Expanding the Diversity of Accumulibacter with a Novel Type and Deciphering the Transcriptional and Morphological Features among Co-Occurring Strains.

"Candidatus Accumulibacter" is the major polyphosphate-accumulating organism (PAO) in global wastewater treatment systems, and its phylogenetic and functional diversity have expanded in recent years. In addition to the widely recognized type I and II sublineages, we discovered a novel type enriched in laboratory bioreactors. Core gene and machine learning-based gene feature profiling supported the assertion that type III "Ca. Accumulibacter" is a potential PAO with the unique function of using dimethyl sulfoxide as an electron acceptor. Based on the correlation between ppk1 and genome similarity, the species-level richness of Accumulibacter was estimated to be over 100, suggesting that the currently recognized species are only the tip of the iceberg. Meanwhile, the interstrain transcriptional and morphological features of multiple "Ca. Accumulibacter" strains co-occurring in a bioreactor were investigated. Metatranscriptomics of seven co-occurring strains indicated that the expression level and interphasic dynamics of PAO phenotype-related genes had minimal correlation with their phylogeny. In particular, the expression of denitrifying and polyphosphate (poly-P) metabolism genes exhibited higher interstrain and interphasic divergence than expression of glycogen and polyhydroxyalkanoate metabolic genes. A strategy of cloning rRNA genes from different strains based on similar genomic synteny was successfully applied to differentiate their morphology via fluorescence in situ hybridization. Our study further expands the phylogenetic and functional diversity of "Ca. Accumulibacter" and proposes that deciphering the function and capability of certain "Ca. Accumulibacter" should be tailored to the environment and population in question. IMPORTANCE In the last 2 decades, "Ca. Accumulibacter" has garnered significant attention as the core functional but uncultured taxon for enhanced biological phosphorus removal due to its phylogenetic and functional diversity and intragenus niche differentiation. Since 2002, it has been widely known that this genus has two sublineages (type I and II). However, in this study, a metagenomic approach led to the discovery of a novel type (type III) with proposed novel functional features. By comparing the average nucleotide identity of "Ca. Accumulibacter" genomes and the similarity of ppk1, a phylogenetic biomarker largely deposited in databases, the global species-level richness of "Ca. Accumulibacter" was estimated for the first time to be over 100. Furthermore, we observed the co-occurrence of multiple "Ca. Accumulibacter" strains in a single bioreactor and found the simultaneous transcriptional divergence of these strains intriguing with regard to their niche differentiation within a single community. Our results indicated a decoupling feature between transcriptional pattern and phylogeny for co-occurring strains.

Life without blood: Molecular and functional analysis of hirudins and hirudin-like factors of the Asian non-hematophagous leech Whitmania pigra.

BACKGROUND: Several leech species of the genera Hirudo, Hirudinaria, and Whitmania are widely used in traditional Chinese medicine (TCM) for the oral treatment of disorders associated with blood stasis. Among them, the non-hematophagous leech Whitmania pigra expresses a variety of components that have the potential to act on the vertebrate blood coagulation system. OBJECTIVE: Whether the thrombin inhibitor hirudin, probably the most prominent leech-derived anticoagulant, is actually present in Whitmania pigra, is still a matter of debate. To answer that open question was the aim of the study. METHODS: We identified several putative hirudin-encoding sequences in transcriptome data of Whitmania pigra. Upon gene synthesis and molecular cloning the respective recombinant proteins were expressed in Escherichia coli, purified, processed, and eventually functionally characterized for thrombin-inhibitory potencies in coagulation assays. RESULTS: We were successful in the identification and functional characterization of several putative hirudins in Whitmania pigra. Some, but not all, of these factors are indeed thrombin inhibitors. Whitmania pigra hence expresses both hirudins (factors that inhibit thrombin) and hirudin-like factors (that do not or only very weakly inhibit thrombin). Furthermore, we revealed the exon/intron structures of the corresponding genes. Coding sequences of some putative hirudins of Whitmania pigra were present also in transcriptome datasets of Hirudo nipponia, a hematophagous leech that is likewise used in TCM. CONCLUSIONS: Based on both structural and functional data we provide very strong evidence for the expression of hirudins in Whitmania pigra. This is the first description of hirudins in a non-hematophagous leech.

Genome-wide identification and expression analyses of the LEA protein gene family in tea plant reveal their involvement in seed development and abiotic stress responses.

Late embryogenesis abundant (LEA) proteins are widely known to be present in higher plants and are believed to play important functional roles in embryonic development and abiotic stress responses. However, there is a current lack of systematic analyses on the LEA protein gene family in tea plant. In this study, a total of 48 LEA genes were identified using Hidden Markov Model profiles in C. sinensis, and were classified into seven distinct groups based on their conserved domains and phylogenetic relationships. Genes in the CsLEA_2 group were found to be the most abundant. Gene expression analyses revealed that all the identified CsLEA genes were expressed in at least one tissue, and most had higher expression levels in the root or seed relative to other tested tissues. Nearly all the CsLEA genes were found to be involved in seed development, and thirty-nine might play an important role in tea seed maturation concurrent with dehydration. However, only sixteen CsLEA genes were involved in seed desiccation, and furthermore, most were suppressed. Additionally, forty-six CsLEA genes could be induced by at least one of the tested stress treatments, and they were especially sensitive to high temperature stress. Furthermore, it was found that eleven CsLEA genes were involved in tea plant in response to all tested abiotic stresses. Overall, this study provides new insights into the formation of CsLEA gene family members and improves our understanding on the potential roles of these genes in normal development processes and abiotic stress responses in tea plant, particularly during seed development and desiccation. These results are beneficial for future functional studies of CsLEA genes that will help preserve the recalcitrant tea seeds for a long time and genetically improve tea plant.