Biotecnologie applicate e
Evoluzione
Sandra Urbanelli - Daniele Porretta
Dipartimento di Biologia Ambientale
Via dei Sardi 70, 4° piano
We live in an exciting time to study Biotechnology
Technological advances have made data collection easier and cheaper than
we could ever have imagined just 10 years ago.
We can now synthesize and analyze large
data sets containing genomes,
transcriptomes, proteomes, and multivariate
phenotypes.
At the same time, society’s need for the results of biological research has
never been greater.
Solutions to many of the world’s most pressing problems will rely heavily on
biologists, collaborating across disciplines.
curing and preventing
diseases
Challenges for the New Biology for the 21st Century
feeding a global population
preserving
ecosystems and biodiversity,
Qual è la differenza di oggi rispetto a ieri?
Il tasso di estinzione attuale è dalle 100 alle 1000 volte maggiore rispetto al
tasso di estinzione di fondo stimato dai dati fossili
Se in passato la durata mediadella vita di una specie dimammiferi o di uccelli eracompresa tra 1 e 10 milioni dianni, nel XX° secolo essa si èridotta a 10.000 anni
La crescita demografica dell’uomo è alla base dell’impatto che osserviamo
La popolazione umana negli ultimi 200 anni è
passata da circa 500 milioni a oltre 7 miliardi
Food and fiber
HealthEnvironment
What is Biotechnology?
UN Convention on Biological Diversity, “biotechnology is technological
application that uses biological systems, living organisms or derivatives
there of, to make or modify products or processes for specific use”
The American Chemical Society defines biotechnology as the application
of biological organisms, systems, or processes by various industries to
learning about the science of life and the improvement of the value of
materials and organisms such as pharmaceuticals, crops, and livestock.
European Federation of Biotechnology, EFB, biotechnology is the
integration of natural science and organisms, cells, parts there of, and
molecular analogues for products and services.
Biotechnology is based on the basic biological sciences
• Biotecnologie tradizionali
1. trasformazione di prodotti di uso quotidiano grazie a processi
biologici (di cui non si conosceva la causa), in particolare i
processi fermentativi
2. selezione di varietà di piante e animali per uso commerciale o
industriale
• Le biotecnologie moderne
3. applicazione di tecnologie geniche, DNA ricombinante, per
modificare la funzione biologica di un organissmo o sviluppare
un nuovo prodotto
4. sviluppo industriale e tecnologico per la produzione di prodotti
chimici specifici (antibiotici, enzimi, aminoacidi, steroidi,
polisaccaridi)
Percorso formativo
Biotecnologie applicate ed Evoluzione
Though Dobzhansky’s statement is sometimes dismissed by biologists in other fields asself-promotion, recent advances in many areas of biology have shown it to be prophetic.
Our ability to apply evolutionary concepts to a wide range ofproblems has never been greater.
Career opportunities for evolutionary biologists may already be more plentiful outside academia than inside it.
Much of the biotechnology industry is concerned with creating biological molecules thathave specific functions. This goal-oriented enterprise has quickly embraced evolutionaryprinciples to direct the evolution of molecules in test tubes and, in so doing, hasprofoundly expanded the horizon and relevance of evolutionary biology. Evolutionaryprinciples are suddenly the material of multimillion dollar patents, leading industrialbiochemists to new drugs and other commercial molecules.
On a different front, the medical establishment, after long ignoring evolution, is faced withan on slaught of drug-resistant microbes, has seen monkey viruses jump into humans andaccelerate into epidemics, and must now use evolutionary principles to understand theworldwide dynamics of pathogens.
Evolution is socially relevant.
First, public perception of evolutionary biology is not up to date with the discipline.“Evolution” is still a bad word to many people, not only because it is perceived asconflicting with some religious views, but also because it is widely viewed as anirrelevant science with no social value. Acceptance of evolutionary biology is farmore likely when the public realizes that it holds the key to many socialimprovements
A second reason is that historical inertia in the training of evolutionary biologists hasresulted in a lack of exposure to socially relevant applications. Individuals trained asevolutionary biologists will have much to offer in solving these problems, but theyneed to be aware of these applications.
This is a recent discorvery because:
L’uomo ha applicato i principi della biologia
evolutiva per produrre prodotti biotecnologici
ancor prima di scoprire l’evoluzione e definire le
biotecnologie
Nearly all the common animals and plants we use today were
domesticated thousands of years ago, some (sheep, goats, dogs, wheat,
and rice) at least 9000 years ago.
the first experiments in applied
evolution.
Their impact was so profound as to make civilization possible by enabling
societies to switch from hunting and gathering to agriculture
Domestication
DomesticationBos taurus
Bos primigenius
Uro_Lascaux
The success of artificial selection is evident in its ultimate creation of selected
phenotypes well outside the extremes of the original species.
The major centers of primary domestication and dates for the earliest
domestication of various plant and animal species.
Riso 2.000 B.P.
Miglio 3.000 B.P.
Sorgo 4.000 BP.
Cotone 5.000 B.P.
Patata 4.500 B.P.
Maranta 8.000 BP.
Patata 7.000 B.P.
Quinoa 5.000 B.P.
Mais 9.000 B.P.
Fagioli 4.000 B.P.
Zucca 10.000 BP.
Marshelder 4.400 B.P.
Girasole 4.800 B.P.
Zucca 10.000 BP.
Manioca 8.000 B.P.
Peperoncino 5.000 B.P.
Orzo 10.000 B.P.
Grano 10.000 B.P.
Miglio 8.000 B.P.
Riso 8.000 B.P.
Quinoa 5.000 B.P.
Riso 8.000 B.P.
Nocciole 8.000 B.P.
Banana 7.000 B.P.
Taro 7.000 B.P.
The distribution of prehistoric agriculture, and early agriculturalist
expansion.
EARLY AGRICULTURALISTPOPULATION DIASPORAS ? FARMING, LANGUAGES, AND GENESPeter Bellwood Annu. Rev. Anthropol. 2001. 30:181–207
The origin and dispersal of domestic livestock species in the Fertile
Crescent.
Shaded areas show the general region and the approximate dates in calibrated years B.P. in which initial
domestication is thought to take place. Dates outside of the shaded areas show the approximate date when
the domesticate first appears in a region. Orange, goats (Capra hircus); blue, sheep (Ovis aries); green, cattle
(Bos taurus); fuscia, pigs (Sus scrofa).
M. A. Zeder, PNAS (2008) 105: 11597–11604.
A model of evolution by natural or artificial selection has three components:
(a) variation,
(b) inheritance, and
(c) differential reproductive success
Domestication probably started as a process of taming, then of captive
breeding, and finally of selecting for specific traits.
Seeking Agriculture’s Ancient Roots M. BALTER (2007) SCIENCE 316
Plant Domestication
Historically, artificial selection
The components of variation and inheritance were present but were often not
manipulated from their natural states, except for the occasional introduction of
novel strains or wild relatives into the breeding stock.
manipulation of differential reproductive
success.
Parents closest to the desired phenotype were chosen to produce the next
generation, and individuals that fell short of the ideal were omitted from the
breeding population.
Domestication
Decrease in genetic diversity in modern
crops during domestication due to
bottleneck events
An example of cotton (Gossypium) evolution under human selection and contemporary breeding
programs.
Plant Domestication
Crop wild relatives. These species represent
sources of novel variation for use in crop
improvement. (a) barley (wild barley (Hordeum
spontaneum) growing near Allepo, Syria); (b) coffee
(Coffea brassii), photograph by Australian Tropical
Herbarium; (c) rice (Oryza A genome wild
mpopulations, growing near Mareeba, Australia); (d)
grape (Cissus antarctica).
Pig domestication has been an integral part of the rise of agriculture and the
adoption of the agricultural practices throughout much of the world.
The domestication of plants and animals led to one of the most important
socioeconomic transitions in human history,
yet little is known about whether the process took place
in a limited number of geographic regions
or was a more widespread innovation
involving multiple, independent events.[
The wild boar is naturally occurring in much of Eurasia
and consists of a number of deeply divergent
populations.
It is the progenitor of the domestic pig, and thought to have
been domesticated on at least two occasions, once in the
Middle East and once in China. More domestications have
been suggested based on mtDNA phylogeography (e.g.
Larson et al. 2005),
L’uomo ha applicato i principi della biologia evolutiva per
produrre prodotti biotecnologici ancor prima di scoprire
l’evoluzione e definire le biotecnologie
Tale applicazione ha innescato effetti a catena sull’uomo stesso
e sull’ambiente alla base dei quali ci sono, ancora una volta, i
processi evolutivi
Dietary shifts
Switch from hunting and gathering to
agriculture
Health consequences
The human genome encompasses only a fraction of the total genetic
diversity found within humans.
The collective microbial communities inhabiting the human body,
contain a vast amount of genetic and functional diversity far
exceeding that of our own nuclear and mitochondrial genomes.
Human microbiome
The human microbiome: an astounding number of bacteria.
the number of bacterial cells (∼1014) in and on the human body
exceeds the number of human cells (∼1013) by at least an order of
magnitude.
the estimated number of unique bacterial genes in our ‘accessory
genome’ (∼3,300,000) exceeds the number of our own genes (∼22,000)
by a factor of 150
the number of cells:
the number of genes:
Despite being 1,000 times smaller than human cells, bacteria still make up
about 2% of adult body mass (1.5 kg), making them collectively equivalent
in size to the human brain (1.4 kg) or liver (1.6 kg), leading some to refer to
our resident microbes as an additional human organ.
The human–microbial relationship has also been compared to that of a
superorganism, like a colony of bees, or that of a holobiont,
like a coral reef.
The human microbiome: an astounding number of bacteria.
the body mass:
Human microbiome: function and dysfunction
Health Human Microbiome Project (HMP) in the United States
Metagenomics of the Human Intestinal Tract (MetaHIT) project in Europe.
Since the early 2000s, numerous studies have investigated the structure
and function of the human microbiome.
2008
previously viewed at best as passive
commensal or nuisances to be scrubbed or
flossed away, we now recognize that the
human oral, gut, skin, and uritogenital
microbiota play critical roles in maintaining
host health by performing essential
functions
in digestion and metabolism
vitamin production
immune system education and maintenance
by restricting the colonization, growth, reproduction, and virulence expression
of exogenous bacterial pathogens through resource competition.
The oral, gut, skin, and uritogenital microbiota play essential functions for
human health
On the other hand, poor diet, illness, stress, antimicrobial drugs, and other
environmental disruptions,
obesity,
type II diabetes,
irritable bowel disease,
colon cancer,
periodontal disease and dental
decay, atherosclerosis and
endocarditis,
autism, anxiety, and depression.
the ecology of the human
microbiome can transition
from a mutualistic to a
dysbiotic state,
The healthy human microbiome also plays host to a number of endemic,
but potentially acute, opportunistic pathogens.
microbiome plays a critical role in making us human, in keeping us
healthy, and in making us sick,
Streptococcus pneumoniae,
Haemophilus influenzae,
Neisseria meningitidis,
Clostridium difficile,
Propionibacterium acnes
Staphylococcus aureus
implicated in hospital and community-
acquired infections and pose particular risk
for the elderly and immunocompromised.
Alarmingly, multiple antibiotic resistant strains are increasingly being
detected in the normal oral and gastrointestinal microbiota of healthy
individuals.
This suggests that the use of antibiotic therapy, either through direct clinical
application or through indirect growth stimulating or prophylactic application in
livestock, impacts non-clinical targets and can result in long-term endogenous
reservoirs of antibiotic resistance.
Antibacterial therapies can themselves be disruptive to healthy bacterial
communities, leading to further complications. Therapeutic courses of broad-
spectrum antibiotics, for example, are known to disrupt gut and uritogenital
microbiota, where they may induce antibiotic-associated colitis and bacterial
vaginosis, respectively.
As a consequence, there is growing interest in probiotic and prebiotic therapies
for treating disrupted microbiomes, but lack of basic knowledge on what
constitutes a healthy microbiome, as well as a clearer understanding of the
transmission and formation of healthy microbiota, are limiting factors in the
development of these therapies.
Need for paleomicrobiology data
Although considerable effort has been invested in characterizing healthy gut and
oral microbiomes, recent investigations of rural, non-Western populations have
raised questions about whether the microbiota we currently define as normal
have been shaped by recent influences of modern Western diet, hygiene,
antibiotic exposure, and lifestyle.
When and how did our bacterial communities become distinctly human?
And what does this mean for our microbiomes today and in the future?
How do we acquire and transmit microbiomes and to what degree is this affected
by our cultural practices and built environments?
How have modern Western diets, hygiene practices, and antibiotic exposure
impacted ‘normal’ microbiome function?
Are we still in mutualistic symbiosis with our microbiomes, or are the so-called
‘diseases of civilization’—heart disease, obesity, type II diabetes, asthma,
allergies, osteoporosis—evidence that our microbiomes are out of ecological
balance and teetering on dysbiosis?
At an even more fundamental level, who are the members of the human
microbiome, how did they come to inhabit us, and how long have they been
there?
Who is ‘our microbial self’?
we know remarkably little about the diversity, variation, and evolution of the human
microbiome both today and in the past.
We are left with many questions:
Even the most extensive sampling of modern microbiota will provide
limited insight into Pre-Industrial microbiomes.
By contrast, the direct investigation of ancient microbiomes
from discrete locations and time points in the past would
provide a unique view into the coevolution of microbes and
hosts, host microbial ecology, and changing human health
states through time.
Ancient microbiome research
Upon death, the ecology of the human microbiome transforms
dramatically through the process of soft tissue decomposition.
With the exception of frozen and mummified remains, only two
microbiomes routinely produce substrates that, under favorable
conditions, persist after death in archaeological contexts: fecal
material of the gut microbiome may desiccate or mineralize to
produce coprolites, and dental plaque of the oral microbiome
calcifies in situ during life such that by the time of death it is already
in a semi-fossilized state known as dental calculus that resists
decomposition and thus continues to preserve after death.
Ancient microbiome research
Dental calculus
Dental calculus is a calcified bacterial biofilm that forms on the
surfaces of teeth, and it is found in all human populations, as
well as Miocene apes (12.5–8.5 Ma), Neanderthals, wild
chimpanzees, and a range of animals.
Among humans, both in the past and today,
the incidence of dental calculus is near-
ubiquitous among adults by age 30 and it is
not uncommon to observe dental calculus
deposits in excess of 100 mg in
archaeological assemblages of agricultural
populations.
a rich source of ancient
biomolecules,
Dental calculus: a rich source of ancient biomolecules,
Extracted DNA yields up to three orders of magnitude greater than from bone or dentine
of the same individual.
Importantly, dental calculus is unique among
ancient microbiome sources in that it does
not shed, remodel, or turnover; rather, it
forms incrementally through serial deposition
and mineralization in situ, making it a
layered record of human life history specific
to each person.
Two of the greatest dietary shifts in human evolution involved
the adoption of carbohydrate-rich Neolithic (farming) diets
(beginning ~10,000 years before the present) and the more
recent advent of industrially processed flour and sugar (in
~1850).
Sampling
34 prehistoric European
human skeletons:
11 males,
11 females
12 of unknown sex,
Dating from before the
Mesolithic period (before
farming) to the medieval
period.
The ancient samples were compared to modern calculus (n = 6)
and plaque (n = 13) samples.
Experimental design
We extracted bacterial DNA from sterilized ancient calculus samples (n = 34) and
generated PCR amplicon libraries of the 16S rRNA gene, targeting three
hypervariable regions (V1, V3 and V6) with barcoded primers
In addition, primers specific to Streptococcus mutans and Porphyromonas
gingivalis were used to detect oral pathogens in ancient dental calculus
Amplicons generated from
extracted samples and
multiple extraction blanks
were sequenced using both
conventional and
pyrosequencing
technology.
At the phylum level, the bacterial composition of ancient calculus was similar to
that of modern oral samples and sequences from the Human Oral Microbiome
Database (HOMD)30 but markedly distinct from the compositions identified for
laboratory reagents (extraction blanks) and environmental samples (soils,
sediments and water) and within the ancient teeth themselves
Results
The archaeological calculus was dominated by Firmicutes (33%), which was
found at a frequency comparable to those in both the HOMD (37%), and modern
oral samples (average 50%). In addition to Firmicutes, the ancient dental calculus
samples contained all 15 phyla commonly found in the modern human oral cavity,
with high percentages of Actinobacteria (19%), as is observed in modern
calculus deposits (7%).
Results
PCA plots of PC1 and PC2 (c) and PC2 and PC3 (d) that only include ancient and
modern oral pyrosequencing samples separated the hunter-gatherer (Mesolithic)
samples from modern, medieval and Neolithic samples.
Changes in the diversity and composition of oral microbiota.
The composition of oral microbiota underwent a distinct shift with the
introduction of farming in the early Neolithic period, with the earlier hunter-
gatherer groups having fewer caries- and periodontal disease–associated
taxa.
Overall, it is clear that
modern Europeans have
much lower oral microbial
diversity than either
Mesolithic or preindustrial
Neolithic groups
(P < 0.001, including fewer
bacteria associated with good
health, Ruminococcaceae).
After the transition to agriculture in the early Neolithic period, there was a
notable consistency in the composition of bacteria through the medieval period
(~400 years before the present), in parallel with the broad similarity of food-
processing technologies during these times. In contrast, today’s oral
environment is much less biodiverse and is dominated by potentially cariogenic
bacteria.
Major changes in carbohydrate intake in human history seem to have affected the
ecosystem of the mouth, opening up pathological niches for periodontal disease in
the early Neolithic period and caries in the recent past.
Overall, it is clear that modern Europeans have periodontal disease–associated
taxa (for example, Porphyromonas gingivalis similar to early agriculturists.
This is consistent with skeletal evidence showing marked increases in
periodontal disease following the transition to an agricultural diet,
suggesting a major impact on the human oral ecosystem around this time.
This is thought to be caused by increased amounts of soft carbohydrate
foods compared with hunter-gatherer diets.
Notably, the frequency of Streptococcus mutans is significantly higher in
modern samples than in preindustrial agricultural samples (P < 0.0001),
indicating that caries-associated bacteria have only become dominant after
medieval times.
This change is most likely associated with the onset of the Industrial Revolution,
which began some 200 years ago and represents the largest change in food
production and processing technology since the shift to farming. The Industrial
Revolution saw the production of refined grain and concentrated sugar from
processed sugar beet and cane, generating mono- and disaccharides, which are
the main substrates for the microbial fermentation that lowers plaque pH and
causes enamel demineralization.
Conclusions
Biotecnologie applicate ed Evoluzione
Scopo del corso: studio dei meccanismi genetici ed evolutivi alla
base delle risposte degli organismi all’ambiente
Approccio: genetico-ecologico
Analisi dei principi di base della genetica ecologica, condotta a diversi livelli
(molecolare, di organismo e di popolazione), attraverso la prospettiva
evolutiva
Obiettivi formativi:
- comprendere i meccanismi che sono alla base della formazione e
mantenimento della diversità genetica
- capire l’importanza dei meccanismi adattativi in ambito biotecnologico
- comprendere il destino dei biotech nei sistemi naturali, recettori ultimi di
tali prodotti
- rilevare costi e benefici dell’applicazione delle biotecnologie.
Prima parte:
comprensione dei processi microevolutivi alla base dell’origine e mantenimento
della diversità biologica
The Modern Synthesis
Prima parte:
comprensione dei processi microevolutivi alla base dell’origine e mantenimento
della diversità biologica
Seconda parte:
applicazione dei principi dell’evoluzione nello sviluppo ed applicazione delle
biotecnologie
Food and fiber
HealthEnvironment
Genetic markers
Genetic population
models
Strumenti e Attività pratiche
Results
Raw data
Input files
Data analyses
QUESTION
Experimental
design
ANSWER