematodes

GHF5

These genes show homology with genes of the bacteria like Erwinia chrysanthemi, Clostridium acetobutylicum and B. subtilis

These genes facilitate the intracellular migration of nematodes through plant roots by partial cell wall degradation

1st report of genes with endogenous cellulase production by animals in absence of cellulolytic microorganisms was in G. rostochinensis and in H. glycines

( Smant et al., 1998 )

Genes - GR-eng-1 and 2, HG-eng-1 and 2 - oesophageal gland Protein- GR-ENG-1 & 2, HG-ENG-1 & 2 – stylet secretion

Cyst Nematodes

Researchers characterized a new β-1,4-endoglucanase gene from the root-knot nematode

They identified the gene - MI-eng-2, which is localized in the secretory oesophageal glands cell of all developmental stages of the nematode and the protein secreted is– MI-ENG-2

This protein is involved in plant cell wall degradation during parasitism

GHF5 genes result from horizontal gene transfer of a bacterial gene with a cellulose-binding domain (CBD)

(Ledger et al.,2006)

RKN (M. incognita)

Results-

(Ledger et al.,2006)

Uninfected tomato roots

Unhatched coiled juveniles

Freshly hatched J2s Females

Males

Lemmi9 primers

RT-PCR

In situ hybridization

This picture shows that MI-eng-2 gene is present in sub-ventral oesophageal glands of M. incognita

Cont…

(Ledger et al.,2006)

(Ledger et al.,2006)

Evolution of Cellulase Genes

Linker domain

CBD

Catalytic domain

GHF16

Large group of nematodes

Worldwide distributed

Most species are solely fungal feeders and all species rely on fungi as a food source at some stage of their life cycle

B. xylophilus - Unique feeding habit i.e. feed on live trees & fungus (both)

(Kikuchi et al., 2005)

Bursaphelenchus

β-1,3-Glucanases catalyse the hydrolysis of β-1,3-D-glucosidic linkages in β-1,3-D-glucan

This polymer (β-1,3-glucan ) is a major structural component of fungal cell walls thus the protein is allowing the nematodes to feed on fungus

This shows that β-1,3-glucanases play an important role in the life cycle of this nematode

β-1,3-Glucanases catalyse the hydrolysis of β-1,3-D-glucosidic linkages in β-1,3-D-glucan

This polymer (β-1,3-glucan ) is a major structural component of fungal cell walls thus the protein is allowing the nematodes to feed on fungus

This shows that β-1,3-glucanases play an important role in the life cycle of this nematode

(Kikuchi et al., 2005)

β-1,3-glucanase in pine wilt nematode

Enzymes with their Glycosil Hydrolase Families-

Bacterial , Nematodes - GHF 16

Plant and fungus- GHF 17

Sequences shows that Pinewood nematode B. xylophilus has acquired endo-β-1,3-glucanase (GHF16) by HGT from bacteria like Xanthomonas axonopodis , Pseudomonas spp. etc

(Kikuchi et al., 2005)

Cont…

RESULTS -

Southern-blot analysis of Bx-lam16A Genomic DNA from B. xylophilus (N) and B. cinerea (F) were digested with EcoRI (lanes 1 and 3) or HindIII (lanes 2 and 4). The blot was hybridized with a probe generated from Bx-lam16A cDNA

(Kikuchi et al., 2005)

Southern-blot analysis

in situ hybridization

Localization by in situ hybridization of Bx-lam16A transcripts in the oesophageal gland cells of B. xylophilus adult female with antisense (A) and sense (B) Bx-lam16A digoxigenin - labelled cDNA probes

Phylogenetic tree of selected GHF16

Phylogenetic tree of selected GHF16 β-1,3-glucanases and β-1,3-glucanase-like proteins generated using maximum-likelihood analysis

(Kikuchi et al., 2005)

GHF45

Life Cycle of B. xylophilus

B. xylophilus is part of clade of PPN (RKN & CN) and not directly related to FFN

As this nematode feed on fungus, some genes are horizontally transmitted to the ancestor by fungus

Researchers identified a cellulase gene: Bx-eng-1 from this nematode which is from GHF 45 and is similar to the fungus

Southern- blot Analysis of Bx-eng-1 digested with EcoR1 or HindIII and the blot was hybridized with a probe generated from Bx-eng-1 cDNA.

(Kikuchi et al., 2004)

GHF 45

Southern- blot Analysis of Bx-eng-1

Localization of Bx-eng-1 transcripts in the oesophageal gland cells of B. xylophilus adult female by in situ hybridization. Nematode sections were hybridized with antisense (A) or sense (B) Bx-eng-1 digoxigenin- labelled cDNA probes

Localization of Bx-eng-1 transcripts in the oesophageal gland cells of B. xylophilus adult female by in situ hybridization. Nematode sections were hybridized with antisense (A) or sense (B) Bx-eng-1 digoxigenin- labelled cDNA probes

in situ hybridization

(Kikuchi et al., 2004)

Endosymbiotic and squid bacteria

mingle

Rhabditid enters in it & quiescence

occurs

Squid with Lux operon came at

seashore

Bacteria start feeding on dead

squid

Nematode feed on bacteria in carcass

(Necromeny)

Lux gene acquired by endosymbiotic

bacteria

Evolution of Heterorhabditis-Photorhabdus symbiosis with a marine Lux operon

(Poinar, 1993)

Wolbachia DNA

Wolbachia is a symbiont in the most filarial worm

Infected filarial nematodes depend on Wolbachia for proper development and survival

Two Filarial species Acanthocheilonema viteae and Onchocerca flexuosa were found without Wolbachia but with DNA of Wolbachia

Researchers identified 49 Wolbachia-like DNA sequences in A. viteae and 114 DNA sequences in O. flexuosa

(McNulty et al., 2010)

Filarial worm with endosymbionts DNA

HGT is an important evolutionary novelty present in nature

Nematode acquired several parasitic genes by horizontal gene transfer from bacteria, fungus & other microbes to become a potential parasite of plant, animals and other invertebrates

Because of HGT Pine wilt nematode got a unique capacity to feed on both live plants and fungi

Heterorhabditid formed a symbiotic association with Photorhabdus and evolved as EPN

Some filarial worms are able to survive without Wolbachia as they have acquired DNA of Wolbachia via HGT which plays an important role in their development & survival