Carriage of linezolid-resistant enterococci (LRE) among humans and animals in Nigeria: coexistence of the cfr, optrA, and poxtA genes in Enterococcus faecium of animal origin.

OBJECTIVES: In contrast to increasing reports of the emergence of linezolid-resistant enterococci (LRE) emanating from many countries in Europe, Asia, and North America, data on its status and dissemination from the African continent remain scarce, with the information available limited to countries in North Africa. This study investigated the carriage of LRE and the genetic mechanism of resistance among Enterococcus faecium and Enterococcus faecalis strains recovered from humans and animals in Makurdi, Nigeria. METHODS: We conducted a cross-sectional study between June 2020 and July 2021 during which 630 non-duplicate human and animal faecal samples were collected and processed for the recovery of LRE. The genetic mechanisms for resistance were investigated using polymerase chain reaction (PCR) and Sanger sequencing. RESULTS: Linezolid-resistant enterococci were recovered from 5.87% (37/630; 95% CI: 4.17-8.00) of the samples, with the prevalence in animals and humans being 6.22% [(28/450); 95% CI: 4.17-8.87] and 5.00% [(9/180); 95% CI: 2.31-9.28], respectively. All isolates remained susceptible to vancomycin. No known point mutation mediating linezolid resistance was detected in the 23S rRNA and ribosomal protein genes; however, acquisition of one or more potentially transferable genes (cfr, optrA, and poxtA) was observed in 26 of the 37 LRE isolates. Co-existence of all three transferable genes in a single isolate was found in four E. faecium strains of animal origin. CONCLUSION: This study provides baseline evidence for the emergence and active circulation of LRE driven majorly by the acquisition of the optrA gene in Nigeria. To the best of our knowledge, our study is the first to report a co-carriage of all three transferable linezolid resistance determinants in E. faecium. Active LRE surveillance is urgently required to understand the extent of LRE spread across sub-Saharan Africa and to develop tailored mitigation strategies.

Concurrent Resistance to Carbapenem and Colistin Among Enterobacteriaceae Recovered From Human and Animal Sources in Nigeria Is Associated With Multiple Genetic Mechanisms.

Resistance to last resort drugs such as carbapenem and colistin is a serious global health threat. This study investigated carbapenem and colistin resistance in 583 non-duplicate Enterobacteriaceae isolates utilizing phenotypic methods and whole genome sequencing (WGS). Of the 583 isolates recovered from humans, animals and the environment in Nigeria, 18.9% (110/583) were resistant to at least one carbapenem (meropenem, ertapenem, and imipenem) and 9.1% (53/583) exhibited concurrent carbapenem-colistin resistance. The minimum inhibitory concentrations of carbapenem and colistin were 2-32 µg/mL and 8 to >64 µg/mL, respectively. No carbapenem resistant isolates produced carbapenemase nor harbored any known carbapenemase producing genes. WGS supported that concurrent carbapenem-colistin resistance was mediated by novel and previously described alterations in chromosomal efflux regulatory genes, particularly mgrB (M1V) ompC (M1_V24del) ompK37 (I70M, I128M) ramR (M1V), and marR (M1V). In addition, alterations/mutations were detected in the etpA, arnT, ccrB, pmrB in colistin resistant bacteria and ompK36 in carbapenem resistant bacteria. The bacterial isolates were distributed into 37 sequence types and characterized by the presence of internationally recognized high-risk clones. The results indicate that humans and animals in Nigeria may serve as reservoirs and vehicles for the global spread of the isolates. Further studies on antimicrobial resistance in African countries are warranted.

Veterinary clinic surfaces as reservoirs of multi-drug- and biocide-resistant Gram-negative bacteria.

This cross-sectional study was carried out to determine the common Gram-negative bacteria (GNB) contaminating veterinary clinic environments, and to evaluate the susceptibility of the isolates to commonly used antibiotics and biocides. A total of 62 swab samples were collected from different frequently touched surfaces in the 4 veterinary clinics visited. The samples were processed for isolation and identification of GNB using standard microbiological procedures. The susceptibility of the isolates to disinfectants and antibiotics was determined using agar dilution and disc diffusion techniques, respectively. A total of 114 GNB were isolated from the 4 clinics with isolation rates of 21.9, 22.8, 23.7 and 31.6% in clinics A, B, C and D, respectively. The surfaces of treatment tables were more contaminated (16.7 %) than receptionist/clinician desks (15.8%), weighing balances (10.5 %), door handles (7.9 %), drip stands (7.9 %), handwashing basins (7.0 %) and client chairs (7.0%). The surface-contaminating isolates were distributed into 20 genera, with members of Enterobacteriaceae predominating (n=97). Fifty-nine per cent of the isolates were resistant to the disinfectant Septol, while 5.3 and 0.9% were resistant to Purit and Dettol disinfectants, respectively. Multiple drug resistance was observed among 99% of the isolates with approximately 100% resistance to beta-lactams. Phenotypic expression of extended-spectrum (3.5 %) and AmpC beta-lactamase (38.6 %) production was detected. These findings highlight the role of clinic environments in serving as reservoirs for potential pathogens and sources for the spread of multi-drug resistant GNB.

Phenotypic detection, antimicrobial susceptibility and virulence profile of staphylococci in the pig production setting, Makurdi, Nigeria.

Livestock, particularly pigs, have increasingly been recognized as important reservoirs for zoonotic transmission of pathogenic bacteria, including staphylococci. Livestock production systems in developing countries of sub-Saharan Africa, including Nigeria, are characterized by high misuse/abuse of antimicrobials and a close association between humans and these animals, which promotes the emergence and transmission of resistant and potentially virulent bacteria. In the present study, we investigated the occurrence and characteristics (species distribution, virulence and resistance profile) of staphylococci from smallholder backyard pig farms, slaughter slabs and pig handlers in Makurdi, Nigeria. A total of 330 nasal swabs originating from 300 pigs and 30 in-contact humans were collected and processed. One hundred and thirteen samples [34.2 %; 95 % confidence interval (CI): 29.1-39.6] comprising 103 (34.3 %; 95 % CI: 29.0-40.0) and 10 (33.3 %; 95 % CI: 17.3-52.8 %) samples from pigs and humans, respectively, were positive for staphylococci, yielding 120 isolates (pigs n=110, humans n=10). The 120 isolates were distributed into 15 species with Staphylococcus aureus (n=25) followed by Staphylococcus cohnii (n=19) and Staphylococcus sciuri (n=14) occurring more frequently. All isolates were resistant to ß-lactam (100 %) antibiotics. Resistance to some critical antimicrobials, including linezolid (22 %), vancomycin (19.2 %), gentamicin (7.5%) and the fluoroquinolones ciprofloxacin (75.8 %) and enrofloxacin (66.7 %), was also observed. Majority (99.2 %) of the isolates displayed a multidrug resistance phenotype with the AMP-C-CIP-E-ENR-FOX-OX-P-S-SXT-TE phenotype being predominant. Overall, 70 % of the isolates expressed the methicillin resistance phenotype, out of which 20 % (n=17) were MRSA. Resistance to serum bactericidal activity and biofilm production were respectively observed in 45 (100 %) and 5 (11.3 %) of the coagulase-positive staphylococci. Our findings demonstrated the occurrence of a high diversity of staphylococci expressing multidrug resistance and potentially virulent phenotypes among healthy swine and pig handlers in small-scale backyard farms in North-Central Nigeria. These findings underscore the potential role of pig production settings in the emergence and dissemination of potentially virulent staphylococci and the importance of the development of antimicrobial resistance monitoring systems/implementation of control measures in developing countries. Proper hygienic practices and control of indiscriminate use and misuse of antibiotics are recommended.

Identification of mobile colistin resistance genes (mcr-1.1, mcr-5 and mcr-8.1) in Enterobacteriaceae and Alcaligenes faecalis of human and animal origin, Nigeria.

Colistin is a last-resort drug used to treat infections caused by multidrug-resistant Gram-negative bacteria that have developed carbapenem resistance. Emergence and rapid dissemination of the nine plasmid-mediated mobile colistin resistance genes (mcr-1 to mcr-9) has led to fear of pandrug-resistant infections worldwide. To date, there is only limited information on colistin resistance in African countries where the drug is widely used in agriculture. In this Nigerian study, 583 non-duplicate bacterial strains were isolated from 1119 samples from humans, camels, cattle, dogs, pigs and poultry using colistin-supplemented MacConkey agar, among which 17.0% (99/583) were colistin-resistant. PCR (mcr-1 to mcr-9) and whole-genome sequencing (WGS) identified mcr in 21.2% (21/99) of colistin-resistant isolates: mcr-1.1 (n = 13), mcr-8.1 (n = 5), mcr-1.1 and mcr-8.1 (n = 2), and mcr-1.1 and mcr-5 (n = 1). Of the 21 mcr-positive strains, 9 were isolated from human samples, with 8 being Klebsiella pneumoniae, and 6 of these human K. pneumoniae had a high colistin MIC (>64 µg/mL). In contrast, 9 of the 12 mcr-positive animal isolates were Escherichia coli, of which only 2 had a colistin MIC of >64 µg/mL. This study is the first to report mcr-1 in Alcaligenes faecalis and the emergence of mcr-5 and mcr-8 in Nigeria. WGS determined that mcr-1 was localised on an IncX4 plasmid and that 95.2% of mcr-1 harbouring isolates (20/21) transferred colistin resistance successfully by conjugation. These findings highlight the global spread of colistin resistance and emphasise the urgent need for co-ordinated global action to combat resistant bacteria.

Identification of Adult Fasciola spp. Using Matrix-Assisted Laser/Desorption Ionization Time-of-Flight (MALDI-TOF) Mass Spectrometry.

Fascioliasis is a neglected trematode infection caused by Fasciola gigantica and Fasciola hepatica. Routine diagnosis of fascioliasis relies on macroscopic identification of adult worms in liver tissue of slaughtered animals, and microscopic detection of eggs in fecal samples of animals and humans. However, the diagnostic accuracy of morphological techniques and stool microscopy is low. Molecular diagnostics (e.g., polymerase chain reaction (PCR)) are more reliable, but these techniques are not routinely available in clinical microbiology laboratories. Matrix-assisted laser/desorption ionization time-of-flight (MALDI-TOF) mass spectrometry (MS) is a widely-used technique for identification of bacteria and fungi; yet, standardized protocols and databases for parasite detection need to be developed. The purpose of this study was to develop and validate an in-house database for Fasciola species-specific identification. To achieve this goal, the posterior parts of seven adult F. gigantica and one adult F. hepatica were processed and subjected to MALDI-TOF MS to create main spectra profiles (MSPs). Repeatability and reproducibility tests were performed to develop the database. A principal component analysis revealed significant differences between the spectra of F. gigantica and F. hepatica. Subsequently, 78 Fasciola samples were analyzed by MALDI-TOF MS using the previously developed database, out of which 98.7% (n = 74) and 100% (n = 3) were correctly identified as F. gigantica and F. hepatica, respectively. Log score values ranged between 1.73 and 2.23, thus indicating a reliable identification. We conclude that MALDI-TOF MS can provide species-specific identification of medically relevant liver flukes.

Characterization of high level ampicillin- and aminoglycoside-resistant enterococci isolated from non-hospital sources.

PURPOSE: High level ampicillin- and aminoglycoside-resistant enterococci are being increasingly reported from non-hospital sources. This study was carried out to characterize these strains from non-hospital sources in Nigeria. METHODOLOGY: A collection of Enterococcus faecium isolated from vegetables, soil, farm animals and manure and observed to be resistant to ampicillin (n=63) and gentamicin (n=37) discs, were screened for resistance to high levels of ampicillin and aminoglycoside using E-test strips. Putative high level ampicillin- and aminoglycoside-resistant strains were screened for pbp5 and aminoglycoside modifying enzyme genes, respectively, by PCR. The C-terminal region of the amplified pbp5 gene was also sequenced. RESULTS: Five (5/63) and thirty-five (35/37) of the ampicillin- and aminoglycoside-resistant strains were identified as high level ampicillin- and aminoglycoside-resistant E. faecium strains, respectively, based on the MIC results. The amplified pbp5 gene from the high level ampicillin-resistant isolates displayed 96-99 % nucleotide sequence similarity with the reference strains and three novel insertions (500Glu→Leu, 502Asp→Arg and 614Ile→Phe) in the amino acid sequence. Aminoglycoside modifying enzyme genes aac(6')-Ie-aph(2″) (100 %), aph(2')-Ic (88.8 %), aph(3')-IIIa (90 %) and ant(4')-Ia (40 %) were detected among the high level aminoglycoside-resistant isolates. CONCLUSION: This is the first report on the characterization of high level ampicillin- and aminoglycoside-resistant Enterococcus faecium among animals and vegetables in Nigeria. The results show that non-hospital sources can constitute a reservoir for potential dissemination of these strains and genes to humans via the food chain or by direct contact.

Equine Dermatophytosis: A Survey of Its Occurrence and Species Distribution among Horses in Kaduna State, Nigeria.

This study was designed to determine the occurrence and species distribution of dermatophyte from cutaneous skin lesions of horses in Kaduna State, Nigeria. A total of 102 skin scrapings were collected from 102 horses with skin lesions. Mycological studies were carried out using conventional techniques. Dermatophytes were isolated from 18 (17.6%) of the 102 samples collected. The 18 dermatophytes were distributed into 10 different species belonging to Microsporum (n = 5) and Trichophyton (n = 5) genera. T. verrucosum (n = 4) was the most predominant species isolated followed by M. equinum (n = 3), T. vanbreuseghemii (n = 2), M. gypseum (n = 2), and M. canis (n = 2). Others include M. fulvum (n = 2), T. mentagrophytes (n = 1), T. equinum (n = 1), T. soudanense (n = 1), and M. gallinae (n = 1). The present study reveals the occurrence of dermatophytes in cutaneous skin lesions of horses in Kaduna State, Nigeria. In addition for the first time in this environment the anthropophilic dermatophyte T. soudanense was isolated from horses. These findings have great economic, veterinary, and public health significance as they relate to the cost of treatment and dissemination of zoonotic dermatophytes.

Draft Genome Sequences of 37 Salmonella enterica Strains Isolated from Poultry Sources in Nigeria.

Here, we report the availability of draft genomes of several Salmonella serotypes, isolated from poultry sources from Nigeria. These genomes will help to further understand the biological diversity of S. enterica and will serve as references in microbial trace-back studies to improve food safety.

Serological prevalence of leptospirosis in cattle slaughtered in the Zango abattoir in Zaria, Kaduna State, Nigeria.

Leptospirosis is an occupational zoonosis caused by pathogenic leptospires. In this study, the presence and prevalence of antibodies specific to Leptospira spp. serovar Hardjo in 142 cattle slaughtered between June and July 2011 was investigated using the enzyme-linked immunosorbent assay (ELISA). Five (3.50%) of the 142 cattle sampled were seropositive for antibodies to Leptospira spp. serovar Hardjo. Despite the fact that there was no significant difference (p>0.05) in seropositivity between sexes and between breeds sampled, there was a significant difference (p<0.05) in sero-positivity between the different age groups examined. Leptospirosis is present in cattle slaughtered in the Zango abattoir; butchers and abattoir workers are exposed to infected animals and are at risk of being infected by Leptospira spp. serovar Hardjo.