MicrobiologyBytes

Lentiviruses are mammalian retroviruses known to infect cattle, cats, horses, sheep, and primates. They are the focus of intense study due to their causative association with AIDS in human. Although our knowledge on the origin and early evolution of HIV has grown exponentially over the past few years, much remains unresolved about the deeper relationships between primate and non-primate lentiviruses, the origin of lentiviruses, and their mode of structural evolution over long periods of evolutionary time. This is because these viruses evolve extremely rapidly, in a conflicting relationship with their hosts, and while their high mutation rate provides a wealth of information documenting their recent history, it also quickly erases evidence of their deeper ancestry. The lifecycle of retroviruses is atypical compared to other viruses in that after appropriate receptor recognition and entry in a specific cell type, their RNA genome is reverse transcribed into double-stranded DNA and integrated into the host genome as a provirus. Occasionally this process can take place in the host germline, and the integrated copy, also called endogenous retrovirus (ERV), may be transmitted vertically from parent to offspring and reach fixation in the host population. As such, ERVs constitute a fossil record of past viral infections that potentially provide an alternative way of gaining insights into the deep evolutionary history of present day exogenous retroviruses.

Although many ERVs have been characterized in mammals (e.g. 8% of the human genome), apparently very few derive from lentiviruses. Two reasons have traditionally been put forward to explain their absence in mammalian genomes: (i) they are of relatively recent evolutionary origin and endogenization has not yet commonly occurred, and/or (ii) they were not able to enter germ cells because of a very specific cell tropism. Recently however, an endogenous lentivirus called RELIK has been identified in the genome of rabbits and hares, whose germline integration was dated at least 12 millions years old. This discovery not only showed that lentiviruses were able to infiltrate mammalian germlines, but also demonstrated that this group of viruses is probably much older than what could previously be inferred based on sequence comparison of extant exogenous lentiviruses. New research now shows that a retrovirus related to HIV became stably integrated into the genomes of lemurs around 4.2 million years ago. The discovery of prosimian immunodeficiency virus (pSIV) offers new insights into the evolution of lentiviruses.

Based on “fossil” sequences collected from different lemur species, the researchers computationally reconstructed an apparently intact and complete DNA sequence for the ancestral prosimian lentivirus. The discovery that two different species of lemurs endemic to Madagascar suffered, independently and quasi-simultaneously, multiple germline infections of pSIV provides evidence that lentiviruses have repeatedly infiltrated the germline of prosimian species. These findings should allow future functional analysis of the extinct virus and advance our understanding of the biology of lentiviruses, including HIV. In addition, the characterization of this ancient lentivirus in lemurs raises the possibility that HIV-like retroviruses are still circulating today in the mammalian fauna of Madagascar.

Parallel Germline Infiltration of a Lentivirus in Two Malagasy Lemurs. 2009 PLoS Genet 5(3): e1000425
Retroviruses normally infect the somatic cells of their host and are transmitted horizontally, i.e., in an exogenous way. Occasionally, however, some retroviruses can also infect and integrate into the genome of germ cells, which may allow for their vertical inheritance and fixation in a given species; a process known as endogenization. Lentiviruses, a group of mammalian retroviruses that includes HIV, are known to infect primates, ruminants, horses, and cats. Unlike many other retroviruses, these viruses have not been demonstrably successful at germline infiltration. Here, we report on the discovery of endogenous lentiviral insertions in seven species of Malagasy lemurs from two different genera – Cheirogaleus and Microcebus. Combining molecular clock analyses and cross-species screening of orthologous insertions, we show that the presence of this endogenous lentivirus in six species of Microcebus is the result of one endogenization event that occurred about 4.2 million years ago. In addition, we demonstrate that this lentivirus independently infiltrated the germline of Cheirogaleus and that the two endogenization events occurred quasi-simultaneously. Using multiple proviral copies, we derive and characterize an apparently full length and intact consensus for this lentivirus. These results provide evidence that lentiviruses have repeatedly infiltrated the germline of prosimian species and that primates have been exposed to lentiviruses for a much longer time than what can be inferred based on sequence comparison of circulating lentiviruses. The study sets the stage for an unprecedented opportunity to reconstruct an ancestral primate lentivirus and thereby advance our knowledge of host–virus interactions.

Related:

• Phoenix from the ashes: The 5 million year old virus
• T Cell Responses to Human Endogenous Retroviruses in HIV-1 Infection
• Human endogenous retroviruses: from ancestral pathogens to bona fide genes
• HIV infection is due to the sins of the fathers
• Rooting the tree

In the last week there has been some fairly wild speculation in the media about viruses which “cause” diabetes. The fuss came from the publication of a paper which claimed to have detected virus proteins in the pancreases of diabetes patients (The prevalence of enteroviral capsid protein vp1 immunostaining in pancreatic islets in human type 1 diabetes. Diabetologia 6 March 2009). At the same time, a separate study found four rare mutations in a gene which is thought to reduce the risk of developing type 1 diabetes and may be involved in the immune response to infection with enteroviruses (Rare Variants of IFIH1, a Gene Implicated in Antiviral Responses, Protect Against Type 1 Diabetes. Science Mar 5 2009).

The press was buzzing with speculation about the chances of a vaccine to prevent diabates. Very good news for diabetics? Well not so fast. Before we look at the science, let me tell you two things about myself. First, I have two close relatives who are affected by diabetes, so this is a disease I care a lot about. Second, I’ve been in the virology business a long time – and we’ve been here before.

Subscribe to podcasts (free):
[iTunes] Enhanced podcasts & videos
[RSS] mp3 podcasts (audio only)
Play this episode: Enhanced version
Audio only

The new paper claimed to have detected “enterovirus capsid protein vp1″ in 44 out of 72 pancreases from children who had died of type-1 diabetes shortly after becoming ill, but in only three out of 50 neonatal and paediatric normal control specimens. Statistically there is a strong correlation in this study between diabetes and the presence of the virus protein, but a correlation does not indicate a cause. Are diabetics more susceptible to enterovirus infection? We don’t know. While it’s not ethically possible to satisfy Koch’s postulates in humans, we need to be very careful in inferring from small scale studies such as this one:

1. The microorganism must be found in abundance in all organisms suffering from the disease.
2. The microorganism must be isolated from a diseased organism and grown in pure culture.
3. The cultured microorganism should cause disease when introduced into a healthy organism.
4. The microorganism must be reisolated from the inoculated, diseased experimental host and identified as being identical to the original specific causative agent.

There are over a hundred different enteroviruses and the antibody used for detection of virus protein in this study (yes, that’s right, just one non-specific antibody) does not identify the virus involved. Vaccine against diabetes? I don’t think so.

But as I said, we’ve been here before. There are reports of viruses associated with diabetes dating from the 1960s, and a very well known model of Coxsackie virus B4 causing diabetes in mice dating from the 1970s (Coxsackie Viruses and Diabetes Mellitus. BMJ 1973 November 3; 4(5887): 260–262). So does the latest work add anything new, and is a vaccine against diabetes just around the corner? No. I wish it was.

Related:

• Can you have confidence in the news?
• Media coverage affects how people perceive the threat of disease

HIV is evolving rapidly to escape the human immune system. Researchers have shown HIV is able to adapt rapidly to counter human genes controlling immune system molecules that can target it for destruction. Progression to AIDS is tied to genes which control production of key immune system molecules called human leucocyte antigens (HLAs). Humans differ in the HLA genes they have, and even small differences can have a big impact on how quickly AIDS develops. Researchers found mutations that enabled HIV effectively to neutralise the effect of a particular HLA gene were more frequent in populations with a high prevalence of that specific gene.

BBC News

Adaptation of HIV-1 to human leukocyte antigen class I. Nature, 25 February 2009
The rapid and extensive spread of the human immunodeficiency virus (HIV) epidemic provides a rare opportunity to witness host–pathogen co-evolution involving humans. A focal point is the interaction between genes encoding human leukocyte antigen (HLA) and those encoding HIV proteins. HLA molecules present fragments (epitopes) of HIV proteins on the surface of infected cells to enable immune recognition and killing by CD8+ T cells; particular HLA molecules, such as HLA-B*57, HLA-B*27 and HLA-B*51, are more likely to mediate successful control of HIV infection1. Mutation within these epitopes can allow viral escape from CD8+ T-cell recognition. Here we analysed viral sequences and HLA alleles from >2,800 subjects, drawn from 9 distinct study cohorts spanning 5 continents. Initial analysis of the HLA-B*51-restricted epitope, TAFTIPSI (reverse transcriptase residues 128–135), showed a strong correlation between the frequency of the escape mutation I135X and HLA-B*51 prevalence in the 9 study cohorts (P = 0.0001). Extending these analyses to incorporate other well-defined CD8+ T-cell epitopes, including those restricted by HLA-B*57 and HLA-B*27, showed that the frequency of these epitope variants (n = 14) was consistently correlated with the prevalence of the restricting HLA allele in the different cohorts (together, P < 0.0001), demonstrating strong evidence of HIV adaptation to HLA at a population level. This process of viral adaptation may dismantle the well-established HLA associations with control of HIV infection that are linked to the availability of key epitopes, and highlights the challenge for a vaccine to keep pace with the changing immunological landscape presented by HIV.

Related:

• Why is HIV a pathogen?
• How does HIV cause AIDS?
• Immune exhaustion in HIV infection
• HIV-2 – a kinder AIDS virus?

Genome recombination is important for its role in unlinking deleterious mutations from those that may be neutral or beneficial, allowing populations at least partial escape from the negative fitness effects of accumulating deleterious mutations. Similarly, recombination allows otherwise asexual organisms such as viruses and bacteria to avoid the evolution-retarding effects of “clonal interference”, which results from competition among distinct beneficial mutations that reside concurrently in multiple genomes – eventually one dominates within the population at the expense of the rest.

In viruses, where genetic exchange between different species or even unrelated taxa is possible, recombination is also capable of generating spectacular genetic diversity. While natural recombination between distantly related genomes has only rarely been shown to occur in double-stranded DNA and RNA viruses, it is apparently quite common amongst most reverse-transcribing, positive-sense single-stranded RNA and single-stranded DNA viruses.

Maize streak virus (MSV), the type strain of the genus Mastrevirus in the family Geminiviridae, has a simple genome consisting of virion sense movement protein (mp) and coat protein (CP) (cp) genes, and a complementary sense replication-associated protein (rep) gene that is expressed in two alternatively spliced isoforms. Separating the complementary and virion sense genes are a long intergenic region (LIR), containing the v-ori and transcriptional promoter elements, and a short intergenic region (SIR), containing the complementary sense ori and transcription termination elements. A recent paper describes a conceptually simple but powerful new experimental system to demonstrate how recombination in geminiviruses such as MSV can be a remarkably efficient mechanism capable of rapidly generating progeny genomes with increased fitness.

Rapid host adaptation by extensive recombination. 2009 J Gen Virol 90: 734-746
Experimental investigations into virus recombination can provide valuable insights into the biochemical mechanisms and the evolutionary value of this fundamental biological process. Here, we describe an experimental scheme for studying recombination that should be applicable to any recombinogenic viruses amenable to the production of synthetic infectious genomes. Our approach is based on differences in fitness that generally exist between synthetic chimaeric genomes and the wild-type viruses from which they are constructed. In mixed infections of defective reciprocal chimaeras, selection strongly favours recombinant progeny genomes that recover a portion of wild-type fitness. Characterizing these evolved progeny viruses can highlight both important genetic fitness determinants and the contribution that recombination makes to the evolution of their natural relatives. Moreover, these experiments supply precise information about the frequency and distribution of recombination breakpoints, which can shed light on the mechanistic processes underlying recombination. We demonstrate the value of this approach using the small single-stranded DNA geminivirus, maize streak virus (MSV). Our results show that adaptive recombination in this virus is extremely efficient and can yield complex progeny genomes comprising up to 18 recombination breakpoints. The patterns of recombination that we observe strongly imply that the mechanistic processes underlying rolling circle replication are the prime determinants of recombination breakpoint distributions found in MSV genomes sampled from nature.

Related:

• A feeling for the molechism
• Rolling down the road

One of the first attempts to use gene therapy to treat HIV has produced promising results in clinical trials. When the therapy was tested on 74 patients, it was shown to be safe and appeared to reduce the effect of the virus on the immune system. In theory, one treatment should be enough to replace the need for a lifetime of antiretroviral therapy. The latest therapy involves giving patients blood stem cells modified to carry a molecule called OZ1, which is designed to stop HIV reproducing itself by targeting two key proteins. The patients in the trial either received the therapy, or a dummy treatment. After 48 weeks the researchers found there was no statistically significant difference in the amount of HIV circulating in the blood of the two groups of patients. However, after 100 weeks the patients who received the gene therapy had higher levels of CD4+ cells – the key cells of the immune system which are specifically destroyed by HIV.

BBC News

Phase 2 gene therapy trial of an anti-HIV ribozyme in autologous CD34+ cells. Nature Medicine, 15 February 2009
Gene transfer has potential as a once-only treatment that reduces viral load, preserves the immune system and avoids lifetime highly active antiretroviral therapy. This study, which is to our knowledge the first randomized, double-blind, placebo-controlled, phase 2 cell-delivered gene transfer clinical trial, was conducted in 74 HIV-1–infected adults who received a tat-vpr–specific anti-HIV ribozyme (OZ1) or placebo delivered in autologous CD34+ hematopoietic progenitor cells. There were no OZ1-related adverse events. There was no statistically significant difference in viral load between the OZ1 and placebo group at the primary end point (average at weeks 47 and 48), but time-weighted areas under the curve from weeks 40–48 and 40–100 were significantly lower in the OZ1 group. Throughout the 100 weeks, CD4+ lymphocyte counts were higher in the OZ1 group. This study indicates that cell-delivered gene transfer is safe and biologically active in individuals with HIV and can be developed as a conventional therapeutic product.

Related:

• HIV “can never be cured”
• Retroviruses
• HIV-2 – a kinder AIDS virus?
• HIV mutation and the immune response
• Immune exhaustion in HIV infection
• How does HIV cause AIDS?

The acquisition of foreign DNA is a fundamental process in the diversification and adaptation of most bacterial species. Horizontal transfer of plasmids, bacteriophages, transposons, integrons, conjugative transposons, integrative conjugative elements (ICEs) and genomic islands is of particular importance to many plant and human pathogens. These integrative elements can encode functions that increase bacterial fitness under different environmental conditions or in different niches. Genomic islands are large chromosomal regions that have aberrant base composition compared to the whole genome, encode an integrase and insert at tRNA loci. The term ‘pathogenicity islands’ was first coined by Hacker to describe unstable regions present in pathogenic isolates of Escherichia coli that were unlike any previously described integrative element. The regions were called pathogenicity islands because they encoded several virulence factors. Subsequently, the term genomic island was introduced to describe regions that contained a diverse range of functions, such as (i) the ability to utilize novel carbon and nitrogen sources (metabolic islands); (ii) the ability to break down novel compounds (degradation islands); (iii) resistance to antibiotic and heavy metals (resistance islands); and (iv) the ability to cause disease (pathogenicity islands). Within each subset of islands, the gene content can vary considerably, even within a species.

Genomic islands are dynamic, ancient integrative elements in bacterial evolution. Trends Microbiol. Jan 20, 2009

Acquisition of genomic islands plays a central part in bacterial evolution as a mechanism of diversification and adaptation. Genomic islands are non-self-mobilizing integrative and excisive elements that encode diverse functional characteristics but all contain a recombination module comprised of an integrase, associated attachment sites and, in some cases, a recombination directionality factor. Here, we discuss how a group of related genomic islands are evolutionarily ancient elements unrelated to plasmids, phages, integrons and integrative conjugative elements. In addition, we explore the diversity of genomic islands and their insertion sites among Gram-negative bacteria and discuss why they integrate at a limited number of tRNA genes.

Related:

• Spread of Pathogenicity Islands in Escherichia coli
• What is a bacterial species?
• The continuing evolution of E. coli O157:H7

The species Escherichia coli comprises non-pathogenic, commensal bacterial strains belonging to the normal gut microbiota of humans and many animals, but also pathogenic strains, which cause different types of intestinal or extraintestinal infections in man and animals. Single factors and mechanisms involved in pathogenesis of extraintestinal pathogenic E. coli (ExPEC) have been analyzed in detail for many years. Horizontal gene transfer is a key step in the evolution of bacterial pathogens. Besides phages and plasmids, pathogenicity islands (PAIs) are subjected to horizontal transfer. The transfer mechanisms of PAIs within a certain bacterial species or between different species are still not well understood.

In a new study the authors applied a phylogenetic approach using multilocus sequence typing on HPI-positive and -negative natural E. coli isolates representative of the species diversity to infer the mechanism of horizontal HPI transfer within the E. coli species. In each strain, the partial nucleotide sequences of 6 HPI–encoded genes and 6 housekeeping genes of the genomic backbone, as well as DNA fragments immediately upstream and downstream of the HPI were compared. This revealed that the HPI is not solely vertically transmitted, but that recombination of large DNA fragments beyond the HPI plays a major role in the spread of the HPI within E. coli species. In support of the results of the phylogenetic analyses, they demonstrated that HPI can be transferred between different E. coli strains by F-plasmid mediated mobilization. Sequencing of the chromosomal DNA regions immediately upstream and downstream of the HPI in the recipient strain indicated that the HPI was transferred and integrated together with HPI–flanking DNA regions of the donor strain. The results of this study demonstrate for the first time that conjugative transfer and homologous DNA recombination play a major role in horizontal transfer of a pathogenicity island within the species E. coli.

Role of Intraspecies Recombination in the Spread of Pathogenicity Islands within the Escherichia coli Species. 2009 PLoS Pathog 5(1): e1000257

Related:

• The continuing evolution of E. coli O157:H7
• What is a bacterial species?

What is genomics? How will it affect our lives? In this primer on the genomics revolution, entrepreneur Barry Schuler says we can at least expect healthier, tastier food. He suggests we start with the pinot noir grape, to build better wines.

MicrobiologyBytes has discussed before Craig Venter’s attempts to create a synthetic microorganism. In 2008, Venter described his work at the TED Conference, and his talk is well worth watching:

Related:

• Mycoplasma laboratorium, the first synthetic organism
• The end of the world? Dr Franken-Venter?