Molecular characterization of antimicrobial resistance in Klebsiella pneumoniae isolated from Brazilian dairy herds

Abstract

In this observational study, phenotypic and genotypic patterns of antimicrobial resistance (AMR) in Klebsiella pneumoniae isolated from intramammary infections, clinical mastitis, fresh feces, rectal swabs, animal hindlimbs, and bulk tank milk samples from Brazilian dairy herds were investigated. In addition, we identified specific genetic variants present among extended-spectrum β-lactamase (ESBL) producers. We obtained 169 isolates of K. pneumoniae from 2009 to 2011 on 24 Brazilian dairy farms located in 4 Brazilian states. The AMR profile of all isolates was determined using disk-diffusion assays. The antimicrobial panel included drugs commonly used as mastitis treatment in Brazilian dairy herds (gentamicin, cephalosporins, sulfamethoxazole-trimethoprim, tetracycline) as well as antimicrobials of critical importance for human health (meropenem, ceftazidime, fluoroquinolones). The K. pneumoniae isolates resistant to tetracycline, fluoroquinolones, sulfamethoxazole-trimethoprim, or chloramphenicol were screened for presence of drug-specific AMR genes [tet, qnr, floR, catA2, cm1A, dfr, sul] using PCR. In addition, we identified ESBL genes present among ESBL-producers by using whole genome sequencing. Genomes were assembled and annotated, and patterns of AMR genes were investigated. Resistance was commonly detected against tetracycline (22.5% of all isolates), streptomycin (20.7%), and sulfamethoxazole-trimethoprim (9.5%). Antimicrobial resistance rates were higher in K. pneumoniae isolated from intramammary infections in comparison with iso-lates from feces (19.2 and 0% of multidrug resistance in intramammary and fecal isolates, respectively). In contrast, no difference in AMR rates was observed when contrasting hind limbs and isolates from intramammary infections. The genes tetA, sul2, and floR were the most frequently observed AMR genes in K. pneumoniae resistant to tetracycline, sulfamethoxazoletrimethoprim, and chloramphenicol, respectively. The tetA gene was present exclusively in isolates from milk. The genes bla CTX-M8 and bla SHV-108 were present in 3 ESBL-producing K. pneumoniae, including an isolate from bulk tank milk. The 3 isolates were of sequence type 281 and had similar mobile genetic elements and virulence genes. Our study reinforced the epidemiological importance and dissemination of bla CTX-M-8 pST114 plasmid in food-producing animals in Brazil.

Epidemiology, Environmental Risks, Virulence, and Resistance Determinants of Klebsiella pneumoniae From Dairy Cows in Hubei, China

“…It has been reported that tetA and floR / cmlA mediate the efflux of tetracycline and florfenicol, respectively, which can pump the drug out of the cell and reduce the intracellular drug concentration to generate resistance ( Acosta-Pérez et al, 2015 ; Wang et al, 2018 ; Graesbøll et al, 2019 ). Notably, tetA and floR are the most frequently observed AMR genes in K. pneumoniae resistant to tetracycline and amide alcohols, respectively, which are consistent with the study by Nobrega et al (2021) . Although the three dihydrofolate synthase genes ( sul1 , sul2 , and sul3 ) were detected to varying degrees, no significant correlation was found between them and SOX and SXT.…”

Antimicrobial resistance and molecular characterization of Klebsiella species causing bovine mastitis in Nghe An province, Vietnam

“…COL is a last resort antibiotic to treat infections caused by K. pneumoniae , and the resistance rate to this antibiotic has increased in recent years from 4.8% in 2013–2018 to 8.2% in 2019–2021 [ 35 ]; our resistance rate still higher, when compared to this finding and might come from the transmission between strains as this antimicrobial, was used for treatment calves with diarrhea. CIP, ENR, PIP, IMI, and STX were highly sensitive in our study, which could be a good choice for mastitis treatment, especially CIP, PIP, and IMI, as these drugs had never been used before at studied farms; this finding was supported by other studies in Brazil [ 6 ], in China [ 31 ]. We found that all isolates were sensitive to GEN and EFT; these drugs also be a good option for veterinarians to deal with mastitis cows.…”

“…This resistance was reported at 45.1% in China [ 33 ], which is quite lower than in our study. Resistance to TET was reported at 32% in China [ 34 ], 19% in Canada [ 32 ], and 41.5% in Brazil [ 6 ]. COL is a last resort antibiotic to treat infections caused by K. pneumoniae , and the resistance rate to this antibiotic has increased in recent years from 4.8% in 2013–2018 to 8.2% in 2019–2021 [ 35 ]; our resistance rate still higher, when compared to this finding and might come from the transmission between strains as this antimicrobial, was used for treatment calves with diarrhea.…”

“…Klebsiella species are found everywhere in nature. This pathogen can be found in water, soil, plants, insects, and other animals, including humans [ 6 ]. Clinical mastitis due to Klebsiella is often severe and does not respond well to antimicrobial treatment, resulting in a prolonged duration of intramammary infection and substantial loss of milk yield [ 7 ].…”

Single-molecule real-time sequencing reveals differences in bacterial diversity in raw milk in different regions and seasons in China

“…Moreover, some pathogens and conditional pathogens are common bacteria with relatively low relative abundance detected in raw milk from all provinces, such as Klebsiella pneumoniae and B. cereus, of which Klebsiella pneumoniae could be sterilized by the subsequent processing of raw milk. Nevertheless, previous studies have revealed that Klebsiella pneumoniae might potentially associate with bacterial antibiotic gene transfer (Hou et al, 2015;Cheng et al, 2021;Nobrega et al, 2021). In addition, existence of Klebsiella pneumoniae in raw milk also remains a nonnegligible problem.…”