It is assumed that intracellular pathogenic bacteria have to cope with DNA alkylating stress within host cells. Here we use single-cell reporter systems to show that the pathogen Brucella abortus does encounter alkylating stress during the first hours of macrophage infection. Genes encoding direct repair and base-excision repair pathways are required by B. abortus to face this stress in vitro and in a mouse infection model. Among these genes, ogt is found to be under the control of the conserved cell-cycle transcription factor GcrA. Our results highlight that the control of DNA repair in B. abortus displays distinct features that are not present in model organisms such as Escherichia coli.

The pathogenic bacterium Brucella abortus invades and multiplies inside host cells. To grow inside host cells, B. abortus requires a functional histidine biosynthesis pathway. Here, we show that a B. abortus histidine auxotroph mutant also displays an unexpected chaining phenotype. The intensity of this phenotype varies according to the culture medium and is exacerbated inside host cells. Chains of bacteria consist of contiguous peptidoglycan, and likely result from the defective cleavage of peptidoglycan at septa. Genetic suppression of the chaining phenotype unearthed two essential genes with a role in B. abortus cell division: dipM and cdlP. Loss of function of dipM and cdlP generates swelling at the division site. While DipM is strictly localized at the division site, CdlP is localized at the growth pole and the division site. Altogether, the unexpected chaining phenotype of a hisB mutant allowed the discovery of new crucial actors in cell division in B. abortus.

The routine work of any molecular biology laboratory includes the daily use of microorganisms, including strains of E. coli, transformed with a variety of plasmids expressing at least one antibiotic resistance gene (ARG). To verify the effectiveness of disinfection methods on laboratory liquid waste, bacteria isolated from laboratory and research institute drains were identified by 16S ribosomal RNA sequencing and tested for the presence of an origin of replication and several ARGs frequently found in laboratory plasmids. Surprisingly, the origin of replication of Enterobacteriaceae plasmids was detected in strains of non-Enterobacteriaceae bacteria suggesting that interspecific transfer of laboratory plasmids had occurred. Using quantitative Polymerase Chain Reaction, we determined the Decimal reduction value (D-value, expressed as concentration of disinfectant or length of physical treatment) of several decontamination methods for their DNA degradation effect on cultures of E. coli Top10 transformed with a kanamycin resistant plasmid (pET28A + or pEGFP-C2). The estimated D-values were 0,7 M for Sulfuric, 6,3% for a commercial P3 disinfectant, 25 minutes for steam sterilization at 121°C and 49 minutes for disinfection by UVC. A 20-minute treatment of bacteria cultures with a final concentration of 1–10% sodium hypochlorite was found to be ineffective in completely destroying a bacteria plasmid gene marker (coding for the pBR322 origin of replication). Residual DNA from HClO treated cells was 60%, while it decreased under 10% using the commercial disinfectant P3 diluted at 5%. As the degradation was uncomplete in both cases, to prevent the horizontal transfer of laboratory ARGs to environmental bacteria, disinfected liquid waste should not be released in sewage without additional plasmid destruction treatment.

Routine work of any molecular biology laboratory includes the daily use of microorganisms, including non-pathogenic E coli strains, transformed with a variety of plasmids expressing at least one antibiotic resistance gene (ARG). To check the efficacy of neutralization methods, bacteria isolated from wastewater from laboratories and research institutes were identified using the 16S ribosomal RNA sequencing and they were tested for the presence of an origin of replication and several ARGs frequently found in laboratory plasmids. Surprisingly, the origin of replication of Enterobacteriaceae was detected in non-Enterobacteriaceae bacteria strains suggesting that an interspecies transfer of laboratory plasmids had occurred. Using quantitative Polymerase Chain Reaction (qPCR), we determined the Decimal reduction value (D-value, expressed as concentration of disinfectant or length of physical treatment) of several neutralization methods for their DNA-bd effect: i.e. 0.7M for Sulfuric, 3.2% for a commercial disinfectant P3, 25 minutes for steam sterilization at 121°C and 49 minutes for UVC. A 20-minute treatment of laboratory liquid waste (LLW) with 1–10% sodium chlorine is ineffective in destroying completely plasmid gene markers intreated cells. Therefore, to prevent the horizontal transfer of laboratory ARGs to environmental bacteria, bleach-treated liquid waste should not be released in sewage without additional plasmid destruction treatment.

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