Browsing by Subject "16S rRNA gene sequencing"

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  • Hyytiainen, Heidi K.; Jayaprakash, Balamuralikrishna; Kirjavainen, Pirkka V.; Saari, Sampo E.; Holopainen, Rauno; Keskinen, Jorma; Hämeri, Kaarle; Hyvarinen, Anne; Boor, Brandon E.; Taubel, Martin (2018)
    Background: Floor dust is commonly used for microbial determinations in epidemiological studies to estimate early-life indoor microbial exposures. Resuspension of floor dust and its impact on infant microbial exposure is, however, little explored. The aim of our study was to investigate how floor dust resuspension induced by an infant's crawling motion and an adult walking affects infant inhalation exposure to microbes. Results: We conducted controlled chamber experiments with a simplified mechanical crawling infant robot and an adult volunteer walking over carpeted flooring. We applied bacterial 16S rRNA gene sequencing and quantitative PCR to monitor the infant breathing zone microbial content and compared that to the adult breathing zone and the carpet dust as the source. During crawling, fungal and bacterial levels were, on average, 8- to 21-fold higher in the infant breathing zone compared to measurements from the adult breathing zone. During walking experiments, the increase in microbial levels in the infant breathing zone was far less pronounced. The correlation in rank orders of microbial levels in the carpet dust and the corresponding infant breathing zone sample varied between different microbial groups but was mostly moderate. The relative abundance of bacterial taxa was characteristically distinct in carpet dust and infant and adult breathing zones during the infant crawling experiments. Bacterial diversity in carpet dust and the infant breathing zone did not correlate significantly. Conclusions: The microbiota in the infant breathing zone differ in absolute quantitative and compositional terms from that of the adult breathing zone and of floor dust. Crawling induces resuspension of floor dust from carpeted flooring, creating a concentrated and localized cloud of microbial content around the infant. Thus, the microbial exposure of infants following dust resuspension is difficult to predict based on common house dust or bulk air measurements. Improved approaches for the assessment of infant microbial exposure, such as sampling at the infant breathing zone level, are needed.
  • Sarhadi, Virinder; Lahti, Leo; Saberi, Farideh; Youssef, Omar; Kokkola, Arto; Karla, Tiina; Tikkanen, Milja; Rautelin, Hilpi; Puolakkainen, Pauli; Salehi, Rasoul; Knuutila, Sakari (2020)
    Background/Aim: Gut microbiota plays an important role in colorectal cancer (CRC) and its composition in CRC patients can be influenced by ethnicity and tumour genomics. Herein, the aim was to study the possible associations of ethnicity and gene mutations with the gut microbiota in CRC patients. Materials and Methods: Bacterial composition in stool samples of 83 CRC patients and 60 controls from Iran and Finland was studied by 16S rRNA gene sequencing. The association of gut microbiota composition with CRC, host mutations in KRAS, NRAS and TP53, and ethnicity analysed. Results: Beta diversity analysis indicated significant differences between the Iranian and Finnish gut microbiota composition, in both controls and patients' groups. The Iranian controls had higher abundance of Prevotella and lower abundance of Bacteroides compared to the Finnish controls, while the Finnish patients had higher abundance of Clostridium compared to Iranian patients. Abundance of Ruminococcus was higher in patients compared to the controls. Higher abundances of Herbaspirillum, Catenibacterium and lower abundances of Barnesiella were associated with mutations in NRAS, TP53, and RAS respectively. Conclusion: A possible link of host gene mutations with gut bacterial composition is suggested.
  • Pezzutto, Denise (Helsingin yliopisto, 2019)
    Antimicrobial resistance is an emerging concern at the global scale, threatening the effectiveness of antibiotics in treating bacterial infections. Among anthropogenically impacted environments, wastewater treatment plants have been indicated as possible reservoirs of antibiotic resistance genes, putative hotspots for their horizontal gene transfer, and a source of their dissemination to the environment. Generally, the abundance of antibiotic resistance genes is reduced during the wastewater treatment process. However, some genes were shown to be enriched in purified effluent water and dried sludge, which are then released to the environment, compared to influent water. Also, the taxonomy of the hosts carrying antibiotic resistance genes could change as a result of horizontal gene transfer events. The aim of this study was to analyse and compare the host range of a series of antibiotic resistance genes in influent water, effluent water and dried sludge collected from the Viikinmäki wastewater treatment plant in Helsinki, Finland, by applying Emulsion, Paired Isolation and Concatenation PCR (epicPCR). EpicPCR is a method that can link a gene of interest to the 16S rRNA gene from the genome of the host bacterium, without any cultivation step. The abundance of the hosts was also evaluated by sequencing the 16S rRNA gene from the whole bacterial community. In several cases, the target antibiotic resistance genes (blaIMP, blaNDM, ermB, ermF, sul1 and strB) were carried in effluent water and dried sludge by taxa that were not hosting them in influent water, suggesting that horizontal gene transfer events might have occurred during the treatment. All the examined genes were detected both in abundant and in rare taxa, including genera that also comprise pathogenic species, such as Arcobacter and Acinetobacter. Some of the detected hosts were not previously known to show resistant phenotypes, namely members of the family Methylophilaceae. These results corroborate the idea that wastewater treatment plants might be hotspots for the horizontal gene transfer of resistance determinants, and potentially disseminate antibiotic resistant pathogens to the environment. However, in order to ensure the accuracy of the results, the limits of epicPCR as a method need to be identified and addressed.
  • Riskumäki, Matilda; Tessas, Ioannis; Ottman, Noora; Suomalainen, Alina; Werner, Paulina; Karisola, Piia; Lauerma, Antti; Ruokolainen, Lasse; Karkman, Antti; Wisgrill, Lukas; Sinkko, Hanna; Lehtimäki, Jenni; Alenius, Harri; Fyhrquist, Nanna (2021)
  • Riskumäki, Matilda; Tessas, Ioannis; Ottman, Noora; Suomalainen, Alina; Werner, Paulina; Karisola, Piia; Lauerma, Antti; Ruokolainen, Lasse; Karkman, Antti; Wisgrill, Lukas; Sinkko, Hanna; Lehtimäki, Jenni; Alenius, Harri; Fyhrquist, Nanna (European Academy of Allergy and Clinical Immunology, 2021)
    Allergy 76: 4, 1280-1284
  • Lundgren, Sara N.; Madan, Juliette C.; Karagas, Margaret R.; Morrison, Hilary G.; Hoen, Anne G.; Christensen, Brock C. (2019)
    The process of breastfeeding exposes infants to bioactive substances including a diversity of bacteria from breast milk as well as maternal skin. Knowledge of the character of and variation in these microbial communities, as well as the factors that influence them, is limited. We aimed to identify profiles of breastfeeding-associated microbial communities and their association with maternal and infant factors. Bilateral milk samples were collected from women in the New Hampshire Birth Cohort Study at approximately 6 weeks postpartum without sterilization of the skin in order to capture the infant-relevant exposure. We sequenced the V4-V5 hypervariable region of the bacterial 16S rRNA gene in 155 human milk samples. We used unsupervised clustering (partitioning around medoids) to identify microbial profiles in milk samples, and multinomial logistic regression to test their relation with maternal and infant variables. Associations between alpha diversity and maternal and infant factors were tested with linear models. Four breastfeeding microbiome types (BMTs) were identified, which differed in alpha diversity and in Streptococcus, Staphylococcus, Acinetobacter, and Pseudomonas abundances. Higher maternal pre-pregnancy BMI was associated with increased odds of belonging to BMT1 [OR (95% CI) = 1.13 (1.02, 1.24)] or BMT3 [OR (95% CI) = 1.12 (1.01, 1.25)] compared to BMT2. Independently, increased gestational weight gain was related to reduced odds of membership in BMT1 [OR (95% CI) = 0.66 (0.44, 1.00) per 10 pounds]. Alpha diversity was positively associated with gestational weight gain and negatively associated with postpartum sample collection week. There were no statistically significant associations of breastfeeding microbiota with delivery mode. Our results indicate that the breastfeeding microbiome partitions into four profiles and that its composition and diversity is associated with measures of maternal weight.
  • Youssef, Omar; Lahti, Leo; Kokkola, Arto; Karla, Tiina; Tikkanen, Milja; Ehsan, Homa; Carpelan-Holmström, Monika Angela; Koskensalo, Selja; Böhling, Tom; Rautelin, Hilpi; Puolakkainen, Pauli; Knuutila, Sakari; Sarhadi, Virinder (2018)
    Background: Microbial ecosystems that inhabit the human gut form central component of our physiology and metabolism, regulating and modulating both health and disease. Changes or disturbances in the composition and activity of this gut microbiota can result in altered immunity, inflammation, and even cancer. Aim: To compare the composition and diversity of gut microbiota in stool samples from patient groups based on the site of neoplasm in the gastrointestinal tract (GIT) and to assess the possible contribution of the bacterial composition to tumorigenesis. Methods: We studied gut microbiota by16S RNA gene sequencing from stool DNA of 83 patients, who were diagnosed with different GIT neoplasms, and 13 healthy individuals. Results: As compared to healthy individuals, stools of patients with stomach neoplasms had elevated levels of Enterobacteriaceae, and those with rectal neoplasms had lower levels of Bifidobacteriaceae. Lower abundance of Lactobacillaceae was seen in patients with colon neoplasms. Abundance of Lactobacillaceae was higher in stools of GIT patients sampled after cancer treatment compared to samples collected before start of any treatment. In addition to site-specific differences, higher abundances of Ruminococcus, Subdoligranulum and lower abundances of Lachnoclostridium and Oscillibacter were observed in overall GIT neoplasms as compared to healthy controls Conclusion: Our study demonstrates that the alterations in gut microbiota vary according to the site of GIT neoplasm. The observed lower abundance of two common families, Lactobacillaceae and Bifidobacteriaceae, and the increased abundance of Enterobacteriaceae could provide indicators of compromised gut health and potentially facilitate GIT disease monitoring.
  • Karnola, Laura (Helsingin yliopisto, 2020)
    Antimicrobial resistance (AMR) is an emerging global health threat with the growing number of antibiotic-resistant bacteria (ARB) having the alarming potential to return humanity to the pre-antibiotic era. Intensive animal production is globally one of the biggest sectors using antibiotics. It has been studied that fertilizing fields with animal manure spreads antimicrobial resistance genes (ARGs) in natural environments. The aim of this study was to determine the host range of three ARGs tetM, strB and qacE∆1 in soil and manure samples collected from a Finnish swine farm. In addition, the microbial communities in the same soil and manure samples were studied and compared. Six different sample types were taken, four from soil and two from manure. Soil samples included unfertilized soil, fertilized soil, soil two weeks after fertilization and soil six weeks after fertilization. Manure samples were taken from fresh and stored manure. Host range analysis was done by using Emulsion, Paired Isolation and Concatenation PCR (epicPCR). EpicPCR enables to link a gene of interest to the 16S rRNA gene of the bacterium that carries the gene in its genome. Microbial communities in soil and manure were analyzed and compared with the traditional 16S rRNA gene sequencing. Host range analysis with epicPCR revealed various bacterial genera as carriers for studied ARGs. Fertilized soil had the highest number of genera carrying the ARGs. This indicates that land application with animal manure increases the ARG load in soil. Microbial communities were found significantly different in soil and manure according to the 16S rRNA gene sequences. The results of epicPCR indicate that epicPCR has also potential for solid samples such as soil and manure as according to publications it has been mainly used for different water samples e.g., wastewaters. As a method epicPCR still requires optimization if applied for these sample materials in the future. A clear reduction in the number of genera carrying the ARGs was observed in six weeks after fertilization. Therefore, fertilizing fields only before cropping season, instead of fertilizing the fields year-round, might be one solution for reducing the ARG dissemination in soil in countries with high antibiotic consumption.