The use of antibiotics at subtherapeutic concentrations for agricultural applications is believed to be an important factor in the proliferation of antibiotic-resistant bacteria. The goal of this study was to determine if the application of manure onto agricultural land would result in the proliferation of antibiotic resistance among soil bacteria. Chlortetracycline-resistant bacteria were enumerated and characterized from soils exposed to the manure of animals fed subtherapeutic concentrations of antibiotics and compared to the chlortetracycline-resistant bacteria from soils at farms with restricted antibiotic use (dairy farms) and from non-agricultural soils. No significant differences were observed at nine different study sites with respect to the numbers and types of cultivated chlortetracycline-resistant bacteria. Genes encoding for tetracycline resistance were rarely detected in the resistant bacteria from these sites. In contrast, soils collected from a tenth farm, which allowed manure to indiscriminately accumulate outside the animal pen, had significantly higher chlortetracycline-resistance levels. These resistant bacteria frequently harbored one of 14 different genes encoding for tetracycline resistance, many of which (especially tet(A) and tet(L)) were detected in numerous different bacterial species. Subsequent bacterial enumerations at this site, following the cessation of farming activity, suggested that this farm remained a hotspot for antibiotic resistance. In conclusion, we speculate that excessive application of animal manure leads to the spread of resistance to soil bacteria (potentially by lateral gene transfer), which then serve as persistent reservoir of antibiotic resistance.

In this study, the abilities of two anaerobic digestion processes used for sewage sludge stabilization were compared for their ability to reduce the quantities of three genes that encode resistance to tetracycline (tet(A), tet(O), and tet(X)) and one gene involved with integrons (intI1). A two-stage, thermophilic/mesophilic digestion process always resulted in significant decreases in the quantities of tet(X) and intI1, less frequently in decreases of tet(O), and no net decrease in tet(A). The thermophilic stage was primarily responsible for reducing the quantities of these genes, while the subsequent mesophilic stage sometimes caused a rebound in their quantities. In contrast, a conventional anaerobic digestion process rarely caused a significant decrease in the quantities of any of these genes, with significant increases occurring more frequently. Our results demonstrate that anaerobic thermophilic treatment was more efficient in reducing quantities of genes associated with the spread of antibiotic resistance compared to mesophilic digestion.

The microbial burden of the Phoenix spacecraft assembly environment was assessed in a systematic manner via several cultivation-based techniques and a suite of NASA-certified, cultivation-independent biomolecule-based detection assays. Extremotolerant bacteria that could potentially survive conditions experienced en route to Mars or on the planet's surface were isolated with a series of cultivation-based assays that promoted the growth of a variety of organisms, including spore formers, mesophilic heterotrophs, anaerobes, thermophiles, psychrophiles, alkaliphiles, and bacteria resistant to UVC radiation and hydrogen peroxide exposure. Samples were collected from the clean room where Phoenix was housed at three different time points, before (1P), during (2P), and after (3P) Phoenix's presence at the facility. There was a reduction in microbial burden of most bacterial groups, including spore formers, in samples 2P and 3P. Analysis of 262 isolates from the facility demonstrated that there was also a shift in predominant cultivable bacterial populations accompanied by a reduction in diversity during 2P and 3P. It is suggested that this shift was a result of increased cleaning when Phoenix was present in the assembly facility and that certain species, such as Acinetobacter johnsonii and Brevundimonas diminuta, may be better adapted to environmental conditions found during 2P and 3P. In addition, problematic bacteria resistant to multiple extreme conditions, such as Bacillus pumilus, were able to survive these periods of increased cleaning.

Aims:  The tet(X) gene has previously been found in obligate anaerobic Bacteroides spp., which is curious because tet(X) encodes for a NADP‐dependent monooxygenase that requires oxygen to degrade tetracycline. In this study, we characterized a tetracycline resistant, aerobic, Gram‐negative Sphingobacterium sp. strain PM2‐P1‐29 that harbours a tet(X) gene. Methods and Results:  Sphingobacterium sp. PM2‐P1‐29 demonstrated the ability to transform tetracycline compared with killed controls. The presence of the tet(X) gene was verified by PCR and nucleotide sequence analysis. Additional nucleotide sequence analysis of regions flanking the tet(X) gene revealed a mobilizable transposon‐like element (Tn6031) that shared organizational features and genes with the previously described Bacteroides conjugative transposon CTnDOT. A circular transposition intermediate of the tet(X) region, characteristic of mobilizable transposons, was detected. However, we could not demonstrate the conjugal transfer of the tet(X) gene using three different recipient strains and numerous experimental conditions. Conclusions:  This study suggests that Sphingobacterium sp. PM2‐P1‐29 or a related bacterium may be an ancestral source of the tet(X) gene. Significance and Impact of the Study:  This study demonstrates the importance of environmental bacteria and lateral gene transfer in the dissemination and proliferation of antibiotic resistance among bacteria.