Analysis of the salivary microbiome using culture-independent techniques

The salivary microbiota is a potential diagnostic indicator of several diseases. Culture-independent techniques are required to study the salivary microbial community since many of its members have not been cultivated

Analysis of the salivary microbiome using culture-independent techniques

The salivary microbiota is a potential diagnostic indicator of several diseases. Culture-independent techniques are required to study the salivary microbial community since many of its members have not been cultivated

Инфекция вируса герпеса человека 7 типа у детей. Клинико-лабораторные аспекты

Objective: to study the activity of HSV 1,2, EBV, CMV, HHV6, HHV7 in children of different age groups, to determine the frequency of various symptoms and syndromes in the examined children depending on hhv7 monoinfection and combined herpetic infection in the age aspect.Materials and methods: 56 children aged 2 to 18 years were examined at the state MEDICAL research Institute named after Prof. A. F. Agafonov in Kazan, at the BIOMED medical and diagnostic center in Kazan and at the LUCH children’s clinic in Naberezhnye Chelny (boys accounted for 62.5% of cases, girls-37.5%). Verification of the diagnosis and evaluation of the dynamics of etiological markers of the infectious process in all observed children was performed by PCR. Blood and saliva were examined to detect markers of human herpes virus type 7 (in qualitative PCR), and blood, oropharyngeal flushing, and urine were examined in semiquantitative PCR to determine other viruses from the herpes family (HHV6, CMV, EBV). In enzyme immunoassay is determined At the same time, we studied options for combining laboratory markers of herpesvirus activity and changes in the main laboratory parameters used in routine practice. We studied the dynamics of the observed parameters against the background of complex antiviral therapy for the next 3 months after the initial examination. The frequency of occurrence of various symptoms and syndromes was determined depending on the activity of various representatives of the herpes family, their combination, and the age of the subjects.Results of the study: HHV7-monoinfection is more typical for the age group of 7-14 years. The significance of mixed HHV6+HHV7 infection decreased with age. The combination of activity of 4 herpesviruses was more often registered in the age categories 2-7 years and 14-18 years. More often, patients had tonsillitis, generalized cervical lymphadenopathy, periodic temperature rise to subfebrile or febrile numbers, pharyngitis, recurrent ARI, rhinitis, which allows us to consider them as the main manifestations of HHV7 infection.Conclusion: HHV7 infection is significant in the modern pathology of the child. The presence of such syndromes as recurrent tonsillitis, cervical lymphadenopathy or isolated enlargement of submandibular lymph nodes, periodic fever, pharyngitis, recurrent ARI in combination with hepato - and/ or splenomegaly, fatigue, and headache is often associated with HHV7. HHV7-viremia is relatively rare, despite obvious clinical and laboratory signs of inflammation indicating the presence of a viral disease, which, apparently, and is the cause of underdiagnosis and lack of proper attention to this infection. Цель: изучение активности ВПГ 1,2, ЭБВ, ЦМВ, ВГЧ6, ВГЧ7 у детей различных возрастных групп, определение частоты встречаемости различных симптомов и синдромов у обследованных детей в зависимости от моноинфекции ВГЧ7 и сочетанной герпетической инфекции в возрастном аспекте.Материалы и методы: обследовано 56 детей в возрасте от 2 до 18 лет в Республиканской клинической инфекционной больнице им. проф. А.Ф. Агафонова г. Казани, ООО «Лечебно-диагностическом центре БИОМЕД» г. Казани и детской клинике «ЛУЧ» г. Набережные Челны (мальчики составили 62,5% наблюдений, девочки – 37,5%). Верификация диагноза и оценка динамики этиологических маркеров инфекционного процесса у всех наблюдавшихся детей производилась методикой ПЦР. Исследовали кровь и слюну для обнаружения маркеров вируса герпеса человека 7 типа (в качественной ПЦР), в полуколичественной методике ПЦР исследовали кровь, смыв из ротоглотки, мочу с целью определения других вирусов из семейства герпесов (ВГЧ6, ЦМВ, ЭБВ). В иммуноферментном анализе определяли IgM к капсидному и IgG к раннему антигену ЭБВ, IgM к ЦМВ, IgG к ВГЧ 6 с коэффициентом позитивности. Параллельно исследовали варианты сочетания лабораторных маркеров активности герпес-вирусов и изменения основных лабораторных показателей, используемых в рутинной практике. Изучалась динамика наблюдаемых параметров на фоне комплексной противовирусной терапии в течение последующих после первичного обследования 3 месяцев. Определялась частота встречаемости различных симптомов и синдромов в зависимости от активности различных представителей семейства герпесов, их сочетания, возраста исследуемых.Результаты: ВГЧ7-моноинфекция более характерна для возрастной категории 7–14 лет. Значимость смешанной инфекции ВГЧ6+ВГЧ7 с возрастом уменьшалась. Сочетание активности 4 герпес-вирусов чаще регистрировалось в возрастных категориях 2–7 лет и 14–18 лет. Чаще у пациентов отмечались тонзиллит, генерализованная шейная лимфаденопатия, периодическое повышение температуры до субфебрильных или фебрильных цифр, фарингит, рекуррентные острые респираторные инфекции, ринит, что позволяет рассматривать их как основные проявления ВГЧ7-инфекции.Заключение: ВГЧ7-инфекция значима в современной патологии ребенка. Наличие таких синдромов, как рецидивирующий тонзиллит, шейная лимфаденопатия или изолированное увеличение подчелюстных лимфоузлов, периодическое повышение температуры, фарингит, рекуррентные острые респираторные инфекции в сочетании с гепато- и/или спленомегалией, усталостью, головной болью, часто ассоциировано с ВГЧ7. ВГЧ7-виремия регистрируется сравнительно редко, несмотря на очевидные клинико-лабораторные признаки воспаления, указывающие на наличие вирусного заболевания, что, повидимому, и является причиной гиподиагностики и отсутствия должного внимания к этой инфекции.

Comparative analysis of bacterial profiles in unstimulated and stimulated saliva samples

Background and objective: The microbial profiles of stimulated saliva samples have been shown to differentiate between patients with periodontitis, patients with dental caries, and orally healthy individuals. Saliva was stimulated to allow for easy and rapid collection; however, microbial composition may not reflect the more natural, unstimulated state. The purpose of this study was to validate whether stimulated saliva is an adequate surrogate for unstimulated saliva in determining salivary microbiomes. Design: Unstimulated (n=20) and stimulated (n=20) saliva samples were collected from 20 orally and systemically healthy, non-smoking participants. Salivary bacterial profiles were analyzed by means of the Human Oral Microbe Identification using Next Generation Sequencing (HOMINGS), and statistical analysis was performed using Mann–Whitney test with Benjamini–Hochberg's correction for multiple comparison, cluster analysis, principal component analysis, and correspondence analysis. Results: From a total of 40 saliva samples, 496 probe targets were identified with a mean number of targets per sample of 203 (range: 146–303), and a mean number of probe targets of 206 and 200 in unstimulated and stimulated saliva samples, respectively (p=0.62). Based on all statistical methods used for this study, the microbial profiles of unstimulated and stimulated saliva samples collected from the same person were not statistically significantly different. Conclusions: Analysis of bacterial salivary profiles in unstimulated and stimulated saliva samples collected from the same individual showed comparable results. Thus, the results verify that stimulated saliva is an adequate surrogate of unstimulated saliva for microbiome-related studies

Salivary bacterial fingerprints of established oral disease revealed by the Human Oral Microbe Identification using Next Generation Sequencing (HOMINGS) technique

Background and objective: The composition of the salivary microbiota, as determined using various molecular methods, has been reported to differentiate oral health from diseases. Thus, the purpose of this study was to utilize the newly developed molecular technique HOMINGS (Human Oral Microbe Identification using Next Generation Sequencing) for comparison of the salivary microbiota in patients with periodontitis, patients with dental caries, and orally healthy individuals. The hypothesis was that this method could add on to the existing knowledge on salivary bacterial profiles in oral health and disease. Design: Stimulated saliva samples (n=30) were collected from 10 patients with untreated periodontitis, 10 patients with untreated dental caries, and 10 orally healthy individuals. Salivary microbiota was analyzed using HOMINGS and statistical analysis was performed using Kruskal–Wallis test with Benjamini–Hochberg's correction. Results: From a total of 30 saliva samples, a mean number of probe targets of 205 (range 120–353) were identified, and a statistically significant higher mean number of targets was registered in samples from patients with periodontitis (mean 220, range 143–306) and dental caries (mean 221, range 165–353) as compared to orally healthy individuals (mean 174, range 120–260) (p=0.04 and p=0.04). Nine probe targets were identified with a different relative abundance between groups (p<0.05). Conclusions: Cross-sectional comparison of salivary bacterial profiles by means of HOMINGS analysis showed that different salivary bacterial profiles were associated with oral health and disease. Future large-scale prospective studies are needed to evaluate if saliva-based screening for disease-associated oral bacterial profiles may be used for identification of patients at risk of acquiring periodontitis and dental caries

Герпес-вирусы 6 и 7 типа: взгляд педиатра

Herpetic diseases are widespread among cancer patients and are of interest to specialists in various fields. It is herpetic infections that are often common among cases of infant mortality. The main causative agents of herpetic phenomena in humans are manifested by the 8 most studied types of the virus. The article presents a review of modern scientific literature on herpetic diseases caused by herpes viruses 6 and 7. The issues of the history of the study of diseases, etiology, features of the distribution, the pathogenesis of diseases, a wide manifestation of manifestations in childhood, diagnosis, and tactics of managing patients are outlined.Герпетические инфекции широко распространены в человеческой популяции, и интерес к ним среди специалистов разного профиля постоянно возрастает, так как они являются причиной развития большого спектра соматических и онкологических заболеваний. Именно герпетические инфекции часто являются причинами преждевременных родов, младенческой смертности. Основные возбудители герпетических инфекций у человека представлены 8 наиболее изученными типами вируса. В статье представлен обзор современной научной литературы по вопросам герпетических инфекций, вызванных герпес-вирусами 6-го и 7-го типа. Изложены вопросы истории изучения заболевания, этиология, особенности распространения, патогенез заболевания, основные клинические проявления в детском возрасте, диагностика и тактика ведения пациентов

Metaproteomics of saliva identifies human protein markers specific for individuals with periodontitis and dental caries compared to orally healthy controls

Background The composition of the salivary microbiota has been reported to differentiate between patients with periodontitis, dental caries and orally healthy individuals. To identify characteristics of diseased and healthy saliva we thus wanted to compare saliva metaproteomes from patients with periodontitis and dental caries to healthy individuals. Methods Stimulated saliva samples were collected from 10 patients with periodontitis, 10 patients with dental caries and 10 orally healthy individuals. The proteins in the saliva samples were subjected to denaturing buffer and digested enzymatically with LysC and trypsin. The resulting peptide mixtures were cleaned up by solid-phase extraction and separated online with 2 h gradients by nano-scale C18 reversed-phase chromatography connected to a mass spectrometer through an electrospray source. The eluting peptides were analyzed on a tandem mass spectrometer operated in data-dependent acquisition mode. Results We identified a total of 35,664 unique peptides from 4,161 different proteins, of which 1,946 and 2,090 were of bacterial and human origin, respectively. The human protein profiles displayed significant overexpression of the complement system and inflammatory markers in periodontitis and dental caries compared to healthy controls. Bacterial proteome profiles and functional annotation were very similar in health and disease. Conclusions Overexpression of proteins related to the complement system and inflammation seems to correlate with oral disease status. Similar bacterial proteomes in healthy and diseased individuals suggests that the salivary microbiota predominantly thrives in a planktonic state expressing no disease-associated characteristics of metabolic activity

High throughput DNA sequencing to detect differences in the subgingival plaque microbiome in elderly subjects with and without dementia

BACKGROUND: To investigate the potential association between oral health and cognitive function, a pilot study was conducted to evaluate high throughput DNA sequencing of the V3 region of the 16S ribosomal RNA gene for determining the relative abundance of bacterial taxa in subgingival plaque from older adults with or without dementia. METHODS: Subgingival plaque samples were obtained from ten individuals at least 70 years old who participated in a study to assess oral health and cognitive function. DNA was isolated from the samples and a gene segment from the V3 portion of the 16S bacterial ribosomal RNA gene was amplified and sequenced using an Illumina HiSeq1000 DNA sequencer. Bacterial populations found in the subgingival plaque were identified and assessed with respect to the cognitive status and oral health of the participants who provided the samples. RESULTS: More than two million high quality DNA sequences were obtained from each sample. Individuals differed greatly in the mix of phylotypes, but different sites from different subgingival depths in the same subject were usually similar. No consistent differences were observed in this small sample between subjects separated by levels of oral health, sex, or age; however a consistently higher level of Fusobacteriaceae and a generally lower level of Prevotellaceae was seen in subjects without dementia, although the difference did not reach statistical significance, possibly because of the small sample size. CONCLUSIONS: The results from this pilot study provide suggestive evidence that alterations in the subgingival microbiome are associated with changes in cognitive function, and provide support for an expanded analysis of the role of the oral microbiome in dementia

Changes and Oral Microbiome Shifts in HIV+ Patients Following Periodontal Therapy

The objective of this study was to compare clinical outcomes and oral microbiome changes following comprehensive dental care in HIV+ patients. Twenty-seven HIV+ patients received periodontal treatment, oral hygiene instructions, caries control, and extraction of hopeless teeth. Systemic (viral load, CD4 counts, ART regimen) and oral clinical parameters (presence of caries, gingival and plaque indices) were measured at baseline (BL), 12-months (12M), and 24-months (24M) after therapy. Differences between clinical and microbial parameters between BL, to 12M, or 24M were determined. Factors associated with changes in relative abundance of bacterial genera include smoking status, CD4 counts, and anti-retroviral mediation status. Several genera appear to be driving changes including Megasphaera, Kingella, Veillonella, and Prevotella. After one year of dental treatment, the oral microbiome shifts to a more HIV-negative, health-associated microbiome with increased alpha diversity of genera. This change in diversity plateaus in the second year of treatment in a well-maintained population.Master of Scienc