Effects of sea-level rise on physiological ecology of populations of a ground-dwelling ant.

INTRODUCTION: Sea-level rise is a consequence of climate change that can impact the ecological and physiological changes of coastal, ground-dwelling species. Sea-level rise has a potential to inundate birds, rodents, spiders, and insects that live on the ground in coastal areas. Yet, there is still much to be learned concerning the specifics of these impacts. The red imported fire ant Solenopsis invicta (Buren) excavates soil for its home and is capable of surviving flooding. Because of their ground-dwelling life history and rapid reproduction, fire ants make an ideal model for discovery and prediction of changes that may be due to sea-level rise. There are up to 500,000 individuals in a colony, and these invasive ants naturally have a painful sting. However, observations suggest that colonies of fire ants that dwell in tidally-influenced areas are more aggressive with more frequent stings and more venom injected per sting (behavioral and physiological changes) than those located inland. This may be an adaption to sea-level rise. Therefore, the objective of this study is to elucidate differences in inland and coastal defensiveness via micro-dissection and comparison of head width, head length, stinger length, and venom sac volume. But first because fire ants' ability to raft on brackish tidal water is unknown, it had to be determined if fire ants could indeed raft in brackish water and examine the behavior differences between those flooded with freshwater vs. saltwater. METHODS: To test the coastal-aggression hypothesis, inland colonies and coastal colonies, which experience relatively greater amounts of flooding, specifically regular tidal and windblown water and oscillations (i.e. El Nino Southern Oscillation) from the Gulf of Mexico, were collected. To mimic sea-level rise, the colonies were flooded in salinities that correspond to both their collection site and conditions found in a variety of locales and situations (such as storm surge from a tropical storm). Individual ants were immediately taken from each colony for dissection before flooding, 1-hour into flooding, and 24-hours into flooding. RESULTS AND DISCUSSION: Fire ants use their venom to defend themselves and to communicate alarm or aggression. Dissections and measurement of heads, venom sacs, and stingers revealed both coastal and inland colonies experience an increase in venom sac volume after 24 hours; in fact coastal colonies increased their venom volume by 75% after 24 h of flooding Whether this venom sac enlargement is due to diffusion of water or venom sac production is unknown. These ground-dwelling ants exhibit physiological and behavioral adaptations to ongoing sea-level rise possibly indicating that they are responding to increased flooding. Fire ants will raft on high-salinity water; and sea-level rise may cause stings by flooded ants to be more severe because of increased venom volume.

Immune homeostasis, dysbiosis and therapeutic modulation of the gut microbiota.

The distal gut harbours ∼10(13) bacteria, representing the most densely populated ecosystem known. The functional diversity expressed by these communities is enormous and relatively unexplored. The past decade of research has unveiled the profound influence that the resident microbial populations bestow to host immunity and metabolism. The evolution of these communities from birth generates a highly adapted and highly personalized microbiota that is stable in healthy individuals. Immune homeostasis is achieved and maintained due in part to the extensive interplay between the gut microbiota and host mucosal immune system. Imbalances of gut microbiota may lead to a number of pathologies such as obesity, type I and type II diabetes, inflammatory bowel disease (IBD), colorectal cancer (CRC) and inflammaging/immunosenscence in the elderly. In-depth understanding of the underlying mechanisms that control homeostasis and dysbiosis of the gut microbiota represents an important step in our ability to reliably modulate the gut microbiota with positive clinical outcomes. The potential of microbiome-based therapeutics to treat epidemic human disease is of great interest. New therapeutic paradigms, including second-generation personalized probiotics, prebiotics, narrow spectrum antibiotic treatment and faecal microbiome transplantation, may provide safer and natural alternatives to traditional clinical interventions for chronic diseases. This review discusses host-microbiota homeostasis, consequences of its perturbation and the associated challenges in therapeutic developments that lie ahead.

Involvement of Streptococcus mutans regulator RR11 in oxidative stress response during biofilm growth and in the development of genetic competence.

AIMS: To identify the genes regulated by RR11, the regulator of the Streptococcus mutans HK/RR11 two-component system. METHODS AND RESULTS: The S. mutans RR11-encoding gene was inactivated, and the effects of gene disruption on the cell's ability to form biofilms under stresses and acquire extracellular DNA were tested. Biofilm was reduced in cells lacking RR11 following exposure to oxidative stress. RR11-defective cells showed approx. 20-fold reduction in transformation efficiency. Microarray used to decipher the RR11-regulated genes in biofilm showed that approx. 5% of the UA159 genome underwent a significant change in expression. RR11 was found to regulate 174 genes, including genes involved in competence, stress-response and cell division. CONCLUSIONS: Target genes controlled by RR11during biofilm growth have been identified by a comparison of transcriptional profiles between an RR11 defective mutant and the parental strain. The results demonstrated that RR11 is involved in the control of diverse cellular processes, including the formation of biofilm under oxidative stress and development of genetic competence. SIGNIFICANCE AND IMPACT OF THE STUDY: The regulator of HK/RR11 system controls a large regulon and is an important regulator involved in stress response during S. mutans biofilm growth enabling the survival and persistence of its progeny in the microbial community.

Mutation screening of the TNFRSF11A gene encoding receptor activator of NF kappa B (RANK) in familial and sporadic Paget's disease of bone and osteosarcoma.

Paget's disease of bone (PDB) is a common disorder characterized by focal areas of increased and disorganized osteoclastic bone resorption, leading to bone pain, deformity, pathological fracture, and an increased risk of osteosarcoma. Genetic factors play an important role in the pathogenesis of Paget's disease. In some families, the disease has been found to be linked to a susceptibility locus on chromosome 18q21-22, which also contains the gene responsible for familial expansile osteolysis (FEO)--a rare bone dysplasia with many similarities to Paget's disease. Insertion mutations of the TNFRSF11A gene encoding Receptor Activator of NF kappa B (RANK) have recently been found to be responsible for FEO and rare cases of early onset familial Paget's disease. Loss of heterozygosity (LOH) affecting the PDB/FEO critical region has also been described in osteosarcomas suggesting that TNFRSF11A might also be involved in the development of osteosarcoma. In order to investigate the possible role of TNFRSF11A in the pathogenesis of Paget's disease and osteosarcoma, we conducted mutation screening of the TNFRSF11A gene in patients with familial and sporadic Paget's disease as well as DNA extracted from Pagetic bone lesions, an osteosarcoma arising in Pagetic bone and six osteosarcoma cell lines. No specific abnormalities of the TNFRSF11A gene were identified in a Pagetic osteosarcoma, the osteosarcoma cell lines, DNA extracted from Pagetic bone lesions, or DNA extracted from peripheral blood in patients with familial or sporadic Paget's disease including several individuals with early onset Paget's disease. These data indicate that TNFRSF11A mutations contribute neither to the vast majority of cases of sporadic or familial PDB, nor to the development of osteosarcoma.

The complexity of simplicity.

What is the minimum number of genes or functions necessary to support cellular life? The concept of a 'minimal genome' has become popular, but is it a useful concept, and if so, what might a minimal genome encode? We argue that the concept may be useful, even though the goal of defining a general minimal genome may never be attained.

Global transposon mutagenesis and a minimal Mycoplasma genome.

Mycoplasma genitalium with 517 genes has the smallest gene complement of any independently replicating cell so far identified. Global transposon mutagenesis was used to identify nonessential genes in an effort to learn whether the naturally occurring gene complement is a true minimal genome under laboratory growth conditions. The positions of 2209 transposon insertions in the completely sequenced genomes of M. genitalium and its close relative M. pneumoniae were determined by sequencing across the junction of the transposon and the genomic DNA. These junctions defined 1354 distinct sites of insertion that were not lethal. The analysis suggests that 265 to 350 of the 480 protein-coding genes of M. genitalium are essential under laboratory growth conditions, including about 100 genes of unknown function.

bca: an activation-related B-cell gene.

We have identified a novel activation related B-cell gene (bca) through differential hybridization screening of a murine B cell cDNA library. The deduced amino acid sequence predicted a protein of 482 amino acids with strong sequence similarity to the SH2 and SH3 domains present within the non-catalytic regions of several protein tyrosine kinases. Northern analysis of RNA from several murine B-cell lines revealed a transcript of 1.8 kb, which was not detected in T-cell and non-lymphoid cell lines. bca was transcribed at low levels in resting spleen cells from a variety of normal mouse strains and was strongly expressed in kidney RNA. bca expression was markedly increased in RNA prepared from mitogen activated B cells, and in freshly isolated spleen and lymph node cells of MRL/lpr and NZB autoimmune strains. The unique sequence of bca, which bears no obvious similarity to any specific class of proteins containing SH2 and SH3 domains, suggests that this gene encodes a novel protein potentially involved in B-cell signal transduction.

Identification of a novel RalGDS-related protein as a candidate effector for Ras and Rap1.

Although Ras and Rap1 share interaction with common candidate effector proteins, Rap1 lacks the transforming activity exhibited by Ras proteins. It has been speculated that Rap antagonizes Ras transformation through the formation of nonproductive complexes with critical Ras effector targets. To understand further the distinct biological functions of these two closely related proteins, we searched for Rap1b-binding proteins by yeast two-hybrid screening. We identified multiple clones that encode the COOH-terminal sequences of a protein that shares sequence identity with RalGDS and RGL, which we have designated RGL2. A 158-amino acid COOH-terminal fragment of RGL2 (RGL2 C-158) bound to Ras superfamily proteins which shared identical effector domain sequences with Rap1 (Ha-Ras, R-Ras, and TC21). RGL2 C-158 binding was impaired by effector domain mutations in Rap1b and Ha-Ras. Furthermore, RGL2 C-158 bound exclusively to the GTP-, but not the GDP-bound form of Ha-Ras. Finally, coexpression of RGL2 C-158 impaired oncogenic Ras activation of transcription from a Ras-responsive promoter element and focus-forming activity in NIH 3T3 cells. We conclude that RGL2 may be an effector for Ras and/or Rap proteins.

Characterization of repetitive DNA in the Mycoplasma genitalium genome: possible role in the generation of antigenic variation.

We have characterized a family of repetitive DNA elements with homology to the MgPa cellular adhesion operon of Mycoplasma genitalium, a bacterium that has the smallest known genome of any free-living organism. One element, 2272 bp in length and flanked by DNA with no homology to MgPa, was completely sequenced. At least four others were partially sequenced. The complete element is a composite of six regions. Five of these regions show sequence similarity with nonadjacent segments of genes of the MgPa operon. The sixth region, located near the center of the element, is an A+T-rich sequence that has only been found in this repeat family. Open reading frames are present within the five individual regions showing sequence homology to MgPa and the adjacent open reading frame 3 (ORF3) gene. However, termination codons are found between adjacent regions of homology to the MgPa operon and in the A+T-rich sequence. Thus, these repetitive elements do not appear to be directly expressible protein coding sequences. The sequence of one region from five different repetitive elements was compared with the homologous region of the MgPa gene from the type strain G37 and four newly isolated M. genitalium strains. Recombination between repetitive elements of strain G37 and the MgPa operon can explain the majority of polymorphisms within our partial sequences of the MgPa genes of the new isolates. Therefore, we propose that the repetitive elements of M. genitalium provide a reservoir of sequence that contributes to antigenic variation in proteins of the MgPa cellular adhesion operon.

The minimal gene complement of Mycoplasma genitalium.

The complete nucleotide sequence (580,070 base pairs) of the Mycoplasma genitalium genome, the smallest known genome of any free-living organism, has been determined by whole-genome random sequencing and assembly. A total of only 470 predicted coding regions were identified that include genes required for DNA replication, transcription and translation, DNA repair, cellular transport, and energy metabolism. Comparison of this genome to that of Haemophilus influenzae suggests that differences in genome content are reflected as profound differences in physiology and metabolic capacity between these two organisms.

Genetic map of the Mycoplasma genitalium chromosome.

At 600 kb, the genome of Mycoplasma genitalium is among the smallest known for cellular organisms capable of independent replication. As such, elucidation of the genetic makeup and chromosome architecture of this organism is of considerable interest. We have located 631 markers on the physical map of M. genitalium. The clones have been mapped by hybridizing 20 overlapping cosmid and lambda clones which encompass the entire M. genitalium chromosome to replica filters containing 856 genomic DNA clones. Three hundred fifty-six of these clones represent sequence tag sites, which were previously characterized by database searches. The remaining markers represent clones with an average size of 2.5 kb derived from Sau3A1 partial digestion of genomic DNA. The hybridization data can be divided into three classes: clones which hybridized to only one cosmid; clones which hybridized to two adjacent and overlapping cosmids; and clones which hybridized to several cosmids, which represent repetitive DNA. This rapid approach for placing clones on the physical map has allowed useful comparisons to be made with other bacterial chromosomes, especially that of the closely related organism M. pneumoniae, and has provided insight to the types of events which may have led to the reduction in size of this genome. Future use of these data is discussed.

Cyclic AMP-dependent activation of Rap1b.

Rap1 proteins belong to the Ras superfamily of small molecular weight GTP-binding proteins. Although Rap1 and Ras share approximately 50% overall amino acid sequence identity, the effector domains of the two proteins are identical, suggesting either similar or antagonistic signaling roles. Several pathways leading to Ras activation have been defined, including those initiated by agonist binding to tyrosine kinase or Gi-coupled receptors. Nothing is known about such events for Rap1 proteins. The cAMP-mediated inhibition of Ras-dependent MAP kinase activation is well documented and resembles that caused by expression of GTPase-deficient Rap1. We have developed a system whereby signals leading to Rap1b activation, i.e. an increase in Rap1b-bound GTP/GDP ratio, can be measured. We report here that treatment of cells with agents that elevate intracellular cAMP levels result in Rap1b activation. These results demonstrate for the first time agonist-dependent activation of Rap1 proteins.

Construction of an ordered genomic library of Mycoplasma genitalium.

As a first step towards sequencing the chromosome of the suspected human pathogen Mycoplasma genitalium, we attempted to clone its entire genome in a set of ordered cosmids. Cosmid libraries were established by partial digestion of M. genitalium genomic DNA with Sau3AI or EcoRI. A chromosome-walking strategy was used to identify 20 overlapping cosmid clones which contained over 99% of the genome. The final 5.1 kb could not be cloned in cosmids, and was eventually obtained from a genomic library established in a lambda vector. Correspondence of cloned and genomic EcoRI fragments indicated no detectable major deletions or rearrangements in the library. The library was oriented on established XhoI and SmaI physical maps of the chromosome with restriction sites present at the expected locations in the library. The genome contained 74 EcoRI fragments which added up to a total genome size of 578 kb. These were arranged in a partial EcoRI physical map, and those containing the MgPa major attachment protein-encoding operon and its repeat sequences were identified. The existence of this ordered genomic library, which accurately and completely encompasses the entire M. genitalium genome, should serve as a valuable tool for many future studies of this organism and facilitate our long-term goal of sequencing its genome.

Rodent L1 evolution has been driven by a single dominant lineage that has repeatedly acquired new transcriptional regulatory sequences.

All mammalian genomes contain approximately 100,000 copies of the transposable element LINES-1 (L1). Phylogenetic analysis indicates that the L1 progenitor predates the mammalian radiation; since that time, the open reading frames encoded in L1 have evolved under selection. The least conserved regions within L1 are the 5'-terminal transcriptional regulatory sequences. In rodents, four types of L1 elements (A, F, and V from mouse and R from rat) have been defined according to the type of apparently nonhomologous promoter sequence present at the 5' end. In this study, we investigate the relationships between these four types of promoters. DNA sequence was determined from approximately 1.5-kb regions from the 5' ends of seven F- and three V-type L1 elements. These sequences were aligned with 29 previously reported L1 elements. Phylogenetic analysis was then performed on the homologous regions of the alignment. The results indicate that in mouse all of the A-, F-, and V-type elements belong to a single dominant lineage but were inserted into the genome during different time periods; V-type elements are the oldest, while A-type elements are the most recently inserted. V-type elements also appear ancestral to the R-type elements found in rat and therefore were replicatively competent prior to the divergence of rat and mouse. Analysis of sequence identity indicates that the different 5' promoters did not derive from a common ancestor. Therefore, the dominant L1 lineage appears to have acquired novel promoter sequences from non-L1 sources. Transposable elements from a wide range of species show similar structural rearrangements, suggesting that acquisition of new sequences may be a common theme in their evolution.

A survey of the Mycoplasma genitalium genome by using random sequencing.

A total of 508 random clones from five Mycoplasma genitalium genomic libraries were partially sequenced and analyzed. This resulted in the identification of 291 unique contigs. Sequence information from these clones (100,993 nucleotides), representing approximately 17% of this pathogen's genome, was analyzed by comparison to the DNA and protein sequence data bases. The frequency with which clones could be identified, by virtue of possessing homology to another data base entry, was 46%. Sequence analysis indicated the following. (i) The M. genitalium genome contains many genes involved in various metabolic processes. (ii) Repetitive DNA may comprise as much as 4% of this genome. (iii) The MgPa adhesin gene may be the result of horizontal transfer from an unknown origin. (iv) Not all dinucleotide pairs are present in this genome at the expected frequency. (v) This genome potentially encodes approximately 390 proteins and makes very efficient use of its limited amount of DNA. In addition, this study allowed us to estimate the number of genes involved with various cellular functions.

A random sequencing approach for placing markers on the physical map of Mycoplasma genitalium.

A physical map of the Mycoplasma genitalium genome has been prepared using pulsed-field gel electrophoresis. This report details recent efforts made to add markers or specific loci to this map in the absence of any mutants or system of genetic exchange. A total of 44 random clones were partially sequenced. Computer analysis was performed in an attempt to identify homologies with genes already recorded in the DNA sequence database. Clones with a large extent of homology to genes from other microorganisms have been assigned to specific loci on the M. genitalium map by hybridization to selected restriction digests. The additional data has facilitated an updated version of the physical map, and verified this random sequencing method as a useful mapping procedure as well as offering new insight into the physiological processes of this fastidious organism.