Hafnia, an enterobacterial genus naturally resistant to colistin revealed by three susceptibility testing methods.

Objectives: To determine the susceptibility to colistin of Hafnia alvei and Hafnia paralvei, and to compare methods for colistin resistance detection in the Hafnia genus. Methods: A collection of 25 Hafnia isolates was studied. Species were identified by using 16S rRNA gene sequencing with subsequent phylogeny analysis. Susceptibility to colistin was determined using the broth microdilution (BMD) reference method, the Phoenix automated system, the Rapid Polymyxin NP test, the Etest system and the disc diffusion method. Results: The collection consisted of 15 H. alvei and 10 H. paralvei isolates. Based on the 16S rRNA analysis, a close relationship of the Hafnia genus with naturally colistin-resistant enterobacterial genera (Proteus, Morganella, Providencia and Serratia) was identified. Susceptibility testing performed using the BMD method, the Phoenix automated system and the Rapid Polymyxin NP test revealed a high rate of colistin resistance (96%). Underestimation of colistin resistance using Etest strips (72%) and the disc diffusion method (0%) was observed. Conclusions: The high rate of colistin resistance observed within the Hafnia genus and its close phylogenetic relationship with naturally colistin-resistant genera suggest that Hafnia is a naturally colistin-resistant enterobacterial genus.

Binary combination of epsilon-poly-L-lysine and isoeugenol affect progression of spoilage microbiota in fresh turkey meat, and delay onset of spoilage in Pseudomonas putida challenged meat.

Proliferation of microbial population on fresh poultry meat over time elicits spoilage when reaching unacceptable levels, during which process slime production, microorganism colony formation, negative organoleptic impact and meat structure change are observed. Spoilage organisms in raw meat, especially Gram-negative bacteria can be difficult to combat due to their cell wall composition. In this study, the natural antimicrobial agents ε-poly-L-lysine (ε-PL) and isoeugenol were tested individually and in combinations for their activities against a selection of Gram-negative strains in vitro. All combinations resulted in additive interactions between ε-PL and isoeugenol towards the bacteria tested. The killing efficiency of different ratios of the two antimicrobial agents was further evaluated in vitro against Pseudomonas putida. Subsequently, the most efficient ratio was applied to a raw turkey meat model system which was incubated for 96 h at spoilage temperature. Half of the samples were challenged with P. putida, and the bacterial load and microbial community composition was followed over time. CFU counts revealed that the antimicrobial blend was able to lower the amount of viable Pseudomonas spp. by one log compared to untreated samples of challenged turkey meat, while the single compounds had no effect on the population. However, the compounds had no effect on Pseudomonas spp. CFU in unchallenged meat. Next-generation sequencing offered culture-independent insight into population diversity and changes in microbial composition of the meat during spoilage and in response to antimicrobial treatment. Spoilage of unchallenged turkey meat resulted in decreasing species diversity over time, regardless of whether the samples received antimicrobial treatment. The microbiota composition of untreated unchallenged meat progressed from a Pseudomonas spp. to a Pseudomonas spp., Photobacterium spp., and Brochothrix thermosphacta dominated food matrix on the expense of low abundance species. We observed a similar shift among the dominant species in meat treated with ε-PL or the antimicrobial blend, but the samples differed markedly in the composition of less abundant species. In contrast, the overall species diversity was constant during incubation of turkey meat challenged with P. putida although the microbiota composition did change over time. Untreated or ε-PL treated samples progressed from a Pseudomonas spp. to a Pseudomonas spp. and Enterobacteriaceae dominated food matrix, while treatment with the antimicrobial blend resulted in increased relative abundance of Hafnia spp., Enterococcaceae, and Photobacterium spp. We conclude that the blend delayed the onset of spoilage of challenged meat, and that all antimicrobial treatments of unchallenged or challenged meat affect the progression of the microbial community composition. Our study confirms that the antimicrobial effects observed in vitro can be extrapolated to a food matrix such as turkey meat. However, it also underlines the consequence of species-to-species variation in susceptibility to antimicrobials, namely that the microbial community change while the CFU remains the same. Addition of antimicrobials may thus prevent the growth of some microorganisms, allowing others to proliferate in their place.

Detection of ticarcillin-clavulonic acid susceptibility with microdilution method in Citrobacter, Hafnia, Proteus and some gram negative bacteria.

The broth dilution method has been regarded as a good alternative test for detection of susceptibilities to antimicrobial agents. In this study, the antimicrobial activity of ticarcillin-clavulonic acid (TIM) was investigated by the minimal inhibitory concentration (MIC) method on strains of Aeromonas, Citrobacter, Hafnia, Morganella, Proteus, Pseudomonas and gram negative bacteria isolated from raw milk. The isolate collection included 91 gram negative strains. Fifty-one (56.04%) isolates were found sensitive (MIC < or = 8 microg/ml), 12 (13.19%) isolates were found intermediately sensitive (MIC 16-32 microg/ml), and 28 (30.77%) isolates were found resistant (MIC > or = 64 microg/ml) to TIM.