Stereochemical aspects of the metabolism of the isomeric methylcyclohexanols and methylcyclohexanones

1. The seven isomeric optically inactive forms of methylcyclohexanol (i.e. 1-, and cis- and trans-2-, -3- and -4-) are excreted by rabbits mainly as glucuronides of the thermodynamically more stable forms of the alcohols. The eighth isomer, cyclohexylmethanol, however, undergoes aromatization in vivo, giving rise to benzoic acid and hippuric acid. The (±)-2-, (±-3- and 4-methylcyclohexanones are reduced in the rabbit and excreted mainly as the glucuronides of the thermodynamically more stable forms of the corresponding methylcyclohexanols. 2. Racemic cis- and trans-2-methylcyclohexanol and 2-methylcyclohexanone are all excreted as conjugated trans-2-methylcyclohexanol. However, when the (±)-cis-alcohol or the (±)-ketone is fed, (+)-trans-2-methylcyclohexanol is excreted, whereas when the (±)-trans-alcohol is fed it is excreted as the (±)-trans-alcohol. 3. Racemic cis- and trans-3-methylcyclohexanol and 3-methylcyclohexanone are all excreted as conjugated racemic cis-3-methylcyclohexanol. cis- and trans-4-Methylcyclohexanol and 4-methylcyclohexanone are all excreted as conjugated trans-4-methylcyclohexanol. 4. The metabolic differences between the various methylcyclohexanols are explicable in terms of their conformations and of Vennesland's (1958) hypothesis of the role of NADH in dehydrogenation reactions.     

Mutations in the Gene Encoding the Ancillary Pilin Subunit of the Streptococcus suis srtF Cluster Result in Pili Formed by the Major Subunit Only

Pili have been shown to contribute to the virulence of different Gram-positive pathogenic species. Among other critical steps of bacterial pathogenesis, these structures participate in adherence to host cells, colonization and systemic virulence. Recently, the presence of at least four discrete gene clusters encoding putative pili has been revealed in the major swine pathogen and emerging zoonotic agent Streptococcus suis. However, pili production by this species has not yet been demonstrated. In this study, we investigated the functionality of one of these pili clusters, known as the srtF pilus cluster, by the construction of mutant strains for each of the four genes of the cluster as well as by the generation of antibodies against the putative pilin subunits. Results revealed that the S. suis serotype 2 strain P1/7, as well as several other highly virulent invasive S. suis serotype 2 isolates express pili from this cluster. However, in most cases tested, and as a result of nonsense mutations at the 5′ end of the gene encoding the minor pilin subunit (a putative adhesin), pili were formed by the major pilin subunit only. We then evaluated the role these pili play in S. suis virulence. Abolishment of the expression of srtF cluster-encoded pili did not result in impaired interactions of S. suis with porcine brain microvascular endothelial cells. Furthermore, non-piliated mutants were as virulent as the wild type strain when evaluated in a murine model of S. suis sepsis. Our results show that srtF cluster-encoded, S. suis pili are atypical compared to other Gram-positive pili. In addition, since the highly virulent strains under investigation are unlikely to produce other pili, our results suggest that pili might be dispensable for critical steps of the S. suis pathogenesis of infection.     

The response of phytoplankton community to anthropogenic pressure gradient in the coastal waters of the eastern Adriatic Sea

In order to test the response of phytoplankton to anthropogenic pressure, data of chlorophyll a concentration, phytoplankton abundance, and composition are analyzed in relation to anthropogenic pressure gradient and environmental variables such as temperature, salinity and nutrients. Investigated sites encompassed wide tropic range according to a preliminary determination of anthropogenic pressure, quantified through the LUSI index. Statistical analyses indicated nitrates and silicates as proxies of freshwater influence, and phytoplankton single metrics such as concentrations of chlorophyll a and abundances as indicators of anthropogenic pressure. Boundary values for different water quality classes for coastal waters under indirect freshwater influence (Type II) are obtained according to gradient between concentration of chlorophyll a and pressure index (LUSI), which empirically fit to exponential equation. The response of phytoplankton diversity was not linear, as the highest diversity was observed in the area with intermediate disturbance level. CCA analysis identified Skeletonema marinoii, Scrippsiella trochoidea, Guinardia flaccida, Leptocylindrus spp., Prorocentrum spp., Proboscia alata, Eutreptiella spp., and Pseudonitzschia spp. as local eutrophication indicators, whose abundances increased with nutrients loads.     

Revisiting the taxonomy of the “Dinophysis acuminata complex’’ (Dinophyta)’

Marine dinoflagellates of the genus Dinophysis are well known for producing diarrhetic shellfish poisoning (DSP) toxins and/or pectenotoxins which have a significant impact on public health as well as on marine aquaculture. Out of more than 80 Dinophysis species recorded so far, D. cf. acuminata is the most commonly observed in coastal areas worldwide. Due to their highly similar morphological features, however, an accurate discrimination of the various D. cf. acuminata species such as D. acuminata, D. ovum, and D. sacculus under light microscopy has proven to be a difficult task to accomplish. Hence, these species have thus far been referred to as the “Dinophysis acuminata complex”. Recent studies showed a discrimination between local strains of D. acuminata and D. ovum from Galician, northwestern Spain, using the mitochondrial cox1 gene as a genetic marker in addition to commonly used morphological features such as size and contour of the large hypothecal plates, shape of the small cells formed as part of their polymorphic life-cycle, development of the left sulcal list and ribs, and length of the right sulcal list. In the present study, attempts were made to discriminate between D. acuminata and D. ovum following single-cell isolation of 54 “D. acuminata complex” collected from Korean coastal waters, based on the abovementioned traits. Morphological data showed that all the traits analyzed overlapped between the two species. The mitochondrial cox1 (cytochrome c oxidase subunit I) gene sequences of every isolate were also determined, but a genetic distinction between D. acuminata and D. ovum could not be confirmed, suggesting that the cox1 gene is not a suitable genetic marker for discrimination between the two species. The results of this study suggest that the morphological variations observed within the “D. acuminata complex” may have been caused by several factors (e.g. different geographical locations, seasonal changes, and different environmental conditions), and that D. acuminata and D. ovum may be the same species.     

Molecular analysis of the interaction between the Bacillus subtilis trehalose repressor TreR and the tre operator

The trehalose operon of Bacillus subtilis is subject to regulation by induction, mediated by the repressor TreR, and by carbon catabolite repression (CCR). For in vitro investigations, TreR from B. subtilis was overproduced and purified. Its molecular mass, as estimated by SDS-PAGE, is 27?kDa. Size fractionation under native conditions yielded a size estimate of 56?kDa, indicating that TreR exists as a dimer in its native state. Analysis of its interaction with various DNA fragments shows that TreR is able to recognize two tre operators with different efficiencies, and indicates cooperative binding. Previous results have suggested that CCR of the tre operon occurs by a mechanism in which the specific regulator, TreR, may be involved independently of the central component, CcpA. The data presented here indicate that the TreR-tre operator interaction is influenced by several effectors. Thus, the presence of trehalose-6-phosphate, as well as glucose-1-phosphate and sodium chloride, inhibits tre operator binding. Glucose-6-phosphate can act as an anti-inducer, which might reflect its additional role in CCR exerted by glucose.     

Genome Dynamics Explain the Evolution of Flowering Time CCT Domain Gene Families in the Poaceae

Numerous CCT domain genes are known to control flowering in plants. They belong to the CONSTANS-like (COL) and PREUDORESPONSE REGULATOR (PRR) gene families, which in addition to a CCT domain possess B-box or response-regulator domains, respectively. Ghd7 is the most recently identified COL gene to have a proven role in the control of flowering time in the Poaceae. However, as it lacks B-box domains, its inclusion within the COL gene family, technically, is incorrect. Here, we show Ghd7 belongs to a larger family of previously uncharacterized Poaceae genes which possess just a single CCT domain, termed here CCT MOTIF FAMILY (CMF) genes. We molecularly describe the CMF (and related COL and PRR) gene families in four sequenced Poaceae species, as well as in the draft genome assembly of barley (Hordeum vulgare). Genetic mapping of the ten barley CMF genes identified, as well as twelve previously unmapped HvCOL and HvPRR genes, finds the majority map to colinear positions relative to their Poaceae orthologues. Combined inter-/intra-species comparative and phylogenetic analysis of CMF, COL and PRR gene families indicates they evolved prior to the monocot/dicot divergence ∼200 mya, with Poaceae CMF evolution described as the interplay between whole genome duplication in the ancestral cereal, and subsequent clade-specific mutation, deletion and duplication events. Given the proven role of CMF genes in the modulation of cereals flowering, the molecular, phylogenetic and comparative analysis of the Poaceae CMF, COL and PRR gene families presented here provides the foundation from which functional investigation can be undertaken.     

Analysis of spermiogenesis like a tool in the study of the triatomines of the Brasiliensis subcomplex

The specific identification and systematic of triatomines have been based fundamentally on morphological observations. These organisms are classified into complexes and specific subcomplexes, principally for morphological parameters and geographical disposition. The use of cytogenetic analyzes has been represented as a tool in systematic and taxonomy of triatomines. Thus, the present work, through the analysis of spermiogenesis, aims to characterize this stage of spermatogenesis in triatomines little studied, and especially to compare it among the species Triatoma lenti and T. sherlocki, to assist in the diagnosis of differentiation of these insects. The presence of the heteropyknotic corpuscle is shown as a diagnostic tool to differentiate T. sherlocki and T. lenti, since it is absent in T. lenti. The analysis of the spermiogenesis in T. sherlocki also allowed us to address morphological differences between elongating cells, which were relatively smaller and more filamentous when compared to T lenti. Furthermore, the flagellum was observed in all stages of cell differentiation and elongation. This structure, which helps in the locomotion of the sperm, is hardly observed in cytogenetic analysis, especially throughout spermiogenesis. Thus, although other comparative approaches should be taken, this paper allowed emphasizing the analysis of spermiogenesis as an important cytotaxonomic tool that assists in the differentiation of morphologically related species, such as T. lenti and T. sherlocki.     

A numerical study of the common narrow-leaved taxa of chenopodium occurring in the Western United States

A numerical taxonomic study was conducted on the common narrow-leaved members of the genusChenopodium that occur in the western United States. As many as six and as few as two species have been recognized. Thirty-five individual popu lations were used as OTU’s, and 35 characters were considered. The data were subjected to cluster (both the weighted-pair group method and the within-group dispersion method) and principal component analysis. The results of both analyses are in general agreement and suggest phenetic relationships that differ from current interpretations in the literature. It is concluded thatC. desiccatum var.leptophylloides is much more closely related phenetically toC. atrovirens (and may not be distinct from it) than it is toC. desiccatum var.desiccatum. Chenopodium hians andC. leptophyllum appear to be phenetically distinct despite the fact that they have sometimes been viewed as conspecific.Cheno podium subglabrum is not very similar toC. leptophyllum, and should probably be treated as a separate species rather than as a variety of the latter. Popu lations referable toC. incognitum form a rather loose cluster that appears some what intermediate between populations ofC. desiccatum var.leptophylloides,C. atrovirens, C. hians, andC. leptophyllum.     

Biodegradation of the Allelopathic Chemical m-Tyrosine by Bacillus aquimaris SSC5 Involves the Homogentisate Central Pathway

m-Tyrosine is an amino acid analogue, exuded from the roots of fescue grasses, which acts as a potent allelopathic and a broad spectrum herbicidal chemical. Although the production and toxic effects of m-tyrosine are known, its microbial degradation has not been documented yet. A soil microcosm study showed efficient degradation of m-tyrosine by the inhabitant microorganisms. A bacterial strain designated SSC5, that was able to utilize m-tyrosine as the sole source of carbon, nitrogen, and energy, was isolated from the soil microcosm and was characterized as Bacillus aquimaris. Analytical methods such as HPLC, GC-MS, and ¹H-NMR performed on the resting cell samples identified the formation of 3-hydroxyphenylpyruvate (3-OH-PPA), 3-hydroxyphenylacetate (3-OH-PhAc), and homogentisate (HMG) as major intermediates in the m-tyrosine degradation pathway. Enzymatic assays carried out on cell-free lysates of m-tyrosine-induced cells confirmed transamination reaction as the first step of m-tyrosine degradation. The intermediate 3-OH-PhAc thus obtained was further funneled into the HMG central pathway as revealed by a hydroxylase enzyme assay. Subsequent degradation of HMG occurred by ring cleavage catalyzed by the enzyme homogentisate 1, 2-dioxygenase. This study has significant implications in terms of understanding the environmental fate of m-tyrosine as well as regulation of its phytotoxic effect by soil microorganisms.     

Phyllocarida and the origin of the Malacostraca

Tentative homologies with Decapoda are proposed for grooves on the carapace of Echinocaris and Montecaris. Together with F.R. Schram's hoploid trend, reinforced by the discovery of a raptorial limb in Sairocaris, this may strengthen a phyllocarid origin for Eumalacostraca. Both these trends, however, may be parallelisms, and phylogenetic analysis is required. Pleopods are here proved to have been present in additional Archaeostraca, which improves their ancestral condition. E. Dahl's view of the phyllocarids as an evolutionary dead end is falsified from fossils, and the phyllocarids are supported as a stem group of the Malacostraca.     

A duplicated amh is the master sex-determining gene for Sebastes rockfish in the Northwest Pacific

Teleost fish are the most diverse group of vertebrates and provide opportunities to study the evolution of sex determination (SD) systems. Using genomic and functional analyses, we identified a male-specific duplication of anti-Müllerian hormone (amh) gene as the male master sex-determining (MSD) gene in Sebastes schlegelii. By resequencing 10 males and 10 females, we characterized a 5 kb-long fragment in HiC_Scaffold_12 as a male-specific region, which contained an amh gene (named amhy). We then demonstrated that amhy is a duplication of autosomal amh that was later translocated to the ancestral Y chromosome. amha and amhy shared high-nucleotide identity with the most significant difference being two insertions in intron 4 of amhy. Furthermore, amhy overexpression triggered female-to-male sex reversal in S. schlegelii, displaying its fundamental role in driving testis differentiation. We developed a PCR assay which successfully identified sexes in two species of northwest Pacific rockfish related to S. schlegelii. However, the PCR assay failed to distinguish the sexes in a separate clade of northeast Pacific rockfish. Our study provides new examples of amh as the MSD in fish and sheds light on the convergent evolution of amh duplication as the driving force of sex determination in different fish taxa.     

Investigating the Roles of the C-Terminal Domain of Plasmodium falciparum GyrA

Malaria remains as one of the most deadly diseases in developing countries. The Plasmodium causative agents of human malaria such as Plasmodium falciparum possess an organelle, the apicoplast, which is the result of secondary endosymbiosis and retains its own circular DNA. A type II topoisomerase, DNA gyrase, is present in the apicoplast. In prokaryotes this enzyme is a proven, effective target for antibacterial agents, and its discovery in P. falciparum opens up the prospect of exploiting it as a drug target. Basic characterisation of P. falciparum gyrase is important because there are significant sequence differences between it and the prokaryotic enzyme. However, it has proved difficult to obtain soluble protein. Here we have predicted a new domain boundary in P. falciparum GyrA that corresponds to the C-terminal domain of prokaryotic GyrA and successfully purified it in a soluble form. Biochemical analyses revealed many similarities between the C-terminal domains of GyrA from E. coli and P. falciparum, suggesting that despite its considerably larger size, the malarial protein carries out a similar DNA wrapping function. Removal of a unique Asn-rich region in the P. falciparum protein did not result in a significant change, suggesting it is dispensable for DNA wrapping.     

Induction of the lac promoter in the absence of DNA loops and the stoichiometry of induction

In vivo induction of the Escherichia coli lactose operon as a function of inducer concentration generates a sigmoidal curve, indicating a non-linear response. Suggested explanations for this dependence include a 2:1 inducer–repressor stoichiometry of induction, which is the currently accepted view. It is, however, known for decades that, in vitro, operator binding as a function of inducer concentration is not sigmoidal. This discrepancy between in vivo and in vitro data has so far not been resolved. We demonstrate that the in vivo non-linearity of induction is due to cooperative repression of the wild-type lac operon through DNA loop formation. In the absence of DNA loops, in vivo induction curves are hyperbolic. In the light of this result, we re-address the question of functional molecular inducer–repressor stoichiometry in induction of the lac operon.     

Xylella fastidiosa in Europe: From the Introduction to the Current Status

Xylella fastidiosa is xylem-limited bacterium capable of infecting a wide range of host plants, resulting in Pierce’s disease in grapevine, citrus variegated chlorosis, olive quick decline syndrome, peach phony disease, plum leaf scald, alfalfa dwarf, margin necrosis and leaf scorch affecting oleander, coffee, almond, pecan, mulberry, red maple, oak, and other types of cultivated and ornamental plants and forest trees. In the European Union, X. fastidiosa is listed as a quarantine organism. Since its first outbreak in the Apulia region of southern Italy in 2013 where it caused devastating disease on Olea europaea (called olive leaf scorch and quick decline), X. fastidiosa continued to spread and successfully established in some European countries (Corsica and PACA in France, Balearic Islands, Madrid and Comunitat Valenciana in Spain, and Porto in Portugal). The most recent data for Europe indicates that X. fastidiosa is present on 174 hosts, 25 of which were newly identified in 2021 (with further five hosts discovered in other parts of the world in the same year). From the six reported subspecies of X. fastidiosa worldwide, four have been recorded in European countries (fastidiosa, multiplex, pauca, and sandyi). Currently confirmed X. fastidiosa vector species are Philaenus spumarius, Neophilaenus campestris, and Philaenus italosignus, whereby only P. spumarius (which has been identified as the key vector in Apulia, Italy) is also present in Americas. X. fastidiosa control is currently based on pathogen-free propagation plant material, eradication, territory demarcation, and vector control, as well as use of resistant plant cultivars and bactericidal treatments.     

Structures of the oligosaccharides isolated from milk of the platypus

Eight major oligosaccharides were isolated from platypus milk. By sequential exoglycosidase digestion and methylation study, their structures were elucidated as shown in Fig. 9 of this paper. The characteristics feature of the platypus milk oligosaccharides is that lacto-N-neotetraose and lacto-N-neohexaose are the major cores in contrast to human milk oligosaccharides in which lacto-N-tetraose and lacto-N-hexaose are found as the major core.     

Unravelling the Wolbachia evolutionary role: the reprogramming of the host genomic imprinting

Environmental factors can induce significant epigenetic changes that may also be inherited by future generations. The maternally inherited symbiont of arthropods Wolbachia pipientis is an excellent candidate as an ‘environmental’ factor promoting trans-generational epigenetic changes: by establishing intimate relationships with germ-line cells, epigenetic effects of Wolbachia symbiosis would be manifested as a ‘maternal effect’, in which infection of the mother modulates the offspring phenotype. In the leafhopper Zyginidia pullula, Wolbachia feminizes genetic males, leaving them as intersexes. With the exception of male chitinous structures that are present in the last abdominal segment, feminized males display phenotypic features that are typical of females. These include ovaries that range from a typical histological architecture to an altered structure. Methylation-sensitive random amplification of polymorphic DNA profiles show that they possess a female genomic imprint. On the other hand, some rare feminized males bear testes instead of ovaries. These specimens possess a Wolbachia density approximately four orders of magnitude lower than feminized males with ovaries and maintain a male genome—methylation pattern. Our results indicate that Wolbachia infection disrupts male imprinting, which dramatically influences the expression of genes involved in sex differentiation and development, and the alteration occurs only if Wolbachia exceeds a density threshold. Thus, a new Wolbachia''s role as an environmental evolutionary force, inducing epigenetic trans-generational changes, should now be considered.