Do Pathogenic Escherichia coli Isolated from Gallus gallus in South Africa Carry Co-Resistance Toward Colistin and Carbapenem Antimicrobials?

Colistin and carbapenems are critically important antimicrobials often used as a last resort to manage multidrug-resistant bacterial infections in humans. With limited alternatives, resistance to these antimicrobials is of concern as organisms could potentially spread horizontally rendering treatments ineffective. The aim of this study was to investigate co-resistance to colistin and carbapenems among Escherichia coli isolated from poultry in South Africa. Forty-six E. coli strains obtained from clinical cases of breeder and broiler chickens were used. In addition to other antibiotics, all the isolates were tested against colistin and carbapenems using broth microdilution. Multiplex polymerase chain reactions were used to investigate the presence of colistin (mcr-1 to 5) and carbapenem (blaOXA-48, blaNDM-1, and blaVIM) resistance genes. Isolates exhibiting colistin resistance (>2 µg/mL) underwent a whole-genome sequencing analysis. Resistance to colistin (10.9%) and cefepime (6.5%) was noted with all colistin-resistant strains harboring the mcr-1 gene. None of the E. coli isolates were resistant to carbapenems nor carried the other resistant genes (mcr-2 to 5, blaOXA-48, blaNDM-1, and blaVIM). The mcr-1-positive strains belonged to sequence types ST117 and ST156 and carried virulence genes ompA, aslA, fdeC, fimH, iroN, iutA, tsh, pic, ast A and set 1A/1B. In conclusion, clinical E. coli strains from chickens in this study possessed mobile resistance genes for colistin and several other clinically relevant antimicrobials but not carbapenems. Additionally, they belonged to sequence types in addition to carrying virulence factors often associated with human extraintestinal pathogenic E. coli infections. Thus, the potential risk of transmitting these strains to humans cannot be underestimated especially if sick birds are dispatched into the thriving poorly regulated Cornish hen industry. The need for routine veterinary surveillance and monitoring of antimicrobial resistance, antimicrobial use and the importance of strengthening regulations guiding the informal poultry sector remains important.

Diclofenac toxicity in susceptible bird species results from a combination of reduced glomerular filtration and plasma flow with subsequent renal tubular necrosis.

Diclofenac caused the death of millions of vultures on the Asian subcontinent. Other non-steroidal anti-inflammatory drugs (NSAIDs) have since also been shown to be toxic to vultures with the exception of meloxicam. For this study, we evaluated the effect of diclofenac on renal uric acid transport and glomerulus filtration in an acute toxicity model. In a two-phase study with the same birds, healthy chickens (a validated model species) were treated intravenously with para-amino hippuric acid (PAH) and iohexol (IOH) in combination in phase 1. In phase 2, the same PAH and IOH combination was then combined with diclofenac (10 mg/kg). In both phases, blood and faeces were sequentially collected. In phase 1, the birds showed no signs of ill health. Moreover, PAH, IOH and uric acid clearance was rapid. In phase 2, two chickens showed early signs of hyperuricemia 8 hours after exposure and died approximately 24h later. Necropsy showed classic signs of renal damage and gout. Diclofenac had a rapid plasma half-life of elimination of less than 2 hours indicating that toxicity was likely due to an irreversible destruction of a physiological process. All the birds in phase 2 had decreased uric acid, PAH and IOH clearance in comparison to phase 1. The decrease in PAH clearance was variable between the birds (average of 71%) but was near 98% reduced in the two birds that died. It is concluded that diclofenac alters both renal perfusion and renal plasma flow, with death associated with tubular secretion being reduced to negligible functionality for a prolonged period. This would support previous in vitro findings of early cell death from ROS accumulation. However, further evaluation is needed to elucidate this final step.

Could the environmental toxicity of diclofenac in vultures been predictable if preclinical testing methodology were applied?

Diclofenac, a non-steroidal anti-inflammatory pharmaceutical agent was responsible for the death of millions of Gyps vulture's in the Indian sub-region with the safety of the other non-steroidal anti-inflammatory drugs (NSAIDs) being questionable. With preclinical safety testing not well established for avian species unlike for mammalian and environmental toxicity, we ask the question if a preclinical model could have predicted the toxic effect of the drug. For this study, we test an Organisation for Economic Co-operation and Development (OECD) guideline 223 for assessing the acute toxic potential of pesticides in birds by exposing three avian species to the drug. Exposed Japanese quails (Coturnix japonica) and Muscovy ducks (Cairina moschata) demonstrated clinical signs and pathology similar to those previously reported in vultures viz. hyperuricemia, depression, death, visceral gout and nephrosis. However, exposed domestic pigeons (Columba livia domestica) were insensitive. Following a pharmacokinetic analysis, the drug was well absorbed and distributed in the pigeons with a half-life below 6 h. A toxicokinetic evaluation in quails showed poisoning was due to metabolic constraint, with a half-life and mean residence time above 6 h and 8 h respectively resulting in death. Toxicity seen in the ducks was however not related to metabolic constraint but hyperuricemia as metabolism was rapid [half-life (1-2 h) and mean residence time (2-3 h)] irrespective of survival or death. Despite succumbing to diclofenac, the established oral median lethal dose (LD50) of 405.42 mg/kg and 189.92 mg/kg in Japanese quails and Muscovy ducks respectively from this study were substantially higher than those reported for Gyps vultures (0.098 mg/kg) which is as a result of the rapid elimination of the drug from the body in the former species. More importantly, it suggests that these species are not suitable as surrogates for non-steroidal anti-inflammatory drug toxicity testing and that the toxicity of diclofenac in vultures is idiosyncratic most likely as a result of species specific metabolism.