Performance of a Pilot-Scale Continuous Flow Ozone-Based Hospital Wastewater Treatment System.

Antimicrobial resistance (AMR) is becoming a global concern. Recently, research has emerged to evaluate the human and environmental health implications of wastewater from medical facilities and to identify acceptable wastewater treatment methods. In this study, a disinfection wastewater treatment system using an ozone-based continuous flow system was installed in a general hospital located in Japan. The effectiveness of antimicrobial-resistant bacteria (ARB) and antimicrobials in mitigating the environmental impact of hospital wastewater was evaluated. Metagenomic analysis was conducted to characterize the microorganisms in the wastewater before and after treatment. The results demonstrated that ozone treatment enables effective inactivation of general gut bacteria, including Bacteroides, Prevotella, Escherichia coli, Klebsiella, DNA molecules, and ARGs, as well as antimicrobials. Azithromycin and doxycycline removal rates were >99% immediately after treatment, and levofloxacin and vancomycin removal rates remained between 90% and 97% for approximately one month. Clarithromycin was more readily removed than the other antimicrobials (81-91%), and no clear removal trend was observed for ampicillin. Our findings provide a better understanding of the environmental management of hospital wastewater and enhance the effectiveness of disinfection wastewater treatment systems at medical facilities for mitigating the discharge of pollutants into aquatic environments.

Early candidacy for differentiation into heterocysts in the filamentous cyanobacterium Anabaena sp. PCC 7120.

The filamentous cyanobacterium Anabaena sp. PCC 7120 fixes dinitrogen facultatively. Upon depletion of combined nitrogen, about 10% of vegetative cells within the filaments differentiate terminally into nitrogen-fixing cells. The heterocyst has been studied as a model system of prokaryotic cell differentiation, with major focus on signal transduction and pattern formation. The fate of heterocyst differentiation is determined at about the eighth hour of induction (point of no return), well before conspicuous morphological or metabolic changes occur. However, little is known about how the initial heterocysts are selected after the induction by nitrogen deprivation. To address this question, we followed the fate of every cells on agar plates after nitrogen deprivation with an interval of 4 h. About 10% of heterocysts were formed without prior division after the start of nitrogen deprivation. The intensity of fluorescence of GFP in the transformants of hetR-gfp increased markedly in the future heterocysts at the fourth hour with respect to other cells. We also noted that the growing filaments consisted of clusters of four consecutive cells that we call quartets. About 75% of initial heterocysts originated from either of the two outer cells of quartets at the start of nitrogen deprivation. These results suggest that the future heterocysts are loosely selected at early times after the start of nitrogen deprivation, before the commitment. Such early candidacy could be explained by different properties of the outer and inner cells of a quartet, but the molecular nature of candidacy remains to be uncovered.

The first symbiont-free genome sequence of marine red alga, Susabi-nori (Pyropia yezoensis).

Nori, a marine red alga, is one of the most profitable mariculture crops in the world. However, the biological properties of this macroalga are poorly understood at the molecular level. In this study, we determined the draft genome sequence of susabi-nori (Pyropia yezoensis) using next-generation sequencing platforms. For sequencing, thalli of P. yezoensis were washed to remove bacteria attached on the cell surface and enzymatically prepared as purified protoplasts. The assembled contig size of the P. yezoensis nuclear genome was approximately 43 megabases (Mb), which is an order of magnitude smaller than the previously estimated genome size. A total of 10,327 gene models were predicted and about 60% of the genes validated lack introns and the other genes have shorter introns compared to large-genome algae, which is consistent with the compact size of the P. yezoensis genome. A sequence homology search showed that 3,611 genes (35%) are functionally unknown and only 2,069 gene groups are in common with those of the unicellular red alga, Cyanidioschyzon merolae. As color trait determinants of red algae, light-harvesting genes involved in the phycobilisome were predicted from the P. yezoensis nuclear genome. In particular, we found a second homolog of phycobilisome-degradation gene, which is usually chloroplast-encoded, possibly providing a novel target for color fading of susabi-nori in aquaculture. These findings shed light on unexplained features of macroalgal genes and genomes, and suggest that the genome of P. yezoensis is a promising model genome of marine red algae.

Genomic structure of the cyanobacterium Synechocystis sp. PCC 6803 strain GT-S.

Synechocystis sp. PCC 6803 is the most popular cyanobacterial strain, serving as a standard in the research fields of photosynthesis, stress response, metabolism and so on. A glucose-tolerant (GT) derivative of this strain was used for genome sequencing at Kazusa DNA Research Institute in 1996, which established a hallmark in the study of cyanobacteria. However, apparent differences in sequences deviating from the database have been noticed among different strain stocks. For this reason, we analysed the genomic sequence of another GT strain (GT-S) by 454 and partial Sanger sequencing. We found 22 putative single nucleotide polymorphisms (SNPs) in comparison to the published sequence of the Kazusa strain. However, Sanger sequencing of 36 direct PCR products of the Kazusa strains stored in small aliquots resulted in their identity with the GT-S sequence at 21 of the 22 sites, excluding the possibility of their being SNPs. In addition, we were able to combine five split open reading frames present in the database sequence, and to remove the C-terminus of an ORF. Aside from these, two of the Insertion Sequence elements were not present in the GT-S strain. We have thus become able to provide an accurate genomic sequence of Synechocystis sp. PCC 6803 for future studies on this important cyanobacterial strain.

CyanoClust: comparative genome resources of cyanobacteria and plastids.

Cyanobacteria, which perform oxygen-evolving photosynthesis as do chloroplasts of plants and algae, are one of the best-studied prokaryotic phyla and one from which many representative genomes have been sequenced. Lack of a suitable comparative genomic database has been a problem in cyanobacterial genomics because many proteins involved in physiological functions such as photosynthesis and nitrogen fixation are not catalogued in commonly used databases, such as Clusters of Orthologous Proteins (COG). CyanoClust is a database of homolog groups in cyanobacteria and plastids that are produced by the program Gclust. We have developed a web-server system for the protein homology database featuring cyanobacteria and plastids. Database URL: http://cyanoclust.c.u-tokyo.ac.jp/.

Elucidating genome structure evolution by analysis of isoapostatic gene clusters using statistics of variance of gene distances.

Identifying genomic regions that descended from a common ancestor is important for understanding the function and evolution of genomes. In related genomes, clusters of homologous gene pairs serve as evidence for candidate homologous regions, which make up genomic core. Previous studies on the structural organization of bacterial genomes revealed that basic backbone of genomic core is interrupted by genomic islands. Here, we applied statistics using variance of distances as a measure to classify conserved genes within a set of genomes according to their "isoapostatic" relationship, which keeps nearly identical distances of genes. The results of variance statistics analysis of cyanobacterial genomes including Prochlorococcus, Synechococcus, and Anabaena indicated that the conserved genes are classified into several groups called "virtual linkage groups (VLGs)" according to their positional conservation of orthologs over the genomes analyzed. The VLGs were used to define mosaic domain structure of the genomic core. The current model of mosaic genomic domains can explain global evolution of the genomic core of cyanobacteria. It also visualizes islands of lateral gene transfer. The stability and the robustness of the variance statistics are discussed. This method will also be useful in deciphering the structural organization of genomes in other groups of bacteria.