The Re-Emergence of H1N1 Influenza Virus in 1977: A Cautionary Tale for Estimating Divergence Times Using Biologically Unrealistic Sampling Dates

In 1977, H1N1 influenza A virus reappeared after a 20-year absence. Genetic analysis indicated that this strain was missing decades of nucleotide sequence evolution, suggesting an accidental release of a frozen laboratory strain into the general population. Recently, this strain and its descendants were included in an analysis attempting to date the origin of pandemic influenza virus without accounting for the missing decades of evolution. Here, we investigated the effect of using viral isolates with biologically unrealistic sampling dates on estimates of divergence dates. Not accounting for missing sequence evolution produced biased results and increased the variance of date estimates of the most recent common ancestor of the re-emergent lineages and across the entire phylogeny. Reanalysis of the H1N1 sequences excluding isolates with unrealistic sampling dates indicates that the 1977 re-emergent lineage was circulating for approximately one year before detection, making it difficult to determine the geographic source of reintroduction. We suggest that a new method is needed to account for viral isolates with unrealistic sampling dates

ИСПОЛЬЗОВАНИЕ ФЛУОРИМЕТРИИ В БЛИЖНЕЙ ИК-ОБЛАСТИ С ФОТОГРАФИЧЕСКОЙ РЕГИСТРАЦИЕЙ СИГНАЛА В МЕТОДЕ «ОТПЕЧАТКОВ ПАЛЬЦЕВ» С ДОБАВКОЙ ФЛУОРОФОРА К ОБЪЕКТУ: ДИСКРИМИНАЦИЯ ЯБЛОЧНЫХ СОКОВ

The application of dyes, that fluoresce in the near infrared (NIR, 700-800 nm) region, for the recognition of samples using a fingerprinting method with the addition of fluorophores to the samples (“fluorescent eye”) is proposed. The technique has been successfully applied to the classification of samples of various nature. In the current work, this strategy has been tested on the example of discrimination of 17 samples of apple juice from different manufacturers, purchased at different times. An indolenine series heptamethine carbocyanine dye in the presence of surfactants was used as the added fluorophore, red LEDs were used as an excitation source, and the signal was recorded using a digital camera with an additional IR filter installed; a spectrofluorimeter with a 96-well plate accessory was used to record the spectra. Photographic images were processed using Unscrambler X and Excel software. The results were presented using the following coordinates: intensity of NIR fluorescence - intensity of visible light reflection (using the photographic images). It was found that such presentation allowed the samples to be divided into groups associated with the manufacturer. We have also obtained intrinsic fluorescence spectra, including those with the addition of NIR dye, and these results were processed by the principal component analysis. It was possible to distinguish 5–6 groups of samples by their intrinsic emission, not counting the blank, while the spectra with the addition of the dye allowed to isolate the largest number of groups of samples (9). At the same time, the classification using spectra did not allow juices to be grouped by the producer. Also, obtaining photographs using a visualizer was easier and faster than recording the fluorescence spectra. The joint processing of emission spectra and photographs did not improve the quality of discrimination.Keywords: fingerprinting method, NIR fluorimetry, apple juice, principal component analysisDOI: http://dx.doi.org/10.15826/analitika.2022.26.1.005E.V. Skorobogatov, I.A. Stepanova, V.S. Orekhov, M.K. Beklemishev Department of Chemistry, Lomonosov Moscow State University, Russin Federation, 119991, GSP-1, Moscow, Leninskie gory, 1, building 3Предложено использовать красители, флуоресцирующие в ближней ИК (БИК) области спектра (700–800 нм), для распознавания объектов методом «отпечатков пальцев», основанным на добавке флуорофоров к объекту («флуоресцентный глаз»). Метод успешно применяется в классификации объектов различной природы. В данной работе метод опробован на примере дискриминации 17 образцов яблочного сока разных производителей, выпущенных в разное время. В качестве добавляемого флуорофора использовали гептаметиновый карбоцианиновый краситель индоленинового ряда в присутствии ПАВ, в качестве источника излучения – красные светодиоды, а сигнал регистрировали с помощью цифрового фотоаппарата с дополнительным ИК-светофильтром; для записи спектров применяли спектрофлуориметр с приставкой для 96-луночного флуориметрического планшета. Фотографические изображения обрабатывали с помощью стандартного программного обеспечения Unscrambler X и Excel. Результаты представили в координатах: интенсивность БИК-флуоресценции – интенсивность отражения видимого света (с использованием соответствующих фотографий). Обнаружили, что такое представление позволяет разделить образцы на группы, связанные с производителем. Получали также спектры собственной флуоресценции, в том числе с добавкой БИК-красителя, обрабатывая эти результаты методом главных компонент. По собственной эмиссии можно выделить 5–6 групп образцов, не считая контрольного, тогда как по спектрам с добавкой красителя удается добиться выделения наибольшего числа групп образцов (девять). При этом классификация с использованием спектров не позволяет группировать соки по производителям. Кроме того, получение фотографий с помощью визуализатора проще и экспресснее, чем регистрация спектров флуоресценции. Совместная обработка эмиссионных спектров и фотографий не позволяет повысить качество дискриминации образцов.Ключевые слова: метод «отпечатков пальцев», флуориметрия в ближней ИК-области, яблочный сок, метод главных компонентDOI: http://dx.doi.org/10.15826/analitika.2022.26.1.00

The use of NIR Fluorimetry with photographic data acquisition in the fingerprinting method with the addition of fluorophores to the samples: discrimination of apple juices

Предложено использовать красители, флуоресцирующие в ближней ИК (БИК) области спектра (700-800 нм), для распознавания объектов методом «отпечатков пальцев», основанным на добавке флуорофоров к объекту («флуоресцентный глаз»). Метод успешно применяется в классификации объектов различной природы. В данной работе метод опробован на примере дискриминации 17 образцов яблочного сока разных производителей, выпущенных в разное время. В качестве добавляемого флуорофора использовали гептаметиновый карбоцианиновый краситель индоленинового ряда в присутствии ПАВ, в качестве источника излучения - красные светодиоды, а сигнал регистрировали с помощью цифрового фотоаппарата с дополнительным ИК-светофильтром; для записи спектров применяли спектрофлуориметр с приставкой для 96-луночного флуориметрического планшета. Фотографические изображения обрабатывали с помощью стандартного программного обеспечения Unscrambler X и Excel. Результаты представили в координатах: интенсивность БИК-флуоресценции - интенсивность отражения видимого света (с использованием соответствующих фотографий). Обнаружили, что такое представление позволяет разделить образцы на группы, связанные с производителем. Получали также спектры собственной флуоресценции, в том числе с добавкой БИК-красителя, обрабатывая эти результаты методом главных компонент. По собственной эмиссии можно выделить 5-6 групп образцов, не считая контрольного, тогда как по спектрам с добавкой красителя удается добиться выделения наибольшего числа групп образцов (девять). При этом классификация с использованием спектров не позволяет группировать соки по производителям. Кроме того, получение фотографий с помощью визуализатора проще и экспресснее, чем регистрация спектров флуоресценции. Совместная обработка эмиссионных спектров и фотографий не позволяет повысить качество дискриминации образцов.The application of dyes, that fluoresce in the near infrared (NIR, 700-800 nm) region, for the recognition of samples using a fingerprinting method with the addition of fluorophores to the samples (“fluorescent eye”) is proposed. The technique has been successfully applied to the classification of samples of various nature. In the current work, this strategy has been tested on the example of discrimination of 17 samples of apple juice from different manufacturers, purchased at different times. An indolenine series heptamethine carbocyanine dye in the presence of surfactants was used as the added fluorophore, red LEDs were used as an excitation source, and the signal was recorded using a digital camera with an additional IR filter installed; a spectrofluorimeter with a 96-well plate accessory was used to record the spectra. Photographic images were processed using Unscrambler X and Excel software. The results were presented using the following coordinates: intensity of NIR fluorescence - intensity of visible light reflection (using the photographic images). It was found that such presentation allowed the samples to be divided into groups associated with the manufacturer. We have also obtained intrinsic fluorescence spectra, including those with the addition of NIR dye, and these results were processed by the principal component analysis. It was possible to distinguish 5-6 groups of samples by their intrinsic emission, not counting the blank, while the spectra with the addition of the dye allowed to isolate the largest number of groups of samples (9). At the same time, the classification using spectra did not allow juices to be grouped by the producer. Also, obtaining photographs using a visualizer was easier and faster than recording the fluorescence spectra. The joint processing of emission spectra and photographs did not improve the quality of discrimination.Работа выполнена при поддержке РНФ (грант № 20-13-00330). Авторы благодарят доцента Т.А.Подругину за предоставление карбоцианинового красителя и А.Добротворского за предоставление ИК-фотоаппарата.Current work was supported by the Russian Science Foundation (grant No 20-13-00330). The authors are grateful to Associate Professor T.A. Podrugina for providing the carbocyanine dye and A. Dobrotvorsky for providing the IR photo camera

Rationality of the moduli spaces of plane curves of sufficiently large degree

We prove that the moduli space of plane curves of degree d is rational for all sufficiently large d.Comment: 18 pages; 1 figure; Macaulay2 scripts used can be found at http://www.uni-math.gwdg.de/bothmer/rationality/ or at the end of the latex source fil

Colorimetric determination of cetyltrimethylammonium bromide by using aggregation with a carbocyanine dye

Обнаружено, что коммерческий карбоцианиновый краситель IR-783, содержащий сульфогруппы, в слабощелочной среде образует агрегаты с бромидом цетилтриметиламмония (ЦТАБ) с появлением новой полосы поглощения, изменением цвета раствора с синего на желтый (Dl = 350 нм) и изменением интенсивности флуоресценции в ближней ИК области. ЦТАБ определяли фотометрическим методом, фотографируя реакционную смесь в 96-луночном планшете камерой смартфона или в визуализаторе Camag. В качестве аналитического сигнала использовали разность интенсивностей красного и синего каналов (R - B), соответствующую желтому цвету. Диапазон определяемых концентраций в водном растворе составляет (3 - 25)·10-6 М, предел обнаружения 1.6·10-6 М, относительное стандартное отклонение 2-5 %. Определению не мешает ряд неионных ПАВ, неорганических солей и полимеров, мешают анионные ПАВ. Другие катионные ПАВ также дают сигналы, но с разной чувствительностью. Характеристики литературной методики определения ЦТАБ на основе красителя кумасси бриллиантовый синий G-250 и предлагаемой сопоставимы. Проанализирован образец лизирующего буфера, содержащего ЦТАБ.It was found that the commercial carbocyanine dye IR-783 containing sulfo groups forms aggregates with cetyltrimethylammonium bromide (CTAB) in a slightly alkaline medium yielding a new absorption band, a change in the solution color from blue to yellow (Dl = 350 nm), and a change in the fluorescence intensity in the near-IR region. CTAB was determined by the photometric method by photographing the reaction mixture in a 96-well plate with a smartphone camera or in a Camag visualizer. The difference between the intensities of the red and blue channels (R - B), corresponding to the yellow color, was used as an analytical signal. The linear range in an aqueous solution is (3 - 25)·10-6 M, the detection limit is 1.6·10-6 M, and the relative standard deviation is 2-5%. The determination is not affected with a number of non-ionic surfactants, inorganic salts and polymers; the anionic surfactants interfere. Other cationic surfactants also give analytical signals, but with different sensitivities. The characteristics of the literature method for the determination of CTAB based on the Coomassie brilliant blue G-250 dye and the proposed method are comparable. A sample of CTAB-containing lysing buffer solution was analyzed.Работа выполнена при поддержке Российского научного фонда (проект № 20-13-00330). Авторы благодарят Т.А. Подругину и И.А. Дорошенко за предоставленный краситель IR-783 и Н.С. Мелик-Нубарова за краситель кумасси. Исследование проведено в рамках Программы развития Междисциплинарной научно-образовательной школы Московского университета «Будущее планеты и глобальные изменения окружающей среды».This work was supported by the Russian Science Foundation (grant No 20-13-00330). The authors are grateful to T.A. Podrugina and I.A. Doroshenko for providing the IR-783 dye and N.S. Melik-Nubarov for the Coomassie dye. The study was conducted within the framework of the Development Program of the Interdisciplinary Scientific and Educational School of Moscow University “The Future of the Planet and Global Environmental Changes”

Tools for crushing diatoms – opal teeth in copepods feature a rubber-like bearing composed of resilin

Diatoms are generally known for superior mechanical properties of their mineralised shells. Nevertheless, many copepod crustaceans are able to crush such shells using their mandibles. This ability very likely requires feeding tools with specific material compositions and properties. For mandibles of several copepod species silica-containing parts called opal teeth have been described. The present study reveals the existence of complex composite structures, which contain, in addition to silica, the soft and elastic protein resilin and form opal teeth with a rubber-like bearing in the mandibles of the copepod Centropages hamatus. These composite structures likely increase the efficiency of the opal teeth while simultaneously reducing the risk of mechanical damage. They are supposed to have coevolved with the diatom shells in the evolutionary arms race, and their development might have been the basis for the dominance of the copepods within today's marine zooplankton

The maternal and early embryonic transcriptome of the milkweed bug Oncopeltus fasciatus

<p>Abstract</p> <p>Background</p> <p>Most evolutionary developmental biology ("evo-devo") studies of emerging model organisms focus on small numbers of candidate genes cloned individually using degenerate PCR. However, newly available sequencing technologies such as 454 pyrosequencing have recently begun to allow for massive gene discovery in animals without sequenced genomes. Within insects, although large volumes of sequence data are available for holometabolous insects, developmental studies of basally branching hemimetabolous insects typically suffer from low rates of gene discovery.</p> <p>Results</p> <p>We used 454 pyrosequencing to sequence over 500 million bases of cDNA from the ovaries and embryos of the milkweed bug <it>Oncopeltus fasciatus</it>, which lacks a sequenced genome. This indirectly developing insect occupies an important phylogenetic position, branching basal to Diptera (including fruit flies) and Hymenoptera (including honeybees), and is an experimentally tractable model for short-germ development. 2,087,410 reads from both normalized and non-normalized cDNA assembled into 21,097 sequences (isotigs) and 112,531 singletons. The assembled sequences fell into 16,617 unique gene models, and included predictions of splicing isoforms, which we examined experimentally. Discovery of new genes plateaued after assembly of ~1.5 million reads, suggesting that we have sequenced nearly all transcripts present in the cDNA sampled. Many transcripts have been assembled at close to full length, and there is a net gain of sequence data for over half of the pre-existing <it>O. fasciatus </it>accessions for developmental genes in GenBank. We identified 10,775 unique genes, including members of all major conserved metazoan signaling pathways and genes involved in several major categories of early developmental processes. We also specifically address the effects of cDNA normalization on gene discovery in <it>de novo </it>transcriptome analyses.</p> <p>Conclusions</p> <p>Our sequencing, assembly and annotation framework provide a simple and effective way to achieve high-throughput gene discovery for organisms lacking a sequenced genome. These data will have applications to the study of the evolution of arthropod genes and genetic pathways, and to the wider evolution, development and genomics communities working with emerging model organisms.</p> <p>[The sequence data from this study have been submitted to GenBank under study accession number SRP002610 (<url>http://www.ncbi.nlm.nih.gov/sra?term=SRP002610</url>). Custom scripts generated are available at <url>http://www.extavourlab.com/protocols/index.html</url>. Seven Additional files are available.]</p

Gene expression patterns associated with blood-feeding in the malaria mosquito Anopheles gambiae

BACKGROUND: Blood feeding, or hematophagy, is a behavior exhibited by female mosquitoes required both for reproduction and for transmission of pathogens. We determined the expression patterns of 3,068 ESTs, representing ~2,000 unique gene transcripts using cDNA microarrays in adult female Anopheles gambiae at selected times during the first two days following blood ingestion, at 5 and 30 min during a 40 minute blood meal and at 0, 1, 3, 5, 12, 16, 24 and 48 hours after completion of the blood meal and compared their expression to transcript levels in mosquitoes with access only to a sugar solution. RESULTS: In blood-fed mosquitoes, 413 unique transcripts, approximately 25% of the total, were expressed at least two-fold above or below their levels in the sugar-fed mosquitoes, at one or more time points. These differentially expressed gene products were clustered using k-means clustering into Early Genes, Middle Genes, and Late Genes, containing 144, 130, and 139 unique transcripts, respectively. Several genes from each group were analyzed by quantitative real-time PCR in order to validate the microarray results. CONCLUSION: The expression patterns and annotation of the genes in these three groups (Early, Middle, and Late genes) are discussed in the context of female mosquitoes' physiological responses to blood feeding, including blood digestion, peritrophic matrix formation, egg development, and immunity