Microbes from oil reservoirs shape petroleum composition through processes such as biodegradation or souring. Such processes are considered economically detrimental and might pose health and safety hazards. It is therefore crucial to understand the composition of a reservoir\'s microbial community and its metabolic capabilities. However, such analyses are hindered by difficulties in extracting DNA from such complex fluids as crude oil. Here, we present a novel DNA extraction method from oils with a wide American Petroleum Institute (API) gravity (density) range. We investigated the ability to extract cells from oils with different solvents and surfactants, the latter both nonionic and ionic. Furthermore, we evaluated three DNA extraction methods. Overall, the best DNA yields and the highest number of 16S rRNA reads were achieved with isooctane as a solvent, followed by an ionic surfactant treatment using sodium dodecyl sulfate and DNA extraction using the PowerSoil Pro Kit (Qiagen). The final method was then applied to various oils from oil reservoirs collected in aseptic conditions. Despite the expected low cell density of 101-103 cells/ml, the new method yielded reliable results, with average 16S rRNA sequencing reads in the order of 41431 (±8860) per sample. Thermophilic, halophilic, and anaerobic taxa, which are most likely to be indigenous to the oil reservoir, were found in all samples. API gravity and DNA yield, despite the sufficient DNA obtained, did not show a correlation.

DNA-based methods are reliable for a precise identification of species in processed products. In this study, we assessed five typical DNA extraction methods from multiple aspects. Full-length and mini-length DNA barcoding were performed to detect the species substitution and mislabeling of 305 processed fish products from the Chinese market covering six processed fish products. The salt extraction method that exhibited the best overall performance was applied. All samples were successfully extracted; however, only 19.3 % of samples could be amplified using the full-DNA barcode primer set, and 90.2 % of samples could be amplified using the newly designed mini-DNA barcode primer sets (401 and 320 bp). Overall, the molecular identification results revealed that 36.4 % (111/305) of the samples were inconsistent with the labels, with commercial fraud observed in all six types of processed fish products. The survey findings provide technical references for effective fish authentication monitoring, offering insights into the seafood safety in markets.

Environmental waters (EW) substantially lend to the transmission of Helicobacter pylori (Hp). But the increase in Hp infections and antimicrobial resistance is often attributed to socioeconomic status. The connection between socioeconomic status and Hp prevalence in EW is however yet to be investigated. This study aimed to assess the impacts of socioeconomic indices (SI: continent, world bank region (WBR), world bank income (WBI), WHO region, Socio-demographic Index (SDI quintile), Sustainable Development Index (SuDI), and Human Development Index (HDI)) on the prevalence of Hp in EW. Hp-EW data were fitted to a generalized linear mixed-effects model and SI-guided meta-regression models with a 1000-resampling test. The worldwide prevalence of Hp in EW was 21.76% [95% confidence interval [CI]: 10.29-40.29], which declined significantly from 59.52% [43.28-74.37] in 1990-99 to 19.36% [3.99-58.09] in 2010-19 and with increasing trend in 2020-22 (33.33%, 22.66-45.43). Hp prevalence in EW was highest in North America (45.12%, 17.07-76.66), then Europe (22.38%, 5.96-56.74), South America (22.09%, 13.76-33.49), Asia (2.98%, 0.02-85.17), and Africa (2.56%, 0.00-99.99). It was negligibly different among sampling settings, WBI, and WHO regions demonstrating highest prevalence in rural location [42.62%, 3.07-94.56], HIEs [32.82%, 13.19-61.10], and AMR [39.43%, 19.92-63.01], respectively. However, HDI, sample size, and microbiological method robustly predict Hp prevalence in EW justifying 26.08%, 21.15%, and 16.44% of the true difference, respectively. In conclusion, Hp is highly prevalence in EW across regional/socioeconomic strata and thus challenged the uses of socioeconomic status as surrogate for hygienic/sanitary practices in estimating Hp infection prevalence.

BACKGROUND: Although wild ungulate populations are heavily monitored throughout Europe, we understand little of how parasites affect population dynamics, and there is no systematic, long-term monitoring of parasite diversity and parasite loads. Such monitoring is in part hampered by a lack of time- and cost-effective assay methodologies with high sensitivity and good taxonomic resolution. DNA metabarcoding has been successfully used to characterize the parasitic nemabiome with high taxonomic resolution in a variety of wild and domestic hosts. However, in order to implement this technique in large-scale, potentially non-invasive monitoring of gastrointestinal parasitic nematodes (GIN), protocol optimization is required to maximize biodiversity detection, whilst maintaining time- and cost-effectiveness.
METHODS: Faecal samples were collected from a wild moose population and GIN communities were characterized and quantified using both parasitological techniques (egg and larva counting) and DNA metabarcoding of the ITS2 region of rDNA. Three different isolation methods were compared that differed in the volume of starting material and cell lysis method.
RESULTS: Similar nematode faunas were recovered from all samples using both parasitological and metabarcoding methods, and the approaches were largely congruent. However, metabarcoding assays showed better taxonomic resolution and slightly higher sensitivity than egg and larvae counts. The metabarcoding was not strictly quantitative, but the proportion of target nematode sequences recovered was correlated with the parasitologically determined parasite load. Species detection rates in the metabarcoding assays were maximized using a DNA isolation method that included mechanical cell disruption and maximized the starting material volume.
CONCLUSIONS: DNA metabarcoding is a promising technique for the non-invasive, large-scale monitoring of parasitic GINs in wild ungulate populations, owing to its high taxonomic resolution, increased assay sensitivity, and time- and cost-effectiveness. Although metabarcoding is not a strictly quantitative method, it may nonetheless be possible to create a management- and conservation-relevant index for the host parasite load from this data. To optimize the detection rates and time- and cost-effectiveness of metabarcoding assays, we recommend choosing a DNA isolation method that involves mechanical cell disruption and maximizes the starting material volume.

Helicobacter pylori (Hp) transmission dynamics via drinking water (DW) has a far much higher direct and indirect public health disease burden than previously thought. This study aimed to assess the global prevalence of Hp in DW, distributions across regions and socioeconomic indices (continent, world bank income, Human Development Index (HDI), Sustainable Development Index (SuDI), Socio-Demographic Index (SDI) quintile, and WHO regions). Hp-DW related data mined from five databases until 10/12/2022 according to PRISMA standard were quality-appraised and fitted to a generalized linear mixed-effects model. Sub-group analysis and meta-regression-modelling coupled with a 1000-permutation test (⁎) were conducted. The global prevalence of Hp in DW was 15.7% (95% confidence interval [CI]: 7.98-27.5), which varied significantly by sampling methods (Moore swabbing (61.0% [0.00-100.0]) vs. grab sampling (13.68%[6.99-25.04])) and detection technique (non-culture (21.35%[9.13-42.31]) vs. cultured-based methods (Psubgroup < 0.01)). The period 1990-99 had the highest prevalence (41.24% [0.02-99.97]). Regarding regional designations, Hp prevalence in DW was significantly different being highest in North America (61.82% [41.03-79.02]) by continents, AMR (42.66% [20.81-67.82]) by WHO group, high HDI (24.64% [10.98-46.43]) by HDI group and North America (61.90% [2.79-98.93]) by world bank region (Psubgroup < 0.01). Generally, sample preparation, SuDI grouping, and detection/confirmation techniques, have significant effects on the detection/prevalence of Hp in DW (Psubgroup < 0.01). Hp prevalence in DW was not significantly different among rural and urban DW (Psubgroup = 0.90), world bank income groups (Psubgroup = 0.15), and SDI quintiles (Psubgroup = 0.07). Among the predictors examined, only sample size (p < 0.1, R∗2(coefficient of determinant) = 15.29%), continent (p∗val = 0.04), HDI (p∗val = 0.02), HDI group (p∗val = 0.05), and microbiological methods (p < 0.1; R∗2=28.09 %) predicted Hp prevalence in DW robustly. In conclusion, Hp prevalence is still endemic in DW regardless of the regional designations/improve DW supplies.

Collecting and obtaining sufficient amount of airborne particles for multiple microbial component assessments can be challenging. A passive dust sampling device, the electrostatic dust fall collector (EDC) has been established for assessing airborne exposures including endotoxin and glucans. Recently, with advances in next-generation sequencing techniques, EDCs were used to collect microbial cells for DNA sequencing analysis to promote the study of airborne bacterial and fungal communities. However, low DNA yields have been problematic when employing passive sampling with EDC. To address this challenge, we attempted to increase the efficiency of extraction. We compared DNA extraction efficiency of bacterial components from EDCs captured on filters through filtration using five extraction techniques. By measuring the abundance, diversity and structure of bacterial communities using qPCR and amplicon sequencing targeting 16S rRNA genes, we found that two techniques outperformed the rest. Furthermore, we developed protocols to simultaneously extract both DNA and endotoxin from a single EDC cloth. Our technique promotes a high quality to price ratio and may be employed in large epidemiological studies addressing airborne bacterial exposure where a large number of samples is needed.

With the advent of new molecular diagnostic techniques, retrieving DNA from the formalin-fixed paraffin-embedded (FFPE) tissues has become an essential yet challenging step for efficient downstream processes. Owing to low quality and quantity of DNA retrieved from the FFPE sections, the process is often impractical and needs significant improvements. Here, we established an efficient method for the purification of DNA from FFPE specimens by optimizing incubation temperature, incubation time, and the concentration of a formalin scavenger tris(hydroxymethyl)aminomethane (Tris) for reverse-crosslinking. The optimized method, named \"Highly concentrated Tris-mediated DNA extraction\" (HiTE), yielded three times the DNA yield per tissue slice compared with a representative DNA extraction kit. Moreover, the use of HiTE-extracted DNA increased the yield of the sequencing library three times and accordingly yielded a log higher and more reproducible sequencing library compared with that obtained using the commonly used commercial kit. The sequencing library prepared from HiTE-extracted FFPE-DNA had longer inserts and produced reads that evenly covered the reference genome. Successful application of HiTE-extracted FFPE-DNA for whole-genome and targeted gene panel sequencing indicates its practical usability.

UNASSIGNED: DNA extraction is an important step of any molecular experiment. DNA could not be easily extracted from members of actinomycetes by the usual methods of lysis. Due to the low efficiency of the conventional DNA extraction methods, development of an effective technique for DNA extraction of actinobacteria in emergency cases seems to be necessary. Since most of the DNA extraction techniques and commercial kits are not efficient enough to extract DNA from actinobacteria, this study was conducted to improve an efficient method obtained from conventional one to extract DNA from this group of bacteria.
UNASSIGNED: DNA extraction was performed using five methods (an improved method, Invisorb Spin Plant Mini Kit, EZ-10 Spin Column, Sarrbrucken method (HZI, Germany) and Kirby Bauer\'s method). To evaluate the quantity and quality of extracted genomic DNA, UV absorbance of all samples and efficiency of polymerase chain reaction (PCR) were evaluated.
UNASSIGNED: Overall, the results revealed that the highest quantity (up to 4000 ng/μl) and good quality of DNA was obtained using introduced DNA extraction method.
UNASSIGNED: Results indicated that recently introduced improved method was more efficient for extraction of DNA from actinobacteria for DDH (DNA-DNA hybridization) test and for those require the high concentration of DNA.

BACKGROUND: Liquid biopsy is gaining increasing popularity in cancer screening and diagnosis. However, there is no relatively mature DNA isolation method or commercial kit available that is compatible with different LB sample types. This study developed a PAN-sample DNA isolation method (PAN method) for liquid biopsy samples.
METHODS: The PAN method has two key steps, including biosample-specific pretreatments for various LB sample types and high concentration guanidine thiocyanate buffer for lysis and denaturation procedure. Subsequently, the performance of PAN method was validated by a series of molecular analyses.
RESULTS: The PAN method was used to isolate DNA from multiple sample types related to LB, including plasma, serum, saliva, nasopharyngeal swab, and stool. All purified DNA products showed good quality and high quantity. Comparison of KRAS mutation analysis using DNA purified using PAN method versus QIAamp methods showed similar efficiency. Epstein-Barr virus DNA was detected via Q-PCR using DNA purified from serum, plasma, nasopharyngeal swab, and saliva samples collected from nasopharyngeal carcinoma patients. Similarly, methylation sequencing of swab and saliva samples revealed good coverage of target region and high methylation of HLA-DPB1 gene. Finally, 16S rDNA gene sequencing of saliva, swab, and stool samples successfully defines the relative abundance of microbial communities.
CONCLUSIONS: This study developed and validated a PAN-sample DNA isolation method that can be used for different LB samples, which can be applied to molecular epidemiological research and other areas.

DNA studies of endangered or extinct species often rely on ancient or degraded remains. The majority of ancient DNA (aDNA) extraction protocols focus on skeletal elements, with skin and hair samples rarely explored. Similar to that found in bones and teeth, DNA extracted from historical or ancient skin and fur samples is also extremely fragmented with low endogenous content due to natural degradation processes. Thus, the development of effective DNA extraction methods is required for these materials. Here, we compared the performance of two DNA extraction protocols (commercial and custom laboratory aDNA methods) on hair and skin samples from decades-old museum specimens to Iron Age archaeological material. We found that apart from the impact sample-specific taphonomic and handling history has on the quantity and quality of DNA preservation, skin yielded more endogenous DNA than hair of the samples and protocols tested. While both methods recovered DNA from ancient soft tissue, the laboratory method performed better overall in terms of DNA yield and quality, which was primarily due to the poorer performance of the commercial binding buffer in recovering aDNA.
濒危或已灭绝动物的DNA研究通常依据古代或已严重降解的标本遗存。比较成熟的古DNA（ancient DNA，aDNA）提取方法主要针对于骨骼样本，对于皮张和毛发的研究较为少见。与古代的骨骼和牙齿情况相似，陈旧皮张样品中的DNA也由于自然降解等情况的存在，使得获取到的DNA片段长度极短、内源DNA含量极低。因此，针对这些样品，需要开发有效的DNA提取方法。本研究以几十年前到铁器时代的动物皮张和毛发样品为材料，比较2种提取方法（试剂盒法和古DNA实验室法）及相互组合的2种方法的DNA提取效果。研究发现除样品本身的差别（如年代等），皮张样品比毛发样品保留更多的内源DNA。所有方法都能够从陈旧的皮张样品中获取内源DNA，但是实验室的方法在DNA产量和质量上整体优于其它方法，实验室方法在纯化上的表现优于试剂盒方法。.