Extended-spectrum beta-lactamase (ESBL)- and non-ESBL producing Escherichia coli surveillance in surface water sources in Edo State, Nigeria: a public health concern.

This research explores the antimicrobial resistance (AMR) profiles and prevalence of extended-spectrum beta-lactamase (ESBL) and non-ESBL-producing Escherichia coli in Ojerame Dam and Ovokoto Spring, Edo State, Nigeria. Over 12 months, water was systematically sampled to accommodate seasonal variations and analyzed by employing an ESBL-selective medium for bacterial species. Additionally, bacterial isolates underwent identification and characterization using polymerase chain reaction (PCR) and disk diffusion methods to evaluate their susceptibility to antimicrobials. Results indicated significant prevalence of ESBL-producing E. coli, which exhibited complete resistance to common antimicrobials like ceftriaxone, ceftazidime, cefotaxime, and ampicillin while demonstrating 100% sensitivity to ertapenem, imipenem, meropenem, and nitrofurantoin. Non-ESBL-producing E. coli were resistant to ampicillin but sensitive to other antimicrobials mentioned earlier. Furthermore, both ESBL and non-ESBL-producing E. coli displayed multidrug resistance to varying degrees. Specific ESBL genes, including blaTEM, blaCTX-M-1, and blaCTX-M-15, were identified, alongside resistance genes like tetA, tetM, sul1, sul2, sul3, qnrA, qnrB, and qnrS in E. coli. This study pioneers the documentation of ESBL-producing E. coli in surface water in the region. This signals impending health risks associated with water being a reservoir of resistant genes while emphasizing the urgency for further research and public awareness concerning the quality of surface water.

Draft genome sequences of a blaCTX-M-15-harboring and an atypical enteropathogenic Escherichia coli isolate recovered from slaughterhouses in Nigeria.

We present the draft genome sequences of two Escherichia coli strains isolated from slaughterhouses in Edo State, Nigeria, in 2019. The isolates were identified as blaCTX-M-15-harboring (19-47-58) and atypical enteropathogenic E. coli (aEPEC) (19-47-66), belonging to multilocus sequence types (MLST) ST46 and ST2089, respectively.

Characterization of resistance and virulence factors in livestock-associated methicillin-resistant Staphylococcus aureus.

The study investigated the economic concerns associated with livestock-associated methicillin-resistant Staphylococcus aureus (LA-MRSA) in livestock (cow), examining its connection to severe infections, antimicrobial resistance (AMR), and virulence factors. The research, conducted in Edo State, Nigeria, analyzed 400 samples (200 rectal and 200 nasal swabs) collected between March 2018 and February 2019. MRSA prevalence was identified using conventional culture-based methods and polymerase chain reaction (PCR) techniques, revealing 63.5% (n = 254) for Staphylococcus aureus and 55% (n = 220) for MRSA. Of the 76 mecA-positive MRSA isolates, 64.5% (n = 49) exhibited multidrug resistance (MDR) while the remaining were sensitive to specific antimicrobials. Key virulence genes, such as PVL (81.6%; n = 62) and tsst-1 (44.7%; n = 34), were prevalent, along with AMR genes like mecC, tetM, ermA, ermC, vanA, and vanC. Staphylococcal chromosomal cassette mec (SCCmec) typing identified different types, notably II, IVa, and IVb. Biofilm formation, a crucial virulence factor varied in strength, is associated with icaA and icaB genes (p < 0.01). The findings highlighted substantial AMR and biofilm-forming capacity within LA-MRSA isolates, emphasizing the importance of ongoing surveillance for informed treatment strategies, AMR policies, and control measures against MDR staphylococcal infections.

Machine learning-guided determination of Acinetobacter density in waterbodies receiving municipal and hospital wastewater effluents.

A smart artificial intelligent system (SAIS) for Acinetobacter density (AD) enumeration in waterbodies represents an invaluable strategy for avoidance of repetitive, laborious, and time-consuming routines associated with its determination. This study aimed to predict AD in waterbodies using machine learning (ML). AD and physicochemical variables (PVs) data from three rivers monitored via standard protocols in a year-long study were fitted to 18 ML algorithms. The models' performance was assayed using regression metrics. The average pH, EC, TDS, salinity, temperature, TSS, TBS, DO, BOD, and AD was 7.76 ± 0.02, 218.66 ± 4.76 µS/cm, 110.53 ± 2.36 mg/L, 0.10 ± 0.00 PSU, 17.29 ± 0.21 °C, 80.17 ± 5.09 mg/L, 87.51 ± 5.41 NTU, 8.82 ± 0.04 mg/L, 4.00 ± 0.10 mg/L, and 3.19 ± 0.03 log CFU/100 mL respectively. While the contributions of PVs differed in values, AD predicted value by XGB [3.1792 (1.1040-4.5828)] and Cubist [3.1736 (1.1012-4.5300)] outshined other algorithms. Also, XGB (MSE = 0.0059, RMSE = 0.0770; R2 = 0.9912; MAD = 0.0440) and Cubist (MSE = 0.0117, RMSE = 0.1081, R2 = 0.9827; MAD = 0.0437) ranked first and second respectively, in predicting AD. Temperature was the most important feature in predicting AD and ranked first by 10/18 ML-algorithms accounting for 43.00-83.30% mean dropout RMSE loss after 1000 permutations. The two models' partial dependence and residual diagnostics sensitivity revealed their efficient AD prognosticating accuracies in waterbodies. In conclusion, a fully developed XGB/Cubist/XGB-Cubist ensemble/web SAIS app for AD monitoring in waterbodies could be deployed to shorten turnaround time in deciding microbiological quality of waterbodies for irrigation and other purposes.

Prevalence, multiple antibiotic resistance and virulence profile of methicillin-resistant Staphylococcus aureus (MRSA) in retail poultry meat from Edo, Nigeria.

Introduction: Staphylococcus aureus causes staphylococcal food poisoning and several difficult-to-treat infections. The occurrence and dissemination of methicillin-resistance S. aureus (MRSA) in Nigeria is crucial and well documented in hospitals. However, findings on MRSA from meat in the country are yet to be adequately reported. The current study determined the prevalence, virulence profile and antibiogram characteristics of MRSA from a raw chicken product from retail outlets within Edo. Methods: A total of 368 poultry meat samples were assessed for MRSA using a standard culture-based approach and characterized further using a molecular method. The antimicrobial susceptibility profile of the isolates was determined using the disc diffusion method. The biofilm profile of the isolates was assayed via the crystal violet microtitre-plate method. Virulence and antimicrobial resistance genes were screened using polymerase chain reaction via specific primers. Results: Of the samples tested, 110 (29.9%) were positive for MRSA. All the isolates were positive for deoxyribonuclease (DNase), coagulase and beta-hemolysis production. Biofilm profile revealed 27 (24.55%) weak biofilm formers, 18 (16.36%) moderate biofilm formers, and 39 (35.45%) strong biofilm formers. The isolates harboured 2 and ≤17 virulence genes. Enterotoxin gene profiling revealed that 100 (90.9%) isolates harboured one or more genes. Resistance against the tested antibiotics followed the order: tetracycline 64(58.2%), ciprofloxacin 71(64.6%), trimethoprim 71(64.6%) and rifampin 103(93.6%). A total of 89 isolates were multidrug-resistant, while 3 isolates were resistant to all 22 antibiotics tested. The isolates harboured antimicrobial-resistant determinants such as methicillin-resistant gene (mecA), tetracycline resistance genes (tetK, tetL), erythromycin resistance genes (ermA, ermC), trimethoprim resistance gene (dfrK). All the staphylococcal cassette chromosome mec (SCCmec) IVa and SCCmec V positive isolates harboured the Panton-Valentine Leukocidin Gene (PVL). Conclusion: In conclusion, S. aureus was resistant to commonly used antibiotics; a concern to public health concerning the transmission of these pathogens after consuming these highlight the significance of antimicrobial and enterotoxigenic monitoring of S. aureus in food chains.

Multidrug-resistant extended spectrum ß-lactamase (ESBL)-producing Escherichia coli from farm produce and agricultural environments in Edo State, Nigeria.

Antimicrobial resistance (AMR) is a major public health concern, especially the extended-spectrum ß-lactamase-producing (ESBL) Escherichia coli bacteria are emerging as a global human health hazard. This study characterized extended-spectrum ß-lactamase Escherichia coli (ESBL-E. coli) isolates from farm sources and open markets in Edo State, Nigeria. A total of 254 samples were obtained in Edo State and included representatives from agricultural farms (soil, manure, irrigation water) and vegetables from open markets, which included ready-to-eat (RTE) salads and vegetables which could potentially be consumed uncooked. Samples were culturally tested for the ESBL phenotype using ESBL selective media, and isolates were further identified and characterized via polymerase chain reaction (PCR) for ß-lactamase and other antibiotic resistance determinants. ESBL E. coli strains isolated from agricultural farms included 68% (17/25) from the soil, 84% (21/25) from manure and 28% (7/25) from irrigation water and 24.4% (19/78) from vegetables. ESBL E. coli were also isolated from RTE salads at 20% (12/60) and vegetables obtained from vendors and open markets at 36.6% (15/41). A total of 64 E. coli isolates were identified using PCR. Upon further characterization, 85.9% (55/64) of the isolates were resistant to ≥ 3 and ≤ 7 antimicrobial classes, which allows for characterizing these as being multidrug-resistant. The MDR isolates from this study harboured ≥1 and ≤5 AMR determinants. The MDR isolates also harboured ≥1 and ≤3 beta-lactamase genes. Findings from this study showed that fresh vegetables and salads could be contaminated with ESBL-E. coli, particularly fresh produce from farms that use untreated water for irrigation. Appropriate measures, including improving irrigation water quality and agricultural practices, need to be implemented, and global regulatory guiding principles are crucial to ensure public health and consumer safety.

Surveillance of Vibrio parahaemolyticus pathogens recovered from ready-to-eat foods.

This study examined the occurrence of V. parahaemolyticus from ready-to-eat (RTE) food in Delta State, Nigeria. It also characterized antibiotic resistance and virulence gene profile patterns to determine the associated health risk hazard. Food samples total of 380 were collected randomly and assessed for V. parahaemolyticus. V. parahaemolyticus isolates were characterized for their virulence and antibiogram potentials using a phenotypic and polymerase chain reaction (PCR) approach. A total of 42 (11.1%) samples were contaminated with V. parahaemolyticus. In 17/42 (40.5%) of the V. parahaemolyticus-positive samples, the densities were < 10 MPN/g. However, 19/42 (45.2%) and 6/42 (14.3%) of the samples had densities of 10 - 102 and > 102 MPN/g, respectively. A total of 67 V. parahaemolyticus isolates were identified using PCR; 54(80.6%) isolates were multidrug resistant. A total of 22 (32.8%), 39 (58.2%), and 67 (100%) of the V. parahaemolyticus harbored the tdh, trh, and tlh toxin genes, respectively. The T3SS1 gene (vcrD1) was detected in 67 (100%) of the isolates. The T3SS2α genes which were vcrD2, vopB2, and vopT were detected in 21 (31.3%), 11 (16.4%) and 30 (44.8%) of the isolates respectively. Some of the V. parahaemolytics strains harbored the orf8 gene 20 (29.9%), and a combination of orf8 + tdh genes 12 (17.9%), categorized as pandemic strains. The antibiotic resistance genes detected in this study include blaTEM 33 (49.3), tetM 19 (28.4), cmlA 32(47.8) and sul1 14 (20.9). The concentration levels and prevalence of V. parahaemolyticus in RTE foods indicate contamination of ready-to-eat foods, particularly street foods consumed in the Delta State of Nigeria, threatening public health and consumer safety.

Identification and characterization of MDR virulent Salmonella spp isolated from smallholder poultry production environment in Edo and Delta States, Nigeria.

Salmonella is responsible for some foodborne disease cases worldwide. It is mainly transmitted to humans through foods of animal origin through the consumption of poultry products. The increased international trade and the ease of transboundary movement could propel outbreaks of local origin to translate into severe global threats. The present study aimed to characterize Salmonella serovars isolated from poultry farms in Edo and Delta States, Nigeria. A total of 150 samples (faecal, water and feed) were collected from ten poultry farms between January and August 2020 and analyzed for Salmonella characterization using standard bacteriological and molecular methods. Salmonella serovars identified include: Salmonella Enteritidis [n = 17 (39.5%)], Salmonella Typhimurium [n = 13 (30.2%)] and other Salmonella serovars [n = 13 (30.2%)]. All Salmonella serovars were cefotaxime and ampicillin resistant. The presence of the invA gene ranged from 9(69.2%) to 15(88.2%). The spvC gene ranged from 2(14.4%) to 10(58.8%). All Salmonella serovars had sdiA gene. The Salmonella isolates produced some extracellular virulence factors (such as protease, lipase, ß-hemolytic activity, and gelatinase), while 13(30.2%) of the overall isolates formed strong biofilms. In conclusion, the detection of multiple antibiotic-resistant Salmonella serovars in faecal sources, which also exhibited virulence determinants, constituted a public health risk as these faecal samples have the potential as manure in the growing of crops. These pathogens can be transmitted to humans nearby and through poultry products, resulting in difficult-to-treat infections and economic loss.

Using machine learning models to predict the effects of seasonal fluxes on Plesiomonas shigelloides population density.

Seasonal variations (SVs) affect the population density (PD), fate, and fitness of pathogens in environmental water resources and the public health impacts. Therefore, this study is aimed at applying machine learning intelligence (MLI) to predict the impacts of SVs on P. shigelloides population density (PDP) in the aquatic milieu. Physicochemical events (PEs) and PDP from three rivers acquired via standard microbiological and instrumental techniques across seasons were fitted to MLI algorithms (linear regression (LR), multiple linear regression (MR), random forest (RF), gradient boosted machine (GBM), neural network (NN), K-nearest neighbour (KNN), boosted regression tree (BRT), extreme gradient boosting (XGB) regression, support vector regression (SVR), decision tree regression (DTR), M5 pruned regression (M5P), artificial neural network (ANN) regression (with one 10-node hidden layer (ANN10), two 6- and 4-node hidden layers (ANN64), and two 5- and 5-node hidden layers (ANN55)), and elastic net regression (ENR)) to assess the implications of the SVs of PEs on aquatic PDP. The results showed that SVs significantly influenced PDP and PEs in the water (p < 0.0001), exhibiting a site-specific pattern. While MLI algorithms predicted PDP with differing absolute flux magnitudes for the contributing variables, DTR predicted the highest PDP value of 1.707 log unit, followed by XGB (1.637 log unit), but XGB (mean-squared-error (MSE) = 0.0025; root-mean-squared-error (RMSE) = 0.0501; R2 =0.998; medium absolute deviation (MAD) = 0.0275) outperformed other models in terms of regression metrics. Temperature and total suspended solids (TSS) ranked first and second as significant factors in predicting PDP in 53.3% (8/15) and 40% (6/15), respectively, of the models, based on the RMSE loss after permutations. Additionally, season ranked third among the 7 models, and turbidity (TBS) ranked fourth at 26.7% (4/15), as the primary significant factor for predicting PDP in the aquatic milieu. The results of this investigation demonstrated that MLI predictive modelling techniques can promisingly be exploited to complement the repetitive laboratory-based monitoring of PDP and other pathogens, especially in low-resource settings, in response to seasonal fluxes and can provide insights into the potential public health risks of emerging pathogens and TSS pollution (e.g., nanoparticles and micro- and nanoplastics) in the aquatic milieu. The model outputs provide low-cost and effective early warning information to assist watershed managers and fish farmers in making appropriate decisions about water resource protection, aquaculture management, and sustainable public health protection.

Polymyxin sensitivity/resistance cosmopolitan status, epidemiology and prevalence among O1/O139 and non-O1/non-O139 Vibrio cholerae: A meta-analysis.

Resistance/sensitivity to polymyxin-B (PB) antibiotic has been employed as one among other epidemiologically relevant biotyping-scheme for Vibrio cholerae into Classical/El Tor biotypes. However, recent studies have revealed some pitfalls bordering on PB-sensitivity/resistance (PBR/S) necessitating study. Current study assesses the PBR/S cosmopolitan prevalence, epidemiology/distribution among O1/O139 and nonO1/nonO139 V. cholerae strains. Relevant databases (Web of Science, Scopus and PubMed) were searched to retrieve data from environmental and clinical samples employing the Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA). Random-effect-model (REM) and common-effect-model (CEM) of meta-analysis was performed to determine prevalence of PBR/S V. cholerae strains, describe the cosmopolitan epidemiological potentials and biotype relevance. Heterogeneity was determined by meta-regression and subgroup analyses. The pooled analyzed isolates from articles (7290), with sensitive and resistance are 2219 (30.44%) and 5028 (69.56%). Among these PB-sensitive strains, more than 1944 (26.67%) were O1 strains, 132 (1.81%) were nonO1 strains while mis-reported Classical biotype were 2080 (28.53) respectively indicating potential spread of variant/dual biotype. A significant PB-resistance was observed in the models (CEM = 0.66, 95% CI [0.65; 0.68], p-value = 0.001; REM = 0.83 [0.74; 0.90], p = 0.001) as both models had a high level of heterogeneity (I2 = 98.0%; df=332=1755.09,Qp=2.4932). Egger test (z = 5.4017, p < 0.0001) reveal publication bias by funnel plot asymmetry. The subgroup analysis for continents (Asia, Africa) and sources (acute diarrhea) revealed (98% CI (0.73; 0.93); 55% CI (0.20; 0.86)), and 92% CI (0.67; 0.98). The Epidemiological prevalence for El tor/variant/dual biotype showed 88% CI (0.78; 0.94) with O1 strains at 88% CI (0.78; 0.94). Such global prevalence, distribution/spread of phenotypes/genotypes necessitates updating the decades-long biotype classification scheme. An antibiotic stewardship in the post antibiotic era is suggestive/recommended. Also, there is need for holistic monitoring/evaluation of clinical/epidemiological relevance of the disseminating strains in endemic localities.

Processed ready-to-eat (RTE) foods sold in Yenagoa Nigeria were colonized by diarrheagenic Escherichia coli which constitute a probable hazard to human health.

The study aimed to recover diarrheagenic Escherichia coli strains from processed ready-to-eat (RTE) foods in Yenagoa, Nigeria and characterize them using culture-based and molecular methods. Three hundred RTE food samples were collected randomly from different food outlets between February 2021 and August 2021 and assessed for the occurrence of E. coli using standard bacteriological procedures. The virulence factor formation and antibiotic susceptibility profile of the isolates was carried out using standard microbiological procedures. Polymerase chain reaction (PCR) was used to confirm the identity of the isolates via specific primers and further used to assay the diarrheagenic determinants of the E. coli isolates. The prevalence of E. coli positive samples based on the proliferation of E. coli on Chromocult coliform agar forming purple to violet colonies was 80(26.7%). The population density of E. coli from the RTE foods ranged from 0-4.3 × 104 ± 1.47 CFU/g. The recovered E. coli isolates (n = 62) were resistant to antibiotics in different proportions such as ampicillin 62(100%), aztreonam 47(75.81%) and chloramphenicol 43(69.35%). All the recovered E. coli isolates were resistant to ≥ 2 antibiotics. The multiple antibiotic-resistant index (MARI) ranged from 0.13-0.94 with 47(75.8%) of isolates having MARI >2. A total of 48(77.4%) of the isolates were multidrug-resistant (MDR). The proportion of extracellular virulence factor formation is as follows: protease 12(19.35%), curli 39(62.9%), cellulose 21(33.89%), ornithine decarboxylase 19(30.65%) and aesculin hydrolysis 14(22.58%). The overall proportion of diarrheagenic E. coli was 33/62(53.2%). The distributions of typical diarrheagenic E. coli includes: tETEC 9(14.5%), tEPEC 13(20.9%), tEAEC 6(9.7%), tEIEC 2(3.2%) and tEHEC 3(4.8%). The proportions of atypical strains include aETEC 10(16.1%), aEAEC 5(8.1%), aEPEC 1(1.6%) and aEIEC 3(4.8%). This study demonstrated that some RTE foods sold in Yenagoa, Nigeria, are contaminated and constitute a probable human health hazard. Thus, there is a need for intensive surveillance of this isolate in RTE foods variety to spot evolving AMR phenotypes and avert food-borne infections.

Prevalence and Characterization of Food-Borne Vibrio parahaemolyticus From African Salad in Southern Nigeria.

The demand for minimally processed vegetables (African salad) has increased partly due to its inclusion in ready-to-eat foods. Nevertheless, the associated risk of the presence of emergent foodborne pathogens, such as Vibrio parahaemolyticus might be underestimated. The present study was designed to isolate and characterize foodborne V. parahaemolyticus from minimally processed vegetables using culture-based methods and molecular approach. A total of 300 samples were examined from retail outlets between November 2018 and August 2019 from Southern Nigeria. The prevalence of vibrios from the overall samples based on the colonial proliferation of yellow, blue-green and/or green colonies on thiosulfate citrate bile salts sucrose agar was 74/300 (24.6%). An average of two green or blue-green colonies from respective plates was screened for V. parahaemolyticus using analytical profile index (API) 20 NE. Polymerase chain reaction further confirmed the identity of positive V. parahaemolyticus. The counts of V. parahaemolyticus ranged from 1.5 to 1,000 MPN/g. A total of 63 recovered V. parahaemolyticus were characterized further. The resistance profile of the isolates include ampicillin 57/63 (90.5%), cefotaxime 41/63 (65.1%), ceftazidime 30/63 (47.6%), amikacin 32/63 (50.8%), kanamycin 15/63 (23.8%), and oxytetracycline 16/63 (25.4%). The multiple antibiotic index ranged from 0-0.81. The formation of biofilm by the isolates revealed the following: strong formation 15/63 (23.8%), moderate formation 31/63 (49.2%), weak formation 12/63 (19.1%), and no formation 5/63 (7.9%). A total of 63/63 (100%), 9/63 (14.3%), and 20/63 (31.8%) of the isolates harbored the tox R gene, TDH-related hemolysin (trh) and thermostable direct hemolysin (tdh) determinants respectively. The isolates with O2 serogroup were most prevalent via PCR. Isolates that were resistant to tetracycline, kanamycin, and chloramphenicol possessed resistant genes. The presence of multidrug-resistant vibrios in the minimally processed vegetables constitutes a public health risk and thus necessitates continued surveillance.

Draft Genome Sequences of Five Staphylococcus saprophyticus Strains Isolated from African Fermented Nono in Nigeria.

Five Staphylococcus saprophyticus strains were isolated from the fermented milk product nono in Nigeria and were sequenced using an Illumina MiSeq platform. The genome sizes ranged from 2.53 to 2.60 Mbp, while the GC contents ranged from 32.99 to 33.07 mol%. The genomes possessed 2,505 to 2,687 protein-coding sequences.

Polyphasic study of antibiotic-resistant enterobacteria isolated from fresh produce in Germany and description of Enterobacter vonholyi sp. nov. isolated from marjoram and Enterobacter dykesii sp. nov. isolated from mung bean sprout.

Forty-two antibiotic-resistant enterobacteria strains were isolated from fresh produce obtained from the northern German retail market. A polyphasic characterization based on both phenotypic and genotypic methods was used to identify predominant strains as Citrobacter (C.) gillenii, C. portucalensis, Enterobacter (En.) ludwigii, Escherichia (E.) coli and Klebsiella (K.) pneumoniae. 38.1% of the enterobacteria strains were resistant to tetracycline, while 23.8% and 9.5% of strains were resistant to streptomycin and chloramphenicol, respectively. A high percentage of Klebsiella (100%), Enterobacter (57.1%) and Citrobacter (42.9%) strains were also resistant to ampicillin, with some strains showing multiple resistances. For unequivocal species identification, the genomes of thirty strains were sequenced. Multilocus sequence analysis, average nucleotide identity and digital DNA-DNA hybridization showed that Enterobacter strains E1 and E13 were clearly clustered apart from Enterobacter species type strains below the species delineation cutoff values. Thus, strains E1T (=DSM 111347T, LMG 31875T) represents a novel species proposed as Enterobacter dykesii sp. nov., while strain E13T (=DSM 110788T, LMG 31764T) represent a novel species proposed as Enterobacter vonholyi sp. nov. Strains often possessed different serine ß-lactamase genes, tet(A) and tet(D) tetracycline resistance genes and other acquired antibiotic resistance genes. Typical plasmid replicon types were determined. This study thus accurately identified the enterobacteria from fresh produce as species belonging to the genera Citrobacter, Enterobacter, Escherichia and Klebsiella, but also showed that these can carry potentially transferable antibiotic resistance genes and may thus contribute to the spread of these via the food route.

Identification and Characterization of Salmonella Serovars Isolated from Pig Farms in Benin City, Edo State, Nigeria: One Health Perspective.

The present study was carried out to characterize Salmonella serovars from commercial pig farms in Benin City, Nigeria. A total of 81 samples were collected from the Agricultural Development Program farms between January and June 2017. Standard culture-based and polymerase chain reaction procedures were adopted in the isolation and identification of Salmonella serovars. Antibiotic susceptibility profiles of the isolates were conducted using the Kirby-Bauer disc diffusion method with prominent resistance determinants screened for using specific primer sets. The 84 identified Salmonella serovars include 15 Salmonella ser. Enteritidis, 11 Salmonella ser. Typhimurium, and 58 other Salmonella serovars. Phenotypic virulence factors include: hemolytic activity (51.7-100%), lipase activity (48.3-81.8%), protease activity (60.3-100%), gelatinase production (50-90.9%), DNA degrading activity (55.2-90.9%), and S-layer formation (63.8-100%). The biofilm formation profiles include nonbiofilm producers (0-12.1%), weak biofilm producers (0-20%), moderate biofilm producers (24.1-27.3%), and strong biofilm producers (48.3-72.7%). Salmonella serovars in this study harbored resistant determinants, such as tetA, tetC, ampC, sul1, sul2, sul3, floR, ermA, and ermC. The occurrence of resistance phenotype and determinants in pathogenic Salmonella serovars from pig farms is a significant public health concern, which could result in the dissemination of resistant elements within the environment.

Microarray-based detection of resistance genes in coagulase-negative staphylococci isolated from cattle and buffalo with mastitis in Egypt.

The present study aimed to provide a detailed characterization of coagulase-negative staphylococci (CoNS) isolated from cows and buffaloes with mastitis. The study included seventy-five CoNS isolates (60 came from cattle and 15 from buffaloes) originating from 68 individual quarters of 67 dairy cows (53 cattle and 14 buffaloes). The animals belonged to five different small holding dairy herds (n = 140 cows) that show clinical or subclinical mastitis. CoNS isolates were phenotypically characterized using MALDI-TOF-MS and were further genotypically characterized by microarray-based assays. Furthermore, the antimicrobial susceptibility of CoNS strains which carried the mecA gene was examined by broth microdilution. The occurrence of CoNS in the respective five herds was 10.5%, 14.7%, 14.8%, 12.8%, and 9.9%, with an average of 12.4%. Six different CoNS species were identified: S. sciuri (n = 37; 30 from cattle and 7 from buffaloes), S. chromogenes (n = 14; 8 from cattle and 6 from buffaloes), S. haemolyticus (n = 10; nine from cattle and one buffalo), S. xylosus (n = 10; nine from cattle and one buffalo), S. hyicus (n = 2), S. warneri (n = 1), and unidentified CoNS (n = 1). Twenty percent (20%) of CoNS isolates (17.3% of cattle origin) carried at least one antimicrobial resistance gene, while 4% of the isolate including two isolates of S. haemolyticus and one S. warneri of cattle origin carried the mecA gene and were phenotypically identified as methicillin-resistant strains. The genes detected were blaZ (16%), followed by tet(K) (8%), aacA-aphD (4%), aphA3 (2.6%), msr(A) (2.6%), [far1 (2.6%), and fusC (2.6%)], sat (2.6%), and cat (1.3%) conferring resistance to penicillin, tetracycline, gentamicin, neomycin/kanamycin, erythromycin, fusidic acid, streptothricin, and chloramphenicol, respectively. The majority of investigated CoNS strains displayed considerably low prevalence of resistance genes, while resistance to more than three antibiotics was found in S. haemolyticus and S. warneri. Implementing effective preventive measures is, therefore, important for limiting the transmission of CoNS, rather than using antibiotics to control mastitis in bovines.

Mechanism of action of secondary metabolites from marine-derived Streptoymces on bacterial isolates by membrane permeability.

This study was designed to further investigate the potential mechanism of action of our previously characterized and identified marine-derived Streptomyces extracts (ESC003 and ESC012) on selected bacterial isolates from our culture bank. Time-kill kinetics, protein and lipid leakages assay, cell membrane permeability, phosphate and potassium ions efflux assay, extracellular adenosine triphosphate (ATP) concentration and membrane potential (MP) were all carried out using the marine-derived Streptomyces extract to determine and understand the probable mode of action at which the extract inhibit or kill bacterial cell. The MIC of ESC003 and ESC012 ranged from 0.16 to 6.25 mg/mL while the MBC ranged from 1.25 to >10 mg/mL. On the time-kill kinetics, a reduction in mean viable cell amount was detected at respective time studied. For the impermeability of the bacterial isolates, the relative electric conductivity increased with increase in concentration and time interval of exposure. As regards the protein leakage, lipid leakage, 260 nm absorbing materials leakage, phosphate and potassium ions efflux; considerable amount of these products were leaked with increase in concentration and time of exposure to the bacterial isolates. The extracellular ATP concentration from respective bacterial isolates increased appreciably with increased concentration of exposure with a simultaneous decrease in membrane potential. Findings from this study revealed that the Streptomyces extracts revealed a significant breakthrough against susceptible bacterial isolates via the permeability of the bacterial cell membrane, and thus resulted in the outflow of ATP, electrolytes, DNA materials, and proteins. These changes lead to disruption, and eventually cell death, which were proportional to a concurrent decrease in the viability of the bacterial cell.

Antibiotics resistance and toxin profiles of Bacillus cereus-group isolates from fresh vegetables from German retail markets.

BACKGROUND: This study aimed to evaluate the safety of raw vegetable products present on the German market regarding toxin-producing Bacillus cereus sensu lato (s.l.) group bacteria. RESULTS: A total of 147 B. cereus s.l. group strains isolated from cucumbers, carrots, herbs, salad leaves and ready-to-eat mixed salad leaves were analyzed. Their toxinogenic potential was assessed by multiplex PCR targeting the hemolysin BL (hbl) component D (hblD), non-hemolytic enterotoxin (nhe) component A (nheA), cytotoxin K-2 (cytK-2) and the cereulide (ces) toxin genes. In addition, a serological test was used to detect Hbl and Nhe toxins. On the basis of PCR and serological results, none of the strains were positive for the cereulide protein/genes, while 91.2, 83.0 and 37.4% were positive for the Hbl, Nhe and CytK toxins or their genes, respectively. Numerous strains produced multiple toxins. Generally, strains showed resistance against the ß-lactam antibiotics such as penicillin G and cefotaxim (100%), as well as amoxicillin/clavulanic acid combination and ampicillin (99.3%). Most strains were susceptible to ciprofloxacin (99.3%), chloramphenicol (98.6%), amikacin (98.0%), imipenem (93.9%), erythromycin (91.8%), gentamicin (88.4%), tetracycline (76.2%) and trimethoprim/sulfamethoxazole combination (52.4%). The genomes of eight selected strains were sequenced. The toxin gene profiles detected by PCR and serological test mostly agreed with those from whole-genome sequence data. CONCLUSIONS: Our study showed that B. cereus s.l. strains encoding toxin genes occur in products sold on the German market and that these may pose a health risk to the consumer if present at elevated levels. Furthermore, a small percentage of these strains harbor antibiotic resistance genes. The presence of these bacteria in fresh produce should, therefore, be monitored to guarantee their safety.

Prevalence of Antimicrobial Resistance and Virulence Gene Elements of Salmonella Serovars From Ready-to-Eat (RTE) Shrimps.

Gastrointestinal illnesses continue to be a global public health risk. Exposure to foodborne Salmonella directly or indirectly through consumption of ready-to-eat seafood can be an important route of infection to humans. This study was designed to estimate the population cell density, prevalence, virulence gene signatures, and antibiotic resistance of Salmonella serovars from ready-to-eat shrimps. Ready-to-eat (RTE) shrimp samples were obtained from different open markets in Delta and Edo States, Nigeria from November 2016 to October 2017. We employed classical and polymerase chain reaction (PCR) approaches. The mean Salmonella species enumerated from the RTE shrimps ranged from -0.301 to 5.434 log10 cfu/g with 210/1440 (14.58%) of the RTE shrimp samples harbored Salmonella species. After biochemical and PCR approach, the identified isolates were Salmonella Enteritidis 11(24.4%), Salmonella Typhimurium 14 (31.1%) and other Salmonella spp. 20 (44.4%). All Salmonella species recovered were resistant to penicillin and erythromycin with 100% sensitivity to cefotaxime, cephalothin, colistin, and polymyxin B. Findings on the multidrug-resistant (MDR) profile showed that a total of 9/14 (64.3%) of Salmonella Enteritidis were resistant to 5 antibiotics which belongs to 3 different groups of antimicrobials with a multiple antibiotic-resistant (MAR) index of 0.21; while 3/11 (27.3%) of Salmonella Typhimurium were resistant to 11 antibiotics which belongs to 7 different groups of antimicrobials with a MAR index of 0.46. Virulence genes (spiA, sipB, invA, sif A, fljB, and sefA) and resistance genes (class 1 and II integrase, sul2, catB3, flor, tmp, bla TEM, strB, dfr1, and tetC) were also detected in some of the Salmonella species with variable percentage. This study indicates that ready-to-eat shrimps are probable reservoirs harboring Salmonella strains. The identified Salmonella isolates which exhibited virulence determinants and antibiotic-resistant coupled with high MAR index constitute a consumer health risk to the communities.

Antimicrobial Resistance, Virulence Determinants, and Biofilm Formation of Enterococcus Species From Ready-to-Eat Seafood.

Enterococcus species form an important population of commensal bacteria and have been reported to possess numerous virulence factors considered significantly important in exacerbating diseases caused by them. The present study was designed to characterize antibiotic-resistant and virulent enterococci from ready-to-eat (RTE) seafood. A total of 720 RTE shrimp samples comprising sauced shrimp (n = 288), boiled shrimp (n = 216), and smoked shrimp (n = 216) obtained from open markets in Delta State, Nigeria, were assessed. Standard classical methods and polymerase chain reaction (PCR) were used in identifying the Enterococcus species. Potential virulence factors (ß-hemolysis, gelatinase activity, S-layer, and biofilm formation) were assessed using standard procedures. The antibiotic susceptibility profile of the identified enterococci isolates was assayed using the Kirby-Bauer disc diffusion method. PCR was further used to screen selected antibiotic resistance and virulence genes. Prevalence of Enterococcus species from shrimp varieties is as follows: sauced, 26 (9.03%); boiled, 6 (2.78%); and smoked, 27 (12.50%), with an overall prevalence of 59 (8.19%) based on the occurrence of black hallow colonies after incubation. Enterococcus species detected include E. faecalis, 17 (28.8%); E. faecium, 29 (49.2%); E. gallinarum, 6 (10.2%); E. casseliflavus, 2 (3.4%); E. hirae, 3 (5.1%); and E. durans, 2 (3.4%). Biofilm occurrence among the shrimp varieties is as follows: 19/26 (73.1%) for sauced shrimps, 5/6 (83.3%) for boiled shrimps, and 16/27 (59.3%) for smoked shrimps. The phenotypic expression of the enterococci virulence revealed the following: S-layer, 59 (100%); gelatinase production, 19 (32.2%); and ß-hemolysis, 21 (35.6%). An average of 3-11 virulence genes were detected in the Enterococcus species. The resistance profile of Enterococcus species is as follows: erythromycin, 29 (49.2%); vancomycin, 22 (37.3%); and tetracycline, 27 (45.8%). The frequency of occurrence of antibiotic resistance genes from the phenotypic resistant enterococci isolates to the macrolide, glycopeptide, and tetracycline antibiotics is as follows: ermA, 13/29 (44.8%); vanA, 14/22 (63.6%); tetA, 14/27 (51.9%); tetM, 15/27 (55.6%); ermB, 4/29 (13.8%); and vanB, 5/22 (22.7%). Findings from this study reveal the antibiotic resistance of enterococci strains of such species as E. durans, E. casseliflavus, E. gallinarum, and E. hirae. This study further revealed that RTE food products are reservoirs of potential virulent enterococci with antibiotic-resistant capabilities. This provides useful data for risk assessment and indicates that these foods may present a potential public health risk to consumers.