Transcript Discover the Microbes Within! The Wolbachia Project

Discover the Microbes Within!
The Wolbachia Project
Real-world Research in the Classroom
Discover the Microbes Within!
The Wolbachia Project
 1,000,000,000,000,000,000 insects on the planet
 Insects present on earth for ~500 MY
 85% of all animal species are insects (1-30 million species)
 20% of all insect species harbor heritable symbionts called
Wolbachia
 Scientists cant discover them all on their own.
 YOU ARE OUR BIGGEST ASSET TO SCIENTIFIC
DISCOVERY AND REAL RESEARCH!
What is symbiosis?
• The living together of dissimilar organisms
(de Bary 1859)
• Often for mutual benefit, but also parasitism
and commensalism
• Endosymbiosis is when one organism lives
and replicates inside another one
Vertically-transmitted
(i.e., Inherited)
Symbionts
Insect egg with
symbiotic bacteria
Insect egg
Credit: Michael Clark & Seth Bordenstein
Ways that vertically transmitted microbes
can increase in frequency
• Increase host survival & reproduction (mutualism)
– Very common
Why might vertical transmission be associated with mutualistic
effects on hosts?
• Most famous cases are the lineages leading to organelles
– Mitochondria evolved from the alpha-Proteobacteria about 2 billion
years ago
– Chloroplasts evolved from cyanobacteria about 1 billion years ago
Must heritable symbionts always
be beneficial?
Reproductive
Parasites
Asymmetric (Uniparental) Inheritance
Dead end
Distortion of
the sex-ratio
Wolbachia are Infectious Widowmakers!
Male-Killing
Parthenogenesis
Feminization
Reproductive Parasitism:
each of these reproductive
distortions leads to more
infected females in the insect
host species
Phylogeny of Wolbachia
NATURE REVIEWS | microbiology Vol OCT 2008, p741
Wolbachia Induced Phenotypes
NATURE REVIEWS | microbiology Vol OCT 2008, p741
Cytoplasmic Incompatibility (CI)
CI
Testes
x
x
x
x
=
X
=
Wolbachiainfected
offspring
=
=
Late prophase
Prometaphase
Embryo
Metaphase
Telophase
Uninfected
offspring
Wolbachiainfected
offspring
Courtesy of
U. Tram
Wolbachia
Host
Wolbachia-to-host lateral gene
transfer in Drosophila
ananassae
NATURE REVIEWS | microbiology Vol OCT 2008, p741
Imagine…
…physicians across the United States…documenting similar events. In each case, an unfertilized egg in a
woman had spontaneously begun to develop, ultimately producing a healthy female baby.
One young researcher, who had analyzed the timing and locales of the virgin births, suggested a spreading
infection might be causing the incidents. The Federal Centers for Disease Control and Prevention in Atlanta
quickly dismissed the idea, calling it "ridiculous."
Several months later came a well-publicized report in the Journal of the American Medical Association concluding
that the number of infertile couples was rising rapidly worldwide. The international uproar intensified when
physicians began to observe another reproductive curiosity: Some newborns that were genetically male
appeared to be female. One week, the New England Journal of Medicine and the National Enquirer ran articles
with the headline, "Is this the end of mankind, or just men?"
Science fiction? Definitely. For many insect species and other arthropods, however, the truth can be as strange as
fiction when bacteria known as Wolbachia are around.
By JOHN TRAVIS
Undesirable Sex Partners
Bacteria manipulate reproduction ofinsects and other species
The Wolbachia
Pandemic
The Wolbachia Pandemic
Insects
(at least 20% !)
Filarial nematodes
Crustaceans
(family Onchocercidae)
Chelicerates
2-6 million insect
species are infected
with Wolbachia!!
Arthropods
Nematodes
Number of scientific papers with Wolbachia in title
150
120
90
PCR detection of Wolbachia in ~20% of all insects
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Nobel prize awarded for PCR
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How Important Are
Wolbachia?
Human Health: Wolbachia may help control
the transmission of arthropod-borne diseases
(Malaria, Dengue fever, Filariasis, Trypanosomiasis, West Nile, Chagas)
Does Not
Transmit
Disease
Transmits
Disease
Reinfect vector
with transgenic
symbiont
Isolate and
culture
symbiont
Transform
symbiont
Anti-pathogen
gene
Drive via Cytoplasmic Incompatibility
Male
Female
X
X
X
X
Progeny
None (Incompatible)
Population Replacement
Transmits Disease
Does Not Transmit Disease
Human Health: Bill and Melinda Gates Foundation
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Wolbachia are Chemotherapy Targets for Curing River Blindness
& Elephantiasis Caused By Filarial Nematodes
Onchocerca volvlulus
Untreated
11 months post-treatment
Horeauf et al, 2003
Tissue Nematode (Roundworm):
Onchocerca volvulus and River
Blindness
• Transmitted by biting black
flies
• Larvae develop into adults in
subcutaneous tissues
• Adult females migrate via the
blood to the eyes, provoking
inflammatory reactions
• Coinfection with Wolbachia
bacteria causes river blindness
• Treatment: tetracycline and
ivermectin
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Overview of Onchocerciasis
• River blindness is the symptomatic
stage of a filarial infection
• It is a painful and debilitating disease.
• 18 million people in Africa are
infected
Overview of Onchocerciasis
• Symptoms include: visual impairment, rashes, lesions,
intense itching, depigmentation, inflammation of lymph
nodes, and general debilitation.
Overview of Onchocerciasis
• Disease is the result of infection by the parasitic worm,
Onchocerca volvulus.
• Females produce millions of microfilariae that migrate
through the body and cause a manifestation of symptoms
Overview of Onchocerciasis
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Simulans spp. (black flies) are vectors
Female black flies require a blood meal prior to egg laying
Microfilariae are taken in with infected blood
Transfer takes place when the fly bites uninfected person
CDC/DPD Summary Report
2001
• Black flies are infected with
Wolbachia bacteria
• Worldwide distribution
• Infection rates have been found to
be up to 76% of insects in some
regions
• Also found in millipedes,
crustaceans, and mites
• Wolbachia is passed horizontally
and vertically
Lancet 2005; 365: 2116–21
Tissue Nematode (Roundworm):
Wuchereria bancrofti
• Tropical infection spread by
mosquitoes
• Vector deposits larvae
which move into lymphatics
and develop
• Chronic infection causes
blockage of lymphatic
circulation and elephantitis,
massive swelling in the
extremities
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Tissue Nematode (Roundworm):
Filariasis due to W. bancrofti
• Endemic in central Africa, Mediterranean coast, parts of
Asia (China, Korea, Japan, the Philippines)
• Blood specimens may show microfilariae
• Acute symptoms include: fever,
lymphangitis/lymphadenitis
• Result of inflammatory response to molting adolescent
worms and dead adults in lymphatic vessels
• May involve any part of body, blocking lymphatic system
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Tissue Nematode (Roundworm):
Elephantitis
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Elephantiasis: Wucheria
bancrofti
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How do YOU
discover
the Wolbachia
within?
Integrated Set of Lab Exercises
From Organisms to Molecules and Back!
Lab 1 - Insect Identification (Biodiversity)
Lab 2 - Isolation of Insect and Wolbachia DNA (Molecular Biology)
Lab 3 - PCR of Wolbachia 16S rDNA (Molecular Biology)
Lab 4 - Presence/Absence of Amplicon (Molecular Biology)
Lab 5 - Evolutionary Sequence Analysis (Bioinformatics)
Features of These Labs
• Original Research in Lab Exercises
• YOU can make new discoveries
• Integrates across Science
Biodiversity -> Molecular -> Evolution
• Integrates Science and Technology
• Covers Emerging Fields
Microbial diversity, Bioinformatics,
Molecular Phylogeny
Discover the Microbes Within: The Wolbachia Project
Lab 1: Insect Identification
Coleoptera
Collembola
Diptera
Hemiptera
Lepidoptera
Orthoptera
beetles; 370,000
species
spring tails; 6,000
species
flies; 120,000
species
aphids; 67,500
species
butterflies, moths,
skippers; 140,000
species
grasshoppers,
crickets,
katydids; 17,000
species
Hymenoptera
Isoptera
Odonata
bees, wasps, ants;
108,000 species
termites; 1,900
species
dragonflies,
damselflies;
5,000 species
Dermaptera
Dictypotera
Ephemeroptera
earwigs; 1,200
species
cockroaches and
mantids; 6,000
species
mayflies; 2,000
species
Discover the Microbes Within: The Wolbachia Project
Lab 2: DNA Extraction
EXTRACT DNA FROM
YOUR SAMPLES
WHY?
Infected sample (+):
QuickTime™ and a
TIFF (Un co mpress ed ) de compres sor
are n eed ed to se e this p ictu re .
Uninfected Sample (-)
Insect
Insect
Mitochondria
Mitochondria
WOLBACHIA
-
Discover the Microbes Within: The Wolbachia Project
Lab 3: Polymerase Chain Reaction
Discover the Microbes Within: The Wolbachia Project
Lab 4: Gel Electrophoresis
Discover the Microbes Within: The Wolbachia Project
Lab 4: Gel Electrophoresis
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Discover the Microbes Within: The Wolbachia Project
Lab 5: Sequence Analysis
• NCBI
• BLAST
• No programming
skills required!
When It all comes together, it looks like this!
High School
MBL
ATGCGC
Primers, Insect controls,
Where Does Your Data Go?
To…
• Class reports
• Peers, family,
teachers, research
scientists
• Online database
(website repository for
your data)
Consequences
• New discoveries
• Collaborations
between research
scientists and high
schools
• Summer
“envisionships”
• Professional meetings
• Professional societies
• Journal publications