Screening for Antibiotic Resistance Genes in Bacteria and the Presence of Heavy Metals in the Upstream and Downstream Areas of the Wadi Hanifah Valley in Riyadh, Saudi Arabia.

Valley surface water is considered a focal public health concern owing to the presence of multi-drug-resistant bacteria. The distribution of antimicrobial resistance (AMR) bacteria in the surface water is affected by the presence of multiple factors, including antibiotics coming from wastewater discharge or other contaminant sources such as pharmaceuticals, biocides, and heavy metals. Furthermore, there is evidence suggesting that high levels of antibiotic resistance genes (ARGs) can be transferred within bacterial communities under the influence of heavy metal stress. Hence, the primary aim of this study is to investigate the presence of heavy metals and bacterial ARGs in upstream as well as downstream locations of Wadi Hanifah Valley in Riyadh, Saudi Arabia. Sample collection was conducted at eighteen surface water sites within the valley in total. The selection of ARGs was associated with the most common antibiotics, including ß-lactam, tetracycline, erythromycin, gentamicin, sulphonamide, chloramphenicol, vancomycin, trimethoprim, and colistin antibiotics, which were detected qualitatively using polymerase chain reaction (PCR) technology. The tested antibiotic resistance genes (ARGs) included (blaNDM-1 (for the antibiotic class Beta-lactamases), mecA (methicillin-resistant Staphylococcus aureus), tet(M) and tet(B) (for the antibiotic class Tetracycline), ampC (for the antibiotic class Beta-lactamases), vanA (for the antibiotic class vancomycin), mcr-1 (for the antibiotic class colistin), erm(B) (for the antibiotic class erythromycin), aac6'-Ie-aph2-Ia (for the antibiotic class Gentamicin), sulII (for the antibiotic class sulphonamide), catII (for the antibiotic class Chlorophincol), and dfrA1 (for the antibiotic class trimethoprim). Moreover, an assessment of the levels of heavy metals such as lithium (Li), beryllium (Be), chromium (Cr), cobalt (Co), arsenic (As), cadmium (Cd), tin (Sn), mercury (Hg), and lead (Pb) was conducted by using inductively coupled plasma mass spectrometry (ICPMS). According to our findings, the concentrations of sulphonamide, erythromycin, and chloramphenicol ARGs (erm(B), sulII, and catII) were observed to be the most elevated. Conversely, two ARGs, namely mecA and mcr-1, were not detected in the samples. Moreover, our data illustrated a significant rise in ARGs in the bacteria of water samples from the upstream sites as compared with the water samples from the downstream sites of Wadi Hanifah Valley. The mean concentration of Li, Be, Cr, Co, As, Cd, Sn, Hg, and Pb in the water samples was estimated to be 37.25 µg/L, 0.02 µg/L, 0.56 µg/L,0.32 µg/L, 0.93 µg/L, 0.01 µg/L, 200.4 µg/L, 0.027 µg/L, and 0.26 µg/L, respectively, for the selected 18 sites. Furthermore, it was revealed that the concentrations of the screened heavy metals in the water samples collected from various sites did not surpass the maximum limits set by the World Health Organization (WHO). In conclusion, this study offers a concise overview of the presence of heavy metals and ARGs in water samples obtained from the Wadi Hanifah Valley in Riyadh, KSA. Such findings will contribute to the ongoing monitoring and future risk assessment of ARGs spread in surface water.

Evaluating the effectiveness of the Ministry of Health restriction policy on seasonal antibiotic consumption trends in Saudi Arabia, 2016-2020.

Background: Understanding antibiotic consumption patterns over time is essential to optimize prescribing practices and minimizing antimicrobial resistance. This study aimed to determine whether the antibiotics restriction policy launched by the Saudi Ministry of Health in April 2018 has impacted antibiotic use by assessing changes and seasonal variations following policy enforcement. Methods: Quarterly sales data of J01 antibacterial for systemic use in standard units were obtained from the IQVIA-MIDAS database, spanning from the first quarter of 2016 to the last quarter of 2020. Antibiotics consumption was measured in defined daily doses per 1,000 inhabitant per day- in a quarter (DDDdq). A comparative analysis of antibiotic consumption pre- and post-policy periods introduction was conducted by computing the average consumption values for each period. Statistical comparison of the mean differences between the two periods were then made using independent samples t-test, Mann-Whitney U Test where needed. Time series analysis was employed to estimate the projected antibiotic consumption in the post-policy period if the restriction policy had not been implemented, which was then compared to actual consumption values to evaluate the effectiveness of the restriction policy. Results: During the pre-policy, there were seasonal trends of the total and oral antibiotic consumption through quarters, with higher consumption observed in the first and fourth quarters. In contrast, parenteral antibiotic consumption did not appear to follow a clear seasonal pattern. Following the restriction policy, there was a significant reduction in total and oral antibiotic use, with mean reductions of -96.9 DDDdq (p-value = 0.002) and -98 DDDdq (p-value = 0.002), respectively. Conversely, a significant increase in parenteral antibiotic consumption was observed with a mean increase of +1.4 DDDdq (p-value < 0.0001). The comparison between the forecasted and actual models showed that the actual antibiotics consumption for total, oral, and parenteral were lower than the corresponding forecasted values by 30%, 31%, and 34%, respectively. Conclusion: Overall, our analysis of antibiotics consumption from 2016 to 2020 displays great success for the policy implemented by the Saudi Ministry of Health in significantly reducing the total and oral use of antibiotics. However, future studies are needed to explore the increased consumption of the parenteral antibiotics as well as the persistent high consumption patterns during the fall and winter months even after the implementation of the restriction policy.

Clonal Flux and Spread of Staphylococcus aureus Isolated from Meat and Its Genetic Relatedness to Staphylococcus aureus Isolated from Patients in Saudi Arabia.

In this study, we investigated both meat-derived and methicillin-resistant Staphylococcus aureus (MRSA), exploring their genetic relatedness to patient-derived MRSA isolates in Saudi Arabia. We collected 250 meat samples and identified 53 S. aureus isolates, with 79% being methicillin-sensitive Staphylococcus aureus (MSSA) and 21% being MRSA. Moreover, we included 80 clinically confirmed patient-derived MRSA isolates. We identified the most common S. aureus clone in both patients and retail meat. In meat, ST6 and ST97 were the most common clones in 55% of the MRSA isolates, and ST1153 and ST672 were the most common in 21% and 17% of the MSSA isolates. In patients, ST5 and ST6 were the predominant clones in 46% of the S. aureus isolates. CC5/ST5-SCCmecVc-t311 and CC361/ST672-SCCmecV-t3841 were common MRSA clones in both meat and patients. CC97 and CC361 clones were the second most prevalent S. aureus clones in meat and were relatively common in patients. Furthermore, we sequenced and characterized novel S. aureus strains ST8109, ST8110, and ST8111. The genomic similarities between meat- and patient-derived S. aureus isolates suggest that retail meat might be a reservoir for S.aureus and MRSA transmission. Therefore, a structured One Health approach is recommended for S. aureus dissemination, genetic characterization, antibiotic resistance, and impact on human health.

Interception of Epoxide ring to quorum sensing system in Enterococcus faecalis and Staphylococcus aureus.

Quorum sensing inhibitor (QSI) has been attracting attention as anti-virulence agent which disarms pathogens of their virulence rather than killing them. QSI marking cyclic peptide-mediated QS in Gram-positive bacteria is an effective tool to overcome the crisis of antibiotic-dependent chemotherapy due to the emergence of drug resistance strain, e.g., methicillin resistant Staphylococcus aureus (MRSA) and Vancomycin resistant Enterococci (VRE). From a semi-large-scale screening thus far carried out, two Epoxide compounds, Ambuic acid and Synerazol, have been found to efficiently block agr and fsr QS systems, suggesting that the Epoxide group is involved in the mode of action of these QSIs. To address this notion, known natural Epoxide compounds, Cerulenin and Fosfomycin were examined for QSI activity for the agr and fsr systems in addition to in silico and SAR studies. As a result, most of investigated Epoxide containing antibiotics correlatively interfere with QSI activity for the agr and fsr systems under sublethal concentrations.

The Silent Threat: Antimicrobial-Resistant Pathogens in Food-Producing Animals and Their Impact on Public Health.

The emergence of antimicrobial resistance (AMR) is a global health problem without geographic boundaries. This increases the risk of complications and, thus, makes it harder to treat infections, which can result in higher healthcare costs and a greater number of deaths. Antimicrobials are often used to treat infections from pathogens in food-producing animals, making them a potential source of AMR. Overuse and misuse of these drugs in animal agriculture can lead to the development of AMR bacteria, which can then be transmitted to humans through contaminated food or direct contact. It is therefore essential to take multifaceted, comprehensive, and integrated measures, following the One Health approach. To address this issue, many countries have implemented regulations to limit antimicrobial use. To our knowledge, there are previous studies based on AMR in food-producing animals; however, this paper adds novelty related to the AMR pathogens in livestock, as we include the recent publications of this field worldwide. In this work, we aim to describe the most critical and high-risk AMR pathogens among food-producing animals, as a worldwide health problem. We also focus on the dissemination of AMR genes in livestock, as well as its consequences in animals and humans, and future strategies to tackle this threat.

Detection of Escherichia coli O157:H7 in imported meat products from Saudi Arabian ports in 2017.

Escherichia coli O157:H7 is a foodborne pathogen, which causes various health conditions in humans, including fatigue, nausea, bloody diarrhoea and in some cases, even death. In 2017, 15.71% of the total imported food products in Saudi Arabia (SA) were meat-based. India and Brazil are two of the top five countries from where SA imports meat. According to the Saudi Food and Drug Authority, in 2017, at least 562, 280, and 50 samples of imported beef, chicken and sheep meat, respectively, were tested for the presence of E. coli O157:H7. Amongst these, E. coli O157:H7 was detected in respectively 6.80% and 2.20% of the tested beef meat samples imported from India and Brazil as well as in respectively 6.96% and 3.57% of the tested chicken samples imported from Brazil and Ukraine. Moreover, the pathogen was detected in 2.13% of the tested sheep meat samples imported from India. The present report provides evidence that imported meat can serve as the carrier of E. coli O157:H7, which may lead to epidemics within the Kingdom of Saudi Arabia.

Comparative genomic analysis of antibiotic resistance and virulence genes in Staphylococcus aureus isolates from patients and retail meat.

Introduction: Staphylococcus aureus is a significant human pathogen that poses a threat to public health due to its association with foodborne contamination and a variety of infections. The factors contributing to the pathogenicity of S. aureus include virulence, drug resistance, and toxin production, making it essential to monitor their prevalence and genetic profiles. This study investigated and compared the genomic characteristics of S. aureus isolates from retail meat and patients in Saudi Arabia. Methods: A total of 136 S. aureus isolates were obtained between October 2021 and June 2022:84 from patients and 53 from meat samples in Riyadh, Saudi Arabia. S. aureus isolates were identified using conventional methods and MALDI-TOF MS, and methicillin-resistant S. aureus (MRSA) was identified using VITEK2 and BD Phoenix systems. MRSA was confirmed phenotypically using chromogenic agar, and genotypically by detecting mecA. Genomic data were analyzed using BactopiaV2 pipeline, local BLAST, and MLST databases. Results: Antibiotic resistance genes were prevalent in both meat and patient S. aureus isolates, with high prevalence of tet38, blaZ, and fosB. Notably, all S. aureus isolates from patients carried multidrug-resistant (MDR) genes, and a high percentage of S. aureus isolates from meat also harbored MDR genes. Phenotypically, 43% of the S. aureus isolates from meat and 100% of the patients' isolates were MDR. Enterotoxin genes, including selX, sem, and sei, exhibited high compatibility between meat and patient S. aureus isolates. Virulence genes such as cap, hly/hla, sbi, and isd were found in all S. aureus isolates from both sources. Conclusion: Our study established a genetic connection between S. aureus isolates from meat and patients, showing shared antibiotic resistance and virulence genes. The presence of these genes in meat derived isolates underscores its role as a reservoir. Genomic relatedness also suggests potential transmission of resistance between different settings. These findings emphasize the necessity for a comprehensive approach to monitor and control S. aureus infections in both animals and humans.

Antimicrobial Sales Comparison before and after the Implementation of Nationwide Restriction Policy in Saudi Arabia.

Antimicrobial dispensing without a prescription has been identified as a significant contributor to the burgeoning crisis of antimicrobial resistance. To combat this, the Saudi Ministry of Health introduced a stringent antimicrobial restriction policy in mid-2018, mandating prescriptions for all antimicrobial drug dispensations at pharmacies. Therefore, this study aimed to assess the immediate impact of this policy on retail antimicrobial sales. To do so, we analyzed annual sales data from 2017 to 2019 sourced from the IQVIA-MIDAS® database, which included a range of antimicrobials, such as antibiotics, antifungals, and other related agents. The analysis revealed a notable reduction in overall antimicrobial sales by 23.2%, decreasing from 818.9 million SAR in 2017 to 648.4 million SAR in 2019. While the Wilcoxon signed-rank test indicated a statistically significant median reduction in total antimicrobial sales post-policy implementation (p = 0.0397), it is important to acknowledge that the long-term effects and adherence to the policy require further investigation. Notably, sales of amoxicillin dropped by 70% in 2019 compared to 2017, contributing largely to the decline. Conversely, a continuous increase in sales of some antimicrobial drugs following the restriction policy was observed, led by amoxicillin/clavulanic acid. Our data support the implementation of antimicrobial restriction measures as an effective means of controlling excessive antimicrobial sales and dispensing without prescriptions.

Draft Genome Sequence of Two Salmonella enterica Subspecies enterica Serovar Minnesota Strains Harboring Mcr-1.1 Gene Isolated from Chicken Meat in Saudi Arabia.

Salmonella has been reported as a high-priority pathogen by the World Health Organization. Here, we report the draft genome sequence of 2 Salmonella enterica serovar Minnesota strains isolated from chicken meat in Saudi Arabia (named SA49317 and SA49319) belonging to sequence type (ST) 548, revealing mobile colistin resistance (mcr)-1.1 gene for colistin resistance.

Assessment of disinfectant efficacy in reducing microbial growth.

The incidence of hospital- and community-acquired infections has been dramatically increased worldwide. Accordingly, hands hygiene and the use of disinfectants have been increased leading to the expansion in hand sanitizers production to meet public demand. This study was conducted to assess the efficiency of common disinfectants in the market of Riyadh, Saudi Arabia in inhibiting the microbial growth during the time of Coronavirus disease 2019 (COVID-19) pandemic. Five bacterial strains of commonly hospital-acquired infections (Pseudomonas aeruginosa, Escherichia coli, Salmonella enteritidis, Staphylococcus aureus, and Enterococcus faecalis) (ATCC reference strains and clinical isolates) were examined for their susceptibility against 18 disinfectants collected from the Saudi market. The tested 18 disinfectants were broadly clustered into different groups based on their active chemical composition as following: 12 products contained alcohol, 2 products had chlorhexidine, 3 products contained mixed concentration of alcohol/chlorhexidine and 1 product had a mixture of chlorhexidine/Hexamidine/Chlorocresol. By measuring the minimum inhibitory concentration (MIC) and the minimum bactericidal concentration (MBC), our results revealed that all the 18 disinfectants have reduced the microbial growth of all the tested strains. Generally, the MICs and the MBCs for the clinical strains are higher than those of the reference strains. Taken together, our findings showed that all tested products have high disinfectants' killing rate against microbes of different origins, which suggest the high quality of these disinfectants and the good surveillance practice by the local authorities in Saudi Arabia.

Complete Genome Sequence of a Colistin-Susceptible Salmonella enterica Serovar Minnesota Strain Harboring mcr-9 on an IncHI2/IncHI2A Plasmid, Isolated from Chicken Meat.

The recent emergence and dissemination of mobilized colistin resistance (mcr) genes have triggered extensive concerns globally. Here, we report the complete genome sequence of a colistin-susceptible Salmonella enterica serotype Minnesota strain (named SA18578), belonging to sequence type 548 (ST548) and carrying the mcr-9 gene on an IncHI2/IncHI2A plasmid, that was isolated from chicken meat in Saudi Arabia in 2020.