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Abstract Circoviruses belong to the family Circoviridae, which is classified into two genera, Circovirus and Cyclovirus. Some circoviruses have been identified in various organisms and/or their fecal samples and might be associated with diseases in their hosts. However, few circoviruses are reported in human. Recently, two novel circoviruses HCirV-1 and HCirV-2 were identified in humans through next-generation sequencing and were defined as the species of Circovirus human in the family Circoviridae. Both viruses are suspected to be associated with liver diseases, particularly among immunosuppressed people, showing potential health implication. Investigation on the prevalence of both viruses, and identification of vulnerable population are important and need a rapid, accurate, and specific assay for detecting and screening the two viruses. In this study, we developed a duplex HiFi-LAMP assay for simultaneous detection of HCirV-1 and HCirV-2. The assay exhibits high sensitivity with LOD of 64 and 49 copies per 25 µL reaction for HCirV-1 and HCirV-2, respectively. The duplex assay was demonstrated to have a rapid reaction time within 35 min. Clinical screening tests showed that neither HCirV-1 nor HCirV-2 were detected among 875 patients with infection of HBV, HIV-1 or other viruses. Large-scale screening of both viruses in diverse populations is encouraged to enhance our understanding of their relevance to various diseases.

ABSTRACT Candida albicans (C. albicans) and Lactiplantibacillus plantarum subsp. plantarum (L. plantarum) are frequently identified in various niches, but their dual-species interaction, especially with C. albicans in yeast form, remains unclear. This study aimed to investigate the dual-species interaction of L. plantarum and C. albicans, including proliferation, morphology, and transcriptomes examined by selective agar plate counting, microscopy, and polymicrobial RNA-seq, respectively. Maintaining a stable and unchanged growth rate, L. plantarum inhibited C. albicans yeast cell proliferation but not hyphal growth. Combining optical microscopy and atomic force microscopy, cell-to-cell direct contact and co-aggregation with L. plantarum cells surrounding C. albicans yeast cells were observed during dual-species interaction. Reduced C. albicans yeast cell proliferation in mixed culture was partially due to L. plantarum cell-free culture supernatant but not the acidic environment. Upon polymicrobial transcriptomics analysis, interesting changes were identified in both L. plantarum and C. albicans gene expression. First, two L. plantarum quorum-sensing systems showed contrary changes, with the activation of lamBDCA and repression of luxS. Second, the upregulation of stress response-related genes and downregulation of cell cycle, cell survival, and cell integrity-related pathways were identified in C. albicans, possibly connected to the stress posed by L. plantarum and the reduced yeast cell proliferation. Third, a large scale of pathogenesis and virulence factors were downregulated in C. albicans, indicating the potential interruption of pathogenic activities by L. plantarum. Fourth, partial metabolism and transport pathways were changed in L. plantarum and C. albicans. The information in this study might aid in understanding the behavior of L. plantarum and C. albicans in dual-species interaction.IMPORTANCEThe anti-Candida albicans activity of Lactiplantibacillus plantarum has been explored in the past decades. However, the importance of C. albicans yeast form and the effect of C. albicans on L. plantarum had also been omitted. In this study, the dual-species interaction of L. plantarum and C. albicans was investigated with a focus on the transcriptomes. Cell-to-cell direct contact and co-aggregation with L. plantarum cells surrounding C. albicans yeast cells were observed. Upon polymicrobial transcriptomics analysis, interesting changes were identified, including contrary changes in two L. plantarum quorum-sensing systems and reduced cell survival-related pathways and pathogenesis determinants in C. albicans.

Abstract Background Kluyveromyces marxianus is a promising cell factory for producing bioethanol and that raised a demand for a high yield of heterologous proteins in this species. Expressions of heterologous proteins usually lead to the accumulation of misfolded or unfolded proteins in the lumen of the endoplasmic reticulum (ER) and then cause ER stress. To cope with this problem, a group of ER stress response target genes (ESRTs) are induced, mainly through a signaling network called unfolded protein response (UPR). Characterization and modulation of ESRTs direct the optimization of heterologous expressions. However, ESRTs in K. marxianus have not been identified so far. Results In this study, we characterized the ER stress response in K. marxianus for the first time, by using two ER stress-inducing reagents, dithiothreitol (DTT) and tunicamycin (TM). Results showed that the Kar2–Ire1–Hac1 pathway of UPR is well conserved in K. marxianus. About 15% and 6% of genes were upregulated during treatment of DTT and TM, respectively. A total of 115 upregulated genes were characterized as ESRTs, among which 97 genes were identified as UPR target genes and 37 UPR target genes contained UPR elements in their promoters. Genes related to carbohydrate metabolic process and actin filament organization were identified as new types of UPR target genes. A total of 102 ESRTs were overexpressed separately in plasmids and their effects on productions of two different lignocellulolytic enzymes were systematically evaluated. Overexpressing genes involved in carbohydrate metabolism, including PDC1, PGK and VID28, overexpressing a chaperone gene CAJ1 or overexpressing a reductase gene MET13 substantially improved secretion expressions of heterologous proteins. Meanwhile, overexpressing a novel gene, KLMA_50479 (named ESR1), as well as overexpressing genes involved in ER-associated protein degradation (ERAD), including HRD3, USA1 andYET3, reduced the secretory expressions. ESR1 and the aforementioned ERAD genes were deleted from the genome. Resultant mutants, except the yet3Δ mutant, substantially improved secretions of three different heterologous proteins. During the fed-batch fermentation, extracellular activities of an endoxylanase and a glucanase in hrd3Δ cells improved by 43% and 28%, respectively, compared to those in wild-type cells. Conclusions Our results unveil the transcriptional scope of the ER stress response in K. marxianus and suggest efficient ways to improve productions of heterologous proteins by manipulating expressions of ESRTs.

ABSTRACT The complexity of genome-wide gene expression has not yet been adequately addressed due to a lack of comprehensive statistical analyses. In the present study, we introduce degree of freedom (DOF) as a summary statistic for evaluating gene expression complexity. Because DOF can be interpreted by a state-space representation, application of the DOF is highly useful for understanding gene activities. We used over 11,000 gene expression data sets to reveal that the DOF of gene expression in Saccharomyces cerevisiae is not greater than 450. We further demonstrated that various degrees of freedom of gene expression can be interpreted by different sequence motifs within promoter regions and Gene Ontology (GO) terms. The well-known TATA box is the most significant one among the identified motifs, while the GO term “ribosome genesis” is an associated biological process. On the basis of transcriptional freedom, our findings suggest that the regulation of gene expression can be modeled using only a few state variables. IMPORTANCE Yeast works like a well-organized factory. Each of its components works in its own way, while affecting the activities of others. The order of all activities is largely governed by the regulation of gene expression. In recent decades, biologists have recognized many regulations for yeast genes. However, it is not known how closely the regulation links each gene together to make all components of the cell work as a whole. In other words, biologists are very interested in how many independent control factors are needed to operate an artificial “cell” that works the same as a real one. In this work, we suggested that only 450 control factors were sufficient to represent the regulation of all 5800 yeast genes.

ObjectivesUmbilical cord blood transplantation (UCBT) is associated with a relatively high rate of engraftment failure. This study aimed at exploring whether any fecal microbiota could be associated with engraftment failure following UCBT in Crohn’s disease patients with IL10RA deficiency.MethodsThirteen patients were recruited and their 230 fecal samples were collected longitudinally from immediately before conditioning chemotherapy to 8 weeks post the UCBT. The V3-V4 regions of the bacterial 16S rRNA gene were amplified by PCR and sequenced, followed by bioinformatics analyses.ResultsFollowing the UCBT, 7 out of 13 patients achieved neutrophil and platelet engraftment with a median of 21 and 28 days, respectively (S group), while 6 patients failed to achieve engraftment (F group). In comparison with that in the S group, significantly lower Shannon diversity values on the UCBT day (P = 0.0176) and less abundance of Bifidobacterium longum, Bifidobacterium pseudolongum, Enterobacteriaceae_538000, and one taxon of Lachnospiraceae family was detected in the F group, accompanied by significantly higher abundances of four taxa including Lautropia, Pseudomonas, and species Microvirgula aerodenitrificans during the chemotherapy period as well as UCBT. The abundances of thirty OTUs were correlated significantly with clinical indices.ConclusionsMicrobial indicators of reduced diversity of microbiota and signatures of specific bacterial abundances, such as a lower abundance of Bifidobacterium longum, for engraftment failure would require validation. These indicators may help for the risk stratification in patients with IL10RA deficiency undergoing UCBT.

Abstract Background Cyclic vomiting syndrome (CVS) is characterized by repeated, stereotypical vomiting episodes. It is possibly associated with mitochondrial DNA (mtDNA) variants. We examined the phenotype, disease burden, treatment and performed mtDNA analysis in pediatric CVS. Methods This retrospective study included 42 children with CVS in a tertiary care center. Information regarding medical history, clinical features, laboratory tests, and treatment were collected. mtDNA sequencing was performed among 13 patients. Results Mean age of onset among patients was 4.0±3.4 years, and mean age at diagnosis was 6.7±4.2 years. CVS episodes in onset and features were stereotypic. Recognizable prodromes were reported in 54.8% patients. Neuroimaging showed previously unknown intracranial abnormalities. Gastrointestinal infection was found in four patients. Mean duration of hospitalization was 7.0±2.4 days, and mean hospitalization cost was 10,891 RMB. Sequencing showed that 4/13 patients had C16519T mtDNA polymorphism, and 2/13 patients had G3010A mtDNA polymorphism. Conclusions Cyclic vomiting syndrome is a disabling disorder, which causes huge disease burdens to the patients and their families. Early clinical suspicion and prompt diagnosis are crucial. mtDNA polymorphisms were found in some patients, but they were not significantly associated with pediatric CVS.