Investigation of the Relationship Between Akkermansia Genomic Diversity in Gut Microbiota and Parkinson’s Disease Dementia

Parkinson hastalığında (PH), genellikle sağlıkla ilişkilendirilen bir bakteri cinsi olan Akkermansia’nın bağırsak mikrobiyotasında artış gösterdiği bilinse de bu artışın nedeni tam olarak anlaşılamamıştır. Bu çalışmada Türkiye’deki PH hastalarında, bağırsak mikrobiyotasındaki muhtemel Akkermansia değişimlerinin belirlenmesi amaçlanmıştır. Bu amaçla, ilk kez shotgun metagenomik ve Akkermansia cinsine özgül bir yeni nesil dizileme (NGS) tekniği kullanılarak PH’de bilişsel bozukluk evreleriyle ilişkili olabilecek belirli Akkermansia suşlarının varlığı ve bu suşlarda bulunan potansiyel genler incelenmiştir. Bu kapsamda Türkiye’de toplanmış dört bağırsak mikrobiyotası örneği -üç demanslı PH (PHD) ve bir bilişsel bozukluğu olmayan sağlıklı kontrol (SK)- shotgun metagenomik dizileme yoluyla analiz edilmiş ve örneklerdeki Akkermansia cinsine ait genomlar yeniden inşa edilmiştir. Bu genomlar, veri tabanlarındaki Akkermansia cinsine ait genomlarla bir araya getirilerek özel bir veri tabanı oluşturulmuş ve Akkermansia cinsine özgül NGS uyumlu primerler bu veri tabanı kullanılarak tasarlanmıştır. Hedef gen bölgesinin çoğaltılması ve cins özgül yeni nesil dizileme için kütüphane hazırlama basamaklarının optimize edilmesinden sonra, 64 PH hastası [32 PHD ve 32 hafif bilişsel bozukluk gösteren PH (PH-MCI)] ile 26 SK’ye ait bağırsak mikrobiyotası örnekleri cins özgül amplikon dizileme ile analiz edilmiştir. Analizler sonucunda, bağırsak mikrobiyotası örneklerinde Akkermansia muciniphila türüne ait oldukları belirlenen yedi suşun varlığı tespit edilmiş ve iki suşun demanslı (PHD) ve demansı olmayan (PH-MCI, HC) gruplar arasındaki dağılımının anlamlı farklılık gösterdiği (p< 0.05) belirlenmiştir. Tespit edilen suşlara ait genomların gen içerikleri, karşılaştırmalı genomik analizler yoluyla incelediğinde yalnızca dağılımı demanslı ve demansı olmayan gruplar arasında anlamlı farklılık gösteren iki suşta bulunan 12 genin varlığı tahmin edilmiştir. Bu genlerin annotasyonları yapıldığında ise daha önce rapor edilmemiş ve işlevi bilinmeyen genler oldukları görülmüştür. Bu çalışmada, ilk kez Türkiye’de toplanmış PH hastalarına ait bağırsak mikrobiyotası örneklerinin shotgun metagenomik analizleri gerçekleştirilmiş, özel olarak Akkermansia cinsinin analizi için cins-özgül bir amplikon dizileme yöntemi geliştirilmiş ve bu yöntem kullanılarak PH’de bilişsel bozukluk evreleriyle ile ilişkili olabilecek Akkermansia suşları ve genleri tespit edilmiştir. Elde edilen sonuçlar, tür ya da suş düzeyindeki farklılıkların araştırılmasının, bağırsak mikrobiyotasındaki PH ile ilişkili değişimlerin daha iyi anlaşılmasına yardımcı olabileceğine işaret etmektedir.Although it is known that the relative abundance of Akkermansia, a bacterial genus commonly associated with health, increases in the gut microbiota of Parkinson’s disease (PD) patients, the exact reason for this increase remains unclear. This study was aimed to identify potential changes in Akkermansia within the gut microbiota of PD patients in Türkiye. For this purpose, shotgun metagenomics and a novel Akkermansia genus-specific amplicon sequencing technique was used to investigate the presence of specific Akkermansia strains associated with cognitive impairment (CI) stages in PD and to examine potential genes within these strains. In this context, four gut microbiota samples from Türkiye -three PD with dementia (PDD) and one healthy control without CI (HC)- were analyzed by shotgun metagenomics and metagenome-assembled genomes assigned to Akkermansia genus were reconstructed. Then, a custom database was created by combining these genomes with the Akkermansia genomes in public databases and next generation sequencing (NGS) compatible primers specific to the genus Akkermansia were designed using this database. After optimization of amplification and library preparation steps for genus-specific next generation sequencing, gut microbiota samples from 64 PD patients [32 PDD and 32 PD with mild CI (PD-MCI)] and 26 HCs were analyzed by genus-specific amplicon sequencing. The results revealed the presence of seven strains assigned to Akkermansia muciniphila in gut microbiota samples, two of which showed significant distribution differences (p< 0.05) between demented (PDD) and non-demented groups (PD-MCI, HC). When gene contents of the detected Akkermansia genomes were examined through comparative genomic analysis, the presence of 12 genes only in Akkermansia genomes specific to non-demented groups were predicted. The annotations of these genes showed that they were not reported before with unknown functions. In this study, for the first time, gut microbiota samples from PD patients in Türkiye were analyzed using shotgun metagenomics, a novel genus-specific amplicon sequencing method was developed specifically for the analysis of Akkermansia genus, and then Akkermansia strains and genes potentially associated with CI stages in PD were identified using this method. The results underscore that investigating the species or strain level differences could help better understanding of the changes associated with PD in the human gut microbiota

Metaproteogenomic analysis of saliva samples from Parkinson's disease patients with cognitive impairment

Cognitive impairment (CI) is very common in patients with Parkinson's Disease (PD) and progressively develops on a spectrum from mild cognitive impairment (PD-MCI) to full dementia (PDD). Identification of PD patients at risk of developing cognitive decline, therefore, is unmet need in the clinic to manage the disease. Previous studies reported that oral microbiota of PD patients was altered even at early stages and poor oral hygiene is associated with dementia. However, data from single modalities are often unable to explain complex chronic diseases in the brain and cannot reliably predict the risk of disease progression. Here, we performed integrative metaproteogenomic characterization of salivary microbiota and tested the hypothesis that biological molecules of saliva and saliva microbiota dynamically shift in association with the progression of cognitive decline and harbor discriminatory key signatures across the spectrum of CI in PD. We recruited a cohort of 115 participants in a multi-center study and employed multi-omics factor analysis (MOFA) to integrate amplicon sequencing and metaproteomic analysis to identify signature taxa and proteins in saliva. Our baseline analyses revealed contrasting interplay between the genus Neisseria and Lactobacillus and Ligilactobacillus genera across the spectrum of CI. The group specific signature profiles enabled us to identify bacterial genera and protein groups associated with CI stages in PD. Our study describes compositional dynamics of saliva across the spectrum of CI in PD and paves the way for developing non-invasive biomarker strategies to predict the risk of CI progression in PD.FEMS Research and Training Gran

The relationship of memory and microbiota in parkinson's dementia

Parkinson hastalığı (PH) her 1000 kişide 1-2 görülen en yaygın 2. Nörodejeneratif hastalıktır. Non-motor semptomlar hastaların %80'den fazlasını etkilemesiyle hastalığın yaşam kalitesinde motor semptomlara kıyasla daha büyük önem taşımaktadır. Özellikle bilişsel bozukluk, hasta ve hasta yakınlarına etkisin büyük olması ve erken mortalitede risk faktörü olması sebebiyle kritiktir. PH'de mikrobiyata profilini geniş çerçevede gösteren çalışmalar kısıtlıdır, mikrobiyotanın bellek performansı üzerine ilişkisini inceleyen araştırma ise henüz bulunmamaktadır. Bu tez çalışmasında Parkinsonyen demans ve hafif bilişsel bozuklukta mikrobiyotada izlenen özelliklerin bellek ile ilişkisinin araştırılması amaçlandı. Çalışmamıza 111 katılımcı dahil edildi, bunların 46'sı Parkinsonyen Demans, 41'i Parkinsonyel Hafif Bilişsel Bozukluk ve 24'ü sağlıklı kontrollerden oluşmaktaydı. Katılımcılardan fekal örnek alınarak -80 °C'de analize kadar bekletildi. Tüm katılımcıların bellek değerlendirmesinde verbal bellek testi olan Sözel Bellek Süreçleri Testi (SBST) uygulandı. Klinik değerlendirmeler CDR, motor değerlendirmeler ise HYE ve BPHDÖ tesleri ile yapıldı. Tüm klinik testler gruplar arası anlamlı farklı bulundu. Fekal mikrobiyota analizinde birçok bakteri her seviyede ayrışma gösterdi. Filum seviyesinde Actinobacteria özellikle demans grubunda artmıştı. Isı tablolarında Ruminococcaceae ailesine ait birçok ASV SBST kendiliğinden geri getirme puanı ile pozitif korele bulunurken, Lachnospiraceae bakteri ailesine ait birçok ASV SBST alt testleri ve MMSE testi ile, özellikle SBST toplam hatırlama puanı ile güçlü pozitif korelasyon gösterdi. Bacteroidaceae ve Enterobacteriaceae ailelerine ait ASV'ler ise tüm bu testlerle negatif ilişki içerisinde bulundu. Yapılan birçok analizde Feacalibacterium, Lachnoclostridium, Anaerostipes ve Agathobacter gibi birçoğu SCFA üreten bazı bakteriler birden fazla analizde bellek ile ilişkileri ortak olarak anlamlı bulunmuştur. Sonuç olarak, bu bulgular PH'de mikrobiyotanın belleğe etki edebileceğini düşündürmüştür.Parkinson's disease (PD) is the second most common neurodegenerative disease, occurring 1-2 in every 1000 people. Since non-motor symptoms affect more than 80% of patients, they are of greater importance in the quality of life of the disease compared to motor symptoms. Cognitive impairment is especially critical as it has a large impact on patients and their relatives and is a risk factor for early mortality. Studies showing the microbiota profile in PD in a broad framework are limited, and there is no research examining the relationship of microbiota on memory performance yet. In this thesis, it was aimed to investigate the relationship between memory and microbiota features in Parkinson's dementia and mild cognitive impairment. Our study included 111 participants, of whom 46 were Parkinson's Dementia, 41 Parkinson's Mild Cognitive Disorder, and 24 healthy controls. Fecal samples were taken from the participants and kept at -80 °C until analysis. Verbal Memory Processes Test (SBST), which is a verbal memory test, was used in the memory evaluation of all participants. Clinical evaluations were made with CDR, motor evaluations were made with HYE and BPHDS tests. All clinical tests were found to be significantly different between groups. In the fecal microbiota analysis, many bacteria showed degradation at all levels. Actinobacteria was increased at the phylum level, especially in the dementia group. While many ASV belonging to the Ruminococcaceae family were positively correlated with the SBST spontaneous retrieval scores in the temperature tables, many ASVs belonging to the Lachnospiraceae family of bacteria showed a strong positive correlation with the SBST subtests and the MMSE test, especially with the SBST total recall score. ASVs belonging to the Bacteroidaceae and Enterobacteriaceae families were negatively correlated with all these tests. In many analyzes, some bacteria, most of which produce SCFA, such as Feacalibacterium, Lachnoclostridium, Anaerostipes and Agathobacter, were found to be associated with memory in more than one analysis. In conclusion, these findings suggested that the microbiota may affect memory in PD

Axillary microbiota is associated with cognitive impairment in parkinson's disease patients

Cognitive impairment (CI) is among the most common non-motor symptoms of Parkinson's disease (PD), with a substantially negative impact on patient management and outcome. The development and progression of CI exhibits high interindividual variability, which requires better diagnostic and monitoring strategies. PD patients often display sweating disorders resulting from autonomic dysfunction, which has been associated with CI. Because the axillary microbiota is known to change with humidity level and sweat composition, we hypothesized that the axillary microbiota of PD patients shifts in association with CI progression, and thus can be used as a proxy for classification of CI stages in PD. We compared the axillary microbiota compositions of 103 PD patients (55 PD patients with dementia [PDD] and 48 PD patients with mild cognitive impairment [PD-MCI]) and 26 cognitively normal healthy controls (HC). We found that axillary microbiota profiles differentiate HC, PD-MCI, and PDD groups based on differential ranking analysis, and detected an increasing trend in the log ratio of Corynebacterium to Anaerococcus in progression from HC to PDD. In addition, phylogenetic factorization revealed that the depletion of the Anaerococcus, Peptoniphilus, and W5053 genera is associated with PD-MCI and PDD. Moreover, functional predictions suggested significant increases in myo-inositol degradation, ergothioneine biosynthesis, propionate biosynthesis, menaquinone biosynthesis, and the proportion of aerobic bacteria and biofilm formation capacity, in parallel to increasing CI. Our results suggest that alterations in axillary microbiota are associated with CI in PD. Thus, axillary microbiota has the potential to be exploited as a noninvasive tool in the development of novel strategies.Suleyman Yildirim from the Scientific and Technological Research Council of Turkey (TUBITAK

Metaproteogenomic analysis of saliva samples from Parkinson's disease patients with cognitive impairment

Cognitive impairment (CI) is very common in patients with Parkinson's Disease (PD) and progressively develops on a spectrum from mild cognitive impairment (PD-MCI) to full dementia (PDD). Identification of PD patients at risk of developing cognitive decline, therefore, is unmet need in the clinic to manage the disease. Previous studies reported that oral microbiota of PD patients was altered even at early stages and poor oral hygiene is associated with dementia. However, data from single modalities are often unable to explain complex chronic diseases in the brain and cannot reliably predict the risk of disease progression. Here, we performed integrative metaproteogenomic characterization of salivary microbiota and tested the hypothesis that biological molecules of saliva and saliva microbiota dynamically shift in association with the progression of cognitive decline and harbor discriminatory key signatures across the spectrum of CI in PD. We recruited a cohort of 115 participants in a multi-center study and employed multi-omics factor analysis (MOFA) to integrate amplicon sequencing and metaproteomic analysis to identify signature taxa and proteins in saliva. Our baseline analyses revealed contrasting interplay between the genus Neisseria and Lactobacillus and Ligilactobacillus genera across the spectrum of CI. The group specific signature profiles enabled us to identify bacterial genera and protein groups associated with CI stages in PD. Our study describes compositional dynamics of saliva across the spectrum of CI in PD and paves the way for developing non-invasive biomarker strategies to predict the risk of CI progression in PD