Reliable differentiation of Meyerozyma guilliermondii from Meyerozyma caribbica by internal transcribed spacer restriction fingerprinting

BACKGROUND: Meyerozyma guilliermondii (anamorph Candida guilliermondii) and Meyerozyma caribbica (anamorph Candida fermentati) are closely related species of the genetically heterogenous M. guilliermondii complex. Conventional phenotypic methods frequently misidentify the species within this complex and also with other species of the Saccharomycotina CTG clade. Even the long-established sequencing of large subunit (LSU) rRNA gene remains ambiguous. We also faced similar problem during identification of yeast isolates of M. guilliermondii complex from indigenous bamboo shoot fermentation in North East India. There is a need for development of reliable and accurate identification methods for these closely related species because of their increasing importance as emerging infectious yeasts and associated biotechnological attributes. RESULTS: We targeted the highly variable internal transcribed spacer (ITS) region (ITS1-5.8S-ITS2) and identified seven restriction enzymes through in silico analysis for differentiating M. guilliermondii from M. caribbica. Fifty five isolates of M. guilliermondii complex which could not be delineated into species-specific taxonomic ranks by API 20 C AUX and LSU rRNA gene D1/D2 sequencing were subjected to ITS-restriction fragment length polymorphism (ITS-RFLP) analysis. TaqI ITS-RFLP distinctly differentiated the isolates into M. guilliermondii (47 isolates) and M. caribbica (08 isolates) with reproducible species-specific patterns similar to the in silico prediction. The reliability of this method was validated by ITS1-5.8S-ITS2 sequencing, mitochondrial DNA RFLP and electrophoretic karyotyping. CONCLUSIONS: We herein described a reliable ITS-RFLP method for distinct differentiation of frequently misidentified M. guilliermondii from M. caribbica. Even though in silico analysis differentiated other closely related species of M. guilliermondii complex from the above two species, it is yet to be confirmed by in vitro analysis using reference strains. This method can be used as a reliable tool for rapid and accurate identification of closely related species of M. guilliermondii complex and for differentiating emerging infectious yeasts of the Saccharomycotina CTG clade

High-Throughput Illumina MiSeq Amplicon Sequencing of Yeast Communities Associated With Indigenous Dairy Products From Republics of Benin and Niger

Traditional Wagashi cheese and fermented cow milk are among the most popular dairy products appreciated by people from Benin, Niger, and the neighboring region. These products are the main source of protein in the diet of the low-income population in the region. The fermented milk is prepared by spontaneous fermentation without back-slopping. Whereas, the leaf extract of Calotropis procera is used for curdling the milk to prepare the soft Wagashi cheese. The present study aims to provide in-depth analysis of yeast communities associated with these traditional milk products by high-throughput Illumina MiSeq amplicon sequencing of internal transcribed spacer (ITS) region of fungal rRNA genes. A total of 60 samples, 20 samples of fermented milk each from Benin and Niger, and 20 samples of Wagashi cheese from Benin were used for analysis. The metagenomic investigation revealed that Kluyveromyces marxianus, Saccharomyces cerevisiae, Candida parapsilosis, and Sagenomella keratitidis were the predominant yeast species present in the traditional milk products. Furthermore, we noticed a high presence of K. marxianus (61.1% relative abundance) in the Wagashi cheese and S. cerevisiae (28.4% relative abundance) in the fermented milk of Niger. The presence of potential pathogenic yeast C. parapsilosis and S. keratitidis in these African milk products calls for further investigation to assess their safety. The predominant yeast K. marxianus and S. cerevisiae, recognized with generally regarded as safe (GRAS) status, could be further selected as starter culture along with lactic acid bacteria for developing controlled fermentation processes with enhanced product quality and safety

Comparative analyses of the bacterial communities present in the spontaneously fermented milk products of Northeast India and West Africa

IntroductionSpontaneous fermentation of raw cow milk without backslopping is in practice worldwide as part of the traditional food culture, including “Doi” preparation in earthen pots in Northeast India, “Kindouri” of Niger and “Fanire” of Benin prepared in calabash vessels in West Africa. Very few reports are available about the differences in bacterial communities that evolved during the spontaneous mesophilic fermentation of cow milk in diverse geographical regions.MethodsIn this study, we used high throughput amplicon sequencing of bacterial 16S rRNA gene to investigate 44 samples of naturally fermented homemade milk products and compared the bacterial community structure of these foods, which are widely consumed in Northeast India and Western Africa.Results and discussionThe spontaneous milk fermentation shared the lactic acid bacteria, mainly belonging to Lactobacillaceae (Lactobacillus) and Streptococcaceae (Lactococcus) in these two geographically isolated regions. Indian samples showed a high bacterial diversity with the predominance of Acetobacteraceae (Gluconobacter and Acetobacter) and Leuconostoc, whereas Staphylococcaceae (Macrococcus) was abundant in the West African samples. However, the Wagashi cheese of Benin, prepared by curdling the milk with proteolytic leaf extract of Calotrophis procera followed by natural fermentation, contained Streptococcaceae (Streptococcus spp.) as the dominant bacteria. Our analysis also detected several potential pathogens, like Streptococcus infantarius an emerging infectious foodborne pathogen in Wagashi samples, an uncultured bacterium of Enterobacteriaceae in Kindouri and Fanire samples, and Clostridium spp. in the Doi samples of Northeast India. These findings will allow us to develop strategies to address the safety issues related to spontaneous milk fermentation and implement technological interventions for controlled milk fermentation by designing starter culture consortiums for the sustainable production of uniform quality products with desirable functional and organoleptic properties

Comparative analysis of the gut microbiota in centenarians and young adults shows a common signature across genotypically non-related populations

Gut microbiota is among the factors that may be involved in healthy aging. Broader and geographically spread studies on gut microbiota of centenarians can help in identifying a common signature of longevity. We identified an endogamous Indian population with high centenarian prevalence. Here, we compared the gut microbiota composition and fecal metabolites of a centenarians group (\u2dc100 years) with young people (25-45 years) of the region with the high centenarian prevalence and the nearby region of low centenarian prevalence to decipher microbial-related longevity signatures. Also, we compared our results with publicly available datasets of similar groups including 125 centenarians from three countries (Italy, Japan, China). Our comparative analysis resulted in higher biodiversity within Ruminococcaceae in centenarians, with respect to younger adults, irrespective of their nationality. We observed bacterial signatures that are common among extremely old people of different nationality. Comparative metabolites profiling identified the fecal metabolic signature of extreme aging in the Indian study population. Our analysis of the co-occurrence network and bimodal distribution of several taxa suggested the establishment of a pervasive change in the gut ecology during extreme aging. Our study might pave the way to develop gut microbiota based biomarkers for healthy aging

Discovery of buried historical structures in the Kaveri–Kollidam interfluve, southern India

The Srirangam interfluve, formed between the Kaveri and Kollidam Rivers in southern India, has been a religious and cultural human settlement for over 2000 years of recorded history. It contains thousands of ancient, yet in use, man-made structures. Owing to its geographic location, and episodic catastrophic floods from the Kaveri and Kollidam Rivers, the region has been inundated frequently leading to the burial of many smaller structures. Thematic maps of geomorphology, vegetation index, land use, digital elevation model, slope, etc., have been generated through multi-temporal remote sensing satellite images. A compilation of historic (stone inscriptions, temple documents) and literature data followed by detailed survey of the region using vertical electrical sounding (VES) and ground-penetrating radar (GPR) through 200 and 400 MHz shielded antennas has led to the discovery of several buried manmade structures at various depths ranging from 0.54 to 3.55 m below ground level (b.g.l.) that were known only in the literature, hearsay and folklore. The VES, GPR and field surveys delineated four major soil/sediment horizons below the ground surface. Sedimentological characteristics of a massive river sand bed found overlying a weathered zone suggest deposition during a major catastrophic flood. The occurrence of flood deposits burying historic structures especially adjoining the regional tectonic structure indicate episodically active climatic and tectonic processes that played a significant role in the burial of ancient-historic land surface and manmade structures that were under desolation after invasion and desecration by war-lords of yore. This study demonstrates the potential of combining traditional knowledge and modern technologies to gain insights to locate historic/archaeological sites and discover buried structures

Neolithic cultural sites and extreme climate related channel avulsion: Evidence from the Vaigai River Basin, southern India

We document evidence for the role of fluvial dynamics-stream channel avulsion and flooding, tectonics and climatic conditions on the occurrence, shifting and burial of ancient cultural sites in the Vaigai River Basin in southern India. Systematic regional mapping, geophysical investigations and archaeological excavations in the flood plains of the Vaigai River led to the unearthing of an ancient culture that thrived at least since 752 BCE (2435 +/- 34 BP). Evidence for early human activities include sites of ancient and now buried industries that were manufacturing steel, glass bead, dye, gold jewelry and earthenware, in addition to dwelling-habitation, sites of religious sacrificial ceremonies, and places of worship, and burial. These information together with the occurrences of microlithic age stone tools in the vicinity led to the proposition of continued habitation of the Vaigai River Basin until today. Radiometric dating shows oldest age of 2435 +/- 34 BP (752 BCE) obtained at 2.5 m depth in a pit excavated at Keezhadi, a location situated far from the present day channel course whereas the youngest age of 689 +/- 24 BP is obtained at Agaram, a location situated nearest to the present day channel course. The bone fragments were dated as 1115 +/- 39 BP (778 CE - 1016 CE) and 1229 +/- 30 BP (689 CE - 883 CE). The organic carbon/peat layers recovered between flood deposits at the upper reaches of the Vaigai River are dated as 830 +/- 30 BP and 1250 +/- 30 BP. The natural levee deposits show evidences of habitation-related disturbances in the internal sedimentary structure, texture and packing. In addition to occurrences of hearth etc., re-currences of overbank-channel deposits over natural levee-habitation surfaces indicate habitation-colonization over relatively highgrounds along river bank, followed by marooning/burial under overbank-channel deposit and abandon -ment of habitation site and/or shifting of habitat. The same sequence of events occurred again, showing two habitation surfaces that are buried under surface currently. Together with facies characteristics and other artifact occurrences, the ages obtained in our study indicate shifting of the Vaigai River from prior to 2435 BP and colonization of ancient population, which might have been affected by a catastrophic flood and stream avulsion at about 1250-830 BP. These resulted in the ancient habitation getting marooned, industries and other structural infrastructure destroyed, and were transported and deposited as interbeds in flood deposits. Following a reduction in the stream energy, the point bar-natural levee environmental setting paved way for the deposition of organic carbon layer at Agaram at about 690 BP. This was followed by lateral shifting of the channel from Agaram towards the present-day course