Adaptive functions of structural variants in human brain development.

Quantifying the structural variants (SVs) in nonhuman primates could provide a niche to clarify the genetic backgrounds underlying human-specific traits, but such resource is largely lacking. Here, we report an accurate SV map in a population of 562 rhesus macaques, verified by in-house benchmarks of eight macaque genomes with long-read sequencing and another one with genome assembly. This map indicates stronger selective constrains on inversions at regulatory regions, suggesting a strategy for prioritizing them with the most important functions. Accordingly, we identified 75 human-specific inversions and prioritized them. The top-ranked inversions have substantially shaped the human transcriptome, through their dual effects of reconfiguring the ancestral genomic architecture and introducing regional mutation hotspots at the inverted regions. As a proof of concept, we linked APCDD1, located on one of these inversions and down-regulated specifically in humans, to neuronal maturation and cognitive ability. We thus highlight inversions in shaping the human uniqueness in brain development.

Origin and evolution of pentanucleotide repeat expansions at the familial cortical myoclonic tremor with epilepsy type1 - SAMD12 locus.

Familial cortical myoclonic tremor with epilepsy type 1 (FCMTE1) is caused by (TTTTA)exp(TTTCA)exp repeat expansions in SAMD12, while pure (TTTTA)exp is polymorphic. Our investigation focused on the origin and evolution of pure (TTTTA)exp and (TTTTA)exp(TTTCA)exp at this locus. We observed a founder effect between them. The phylogenetic analysis suggested that the (TTTTA)exp(TTTCA)exp might be generated from pure (TTTTA)exp through infrequent transformation events. Long-read sequencing revealed somatic generation of (TTTTA)exp(TTTCA)exp from pure (TTTTA)exp, likely via long segment (TTTCA) repeats insertion. Our findings indicate close relationships between the non-pathogenic (TTTTA)exp and the pathogenic (TTTTA)exp(TTTCA)exp, with dynamic interconversions. This sheds light on the genesis of pathogenic repeat expansions from ancestral premutation alleles. Our results may guide future studies in detecting novel repeat expansion disorders and elucidating repeat expansion mutational processes, thereby enhancing our understanding of human genomic variation.

The diagnostic utility of nanopore targeted sequencing in suspected endophthalmitis.

PURPOSE: This paper aimed to assess the diagnostic utility of a newly developed gene-based technology-nanopore targeted sequencing (NTS) in suspected endophthalmitis patients. METHODS: This retrospective study included 43 patients (44 eyes) with suspected endophthalmitis. NTS was applied along with microbiological culture to detect unknown pathogens in intraocular fluid samples. The diagnostic utility of NTS was mainly evaluated from three aspects, including the positivity rate of bacterial/fungal presence, diagnostic turnaround time and the frequency of change in treatment based on etiology test results. Non-parametric, two-sided Wilcoxon rank sum test, the McNemar's test and the kappa statistic were used for statistical comparisons. RESULTS: NTS showed significant advantages over traditional culture in positivity rates and diagnostic time (P < 0.001, kappa = 0.082; Z = -5.805, P < 0. 001). As regards antibiotic strategy, 17 patients (39.53%) and 5 patients (11.63%) underwent medication change following NTS and culture results respectively (P < 0.001, kappa = 0.335). With reasonable use of antibiotic and surgical intervention, most patients responded favorably, judged by significantly improved visual acuity (Z = -4.249, P < 0.001). The mean duration of hospitalization was 8.49 ± 2.45 days (range, 1-16 days). CONCLUSION: The high efficiency feature of NTS in pathogen detection renders it a valuable supplementary to traditional culture. Additionally, it has facilitated patients' management for the early and precise diagnosis of endophthalmitis.

Species-Level Characterization of the Microbiome in Breast Tissues with Different Malignancy and Hormone-Receptor Statuses Using Nanopore Sequencing.

Unambiguous evidence indicates that microbes are closely linked to various human diseases, including cancer. Most prior work investigating the microbiome of breast tissue describes an association between compositional differences of microbial species in benign and malignant tissues, but few studies have examined the relative abundance of microbial communities within human breast tissue at the species level. In this work, a total of 44 breast tissue samples including benign and malignant tissues with adjacent normal breast tissue pairs were collected, and Oxford Nanopore long-read sequencing was employed to assess breast tissue microbial signatures. Nearly 900 bacterial species were detected from the four dominant phyla: Proteobacteria, Firmicutes, Actinobacteria and Bacteroidetes. The bacteria with the highest abundance in all breast tissues was Ralstonia pickettii, and its relative abundance increased with decreasing malignancy. We further examined the breast-tissue microbiome composition with different hormone-receptor statuses, and the relative abundance of the genus Pseudomonas increased most significantly in breast tissues. Our study provides a rationale for exploring microbiomes associated with breast carcinogenesis and cancer development. Further large-cohort investigation of the breast microbiome is necessary to characterize a microbial risk signature and develop potential microbial-based prevention therapies.

The application of nanopore targeted sequencing in the diagnosis and antimicrobial treatment guidance of bloodstream infection of febrile neutropenia patients with hematologic disease.

Traditional microbiological methodology has limited sensitivity, detection range, and turnaround times in diagnosis of bloodstream infection in Febrile Neutropenia (FN) patients. A more rapid and sensitive detection technology is urgently needed. Here we used the newly developed Nanapore targeted sequencing (NTS) to diagnose the pathogens in blood samples. The diagnostic performance (sensitivity, specificity and turnaround time) of NTS detection of 202 blood samples from FN patients with hematologic disease was evaluated in comparison to blood culture and nested Polymerase Chain Reaction (PCR) followed by sanger sequence. The impact of NTS results on antibiotic treatment modification, the effectivity and mortality of the patients under the guidance of NTS results were assessed. The data showed that NTS had clinical sensitivity of 92.11%, clinical specificity of 78.41% compared with the blood culture and PCR combination. Importantly, the turnaround time for NTS was <24 h for all specimens, and the pre-report time within 6 h in emergency cases was possible in clinical practice. Among 118 NTS positive patients, 98.3% patients' antibiotic regimens were guided according to NTS results. There was no significant difference in effectivity and mortality rate between Antibiotic regimen switched according to NTS group and Antibiotic regimen covering pathogens detected by NTS group. Therefore, NTS could yield a higher sensitivity, specificity and shorter turnaround time for broad-spectrum pathogens identification in blood samples detection compared with traditional tests. It's also a good guidance in clinical targeted antibiotic treatment for FN patients with hematologic disease, thereby emerging as a promising technology for detecting infectious disease.

3ß-Hydroxysteroid dehydrogenase expressed by gut microbes degrades testosterone and is linked to depression in males.

Testosterone deficiency can lead to depressive symptoms in humans; however, the causes of this deficiency are incompletely understood. Here, we isolated Mycobacterium neoaurum from the fecal samples of testosterone-deficient patients with depression and showed that this strain could degrade testosterone in vitro. Furthermore, gavaging rats with M. neoaurum reduced their serum and brain testosterone levels and induced depression-like behaviors. We identified the gene encoding 3ß-hydroxysteroid dehydrogenase (3ß-HSD) as the enzyme causing testosterone degradation. Introducing 3ß-HSD into Escherichia coli enhanced its ability to degrade testosterone. Gavaging rats with 3ß-HSD-producing E. coli reduced their serum and brain testosterone levels and caused depression-like behaviors. Finally, compared with 16.67% of participants without depression, 42.99% (46/107) of the fecal samples of patients with depression harbored 3ß-HSD, and 60.87% (28/46) of these fecal samples expressed 3ß-HSD. These results suggest that 3ß-HSD expressed by gut microbes may be associated with depressive symptoms due to testosterone degradation.

New Insights of Corynebacterium kroppenstedtii in Granulomatous Lobular Mastitis based on Nanopore Sequencing.

BACKGROUND: The etiology of granulomatous lobular mastitis (GLM) remains unknown. This study aimed to detect bacteria in GLM using Nanopore sequencing and identify the relationship between GLM and Corynebacterium kroppenstedtii. METHODS AND MATERIALS: The bacterial detection on fresh samples (including breast pus and tissue) of 50 GLM patients using nanopore sequencing and culture methods. The bacterial detection rate of participants with different stages were compared and analyzed. Formalin-fixed and paraffin-embedded (FFPE) tissues from 39 patients were performed on Gram staining to identify Gram-positive bacilli (GPB) within lipid vacuoles. Moreover, the clinicopathological characteristics of GLM patients in different bacterial subgroups were also conducted. RESULTS: In 50 GLM patients, the detection rate of bacteria was 78% using nanopore sequencing method, especially in the early stage of GLM (over 80%), which was significantly higher than that using culture methods (24%, p < 0.001). The dominant bacteria were Corynebacterium species (64%), especially for the Corynebacterium kroppenstedtii. The detection rate of C. kroppenstedtii in nanopore sequencing method (56%) was higher than that in culture methods (16%, p < 0.001). Gram staining positive of bacteria in 7 patients, and 5 of them were C. kroppenstedtii. Thirty-one patients (31/39, 79.5%) exhibited typical histological structure of cystic neutrophilic granulomatous mastitis (CNGM), and eighteen patients detected with C. kroppenstedtii. CONCLUSION: Nanopore sequencing showed rapid and accurate bacteria detection over culture method in GLM patients. GLM is not sterile inflammation and closely related to C. kroppenstedtii. CNGM was associated with Corynebacterium infection, especially for C. kroppenstedtii.

Microbiological diagnosis of endophthalmitis using nanopore targeted sequencing.

BACKGROUND: Microorganism identification is critical for the early diagnosis and management of infectious endophthalmitis, but traditional culture can yield false-negative results. Nanopore targeted sequencing (NTS) is a third-generation sequencing technique with multiple advantages. This study aimed to test aqueous humour or vitreous fluid samples from presumed cases of infectious endophthalmitis using NTS to evaluate the feasibility of NTS in diagnosing endophthalmitis, especially for culture-negative cases. METHODS: This prospective study enrolled patients who presented to the Department of Ophthalmology of Union Hospital (Wuhan, China) between June 2018 and December 2020. The samples were sent immediately for routine microbiology culture processing and NTS assay. RESULTS: NTS identified microorganisms in 17 of 18 cases (94.4%) (eight culture-positive cases, nine culture-negative cases, and one case unavailable for culture). There was a high-quality match between culture and NTS for culture-positive cases. In the eight culture-negative cases and the case unavailable for culture, NTS detected either bacteria, fungi, or a mixture of bacteria and fungi in the intraocular fluids. The average waiting times for the results of bacterial and fungal cultures were 48 and 72 h, respectively. The average time for the NTS results was 12 h. CONCLUSIONS: NTS appears to be a promising diagnostic platform for diagnosing infectious endophthalmitis, even for culture-negative cases.