MicrobiologyBytes

Eucaryotic organisms depend on networks of gene regulatory pathways. Small RNAs (sRNAs), are key components of these networks. sRNAs are short (21–24 nt in length), endogenously expressed, and are processed from double stranded (ds)RNAs or dsRNA-like precursors. In both plants and animals, sRNAs exert their functions upon incorporation into ribonucleoprotein silencing complexes and through their base-paring capacity. They are implicated in a variety of processes, including post-transcriptional regulation of mRNA, mRNA stability and availability for translation, establishment of heterochromatin and silencing of transposons. Different classes of sRNAs differ in the proteins required in their biogenesis, the constitution of ribonucleoprotein complexes that mediate their regulatory functions, their type of gene regulation, and the biological functions in which they are implicated. Plants display a remarkable diversity of sRNA types and sRNA pathways, likely needed for managing multiple environmental stimuli, including biotic and abiotic stresses. Several lines of evidence suggest that plant sRNAs play critical roles in plant–pathogen interactions. Indeed, upon infection, most plant pathogens can interfere with the expression of endogenous sRNAs, thus altering the expression of specific host factors implicated in the suppression or in the activation of plant defences. Evidence for these phenomena has been reported for bacterial and fungal pathogens.

Viruses are obligate infectious agents, whose life cycle (expression of viral proteins, viral genome replication and virion assembly) is integrated with host cell functions. Plant viruses can both modify the profiles of endogenous sRNAs (in common with bacteria and fungi) and induce the production of additional sRNAs derived from their own genomes (viral sRNAs; vsiRNAs). The latter gives a clear indication of the activation of RNA silencing-based responses of the plant. In some cases, this results in reduction of the titre of the invading virus and, in recovery of upper, non-inoculated leaves. To counteract RNA silencing, many plant viruses have evolved proteins (viral suppressors of RNA silencing: VSR) that target various components of the plant silencing machinery. Viruses can induce specific symptoms resembling developmental anomalies and affecting organs and tissues such as leaves, flowers and fruits. These anomalies are often reconcilable with virus-induced alterations of RNA silencing-based endogenous pathways, due to: i) the direct activity of VSRs on endogenous sRNAs or on silencing related effectors; ii) the abundance of vsiRNAs in competition with endogenous sRNAs; iii) the action of specific vsiRNAs entering into RNA silencing complexes and targeting specific host genes.

This review provides an overview of the major cellular RNA silencing pathways in plants with particular reference to those involved in antiviral functions and highlights examples of the complex interactions between viral molecular processes and host RNA processes.

Viral induction and suppression of RNA silencing in plants. Biochim Biophys Acta. Apr 30 2011
RNA silencing in plants and insects can function as a defence mechanism against invading viruses. RNA silencing-based antiviral defence entails the production of virus-derived small interfering RNAs which guide specific antiviral effector complexes to inactivate viral genomes. As a response to this defence system, viruses have evolved viral suppressors of RNA silencing (VSRs) to overcome the host defence. VSRs can act on various steps of the different silencing pathways. Viral infection can have a profound impact on the host endogenous RNA silencing regulatory pathways; alterations of endogenous short RNA expression profile and gene expression are often associated with viral infections and their symptoms. Here we discuss our current understanding of the main steps of RNA-silencing responses to viral invasion in plants and the effects of VSRs on endogenous pathways. This article is part of a Special Issue entitled: MicroRNAs in viral gene regulation.

Tweet

Staphylococcus aureus is an important human pathogen that is found in the nasal passages of approximately 1/3 of the population. The nose serves as a reservoir for spread of this pathogen and predisposes the host to potential infection. Factors contributing to S. aureus nasal colonization are only beginning to be understood. The collection and analysis of human nasal secretions provided evidence that the presence of haemoglobin in nasal secretions can promote S. aureus nasal colonization. Hemoglobin reduced expression of the S. aureus agr quorum sensing regulatory system known to be involved in surface colonization, and it was found that induction of the agr system reduced nasal colonization. These findings suggest that individuals experiencing frequent nosebleeds would be prone to S. aureus colonization and epidemiological data supports these findings. By understanding host factors and bacterial molecular mechanisms involved in nasal colonization we may one day be able to design novel decolonization strategies.

Hemoglobin Promotes Staphylococcus aureus Nasal Colonization. (2011) PLoS Pathog 7(7): e1002104. doi:10.1371/journal.ppat.1002104
Staphylococcus aureus nasal colonization is an important risk factor for community and nosocomial infection. Despite the importance of S. aureus to human health, molecular mechanisms and host factors influencing nasal colonization are not well understood. To identify host factors contributing to nasal colonization, we collected human nasal secretions and analyzed their ability to promote S. aureus surface colonization. Some individuals produced secretions possessing the ability to significantly promote S. aureus surface colonization. Nasal secretions pretreated with protease no longer promoted S. aureus surface colonization, suggesting the involvement of protein factors. The major protein components of secretions were identified and subsequent analysis revealed that hemoglobin possessed the ability to promote S. aureus surface colonization. Immunoprecipitation of hemoglobin from nasal secretions resulted in reduced S. aureus surface colonization. Furthermore, exogenously added hemoglobin significantly decreased the inoculum necessary for nasal colonization in a rodent model. Finally, we found that hemoglobin prevented expression of the agr quorum sensing system and that aberrant constitutive expression of the agr effector molecule, RNAIII, resulted in reduced nasal colonization of S. aureus. Collectively our results suggest that the presence of hemoglobin in nasal secretions contributes to S. aureus nasal colonization.

Tweet

Noroviruses (NoVs) are a major cause of epidemic acute gastroenteritis affecting millions of people worldwide. Infection by NoVs relies on recognition of human histo-blood group antigens (HBGAs) as ligands or receptors for attachment, an early infection event that most likely controls host susceptibility and resistance to NoVs. HBGAs are complex carbohydrates on red blood cells, mucosal epithelia, saliva, milk and other body fluids, which are highly polymorphic and are related to the ABO, secretor and Lewis families. The interaction between NoVs and HBGAs has been extensively studied since it was discovered in 2002. Early studies using variable in vitro binding assays to measure the binding of NoV-like particles (VLPs) with HBGAs revealed diverse binding patterns. Further studies in human volunteer challenge and outbreak investigations provided direct evidence on the linkage between host blood types and susceptibility to NoV infection and, therefore, implicated HBGAs as NoV receptors or co-receptors, although undefined results were also reported.

Recent structural and functional analysis of the HBGA binding interfaces of NoVs has significantly advanced our understanding of the complicated interaction between NoVs and human HBGAs. For the first time, crystal structures of the HBGA binding interfaces of representative NoVs in complex with different HBGAs precisely elucidated interactions between individual amino acids in the binding interfaces of NoVs with specific saccharides of HBGAs. This article summarizes these advancements, proposes a new model of NoV–HBGA interaction, and discusses the role of HBGAs in NoV evolution and the resulting impacts on epidemiology and classification of human NoVs.

Norovirus-host interaction: Multi-selections by human histo-blood group antigens. Trends Microbiol. Jun 24 2011
The discovery of human histo-blood group antigens (HBGAs) as receptors or ligands of noroviruses (NoVs) raises a question about the potential role of host factors in the evolution and diversity of NoVs. Recent structural analysis of selected strains in the two major genogroups of human NoVs (GI and GII) demonstrated highly conserved HBGA binding interfaces within the two groups but not between them, indicating convergent evolution of GI and GII NoVs. GI and GII NoVs are probably introduced to humans from different non-human hosts with the HBGAs as a common niche. Each genogroup has further diverged into multiple sub-lineages (genotypes) through selections by the polymorphic HBGAs of the hosts. An elucidation of such pathogen–host interaction, including determination of the phenotypes of NoV–HBGAs interaction for each genotype, is important in understanding the epidemiology, classification and disease control and prevention of NoVs. A model of this multi-selection of NoVs by HBGAs is proposed.

Tweet

Before the genomic era there was already a longstanding interest in understanding the origins of bacterial pathogens and the molecular attributes of virulence. Large-scale genome sequencing has provided a rapid and unbiased means of uncovering the evolution of many pathogens, contributing to both fundamental microbiological insights and the development of new disease-control strategies. For these reasons, the evolution of one of the most devastating human pathogens, Mycobacterium tuberculosis, has captivated researchers since its discovery in 1882. This interest was stimulated not only by the epidemiologic importance of the pathogen but also by the lack of consensus on its origins and its apparent exception to the stereotypes of bacterial evolution (e.g. acquisition of pathogenicity islands). So where did M. tuberculosis come from?

The rise and fall of the Mycobacterium tuberculosis genome. Trends Microbiol. 2011 19(4): 156-161
When studied from the perspective of non-tuberculous mycobacteria (NTM) it is apparent that Mycobacterium tuberculosis has undergone a biphasic evolutionary process involving genome expansion (gene acquisition and duplication) and reductive evolution (deletions). This scheme can instruct descriptive and experimental studies that determine the importance of ancestral events (including horizontal gene transfer) in shaping the present-day pathogen. For example, heterologous complementation in an NTM can test the functional importance of M. tuberculosis-specific genetic insertions. An appreciation of both phases of M. tuberculosis evolution is expected to improve our fundamental understanding of its pathogenicity and facilitate the evaluation of novel diagnostics and vaccines.

Tweet

MicroRNAs (miRNAs) are the subject of enormous interest. They are small non-coding RNAs that play a regulatory role in numerous and diverse cellular processes such as immune function, apoptosis and tumorigenesis. Several virus families have been shown to encode miRNAs, and an appreciation for their roles in the viral infectious cycle continues to grow. Despite the identification of numerous (>225) viral miRNAs, an in depth functional understanding of most virus-encoded miRNAs is lacking. This review focuses on a few viral miRNAs with well-defined functions and uses these examples to extrapolate general themes of viral miRNA activities including autoregulation of viral gene expression, avoidance of host defenses, and a likely important role in maintaining latent and persistent infections. Although the molecular mechanisms and machinery are similar, the majority of viral miRNAs may utilize a target strategy that differs from host miRNAs. That is, many viral miRNAs may have evolved to regulate viral-encoded transcripts or networks of host genes that are unique to viral miRNAs. Included in this latter category is a likely abundant class of viral miRNAs that may regulate only one or a few principal host genes. Key steps forward for the field are discussed, including the need for additional functional studies that utilize surgical viral miRNA mutants combined with relevant models of infection.

Virus-encoded microRNAs. Virology. 2011 411(2):325-343

Papillomaviruses (PV) replicate exclusively in the terminally differentiating epidermal cells of skin and mucosa in a strictly species-specific manner. They infect the basal cells of this tissue, to which they probably gain access via microlesions. Following delivery of virus DNA to the nucleus of infected cells, infection is established by the initial amplification of the viral genome as nuclear minichromosomes. The infection spreads by cell division during which the viral genome copy number per cell remains constant. The viral life cycle is completed by amplification of the viral genome in fully differentiated cells, structural (late) gene expression, and viral assembly. Progeny virions are shed within the dead squames of the terminally differentiating keratinocytes. Human PV (HPV) comprises a large group of viruses with more than 100 genotypes identified so far. They usually induce benign self limiting tumors of skin and mucosa, which rarely progress to carcinomas of the cervix, vagina, penis, anus and others. Progression usually requires persistent infection by high risk types, e.g. HPV16 and HPV18. Due to the high incidence of HPV in the general population, HPV infection is associated with more than 7% and 1% of cancers in women and men respectively.

The strict dependence of PV on terminally differentiating keratinocytes for completion of their replication cycle initially made the study of entry processes difficult. It was impossible to propagate virions in cell culture, and virus yields from natural lesions were low. In vitro data backed by in vivo studies suggest an elaborate sequence of cell surface events that includes retrograde flow along actin protrusions towards the cell body. In addition, it has become obvious that PV have evolved unique strategies for internalization and intracellular trafficking to overcome the challenges it faces by replicating in this terminally differentiating, stratified epithelium. This mini-review summarizes recent advances in our understanding of papillomavirus’ interactions with the host cell cytoskeletal elements.

The Cytoskeleton in Papillomavirus Infection. Viruses 2011, 3, 260-271
Cytoskeleton defines the shape and structural organization of the cell. Its elements participate in cell motility, intracellular transport and chromosome movement during mitosis. Papillomaviruses (PV) are strictly epitheliotropic and induce self-limiting benign tumors of skin and mucosa, which may progress to malignancy. Like many other viruses, PV use the host cytoskeletal components for several steps during their life cycle. Prior to internalization, PV particles are transported along filopodia to the cell body. Following internalization, retrograde transport along microtubules via the dynein motor protein complex is observed. In addition, viral minichromosomes depend on the host cell machinery for partitioning of viral genomes during mitosis, which may be affected by oncoproteins E6 and E7 of high-risk human PV types. This mini-review summarizes recent advances in our understanding of papillomavirus’ interactions with the host cell cytoskeletal elements.

Tweet

A novel virus that spread through a California New World titi monkey colony in late 2009 has been shown to have also infected a human researcher and a household family member, in a documented example of an adenovirus “jumping” from one species to another and remaining contagious after the jump. Adenoviruses are known to cause a wide range of clinical illnesses in humans, from cold-like symptoms to diarrhea and pneumonia. Unlike influenza or coronaviruses, adenoviruses had previously not been known to spread from one species to another. Now adenoviruses can be added to the list of pathogens that have the ability to cross species.

The virus, which researchers have named titi monkey adenovirus (TMAdV), infected titi monkeys in the California National Primate Research Center (CNPRC) in 2009. At the time of the outbreak, a researcher caring for the sick monkeys also developed an upper respiratory infection with fever, as did two members of the researchers’ family who had no contact with the monkey colony. But which direction the virus spread – from monkeys to humans or vice versa – remains a mystery. The viral center is now conducting further studies in both humans and monkeys in Brazil and Africa to determine whether TMAdV is common in wild populations of monkeys, as well as whether it has crossed species in those settings to humans who live nearby.

Cross-Species Transmission of a Novel Adenovirus Associated with a Fulminant Pneumonia Outbreak in a New World Monkey Colony. (2011) PLoS Pathog 7(7): e1002155. doi:10.1371/journal.ppat.1002155
Adenoviruses are DNA viruses that naturally infect many vertebrates, including humans and monkeys, and cause a wide range of clinical illnesses in humans. Infection from individual strains has conventionally been thought to be species- specific. Here we applied the Virochip, a pan-viral microarray, to identify a novel adenovirus (TMAdV, titi monkey adenovirus) as the cause of a deadly outbreak in a closed colony of New World monkeys (titi monkeys; Callicebus cupreus) at
the California National Primate Research Center (CNPRC). Among 65 titi monkeys housed in a building, 23 (34%) developed upper respiratory symptoms that progressed to fulminant pneumonia and hepatitis, and 19 of 23 monkeys, or 83% of those infected, died or were humanely euthanized. Whole-genome sequencing of TMAdV revealed that this adenovirus is a new species and highly divergent, sharing ,57% pairwise nucleotide identity with other adenoviruses. Cultivation of TMAdV was successful in a human A549 lung adenocarcinoma cell line, but not in primary or established monkey kidney cells. At the onset of the outbreak, the researcher in closest contact with the monkeys developed an acute respiratory illness, with symptoms persisting for 4 weeks, and had a convalescent serum sample seropositive for TMAdV. A clinically ill family member, despite having no contact with the CNPRC, also tested positive, and screening of a set of 81 random adult blood donors from the Western United States detected TMAdV-specific neutralizing antibodies in 2 individuals (2/81, or 2.5%). These findings raise the possibility of zoonotic infection by TMAdV and human-to-human transmission of the virus in the population. Given the unusually high case fatality rate from the outbreak (83%), it is unlikely that titi monkeys are the native host species for TMAdV, and the natural reservoir of the virus is still unknown. The discovery of TMAdV, a novel adenovirus with the capacity to infect both monkeys and humans, suggests that adenoviruses should be monitored closely as potential causes of cross-species outbreaks.

Tweet

A new study has confirmed the existence of a “trial effect” in clinical trials for treatment of HIV. Trial effect is an umbrella term for the benefit experienced by study participants simply by virtue of their participating in the trial. It includes the benefit of newer and more effective treatments, the way those treatments are delivered, increased care and follow-up, and the patient’s own behavior change as a result of being under observation.

Researchers compared HIV suppression among patients who began highly active antiretroviral therapy (HAART) in a clinical trial with patients who received HAART in routine clinical care in two different time periods, 1996-1999 and 2000-2006. They found clear evidence of a trial effect during the earlier period, but not during the later period. Researchers offer that improvements to antiretroviral therapy (fewer pills and fewer side effects), and the change in attitude to HIV, which has come to be seen by many as a chronic, but treatable infection, may be among the explanations for the lack of demonstrable trial effect in the later period.

This is the first study to clearly demonstrate a trial effect in HIV clinical trials, and this has important implications moving forward. Documentation of a clinical trial effect should be considered when interpreting the generalizability of clinical trial results. At the same time, the fact that no trial effect was observed in the current HAART period argues that the efficacy demonstrated in clinical trials is likely to predict the effectiveness of the therapy in broader treatment populations. Clinicians and public health officials may have increased confidence that treatment guidelines based on clinical trial data are relevant to routine clinical care.

Does HAART Efficacy Translate to Effectiveness? Evidence for a Trial Effect. PLoS ONE 6(7): e21824. doi:10.1371/journal.pone.0021824
Background: Patients who participate in clinical trials may experience better clinical outcomes than patients who initiate similar therapy within clinical care (trial effect), but no published studies have evaluated a trial effect in HIV clinical trials.
Methods: To examine a trial effect we compared virologic suppression (VS) among patients who initiated HAART in a clinical trial versus in routine clinical care. VS was defined as a plasma HIV RNA #400 copies/ml at six months after HAART initiation and was assessed within strata of early (1996–99) or current (2000–06) HAART periods. Risk ratios (RR) were estimated using binomial models.
Results: Of 738 persons initiating HAART, 30.6% were women, 61.7% were black, 30% initiated therapy in a clinical trial and 67% (n = 496) had an evaluable six month HIV RNA result. HAART regimens differed between the early and current periods (p,0.001); unboosted PI regimens (55.6%) were more common in the early and NNRTI regimens (46.4%) were more common in the current period. Overall, 78% (95%CI 74, 82%) of patients achieved VS and trial participants were 16% more likely to achieve VS (unadjusted RR 1.16, 95%CI 1.06, 1.27). Comparing trial to non-trial participants, VS differed by study period. In the early period, trial participants initiating HAART were significantly more likely to achieve VS than non-trial participants (adjusted RR 1.33; 95%CI 1.15, 1.54), but not in the current period (adjusted RR 0.98; 95%CI 0.87, 1.11).
Conclusions: A clear clinical trial effect on suppression of HIV replication was observed in the early HAART period but not in the current period.

Tweet

Iron is required for Staphylococcus aureus growth and persistence and hence must be acquired during infection. Most vertebrate iron is utilized as a cofactor in biochemical reactions that occur intracellularly. This intracellular pool of iron is generally not available to extracellular pathogens such as S. aureus. Moreover, the amount of free iron found within the serum is negligible, as it is usually complexed to high-affinity iron-binding proteins. This process of iron sequestration by the host, also referred to as nutritional immunity, inhibits the growth of invading microorganisms. In response to this severe iron limitation, S. aureus has evolved sophisticated strategies to obtain iron required to proliferate within vertebrates. This review provides a comprehensive analysis of the pathways S. aureus utilizes to obtain iron during infection.

Molecular Mechanisms of Staphylococcus aureus Iron Acquisition. Annual Review of Microbiology June 2, 2011 doi: 10.1146/annurev-micro-090110-102851
The unique redox potential of iron is an ideal cofactor in diverse biochemical reactions. Iron is therefore vital for the growth and proliferation of nearly all organisms, including pathogenic bacteria. Vertebrates sequester excess iron within proteins in order to alleviate toxicity and restrict the amount of free iron available for invading pathogens. Restricting the growth of infectious microorganisms by sequestering essential nutrients is referred to as nutritional immunity. In order to circumvent nutritional immunity, bacterial pathogens have evolved elegant systems that allow for the acquisition of iron during infection. The gram-positive extracellular pathogen Staphylococcus aureus is a commensal organism that can cause severe disease when it gains access to underlying tissues. Iron acquisition is required for S. aureus colonization and subsequent pathogenesis. Herein we review the strategies S. aureus employs to obtain iron through the production of siderophores and the consumption of host heme.

Tweet