bluuebunny/crossref_metadata_2025 huggingface dataset:
Why Existence Does Not Emerge as a Distinct Concept in Greek Philosophy*
Authors: Kahn Charles H
Published: November 2023

In the extended discussion of the concept (or concepts) of Being in Greek philosophy from Parmenides to Aristotle, the theme of existence does not figure as a distinct topic for philosophical reflection. My aim here is to defend and illustrate this claim, and at the same time to suggest some of the reasons why it is that the concept of existence does not get singled out as a topic in its own right. Finally, I shall raise in a tentative way the question whether or not the neglect of this topic was necessarily a philosophical disadvantage.
The Manambu Language of East Sepik, Papua New Guinea
Authors: Aikhenvald Alexandra Y , Ala Jacklyn Yuamali , Agnes Pauline , Laki Yuaneng Luma
Published: November 2023

This book presents the first comprehensive description of the Manambu language of Papua New Guinea. Manambu belongs to the Ndu language family, and is spoken by about 2,500 people in five villages: Avatip, Yawabak, Malu, Apa:n, and Yambon (Yuanab) in East Sepik Province, Ambunti district. About 200-400 speakers live in the cities of Port Moresby, Wewak, Lae, and Madang; and a few live in Kokopo and Mount Hagen. The book is based entirely on the author’s fieldwork. After an introductory account of the language and its speakers, Professor Aikhenvald devotes chapters to phonology, grammatical relations, word classes, gender, semantics, number, case, possession, derivation and compounding, pronouns, morphology, verbs, mood and modality, negation, clauses, pragmatics, discourse, semantics, the lexicon, current directions of change, and genetic relationship to other languages. The description is presented in a clear style in a framework that will be comprehensible to all linguists and linguistic anthropologists.
Phonology
Authors: Aikhenvald Alexandra Y , Ala Jacklyn Yuamali , Agnes Pauline , Laki Yuaneng Luma
Published: November 2023

Manambu is more complex in its phonology than most other Ndu languages. It has a large inventory of phonemes (twenty-one consonants and nine vowels), contrastive stress, and an array of phonological processes. Most loans are assimilated to the Manambu phonology. Morphophonological processes relevant for nouns are discussed in §4.1.1, and those relevant for verbs in §11.3. Examples throughout the grammar are given in their phonological representation. The phonological sequences y@ and w@ are rendered as yi and wu, to reflect their most frequent surface realization.
Gender Marking, Semantics, and Agreement
Authors: Aikhenvald Alexandra Y , Ala Jacklyn Yuamali , Agnes Pauline , Laki Yuaneng Luma
Published: November 2023

Manambu is among the few languages of the world where the gender assignment involves animacy and sex, and also correlates with the shape and size of the referent. ‘Shape-based’ gender assignment appears to be an areal feature of the Sepik area (a somewhat similar system has been described for Alamblak by Bruce 1984; also see Aikhenvald 2000: 276–8 for a discussion of Manambu in a typological perspective). Manambu gender is, by and large, ‘covert’: that is, it is not marked on the noun itself. Two genders, masculine and feminine, are distinguished in singular. They are realized through agreement within a noun phrase and on the predicate.
Directionals and Valency-Changing Devices
Authors: Aikhenvald Alexandra Y , Ala Jacklyn Yuamali , Agnes Pauline , Laki Yuaneng Luma
Published: November 2023

Manambu verbs fall into four categories depending on which directional markers they take, if any. We distinguish four classes of verbs, two closed and two open. The forms and the meanings of the inherently directional verbs, the directional markers with intrinsically directional verbs, and bound directional forms with optionally directional verbs (I–III below) are given in Table 16.1. This table also contains directional forms used with demonstratives (see Table 10.5, which includes some verbal directionals). An etymological connection between the roots of inherently directional verbs, the directional suffixes on intrinsically directional verbs, and bound directionals is apparent. The shared morphemes are in bold. Table 16.2 features the directional suffixes used with intrinsically directional verbs.
Clause Linking and Dependent Clauses
Authors: Aikhenvald Alexandra Y , Ala Jacklyn Yuamali , Agnes Pauline , Laki Yuaneng Luma
Published: November 2023

Clause linking in Manambu is achieved using a variety of means. The major strategy is ‘clause chaining’ via medial dependent clauses which in most cases involves switch-reference marking. Other clause-linking devices are considered in Chapter 19. Further issues of semantics of clause linking are addressed in Aikhenvald (forthcoming g). Neither dependent medial clauses nor dependent clauses of any other sort can be considered core arguments, or complements, of the main clause. They are akin to peripheral arguments in that they provide temporal, conditional, locative, and manner specification to the main clause. That is, they cannot be considered embedded clauses in the way complement clauses would be. Their further affinity with peripheral arguments is corroborated by the fact that their marking may involve one case and the suffix ‘like’ also used on nouns and noun phrases (§19.3).
Other Dependent Clauses and Further Features of Clause Linking
Authors: Aikhenvald Alexandra Y , Ala Jacklyn Yuamali , Agnes Pauline , Laki Yuaneng Luma
Published: November 2023

We will now discuss clause-linking devices other than clause chaining via medial dependent clauses (see Aikhenvald forthcoming g, on their semantics). These include: juxtaposition of a dependent clause and a main clause (§19.1); relative clauses (§19.2); clause linking via a case marker ‘instead’ and a suffix ‘like’ (§19.3); purposive and desiderative clauses (§19.4); speech reports (§19.5); clause linking involving connectives (§19.6); and juxtaposition of main clauses (§19.7). Manambu does not have complement clauses as a type on their own. Some types of medial dependent clauses, juxtaposed clauses, purposives, desideratives, speech reports, and nominalizations can be used as complementation strategies—see §19.8. Dependent clauses can be used as main clauses. This phenomenon, known as desubordination, is addressed in §19.9, together with grammaticalization and reinterpretation of ‘medial’ verbal forms.
Falling Back Together in the 1980s: The Continuing Voyage of the Mary Rose
Authors: Wright Patrick , Krauze Andrzej
Published: November 2023

Poor old Mary Rose has had quite a bit to contend with these last five centuries. When she went down, on a sparkling summer’s morning in 1545, she was the jewel in the crown of Henry VIII’s Navy. But her 91 guns were no match for the ruthless onslaught of the murderous currents that flow beneath the Solent. A deep scourpit was eroded on the port side of the hull. The currents then relentlessly shrouded her in a thick layer of silt. By the early eighteenth century a hard layer of shelly clay had sealed the once proud warship in her watery grave. On October 11th 1982, a salvage operation on a scale never before attempted brought the forlorn remains of the Mary Rose to the surface.
From Markers To Mechanisms: Using Psychophysiological Measures To Elucidate Basic Processes Underlying Aggressive Externalizing Behaviour
Authors: Patrick Christopher J , Bernat Edward M
Published: November 2023

The brain is the essential common pathway through which heredity and experience operate to shape core psychological processes that determine behaviour. Understanding the behaviour of persistently violent individuals thus requires consideration of phenomena at varying levels of analysis: genes and environments; nervous system anatomy and physiology; neural representations and networks; perceptual, cognitive, and affective functions; and overt action and its regulation. Psychophysiological approaches to the study of aggression fill a crucial niche in this analytic hierarchy by providing information about brain and bodily responses associated with ongoing psychological and behavioural processes in time. The aim of this chapter is to provide a framework for thinking about findings from existing published studies of this kind, and to highlight in particular the unique contribution that advanced electrocortical (EEG/ERP) measurement techniques can make to our understanding of neuropsychological processing dynamics underlying impulsive aggressive behaviour.
Introduction: perspectives on the evolution of language in Africa
Authors: Knight Chris
Published: November 2023

Africa was the cradle of language, mind, and culture. Until recently, the evidence for this remained little known. The prevailing ‘‘human revolution’’ theory saw modern language and cognition emerging suddenly and nearly simultaneously throughout the Old World some 40 to 50 thousand years ago. This ‘‘Great Leap Forward’’ for humanity (Diamond 1992) was depicted as a cognitive transition based on a neural mutation yielding syntax and hence true language (Klein 1995, 2000; Tattersall 1995). When modern Homo sapiens evolved in Africa some 200,000–150,000 years ago, according to this theory, our ancestors were modern only ‘‘anatomically’’; mentally and behaviorally, they remained archaic. Only when such humans began migrating out of Africa—triggering the Middle-to-Upper Paleolithic transition in Europe—did the ‘‘leap’’ to cognitive and behavioral modernity occur.
The ideal of my heart
Authors: Larson Frances
Published: November 2023

On 25 June 1901 Henry Wellcome married Syrie Barnardo, daughter of Dr Thomas Barnardo, the great Victorian philanthropist and founder of the Barnardo homes for disadvantaged children. News of the marriage delighted Wellcome’s friends, but most expressed their surprise at his decision to marry after so many years alone. Wellcome was two months shy of his 48th birthday when he wed; his new wife was just two weeks short of her 22nd. They had known each other for four years, during which time Wellcome had become friends with Syrie’s father, but nonetheless, their courtship was short and their marriage rushed. Many of Wellcome’s friends had never met his future wife when they found themselves writing congratulatory letters at the news of their engagement. Only a few days later the same friends were reading announcements of the Wellcomes’ small, family wedding at St. Mark’s Church, Surbiton, in their morning papers.
Excuse me Mr Treve
Authors: Larson Frances
Published: November 2023

On 26 January 1899, Thompson attended a sale at the prestigious auction rooms of Sotheby’s in Covent Garden. It was one of the first auction sales he attended as Wellcome’s agent. He was armed with a list of early medical manuscripts to bid for, and the price limits that Wellcome had agreed for each lot. His visit was a success and he managed to buy nearly everything Wellcome wanted. The only item he missed was a fifteenth-century manuscript, which was bought by the son of a well-known book dealer whom Thompson recognized at the sale. In a calculated act of revenge, once bidding had exceeded the £5 Thompson was willing to spend on the manuscript he continued to bid, driving the price upwards, merely for the satisfaction of knowing that Mr Quaritch would have to pay more for his prize. Thompson’s presence at the sale had caused a stir among London’s established book dealers, particularly as he had successfully outbid them a number of times during the sale, ‘much to their annoyance’. As he was a relatively new player on the London auction circuit, no one recognized Thompson, but the considerable amount of money he was spending was enough to convince Sotheby’s regular clientele that they needed to find out who he was.
The finest historical medical museum in the world
Authors: Larson Frances
Published: November 2023

The Wellcome Historical Medical Exhibition—later Museum—opened in the summer of 1913. Wellcome, who had already postponed the opening for eight years, later said that he had not meant to open his Exhibition for a further ten years, but various ‘eminent medical men insisted’ that he make his collections ‘the centre’ of the historical section of the International Medical Congress that year. Wellcome’s claim that he had been persuaded to exhibit only by his peers was not entirely true: Thompson had suggested synchronizing the Exhibition with the Congress, and had been lobbying the organizers for their support since early 1910. Wellcome liked to think his collection had been propelled into the limelight, against his better instincts, by the medical establishment, because it was the medical establishment that he wanted to impress with his efforts as a collector. The International Medical Congress was held every four years and had not visited London since 1881. In 1913 it attracted around 5,000 delegates from twenty-eight countries, ‘the most remarkable gathering of the world’s doctors that has ever assembled’.
This is the History of Medicine
Authors: Larson Frances
Published: November 2023

Wellcome insisted that his Museum was for research, but very little research went on there. He spoke of his aim to create a museum ‘as an institution for post-graduate study’, with workrooms and laboratories and lecture halls, but he never opened its doors to the researchers that could bring such a place to life. The Historical Medical Museum was nominally part of the Wellcome Bureau of Scientific Research, which had been established in 1914 as an umbrella organization for the Wellcome Research Laboratories and the Museum of Tropical Medicine and Hygiene. The latter was a specialized teaching collection established in London that year in the wake of Wellcome’s successful research programme in Sudan. The Museum of Tropical Medicine and Hygiene was part of the Bureau, both physically and administratively, and was used by army officers, missionaries, and educationalists preparing to work abroad. But the Wigmore Street museum remained independent and selfcontained, and no formal teaching programme was developed there. Moreover, publishing was discouraged, staff were not allowed to discuss Wellcome’s historical collections with anyone outside the institution, and only a few papers were given at conferences. Thompson had drafted one or two booklets each year between 1905 and 1914, but they were designed primarily as promotional literature for the firm and were published anonymously under the Burroughs Wellcome name. After the First World War, these dried up.
Why do the Laws Explain Why?
Authors: Lange Marc
Published: November 2023

What exactly is this criterion, that laws must explain the phenomena? ... What makes laws so well suited to secure us this good? When laws give us ‘satisfying’ explanations, in what does this warm feeling of satisfaction consist? There are indeed philosophical accounts of explanation, and some mention laws very prominently; but they disagree with one another, and in any case I have not found that they go very far toward answering these questions.
The Petrine Era and After 1689–1740
Authors: Alexander John T
Published: November 2023

Peter I is associated with many ‘firsts’ in Russian history. He was the first legitimate Muscovite ruler to have that name in Russian and European languages (Piter in Dutch), the first to use a Roman numeral after his name, the first to travel incessantly by land and water and to venture abroad, the first to be titled emperor and ‘the Great, Most Wise Father of the Father-land’, the first to inspire radical change in diverse spheres of activity, the first to found urban sites sharing his name, the first to be buried in St Petersburg, and the first to imprint his name on an entire era encompassing the birth of modern Russia in an expanded, European context.
Revolutionary Russia 1890–1914
Authors: Zelnik Reginald E
Published: November 2023

Any analysis of imperial Russia at the end of the nineteenth century and beginning of the twentieth must include the obvious but essential reminder that Russia was still an autocracy (samo- derzhavie). This was not simply a garden-variety old regime, with a divinely anointed absolute monarch at the top and a system of legally defined social estates (sosloviia= Stände), headed by the nobility defining its social hierarchy. It was also an old regime whose sovereigns, even the most ‘liberal’ (Alexander II), self-consciously resisted the dilution of their sovereign power, the delegation of that power to intermediary institutions, and its limitation by any constitutional mechanism.
Russia in War and Revolution 1914–1921
Authors: Orlovsky Daniel T
Published: November 2023

The years separating the outbreak of war in 1914 and the announcement of the ‘New Economic Policy’ in 1921 form a critical watershed in modern Russian history. The revolutions of 1917, while not an unbridgeable caesura, fundamentally transformed the polity and social order. This era also had a profound impact on the ‘bourgeois’ West, which in the coming decades had to contend with the spectre of a socialist Prometheus in the East.

In a memorandum of February 1914 P. N. Durnovo, a former Minister of Internal Affairs, implored the emperor to avoid war with Germany.
Urban Trends 400 – 1500
Authors: Clark Peter
Published: November 2023

Most of the foundations of the European urban order were laid in the medieval period, but the process was difficult and uneven, characterized by surges of growth (as between the eleventh and thirteenth centuries) and contraction (such as the late fourteenth and fifteenth centuries), by variations between different European regions, and by the instability of individual cities and towns.
Culture And Landscape 400 – 1500
Authors: Clark Peter
Published: November 2023

The Church was central to the urban identity and to the cultural world of the medieval town. That identity was also shaped by public buildings, especially, in the case of bigger cities, by town walls and gates. However, as towns expanded after the eleventh and twelfth centuries and faced growing pressures from high mobility, social tension, party factionalism, and spatial sprawl, communities (or at least their elites) sought to reinforce their sense of urban identity in further ways— through new patrician houses, new religious institutions, and new civic buildings.
Social Life 1500 – 1800
Authors: Clark Peter
Published: November 2023

Living in the parish of St Michael, Cornhill, in the heart of London as the young servant of a wealthy fishmonger John Bathurst, Robert Smith was infected with the smallpox virus towards the middle of May 1655, probably after talking with a sick neighbour or selling fish to a customer. Not immediately affected, he carried on work but ten days later he showed the classic symptoms of fever, backache, and vomiting, and soon after tiny red spots spread over his body.
Culture and Landscape 1500 – 1800
Authors: Clark Peter
Published: November 2023

On the eve of the Reformation, the iconic image of the European city, portrayed in early maps and landscapes, on civic seals, and as the backcloth in religious paintings or patrician portraits, was that of a Christian community defined by its town walls and gates, and embellished with a spiky coronet of church spires. This urban representation followed a tradition dating back to the early Middle Ages.
Urban Trends 1800 – 2000
Authors: Clark Peter
Published: November 2023

The last part of this book examines the period from the early nineteenth century to the late twentieth century when, step by step, Europe became an urban continent. After the Second World War the majority of the European population, in some countries the great majority of the population, was living in towns rather than villages, and conurbations became increasingly numerous. In 1990, according to one estimate, Europe had 225 cities with over 200,000 inhabitants and forty of its cities counted more than a million people.
Social Life 1800 – 2000
Authors: Clark Peter
Published: November 2023

Caroline Luckhurst was the daughter of a middle-class family in late Victorian London. Her father was a prosperous railway official wearing a top hat on public occasions and residing in a spacious semi-detached villa in the leafy suburbs of Blackheath. A younger child, apparently disadvantaged in the family, Caroline moved out of the capital to the countryside, worked as a servant in a large house, and met there the youngest son of a small farmer. Much to her family’s anger, they married, had several children, and fell into relative poverty.
Culture And Landscape 1800 – 2000
Authors: Clark Peter
Published: November 2023

In Hjalmar Söderberg’s novel Doktor Glas (1905), the Swedish doctor declares: ‘Never have I felt happier and prouder to be from a city than when I returned from the countryside as a child one autumn evening and saw the lights glowing … Now I thought those poor devils back in the country will have to … trudge about in the dark and the dirt’. During the modern period, European city culture, increasingly self-assured and multifaceted, wiped off the last traces of rural tradition, and European cultural life turned urban.
What Causes the Effects of the Pill?
Authors: Guillebaud John , Maccregor Anne
Published: November 2023

All the good and bad effects must have their ultimate explanation in the chemistry of the body. More than 100 different laboratory tests on blood, urine, and other bodily fluids have given abnormal results in women on the pill (Table 4.1).
What if I get Side-Effects?
Authors: Guillebaud John , Maccregor Anne
Published: November 2023

Changes in bleeding patterns are probably the most frequently reported side-effects of the pill, particularly when the pill is first started. Let the pill packet rule your pill taking, not the bleeding pattern—whatever that may be. Do not stop taking pills before the end of a packet because of bleeding, even if it seems like a period. Pregnancies have happened that way.
Popular Opinion in Nazi Germany as a Factor in the Policy of the ‘Solution of the Jewish Question’: The Nuremberg Laws and the Reichskristallnacht
Authors: Kulka Otto Dov
Published: November 2023

This chapter examines two sample cases of the interrelation between radical expressions of German popular opinion and the policy of the totalitarian racialist regime of the Third Reich concerning the ‘Jewish Question’. A systematic examination of this relation became possible following the publication of a digital edition of all available secret Nazi reports on the German population’s attitudes toward the Jews and the anti-Jewish policy of the regime between 1933 and 1945. Here, a text-analytical approach will be combined with quantitative, computer-based research.
The Semantics of Clause Linking in Kham
Authors: Watters David E
Published: November 2023

The language treated in this chapter is the Takale dialect of Kham (spoken in Taka village), something of a lingua franca in Kham territories. Kham is a complex of Tibeto-Burman languages centered primarily around a knot of 4,000-metre ridges at the western end of the Dhaulagiri Massif in the remote highlands of Rukum and Rolpa Districts, ‘Mid-western Nepal’, and in the north-western corner of Baglung District. The total population of all dialects is estimated to be about 60,000 speakers. The language is vigorous in all domains of life; most pre-school children speak only Kham, and there are more speakers now than ever before.
The Semantics of Clause Linking in Konso
Authors: Mous Maarten , Oda Ongaye
Published: November 2023

Konso is a Cushitic language of about 200,000 speakers spoken in southern Ethiopia not far from the Kenyan border. Its closest relatives are Dirayta (or Gidole) and Oromo. Konso society is divided into farmers (etanta) and traders or craftsmen (xawdáa); farmers are in higher esteem. The people live in walled towns and their social organization is both by clan and by locality. The clan rulers also have some religious functions. The Konso are famous for their statues (waakkaa) for deceased heroes and clan leaders. The people live in a hilly area which is heavily overpopulated. Every inch of land is used for agriculture and river banks are protected against erosion to save farm land. The farmers grow many different crops which are often intermingled in the fields. Fields are family owned, fenced with low stone walls and have their own proper names. Konso culture has been extensively studied; some major publications are Hallpike (1972), Amborn (1989), and Watson (1998).
Isolation of anaerobes from clinical material
Authors: Levett P N
Published: November 2023

With the notable exceptions of the clostridial diseases such as tetanus and botulism, infections caused by obligate anaerobes are endogenous in origin. Such infections are invariably caused by non-sporing anaerobes derived from the normal flora of the large intestine, the female genital tract, or the oropharynx (1). Because of their origin these are usually mixed infections involving several species of anaerobes in addition to facultative anaerobes such as colifonns. Experience throughout the world during the last 20 years has shown that obligate anaerobes are usually the principal components of such infections and that effective therapy depends upon eradication of the anaerobes (2).
Methods for biochemical studies
Authors: Forsythe S J
Published: November 2023

Numerous methods of quantitative and qualitative biochemical analysis are now available for the study of anaerobic micro-organisms. Fortunately, most biochemical studies use standard protocols and thus the major problem for the anaerobic microbiologist is being able to grow sufficient quantities of the organism. Certain procedures, however, such as rcductase enzymes, require strictly anaerobic assay conditions, but even this problem can be overcome through the use of anaerobic cabinets, anaerobic cuvettes, and artificial electron carriers. This chapter will concentrate, therefore, on the analysis of oxygen¬ sensitive components from anaerobic micro-organisms.
Methanogens
Authors: Jain Mahendra K , Zeikus J Gregory , bhatnagar Lakshmi
Published: November 2023

Methanogenic bacteria are a morphologically diverse group of bacteria unified by their ability to produce methane. This property has been used to distinguish these bacteria from the other groups of anaerobic bacteria.
Identification of protein sequence motifs
Authors: Saqi Mansoor A S
Published: November 2023

One of the main aims of computational biology is to address the question: “Given a newly-determined sequence what can be said about the structure and function of the corresponding protein”. If a similarity search (1–3) against the sequence database reveals a global similarity with another sequence of known function, much can immediately be inferred (see Chapter 2). If the similarity is with a sequence of known three-dimensional structure homology modelling studies can be initiated (4) (see Chapter 6).
Identification of common post-translational modifications
Authors: Krishna Radha G , Wold Finn
Published: November 2023

Considering that some of the 20 primary, encoded amino (imino) acids can be converted to something close to 200 secondary amino acids in post-translational reactions (1), it should not be surprising to find that the number and complexity of the methods involved in recognizing and characterizing these secondary amino acids are quite impressive. The chemical properties of the secondary amino acid derivatives vary broadly and complicate any attempts to extract some general rules and approaches.
Immunochemical analysis of protein conformation
Authors: Friguet Bertrand , Djavadi-Ohaniance, Lisa , Goldberg Michel E
Published: November 2023

Antibodies can be used to study protein conformation in two ways. One, already outlined in Chapter 3, is to take advantage of their specificity and of the sensitivity of immunochemical detection methods to trace the protein after an appropriate analytical procedure. This approach is particularly useful when the biological activity of the protein is not easy to test. This is the case for instance with engineered proteins that have lost some of their original properties. Using immunochemical detection, it is possible to determine, even with crude extracts, such physicochemical parameters as molecular weight (by electrophoresis; see Chapters 1 and 8), diffusion coefficient (by gel filtration; see Chapter 9), or sedimentation coefficient (by sucrose gradient), ligand fixation (by immunoprecipitation of the ligand-bound protein), and stability (by irreversible heat precipitation). In this approach, polyclonal as well as monoclonal antibodies can be used, as long as the antibodies are able to detect the protein with sufficient efficiency and specificity.
Fusion protein purification methods
Authors: Brewer Stephen J , Glatz Charles E , Dickerson Charles
Published: November 2023

Genetic engineering has provided scientists and technologists with the opportunity to change the properties of proteins so that their purification properties can be improved. By the insertion of DNA sequences on either the 3’ or 5’ ends of translated DNA, the amino acid sequences on either end of a protein can be changed to provide a purification fusion. These fusions may be used to simply stabilize the protein from attack by proteases by helping the protein form an inclusion body. The fusion might also impart specific purification properties on the protein, making it suitable for immunoaffinity, metal chelate, ion exchange, hydrophobic chromatography, or partition. Alternatively, for well-characterized proteins, specific amino acids may be changed within the protein to introduce patches with a specific affinity for an adsorption matrix.
Initial purification of inclusion bodies
Authors: Violand Bernard N
Published: November 2023

Recombinant DNA technology has made possible the production of many proteins which were previously available in only limited quantities from natural sources. This technology has resulted in the generation of many therapeutically important protein products and large numbers of other commercially significant proteins. High level expression of both heterologous and homologous proteins frequently results in the formation of dense amorphous insoluble aggregates, named inclusion bodies, in the host cytoplasm. This chapter discusses some of the properties of inclusion bodies and also describes the most frequently used methods for preparing them. Because the majority of this data has been generated in E. coli, this chapter will concentrate on production of inclusion bodies from this host cell.
Protein purification from milk
Authors: Burr Richard
Published: November 2023

This chapter illustrates examples of techniques used in the purification of common milk proteins. It outlines some of the specific factors to consider in the handling, purification, and analysis of milk components. Simple protocols for the purification of major bovine milk proteins are provided and some newer developments that are applicable to the isolation of milk proteins are discussed.
Efficacy and Safety of a Polypill to Reduce Cardiovascular Events: A Review of Clinical Trials
Authors: Kelly Michael S. , Dacey Ana , Siana Audrey , Ojeda Jason
Published: November 2023

Cardiovascular disease continues to be the leading cause of mortality globally. Modifiable risk factors, such as hypertension and dyslipidemia, can be managed through lifestyle and pharmacotherapy treatments to reduce the risk of primary and secondary major cardiovascular events in patients with elevated risk. Despite effective and available medications to manage and mitigate cardiovascular risk factors, control rates of hypertension and dyslipidemia are suboptimal, and greater efforts are needed to reduce cardiovascular event rates worldwide. A polypill containing several classes of medications proven to lower cardiovascular risk in a single-dose form has been associated with improved medication adherence over multiple single-ingredient medications and may lead to reduced cardiovascular events. The goal of this article is to review available data from clinical trials assessing the efficacy and safety of polypills compared with placebo or usual care for cardiovascular risk reduction. Three databases were searched (PubMed/MEDLINE, CINAHL, and ScienceDirect) for randomized trials that compared a single polypill with usual care or placebo and reported major adverse cardiovascular events for each study group. A total of 6 trials were selected for inclusion. Several polypill formulations were compared with placebo or usual care with multiple single-ingredient medications in study populations consisting of both primary and secondary prevention patients. Overall, the polypill seems to be associated with reduced major adverse cardiovascular event and comparable safety with usual care treatment with an added benefit of improved adherence over multiple single-ingredient medications. The polypill has potential to be a cost-effective intervention to reduce the global burden of cardiovascular disease.
Attentional bias toward fearful faces is associated with maternal postnatal distress and alterations in white matter microstructure in 5-year-old females
Authors: Kumpulainen Venla , Kataja Eeva-Leena , Pulli Elmo P. , Copeland Anni M. , Silver Eero A. , Haikio Tuomo , Saukko Ekaterina , Korja Riikka , Karlsson Linnea , Karlsson Hasse , Tuulari Jetro J.
Published: November 2023

The development of face processing system is paramount for social interaction and communication skills. Eye-tracking provides a viable means to investigate face processing even in very young children and can be used to derive indices of affective processing biases that are adaptive in supporting preparedness for emotional encoding. The neural correlates of early-emerging attentional biases have not been explored.

In the current study, we gathered diffusion tensor imaging (DTI) and eye-tracking data (attention disengagement from neutral, happy, fearful faces, and control pictures towards distractor stimuli) to investigate the association of white matter (WM) microstructural alterations and attentional biases in 117 (55 female) typically developing 5-year-old children. We estimated fractional anisotropy (FA) and used tract-based spatial statistics (TBSS) before conducting voxel-wise analyses testing associations between attentional biases and brain diffusivity.

We found that reduced WM integrity (indexed as decrease in FA) in widespread WM regions, including the splenium of corpus callosum, left anterior limb of internal capsule (ALIC), left posterior limb of internal capsule (PLIC), left posterior thalamic radiation and optic tract, predicted higher attentional bias toward fearful expressions in females (adjusted for potential confounders). Further, maternal postnatal anxiety and depressive symptoms were detected to associate positively with attentional bias toward fearful expression, but only in females.

Based on these findings and prior results on association between maternal distress and reduced WM integrity in female offspring, it is possible that alterations in WM microstructure may transmit the long-term effects of maternal mental distress during early life to increased vigilance towards negative emotional expressions. These hypotheses and sexual dimorphism in the observed associations require replication and are potential focus areas for future studies.
Transferability of ecological forecasting models to novel biotic conditions in a long-term experimental study
Authors: Dumandan Patricia Kaye T. , Simonis Juniper L. , Yenni Glenda M. , Ernest S. K. Morgan , White Ethan P.
Published: November 2023

Ecological forecasting models play an increasingly important role for managing natural resources and assessing our fundamental knowledge of processes driving ecological dynamics. As global environmental change pushes ecosystems beyond their historical conditions, the utility of these models may depend on their transferability to novel conditions. Because species interactions can alter resource use, timing of reproduction, and other aspects of a species’ realized niche, changes in biotic conditions, which can arise from community reorganization events in response to environmental change, have the potential to impact model transferability. Using a long-term experiment on desert rodents, we assessed model transferability under novel biotic conditions to better understand the limitations of ecological forecasting. We show that ecological forecasts can be less accurate when the models generating them are transferred to novel biotic conditions, and that the extent of model transferability can depend on the species being forecast. We also demonstrate the importance of incorporating uncertainty in forecast evaluation with transferred models generating less accurate and more uncertain forecasts. These results suggest that how a species perceives its competitive landscape can influence model transferability, and that when uncertainties are properly accounted for, transferred models may still be appropriate for decision making. Assessing the extent of the transferability of forecasting models is a crucial step to increase our understanding of the limitations of ecological forecasts.

Open Research Statement

Data (10.5281/zenodo.8436468) and code (10.5281/zenodo.11053744) used to conduct data analyses are archived in Zenodo and supplementary results (10.6084/m9.figshare.25733691.v1) in Figshare.
Ciliary intrinsic mechanisms regulate dynamic ciliary extracellular vesicle release from sensory neurons
Authors: Wang Juan , Saul Josh , Nikonorova Inna A. , Cruz Carlos Nava , Power Kaiden M. , Nguyen Ken C. , Hall David H. , Barr Maureen M.
Published: November 2023

Cilia-derived extracellular vesicles (EVs) contain signaling proteins and act in intercellular communication. Polycystin-2 (PKD-2), a transient receptor potential channel, is a conserved ciliary EVs cargo.Caenorhabditis elegansserves as a model for studying ciliary EV biogenesis and function.C. elegansmales release EVs in a mechanically-induced manner and deposit PKD-2-labeled EVs onto the hermaphrodite vulva during mating, suggesting an active release process. Here, we study the dynamics of ciliary EV release using time-lapse imaging and find that cilia can sustain the release of PKD-2-labeled EVs for a two-hour duration. Intriguingly, this extended release doesn’t require neuronal synaptic transmission. Instead, ciliary intrinsic mechanisms regulate PKD-2 ciliary membrane replenishment and dynamic EV release. The ciliary kinesin-3 motor KLP-6 is necessary for both initial and extended ciliary EV release, while the transition zone protein NPHP-4 is required only for sustained EV release. The dihydroceramide desaturase DEGS1/2 ortholog TTM-5 is highly expressed in the EV-releasing sensory neurons, localizes to cilia, and is required for sustained but not initial ciliary EV release, implicating ceramide in ciliary ectocytosis. The study offers a comprehensive portrait of real-time ciliary EV release, and mechanisms supporting cilia as proficient EV release platforms.
One-carbon unit supplementation fuels tumor-infiltrating T cells and augments checkpoint blockade
Authors: Xu Xincheng , Chen Zihong , Bartman Caroline R. , Xing Xi , Olszewski Kellen , Rabinowitz Joshua D.
Published: November 2023

Nucleotides perform important metabolic functions, carrying energy and feeding nucleic acid synthesis. Here, we use isotope tracing-mass spectrometry to quantitate the contributions to purine nucleotides of salvage versusde novosynthesis. We further explore the impact of augmenting a key precursor for purine synthesis, one-carbon (1C) units. We show that tumors and tumor-infiltrating T cells (relative to splenic T cells) synthesize purinesde novo. Purine synthesis requires two 1C units, which come from serine catabolism and circulating formate. Shortage of 1C units is a potential bottleneck for anti-tumor immunity. Elevating circulating formate drives its usage by tumor-infiltrating T cells. Orally administered methanol functions as a formate pro-drug, with deuteration enabling control of formate-production kinetics. In MC38 tumors, safe doses of methanol raise formate levels and augment anti-PD-1 checkpoint blockade, tripling durable regressions. Thus, 1C deficiency can gate antitumor immunity and this metabolic checkpoint can be overcome with pharmacological 1C supplementation.

Statement of significance

Checkpoint blockade has revolutionized cancer therapy. Durable tumor control, however, is achieved in only a minority of patients. We show that the efficacy of anti-PD-1 blockade can be enhanced by metabolic supplementation with one-carbon donors. Such donors support nucleotide synthesis in tumor-infiltrating T cells and merit future clinical evaluation.
Partitioning the Impacts of Spatial-Temporal Variation in Demography and Dispersal on Metapopulation Growth Rates
Authors: Schreiber Sebastian J.
Published: November 2023

Abstract

Spatial-temporal variation in environmental conditions is ubiquitous in nature. This variation simultaneously impacts survival, reproduction, and movement of individuals and, thereby, the rate at which metapopulations grow. Using the tools of stochastic demography, the metapopulation growth rate is decomposed into five components corresponding to temporal, spatial, and spatial-temporal variation in fitness, and spatial and spatial-temporal covariation in dispersal and fitness. While temporal variation in fitness always reduces the metapopulation growth rate, all other sources of variation can either increase or reduce the metapopulation growth rate. Increases occur either by reducing the impacts of temporal variation or by generating a positive fitness-density covariance where individuals tend to concentrate in higher-quality patches. For example, positive auto-correlations in spatial-temporal variability in fitness generate this positive fitness-density covariance for less dispersive populations, but decrease it for highly dispersive populations, e.g. migratory species. Negative auto-correlations in spatialtemporal variability have the opposite effects. Positive covariances between movement and future fitness, on short or long time scales, increase growth rates. These positive covariances can arise is unexpected ways. For example, the win-stay, lose-shift dispersal strategy in negatively autocorrelated environments can generate positive spatial covariances that exceed negative spatial-temporal covariances. This decomposition of the metapopulation growth rate provides a way to quantify the relative importance of fundamental sources of variation on metapopulation persistence.
A Massively Parallel Screen of 5′UTR Mutations Identifies Variants Impacting Translation and Protein Production in Neurodevelopmental Disorder Genes
Authors: Plassmeyer Stephen P. , Florian Colin P. , Kasper Michael J. , Chase Rebecca , Mueller Shayna , Liu Yating , White Kelli McFarland , Jungers Courtney F. , Djuranovic Slavica Pavlovic , Djuranovic Sergej , Dougherty Joseph D.
Published: November 2023

De novomutations cause a variety of neurodevelopmental disorders including autism. Recent whole genome sequencing from individuals with autism has shown that manyde novomutations also occur in untranslated regions (UTRs) of genes, but it is difficult to predict from sequence alone which mutations are functional, let alone causal. Therefore, we developed a high throughput assay to screen the transcriptional and translational effects of 997 variants from 5′UTR patient mutations. This assay successfully enriched for elements that alter reporter translation, identifying over 100 potentially functional mutations from probands. Studies in patient-derived cell lines further confirmed that these mutations can alter protein production in individuals with autism, and some variants fall in genes known to cause syndromic forms of autism, suggesting a diagnosis for these individual patients. Since UTR function varies by cell type, we further optimized this high throughput assay to enable assessment of mutations in neuronsin vivo. First, comparingin cellulotoin vivoresults, we demonstrate neurons have different principles of regulation by 5′UTRs, consistent with a more robust mechanism for reducing the impact of RNA secondary structure. Finally, we discovered patient mutations specifically altering the translational activity of additional known syndromic genesLRRC4andZNF644in neurons of the brain. Overall our results highlight a new approach for assessing the impact of 5′UTR mutations across cell types and suggest that some cases of neurodevelopmental disorder may be caused by such variants.
Strong Effect of Demographic Changes on Tuberculosis Susceptibility in South Africa
Authors: Oyageshio Oshiomah P. , Myrick Justin W. , Saayman Jamie , van der Westhuizen Lena , Al-Hindi Dana , Reynolds Austin W. , Zaitlen Noah , Uren Caitlin , Möller Marlo , Henn Brenna M.
Published: November 2023

South Africa is among the world’s top eight TB burden countries, and despite a focus on HIV-TB co-infection, most of the population living with TB are not HIV co-infected. The disease is endemic across the country with 80-90% exposure by adulthood. We investigated epidemiological risk factors for tuberculosis (TB) in the Northern Cape Province, South Africa: an understudied TB endemic region with extreme TB incidence (645/100,000) and the lowest provincial population density. We leveraged the population’s high TB incidence and community transmission to design a case-control study with population-based controls, reflecting similar mechanisms of exposure between the groups. We recruited 1,126 participants with suspected TB from 12 community health clinics, and generated a cohort of 878 individuals (cases =374, controls =504) after implementing our enrollment criteria. All participants were GeneXpert Ultra tested for active TB by a local clinic. We assessed important risk factors for active TB using logistic regression and random forest modeling. Additionally, a subset of individuals were genotyped to determine genome-wide ancestry components. Male gender had the strongest effect on TB risk (OR: 2.87 [95% CI: 2.1-3.8]); smoking and alcohol consumption did not significantly increase TB risk. We identified two interactions: age by socioeconomic status (SES) and birthplace by residence locality on TB risk (OR = 3.05, p = 0.016) – where rural birthplace but town residence was the highest risk category. Finally, participants had a majority Khoe-San ancestry, typically greater than 50%. Epidemiological risk factors for this cohort differ from other global populations. The significant interaction effects reflect rapid changes in SES and mobility over recent generations and strongly impact TB risk in the Northern Cape of South Africa. Our models show that such risk factors combined explain 16% of the variance (r2) in case/control status.