SARS transmission pattern in Singapore reassessed by viral sequence variation analysis

Background

Epidemiological investigations of infectious disease are mainly dependent on indirect contact information and only occasionally assisted by characterization of pathogen sequence variation from clinical isolates. Direct sequence analysis of the pathogen, particularly at a population level, is generally thought to be too cumbersome, technically difficult, and expensive. We present here a novel application of mass spectrometry (MS)–based technology in characterizing viral sequence variations that overcomes these problems, and we apply it retrospectively to the severe acute respiratory syndrome (SARS) outbreak in Singapore.

Methods and Findings

The success rate of the MS-based analysis for detecting SARS coronavirus (SARS-CoV) sequence variations was determined to be 95% with 75 copies of viral RNA per reaction, which is sufficient to directly analyze both clinical and cultured samples. Analysis of 13 SARS-CoV isolates from the different stages of the Singapore outbreak identified nine sequence variations that could define the molecular relationship between them and pointed to a new, previously unidentified, primary route of introduction of SARS-CoV into the Singapore population. Our direct determination of viral sequence variation from a clinical sample also clarified an unresolved epidemiological link regarding the acquisition of SARS in a German patient. We were also able to detect heterogeneous viral sequences in primary lung tissues, suggesting a possible coevolution of quasispecies of virus within a single host.

Conclusion

This study has further demonstrated the importance of improving clinical and epidemiological studies of pathogen transmission through the use of genetic analysis and has revealed the MS-based analysis to be a sensitive and accurate method for characterizing SARS-CoV genetic variations in clinical samples. We suggest that this approach should be used routinely during outbreaks of a wide variety of agents, in order to allow the most effective control.     

Long-range air transmission of bacteria.

Bacterial spores from a sandstorm area north of the Black Sea were transmitted to Sweden by air, giving increased concentrations of viable bacterial spores at two air sampling stations in Sweden.     

Watching liquid droplets of TDP-43CTD age by Raman spectroscopy

Liquid–liquid phase separation (LLPS) is a biological phenomenon wherein a metastable and concentrated droplet phase of biomolecules spontaneously forms. A link may exist between LLPS of proteins and the disease-related process of amyloid fibril formation; however, this connection is not fully understood. Here, we investigated the relationship between LLPS and aggregation of the C-terminal domain of TAR DNA-binding protein 43, an amyotrophic lateral sclerosis–related protein known to both phase separate and form amyloids, by monitoring conformational changes during droplet aging using Raman spectroscopy. We found that the earliest aggregation events occurred within droplets as indicated by the development of β-sheet structure and increased thioflavin-T emission. Interestingly, filamentous aggregates appeared outside the solidified droplets at a later time, suggestive that amyloid formation is a heterogeneous process under LLPS solution conditions. Furthermore, the secondary structure content of aggregated structures inside droplets is distinct from that in de novo fibrils, implying that fibril polymorphism develops as a result of different environments (LLPS versus bulk solution), which may have pathological significance.     

Quantitative studies of liquid food transmission in ants

Summary Tranmission of honey in several species of ants was studied using radioactive iodide as a tracer. Great variation in transmission rates between species was noted, ranging from negligible transmission over a ten-day period (inPogonomyrmex badius) to complete colony saturation within thirty hours (inFormica spp.). The honey was passed mostly among workers, very little being given to the queens or larvae. Indirect evidence is cited which suggests the occurrrence of chain transmission beyond the primary donations given by the original foragers.

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Zusammenfassung Die Uebertragung von Honig bei verschiedenen Arten von Ameisen wurde mit Hilfe von Zugabe radioaktiven Iodids untersucht. Eine mit der Mischung gefütterte Arbeiterin wurde in das Nest gesetzt, und die Verteilung des von ihr abgegebenen Honigs gemessen. Die Verteilungs-geschwindigkeit war sehr verschieden je nach der untersuchten Art: praktisch zu vernachlässigende Werte, sogar nach 10 Tagen, beiPogonomyrmex badius; vollständige Sättigung des Staates, nach 30 Stunden, beiFormica spp. Der Honig wurde hauptsächlich unter Arbeiterinnen verteilt; die Königinnen und Larven bekamen, wenn überhaubt, nur minimale Mengen. Die Resultate schienen darauf hinzuweisen, daß zur Verteilung im Staate nicht nur die Abgaben der ursprünglich gefütterten Arbeiterin verantwortlich waren, sondern auch Uebertragung der Empfänger untereinander.

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Résumé On a étudié la transmission du miel chez plusieurs espèces de Fourmis en se servant d'iodure radio-actif comme traceur. On observe des différences considérables dans le taux de la transmission d'une espèce à l'autre, allant d'une transmission négligeable pour une période de dix jours (chezPogonomyrmex badius) à une saturation intégrale de la colonie en trente heures (chez certaines espèces deFormica). Le plus souvent, le miel est passé d'une ouvrière à une autre, les reines et les larves recevant très peu de miel. Une preuve indirecte suggère une transmission en chaîne au-delà des dons primaires par les ouvrières fourragères nourries à la source.

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The authors wish to express their appreciation to Mr. E. W. Samuel, of the Harvard Biological Laboratories, whose expert aid and advice in the use of radioactive tracer technique made this study possible. Research was supported in part by a U. S. Public Health Service grant held by Eisner.     

Volume transmission signalling via astrocytes

The influence of astrocytes on synaptic function has been increasingly studied, owing to the discovery of both gliotransmission and morphological ensheathment of synapses. While astrocytes exhibit at best modest membrane potential fluctuations, activation of G-protein coupled receptors (GPCRs) leads to a prominent elevation of intracellular calcium which has been reported to correlate with gliotransmission. In this review, the possible role of astrocytic GPCR activation is discussed as a trigger to promote synaptic plasticity, by affecting synaptic receptors through gliotransmitters. Moreover, we suggest that volume transmission of neuromodulators could be a biological mechanism to activate astrocytic GPCRs and thereby to switch synaptic networks to the plastic mode during states of attention in cerebral cortical structures.     

Evidence of scrapie transmission via milk

Background

The risk of scrapie infection increases with increased duration and proximity of contact between sheep at lambing. Scrapie infectivity has not been detected in milk but cellular prion protein, the precursor of disease-associated prion protein PrP^d, has been found in milk from ruminants. To determine whether milk is able to transmit scrapie, 18 lambs with a prion protein genotype associated with high susceptibility to scrapie (VRQ/VRQ) were fed milk from twelve scrapie-affected ewes of the same genotype, and 15 VRQ/VRQ sheep reared on scrapie-free dams served as controls.

Results

Three lambs fed milk from scrapie-affected ewes were culled due to intercurrent diseases at 43, 44 and 105 days of age respectively, and PrP^d was detected in the distal ileum of the first two lambs, whilst PrP^d was not found in lymphoreticular tissues in the third lamb. A control lamb, housed in a separate pen and culled at 38 days of age, was also negative for PrP^d in a range of tissues. Samples of recto-anal mucosa associated lymphoid tissue collected from the remaining 15 live lambs at seven months of age (between five to seven months after mixing) were positive for PrP^d in the scrapie milk recipients, whereas PrP^d was not detected in the remaining 14 controls at that time. A subsequent sample collected from control lambs revealed PrP^d accumulation in two of five lambs eight months after mixing with scrapie milk recipients suggestive of an early stage of infection via lateral transmission. By contrast, the control sheep housed in the same building but not mixed with the scrapie milk recipients were still negative for PrP^d.

Conclusion

The presence of PrP^d in distal ileum and rectal mucosa indicates transmission of scrapie from ewe to lamb via milk (or colostrum) although it is not yet clear if such cases would go on to develop clinical disease. The high level of infection in scrapie-milk recipients revealed by rectal mucosal testing at approximately seven months of age may be enhanced or supplemented by intra-recipient infection as these lambs were mixed together after feeding with milk from scrapie-affected ewes and we also observed lateral transmission from these animals to lambs weaned from scrapie-free ewes.

     

Mother-to-infant transmission of SIV via breast-feeding in rhesus macaques

To decipher the mechanisms involved in oral transmission of human immunodeficiency virus/simian immunodeficiency virus (HIV/SIV) through breast-feeding, we have developed an animal model using SIV-infected lactating rhesus macaques (Macaca mulatta) and their infants. Five of eight macaque infants became infected during a 10-month study course after SIV inoculation of lactating dams. In a second study, three of four chronically infected female macaques transmitted virus to their infants through breast-feeding within 4 months of birth. Transmission of virus to infants did not correlate with viral loads in either milk or plasma. Infants were infected with homogeneous virus populations, while milk samples near the time of transmission were more diverse. These studies suggest that specific viral phenotypes are selectively transmitted through breast-feeding.     

Epidemiology, transmission dynamics and control of SARS: the 2002-2003 epidemic

This paper reviews current understanding of the epidemiology, transmission dynamics and control of the aetiological agent of severe acute respiratory syndrome (SARS). We present analyses of data on key parameters and distributions and discuss the processes of data capture, analysis and public health policy formulation during the SARS epidemic are discussed. The low transmissibility of the virus, combined with the onset of peak infectiousness following the onset of clinical symptoms of disease, transpired to make simple public health measures, such as isolating patients and quarantining their contacts, very effective in the control of the SARS epidemic. We conclude that we were lucky this time round, but may not be so with the next epidemic outbreak of a novel aetiological agent. We present analyses that help to further understanding of what intervention measures are likely to work best with infectious agents of defined biological and epidemiological properties. These lessons learnt from the SARS experience are presented in an epidemiological and public health context.     

Kibizu concentrated liquid suppresses the accumulation of lipid droplets in 3T3-L1 cells

Adipocyte size is closely related to the occurrence of diabetes, metabolic syndrome, and insulin resistance. Thus, researchers are searching for active substances that function to reduce adipocyte size. In the present study, we focused on sugar cane vinegar, Kibizu, and evaluated the function of Kibizu to reduce adipocyte size by using an in vitro model system, because people in Amami Oshima famous for longevity regularly consume Kibizu. Results showed that Kibizu treatment significantly reduced the size and number of lipid droplets in 3T3-L1 cells, relative to treatment with Kurozu, another traditional vinegar. Results of an extraction experiment suggest that the active components in Kibizu are lipophilic and hydrophobic. In addition, an in vivo experiment on rats treated with Kibizu showed that the active components were contained in large vein blood. Results of an additional in vivo experiment suggest that metabolites generated by Kibizu-treated rats are primarily contained or modified specifically in the large vein blood.     

严重急性呼吸综合征(SARS)病人血清抗体水平的初步探讨

为了了解严重急性呼吸综合征(SARS)病人血清中的抗体产生规律,对56份病人血清和36份健康人血清,分别采用间接酶联免疫吸附试验(EIA)检测IgG抗体和捕获法检测IgM抗体。结果显示：56份病人血清中有48份IgG抗体阳性,7份IgM抗体阳性。     

Fermentation process air filtration via cartridge filters

The use of cartridge air filters for process air sterilization in commercial fermentation has proliferated in recent years. Sterile air cartridge filter performance is discussed. The use of dispersed oil particle (DOP) tests for in situ integrity testing is described and discussed.     

Structure of the nanobubble clusters of dissolved air in liquid media

A qualitative model of the nucleation of stable bubbles in water at room temperature is suggested. This model is completely based on the property of the affinity of water at the nanometer scale; it is shown that under certain conditions the extent of disorder in a liquid starts growing, which results in a spontaneous decrease of the local density of the liquid and in the formation of nanometer-sized voids. These voids can serve as nuclei for the following generation of the so-called bubstons (the abbreviation for bubbles, stabilized by ions). The model of charging the bubstons by the ions, which are capable of adsorption, and the screening by a cloud of counter-ions, which are incapable of adsorption, is analyzed. It was shown that, subject to the charge of bubston, two regimes of such screening can be realized. At low charge of bubston the screening is described in the framework of the known linearized Debye–Huckel approach, when the sign of the counter-ion cloud preserves its sign everywhere in the liquid surrounding the bubston, whereas at large charge this sign is changed at some distance from the bubston surface. This effect provides the mechanism of the emergence of two types of compound particles having the opposite polarity, which leads to the aggregation of such compound particles by a ballistic kinetics.     

The effect of air ions on liquid evaporation rates

The evaporation rates of water, heptane, ethyl alcohol and physiological saline were investigated as a function of atmospheric ion density.Ion concentrations up to about 10⁶ ions/cm³ were employed. In general evaporation rates decreased with increasing ion density (of either sign)to a minimum after which evaporation increased steadily. For water a reduction of about 15% occurred.The different liquids exhibited some deviations from this general pattern. The effects are specific to the type of gas ion, being exceptionally great for ionized oxygen. They also seem to be associated with the polar nature of the liquid molecules. The results are qualitatively explained on the basis of surface charging.The modification of evaporation may be a mechanism by which biological effects are produced by air ions.

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Zusammenfassung Die Verdunstungsquoten von Wasser,Heptan,Äthylalkohol und physiologischer Kochsalzlösung wurden als Funktion der atmosphärischen Ionenkonzentration untersucht. Es wurden Ionenkonzentrationen oberhalb von etwa 10⁶ Ionen/cm³ benutzt. Im allgemeinen nahmen die Verdunstungsquoten mit steigender Ionenkonzentration (jeden Vorzeichens) bis zu einem Minimum ab,nach dem die Verdunstung wieder allmählich anstieg. Für Wasser ergab sich eine Abnahme um etwa 15%. Die verschiedenen Flüssigkeiten zeigten einige Abweichungen von diesem allgemeinen Trend. Die Wirkungen sind für Gasionen spezifisch und besonders gross für ionisierten Sauerstoff.Sie scheinen auch mit dem Vorzeichen der flüssigen Moleküle gekoppelt zu sein.Die Ergebnisse werden qualitativ mit der Oberflächenladung erklärt. Die Änderung der Verdunstung ist möglicherweise ein Mechanismus, durch den biologische Wirkungen durch Luftionen hervorgerufen werden.

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Resume On a étudié le taux d'évaporation de l'eau, de l'heptane, de l'alcool éthylique et du sérum physiologique (solution acqueuse de sel de cuisine)en fonction de la densité des ions dans l'atmosphère. Pour ce faire, on a utilisé des concentrations en ions allant jusqu'à 10⁶ ions/cm³. En général, le taux d'évaporation diminue parallèlement à l'augmentation de la densité des ions (quel que soit leur signe) et cela jusqu'à un minimum au-delà duquel il augmente de nouveau. Pour l'eau,la réduction est d'environ 15%. Les différents liquides examinés présentent quelques variantes de cette tendance générale. L'effet constaté est spécifique pour les ions gazeux et spécialement important pour l'oxygène ionisé. Il paraît aussi lié à la polarité de la molécule liquide. Les résultats obtenus sont qualitativement explicables en tenant compte de la charge superficielle du liquide. La variation de l'évaporation est peut-être le mécanisme par le truchement duquel les ions de l'air exercent une influence biologique.

     

The transmission of genes via pollen and ovules in gynodioecious angiosperms

Gynodioecy is defined as the class of sexual dimorphism in which females are constant (functionally unisexual) and males are inconstant (bisexual). Quantitative measures describe the sexual performance of males in gynodioecious populations. The relative (male/female) ovule contribution compares the ovule contributions of the sexes to the next generation. Male constancy and inconstancy, the proportions of male genes transmitted via pollen and ovules, are direct measure of maleness and femaleness. Equations permitting the values of the sex parameters to be estimated are derived. The values obtained for twelve species show that gynodioecy is not a single breeding system, but merges into hermaphroditism and dioecy at opposite extremes. The distribution of inconstancies among individual males is described for six species. In most species, seed counts overestimate the ovule contributions of males.     

Cholinergic transmission via central synapses in the locust nervous system

In this study we examine the nature of chemical synaptic transmission between identified filiform hair receptors on the prothoracic segment of a locust and the identified postsynaptic projection interneuron (A4I1). The effects of pressure ejected acetylcholine, and various ligands of acetylcholine receptors on the activity of the postsynaptic neuron A4I1, or on wind-elicited responses in A4I1 are reported. It is suggested that the transmitter of the afferent fibers is acetylcholine, and that fast transmission is mediated by nicotinic acetylcholine-receptors. Both nicotine and carbachol act as agonists, whereas d-tubocurarine and alpha-bungarotoxin act as antagonists. The presence of muscarinic acetylcholine receptors was also evident from the modulatory effects of muscarine, oxotremorine and pilocarpine, which were blocked by bath application of atropine. GABA, and its agonists muscimol and cis-4-amino-crotonic-acid lead to inhibition of A4I1 responses. This inhibition was prevented by the additional application of picrotoxin. This suggests involvement of a ligand-gated GABA receptor which, most likely, increases chloride conductance. Metabotropic GABA-receptors do not seem to be involved, since baclofene, diazepam and bicuculline ejections had no effects. Glutamate also inhibits wind elicited A4I1 responses. Although attempts were made to further characterize the receptor involved, tested substances such as kainic acid, glycine, CNQX or GDEE had no effect.