Veterinary Drug Residues in the Food Chain as an Emerging Public Health Threat: Sources, Analytical Methods, Health Impacts, and Preventive Measures.

Veterinary medications are necessary for both contemporary animal husbandry and food production, but their residues can linger in foods obtained from animals and pose a dangerous human risk. In this review, we aim to highlight the sources, occurrence, human exposure pathways, and human health effects of drug residues in food-animal products. Following the usage of veterinary medications, pharmacologically active compounds known as drug residues can be found in food, the environment, or animals. They can cause major health concerns to people, including antibiotic resistance development, the development of cancer, teratogenic effects, hypersensitivity, and disruption of normal intestinal flora. Drug residues in animal products can originate from variety of sources, including water or food contamination, extra-label drug use, and ignoring drug withdrawal periods. This review also examines how humans can be exposed to drug residues through drinking water, food, air, and dust, and discusses various analytical techniques for identifying these residues in food. Furthermore, we suggest some potential solutions to prevent or reduce drug residues in animal products and human exposure pathways, such as implementing withdrawal periods, monitoring programs, education campaigns, and new technologies that are crucial for safeguarding public health. This review underscores the urgency of addressing veterinary drug residues as a significant and emerging public health threat, calling for collaborative efforts from researchers, policymakers, and industry stakeholders to develop sustainable solutions that ensure the safety of the global food supply chain.

Nanoengineered Silica-Based Biomaterials for Regenerative Medicine.

The paradigm of regenerative medicine is undergoing a transformative shift with the emergence of nanoengineered silica-based biomaterials. Their unique confluence of biocompatibility, precisely tunable porosity, and the ability to modulate cellular behavior at the molecular level makes them highly desirable for diverse tissue repair and regeneration applications. Advancements in nanoengineered silica synthesis and functionalization techniques have yielded a new generation of versatile biomaterials with tailored functionalities for targeted drug delivery, biomimetic scaffolds, and integration with stem cell therapy. These functionalities hold the potential to optimize therapeutic efficacy, promote enhanced regeneration, and modulate stem cell behavior for improved regenerative outcomes. Furthermore, the unique properties of silica facilitate non-invasive diagnostics and treatment monitoring through advanced biomedical imaging techniques, enabling a more holistic approach to regenerative medicine. This review comprehensively examines the utilization of nanoengineered silica biomaterials for diverse applications in regenerative medicine. By critically appraising the fabrication and design strategies that govern engineered silica biomaterials, this review underscores their groundbreaking potential to bridge the gap between the vision of regenerative medicine and clinical reality.

Genetic Mutations in FKS1 Gene Associated with Acquired Echinocandin Resistance in Candida parapsilosis Complex.

Candida parapsilosis complex has recently received special attention due to naturally occurring FKS1 polymorphism associated with high minimal inhibitory concentrations for echinocandin and the increase of clonal outbreaks of strains resistant to commonly used antifungals such as fluconazole. Despite the previous fact, little is known about the genetic mechanism associated with echinocandin resistance. Therefore, the present study was designed to investigate the mechanism of acquired echinocandin resistance in C. parapsilosis complex strains. A total of 15 clinical C. parapsilosis complex isolates were sub-cultured for 30 days at a low concentration of micafungin at ½ the lowest MIC value of the tested isolates (0.12 µg/ml). After culturing, all the isolates were checked phenotypically for antifungal resistance and genotypically for echinocandin resistance by checking FKS1 gene hot spot one (HS1) and HS2 mutations. In vitro induction of echinocandin resistance confirmed the rapid development of resistance at low concentration micafungin, with no difference among C. parapsilosis, C. metapsilosis, and C. orthopsilosis in the resistance development. For the first time we identified different FKS1 HS1 and or HS2 mutations responsible for echinocandin resistance such as R658S and L1376F in C. parapsilosis, S656X, R658X, R658T, W1370X, X1371I, V1371X, and R1373X (corresponding to their location in C. parapsilosis) in C. metapsilosis, and L648F and R1366H in C. orthopsilosis. Our results are of significant concern, since the rapid development of resistance may occur clinically after short-term exposure to antifungals as recently described in other fungal species with the potential of untreatable infections.

After the Hurricane: Anti-COVID-19 Drugs Development, Molecular Mechanisms of Action and Future Perspectives.

Severe Acute Respiratory Syndrome Coronavirus-2 (SARS-CoV-2) is a new coronavirus in the Coronaviridae family. The COVID-19 pandemic, caused by SARS-CoV-2, has undoubtedly been the largest crisis of the twenty-first century, resulting in over 6.8 million deaths and 686 million confirmed cases, creating a global public health issue. Hundreds of notable articles have been published since the onset of this pandemic to justify the cause of viral spread, viable preventive measures, and future therapeutic approaches. As a result, this review was developed to provide a summary of the current anti-COVID-19 drugs, as well as their timeline, molecular mode of action, and efficacy. It also sheds light on potential future treatment options. Several medications, notably hydroxychloroquine and lopinavir/ritonavir, were initially claimed to be effective in the treatment of SARS-CoV-2 but eventually demonstrated inadequate activity, and the Food and Drug Administration (FDA) withdrew hydroxychloroquine. Clinical trials and investigations, on the other hand, have demonstrated the efficacy of remdesivir, convalescent plasma, and monoclonal antibodies, 6-Thioguanine, hepatitis C protease inhibitors, and molnupiravir. Other therapeutics, including inhaled medicines, flavonoids, and aptamers, could pave the way for the creation of novel anti-COVID-19 therapies. As future pandemics are unavoidable, this article urges immediate action and extensive research efforts to develop potent specialized anti-COVID-19 medications.

Synthesis, Physicochemical Characterization using a Facile Validated HPLC Quantitation Analysis Method of 4-Chloro-phenylcarbamoyl-methyl Ciprofloxacin and Its Biological Investigations.

A novel derivative of ciprofloxacin (Cpx) was synthesized and characterized using various analytical techniques, including FT-IR spectroscopy, UV-Vis spectroscopy, TEM and SEM analysis, 1H NMR, 13C NMR, and HPLC analysis. The newly prepared Cpx derivative (Cpx-Drv) exhibited significantly enhanced antibacterial properties compared to Cpx itself. In particular, Cpx-Drv demonstrated a 51% increase in antibacterial activity against S. aureus and a 30% improvement against B. subtilis. It displayed potent inhibitory effects on topoisomerases II (DNA gyrase and topoisomerase IV) as potential molecular targets, with IC50 values of 6.754 and 1.913 µg/mL, respectively, in contrast to Cpx, which had IC50 values of 2.125 and 0.821 µg/mL, respectively. Docking studies further supported these findings, showing that Cpx-Drv exhibited stronger binding interactions with the gyrase enzyme (PDB ID: 2XCT) compared to the parent Cpx, with binding affinities of -10.3349 and -7.7506 kcal/mole, respectively.

Enumeration, antimicrobial resistance and genomic characterization of extended-spectrum ß-lactamases producing Escherichia coli from supermarket chicken meat in the United Arab Emirates.

The occurrence and counts of extended-spectrum ß-lactamase (ESBL)-producing Escherichia coli in retail chicken sold in the United Arab Emirates (UAE) were investigated in this study. Results indicated that 79.68 % of chicken carcasses (251/315) sampled from UAE supermarkets harbored ESBL-producing E. coli. About half (51.75 % [163/315]) of the tested samples had an ESBL-producing E. coli count range between ≥3 log10 and < 5 log10 CFU/g. The antimicrobial resistance profiles of a subset of 100 isolates showed high rates of non-susceptibility to clinically significant antibiotics, particularly ciprofloxacin (80 %) and cefepime (46 %). Moreover, 7 % of the isolates exhibited resistance to colistin, with PCR-based screening revealing the presence of the mcr-1 gene in all colistin-resistant isolates. Multiplex PCR screening identified blaCTX-M and blaTEM genes as the most frequently presented genes among the phenotypically confirmed ESBL-producing E. coli. Further whole-genome sequencing and bioinformatic analysis of 27 ESBL-producing E. coli isolates showed that the gene family blaCTX group 1 was the most prevalent, notably CTX-M-55 (55.55 % [15/27]), followed by CTX-M-15 (22.22 % [6/27]). The most common sequence types (STs) were ST359 and ST1011, with three evident clusters identified based on phylogenomic analysis, aligned with isolates from specific production companies. Analysis of plasmid incompatibility types revealed IncFIB, IncFII, Incl2, and IncX1 as the most commonly featured plasmids. The findings of this study indicate a noticeable prevalence and high counts of ESBL-producing E. coli in chicken sampled from supermarkets in the UAE. The high rates of antimicrobial resistance to clinically important antibiotics highlight the potential public health risk associated with consuming chicken contaminated with ESBL-producing E. coli. Overall, this study emphasizes the importance of continued antimicrobial resistance monitoring in the UAE food chain and calls for further exposure risk assessment of the consumption of ESBL-producing E. coli via chicken meat.

Antifungal Resistance and Genotyping of Clinical Candida parapsilosis Complex in Japan.

Non-albicans Candida infections have recently gained worldwide attention due to their intrinsic resistance to different antifungal agents and the limited therapeutic options for treating them. Although the Candida parapsilosis complex is reported to be the second or third most prevalent Candida spp., little information is available on the prevalence of antifungal resistance along with genotyping of the C. parapsilosis complex. In this study, we aimed to evaluate the prevalence of antifungal resistance, the genetic basis of such resistance, and the genotyping of C. parapsilosis complex isolates that were recovered from hospitalized patients in Japan from 2005 to 2019. Our results indicated that, with the exception of one single C. metapsilosis isolate that was dose-dependently susceptible to fluconazole, all other isolates were susceptible or showed wild phenotypes to all tested antifungals, including azoles, echinocandins, amphotericin B, and flucytosine. Molecular analyses for azole and echinocandin resistance via evaluating ERG11 mutation and FKS1 hotspot one (HS1) and hotspot two (HS2) mutations, respectively, confirmed the phenotypic results. Genotyping of our isolates confirmed that they belong to 53 different but closely related genotypes, with a similarity percentage of up to 90%. Our results are of significant concern, since understanding the genetic basis of echinocandin resistance in the C. parapsilosis complex as well their genotyping is essential for directing targeted therapy, identifying probable infection sources, and developing strategies for overcoming epidemic spread.

Emergency Vaccination as a Control Strategy against Sheeppox Outbreak in a Highly Susceptible Population.

This study aimed to investigate a sheeppox outbreak in a highly susceptible naive sheep population in Kharsit village, Gharbia Governorate, Egypt. Moreover, to compare commercial sheeppox vaccines, the Romanian strain and RM-65 vaccines, as emergency vaccination against sheeppox under field conditions. In December 2018, a sheeppox outbreak occurred in a flock of 65 sheep upon the purchase of an apparently healthy ewe from outside the village. This ewe showed a systemic disease with cutaneous lesions after a few days, thereafter more cases began to appear. Cutaneous lesions in other sheep in the flock in the form of macules, papules, and scabs were common in wool-less areas of the body, in addition to fever and respiratory disorders. Postmortem findings revealed the congestion of visceral organs with apparent gross pathology of the lung. Biopsies of cutaneous lesions and visceral organs were collected, and sheeppox was identified by histopathology and transmission electron microscopy, which showed the existence of sheeppox cells and intracytoplasmic brick-shape sheeppox virions. The Romanian strain and RM-65 vaccines were used for the emergency vaccination for two different groups of animals and the third group was left as a control group. Serum samples were collected before vaccination as well as 21 days post-vaccination, and serum protein fractionation analysis was performed for all groups. The outbreak ended after 2.5 months, the cumulative incidence was 66.2%, and the overall case fatality was 51.1%. There was significantly higher protection against sheeppox infection and mortalities among RM-65 vaccine immunized group compared to Romanian strain vaccine-immunized animals at p < 0.05. RM-65-vaccinated animals did not show sheeppox cases or mortalities, compared to Romanian strain-vaccinated animals, which had mild pox signs in 78% of animals and case fatality of 35.7%. The serum protein analysis also indicated the superior performance of the RM-65 vaccine; it increased the level of α1-globulin and ß-globulin compared to the Romanian strain, which increased the level of ß-globulin only. The current study shows a better performance of the tested RM-65 than the Romanian strain vaccine for emergency vaccination against sheeppox under field conditions. These findings point to the validity of emergency vaccination against sheeppox and the importance of the comparative field evaluation of vaccines; however, wide-scale studies are required for further evaluation. Future investigation of whether the Romanian strain itself or vaccine-production-related issues are responsible for these findings is required.

Review on Bovine Tuberculosis: An Emerging Disease Associated with Multidrug-Resistant Mycobacterium Species.

Bovine tuberculosis is a serious infectious disease affecting a wide range of domesticated and wild animals, representing a worldwide economic and public health burden. The disease is caused by Mycobacteriumbovis and infrequently by other pathogenic mycobacteria. The problem of bovine tuberculosis is complicated when the infection is associated with multidrug and extensively drug resistant M. bovis. Many techniques are used for early diagnosis of bovine tuberculosis, either being antemortem or postmortem, each with its diagnostic merits as well as limitations. Antemortem techniques depend either on cellular or on humoral immune responses, while postmortem diagnosis depends on adequate visual inspection, palpation, and subsequent diagnostic procedures such as bacterial isolation, characteristic histopathology, and PCR to reach the final diagnosis. Recently, sequencing and bioinformatics tools have gained increasing importance for the diagnosis of bovine tuberculosis, including, but not limited to typing, detection of mutations, phylogenetic analysis, molecular epidemiology, and interactions occurring within the causative mycobacteria. Consequently, the current review includes consideration of bovine tuberculosis as a disease, conventional and recent diagnostic methods, and the emergence of MDR-Mycobacterium species.

Azole and echinocandin resistance mechanisms and genotyping of Candida tropicalis in Japan: cross-boundary dissemination and animal-human transmission of C. tropicalis infection.

OBJECTIVES: To assess the prevalence and genetic basis of antifungal resistance mechanisms as well as the genotyping of Candida tropicalis from clinical and non-clinical sources in Japan. METHODS: Eighty C. tropicalis isolates, including 32 clinical isolates recovered from 29 patients and 48 non-clinical isolates recovered from 24 different sources (animals and the environment) were evaluated. All isolates were tested phenotypically for resistance to a wide range of antifungals and genotypically for resistance mechanisms to azole and echinocandin. Furthermore, all the isolates were genotyped by multilocus sequence typing (MLST). RESULTS: Phenotypically, 30.2% (16/53) of the isolates were azole-resistant, with high levels of azole resistance among clinical isolates (51.7%; 15/29) and low levels (4.2%; 1/24) among non-clinical isolates. None of the isolates were reported as echinocandin resistant, with 60.4% (32/53) of the isolates intermediate to caspofungin. Azole resistance was basically attributed to high expression levels of drug efflux transporter genes (CDR2 and CDR3), transcription factors (TAC1 and UPC2) and ergosterol biosynthesis pathway HMG gene. No FKS1 hot spot 1 (HS1) or HS2 missense mutations were detected in any of the isolates. MLST analysis revealed 36 different sequence types (STs), with the first identification of 23 new STs. Phylogenetic analysis confirmed the close relationship between the clinical and non-clinical isolates, with identifications of ST232 and ST933 among patients and marine mammals. CONCLUSION: Our results confirmed the emergence of azole resistance in C. tropicalis in Japan. Furthermore, phylogenetic analysis confirmed the transboundary dissemination and cross-transmission of C. tropicalis between humans and animals.

Prevalence of Antifungal Resistance, Genetic Basis of Acquired Azole and Echinocandin Resistance, and Genotyping of Candida krusei Recovered from an International Collection.

This study was designed to evaluate the prevalence of antifungal resistance, genetic mechanisms associated with in vitro induction of azole and echinocandin resistance and genotyping of Candida krusei, which is intrinsically resistant to fluconazole and is recovered from clinical and nonclinical sources from different countries. Our results indicated that all the isolates were susceptible or had the wild phenotype (WT) to azoles, amphotericin B, and only 1.27% showed non-WT for flucytosine. Although 70.88% of the isolates were resistant to caspofungin, none of them were categorized as echinocandin-resistant as all were susceptible to micafungin and no FKS1 hot spot 1 (HS1) or HS2 mutations were detected. In vitro induction of azole and echinocandin resistance confirmed the rapid development of resistance at low concentrations of fluconazole (4 µg/ml), voriconazole (0.06 µg/ml), and micafungin (0.03 µg/ml), with no difference between clinical and nonclinical isolates in the resistance development. Overexpression of ABC1 gene and FKS1 HS1 mutations were the major mechanisms responsible for azole and echinocandin resistance, respectively. Genotyping of our 79 isolates coupled with 217 other isolates from different sources and geography confirmed that the isolates belong to two main subpopulations, with isolates from human clinical material and Asia being more predominant in cluster 1, and environmental and animals isolates and those from Europe in cluster 2. Our results are of critical concern, since realizing that the C. krusei resistance mechanisms and their genotyping are crucial for guiding specific therapy and for exploring the potential infection source.

High ß-Lactam and Quinolone Resistance of Enterobacteriaceae from the Respiratory Tract of Sheep and Goat with Respiratory Disease.

During the last decade's increase of antimicrobial resistance (AMR) in animals, animal-human transmission has become a major threat. Therefore, the present study aimed to evaluate the genetic basis of AMR in Gram-negative bacteria recovered from sheep and goats with respiratory disease. Nasal and ocular swabs were collected from 69 diseased animals, and 76 Gram-negative bacterial isolates were identified from 59 animals. All isolates were checked phenotypically for resistance and genotypically for different resistance mechanisms, including ß-lactam, quinolone, and aminoglycoside resistance. Our results demonstrated that 9.2% (95% CI 4.5-17.8%) of the isolates were multidrug-resistant, with high resistance rates to ß-lactams and quinolones, and 11.8% (95% CI 6.4-21%) and 6.6% (95% CI 2.8-14.5%) of the isolates were phenotypically positive for AmpC and ESBL, respectively. Genotypically, blaTEM was the most identified ß-lactamase encoding gene in 29% (95% CI 20-40%) of the isolates, followed by blaSHV (14.5%, 95% CI 8.3-24.1%) and blaCTX-M (4%, 95% CI 1.4-11%). Furthermore, 7.9% (95% CI 3.7-16.2%) of the isolates harbored plasmid-mediated quinolone resistance gene qnrS. Our study revealed for the first time to our knowledge high ß-lactam and quinolone resistance associated with the bacteria recovered from sheep and one goat with respiratory disease. Furthermore, different antimicrobial resistant determinants were identified for the first time from animals in Africa, such as blaLEN-13/55, blaTEM-176 and blaTEM-198/214. This study highlights the potential role of sheep and goats in disseminating AMR determinants and/or resistant bacteria to humans. The study regenerates interest for the development of a One Health approach to combat this formidable problem.

In Vitro Characterization of Twenty-One Antifungal Combinations against Echinocandin-Resistant and -Susceptible Candida glabrata.

This study was designed to analyze the interaction of 21 antifungal combinations consisting of seven major antifungal agents against 11 echinocandin- susceptible and six-resistant C. glabrata isolates. The combinations were divided into five major groups and were evaluated by checkerboard, disc diffusion, and time-killing assays. Synergy based on the fractional inhibitory concentration index of ≤0.50 was observed in 17.65-29.41% of the cases for caspofungin combinations with azoles or amphotericin B. Amphotericin B combination with azoles induced synergistic interaction in a range of 11.76-29.41%. Azole combinations and 5-flucytosine combinations with azoles or amphotericin B did not show synergistic interactions. None of the 21 combinations showed antagonistic interactions. Interestingly, 90% of the detected synergism was among the echinocandin-resistant isolates. Disk diffusion assays showed that the inhibition zones produced by antifungal combinations were equal to or greater than those produced by single drugs. The time-killing assay showed the synergistic action of caspofungin combination with fluconazole, voriconazole, and posaconazole, and the amphotericin B-5-flucytosine combination. Furthermore, for the first time, this assay confirmed the fungicidal activity of caspofungin-voriconazole and amphotericin B-5-flucytosine combinations. The combination interactions ranged from synergism to indifference and, most importantly, no antagonism was reported and most of the synergistic action was among echinocandin-resistant isolates.

High ß-lactam resistance in Gram-negative bacteria associated with kennel cough and cat flu in Egypt.

Antimicrobial resistance within pets has gained worldwide attention due to pets close contact with humans. This report examined at the molecular level, the antimicrobial resistance mechanisms associated with kennel cough and cat flu. 1378 pets in total were assessed for signs of respiratory infection, and nasal and conjunctival swabs were collected across 76 diseased animals. Phenotypically, 27% of the isolates were characterized by multidrug resistance and possessed high levels of resistance rates to ß-lactams. Phenotypic ESBLs/AmpCs production were identified within 40.5% and 24.3% of the isolates, respectively. Genotypically, ESBL- and AmpC-encoding genes were detected in 33.8% and 10.8% of the isolates, respectively, with blaSHV comprising the most identified ESBL, and blaCMY and blaACT present as the AmpC with the highest levels. qnr genes were identified in 64.9% of the isolates, with qnrS being the most prevalent (44.6%). Several antimicrobial resistance determinants were detected for the first time within pets from Africa, including blaCTX-M-37, blaCTX-M-156, blaSHV-11, blaACT-23, blaACT25/31, blaDHA-1, and blaCMY-169. Our results revealed that pets displaying symptoms of respiratory illness are potential sources for pathogenic microbes possessing unique resistance mechanisms which could be disseminated to humans, thus leading to the development of severe untreatable infections in these hosts.

Evaluation of Surveyor nuclease for rapid identification of FKS genes mutations in Candida glabrata.

INTRODUCTION: Infections with Candida glabrata have recently gained worldwide attention owing to its association with long hospitalizations and high mortality rates. This problem is highlighted when the infection is associated with echinocandin resistance, which is used for first-line therapy. Echinocandin resistance is exclusively attributed to functional mutations in FKS genes, and especially in hot spot (HS) regions. Unfortunately, few studies have focused on the rapid identification of FKS mutations associated with echinocandin resistance in C. glabrata. This study was intended to evaluate and validate the use of Surveyor nuclease assay (SN) for detection of FKS gene mutations. METHODS: SN was evaluated against three segments of FKS1 and FKS2 genes including whole gene, regions including all HSs, and the region including only HS1. RESULTS: Our results showed that SN results are basically dependent on the type of gene as well as the segment type. Interestingly, SN can detect mutations in the region containing HS1 in both FKS1 and FKS2 genes. Furthermore, SN can detect mutations in the segment containing all HS regions for FKS1 but not FKS2. SN was unable to detect mutations in the whole FKS1 and FKS2 genes. CONCLUSIONS: As far as we know, this is the first study to validate SN for rapid identification of FKS gene mutations. This assay could be used as a sample for rapid identification of mutations associated with HS1 region in FKS genes, which have a predominant role for echinocandin resistance induction in C. glabrata.

High-Speed Quenching Probe-Polymerase Chain Reaction Assay for the Rapid Detection of Carbapenemase-Producing Gene Using GENECUBE: A Fully Automatic Gene Analyzer.

BACKGROUND: The prevalence of carbapenemase-producing organisms (CPOs) globally poses a public health threat; however, detecting carbapenemases is a challenge because of their variety. METHODS: GENECUBE, a fully automated gene analyzer, detects a target gene in a short time and simultaneously detects its single nucleotide polymorphism. We used this property to develop for the first time a rapid assay for detecting CPOs from cultured bacteria using GENECUBE. The original primer-probe sets were used to detect blaKPC, blaIMP, blaVIM, blaNDM, and blaOXA-48-like from 149 CPOs (nine types) and 61 non-CPOs. RESULTS: The sensitivity, specificity, and positive and negative predictions of the GENECUBE assay were 100%. This assay detected carbapenemase single-producers and carbapenemase co-producers with 100% accuracy. The time required for detects of four types of carbapenemase at one run was about 30 min, but it took about 1 h to detect all five types. In addition, this assay performed the rapid detection and classification of blaOXA-48, blaOXA-181, blaOXA-232, and blaOXA-244 simultaneously. CONCLUSIONS: The GENECUBE assay is a promising tool for controlling the spread of CPOs and helping to select accurate and rapid antibiotic therapies.

Genetic Basis of Azole and Echinocandin Resistance in Clinical Candida glabrata in Japan.

Infections caused by Candida glabrata have caused worldwide concern, especially when they are associated with increasing echinocandin and azole resistance. In this study, we analyzed the molecular mechanisms of azole and echinocandin resistance in C. glabrata isolates obtained from hospitalized patients in Japan from 1997 to 2019. All isolates were checked phenotypically for resistance and genotypically for mutations in PDR1, ERG11, hot spot 1 (HS1), HS2, and HS3 of FKS1, and HS1 and HS2 of FKS2, and all isolates were genotyped by multilocus sequence typing (MLST). Interestingly, 32.6% of the isolates were resistant to caspofungin, and 4.7% were resistant to micafungin. The isolates showed low rates of resistance to azoles, ranging from 2.3% to 9.3%, and only 4.7% of the isolates were non-wild type for flucytosine susceptibility. For the first time in Japan, 4.7% of the isolates were identified as multidrug-resistant strains. Nonsynonymous mutations in PDR1, including two novel mutations associated with azole resistance, were identified in 39.5% of the isolates, and a single nonsynonymous mutation was identified in ERG11 Nine isolates from the same patient harbored nonsynonymous mutations in HS1 of FKS2, and a single isolate harbored a single nonsynonymous mutation in HS1 of FKS1 MLST genotyping revealed 13 different sequence types (STs), with 3 new STs, and ST7 was the most prevalent among the patients (35%) and was associated with high resistance rates. Our results are of crucial clinical concern, since understanding the molecular mechanisms underlying fungal resistance is imperative for guiding specific therapy for efficient patient treatment and promoting strategies to prevent epidemic spread.

Comparative Evaluation of Five Assays for Detection of Carbapenemases with a Proposed Scheme for Their Precise Application.

The escalating problem of the dissemination of carbapenemase-producing bacteria (CPB) has gained worldwide attention. The prompt diagnosis of CPB and precise identification of carbapenemases are imperative to enable specific antibiotic therapy and control the spread of these bacteria. The present study was designed to assess the performance of five important assays for the detection of carbapenemases. The modified carbapenem inactivation method (mCIM), CARBA-5, GeneXpert Carba-R, BD MAX Check-Points CPO, and GeneFields CPE assays were evaluated with an international collection of 159 bacterial isolates, including 93 CPB and 66 non-CPB isolates. The overall accuracy/sensitivity/specificity for carbapenemase detection were 100% (95% CI, 97.7%-100%)/100% (95% CI, 96.1%-100%)/100% (95% CI, 94.6%-100%) for mCIM, 98.7% (95% CI, 95.5%-99.9%)/97.9% (95% CI, 92.5%-99.7%)/100% (95% CI, 94.6%-100%) for CARBA-5, 96.9% (95% CI, 92.8%-99%)/95.7% (95% CI, 89.4%-98.8%)/98.5% (95% CI, 91.8%-99.9%) for GeneXpert Carba-R, 94.3% (95% CI, 89.5%-97.4%)/90.3% (95% CI, 82.4%-95.5%)/100% (95% CI, 94.6%-100%) for BD MAX Check-Points CPO, and 86.2% (95% CI, 79.8%-91.1%)/77.4% (95% CI, 67.6%-85.5%)/98.5% (95% CI, 91.8%-100%) for GeneFields CPE. Interestingly, mCIM and CARBA-5 assays showed 100% accuracy/sensitivity/specificity for detection of the target genes. Furthermore, all the other assays showed comparable high accuracy (96.9% to 100%), sensitivity (100%), and specificity (96.4% to 100%) for the detection of the target genes. On the basis of these results, a new scheme was proposed for their efficient application. These results confirmed the high sensitivity of the evaluated assays, and the proposed scheme is reliable and improves the overall sensitivity and specificity of the assays.