MicrobiologyBytes

Iron is required for many cellular processes, but can be toxic at high concentrations. Thus iron homeostasis is strictly regulated so that iron acquisition, storage, and consumption are geared to iron availability, and that intracellular levels of free iron do not reach toxic levels. Recently, the roles of manganese and its control in cells have been investigated, and it is becoming clear that some aspects of the metabolism of iron and manganese are interrelated.

Control of bacterial iron homeostasis by manganese. PNAS USA May 24 2010. doi: 10.1073/pnas.100234210
Perception and response to nutritional iron availability by bacteria are essential to control cellular iron homeostasis. The Irr protein from Bradyrhizobium japonicum senses iron through the status of heme biosynthesis to globally regulate iron-dependent gene expression. Heme binds directly to Irr to trigger its degradation. Here, we show that severe manganese limitation created by growth of a Mn2+ transport mutant in manganese-limited media resulted in a cellular iron deficiency. In wild-type cells, Irr levels were attenuated under manganese limitation, resulting in reduced promoter occupancy of target genes and altered iron-dependent gene expression. Irr levels were high regardless of manganese availability in a heme-deficient mutant, indicating that manganese normally affects heme-dependent degradation of Irr. Manganese altered the secondary structure of Irr in vitro and inhibited binding of heme to the protein. We propose that manganese limitation destabilizes Irr under low-iron conditions by lowering the threshold of heme that can trigger Irr degradation. The findings implicate a mechanism for the control of iron homeostasis by manganese in a bacterium.

Related:

• Iron Uptake and Virulence
• How bacteria capture iron from heme
• Utilization of iron by Campylobacter jejuni

Bats, which are ‘keystone species’ in many ecosystems, play notable roles in plant pollination, forest regeneration and control of insect populations. Bats are important to human health as they are reservoirs or carriers for rabies and other viruses, parasites, and pathogenic fungi. Hibernation is believed to be an important adaptation in bats that may contribute to their exceptional longevity. The common little brown bat (Myotis lucifugus) hibernates, along with the endangered Indiana bat (Myotis sodalis), in many hibernacula in the Northeastern United States, including caves and mines in upstate New York. Hibernating bats can suffer significant mortality due to adverse environmental conditions such as freezing or flooding, as well as human activities including visitation and pesticide applications. No mass mortality was reported until recently from bat sites that had been surveyed for almost three decades by the New York State Department of Environmental Conservation. Recently, however little brown bats have been found to be dying in large numbers at many hibernation sites in upstate New York. This problem has spread to other States in the Northeastern USA.

Morphological and Molecular Characterizations of Psychrophilic Fungus Geomyces destructans from New York Bats with White Nose Syndrome (WNS). PLoS ONE 5(5): e10783. doi:10.1371/journal.pone.0010783
Massive die-offs of little brown bats (Myotis lucifugus) have been occurring since 2006 in hibernation sites around Albany, New York, and this problem has spread to other States in the Northeastern United States. White cottony fungal growth is seen on the snouts of affected animals, a prominent sign of White Nose Syndrome (WNS). A previous report described the involvement of the fungus Geomyces destructans in WNS, but an identical fungus was recently isolated in France from a bat that was evidently healthy. The fungus has been recovered sparsely despite plentiful availability of afflicted animals.
We have investigated 100 bat and environmental samples from eight affected sites in 2008. Our findings provide strong evidence for an etiologic role of G. destructans in bat WNS. (i) Direct smears from bat snouts, Periodic Acid Schiff-stained tissue sections from infected tissues, and scanning electron micrographs of bat tissues all showed fungal structures similar to those of G. destructans. (ii) G. destructans DNA was directly amplified from infected bat tissues, (iii) Isolations of G. destructans in cultures from infected bat tissues showed 100% DNA match with the fungus present in positive tissue samples. (iv) RAPD patterns for all G. destructans cultures isolated from two sites were indistinguishable. (v) The fungal isolates showed psychrophilic growth. (vi) We identified in vitro proteolytic activities suggestive of known fungal pathogenic traits in G. destructans.
Further studies are needed to understand whether G. destructans WNS is a symptom or a trigger for bat mass mortality. The availability of well-characterized G. destructans strains should promote an understanding of bat–fungus relationships, and should aid in the screening of biological and chemical control agents.

Related:

• Colony Collapse Disorder

Success of a virus infection requires that each infected cell delivers a sufficient number of infectious particles to allow new rounds of infection. In picornaviruses, virus replication is initiated by the viral polymerase and a viral-coded protein, termed VPg, that primes RNA synthesis. Foot-and-mouth disease virus (FMDV) is exceptional among picornaviruses in that its genome encodes three copies of VPg. Why FMDV encodes three VPgs is unknown.

Researchers constructed four mutant FMDVs that encode only one VPg: either VPg1, VPg3, or two chimeric versions containing part of VPg1 and VPg3. All mutants, except that encoding only VPg1, were replication-competent. Unexpectedly, despite being replication-competent, the mutants did not form plaques on BHK-21 cell monolayers. The one-VPg mutant FMDVs released lower amounts of encapsidated viral RNA to the extracellular environment than wild type FMDV, suggesting that deficient plaque formation was associated with insufficient release of infectious progeny. Mutant FMDVs subjected to serial passages in BHK-21 cells regained plaque-forming capacity without modification of the number of copies of VPg. Substitutions in non-structural proteins 2C, 3A and VPg were associated with restoration of plaque formation. Specifically, replacement R55W in protein 2C was repeatedly found in several mutant viruses that had regained competence in plaque development. The results link the VPg copies in the FMDV genome with the cytopathology capacity of the virus, and have unveiled yet another function of 2C: modulation of picornavirus cell-to-cell transmission. these data highlight the role of non–structural proteins in the adaptability to changing environments during picornavirus infections, with clear implications for viral pathogenesis.

Deletion Mutants of VPg Reveal New Cytopathology Determinants in a Picornavirus. 2010 PLoS ONE 5(5): e10735. doi:10.1371/journal.pone.0010735

Related:

• Foot and Mouth Disease

Antibiotic drug–target interactions, and their respective direct effects, are generally well characterized. By contrast, the bacterial responses to antibiotic drug treatments that contribute to cell death are not as well understood and have proven to be complex as they involve many genetic and biochemical pathways. This review discusses the multilayered effects of drug–target interactions, including the essential cellular processes that are inhibited by bactericidal antibiotics and the associated cellular response mechanisms that contribute to killing. It also discusses new insights into these mechanisms that have been revealed through the study of biological networks, and describe how these insights, together with related developments in synthetic biology, could be exploited to create new antibacterial therapies.

How antibiotics kill bacteria: from targets to networks. 2010 Nature Reviews Microbiology 8, 423-435 doi:10.1038/nrmicro2333

Related:

• The cost of antibiotic resistance
• The clinical consequences of antibiotic resistance
• Antibiotics, your gut, and you

Human rhinoviruses (HRV) are the most frequent cause of respiratory infection in humans. These viruses belong to the Picornaviridae, one of the oldest and most diversified human virus family, characterized by a non-enveloped, single positive-stranded RNA genome. Although rhinovirus replication is often restricted to the upper respiratory tract leading to self-limited illnesses of short duration, such as the common cold, HRV can also invade the lower respiratory tract and lead to more serious infections. Similar to many other RNA viruses, the error-prone rhinoviral polymerase can accumulate a large number of nucleotide mutations over a very short period of time, a feature that favors viral adaptation. The error rate of picornavirus RNA polymerases has been estimated to range between 10⁻³ and 10⁻⁴ errors/nucleotide/cycle of replication. This variability is a driving force for virus evolution and results in a large genetic and phenotypic diversity illustrated by the very high number of different HRV serotypes identified to date.

By using ultra-deep sequencing technology, researchers were able to pinpoint HRV evolution at the level of a quasispecies population both in vivo and in vitro. The data illustrate the ability of rhinoviruses to produce several new variants as rapidly as 5 days’ post-infection.

Rhinovirus Genome Evolution during Experimental Human Infection. 2010 PLoS ONE 5(5): e10588. doi:10.1371/journal.pone.0010588
Human rhinoviruses (HRVs) evolve rapidly due in part to their error-prone RNA polymerase. Knowledge of the diversity of HRV populations emerging during the course of a natural infection is essential and represents a basis for the design of future potential vaccines and antiviral drugs. To evaluate HRV evolution in humans, nasal wash samples were collected daily for five days from 15 immunocompetent volunteers experimentally infected with a reference stock of HRV-39. In parallel, HeLa-OH cells were inoculated to compare HRV evolution in vitro. Nasal wash in vivo assessed by real-time PCR showed a viral load that peaked at 48–72 h. Ultra-deep sequencing was used to compare the low-frequency mutation populations present in the HRV-39 inoculum in two human subjects and one HeLa-OH supernatant collected 5 days post-infection. The analysis revealed hypervariable mutation locations in VP2, VP3, VP1, 2C and 3C genes and conserved regions in VP4, 2A, 2B, 3A, 3B and 3D genes. These results were confirmed by classical sequencing of additional samples, both from inoculated volunteers and independent cell infections, and suggest that HRV inter-host transmission is not associated with a strong bottleneck effect. A specific analysis of the VP1 capsid gene of 15 human cases confirmed the high mutation incidence in this capsid region, but not in the antiviral drug-binding pocket. We could also estimate a mutation frequency in vivo of 3.4×10⁻⁴ mutations/nucleotides and 3.1×10⁻⁴ over the entire ORF and VP1 gene, respectively. In vivo, HRV generate new variants rapidly during the course of an acute infection due to mutations that accumulate in hot spot regions located at the capsid level, as well as in 2C and 3C genes.

Related:

• Virus incubation periods
• Sleep and susceptibility to the common cold

Malaria caused by Plasmodium falciparum is a disease that is responsible for 880,000 deaths per year worldwide. Vaccine development has proved difficult and resistance has emerged for most antimalarial drugs. To discover new antimalarial chemotypes, researchers have used a chemical genetic approach to assay 309,474 chemicals. Many chemicals in the library of compounds tested showed potent in vitro activity against drug-resistant P. falciparum strains. A reverse chemical genetic study identified 19 new inhibitors of 4 validated drug targets and 15 novel binders among 61 malarial proteins. Phylochemogenetic profiling in several organisms revealed similarities between Toxoplasma gondii and mammalian cell lines and dissimilarities between P. falciparum and related protozoans. One exemplar compound displayed efficacy in a mouse model. These findings provide the scientific community with new starting points for malaria drug discovery.

Chemical genetics of Plasmodium falciparum. Nature 465, 311–315 (2010) doi:10.1038/nature09099

Related:

• Drug discovery: Priming the antimalarial pipeline
• Nature Collections – Malaria
• New ways to beat malaria

Strategies in modern agriculture aim to enhance harvest yields per acreage and to reduce pre-harvest and post-harvest losses caused by detrimental abiotic and biotic causes. The potential of that reflects an estimate 20–40% reduction of agricultural production worldwide that is taken by pests and diseases. Modern pest management strategies in crop plants include classical and molecular marker-based resistance breeding, genetic engineering of plant immunity and the use of chemicals as pesticides or strengtheners of plant health. While breeding strategies are time-consuming and harbor the problem of ‘linkage drag’ (transfer of undesirable traits that need to be removed after introgression of the desired trait by back-crossing), genetic engineering holds the potential of being reasonably fast and predictable in its consequences because of the targeted introduction of individual, heterologous traits into elite crop lines.

Sequencing of entire plant genomes, systematic plant transcriptome profiling and comprehensive genetic dissection of immune pathways in model plants (Arabidopsis thaliana, rice) has significantly enhanced our understanding of the mechanisms underlying microbial infection and plant immunity. The plant immune system consists of two evolutionarily linked branches. Recognition of invariant microbial surface patterns (pathogen or microbe-associated patterns; PAMP/MAMP) through plant pattern recognition receptors is referred to as PAMP-triggered immunity (PTI) and is the basis for broad-spectrum resistance of plants against host non-adapted microbial pathogens (i.e. all genetic variants of a given microbial species are unable to grow on a given plant species).

Novel insight into plant immunity and disease may now be turned into new tools to engineer durable, broad-spectrum plant disease resistance. This review highlights recent scientific discoveries in plant immunity and discusses their potential for enhancing plant immunity in crop plants with particular emphasis on immunity to bacterial and fungal infection. Saving the world’s food supply constitutes one of the major challenges of the future. As a complement to classical and molecular breeding technologies, novel strategies for biotechnological improvement of plant immunity aim at enhancing host recognition capacities for potential pathogens, at boosting the executive arsenal of plant immunity, and at interfering with virulence strategies employed by microbial pathogens. In addition, chemical and biological priming provides means for triggering plant defenses in a non-transgenic manner. Major advances in our understanding of the molecular basis of plant immunity and of microbial infection strategies have opened new ways for engineering durable disease resistance in crop plants that are highlighted in this review.

Biotechnological concepts for improving plant innate immunity. Curr Opin Biotechnol. Feb 22 2010

Related:

• Toxins for microbial attack and plant defence
• Resistance to Plant Viruses
• Bacteriophages for plant disease control

The incidence and geographic range of dengue and dengue hemorrhagic fever has increased dramatically in recent decades. With 2.5 billion people now living in areas at risk for epidemic transmission, dengue has become the most important mosquito-borne viral disease affecting humans. Dengue virus (DENV) is a positive-strand RNA virus of the family Flaviviridae. It exists as four closely related but antigenically distinct serotypes, all of which have Aedes aegypti mosquitoes as their primary vector, with A. albopictus as a secondary vector.

Mosquitoes, like all insects, are exposed to a variety of microbes in their natural habitats, and possess an innate immune system that is capable of mounting a potent response against microbial challenge. The insect innate immune response is largely regulated by three main immune signaling pathways: the Toll, immune deficiency (IMD) and Janus kinase signal transducer and activator of transcription (JAK-STAT) pathways. The Toll pathway is involved in defense against fungi, Gram-positive bacteria, and viruses, and has been found to be specifically involved in the A. aegypti anti-DENV response.

In order to study the interaction of DENV with the A. aegypti immune response, researchers have characterized the DENV infection-responsive transcriptome of the immune-competent A. aegypti cell line. As in mosquitoes, DENV infection transcriptionally activated the cell line Toll pathway and a variety of cellular physiological systems. Most notably, however, DENV infection down-regulated the expression levels of numerous immune signaling molecules and antimicrobial peptides (AMPs). Functional assays showed that transcriptional induction of AMPs from the Toll and IMD pathways in response to bacterial challenge is impaired in DENV-infected cells. In addition, Escherichia coli, a Gram-negative bacteria species, grew better when co-cultured with DENV-infected cells than with uninfected cells, suggesting a decreased production of AMPs from the IMD pathway in virus-infected cells. Pre-stimulation of the cell line with Gram-positive bacteria prior to DENV infection had no effect on DENV titers, while pre-stimulation with Gram-negative bacteria resulted in an increase in DENV titers. These results indicate that DENV is capable of actively suppressing immune responses in the cells it infects, a phenomenon that may have important consequences for virus transmission and insect physiology.

Dengue Virus Inhibits Immune Responses in Aedes aegypti Cells. 2010 PLoS ONE 5(5): e10678. doi:10.1371/journal.pone.0010678

Related:

• The dengue vector Aedes aegypti
• Dengue Virus
• Rethinking dengue hemorrhagic fever

Staphylococcus aureus is an invasive human pathogen with increasing incidence and morbidity in hospitals and the community. Both healthy persons and those with underlying illness are at risk for diverse skin and soft tissue infections, endocarditis, osteomyelitis, meningitis, bacteremia, and pneumonia (including pneumonia arising as a complication of influenza), with mortality rates ranging from 6–40%. The high frequency of poorly responsive and recurrent S. aureus disease in apparently immunocompetent hosts is a challenging feature of these infections. Groups that are particularly susceptible include children in daycare, sports teams, jailed inmates and military personnel. Moreover, the emergence and rapid spread of methicillin-resistant S. aureus (MRSA) has placed substantial burden on the healthcare system.

Colonization of the nares (nostrils) is a potent and increasingly prevalent risk factor for subsequent S. aureus infection. In at least 80% of S. aureus bacteremia cases in colonized subjects, the infecting strain is identical to a nasal colonizing strain detected prior to onset of bacteremia. Followed longitudinally, approximately 20–30% of persons are colonized persistently with S. aureus, 30% are colonized intermittently, and 50% never, or rarely, are colonized. Why some individuals apparently are resistant to colonization, and thus at lower risk of infection, remains an open question. Understanding the biology of this pathogen, especially its ecological niche in humans and the initial step in infection, colonization, may therefore provide new methods of limiting disease.

The Human Nasal Microbiota and Staphylococcus aureus Carriage. 2010 PLoS ONE 5(5): e10598. doi:10.1371/journal.pone.0010598
Nasal specimens were collected longitudinally from five healthy adults and a cross-section of hospitalized patients (26 S. aureus carriers and 16 non-carriers). Culture-independent analysis of 16S rRNA sequences revealed that the nasal microbiota of healthy subjects consists primarily of members of the phylum Actinobacteria (e.g., Propionibacterium spp. and Corynebacterium spp.), with proportionally less representation of other phyla, including Firmicutes (e.g., Staphylococcus spp.) and Proteobacteria (e.g. Enterobacter spp). In contrast, inpatient nasal microbiotas were enriched in S. aureus or Staphylococcus epidermidis and diminished in several actinobacterial groups, most notably Propionibacterium acnes. Moreover, within the inpatient population S. aureus colonization was negatively correlated with the abundances of several microbial groups, including S. epidermidis. The nares environment is colonized by a temporally stable microbiota that is distinct from other regions of the integument. Negative association between S. aureus, S. epidermidis, and other groups suggests microbial competition during colonization of the nares, a finding that could be exploited to limit S. aureus colonization.

Related:

• Staphylococcus aureus: superbug
• MRSA: Methicillin-resistant Staphylococcus aureus
• Evolution and pathogenesis of Staphylococcus aureus