MicrobiologyBytes

The Archaea possess unique metabolic pathways, distinct from those in Bacteria and Eukarya. Based on the genome sequences of the Archaea, there are many cases in which a particular metabolic pathway seems to be absent or incomplete. The search for these ‘missing’ pathways or enzymes has been an exciting field of research in the Archaea. A representative example was the CO₂-fixing mechanisms in autotrophic Crenarchaeota. Although many autotrophic Crenarchaeota had been isolated, homologs of previously recognized CO₂-fixing pathways could not be identified on their genomes. Genes responsible for the degradation and biosynthesis of various sugars had also been unidentified. This paper describes recent findings in archaeal metabolism, including sugar metabolism, CO₂ fixation and a wide range of biosynthetic pathways. The predicted distributions of these pathways, based on genome sequence analyses, in the Archaea are also discussed. These investigations will help understand how microorganisms use and interact with the many natural and man-made compounds they encounter in their environments and also provide the foundation for many biotechnology developments.

Novel metabolic pathways in Archaea, Curr Opin Microbiol. May 23 2011 doi:10.1016/j.mib.2011.04.014
The Archaea harbor many metabolic pathways that differ to previously recognized classical pathways. Glycolysis is carried out by modified versions of the Embden-Meyerhof and Entner-Doudoroff pathways. Thermophilic archaea have recently been found to harbor a bi-functional fructose-1,6-bisphosphate aldolase/phosphatase for gluconeogenesis. A number of novel pentose-degrading pathways have also been recently identified. In terms of anabolic metabolism, a pathway for acetate assimilation, the methylaspartate cycle, and two CO(₂)-fixing pathways, the 3-hydroxypropionate/4-hydroxybutyrate cycle and the dicarboxylate/4-hydroxybutyrate cycle, have been elucidated. As for biosynthetic pathways, recent studies have clarified the enzymes responsible for several steps involved in the biosynthesis of inositol phospholipids, polyamine, coenzyme A, flavin adeninedinucleotide and heme. By examining the presence/absence of homologs of these enzymes on genome sequences, we have found that the majority of these enzymes and pathways are specific to the Archaea.

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Bacteria can change course almost instantaneously, zipping towards food or away from toxins. How do such simple organisms do something so complex? It’s all in the flagella, a tail-like structure with rotating helical filaments. The flagella work in unison to propel the cell forward by rotating counterclockwise and thus bundling together. When the flagella reverse their rotation to clockwise, they disrupt the bundle and make the cell tumble in place. When the flagella shift back to counterclockwise again, the bacteria set off on a new course.

This description of bacterial locomotion is well known, but the mechanisms that allow the flagella to shift gears from counterclockwise to clockwise have proven difficult to identify. Now, a new study bring us closer to answering this fundamental question and propose a new model describing how flagella manage this switch. Filaments in the flagella are powered by rotary motors that span the cell membrane. Things of beauty, these motors are tooled so precisely that they are nearly 100% efficient, and their direction is set by a rotor that can turn thousands of revolutions per minute. The rotor shifts from the forward-propelling counterclockwise to the tumble-inducing clockwise when chemical gradients tell bacteria they’ve gone astray, for example, away from food. This activates a cytoplasmic signaling protein that binds proteins in the rotor switch, changing the orientation of another switch protein called FliG and thereby reversing the rotor’s spin to clockwise.

Source: How Bacteria Shift Gears. 2011 PLoS Biol 9(5): e1001061. doi:10.1371/journal.pbio.1001061

Related:

• Bacterial motility

Bacterial associates are ubiquitous among insects, including mosquitoes. Wolbachia are obligate endosymbiotic bacteria that infect numerous insects, many of which are vectors of pathogenic microorganisms. Interest has centered around Wolbachia as a means of reducing arthropod-borne disease due to the capacity of the bacteria to manipulate the reproduction of the insect host, which in turn favors their own transmission.

Recent studies show that Wolbachia can directly cause pathogen interference (PI) in their invertebrate hosts, whereby infected insects are less susceptible to pathogens. Infection with Wolbachia bacteria has been shown to reduce pathogen levels in multiple mosquito species. Anopheles mosquitoes (the obligate vectors of human malaria) are naturally uninfected with Wolbachia, and stable artificial infections have not yet succeeded in this genus; however somatic infections can be established that can be used to assess the effect of Wolbachia infection in Anopheles. Here, we show that infection with two different Wolbachia strains can significantly reduce levels of the human malaria parasite Plasmodium falciparum in Anopheles gambiae. After infection, Wolbachia disseminate throughout the mosquito but are notably absent from the gut and ovaries. The mosquito immune system is first induced in response to Wolbachia infection, but is then suppressed as the infection progresses. The Wolbachia strain wMelPop is highly virulent to Anopheles only after blood feeding. If stable infections can be established in Anopheles, and they act in a similar manner to somatic infections, Wolbachia could potentially be used as part of a strategy to control malaria.

Wolbachia Infections Are Virulent and Inhibit the Human Malaria Parasite Plasmodium Falciparum in Anopheles Gambiae. 2011 PLoS Pathog 7(5): e1002043. doi:10.1371/journal.ppat.1002043
Endosymbiotic Wolbachia bacteria are potent modulators of pathogen infection and transmission in multiple naturally and artificially infected insect species, including important vectors of human pathogens. Anopheles mosquitoes are naturally uninfected with Wolbachia, and stable artificial infections have not yet succeeded in this genus. Recent techniques have enabled establishment of somatic Wolbachia infections in Anopheles. Here, we characterize somatic infections of two diverse Wolbachia strains (wMelPop and wAlbB) in Anopheles gambiae, the major vector of human malaria. After infection, wMelPop disseminates widely in the mosquito, infecting the fat body, head, sensory organs and other tissues but is notably absent from the midgut and ovaries. Wolbachia initially induces the mosquito immune system, coincident with initial clearing of the infection, but then suppresses expression of immune genes, coincident with Wolbachia replication in the mosquito. Both wMelPop and wAlbB significantly inhibit Plasmodium falciparum oocyst levels in the mosquito midgut. Although not virulent in non-bloodfed mosquitoes, wMelPop exhibits a novel phenotype and is extremely virulent for approximately 12–24 hours post-bloodmeal, after which surviving mosquitoes exhibit similar mortality trajectories to control mosquitoes. The data suggest that if stable transinfections act in a similar manner to somatic infections, Wolbachia could potentially be used as part of a strategy to control the Anopheles mosquitoes that transmit malaria.

HCMV infection dramatically alters cellular metabolism, this review focuses on how the virus alters glucose and glutamine metabolism and fatty acid synthesis. Changes to glucose metabolism include increased glucose uptake through the induction of the glucose transporter GLUT4, upregulation of glycolytic enzymes and probable allosteric activation of glycolysis. Although these changes are rather dramatic they are likely to represent only a fraction of the metabolic changes wrought by HCMV infection. Such changes are likely to contribute to forms of HCMV pathogenesis that could be very different from those that have been studied and documented to this point. For example, HCMV and other viruses which induce metabolic changes similar to HCMV might play a greater role in oncogenesis than previously recognized. At this point, our understanding of the mechanisms used by HCMV to alter metabolic processes is very limited and requires further study.

Viral effects on metabolism: changes in glucose and glutamine utilization during human cytomegalovirus infection. Trends Microbiol. May 11 2011
Human cytomegalovirus (HCMV) infection causes dramatic alterations of intermediary metabolism, similar to those found in tumor cells. In infected cells, glucose carbon is not completely broken down by the tricarboxylic acid (TCA) cycle for energy; instead, it is used biosynthetically. This process requires increased glucose uptake, increased glycolysis and the diversion of glucose carbon, in the form of citrate, from the TCA cycle for use in HCMV-induced fatty acid biosynthesis. The diversion of citrate from the TCA cycle (cataplerosis) requires induction of enzymes to promote glutaminolysis, the conversion of glutamine to α-ketoglutarate to maintain the TCA cycle (anaplerosis) and ATP production. Such changes could result in heretofore uncharacterized pathogenesis, potentially implicating HCMV as a subtle cofactor in many maladies, including oncogenesis. Recognition of the effects of HCMV, and other viruses, on host cell metabolism will provide new understanding of viral pathogenesis and novel avenues for antiviral therapy.

A number of ecological studies have revealed that microbial viruses predominate in the biosphere and outnumber their hosts by at least one order of magnitude. Due to their abundance and consequent influence on the composition and diversity of microbial communities, viruses can be rightfully considered to be the “major players in the global ecosystem”. Until recently, the majority of viruses in the environment were believed to possess double-stranded DNA genomes. However, technological advances in single-stranded (ss) DNA amplification and sequencing from environmental samples revealed that viruses with ssDNA genomes are more prevalent in both soil and marine environments than previously recognized. This realization precipitated an interest amongst environmental virologists in the diversity and distribution of ssDNA bacterial viruses in nature. Among ssDNA viruses that are most often identified in the environment using metagenomic approach are those belonging to the family Microviridae. However, the host organisms have yet to be determined.

Unexplored diversity and abundance of the Microviridae viruses in the environment fuels interest in this virus group. In order to obtain more information about these viruses, researchers analyzed the genomic sequences available in public databases for the presence of proviruses related to Microviridae. The rationale behind this approach is that a provirus, defective or not, represents a molecular record that a cell has been in contact with a particular virus. This study identified seven proviruses that are related to members of the Microviridae. The proviruses are integrated in the genomes of different species of the order Bacteroidales (phylum Bacteroidetes). The identified proviruses are only distantly related to the previously characterized microviruses and gokushoviruses and may represent a new group or subfamily within the Microviridae. Searches against metagenomic databases suggest that these new viruses might be associated with the human gut microbiota. This extends our knowledge of the evolution, diversity and host range of microviruses.

Microviridae Goes Temperate: Microvirus-Related Proviruses Reside in the Genomes of Bacteroidetes. 2011 PLoS ONE 6(5): e19893. doi:10.1371/journal.pone.0019893
The Microviridae comprises icosahedral lytic viruses with circular single-stranded DNA genomes. The family is divided into two distinct groups based on genome characteristics and virion structure. Viruses infecting enterobacteria belong to the genus Microvirus, whereas those infecting obligate parasitic bacteria, such as Chlamydia, Spiroplasma and Bdellovibrio, are classified into a subfamily, the Gokushovirinae. Recent metagenomic studies suggest that members of the Microviridae might also play an important role in marine environments. In this study we present the identification and characterization of Microviridae-related prophages integrated in the genomes of species of the Bacteroidetes, a phylum not previously known to be associated with microviruses. Searches against metagenomic databases revealed the presence of highly similar sequences in the human gut. This is the first report indicating that viruses of the Microviridae lysogenize their hosts. Absence of associated integrase-coding genes and apparent recombination with dif-like sequences suggests that Bacteroidetes-associated microviruses are likely to rely on the cellular chromosome dimer resolution machinery. Phylogenetic analysis of the putative major capsid proteins places the identified proviruses into a group separate from the previously characterized microviruses and gokushoviruses, suggesting that the genetic diversity and host range of bacteriophages in the family Microviridae is wider than currently appreciated.

Wolbachia are a group of bacteria that infect a major proportion of insect species. They are known for intricate manipulations of their host’s reproduction. The most puzzling manipulation is called Cytoplasmic Incompatibility (CI). In males, CI consists of Wolbachia manipulating the sperm in a yet unknown way – this manipulation is called mod (for modification). DNA from modified sperm cannot properly participate in the first embryonic mitosis, except if Wolbachia action in the egg recovers the functionality of the sperm DNA.

Owing to the nature of CI, a female can only successfully mate with an infected male if she is herself infected by an appropriate Wolbachia strain. If such an infected female mates with an uninfected male, there are no defects. Therefore, infected females have a selective advantage over uninfected ones, helping Wolbachia spread. Considering that CI effectively inhibits certain crosses, Wolbachia infection could lead to reproductive isolation or gene flow reduction between host populations with different infection statuses. Therefore, CI in Wolbachia may play an important role in insect speciation. A deeper insight into the mechanism behind Wolbachia-induced CI is likely to further our understanding of host evolutionary dynamics.

A New Model and Method for Understanding Wolbachia-Induced Cytoplasmic Incompatibility. 2011 PLoS ONE 6(5): e19757. doi:10.1371/journal.pone.0019757
Wolbachia are intracellular bacteria transmitted almost exclusively vertically through eggs. In response to this mode of transmission, Wolbachia strategically manipulate their insect hosts’ reproduction. In the most common manipulation type, cytoplasmic incompatibility, infected males can only mate with infected females, but infected females can mate with all males. The mechanism of cytoplasmic incompatibility is unknown; theoretical and empirical findings need to converge to broaden our understanding of this phenomenon. For this purpose, two prominent models have been proposed: the mistiming-model and the lock-key-model. The former states that Wolbachia manipulate sperm of infected males to induce a fatal delay of the male pronucleus during the first embryonic division, but that the bacteria can compensate the delay by slowing down mitosis in fertilized eggs. The latter states that Wolbachia deposit damaging “locks” on sperm DNA of infected males, but can also provide matching “keys” in infected eggs to undo the damage. The lock-key-model, however, needs to assume a large number of locks and keys to explain all existing incompatibility patterns. The mistiming-model requires fewer assumptions but has been contradicted by empirical results. We therefore expand the mistiming-model by one quantitative dimension to create the new, so-called goalkeeper-model. Using a method based on formal logic, we show that both lock-key- and goalkeeper-model are consistent with existing data. Compared to the lock-key-model, however, the goalkeeper-model assumes only two factors and provides an idea of the evolutionary emergence of cytoplasmic incompatibility. Available cytological evidence suggests that the hypothesized second factor of the goalkeeper-model may indeed exist. Finally, we suggest empirical tests that would allow to distinguish between the models. Generalizing our results might prove interesting for the study of the mechanism and evolution of other host-parasite interactions.

Guardian University Guide 2012: Biosciences

Study at the University of Leicester: Microbiology or Medical Microbiology

Biological membranes represent a major barrier during virus infection. While the mechanism by which an enveloped virus breaches the limiting membrane of a host cell is well-characterized, this membrane penetration process is poorly understood for non-enveloped viruses. Indeed, most available insights on membrane transport of non-enveloped viruses are built upon in vitro studies.

A new paper describes a cell-based assay to explain the molecular mechanism by which the non-enveloped SV40 penetrates the endoplasmic reticulum (ER) membrane to access the cytosol, a critical step in infection. Strikingly, it shows that SV40 breaches the ER membrane as a large and intact virus particle, despite the conformational changes it experiences in the ER lumen. This suggests that the ER membrane can accommodate translocation of a large protein complex, possibly through either a sizeable protein channel or the ER membrane bilayer. This data illuminates the cellular mechanism by which a non-enveloped virus penetrates the limiting membrane of a target cell during infection.

A Large and Intact Viral Particle Penetrates the Endoplasmic Reticulum Membrane to Reach the Cytosol. 2011 PLoS Pathog 7(5): e1002037. doi:10.1371/journal.ppat.1002037
Non-enveloped viruses penetrate host membranes to infect cells. A cell-based assay was used to probe the endoplasmic reticulum (ER)-to-cytosol membrane transport of the non-enveloped SV40. We found that, upon ER arrival, SV40 is released into the lumen and undergoes sequential disulfide bond disruptions to reach the cytosol. However, despite these ER-dependent conformational changes, SV40 crosses the ER membrane as a large and intact particle consisting of the VP1 coat, the internal components VP2, VP3, and the genome. This large particle subsequently disassembles in the cytosol. Mutant virus and inhibitor studies demonstrate VP3 and likely the viral genome, as well as cellular proteasome, control ER-to-cytosol transport. Our results identify the sequence of events, as well as virus and host components, that regulate ER membrane penetration. They also suggest that the ER membrane supports passage of a large particle, potentially through either a sizeable protein-conducting channel or the lipid bilayer.