The chromosomal arsenic resistance genes of the acidophilic, chemolithoautotrophic, biomining bacterium Thiobacillus ferrooxidans were cloned and sequenced. Homologues of four arsenic resistance genes, arsB, arsC, arsH, and a putative arsR gene, were identified. The T. ferrooxidans arsB (arsenite export) and arsC (arsenate reductase) gene products were functional when they were cloned in an Escherichia coli ars deletion mutant and conferred increased resistance to arsenite, arsenate, and antimony. Therefore, despite the fact that the ars genes originated from an obligately acidophilic bacterium, they were functional in E. coli. Although T. ferrooxidans is gram negative, its ArsC was more closely related to the ArsC molecules of gram-positive bacteria. Furthermore, a functional trxA (thioredoxin) gene was required for ArsC-mediated arsenate resistance in E. coli; this finding confirmed the gram-positive ArsC-like status of this resistance and indicated that the division of ArsC molecules based on Gram staining results is artificial. Although arsH was expressed in an E. coli-derived in vitro transcription-translation system, ArsH was not required for and did not enhance arsenic resistance in E. coli. The T. ferrooxidans ars genes were arranged in an unusual manner, and the putative arsR and arsC genes and the arsBH genes were translated in opposite directions. This divergent orientation was conserved in the four T. ferrooxidans strains investigated.Thiobacillus ferrooxidans is an acidophilic (optimum pH, 1.8 to 2.5), obligately chemolithotrophic bacterium that obtains its energy through oxidation of ferrous iron to ferric iron or oxidation of reduced inorganic sulfur compounds to sulfuric acid. It is a member of a consortium of bacteria (which includes Thiobacillus caldus and Leptospirillum ferrooxidans) that is used in commercial biooxidation processes to recover gold from arsenopyrite ores (22). Although recent analysis of microbial populations in continuous-flow biooxidation tanks has revealed that T. ferrooxidans may not be as dominant as was once thought, this organism is nevertheless usually present in such tanks (21). Total arsenic levels greater than 13 g liter Ϫ1 may be present in arsenopyrite biooxidation tanks, and therefore the microorganisms present must have a mechanism of resistance to arsenic (8).Plasmid-associated arsenic efflux resistance mechanisms have been known for many years and have been extensively reviewed (5,23,(30)(31)(32)35). Although the number of components of these systems varies, in the case of Escherichia coli plasmids R773 and R46, as well as Acidiphilium multivorum plasmid pKW301 (34), as many as five genes (arsRDABC) are present. In the case of R773, the genes are transcribed in a single operon. The arsR and arsD genes encode repressors that control the basal and upper levels of ars operon expression, while the arsABC genes encode the structural components of the arsenic resistance mechanism. ArsA is an ATPase which forms a complex with ArsB, the transmembrane arsenite efflux p...

Gastrointestinal (GI) diseases, and in particular those caused by bacterial infections, are a major cause of morbidity and mortality worldwide. Treatment is becoming increasingly difficult due to the increase in number of species that have developed resistance to antibiotics. Probiotic lactic acid bacteria (LAB) have considerable potential as alternatives to antibiotics, both in prophylactic and therapeutic applications. Several studies have documented a reduction, or prevention, of GI diseases by probiotic bacteria. Since the activities of probiotic bacteria are closely linked with conditions in the host's GI-tract (GIT) and changes in the population of enteric microorganisms, a deeper understanding of gut-microbial interactions is required in the selection of the most suitable probiotic. This necessitates a deeper understanding of the molecular capabilities of probiotic bacteria. In this review, we explore how probiotic microorganisms interact with enteric pathogens in the GIT. The significance of probiotic colonization and persistence in the GIT is also addressed.

Acid tolerance is considered an important characteristic of probiotic bacteria. Lactobacillus plantarum 423 tolerates acidic pH and is the ideal candidate in which to study molecular mechanisms that acid-tolerant lactic acid bacteria employ to survive such conditions. In this study we recorded changes in the protein profile of L. plantarum 423 when exposed to pH 2.5 by using a gel-free nanoLC-MS/MS proteomics approach. In total, 97 proteins were detected as more abundant, and 12 proteins were detected solely when strain 423 was exposed to pH 2.5. General stress response proteins, the utilization of a variety of carbohydrate sources in a glucose rich environment, altered pyruvate metabolism, increased lysine biosynthesis, and a significant oxidative stress response was observed in acid-stressed cells. The accumulation of basic compounds also seemed to play an integral role in the response to acid stress. We observed a marked decrease in proteins involved in cell wall and phospholipid biosynthesis, transcription, translation, and cell division. The most abundant protein detected was an uncharacterized protein, JDM1_2142. Functional analysis revealed that this protein plays a role in survival during acid stress. Our results contribute to the growing body of knowledge on the molecular mechanisms employed by lactobacilli, in particular L. plantarum, to ensure survival in acidic conditions.

Two sets of arsenic resistance genes were isolated from the highly arsenic-resistant Leptospirillum ferriphilum Fairview strain. One set is located on a transposon, TnLfArs, and is related to the previously identified TnAtcArs from Acidithiobacillus caldus isolated from the same arsenopyrite biooxidation tank as L. ferriphilum. TnLfArs conferred resistance to arsenite and arsenate and was transpositionally active in Escherichia coli. TnLfArs and TnAtcArs were sufficiently different for them not to have been transferred from one type of bacterium to the other in the biooxidation tank. The second set of arsenic resistance genes conferred very low levels of resistance in E. coli and appeared to be poorly expressed in both L. ferriphilum and E. coli.Processes for the biooxidation of gold-bearing arsenopyrite concentrates were developed in the 1980s and are now used in several countries (17). These are among the largest commercial fermentation processes known. Biooxidation is a mineral pretreatment process during which the molecular structure of the arsenopyrite mineral is broken down, exposing the gold and allowing its extraction by cyanide (7). During this process, large quantities of arsenic are released into continuous-flow aeration tanks in which the biooxidation takes place. The majority of arsenopyrite biooxidation processes operate at 40°C and are dominated by a mixture of the sulfur-oxidizing bacterium Acidithiobacillus caldus and the iron-oxidizing bacterium Leptospirillum ferriphilum. During the first few years of operation, the continuous-flow nature of the processes resulted in the selection of highly arsenic-resistant bacteria.Studies to investigate what genetic changes had taken place that accompanied this high level of arsenic resistance have been carried out with A. caldus. Highly arsenic-resistant strains of A. caldus have been found to contain an unusual Tn21-like ars operon that is not present in less-resistant strains (6,23,24). The 12-kb TnAtcArs is unusual in that the tnpA (transposase) and tnpR (resolvase) genes occur on opposite ends of the transposon, unlike other transposons of the Tn21and Tn3 family, where they form an adjacent unit. These two transposon genes flank a series of genes (arsRCDADA orf7 orf8B) associated with arsenic resistance that are themselves unusual. Genes for an ArsR (negative regulator) (20, 27) and ArsC (arsenate reductase) (10, 11) are followed by a tandem duplication of the genes for ArsD (a second repressor) (28) and ArsA (an ATPase that associates with ArsB and links arsenite export to ATP hydrolysis) (22). These genes are followed by genes encoding ORF7 (an NADH-like oxidoreductase), ORF8 (a cystathione-␤-synthase [CBS] domain-containing protein), and ArsB, the arsenite efflux pump. Deletion of one copy of arsDA readily occurs, but this deletion does not appear to affect resistance when cloned on a multicopy plasmid in Escherichia coli (24). Similarly, the inactivation or deletion of ORF7 and ORF8 did not affect arsenic resistance in E. coli. Some strains of high...

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