Annual Review of Microbiology - Volume 55, 2001

Disulfide bonds in proteins play various important roles. They are either formed as structural features to stabilize the protein or are found only transiently as part of a catalytic or regulatory cycle. In vivo, the formation and reduction of disulfide bonds is catalyzed by specialized thiol-disulfide exchanging enzymes that contain an active site with the sequence motif Cys-X-X-Cys. These proteins have structurally evolved to catalyze predominantly either oxidative reactions or reductive steps. There is mounting evidence that, in addition to the thiol redox potential, the spatial distribution within different cell compartments and the overall redox state of the cell are equally important. In the cytoplasm, multiple pathways play overlapping roles in the reduction of disulfide bonds and additionally, the expression of several components of thiol-redox pathways was shown to respond to the changes in the cellular thiol-redox equilibrium. In the periplasm, two systems coexist, one catalyzing thiol oxidation and the other disulfide reduction. Recent results suggest that two different mechanisms are used to translocate reducing power from the cytoplasm or to dissipate the electrons after oxidation.

The mechanisms responsible for bacterial gliding motility have been a mystery for almost 200 years. Gliding bacteria move actively over surfaces by a process that does not involve flagella. Gliding bacteria are phylogenetically diverse and are abundant in many environments. Recent results indicate that more than one mechanism is needed to explain all forms of bacterial gliding motility. Myxococcus xanthus “social gliding motility” and Synechocystis gliding are similar to bacterial “twitching motility” and rely on type IV pilus extension and retraction for cell movement. In contrast, gliding of filamentous cyanobacteria, mycoplasmas, members of the Cytophaga-Flavobacterium group, and “adventurous gliding” of M. xanthus do not appear to involve pili. The mechanisms of movement employed by these bacteria are still a matter of speculation. Genetic, biochemical, ultrastructural, and behavioral studies are providing insight into the machineries employed by these diverse bacteria that enable them to glide over surfaces.

Toxic shock syndrome (TSS) is an acute onset illness characterized by fever, rash formation, and hypotension that can lead to multiple organ failure and lethal shock, as well as desquamation in patients that recover. The disease is caused by bacterial superantigens (SAGs) secreted from Staphylococcus aureus and group A streptococci. SAGs bypass normal antigen presentation by binding to class II major histocompatibility complex molecules on antigen-presenting cells and to specific variable regions on the β-chain of the T-cell antigen receptor. Through this interaction, SAGs activate T cells at orders of magnitude above antigen-specific activation, resulting in massive cytokine release that is believed to be responsible for the most severe features of TSS. This review focuses on clinical and epidemiological aspects of TSS, as well as important developments in the genetics, biochemistry, immunology, and structural biology of SAGs. From the evolutionary relationships between these important toxins, we propose that there are five distinct groups of SAGs.

A small number of prokaryotic species have a unique physiology or ecology related to their development of unusually large size. The biomass of bacteria varies over more than 10 orders of magnitude, from the 0.2 μm wide nanobacteria to the largest cells of the colorless sulfur bacteria, Thiomargarita namibiensis, with a diameter of 750 μm. All bacteria, including those that swim around in the environment, obtain their food molecules by molecular diffusion. Only the fastest and largest swimmers known, Thiovulum majus, are able to significantly increase their food supply by motility and by actively creating an advective flow through the entire population. Diffusion limitation generally restricts the maximal size of prokaryotic cells and provides a selective advantage for μm-sized cells at the normally low substrate concentrations in the environment. The largest heterotrophic bacteria, the 80 × 600 μm large Epulopiscium sp. from the gut of tropical fish, are presumably living in a very nutrient-rich medium. Many large bacteria contain numerous inclusions in the cells that reduce the volume of active cytoplasm. The most striking examples of competitive advantage from large cell size are found among the colorless sulfur bacteria that oxidize hydrogen sulfide to sulfate with oxygen or nitrate. The several-cm-long filamentous species can penetrate up through the ca 500-μm-thick diffusive boundary layer and may thereby reach into water containing their electron acceptor, oxygen or nitrate. By their ability to store vast quantities of both nitrate and elemental sulfur in the cells, these bacteria have become independent of the coexistence of their substrates. In fact, a close relative, T. namibiensis, can probably respire in the sulfidic mud for several months before again filling up their large vacuoles with nitrate.

There is ample clinical evidence, as well as evidence from animal experiments, that Mycobacterium tuberculosis can persist in tissues for months to decades without replicating, yet with the ability to resume growth and activate disease. Our knowledge of both macrophage physiology and the nature of tuberculous lesions in man and animals suggests that hypoxia is a major factor in inducing nonreplicating persistence (NRP) of tubercle bacilli. In vitro models reinforce this conclusion and provide insights into mechanisms that make NRP possible. There is evidence from in vitro models that the strategies employed by the bacilli to permit hypoxic NRP include restriction of biosynthetic activity to conserve energy, induction of alternative energy pathways, and stabilization of essential cell components to lessen the need for repair or replacement.

Quorum sensing is the regulation of gene expression in response to fluctuations in cell-population density. Quorum sensing bacteria produce and release chemical signal molecules called autoinducers that increase in concentration as a function of cell density. The detection of a minimal threshold stimulatory concentration of an autoinducer leads to an alteration in gene expression. Gram-positive and Gram-negative bacteria use quorum sensing communication circuits to regulate a diverse array of physiological activities. These processes include symbiosis, virulence, competence, conjugation, antibiotic production, motility, sporulation, and biofilm formation. In general, Gram-negative bacteria use acylated homoserine lactones as autoinducers, and Gram-positive bacteria use processed oligo-peptides to communicate. Recent advances in the field indicate that cell-cell communication via autoinducers occurs both within and between bacterial species. Furthermore, there is mounting data suggesting that bacterial autoinducers elicit specific responses from host organisms. Although the nature of the chemical signals, the signal relay mechanisms, and the target genes controlled by bacterial quorum sensing systems differ, in every case the ability to communicate with one another allows bacteria to coordinate the gene expression, and therefore the behavior, of the entire community. Presumably, this process bestows upon bacteria some of the qualities of higher organisms. The evolution of quorum sensing systems in bacteria could, therefore, have been one of the early steps in the development of multicellularity.

Advances in the elaboration of novel genomic types of β-galactosidase-positive Enterobacteriaceae and comprehensive studies of their habitats have resulted in an innovative approach to the assessment of the merits and shortcomings of the thermotrophic and fecal species Escherichia coli and all other coliforms as markers of the microbiological safety of water. As one of the consequences, it is recommended to abolish the “technical” designation fecal coliforms because their current method of detection will result in the isolation of thermotrophic organisms that have been demonstrated, beyond a doubt, to be of environmental, rather than uniquely enteric origin. Additional population studies have demonstrated that none of the coliforms can function as reliable markers for all enteric pathogens (index organisms sensu Ingram), nor be of use in validating adequate processing for safety of raw water, which represents the indicator function of markers, as defined by Ingram. Future studies along these lines will have to provide the data required to assess the suitability of additional markers for the reliable monitoring of drinking water for microbiological safety.

Biological weapons are not new. Biological agents have been used as instruments of warfare and terror for thousands of years to produce fear and harm in humans, animals, and plants. Because they are invisible, silent, odorless, and tasteless, biological agents may be used as an ultimate weapon—easy to disperse and inexpensive to produce. Individuals in a laboratory or research environment can be protected against potentially hazardous biological agents by using engineering controls, good laboratory and microbiological techniques, personal protective equipment, decontamination procedures, and common sense. In the field or during a response to an incident, only personal protective measures, equipment, and decontamination procedures may be available. In either scenario, an immediate evaluation of the situation is foremost, applying risk management procedures to control the risks affecting health, safety, and the environment. The microbiologist and biological safety professional can provide a practical assessment of the biological weapons incident to responsible officials in order to help address microbiological and safety issues, minimize fear and concerns of those responding to the incident, and help manage individuals potentially exposed to a threat agent.

The interferon system is the first line of defense against viral infection in mammals. This system is designed to block the spread of virus infection in the body, sometimes at the expense of accelerating the death of the infected cells. As expected of potent cytokines, in addition to their antiviral effects, interferons have profound effects on many aspects of cell physiology. All these actions of interferons are mediated by hundreds of interferon-induced proteins that are usually not synthesized in resting cells. Interferons induce their synthesis by activating the Jak-STAT pathways, a paradigm of cell signaling used by many cytokines and growth factors. Surprisingly, some of the same genes can also be induced directly by viruses and double-stranded RNA, a common viral by-product. Some of the interferon-induced proteins have novel biochemical properties and some are inactive as such but can be activated by double-stranded RNA produced during virus infection. Finally, almost all viruses have evolved mechanisms to evade the interferon system by partially blocking interferon synthesis or interferon action. Thus, in nature interferons and viruses maintain an equilibrium that allows regulated viral replication.

Bacteriophages of lactic acid bacteria are a threat to industrial milk fermentation. Owing to their economical importance, dairy phages became the most thoroughly sequenced phage group in the database. Comparative genomics identified related cos-site and pac-site phages, respectively, in lactococci, lactic streptococci and lactobacilli. Each group was represented with closely related temperate and virulent phages. Over the structural genes their gene maps resembled that of lambdoid coliphages, suggesting distant evolutionary relationships. Despite a lack of sequence similarity, a number of biochemical characteristics of these dairy phages are lambda-like (genetic switch, DNA packaging, head and tail morphogenesis, and integration, but not excision). These dairy phages thus provide interesting variations to the phage λ paradigm. The structural gene cluster of Lactococcus phage r1t resembled that of phages from mycobacteria. Virulent lactococcal phages with prolate heads (c2-like genus of Siphoviridae), in contrast, have no known counterparts in other bacterial genera.

The increase in drug-resistant pathogenic bacteria has created an urgent demand for new antibiotics. Among the more attractive targets for the development of new antibacterial compounds are the enzymes of fatty acid biosynthesis. Although a number of potent inhibitors of microbial fatty acid biosynthesis have been discovered, few of these are clinically useful drugs. Several of these fatty acid biosynthesis inhibitors have potential as lead compounds in the development of new antibacterials. This review encompasses the known inhibitors and prospective targets for new antibacterials.

Glutathione metabolism is associated with oxygenic cyanobacteria and the oxygen-utilizing purple bacteria, but is absent in many other prokaryotes. This review focuses on novel thiols found in those bacteria lacking glutathione. Included are glutathione amide and its perthiol, produced by phototrophic purple sulfur bacteria and apparently involved in their sulfide metabolism. Among archaebacteria, coenzyme M (2-mercaptoethanesulfonic acid) and coenzyme B (7-mercaptoheptanoylthreonine phosphate) play central roles in the anaerobic production of CH4 and associated energy conversion by methanogens, whereas the major thiol in the aerobic phototrophic halobacteria is γ-glutamylcysteine. The highly aerobic actinomycetes produce mycothiol, a conjugate of N-acetylcysteine with a pseudodisaccharide of glucosamine and myo-inositol, AcCys-GlcNα(1 → 1)Ins, which appears to play an antioxidant role similar to glutathione. Ergothioneine, also produced by actinomycetes, remains a mystery despite many years of study. Available data on the biosynthesis and metabolism of these and other novel thiols is summarized and key areas for additional study are identified.

The ancient and highly evolved mutualism between fungus-growing ants and their fungi is a textbook example of symbiosis. The ants carefully tend the fungus, which serves as their main food source, and traditionally are believed to be so successful at fungal cultivation that they are able to maintain the fungus free of microbial pathogens. This assumption is surprising in light of theories on the evolution of parasitism, especially for those species of ants that have been clonally propagating their cultivars for millions of years. Recent work has established that, as theoretically predicted, the gardens of fungus-growing ants are host to a specialized, virulent, and highly evolved fungal pathogen in the genus Escovopsis. In addition, the ants have evolved a mutualistic association with filamentous bacteria (actinomycetes) that produce antibiotics that suppress the growth of Escovopsis. Thus, the attine symbiosis appears to be a coevolutionary “arms race” between the garden parasite Escovopsis on the one hand and the ant-fungus-actinomycete tripartite mutualism on the other. These recent findings indicate that microbes may be key components in the regulation of other symbiotic associations between higher organisms.

Homology-dependent gene silencing (HDGS) is a ubiquitous phenomenon among fungi, plants, and animals. Gene silencing can be triggered and can affect artificially introduced nucleic acid molecules, both DNA and RNA, and/or can act on endogenous duplicated sequences. Although the various HDGS phenomena may be related each other, probably deriving from an ancestral defense mechanism, relevant differences do exist between different HDGS mechanisms. Especially in fungi, a variety of HDGS phenomena have been uncovered during the past 10 years: Gene inactivation of duplicated sequences can be achieved either through DNA-methylation and block of transcription or through sequence-specific degradation of mRNA. Moreover, duplicated sequences can also be specifically mutagenized. Studying HDGS in fungi gives us the opportunity to study such complex mechanisms in relatively simple organisms in which both genetic and biochemical approaches can be easily used.

Many pathogens must surmount an epithelial cell barrier in order to establish an infection. While much has been learned about the interaction of bacterial pathogens with cultured epithelial cells, the influence of cell polarity on these events has only recently been appreciated. This review outlines bacterial-host epithelial cell interactions in the context of the distinct apical and basolateral surfaces of the polarized epithelium that lines the lumens of our organs.

In 1917, bacteriophages were recognized as epizootic infections of bacteria and were almost immediately deployed for antibacterial therapy and prophylaxis. The early trials of bacteriophage therapy for infectious diseases were confounded, however, because the biological nature of bacteriophage was poorly understood. The early literature reviewed here indicates that there are good reasons to believe that phage therapy can be effective in some circumstances. The advent of antibiotics, together with the “Soviet taint” acquired by phage therapy in the postwar period, resulted in the absence of rigorous evaluations of phage therapy until very recently. Recent laboratory and animal studies, exploiting current understandings of phage biology, suggest that phages may be useful as antibacterial agents in certain conditions.

Leishmania-sand fly interactions are reviewed in the context of the potential barriers to the complete development of the parasite that exist within the midgut environment of phlebotomine flies and the molecular adaptations that the parasite has evolved that permit the development of transmissible infections to proceed. Cell surface and secreted phosphoglycans protect the parasite from the proteolytic activities of the blood-fed midgut, mediate attachment to the gut wall in order to maintain infection during excretion of the bloodmeal, and contribute to the formation of a biological plug in the anterior gut that may promote transmission by bite. The importance of vector saliva in modulating the host response to transmitted parasites is also reviewed.

The eutrophication of many ecosystems in recent decades has led to an increased interest in the ecology of nitrogen transformation. Chemolitho-autotrophic ammonia-oxidizing bacteria are responsible for the rate-limiting step of nitrification in a wide variety of environments, making them important in the global cycling of nitrogen. These organisms are unique in their ability to use the conversion of ammonia to nitrite as their sole energy source. Because of the importance of this functional group of bacteria, understanding of their ecology and physiology has become a subject of intense research over recent years. The monophyletic nature of these bacteria in terrestrial environments has facilitated molecular biological approaches in studying their ecology, and progress in this field has been rapid. The ammonia-oxidizing bacteria of the β-subclass Proteobacteria have become somewhat of a model system within molecular microbial ecology, and this chapter reviews recent progress in our knowledge of their distribution, diversity, and ecology.