![UNAFold 5
(or DG37
) and enthalpy, ∆H, are easily computed. From this, the
entropy, ∆S, is easily computed as:
∆ ×
∆ − ∆
S =
H G
T
1000 ,
where T is the hybridization temperature (K) and the factor of 1000
expresses ∆S in e.u. (entropy units; 1 e.u. = 1cal/mol/K). The
terms “free energy,” “enthalpy,” and “entropy” are really changes in
free energy, enthalpy, and entropy with respect to the “random coil
state.” In reality, “random coil” is a large ensemble of states. The free
energy and enthalpy of this ensemble of states may be (and is) set to
zero. Only entropy cannot be arbitrarily assigned. Thus a ∆S term
is Rln(N/Nref
), where R is the universal gas constant, 1.9872 e.u.;
N is the number of states consistent with hybridization; and Nref
is the
number of states, always larger, in the unconstrained “random coil.”
The simplest model for hybridization assumes that there are
two states: hybridized, where the structure is unique and known,
and “random coil.” It is then a simple matter to deduce a melt-
ing temperature, Tm
, at which half of the dimers have dissociated.
The formula is given by:
T
f
m =
H
S + R
1000
ln(Ct
×
∆
∆ )
, [1]
where ∆H, ∆S (expressed in e.u.) and R have already been defined, Ct
is the total strand concentration, and f = 4 when the two strands are
different and f = 1 when self-hybridization takes place. This formula
implicitly assumes that both strands are present in equal concentra-
tions. Competition with folding and with other possible hybridiza-
tions, such as homodimer formation, is not considered at all.
UNAFold offers a number of increasingly sophisticated ways
to compute melting temperatures and also computes entire melt-
ing profiles: UV absorbance at 260nm, heat capacity (Cp
), and
mole fractions of various single and double-stranded molecu-
lar species as a function of temperature. Even when Equation
[1] is used, the hybridization is computed by minimizing free
energy. The various levels of complexity and choice of methods
are described in the following.
It is important to issue a warning to those who wish to apply
these methods to microarrays. Hybridization on microarrays is
complicated by the fact that one of each hybridizing pair is immo-
bilized. It is difficult to compute the effective “solution concentra-
tion” for such molecules, and diffusion is probably an important
factor in slowing the time necessary to reach equilibrium. How-
ever, even for hybridization in solution, kinetic simulations that
treat different probe-target pairs independently can lead to totally
incorrect results for the time required to reach equilibrium as well
as the equilibrium concentrations themselves (21).](https://image.slidesharecdn.com/38331-250415011217-e4d1556d/85/Bioinformatics-Structure-Function-and-Applications-1st-Edition-Nicholas-R-Markham-22-320.jpg)
![From the quantity of air which the lungs receive, some conception
may be formed of the amount of obnoxious matter which may be
introduced into the system through these portals.
At each inspiration there enter the lungs of an ordinary-sized
person about 20 cubic inches of air. There are 20 respirations in a
minute: 400 cubic inches of air must therefore enter in one minute;
14 cubic feet in one hour, and 366 cubic feet, or 36 hogsheads, in
one day. To meet this the heart sends into the lungs at each
contraction two ounces of blood; there are 75 pulsations in a
minute, during which 150 ounces are propelled into the lungs; a
quantity which gives 562 pounds in one hour and 24 hogsheads in
24 hours.
The main purpose for bringing these enormous quantities of air
and blood together, with such velocity, is to provide for the
enormous waste which is caused by the rapid and unceasing
mutation of organic matter. The activity of an organ is sustained at
the expense of the matter of which it is composed. No thought
passes through the mind, but an equivalent portion of the substance
of the brain is consumed; no nervous current flows along the
nervous conducters, but a corresponding portion of nervous tissue is
used up; no muscular movement, no glandular secretion, takes place
without a proportionate waste of muscle and of gland. What must be
the amount of supply required to meet this waste, when able-bodied
men employed in their ordinary labour lose from 2 lbs. to 5 lbs. and
upwards of their weight twice a day.[8]
Some physiologists of
eminence have estimated that in order to supply that waste, there
passes in the course of every 24 hours as much fluid through the
thoracic duct[9]
as equals the whole quantity of blood in the body.
8. See Experiments on the daily loss of weight sustained by workmen employed
in gas-works.—Philosophy of Health, 11th Edit. p. 284, et seq.
9. The tube which conveys the debris of the body, together with the nutritious
part of the food,—both measures of change or waste.
The results of the highly interesting experiments recently made by
Professor Graham on the part taken by the active agent in all these](https://image.slidesharecdn.com/38331-250415011217-e4d1556d/85/Bioinformatics-Structure-Function-and-Applications-1st-Edition-Nicholas-R-Markham-30-320.jpg)
![Strictly speaking, however, all that we really know is this—that
where certain conditions exist, epidemics break out and spread; that
where those conditions do not exist, epidemics do not break out and
spread; and that where those conditions did exist, but have been
removed, thereupon epidemics cease.
We call those conditions Causes, Predisposing or Localizing
Causes, but how they act, whether by accumulating decomposing
organic matter in the blood, or in what other way, we have no
certain knowledge.
One further fact however is ascertained, that where any one of
these predisposing causes is present, epidemics break out and
spread just as readily as when all are present together.
Where there is overcrowding alone, for example, epidemics break
out and spread. Where there is decomposing filth alone, epidemics
break out and spread; and so of the whole number. The removal of
one of these causes, therefore, or the removal of two or three of
them, will not suffice for safety; every one must be removed before
there can be safety.
This we know; all beyond this is conjecture, but as to the most
probable of these conjectures, some who have thought on this
subject believe that the preponderance of evidence justifies the
conclusion that the predisposing causes may themselves become
efficient causes; that instances in which they actually do so, are
constantly passing before our eyes; that it is practicable to
manufacture fever and even epidemic fever to any amount by
placing a population under certain known conditions; that it is
practicable to prevent the outbreak of epidemics altogether by
placing the population under certain other conditions;[10]
that the
prevalence of the predisposing causes in particular localities, in
certain intensities, is sufficient to produce local epidemic outbreaks;
that the prevalence of such causes in such intensities, joined to
some general conditions of the atmosphere, such as the
meteorological conditions which have been enumerated, particularly
those which favour the accumulation and concentration of the](https://image.slidesharecdn.com/38331-250415011217-e4d1556d/85/Bioinformatics-Structure-Function-and-Applications-1st-Edition-Nicholas-R-Markham-32-320.jpg)
![reigns constantly a kind of half daylight, a sort of obscurity, of which
our forests of pines, oaks, and beech trees afford no example;
forming a carpet of verdure, the dark tint of which augments the
splendour of the aërial light.”
With this luxuriance of vegetation is combined a corresponding
abundance of animal life. The earth and air teem with living
creatures.
“The mould,” observes the same distinguished traveller, “contains
the spoils of innumerable quantities of reptiles, worms, and insects.
Wherever the soil is turned up we are struck with a mass of organic
substances, which by turns are developed, transformed, and
decomposed. Nature in these climates appear more active, more
fruitful, we might say more prodigal of life.”
The air is still more alive than the land. Insects fill the lower strata
of the atmosphere to the height of fifteen or twenty feet, like a
condensed vapour. It is estimated that a cubic foot of air is often
peopled by a million of winged insects, which contain a caustic and
venomous liquid, several species being nearly two lines (1.8) long.
When two persons who have their home in these regions meet in
the morning, the first questions they address to each other are,
“How did you find the zancudoes during the night?” “How are we to-
day for the mosquitoes?” An ancient form of Chinese politeness,
showing the ancient state of that country, was—“Have you been
incommoded in the night by serpents?”
It appears that there are still inhabited places in which the
Chinese compliment on the serpents might be added to that of the
mosquitoes.
Proportionate to this prodigality of organic life is the amount of
organic decomposition, the products of which are poured into the
atmosphere and suspended in the surrounding vapour and fog,[11]
to
which they give a decided and often a highly offensive odour.
11. See note, p. 16.](https://image.slidesharecdn.com/38331-250415011217-e4d1556d/85/Bioinformatics-Structure-Function-and-Applications-1st-Edition-Nicholas-R-Markham-36-320.jpg)
![constitute the rule of its existence. Places, not persons, comprehend
the whole history, the etiology of the disease. Places, not persons!
Let the emphatic words be dinned into the ears of the Lords of the
Treasury, of Trade and Plantations, until they acquire the force of a
creed, which will save them hereafter from the absurdity of
enforcing a quarantine[12]
in England against an amount of solar heat
of which its climate is insusceptible. Let them further be repeated in
the Schools of Medicine until the Professors become ashamed of
imbuing the minds of the young with prejudice and false belief,
which, should they ever visit warmer climates, may cause them to
be eminently mischievous in vexing the commerce and deeply and
injuriously agitating the public mind of whatever community may
have received them.”
12. See Cases of the Eclair, Dygden, &c., post.
Climate differs not only in different countries but in different parts
of the same country. The climate of the country is different from that
of the city. The climate of every city, town, and village, differs from
that of every other. The temperature, the moisture, and the other
meteorological conditions of different districts, nay, even of different
streets in the same town, vary to such a degree as to influence
materially their relative salubrity and the prevalence or absence of
particular classes of disease. These local climatic conditions and their
connection with prevalent diseases, have not as yet received due
attention: when they shall have received it—and they will receive it—
a new light will be shed on local epidemics.
I pass now to Civilization.
We have no sufficient knowledge of the state of the people and of
their diseases, in any of the civilized nations of antiquity, to trace the
relation between them. The authentic history of periods,
comparatively near to our own time, as far as concerns the diseases
of the people, goes scarcely further back than the 14th century. The
first great epidemic, to which I have so often called attention,
occurred in that century, and we have reliable evidence, both of the](https://image.slidesharecdn.com/38331-250415011217-e4d1556d/85/Bioinformatics-Structure-Function-and-Applications-1st-Edition-Nicholas-R-Markham-41-320.jpg)
![thousand were burnt alive. Whoever showed them compassion and
endeavoured to protect them were put upon the rack and burnt with
them. In numerous instances these unhappy people, driven to
despair, assembled in their own habitations, to which they set fire
and consumed themselves with their families. The noble and the
mean bound themselves by an oath to extirpate them from the face
of the earth by fire and sword.
In England this relentless cruelty took particularly the shape of
burning innocent people under the name of witches; an infatuation
which pervaded all classes from the highest to the lowest, affording
a melancholy exemplification of the close alliance between credulity
and cruelty.[13]
13. The number of wretched beings condemned and executed for this imaginary
crime at the Assizes of Suffolk and Essex alone, in the year 1646, amounted
to two hundred. Dr Zachary Gray affirms that he had seen an authentic
account of persons who had so suffered in the whole of England, amounting
to from three to four thousand. So late as the year 1697 seven persons,
three men and four women, were burnt at Paisley for this alleged crime. We
seldom sufficiently consider how near we are to those times of dreadful
superstition and cruelty! How short a period it is since the light of a brighter
day dawned upon us!
But in the midst of these terrible disorders, changes which had
been in silent operation during several centuries began to produce
visible results. The independent power of the nobles had been
suppressed; the feuds that raged between them, filling the country
with disorder and bloodshed, had been put down; the supremacy of
the law had been established; property and life had become more
secure; industry had taken a surprising start; the practical abolition
of serfdom had been to a large extent effected; and at last came the
final breaking up of the feudal system in the reign of Henry VII. by
the passing of the law authorizing the alienation of land.
About the middle of the fifteenth century improvements in the
condition of the people, which had been gradually effected by these
changes, were accelerated by a succession of events that gave an
extraordinary impulse to the human mind, just aroused from the](https://image.slidesharecdn.com/38331-250415011217-e4d1556d/85/Bioinformatics-Structure-Function-and-Applications-1st-Edition-Nicholas-R-Markham-48-320.jpg)
Bioinformatics Structure Function and Applications 1st Edition Nicholas R. Markham
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- 5. Bioinformatics Structure Function and Applications 1st Edition Nicholas R. Markham Digital Instant Download Author(s): Nicholas R. Markham, Michael Zuker (auth.), Jonathan M. Keith PhD (eds.) ISBN(s): 9781603274296, 1603274294 Edition: 1 File Details: PDF, 19.09 MB Year: 2008 Language: english
- 8. 460. Essential Concepts in Toxicogenomics, edited by Donna L. Mendrick and William B. Mattes, 2008 459. Prion Protein Protocols, edited by Andrew F. Hill, 2008 458. Artificial Neural Networks: Methods and Applica- tions, edited by David S. Livingstone, 2008 457. Membrane Trafficking, edited by Ales Vancura, 2008 456. Adipose Tissue Protocols, Second Edition, edited by Kaiping Yang, 2008 455. Osteoporosis, edited by Jennifer J. Westendorf, 2008 454. SARS- and Other Coronaviruses: Laboratory Proto- cols, edited by Dave Cavanagh, 2008 453. Bioinformatics, Volume II: Structure, Function and Applications, edited by Jonathan M. Keith, 2008 452. Bioinformatics, Volume I: Data, Sequence Analysis and Evolution, edited by Jonathan M. Keith, 2008 451. Plant Virology Protocols: From Viral Sequence to Pro- tein Function, edited by Gary Foster, Elisabeth Johansen, Yiguo Hong, and Peter Nagy, 2008 450. Germline Stem Cells, edited by Steven X. Hou and Shree Ram Singh, 2008 449. Mesenchymal Stem Cells: Methods and Protocols, edited by Darwin J. Prockop, Douglas G. Phinney, and Bruce A. Brunnell, 2008 448. Pharmacogenomics in Drug Discovery and Develop- ment, edited by Qing Yan, 2008 447. Alcohol: Methods and Protocols, edited by Laura E. Nagy, 2008 446. Post-translational Modification of Proteins: Tools for Functional Proteomics, Second Edition, edited by Christoph Kannicht, 2008 445. Autophagosome and Phagosome, edited by Vojo Deretic, 2008 444. Prenatal Diagnosis, edited by Sinhue Hahn and Laird G. Jackson, 2008 443. Molecular Modeling of Proteins, edited by Andreas Kukol, 2008. 442. RNAi: Design and Application, edited by Sailen Barik, 2008 441. Tissue Proteomics: Pathways, Biomarkers, and Drug Discovery, edited by Brian Liu, 2008 440. Exocytosis and Endocytosis, edited by AndreiI.Ivanov, 2008 439. Genomics Protocols, Second Edition, edited by Mike Starkey and Ramnanth Elaswarapu, 2008 438. Neural Stem Cells: Methods and Protocols, Second Edition, edited by Leslie P. Weiner, 2008 437. Drug Delivery Systems, edited by Kewal K. Jain, 2008 436. Avian Influenza Virus, edited by Erica Spackman, 2008 435. Chromosomal Mutagenesis, edited by Greg Davis and Kevin J. Kayser, 2008 434. Gene Therapy Protocols: Volume 2: Design and Char- acterization of Gene Transfer Vectors, edited by Joseph M. LeDoux, 2008 433. Gene Therapy Protocols: Volume 1: Production and In Vivo Applications of Gene Transfer Vectors, edited by Joseph M. LeDoux, 2008 432. Organelle Proteomics, edited by Delphine Pflieger and Jean Rossier, 2008 431. Bacterial Pathogenesis: Methods and Protocols, edited by Frank DeLeo and Michael Otto, 2008 430. Hematopoietic Stem Cell Protocols, edited by Kevin D. Bunting, 2008 429. Molecular Beacons: Signalling Nucleic Acid Probes, Methods and Protocols, edited by Andreas Marx and Oliver Seitz, 2008 428. Clinical Proteomics: Methods and Protocols, edited by Antonia Vlahou, 2008 427. Plant Embryogenesis, edited by Maria Fernanda Suarez and Peter Bozhkov, 2008 426. Structural Proteomics: High-Throughput Methods, edited by Bostjan Kobe, Mitchell Guss, and Huber Thomas, 2008 425. 2D PAGE: Sample Preparation and Fractionation, Volume 2, edited by Anton Posch, 2008 424. 2D PAGE: Sample Preparation and Fractionation, Volume 1, edited by Anton Posch, 2008 423. Electroporation Protocols: Preclinical and Clinical Gene Medicine, edited by Shulin Li, 2008 422. Phylogenomics, edited by William J. Murphy, 2008 421. Affinity Chromatography: Methods and Protocols, Sec- ond Edition, edited by Michael Zachariou, 2008 420. Drosophila: Methods and Protocols, edited by Christian Dahmann, 2008 419. Post-Transcriptional Gene Regulation, edited by Jeffrey Wilusz, 2008 418. Avidin–Biotin Interactions: Methods and Applications, edited by Robert J. McMahon, 2008 417. Tissue Engineering, Second Edition, edited by Hannsjörg Hauser and Martin Fussenegger, 2007 416. Gene Essentiality: Protocols and Bioinformatics, edited by Svetlana Gerdes and Andrei L. Osterman, 2008 415. Innate Immunity, edited by Jonathan Ewbank and Eric Vivier, 2007 414. Apoptosis in Cancer: Methods and Protocols, edited by Gil Mor and Ayesha Alvero, 2008 413. Protein Structure Prediction, Second Edition, edited by Mohammed Zaki and Chris Bystroff, 2008 412. Neutrophil Methods and Protocols, edited by Mark T. Quinn, Frank R. DeLeo, and Gary M. Bokoch, 2007 411. Reporter Genes: A Practical Guide, edited by Don Anson, 2007 410. Environmental Genomics, edited by Cristofre C. Martin, 2007 409. Immunoinformatics: Predicting Immunogenicity In Silico, edited by Darren R. Flower, 2007 408. Gene Function Analysis, edited by Michael Ochs, 2007 407. Stem Cell Assays, edited by Vemuri C. Mohan, 2007 406. Plant Bioinformatics: Methods and Protocols, edited by David Edwards, 2007 405. Telomerase Inhibition: Strategies and Protocols, edited by Lucy Andrews and Trygve O. Tollefsbol, 2007 METHODS IN MOLECULAR BIOLOGY™ John M. Walker, SERIES EDITOR
- 9. ME T H O D S I N MO L E C U L A R BI O L O G Y ™ Bioinformatics Volume II Structure, Function and Applications Edited by Jonathan M. Keith, PhD School of Mathematical Sciences, Queensland University of Technology, Brisbane, Queensland,Australia
- 10. ISBN: 978-1-60327-428-9 e-ISBN: 978-1-60327-429-6 ISSN 1064-3745 e-ISSN: 1940-6029 DOI: 10.1007/978-1-60327-429-6 Library of Congress Control Number: 20082922946 © 2008 Humana Press, a part of Springer Science+Business Media, LLC All rights reserved. This work may not be translated or copied in whole or in part without the written permission of the publisher (Humana Press, 999 Riverview Drive, Suite 208, Totowa, NJ 07512 USA), except for brief excerpts in connection with reviews or scholarly analysis. Use in connection with any form of information storage and retrieval, electronic adaptation, computer software, or by similar or dissimilar methodology now known or hereafter developed is forbidden. The use in this publication of trade names, trademarks, service marks, and similar terms, even if they are not identified as such, is not to be taken as an expression of opinion as to whether or not they are subject to proprietary rights. While the advice and information in this book are believed to be true and accurate at the date of going to press, nei- ther the authors nor the editors nor the publisher can accept any legal responsibility for any errors or omissions that may be made. The publisher makes no warranty, express or implied, with respect to the material contained herein. Cover illustration: Fig. 1A, Chapter 23, “Visualization,” by Falk Schreiber (main image); and Fig. 4, Chapter 5, “The Classification of Protein Domains,” by Russell L. Marsden and Christine A. Orengo (surrounding images) Printed on acid-free paper 9 8 7 6 5 4 3 2 1 springer.com Editor Jonathan M. Keith School of Mathematical Sciences Queensland University of Technology Brisbane, Queensland, Australia j.keith@qut.edu.au Series Editor John Walker Hatfield, Hertfordshire AL10 9NP UK
- 11. Preface Bioinformatics is the management and analysis of data for the life sciences. As such, it is inherently interdisciplinary, drawing on techniques from Computer Science, Statis- tics, and Mathematics and bringing them to bear on problems in Biology. Moreover, its subject matter is as broad as Biology itself. Users and developers of bioinformatics methods come from all of these fields. Molecular biologists are some of the major users of Bioinformatics, but its techniques are applicable across a range of life sciences. Other users include geneticists, microbiologists, biochemists, plant and agricultural scientists, medical researchers, and evolution researchers. The ongoing exponential expansion of data for the life sciences is both the major challenge and the raison d’être for twenty-first century Bioinformatics. To give one example among many, the completion and success of the human genome sequencing project, far from being the end of the sequencing era, motivated a proliferation of new sequencing projects. And it is not only the quantity of data that is expanding; new types of biological data continue to be introduced as a result of technological development and a growing understanding of biological systems. Bioinformatics describes a selection of methods from across this vast and expanding discipline. The methods are some of the most useful and widely applicable in the field. Most users and developers of bioinformatics methods will find something of value to their own specialties here, and will benefit from the knowledge and experience of its 86 con- tributing authors. Developers will find them useful as components of larger methods, and as sources of inspiration for new methods. Volume II, Section IV in particular is aimed at developers; it describes some of the “meta-methods”—widely applicable mathematical and computational methods that inform and lie behind other more special- ized methods—that have been successfully used by bioinformaticians. For users of bioin- formatics, this book provides methods that can be applied as is, or with minor variations to many specific problems. The Notes section in each chapter provides valuable insights into important variations and when to use them. It also discusses problems that can arise and how to fix them. This work is also intended to serve as an entry point for those who are just beginning to discover and use methods in bioinformatics. As such, this book is also intended for students and early career researchers. As with other volumes in the Methods in Molecular Biology™ series, the intention of this book is to provide the kind of detailed description and implementation advice that is crucial for getting optimal results out of any given method, yet which often is not incorporated into journal publications. Thus, this series provides a forum for the com- munication of accumulated practical experience. The work is divided into two volumes, with data, sequence analysis, and evolution the subjects of the first volume, and structure, function, and application the subjects of the second. The second volume also presents a number of “meta-methods”: techniques that will be of particular interest to developers of bioinformatic methods and tools. Within Volume I, Section I deals with data and databases. It contains chapters on a selection of methods involving the generation and organization of data, including v
- 12. sequence data, RNA and protein structures, microarray expression data, and func- tional annotations. Section II presents a selection of methods in sequence analysis, beginning with multiple sequence alignment. Most of the chapters in this section deal with methods for discovering the functional components of genomes, whether genes, alternative splice sites, non-coding RNAs, or regulatory motifs. Section III presents several of the most useful and interesting methods in phylogenetics and evolution. The wide variety of topics treated in this section is indicative of the breadth of evolution research. It includes chapters on some of the most basic issues in phylogenet- ics: modelling of evolution and inferring trees. It also includes chapters on drawing infer- ences about various kinds of ancestral states, systems, and events, including gene order, recombination events and genome rearrangements, ancestral interaction networks, lateral gene transfers, and patterns of migration. It concludes with a chapter discussing some of the achievements and challenges of algorithm development in phylogenetics. In Volume II, Section I, some methods pertinent to the prediction of protein and RNA structures are presented. Methods for the analysis and classification of structures are also discussed. Methods for inferring the function of previously identified genomic elements (chiefly protein-coding genes) are presented in Volume II, Section II. This is another very diverse subject area, and the variety of methods presented reflects this. Some well-known techniques for identifying function, based on homology, “Rosetta stone” genes, gene neighbors, phylogenetic profiling, and phylogenetic shadowing are discussed, alongside methods for identifying regulatory sequences, patterns of expres- sion, and participation in complexes. The section concludes with a discussion of a technique for integrating multiple data types to increase the confidence with which functional predictions can be made. This section, taken as a whole, highlights the opportunities for development in the area of functional inference. Some medical applications, chiefly diagnostics and drug discovery, are described in Volume II, Section III. The importance of microarray expression data as a diagnostic tool is a theme of this section, as is the danger of over-interpreting such data. The case study presented in the final chapter highlights the need for computational diagnostics to be biologically informed. The final section presents just a few of the “meta-methods” that developers of bioinformatics methods have found useful. For the purpose of designing algorithms, it is as important for bioinformaticians to be aware of the concept of fixed parameter tractability as it is for them to understand NP-completeness, since these concepts often determine the types of algorithms appropriate to a particular problem. Clustering is a ubiquitous problem in Bioinformatics, as is the need to visualize data. The need to interact with massive data bases and multiple software entities makes the development of computational pipelines an important issue for many bioinformaticians. Finally, the chapter on text mining discusses techniques for addressing the special problems of interacting with and extracting information from the vast biological literature. Jonathan M. Keith vi Preface
- 13. Contents Preface . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . v Contributors. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . ix Content of Volume I . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . xiii SECTION I: STRUCTURES 1. UNAFold: Software for Nucleic Acid Folding and Hybridization . . . . . . . . . . . . . . 3 Nicholas R. Markham and Michael Zuker 2. Protein Structure Prediction . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 33 Bissan Al-Lazikani, Emma E. Hill, and Veronica Morea 3. An Introduction to Protein Contact Prediction . . . . . . . . . . . . . . . . . . . . . . . . . . . 87 Nicholas Hamilton and Thomas Huber 4. Analysis of Mass Spectrometry Data in Proteomics. . . . . . . . . . . . . . . . . . . . . . . . . 105 Rune Matthiesen and Ole N. Jensen 5. The Classification of Protein Domains . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 123 Russell L. Marsden and Christine A. Orengo SECTION II: INFERRING FUNCTION 6. Inferring Function from Homology. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 149 Richard D. Emes 7. The Rosetta Stone Method . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 169 Shailesh V. Date 8. Inferring Functional Relationships from Conservation of Gene Order . . . . . . . . . . 181 Gabriel Moreno-Hagelsieb 9. Phylogenetic Profiling . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 201 Shailesh V. Date and José M. Peregrín-Alvarez 10. Phylogenetic Shadowing: Sequence Comparisons of Multiple Primate Species . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 217 Dario Boffelli 11. Prediction of Regulatory Elements . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 233 Albin Sandelin 12. Expression and Microarrays . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 245 Joaquín Dopazo and Fátima Al-Shahrour 13. Identifying Components of Complexes . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 257 Nicolas Goffard and Georg Weiller 14. Integrating Functional Genomics Data . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 267 Insuk Lee and Edward M. Marcotte SECTION III: APPLICATIONS AND DISEASE 15. Computational Diagnostics with Gene Expression Profiles. . . . . . . . . . . . . . . . . . . 281 Claudio Lottaz, Dennis Kostka, Florian Markowetz, and Rainer Spang vii
- 14. 16. Analysis of Quantitative Trait Loci. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 297 Mario Falchi 17. Molecular Similarity Concepts and Search Calculations . . . . . . . . . . . . . . . . . . . . . 327 Jens Auer and Jürgen Bajorath 18. Optimization of the MAD Algorithm for Virtual Screening . . . . . . . . . . . . . . . . . . 349 Hanna Eckert and Jürgen Bajorath 19. Combinatorial Optimization Models for Finding Genetic Signatures from Gene Expression Datasets . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 363 Regina Berretta, Wagner Costa, and Pablo Moscato 20. Genetic Signatures for a Rodent Model of Parkinson’s Disease Using Combinatorial Optimization Methods. . . . . . . . . . . . . . . . . . . . . . . . . . . . . 379 Mou’ath Hourani, Regina Berretta, Alexandre Mendes, and Pablo Moscato SECTION IV: ANALYTICAL AND COMPUTATIONAL METHODS 21. Developing Fixed-Parameter Algorithms to Solve Combinatorially Explosive Biological Problems . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 395 Falk Hüffner, Rolf Niedermeier, and Sebastian Wernicke 22. Clustering . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 423 Geoffrey J. McLachlan, Richard W. Bean, and Shu-Kay Ng 23. Visualization . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 441 Falk Schreiber 24. Constructing Computational Pipelines. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 451 Mark Halling-Brown and Adrian J. Shepherd 25. Text Mining . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 471 Andrew B. Clegg and Adrian J. Shepherd Index. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 493 viii Contents
- 15. Contributors BISSAN AL-LAZIKANI • Biofocus DPI, London, United Kingdom FÁTIMA AL-SHAHROUR • Department of Bioinformatics, Centro de Investigación Príncipe Felipe (CIPF), Valencia, Spain JENS AUER • Department of Life Science Informatics, Bonn-Aachen International Center for Information Technology (B-IT), Rheinische Friedrich-Wilhelms-University Bonn, Bonn, Germany JÜRGEN BAJORATH • Professor and Chair of Life Science Informatics, Department of Life Science Informatics, Bonn-Aachen International Center for Information Technology (B-IT), Rheinische Friedrich-Wilhelms-University Bonn, Bonn, Germany RICHARD W. BEAN • ARC Centre of Excellence in Bioinformatics, and Institute for Molecular Bioscience, The University of Queensland, Brisbane, Queensland, Australia REGINA BERRETTA • Centre of Bioinformatics, Biomarker Discovery and Information- Based Medicine, The University of Newcastle, Callaghan, New South Wales, Australia DARIO BOFFELLI • Children’s Hospital Oakland Research Institute, Oakland, CA ANDREW B. CLEGG • Institute of Structural Molecular Biology, School of Crystallography, Birkbeck College, University of London, London, United Kingdom WAGNER COSTA • School of Electrical Engineering and Computer Science, The University of Newcastle, Callaghan, New South Wales, Australia SHAILESH V. DATE • PENN Center for Bioinformatics, Department of Genetics, University of Pennsylvania School of Medicine, Philadelphia, PA JOAQUÍN DOPAZO • Department of Bioinformatics, Centro de Investigación Príncipe Felipe (CIPF), Valencia, Spain HANNA ECKERT • Department of Life Science Informatics, Bonn-Aachen International Center for Information Technology (B-IT), Rheinische Friedrich-Wilhelms-University Bonn, Bonn, Germany RICHARD D. EMES • Department of Biology, University College London, London, United Kingdom MARIO FALCHI • Twin Research and Genetic Epidemiology Unit, King’s College London School of Medicine, London, United Kingdom NICOLAS GOFFARD • Research School of Biological Sciences and ARC Centre of Excellence for Integrative Legume Research, The Australian National University, Canberra, Australian Capital Territory, Australia MARK HALLING-BROWN • Institute of Structural Molecular Biology, School of Crystallography, Birkbeck College, University of London, London, United Kingdom NICHOLAS HAMILTON • ARC Centre of Excellence in Bioinformatics, Institute for Molecular Bioscience and Advanced Computational Modelling Centre, The University of Queensland, Brisbane, Queensland, Australia EMMA E. HILL • The Journal of Cell Biology, Rockefeller University Press, New York, NY ix
- 16. MOU’ATH HOURANI • Newcastle Bioinformatics Initiative, School of Electrical Engineer- ing and Computer Science, The University of Newcastle, Callaghan, New South Wales, Australia THOMAS HUBER • School of Molecular and Microbial Sciences and Australian Institute for Bioengineering and Nanotechnology, The University of Queensland, Brisbane, Queensland, Australia FALK HÜFFNER • Institut für Informatik, Friedrich-Schiller-Universität Jena, Jena, Germany OLE N. JENSEN • Department of Biochemistry and Molecular Biology, University of Southern Denmark, Odense, Denmark JONATHAN M. KEITH • School of Mathematical Sciences, Queensland University of Technology, Brisbane, Queensland, Australia DENNIS KOSTKA • Max Planck Institute for Molecular Genetics and Berlin Center for Genome-Based Bioinformatics, Berlin, Germany INSUK LEE • Center for Systems and Synthetic Biology, Institute for Molecular Biology, University of Texas at Austin, Austin, TX CLAUDIO LOTTAZ • Max Planck Institute for Molecular Genetics and Berlin Center for Genome-Based Bioinformatics, Berlin, Germany EDWARD M. MARCOTTE • Center for Systems and Synthetic Biology, and Department of Chemistry and Biochemistry, Institute for Molecular Biology, University of Texas at Austin, Austin, TX NICHOLAS R. MARKHAM • Xerox Litigation Services, Albany, NY FLORIAN MARKOWETZ • Max Planck Institute for Molecular Genetics and Berlin Center for Genome-Based Bioinformatics, Berlin, Germany RUSSELL L. MARSDEN • Biochemistry and Molecular Biology Department, University College London, London, United Kingdom RUNE MATTHIESEN • CIC bioGUNE, Bilbao, Spain GEOFFREY J. MCLACHLAN • ARC Centre of Excellence in Bioinformatics, Institute for Molecular Bioscience, and Department of Mathematics, The University of Queensland, Brisbane, Queensland, Australia ALEXANDRE MENDES • Centre of Bioinformatics, Biomarker Discovery and Information- Based Medicine, The University of Newcastle, Callaghan, New South Wales, Australia VERONICA MOREA • National Research Council (CNR), Institute of Molecular Biology and Pathology (IBPN), Rome, Italy GABRIEL MORENO-HAGELSIEB • Department of Biology, Wilfrid Laurier University, Waterloo, Ontario, Canada PABLO MOSCATO • ARC Centre of Excellence in Bioinformatics, and Centre of Bioin- formatics, Biomarker Discovery and Information-Based Medicine, The University of Newcastle, Callaghan, New South Wales, Australia SHU-KAY NG • Department of Mathematics, The University of Queensland, Brisbane, Queensland, Australia ROLF NIEDERMEIER • Institut für Informatik, Friedrich-Schiller-Universität Jena, Jena, Germany CHRISTINE A. ORENGO • Biochemistry and Molecular Biology Department, University College London, London, United Kingdom JOSÉ M. PEREGRÍN-ALVAREZ • Hospital for Sick Children, Toronto, Ontario, Canada x Contributors
- 17. ALBIN SANDELIN • The Bioinformatics Centre, Department of Molecular Biology and Biotech Research and Innovation Centre, University of Copenhagen, Copenhagen, Denmark FALK SCHREIBER • Leibniz Institute of Plant Genetics and Crop Plant Research (IPK) Gatersleben, Germany and Institute for Computer Science, Martin-Luther University Halle-Wittenberg, Germany ADRIAN J. SHEPHERD • Institute of Structural Molecular Biology, School of Crystallography, Birkbeck College, University of London, London, United Kingdom RAINER SPANG • Max Planck Institute for Molecular Genetics and Berlin Center for Genome-Based Bioinformatics, Berlin, Germany GEORG F. WEILLER • Research School of Biological Sciences and ARC Centre of Excellence for Integrative Legume Research, The Australian National University, Canberra, Australian Capital Territory, Australia SEBASTIAN WERNICKE • Institut für Informatik, Friedrich-Schiller-Universität Jena, Jena, Germany MICHAEL ZUKER • Mathematical Sciences and Biology Department, Rensselaer Polytechnic Institute, Troy, NY Contributors xi
- 18. Contents of Volume I SECTION I: DATA AND DATABASES 1. Managing Sequence Data Ilene Karsch Mizrachi 2. RNA Structure Determination by NMR Lincoln G. Scott and Mirko Hennig 3. Protein Structure Determination by X-Ray Crystallography Andrea Ilari and Carmelinda Savino 4. Pre-Processing of Microarray Data and Analysis of Differential Expression Steffen Durinck 5. Developing an Ontology Midori A. Harris 6. Genome Annotation Hideya Kawaji and Yoshihide Hayashizaki SECTION II: SEQUENCE ANALYSIS 7. Multiple Sequence Alignment Walter Pirovano and Jaap Heringa 8. Finding Genes in Genome Sequence Alice Carolyn McHardy 9. Bioinformatics Detection of Alternative Splicing Namshin Kim and Christopher Lee 10. Reconstruction of Full-Length Isoforms from Splice Graphs Yi Xing and Christopher Lee 11. Sequence Segmentation Jonathan M. Keith 12. Discovering Sequence Motifs Timothy L. Bailey SECTION III: PHYLOGENETICS AND EVOLUTION 13. Modeling Sequence Evolution Pietro Liò and Martin Bishop 14. Inferring Trees Simon Whelan 15. Detecting the Presence and Location of Selection in Proteins Tim Massingham 16. Phylogenetic Model Evaluation Lars Sommer Jermiin, Vivek Jayaswal, Faisal Ababneh, and John Robinson xiii
- 19. 17. Inferring Ancestral Gene Order Julian M. Catchen, John S. Conery, and John H. Postlethwait 18. Genome Rearrangement by the Double Cut and Join Operation Richard Friedberg, Aaron E. Darling, and Sophia Yancopoulos 19. Inferring Ancestral Protein Interaction Networks José M. Peregrín-Alvarez 20. Computational Tools for the Analysis of Rearrangements in Mammalian Genomes Guillaume Bourque and Glenn Tesler 21. Detecting Lateral Genetic Transfer: A Phylogenetic Approach Robert G. Beiko and Mark A. Ragan 22. Detecting Genetic Recombination Georg F. Weiller 23. Inferring Patterns of Migration Paul M.E. Bunje and Thierry Wirth 24. Fixed-Parameter Algorithms in Phylogenetics Jens Gramm, Arfst Nickelsen, and Till Tantau Index xiv Contents of Volume I
- 20. Chapter 1 UNAFold Software for Nucleic Acid Folding and Hybridization Nicholas R. Markham and Michael Zuker Abstract The UNAFold software package is an integrated collection of programs that simulate folding, hybridiza- tion, and melting pathways for one or two single-stranded nucleic acid sequences. The name is derived from “Unified Nucleic Acid Folding.” Folding (secondary structure) prediction for single-stranded RNA or DNA combines free energy minimization, partition function calculations and stochastic sampling. For melting simulations, the package computes entire melting profiles, not just melting temperatures. UV absorbance at 260nm, heat capacity change (Cp ), and mole fractions of different molecular species are computed as a function of temperature. The package installs and runs on all Unix and Linux platforms that we have looked at, including Mac OS X. Images of secondary structures, hybridizations, and dot plots may be computed using common formats. Similarly, a variety of melting profile plots is created when appropriate. These latter plots include experimental results if they are provided. The package is “command line” driven. Underlying compiled programs may be used individually, or in special combina- tions through the use of a variety of Perl scripts. Users are encouraged to create their own scripts to sup- plement what comes with the package. This evolving software is available for download at http:/ /www. bioinfo.rpi.edu/applications/hybrid/download.php. Key words: RNA folding, nucleic acid hybridization, nucleic acid melting profiles (or DNA melting profiles), free energy minimization, partition functions, melting temperature. The UNAFold software predicts nucleic acid foldings, hybridi- zations, and melting profiles using energy-based methods and a general computational technique known as dynamic program- ming (1). Early software for RNA folding predicted minimum 1. Introduction 1. Introduction Jonathan M. Keith (ed.), Bioinformatics, Volume II: Structure, Function and Applications, vol. 453 © 2008 Humana Press, a part of Springer Science+Business Media, Totowa, NJ Book doi: 10.1007/978-1-60327-429-6 Springerprotocols.com 3
- 21. 4 Markham and Zuker free energy foldings only (2–6). It became clear early on that such methods were unreliable in the sense that many different fold- ings, with free energies close to the computed minimum, could exist. Although constraints deduced from experiments or phylo- genetic analyses could be applied to reduce uncertainty, a method to compute a variety of close to optimal foldings was needed. The mfold software (7–10) computes a collection of optimal and suboptimal foldings as well as a triangular-shaped plot called an energy dot plot (EDP). The EDP contains a dot or other symbol in row i and column j (i < j) to indicate that the base pair between the ith and jth nucleotides can occur in a folding within some user prescribed free energy increment from the minimum. The Vienna RNA Package (Vienna RNA) (11–13) differs funda- mentally from mfold because the underlying algorithm computes partition functions, rather than minimum free energies. This leads naturally to the computation of base pair probabilities (14) and what they call “boxplots.” We call these triangular shaped plots probability dot plots (PDPs). In this case, all base pairs with probabilities above a certain threshold are plotted as boxes (or other symbols) whose area is proportional to the probability of that base pair. This software can compute all possible foldings close to optimal. The sfold package (15–17) also computes parti- tion functions, but it uses a simpler algorithm than the Vienna RNA package because base pair probabilities are not computed directly (exactly). Instead, it computes a “statistically valid sam- ple” (Gibbs sample) that permits the estimation of not only base pair probabilities, but probabilities of any desired motif(s). UNA- Fold encompasses all three methods by computing minimum and suboptimal foldings in the mfold style, and full partition func- tions that allow both exact base pair computations as well as sto- chastic sampling. In addition, stochastic sampling may be used to compute both ensemble enthalpy and ensemble heat capacity for single sequence folding. For practical applications, it is common to use crude methods, often ad hoc, to compute melting temperatures for dimers. It is usual to assume that hybridized strands are perfectly complementary. When mismatches are permitted, they are few and isolated. There is no question about base pairs and no computations are required to determine the hybridization. For example, the simple case: 5′-ACGGtCCAGCAA-3′ 3′-TGCCgGGTCGTT-5′ shows the dimerization of two almost complementary 12-mers. Note the single T⋅G wobble pair. A computer is not needed to compute the hybridization. A calculator would be helpful in add- ing up nearest neighbor free energies and enthalpies from pub- lished tables (18–20), and a computer would be needed to process thousands of these dimers. In any case, free energy at 37°C, ∆G
- 22. UNAFold 5 (or DG37 ) and enthalpy, ∆H, are easily computed. From this, the entropy, ∆S, is easily computed as: ∆ × ∆ − ∆ S = H G T 1000 , where T is the hybridization temperature (K) and the factor of 1000 expresses ∆S in e.u. (entropy units; 1 e.u. = 1cal/mol/K). The terms “free energy,” “enthalpy,” and “entropy” are really changes in free energy, enthalpy, and entropy with respect to the “random coil state.” In reality, “random coil” is a large ensemble of states. The free energy and enthalpy of this ensemble of states may be (and is) set to zero. Only entropy cannot be arbitrarily assigned. Thus a ∆S term is Rln(N/Nref ), where R is the universal gas constant, 1.9872 e.u.; N is the number of states consistent with hybridization; and Nref is the number of states, always larger, in the unconstrained “random coil.” The simplest model for hybridization assumes that there are two states: hybridized, where the structure is unique and known, and “random coil.” It is then a simple matter to deduce a melt- ing temperature, Tm , at which half of the dimers have dissociated. The formula is given by: T f m = H S + R 1000 ln(Ct × ∆ ∆ ) , [1] where ∆H, ∆S (expressed in e.u.) and R have already been defined, Ct is the total strand concentration, and f = 4 when the two strands are different and f = 1 when self-hybridization takes place. This formula implicitly assumes that both strands are present in equal concentra- tions. Competition with folding and with other possible hybridiza- tions, such as homodimer formation, is not considered at all. UNAFold offers a number of increasingly sophisticated ways to compute melting temperatures and also computes entire melt- ing profiles: UV absorbance at 260nm, heat capacity (Cp ), and mole fractions of various single and double-stranded molecu- lar species as a function of temperature. Even when Equation [1] is used, the hybridization is computed by minimizing free energy. The various levels of complexity and choice of methods are described in the following. It is important to issue a warning to those who wish to apply these methods to microarrays. Hybridization on microarrays is complicated by the fact that one of each hybridizing pair is immo- bilized. It is difficult to compute the effective “solution concentra- tion” for such molecules, and diffusion is probably an important factor in slowing the time necessary to reach equilibrium. How- ever, even for hybridization in solution, kinetic simulations that treat different probe-target pairs independently can lead to totally incorrect results for the time required to reach equilibrium as well as the equilibrium concentrations themselves (21).
- 23. 6 Markham and Zuker It is important to emphasize that the computations used by the UNAFold software are based on a number of assumptions. 1. The simulations are for molecules in solution. For microar- rays, computing an effective concentration of the immobilized oligos is necessary. One needs to estimate the “surface phase concentration,” c, in units such as “molecules/cm2 ,” and the solution volume per cm2 of surface area, n (L/cm2 ) (21). 2. The system contains one or two different molecules. If, for example, a true system contains three different oligos, A, B and C, then some common sense must be used. If A and B do not hybridize with each other over the temperature range of interest, then A and C could be simulated separately from B and C if, in addition, the concentration of C is a couple of orders of magnitude greater than that for both A and B. 3. The computations assume that the system is in equilibrium. Thus, the melting profile predictions assume that tempera- ture changes are slow enough so that the system is always at equilibrium. When this is not true, observed melting profiles depend on the rate of temperature change. In particular, a hysteresis effect can be observed. For example, even if the temperature is raised at a uniform rate from T0 to T1 , and then brought down to T0 at the same rate, the measured UV absorbance profiles for melting and cooling may differ. The UNAFold package compiles and runs on many platforms. Table 1.1 lists a few operating system/architecture combinations under which the package is known to work. In fact, the UNAFold package is not known not to run on any platform. The software should function on any system having the following basic tools: ● A Unix-style shell. ● Make. ● A C compiler. ● A Perl interpreter. As noted, UNAFold requires only some very basic tools to build and run. However, there are several optional libraries and pro- grams that provide additional functionality when installed: ● gnuplot: If gnuplot is detected, the hybrid2.pl script will use it to produce several plots in Postscript format. ● OpenGL/glut: If OpenGL and the glut library (22) are detected, the hybrid-plot program will be built. 2. Materials: The UNAFold Software 2. Materials: The UNAFold Software 2.1. Supported Platforms 2.1. Supported Platforms 2.2. Dependencies 2.2. Dependencies
- 24. UNAFold 7 ● gd: If the gd library is detected, the hybrid-plot-ng program will use it to create images in GIF, JPEG, and/or PNG for- mats directly. (With or without this library, hybrid-plot-ng creates Postscript files which can be converted to virtually any other image format.) The UNAFold software is available for download at http:/ /www. bioinfo.rpi.edu/applications/hybrid/download.php. Binaries for Linux (in RPM format) and Windows (in EXE format) are available along with the source code. After downloading and unpacking, building the software consists of three steps: ● Configuration: Usually this is as simple as typing ./config- ure. Running ./configure --help lists options that may be used to configure installation locations, specify locations of libraries, and set compiler options. ● Compilation: A single command—make—compiles and links all programs. ● Installation: Typing make install copies the programs, scripts, documentation, and data files to the location set with the configure script. The core programs in UNAFold are written in C and are opti- mized when compiled since they are computationally intensive. Man pages exist for all of these programs. In addition, invoking any program with the --help option will generate an abbreviated set of instructions. Some programs have counterparts formed by 2.3. Downloading and Installing 2.3. Downloading and Installing 2.4. Core Programs 2.4. Core Programs Table 1.1 Some platforms supported by the UNAFold package Operating system Architecture Linux x86 Linux x86_64 FreeBSD x86 SunOS sparc IRIX mips IRIX64 mips AIX powerpc MacOS powerpc MS Windows x86
- 25. 8 Markham and Zuker adding the suffix -same. When simulating a one-sequence ensem- ble (in which the dominant dimer is a homodimer rather than a heterodimer) the -same version replaces the regular one. ● Folding: — hybrid-ss: Computes full partition functions for folding RNA or DNA. It may be run using the --energyOnly option, in which case the probabilities of base pairs and of single-stranded nucleotides and dinucleotides will not be computed, saving significant time and memory. It may also be run with the --tracebacks option to gen- erate a stochastic sample of foldings. UNAFold also includes two simplified versions of hybrid-ss: hybrid- ss-simple and hybrid-ss-noml. hybrid-ss-simple assigns fixed entropic costs to multibranch loops and ignores single-base stacking, while hybrid-ss-noml does not allow multibranch loops at all. — hybrid-ss-min: Computes minimum energy foldings. It can predict a single, minimum free energy folding or, using an extended algorithm, generate an mfold-like collection of foldings and an EDP. For fast folding of many sequences with the same parameters, the --stream option reads sequences, one at a time, from standard input and writes free energies to standard output. ● Hybridization: — hybrid: Computes full partition functions for hybridiz- ing RNA or DNA (without intramolecular base pairs). The --energyOnly and --tracebacks options function as in hybrid-ss. — hybrid-min: Computes minimum energy hybridiza- tions. The --mfold and --stream options function as in hybrid-ss-min. ● Ensemble computations: These programs rely on the output of the core programs. — concentration(-same): Computes the mole fractions of different molecular species using the already com- puted free energies for individual monomer (folded) and dimer (hybridized) species. — ensemble-dg(-same): Computes ensemble free ener- gies using species free energies and computed mole fractions. — ensemble-ext(-same): Computes UV absorbance at 260nm using computed probabilities for single-strand- edness of nucleotides and (optionally) dinucleotides, computed mole fractions, and published extinction coefficients (23).
- 26. Discovering Diverse Content Through Random Scribd Documents
- 27. 3. The quantity of water in the air was 1/20th less than the average, at the same time that the mean weight of a cubic foot of air was 2 grains above the average. 4. An unusual alternation of heat and cold, yet the heat predominating to such an extent that in particular localities it rose as much as from 2° to 8° above the average. These excesses were most striking at night, particularly in the parts of London on a level with the Thames, where the night temperatures ranged from 7°, 8°, 9°, and 10° above the temperature of the country, and even of the suburban districts. These temperatures were highest, especially the night ones, when the mortality was greatest; and the mortality was greatest where the temperatures were highest. 5. A remarkable increase above the average in the temperature of the water of the Thames. From a long series of observations it had been found that the normal temperature of the Thames is 51.7°. During the prevalence of the epidemic it rose to 60°, 66°, and once to 70°. At this temperature the “simmering” water must have poured enormous quantities of vapour into the surrounding atmosphere; not the pure vapour of water, for that cannot arise from a river which is the recipient of the foul contents of all the sewers and cesspools of the metropolis. In some instances there was an excess of 20° of the temperature of the water above that of the air. For twenty-eight continuous nights during the height of the epidemic, the average excess exceeded 16.5°. 6. An unusual prevalence of haze, mist, and fog; the fog being sometimes so dense that London could not be discerned from Greenwich. 7. An extraordinary stillness and stagnation of the air, both by day and night. Sometimes in the low-lying districts not a breath could be observed. Even when at more elevated stations the wind was moving with a force of 1 lb 7 oz., the pressure was only ¼ lb in the heart of London.
- 28. Wind is the ventilator of nature. Artificial ventilation, as far as it is successful, is an imitation of nature’s process. It is stated on undoubted authority (Maitland’s History of London) that for several weeks before the Great Plague broke out in London, there was an uninterrupted calm, so that there was not sufficient motion of the air to stir a vane. Baynard, a contemporary physician, confirms this fact. The like circumstance is mentioned by Diemerbroeck in giving an account of the plague at Nimeguen. At the period when the last plague visited Vienna, according to Sir Gilbert Blane, there had been no wind for three months. The terrific outbreak of the cholera at Kurrachee was preceded for some days by such a stagnation of the atmosphere that an oppression scarcely to be endured affected the whole population. It is obvious that calms must favour the accumulation and concentration of effluvia from every source from which they arise. 8. A general deficiency in the tension of common positive electricity. 9. A deficiency of one fourth of the rain-fall for the year. During 118 consecutive days there was scarcely any rain, and not a single drop for 18 days at the period of the highest mortality. 10. A total absence of ozone at all the stations near the river, while at stations of high elevation it was of general occurrence. These observations relate particularly to the epidemic of 1854, which was more carefully watched than the two former; but the results are similar for each. “The three epidemics,” says Mr Glashier, in summing up the results of his inquiry, “were attended with a particular state of atmosphere, characterized by a prevalent mist, thin in high places, dense in low. During the height of the epidemic, in all cases, the reading of the barometer was remarkably high, the atmosphere thick; and in 1849 and 1854 the temperature above its average. A total absence of rain, and a stillness of air amounting almost to calm, accompanied the progress of the disease on each occasion. In places near the river,
- 29. the night temperatures were high, with small diurnal range, with a dense torpid mist and air charged with the many impurities arising from the exhalations of the Thames, and adjoining marshes; a deficiency of electricity, and, as shown in 1854, a total absence of ozone, most probably destroyed by the decomposition of the organic matter with which the air in these situations is so strongly charged. “In both 1849 and 1854, the first decline of the disease was marked by a decrease in the readings of the barometer, and in the temperature of the air and water; the air, which previously had for a long time continued calm, was succeeded by a strong S. W. wind, which soon dissipated the former stagnant and poisonous atmosphere.” We knew before that such influences were in operation, but they had not been weighed and measured. We now know definitely something of an epidemic atmosphere, and the information obtained is most significant; for it shows that the several meteorological changes that take place during the prevalence of an epidemic concur to produce a heavy, warm, moist, and stagnant atmosphere, with disturbed electricity: conditions highly favourable to the decomposition of organic matter. Under the influence of such an atmosphere, over the moist and warmed surface of every filthy place, over the entire mass of all accumulations of filth in streets, lanes, and courts, and within and about houses, and over the heated surface of all foul water, decomposition goes on with the utmost activity, and the products are poured into the stagnant air. Against such products the human body has no defence. The lungs admit whatever is brought to them—poisonous and salubrious substances alike. They are guarded by none of those protective contrivances which we see in some other parts of the body. Whatever is capable of suspension in the respired air passes with it directly into the current of the circulation, and when once there, is carried with astonishing rapidity into the very substance of the vital organs.
- 30. From the quantity of air which the lungs receive, some conception may be formed of the amount of obnoxious matter which may be introduced into the system through these portals. At each inspiration there enter the lungs of an ordinary-sized person about 20 cubic inches of air. There are 20 respirations in a minute: 400 cubic inches of air must therefore enter in one minute; 14 cubic feet in one hour, and 366 cubic feet, or 36 hogsheads, in one day. To meet this the heart sends into the lungs at each contraction two ounces of blood; there are 75 pulsations in a minute, during which 150 ounces are propelled into the lungs; a quantity which gives 562 pounds in one hour and 24 hogsheads in 24 hours. The main purpose for bringing these enormous quantities of air and blood together, with such velocity, is to provide for the enormous waste which is caused by the rapid and unceasing mutation of organic matter. The activity of an organ is sustained at the expense of the matter of which it is composed. No thought passes through the mind, but an equivalent portion of the substance of the brain is consumed; no nervous current flows along the nervous conducters, but a corresponding portion of nervous tissue is used up; no muscular movement, no glandular secretion, takes place without a proportionate waste of muscle and of gland. What must be the amount of supply required to meet this waste, when able-bodied men employed in their ordinary labour lose from 2 lbs. to 5 lbs. and upwards of their weight twice a day.[8] Some physiologists of eminence have estimated that in order to supply that waste, there passes in the course of every 24 hours as much fluid through the thoracic duct[9] as equals the whole quantity of blood in the body. 8. See Experiments on the daily loss of weight sustained by workmen employed in gas-works.—Philosophy of Health, 11th Edit. p. 284, et seq. 9. The tube which conveys the debris of the body, together with the nutritious part of the food,—both measures of change or waste. The results of the highly interesting experiments recently made by Professor Graham on the part taken by the active agent in all these
- 31. processes—organic membrane, of which the organic cell is the type, demonstrates that all the phenomena known as Endosmose and Exosmose depend on a chemical action involving the destruction of organic membrane. In this process chemical action is set up dependent upon active chemical agents, neutral substances being inoperative. Out of this chemical action a new force is induced, the Osmotic force; a purely chemical being converted into an equivalent mechanical force, which is made subservient to the essential phenomena of organic and animal life: a vis motrix, a force which is to the extra-vascular movements of the body, what the contraction of the heart is to the vascular. In a frame so constructed, any particles contaminating the circulating fluid most rapidly pervade and contaminate every part of the system. It has been sometimes imagined that the quantity of matter suspended in the atmosphere and conveyed into the system in respired air, must be too minute to exert any serious influence upon the body. One single puncture of the finger, so small as not to be visible without the aid of a lens, has introduced into the system a sufficient quantity of putrid matter to cause death with the most violent symptoms. A few drops of the liquid matter obtained by a condensation of the air of a foul locality, introduced into the vein of a dog, is stated to have produced death with the usual phenomena of typhus fever. It is certain that on the introduction into the body of an inappreciable portion of the matter of cow-pox, or of small-pox, those specific forms of fever are produced. From these and similar facts it is inferred, that when putrescent or decomposing organic matter is introduced into the blood it acts as a poison and produces the phenomena of fever, and that all the predisposing causes of epidemics act in this way—by overcharging the blood with the products of decomposing organic matter.
- 32. Strictly speaking, however, all that we really know is this—that where certain conditions exist, epidemics break out and spread; that where those conditions do not exist, epidemics do not break out and spread; and that where those conditions did exist, but have been removed, thereupon epidemics cease. We call those conditions Causes, Predisposing or Localizing Causes, but how they act, whether by accumulating decomposing organic matter in the blood, or in what other way, we have no certain knowledge. One further fact however is ascertained, that where any one of these predisposing causes is present, epidemics break out and spread just as readily as when all are present together. Where there is overcrowding alone, for example, epidemics break out and spread. Where there is decomposing filth alone, epidemics break out and spread; and so of the whole number. The removal of one of these causes, therefore, or the removal of two or three of them, will not suffice for safety; every one must be removed before there can be safety. This we know; all beyond this is conjecture, but as to the most probable of these conjectures, some who have thought on this subject believe that the preponderance of evidence justifies the conclusion that the predisposing causes may themselves become efficient causes; that instances in which they actually do so, are constantly passing before our eyes; that it is practicable to manufacture fever and even epidemic fever to any amount by placing a population under certain known conditions; that it is practicable to prevent the outbreak of epidemics altogether by placing the population under certain other conditions;[10] that the prevalence of the predisposing causes in particular localities, in certain intensities, is sufficient to produce local epidemic outbreaks; that the prevalence of such causes in such intensities, joined to some general conditions of the atmosphere, such as the meteorological conditions which have been enumerated, particularly those which favour the accumulation and concentration of the
- 33. products of organic decomposition, are all that is required to engender wide-spread epidemics. Those who adopt this view contend that the existence of a primary cause as a distinct and separate entity is not necessary to account for the phenomena. The more common opinion however is, that joined to the predisposing causes there must always be present a primary cause, having a distinct existence, capable of travelling from one part of the globe to another; capable of spreading over any space however extended, or of confining itself to any space however small—a district, a street, a house, a room. 10. See Baltimore case, p. 78. It is urged that though we are unacquainted with the physical form or chemical properties of this body, this is no reason why we should not understand its force as a special agent in the production of disease, just as we know the forces of other physical bodies, though not their nature. The existence of such a body being assumed, it is conceived that it exists not in a gaseous but in a liquid state. It is supposed that it cannot exist in a gaseous state because a gas is readily diffused and dissipated; because when organic matter is reduced to a gaseous state, it has passed from the organic into the inorganic kingdom, and there is no evidence that the elementary bodies belonging to this kingdom are capable of producing any form of fever; and because there is indubitable evidence that organic matter in a recent state of putrescence—the more recent the more potent—is capable of producing the most deadly forms of fever. From these considerations it is conjectured that the primary cause, whatever it be, is some subtle fluid which has not wholly lost its organic composition, and that it consists of particles of extreme minuteness, capable of attaching itself to the surfaces of other bodies, and even of increasing under favourable circumstances. It is further thought that this body is not equally diffused through the atmosphere, but is only partially distributed, and that this
- 34. accounts for the local distribution of epidemics, and for their occasional absence from places which apparently present all the conditions favourable to their development. Lastly, the opinion is gaining ground, that this body acts in the manner of a ferment. It is urged in favour of this view, that a ferment being an azotized substance in a state of putrefactive alteration, the body in question must find, in the decomposing organic compounds with which impure blood is charged, precisely the materials for taking on the fermenting process. The advocates for this view think that the term “zymotic” is not only the appropriate name of the whole of this class of diseases, but that it also declares an interesting fact connected with them. Whatever may be the truth with respect to these points, on which at present we have no positive knowledge, one thing is certain, that practically our concern is with the known causes,—the ascertained conditions. These are palpable, definite, and capable of complete removal and prevention. Overcrowding, for example, we can prevent; the accumulation of filth in towns and houses we can prevent; the supply of light, air, and water, together with the several other appliances included in the all-comprehensive word Cleanliness, we can secure. To the extent to which it is in our power to do this, it is in our power to prevent epidemics. The human family have now lived together in communities more than six thousand years, yet they have not learnt to make their habitations clean. At last we are beginning to learn the lesson. When we shall have mastered it, we shall have conquered epidemics. Our duties, then, and our hopes in this respect, I shall proceed to show. The principal constituents of the atmosphere maintain their equilibrium steadily over the whole surface of the globe. There is scarcely any difference in the relative proportion of its oxygen and nitrogen in the torrid zone and in the arctic regions. Whatever influence the atmosphere may have on climate must consequently depend on something adventitious to it and not in anything forming
- 35. a part of it. Possibly therefore that something may be, in some degree, under human control. The main constituents of climate are temperature and moisture, and these are the climatic conditions that exercise the greatest influence on epidemics. Minor but still important conditions are the nature of the soil, the proportion of land that is cleared and under cultivation, the extent of forests, lakes, and rivers, the prevailing winds, the electrical state of the atmosphere, and so on. The temperature is highest where the sun’s rays are vertical, or nearly so; where the sky is cloudless; where the day is longest; and where there is the smallest difference between the fervid noon-tide heat and the temperature of the short night. The moisture is greatest where in addition to all the other sources of humidity there are periodical rains. In the countries subject to these rains, the entire extent of the level and low land is often covered a foot deeper with water than before the rain set in. Elevated temperature and excessive moisture are combined in tropical countries; and they are concentrated in those parts of the tropics in which there are extensive forests having an undergrowth of luxuriant vegetation; in which the tides of the ocean penetrate deeply into the interior of the land, and mix with the waters of the rivers; and in which the rivers constantly overflow their banks and form marshes and swamps. In tropical countries there are tracts such as these that extend in unbroken continuity hundreds of leagues. The western coast of Africa (the Bight of Benin) presents an unbroken area of upwards of 100,000 square miles, consisting of one vast alluvial and densely- wooded forest, irrigated by Atlantic tides, and intersected by numerous rivers and creeks, whose muddy banks are constantly overflowed. In describing a tropical forest, Humboldt says, “Under the bushy, deep, green verdure of trees of stupendous height and size, there
- 36. reigns constantly a kind of half daylight, a sort of obscurity, of which our forests of pines, oaks, and beech trees afford no example; forming a carpet of verdure, the dark tint of which augments the splendour of the aërial light.” With this luxuriance of vegetation is combined a corresponding abundance of animal life. The earth and air teem with living creatures. “The mould,” observes the same distinguished traveller, “contains the spoils of innumerable quantities of reptiles, worms, and insects. Wherever the soil is turned up we are struck with a mass of organic substances, which by turns are developed, transformed, and decomposed. Nature in these climates appear more active, more fruitful, we might say more prodigal of life.” The air is still more alive than the land. Insects fill the lower strata of the atmosphere to the height of fifteen or twenty feet, like a condensed vapour. It is estimated that a cubic foot of air is often peopled by a million of winged insects, which contain a caustic and venomous liquid, several species being nearly two lines (1.8) long. When two persons who have their home in these regions meet in the morning, the first questions they address to each other are, “How did you find the zancudoes during the night?” “How are we to- day for the mosquitoes?” An ancient form of Chinese politeness, showing the ancient state of that country, was—“Have you been incommoded in the night by serpents?” It appears that there are still inhabited places in which the Chinese compliment on the serpents might be added to that of the mosquitoes. Proportionate to this prodigality of organic life is the amount of organic decomposition, the products of which are poured into the atmosphere and suspended in the surrounding vapour and fog,[11] to which they give a decided and often a highly offensive odour. 11. See note, p. 16.
- 37. “On fixing our eyes on the tops of the trees,” describes Humboldt, “we discovered streams of vapour wherever a solar ray penetrated and traversed the dense atmosphere, exhaling, together with the aromatic odour yielded by the flowers, the fruit, and even the wood, that peculiar odour which we perceive in autumn in foggy seasons. It might be said, that notwithstanding the elevated temperature the air cannot dissolve the quantity of water exhaled from the surface of the soil and of the vegetation.” “At the distance of several miles from the coast,” says Dr Daniell, in describing the western shores of Africa, “the peculiar odour arising from swampy exhalations and the decomposition of vegetable matter is very perceptible, and sometimes even offensive. The water also is frequently of a dusky hue, with leaves, branches, and other vegetable debris floating on the surface, brought down from the interior by innumerable narrow channels that empty their turbid streams into the open ocean.” It is under these climatic conditions that the worst forms of epidemics are engendered: the most sudden in their attack, the most rapid in their development, the most general in their prevalence, and the most mortal. The form of the epidemic prevalent in any particular district is dependent on the physical characters of the immediate neighbourhood. Thus intermittents prevail chiefly in marshy and swampy districts: remittents also chiefly there, though not exclusively; while in other localities other forms arise approximating to the continued type of temperate climates. For the most part these epidemics are strictly endemic, and are confined to the particular regions in which they are engendered. They never pass the limit of the equatorial or tropical zone. Yellow Fever, one of the most common and destructive of these diseases, is still more restricted in its range, being confined within a definite line determined by temperature. It is incapable of existing where the average range of the thermometer is greater than from 76° to 86° of Fahrenheit, or where the temperature varies more than from 5° to
- 38. 10° night and day. Extreme heat and moderate cold immediately stop it; nay, even the prevalence of a cold wind for a few hours only. In other instances these epidemics pass beyond the regions in which they are produced, and sometimes extend to all the other quarters of the globe. The Black Death, the range of which we have seen, was engendered in China; the Cholera of our own day, generated in the delta of the Ganges, the great source and centre of Indian epidemics, ravaged that country long before it directed its course to Europe. When these tropical epidemics advance into more temperate climes, they lay aside nothing of their nature; they lose but little of their power. Wherever they go they decimate the populations which they attack. One remarkable peculiarity of some of these epidemics is, that natives of the region in which they prevail are for the most part unsusceptible to them. This is true however only of particular forms of pestilence. Some of them acknowledge no acclimation. Cholera, for example, attacks equally natives and new comers. On the other hand, yellow fever rarely attacks the natives who reside permanently within its zone. Its chief victims are strangers who have recently arrived within its sphere, particularly the inhabitants of northern climates. The susceptibility to its influence appears to be strictly proportionate to the degree of northern latitude from which the stranger has arrived, and the shortness of the interval that has passed since he left the European for the Equatorial regions. We see something of the same kind in the wide-spread epidemics of our own country. During the prevalence of Cholera it was observed over and over again, that persons coming directly from the pure air of the country into the infected part of a town, were seized with the disease. The explanation is not obvious. It would seem, however, to be connected with the suddenness of the shock on the system. Priestley found, that after shutting up a mouse in a given quantity of air a considerable time, it seemed to be weak, and to be slowly dying. If at this period he put a fresh mouse into the same air,
- 39. it instantly died. It seems as if the system can bear a pestiferous atmosphere better when gradually than when suddenly exposed to it. I do not know that I can give a more vivid picture of a tropical epidemic than that which is afforded by the outbreak of Cholera in the 86th regiment at Kurrachee in June, 1846. On this occasion the atmosphere was very peculiar,—damp, hot, stagnant, and oppressive. Not a breath of air was stirring. A few isolated cases of cholera had occurred for some days. The utmost alarm was excited in the minds of experienced persons, who felt certain that an epidemic was at hand. Their fears were too fully realized. On the night of the 15th, upwards of 40 men were seized with cholera in its severest form; in two days more 256 were attacked, of whom 131 were already dead. “The floors of the hospital,” says Dr Thom, the surgeon of the regiment, “were literally strewed with the livid bodies of men labouring under the pangs of premature dissolution. Many were brought in with the cold and clammy damp of death; as if sudden obstruction of every vital function had taken place, and the fountains of life had been arrested by an invisible but instantaneous shock. It was indeed a sight never to be forgotten, to behold the powerful frames of the finest men of a fine corps, who had that morning been in apparent good health, and most of them on the evening parade, as if at once stricken down, and striving, with the last efforts of gigantic strength, to resist a death-call that would not be refused.” In describing a river on the west coast of Africa, Dr Daniell says —“When I visited it, I found two vessels moored a short distance from its mouth, one of which within the space of five months had buried two entire crews, a solitary person alone surviving. The other, which had arrived at a much later period, had been similarly deprived of one-half of its men, and the remainder were in such a debilitated condition as to be incapable of undertaking any active or laborious duty. Immediately before, another vessel had sailed from
- 40. this port in such a deplorable state as to be solely dependent on the aid of Kroomen to perform the voyage.” In the statistical report of Sir Alexander Tulloch it is stated, that out of 1658 white troops sent out to military stations on the western coast of Africa, 1271 perished from climatic diseases; while of the 387 who remained to be sent home, 17 died on their passage; 157 were reported as incapable of further service; and 180 as qualified only for garrison service; thus leaving only 33 out of 1658 men who were fit for active service. As we pass out of the torrid zone a remarkable change takes place in the general character of epidemics. They lose more and more of their intermittent type, and become either remittent or continued. The remittent keeps its hold over the southern part of Europe, and continually breaks out in the form of Yellow Fever. As we proceed northward out of the yellow fever zone, that disease wholly disappears, and typhus and its kindred maladies take its place; typhus commencing precisely at the point where yellow fever ends. There is, indeed, one of the ordinary diseases of temperate climes, and only one, which appears capable of penetrating within the torrid zone, and of committing greater ravages there than in lower temperatures, and that is Small-pox. With this exception, the ordinary epidemics of temperate climates do not enter the tropics, while, on the other hand, the ordinary epidemics of the tropics every now and then decimate the temperate regions. “In these our latitudes,” says Dr William Fergusson, “cold and fatigue, and sorrow and hunger, will generate fever anywhere; but every region, every climate, will exhibit its own form of fever. With us it is Typhus; in the warmer countries of Europe, Remittent; in the upper Mediterranean, Plague; in the Antilles and Western Africa, Yellow Fever; this last being restricted to particular localities, temperatures, and elevation. While typhus fever goes out when you enter the tropics, it is there that yellow fever commences; the pure epidemic of a hot climate that cannot be transported or communicated upon any other ground. Places, not persons,
- 41. constitute the rule of its existence. Places, not persons, comprehend the whole history, the etiology of the disease. Places, not persons! Let the emphatic words be dinned into the ears of the Lords of the Treasury, of Trade and Plantations, until they acquire the force of a creed, which will save them hereafter from the absurdity of enforcing a quarantine[12] in England against an amount of solar heat of which its climate is insusceptible. Let them further be repeated in the Schools of Medicine until the Professors become ashamed of imbuing the minds of the young with prejudice and false belief, which, should they ever visit warmer climates, may cause them to be eminently mischievous in vexing the commerce and deeply and injuriously agitating the public mind of whatever community may have received them.” 12. See Cases of the Eclair, Dygden, &c., post. Climate differs not only in different countries but in different parts of the same country. The climate of the country is different from that of the city. The climate of every city, town, and village, differs from that of every other. The temperature, the moisture, and the other meteorological conditions of different districts, nay, even of different streets in the same town, vary to such a degree as to influence materially their relative salubrity and the prevalence or absence of particular classes of disease. These local climatic conditions and their connection with prevalent diseases, have not as yet received due attention: when they shall have received it—and they will receive it— a new light will be shed on local epidemics. I pass now to Civilization. We have no sufficient knowledge of the state of the people and of their diseases, in any of the civilized nations of antiquity, to trace the relation between them. The authentic history of periods, comparatively near to our own time, as far as concerns the diseases of the people, goes scarcely further back than the 14th century. The first great epidemic, to which I have so often called attention, occurred in that century, and we have reliable evidence, both of the
- 42. phenomena attending this plague and the condition of the people at that time. I assume this period therefore as my starting-point. I take a civilized community to be one in which there exist— 1. A sovereign authority. 2. Laws incorruptibly administered. 3. Physical comfort generally diffused. 4. Intellectual development and activity generally diffused. 5. Recognition of the fundamental principles of religion and morality. Without the two first, there can be no security for life and property, both of which must be placed in absolute and unquestionable safety before a single step can be taken out of the lowest depth of barbarism. Without the two last, none of the others can be acquired. These conditions are therefore the basis of the pyramid of society. Taking these then as the essential constituents of civilization, and applying them as a test to Great Britain, we shall see that at the commencement of the 14th century England was in a state of barbarism, since every one of these elements was wanting, although the foundation of political and social institutions containing the germs of liberty and progress had been already laid. Practically, however, at that period there was no sovereign authority, for the king had no sufficient power to maintain order, to protect the rights and liberties of the people, or to defend his own throne against armed men nominally his subjects; while the lord of every feudal castle exercised a more perfect sovereignty over his vassals than the so-called monarch over the nation. Every town was a fortress, and every house in which it was safe to dwell a castle, the inmates of which, like people in a garrison, constantly held themselves prepared to resist attack, from which they were never secure. They slept with arms at their side.
- 43. Marauders openly encamped on the public roads for the plunder of the wayfarer, which often ended in his murder. Few persons ventured to travel alone, and none without the reasonable apprehension that they might never return alive. Scarcely a third part of the area of the kingdom was under cultivation. The remainder consisted of moor, forest, and fen. Vast tracts were under water during the greater part of the year, and at other times formed morasses, marshes, and swamps. Immediately beyond the walls that encompassed the towns were large stagnant ditches, which being the nearest receptacles for refuse, were full of all sorts of decomposing filth. The streets were narrow, unpaved, undrained, uncleansed, and unlighted. There was no provision for the removal of the town refuse. Gutters were formed at the sides of the streets, as in Bethnal Green and the neglected parts of all our towns at the present time, into which the inhabitants threw the refuse of their houses; forming in dry weather a semi-fluid mass of corrupting animal and vegetable matter, and in rainy weather black turbid rivulets which ultimately poured their contents into some water-course. The houses were mean and squalid, built of wood and wattles, thatched with straw, without chimneys, the windows without glass, the floors without boards, the furniture of the rudest description; the use of linen was scarcely known; common straw formed the king’s bed. “The floors,” says Erasmus, writing two centuries later, “generally are made of nothing but loam, and are strewed with rushes, which being constantly put on fresh, without a removal of the old, remain lying there, in some cases for twenty years; with fish bones, broken victuals, the dregs of tankards, and impregnated with other filth underneath, from dogs and men.” Contemporary writers concur in representing the offensive odour of decaying straw and rushes as universal in the houses. There was no knowledge of the art of collecting, preserving, and storing fodder. The animals for winter food were slaughtered in
- 44. autumn, and their flesh salted or smoked. It was only during three months of the year, from Midsummer to Michaelmas, that any fresh animal food, excepting game and river fish, was tasted even by the nobles of the land. The common people subsisted chiefly on salted beef, veal, and pork, the price of which was one-half less than that of wheat in the time of Henry VIII. There were no fresh vegetables. As late as the 18th century salads were sent from Holland for the table of Queen Caroline. Sir John Pringle, writing in the middle of the last century, states that his father’s gardener told him that in the time of his grandfather cabbages were sold for a crown a-piece. It was not until towards the close of the 16th century (1585) that the potato was first brought to England, where it was limited to the garden for at least a century and a half after it had been planted by Sir Walter Raleigh in his own garden. It was first cultivated as a field crop in Scotland so recently as the year 1752. For many centuries England remained in the condition of country in which no more subsistence is produced than is barely sufficient for the necessities of the people. Consequently every year of scarcity became a year of famine, and such years, about one in ten, occurred for ages with great regularity, and often equalled in their terrible results the worst famines of antiquity. In a cold climate fuel is nearly as important as food, for which indeed it is a substitute. A large portion of our daily food is used up in supporting that internal fire by which the heat of the human body in every climate, and under every variety of external temperature, is maintained at the 98th degree of Fahrenheit. The greater the loss of heat by cooling, the greater the amount of heat which the body itself must generate to maintain its temperature at this elevated point. This demand for additional heat cannot be supplied without additional quantities of food, and unless these supplies are afforded, the substance of the body itself, its very tissues and organs, are consumed; a process which cannot be continued long without exhaustion, disease, and death. The phrase “starved by cold”
- 45. expresses a more literal fact than is commonly understood. Unhappily the circumstances which deprive a population of the means of counteracting cold limit also the supplies of food at their command, and the pressure of the twofold privation, want of food and want of fuel, commonly occurs at the very season when both these indispensable supports of life are most needed. Some conception may be formed of the suffering to which our ancestors were exposed from this cause, from the fact that their prejudice against the use of coal as an article of fuel was such that a law was passed rendering it a capital offence to burn it within the City, and there is a record in the Tower importing that a person was tried, convicted, and executed for this offence in the reign of Edward the First. It was not until the reign of Charles the First that there was a regular supply of coals to London. The habits of the people increased the force of these privations. Intemperance was a national vice. Excessive carousing at home, or days and nights spent in taverns, was the usual practice among all classes, and the physical and moral evils resulting from the custom were neither redeemed nor lessened by the epithet which these habitual convivialities appear to have conferred upon the nation of “Merrie England.” Caius, indeed, one of the most celebrated physicians of the sixteenth century, couples Germany and the Netherlands with England in this common reproach. “These three nations,” he says, “destroy more meats and drynkes without all order, convenient time, reason, and necessitie, than all other countries under the son, to the great annoyance of their bodies and wittes.” This condition of the country and this mode of life themselves constitute the most powerful causes of epidemics; and an extraordinary concurrence and concentration of these causes are manifested in the combination of the circumstances which have been enumerated, namely, in the malarious state of the greater part of the kingdom, in the confined space of the towns, in the deficiency and putrescency of the food, in the inadequacy of the means of protection from cold, and in the intemperance of the people. These
- 46. were the true sources of the malignity and mortality of the pestilences of that age. We have no reliable evidence of the actual mortality produced by these terrible diseases; for no physician has left such an account of the epidemics of which he was an eye-witness as enables us to determine it, and there was no Registrar-General to fill up the momentous columns included in his death-roll. We can therefore only take the statements of the time as we find them. According to the accounts of contemporary writers, the Black Death swept away, within the space of four years, a fourth part of the population of Europe. Some towns in England are stated to have lost two-thirds of their inhabitants, and it is computed that one-half of the entire population of the country perished. Of the Sweating Sickness, Bacon says it “destroyed infinite persons;” Stowe “a wonderful number;” and other writers reckon the deaths in the places attacked by thousands. Similar representations are given of the ravages of the Plague, of the Petechial Fever, and even occasionally of Intermittent Fever; and the substantial correctness of these statements is confirmed by entries in parish registers still extant, which tell the story of the local outbreaks of those days with graphic and touching simplicity. During some of the worst of these visitations, contemporary writers concur in stating that the living were insufficient to bury the dead; business was suspended; the courts of law were closed; the churches were deserted for want of a sufficient number of clergy to perform the service; and ships were seen driving about on the ocean and drifting on shore, whose crews had perished to the last man. We can form no adequate conception of the terror inspired by these events. We have seen alarm in our own day, but then it bordered on maniacal despair. It seemed as if the last judgment had come upon the world, and men abandoned alike their possessions and their friends. The rich gave up their treasures and laid them at the foot of the altars; neighbour abandoned neighbour; parents their
- 47. offspring, and brothers their sisters. “If” says one of the chroniclers, “in a circle of friends any one only by a single word happened to bring the plague to mind, first one and then another of the company was seized with a tormenting anguish; certain that they were attacked with a mortal sickness, they slunk away home, and there soon yielded up the ghost.” These fearful forms of pestilence were accompanied by moral epidemics more appalling than the physical. Of these the two following may serve as examples:— Vast assemblages of men and women formed circles hand in hand, dancing, leaping, shouting, insensible to external impressions; some seeing visions and spirits whose names they shrieked out; others in epileptic convulsions with foaming at the mouth; all continuing to make the most violent muscular exertions for hours together, until they fell to the ground in a state of exhaustion. Lookers-on were seized with an uncontrollable impulse to join in these wild revels. Peasants left their ploughs, mechanics their workshops, servants their masters, boys and girls their parents, women their domestic duties, and men their business, thus to spend days and nights; these infatuated crowds passing furiously through streets, along highways, over fields, and from town to town. This madness pervaded the least barbarous countries of Europe for upwards of two centuries, under the name of the “Dancing Mania.” It was universally attributed to demoniacal possession, and its cure was attempted by exorcism. It was one expression and outlet of the violent passions of that time, imposture and profligacy playing principal parts in this strange drama. More pernicious than this madness was the mania of cruelty, an especial manifestation of which was the ferocious persecution of the Jews, who were put to death by hundreds and thousands, under the accusation that they had poisoned the wells. At Basle a number of this nation, whose European history proves them to have been everywhere amongst the most inoffensive of the people, were enclosed in a wooden building and burnt with it. At Strasburg two
- 48. thousand were burnt alive. Whoever showed them compassion and endeavoured to protect them were put upon the rack and burnt with them. In numerous instances these unhappy people, driven to despair, assembled in their own habitations, to which they set fire and consumed themselves with their families. The noble and the mean bound themselves by an oath to extirpate them from the face of the earth by fire and sword. In England this relentless cruelty took particularly the shape of burning innocent people under the name of witches; an infatuation which pervaded all classes from the highest to the lowest, affording a melancholy exemplification of the close alliance between credulity and cruelty.[13] 13. The number of wretched beings condemned and executed for this imaginary crime at the Assizes of Suffolk and Essex alone, in the year 1646, amounted to two hundred. Dr Zachary Gray affirms that he had seen an authentic account of persons who had so suffered in the whole of England, amounting to from three to four thousand. So late as the year 1697 seven persons, three men and four women, were burnt at Paisley for this alleged crime. We seldom sufficiently consider how near we are to those times of dreadful superstition and cruelty! How short a period it is since the light of a brighter day dawned upon us! But in the midst of these terrible disorders, changes which had been in silent operation during several centuries began to produce visible results. The independent power of the nobles had been suppressed; the feuds that raged between them, filling the country with disorder and bloodshed, had been put down; the supremacy of the law had been established; property and life had become more secure; industry had taken a surprising start; the practical abolition of serfdom had been to a large extent effected; and at last came the final breaking up of the feudal system in the reign of Henry VII. by the passing of the law authorizing the alienation of land. About the middle of the fifteenth century improvements in the condition of the people, which had been gradually effected by these changes, were accelerated by a succession of events that gave an extraordinary impulse to the human mind, just aroused from the
- 49. long and deep sleep of the middle ages—that dark night which was now passing away. Among the most memorable of these was the invention of printing, which the three immortal masters of the art had now completed (1436–1442), giving untiring and undying wings to thought;— The diffusion over the West of Europe of the remains of a former civilization, by the dispersion of the treasures of classical art, literature, and science, which before Constantinople fell into the hands of barbarians (1453) had been confined within the walls of that city;— The cessation of the long and disastrous struggle between the East and the West, by the expulsion of the Moors from Spain (1492); — The discovery of the New World;— And lastly, the Reformation, that stupendous work which with giant strength burst asunder the chain which consummate skill and supreme power had spent ages in forging and riveting: that stupendous work, which was not merely emancipation from spiritual bondage, but the re-communication of the long-lost spirit of religion; the noble men who achieved it being ever, even in their day of triumph, less intent on demolishing the gorgeous edifice that had held the mind enthralled, than on erecting a pure temple in which it might worship with sincerity and freedom. The time when the foundation was laid for this intellectual and spiritual renovation was also that of the commencement of physical improvement. The towns being no longer fortresses, it became unnecessary to maintain their fortifications. Walls were thrown down; stagnant moats were filled up; broader streets were opened; more convenient houses were erected. Forests were cleared; marshes and swamps were drained; more land was brought under cultivation; more vegetable matter was produced; the art of collecting, storing, and preserving fodder was discovered. Fresh
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