Resistance to cisplatin in human lung adenocarcinoma cells: effects on the glycophenotype and epithelial to mesenchymal transition markers.

The characteristics that grant the most malignancy to cancer cells are the ability to evade apoptotic mechanisms and the capacity to migrate beyond the boundaries of the original tissue. Studies by our own group and others show that changes in glycosylation are now considered hallmarks of cancer cells and are also able to impact tumor malignancy. This study aims to evaluate changes in the glycosylation profile of the A549 lung cancer cells brought about by the induction of a MDR phenotype as well as investigate the relationship between drug resistance, the cell glycophenotype and EMT. We induced resistance by employing a continuous treatment with cisplatin. Our results demonstrate overexpression of ABC transporters as well as anti-apoptotic members of the Bcl-2 family, leading to a MDR phenotype. The cells also undergo a classic EMT process, displaying the iconic cadherin switch and increased of both total and oncofetal fibronectin, coupled with increased cell motility. We also managed to show changes in the expression of both glycosyltransferases and the glycan epitopes they are responsible for building. We also suggest that perhaps not only changes in cell sialylation are common during resistance induction but are essential to it.

Interaction Between Near-Infrared Radiation and Temozolomide in a Glioblastoma Multiform Cell Line: A Treatment Strategy?

Photodynamic therapy (PDT) is a potential therapeutic modality against cancer, resulting from the interaction of a photosensitizer (PS) and radiation that generates damage to tumor cells. The use of near-infrared radiation (IR-A) is relevant because presents recognized biological effects, such as antioxidant, neuroprotective and antitumor effects. Glioblastoma is the most aggressive central nervous system (CNS) neoplasm with high proliferation and tissue invasion capacity and is resistant to radio and chemotherapy. Here, we evaluated in vitro the possible interaction of temozolomide (TMZ) with IR-A in a glioblastoma cell line (C6) and in a human keratinocyte cell line (HaCat) how non-tumor cell model, in an attempt to search for a new treatment strategy. The effects of TMZ, IR-A and the interaction between TMZ and IR-A was evaluated by viability exclusion with trypan blue. To perform the interaction experiments, we have chosen 10 µM TMZ and 4.5 J/cm2 of IR-A. From this, we evaluated cytotoxicity, cell proliferation, intracellular reactive oxygen species levels (ROS), as well as the process of cell migration and the P-gp and MRP-1 activity. Cell death mainly due to apoptosis, followed by necrosis, decreased cell proliferation, increased ROS levels, decreased cell migration and decreased P-gp and MRP1 activity were observed only when there was interaction between TMZ and IR-A in the C6 cell line. The interaction between TMZ and IR-A was not able to affect cell proliferation in the HaCat non-tumor cell line. Our results suggest that this interaction could be a promising approach and that in the future may serve as an antitumor strategy for PDT application.

Resistance to paclitaxel induces glycophenotype changes and mesenchymal-to-epithelial transition activation in the human prostate cancer cell line PC-3.

The development of the multidrug resistance phenotype is one of the major challenges faced in the treatment of cancer. The multidrug resistance phenotype is characterized by cross-resistance to drugs with different chemical structures and mechanisms of action. In this work, we hypothesized that the acquisition of resistance in cancer is accompanied by activation of the epithelial-to-mesenchymal transition process, where the tumor cell acquires a more mobile and invasive phenotype; a fundamental step in tumor progression and in promoting the invasion of other organs and tissues. In addition, it is known that atypical glycosylations are characteristic of tumor cells, being used as biomarkers. We believe that the acquisition of the multidrug resistance phenotype and the activation of epithelial-to-mesenchymal transition provoke alterations in the cell glycophenotype, which can be used as glycomarkers for chemoresistance and epithelial-to-mesenchymal transition processes. Herein, we induced the multidrug resistance phenotype in the PC-3 human prostate adenocarcinoma line through the continuous treatment with the drug paclitaxel. Our results showed that the induced cell multidrug resistance phenotype (1) acquired a mixed profile between epithelial and mesenchymal phenotypes and (2) modified the glycophenotype, showing an increase in the level of sialylation and in the number of branched glycans. Both mechanisms are described as indicators of poor prognosis.

Discovery of Potent Non-nucleoside Inhibitors of Dengue Viral RNA-Dependent RNA Polymerase from Fragment Screening and Structure-Guided Design.

Flavivirus NS5 RNA-dependent RNA polymerase (RdRp) is an important drug target. Whilst a number of allosteric inhibitors have been described for Hepatitis C virus RdRp, few have been described for DENV RdRp. In addition, compound screening campaigns have not yielded suitable leads for this enzyme. Using fragment-based screening via X-ray crystallography, we identified a biphenyl acetic acid fragment that binds to a novel pocket of the dengue virus (DENV) RdRp, in the thumb/palm interface, close to its active site (termed "N pocket"). Structure-guided optimization yielded nanomolar inhibitors of the RdRp de novo initiation activity, with low micromolar EC50 in DENV cell-based assays. Compound-resistant DENV replicons exhibited amino acid mutations that mapped to the N pocket. This is the first report of a class of pan-serotype and cell-active DENV RdRp inhibitors and provides a significant opportunity for rational design of novel therapeutics against this proven antiviral target.

Associations between resistance phenotype and gene expression in response to serial exposure to oxacillin and ciprofloxacin in Staphylococcus aureus.

This study was designed to delineate the relationship between resistance phenotypes and gene expression in wild-type (SAWT ), oxacillin-induced (SAOXA ), ciprofloxacin-induced (SACIP ) and clinically acquired antibiotic-resistant Staphylococcus aureus (SACA ) exposed to oxacillin (ß-lactam) and ciprofloxacin (fluoroquinolone). The phenotypic response and gene expression were varied with the antibiotic exposure. SAWT was highly resistant to oxacillin (MIC = 8 µg ml-1 ) after serial exposure to oxacillin, while the oxacillin susceptibility was not changed in SAWT when exposed to ciprofloxacin (MIC = 0·25 µg ml-1 ). The clinical isolate, SACA , was highly resistant to all classes of antibiotics used in this study. The increased resistance of SAOXA and SACIP to penicillinase-labile penicillins was attributed to the production of ß-lactamase, which is in good agreement with the overexpression of blaZ (>2-fold). The overexpression of efflux pump-related genes (norA, norB, norC, mdeA, mepR, mgrA and lmrS) was associated with the increased resistance of SACIP and SACA to aminoglycosides and quinolones. This study confirmed that the linkage between resistance phenotypes and molecular genotypes highly varied depending on intrinsic resistance profile, response to antibiotic exposure and genes conferring resistance. This study provides useful information for understanding the mechanisms of methicillin resistance in S. aureus in association with phenotypic and genotypic resistance determinants. SIGNIFICANCE AND IMPACT OF THE STUDY: The improvement in current standards is essential to accurately detect methicillin-resistant Staphylococcus aureus in consideration of various resistance phenotypes and genotypes. The varied and distinctive expression patterns of antibiotic resistance-related genes were observed in S. aureus exposed to oxacillin and ciprofloxacin. It is worth noting the relationship between resistance phenotype and resistance genotype in terms of MIC values and expression of antibiotic resistance determinants. This study provides useful information for understanding the mechanisms of methicillin resistance in S. aureus in association with phenotypic and genotypic resistance determinants.

Neuronal Trans-Differentiation in Prostate Cancer Cells.

BACKGROUND: Neuroendocrine (NE) differentiation in prostate cancer (PCa) is an aggressive phenotype associated with therapy resistance. The complete phenotype of these cells is poorly understood. Clinical classification is based predominantly on the expression of standard NE markers. METHODS: We analyzed the phenotype of NE carcinoma of the prostate utilizing in vitro methods, in silico, and immunohistochemical analyses of human disease. RESULTS: LNCaP cells, subjected to a variety of stressors (0.1% [v/v] fetal bovine serum, cyclic AMP) induced a reproducible phenotype consistent with neuronal trans-differentiation. Cells developed long cytoplasmic processes resembling neurons. As expected, serum deprived cells had decreased expression in androgen receptor and prostate specific antigen. A significant increase in neuronal markers also was observed. Gene array analysis demonstrated that LNCaP cells subjected to low serum or cAMP showed statistically significant manifestation of a human brain gene expression signature. In an in silico experiment using human data, we identified that only hormone resistant metastatic prostate cancer showed enrichment of the "brain profile." Gene ontology analysis demonstrated categories involved in neuronal differentiation. Three neuronal markers were validated in a large human tissue cohort. CONCLUSION: This study proposes that the later stages of PCa evolution involves neuronal trans-differentiation, which would enable PCa cells to acquire independence from the neural axis, critical in primary tumors. Prostate 76:1312-1325, 2016. © 2016 Wiley Periodicals, Inc.

Resistance testing of clinical herpes simplex virus type 2 isolates collected over 4 decades.

There is only little information about the role of mutations of the thymidine kinase (TK) and DNA polymerase (pol) genes of herpes simplex virus type 2 (HSV-2) for the development of antiviral resistance. In this study, the polymorphism of TK and DNA pol genes was examined in 82 clinical isolates collected routinely between 1973 and 2013. If novel, presently unclear or resistance-related mutations were found, the resistance phenotype against acyclovir (ACV) and foscarnet (FOS) was analyzed. The four novel amino acid changes G150D, A157T, R248W, L342W and the hitherto phenotypically unclear substitution T131M within the TK gene were identified as natural polymorphisms. Within the DNA pol gene, 17 novel substitutions and the to-date unclear change R628C were characterized as part of natural gene polymorphism. Two novel DNA pol mutations were linked to resistance (M910T) and weak susceptibility to ACV (684 insertion ED), respectively. In one isolate, the genomic cause of ACV resistance could not be identified. Phylogenetic analysis including sequences of this study and of the GenBank revealed a hierarchy of mutation clusters in TK displaying G39E as first common mutation step, followed by N78D and L140F. In conclusion, the present findings allow a deeper insight in the variability of HSV-2 TK and DNA pol genes. The most common substitution G39E can be excluded as unique cause of HSV-2 resistance. This study supports once more the importance of phenotypic adjustment of genotypic results to enhance the clinical utility of genotypic findings.

Genotype diversity and antibiotic resistance risk in Aeromonas hydrophila in Sichuan, China.

Sichuan is a significant aquaculture province in China, with a total aquaculture output of 1.72 × 106 tons in 2022. One of the most significant microorganisms hurting the Sichuan aquaculture is Aeromonas hydrophila, whose genotype and antibiotic resistance are yet unknown. This study isolated a total of 64 strains of A. hydrophila from various regions during September 2019 to June 2021 within Sichuan province, China. The technique of Multi-Locus Sequence Typing (MLST) was used for the purpose of molecular typing. Meanwhile, identification of antibiotic resistance phenotype and antibiotic resistance gene was performed. The findings of the study revealed that 64 isolates exhibited 29 sequence types (ST) throughout different regions in Sichuan, with 25 of these ST types being newly identified. Notably, the ST251 emerged as the predominant sequence type responsible for the pandemic. The resistance rate of isolated strains to roxithromycin was as high as 98.3%, followed by co-trimoxazole (87.5%), sulfafurazole (87.5%), imipenem (80%), amoxicillin (60%), and clindamycin (57.8%). Fifteen strains of A. hydrophila exhibited resistance to medicines across a minimum of three categories, suggesting the development of multidrug resistance in these isolates. A total of 63 ARGs were detected from the isolates, which mediated a range of antibiotic resistance mechanisms, with deactivation and efflux potentially serving as the primary mechanisms of antibiotic resistance. This study revealed the diversity of A. hydrophila genotypes and the risk of antibiotic resistance in Sichuan, providing reference for scientific and effective control of A. hydrophila infection.

Toxin gene detection and antibiotic resistance of Clostridium perfringens from aquatic sources.

Clostridium perfringens is a zoonotic opportunistic pathogen that produces toxins that can cause necrotic enteritis and even "sudden death disease". This bacterium is widely distributed in the intestines of livestock and human, but there are few reports of distribution in aquatic animals (Hafeez et al., 2022). In order to explore the isolation rate of C. perfringens and the toxin genes they carry, 141 aquatic samples, including clams (Ruditapes philippinarum), oysters (Ostreidae), and mud snails (Bullacta exerata Philippi), were collected from the coastal areas of Shandong Province, China. C. perfringens strains were tested for cpa, cpb, etx, iap, cpb2, cpe, netB, and tpeL genes. 45 clam samples were boiled at 100 °C for 5 min before bacteria isolation. 80 strains were isolated from 141 samples with the positive rate being 57 %.And the positive rates of cooked clams was 87 % which was higher than the average. In detection of 8 toxin genes, all strains tested cpa positive, 3 strains netB positive, and 2 cpb and cpe, respectively. 64 strains were selected to analyze the antibiotic resistance phenotype of 10 antibiotics. The average antibiotic resistance rates of the strains to tetracycline, clindamycin, and ampicillin were 45 %, 20 %, and 16 % respectively, and the MIC of 4 strains to clindamycin was ≥128 µg/mL. A high isolation rate of C. perfringens from aquatic animals was shown, and it was isolated from boiled clams for the first time, in which cpe and netB toxin genes were detected for the first time too. The toxin encoded by cpe gene can cause food poisoning of human, thus the discoveries of this study have certain guiding significance for food safety. Antibiotics resistant C. perfringens of aquatic origin may arise from transmission in the terrestrial environment or from antibiotic contamination of the aquaculture environment and is of public health significance.

Comparative analysis of antimicrobial resistance phenotype and genotype of Riemerella anatipestifer.

Riemerella anatipestifer is one of the important bacterial pathogens that threaten the waterfowl farming industry. In this study, 157 suspected R. anatipestifer strains were isolated from diseased ducks and geese from seven regions of China during 2019-2020, and identified using multiple polymerase chain reaction (PCR). Antimicrobial susceptibility tests and whole-genome sequence (WGS) analysis were then performed for comparative analysis of antimicrobial resistance phenotypes and genotypes. The results showed that these strains were susceptible to florfenicol, ceftriaxone, spectinomycin, sulfafurazole and cefepime, but resistant to kanamycin, amikacin, gentamicin, and streptomycin, exhibiting multiple antimicrobial resistance phenotypes. WGS analysis revealed a wide distribution of genotypes among the 157 strains with no apparent regional pattern. Through next-generation sequencing analysis of antimicrobial resistance genes, a total of 88 resistance genes were identified. Of them, 19 tetracycline resistance genes were most commonly found, followed by 15 efflux pump resistance genes, 11 glycopeptide resistance genes and seven macrolide resistance genes. The 157 R. anatipestifer strains contained 42-55 resistance genes each, with the strains carrying 47 different resistance genes being the most abundant. By comparing the antimicrobial resistance phenotype and genotype, it was observed that a high correlation between them for most antimicrobial resistance properties was detected, except for a difference in aminoglycoside resistance phenotype and genotype. In conclusion, 157 R. anatipestifer strains exhibited severe multiple antimicrobial resistance phenotypes and genotypes, emphasizing the need for improved antimicrobial usage guidelines. The wide distribution and diverse types of resistance genes among these strains provide a foundation for studying novel mechanisms of antimicrobial resistance.

Characteristics of Nontyphoid Salmonella Isolated from Human, Environmental, Animal, and Food Samples in Burkina Faso: A Systematic Review and Meta-Analysis.

Salmonella is one of the world's leading causes of zoonotic and foodborne illnesses. Recently, antimicrobial resistance (AMR) has become one of the most critical challenges to public health and food safety. Herein, we employed a meta-analysis to determine the pooled prevalence and spatiotemporal distribution of serovars and antimicrobial resistance in NTS in Burkina Faso. To find eligible articles, a comprehensive literature search of PubMed, African Journals Online, ScienceDirect, Google Scholar, and the gray literature (university libraries) in Burkina was conducted for the period from 2008 to 2020. Studies meeting the inclusion criteria were selected and assessed for risk of bias. To assess the temporal and spatial relationships between serotypes and resistant strains from humans, animals, food, and the environment, a random-effects statistical model meta-analysis was carried out using the Comprehensive Meta-Analysis Version 3.0 program. The NTS prevalence rates were 4.6% (95% CI: 3-7) and 20.1% (95% CI: 6.6-47.4) in humans and animals, respectively, and 16.8% (95% CI: 10.5-25.8) and 15.6% (95% CI: 8.2-27.5) in food and the environment, respectively. Most NTS serovars were S. Derby, reported both in food and animals, and S. Typhimurium, reported in humans, while S. Croft II, S. Jodpur II, and S. Kentucky were the most prevalent in the environment. NTS isolates were highly resistant to erythromycin, amoxicillin, cefixime, and cephalothin, with a pooled prevalence of multidrug resistance of 29% (95% CI: 14.5-49.5). The results of this review show a high diversity of Salmonella serotypes, as well as high antibiotic resistance in Salmonella isolates from animal, human, food, and environmental samples in Burkina, calling for a consolidated "One Health" approach to better understand the drivers of pathogen emergence, spread, and antimicrobial resistance, as well as the formulation of intervention measures needed to limit the risk associated with the disease.

Resistance phenotype and genetic features of a heterogeneous vancomycin intermediate-resistant Staphylococcus aureus strain from an immunocompromised patient.

Strain C1 was successfully isolated from an immunosuppressed patient with persistent bacteremia, who had not previously been exposed to glycopeptide antibiotics. This strain was found to be a heterogeneous vancomycin intermediate-resistant Staphylococcus aureus (hVISA). It is noteworthy that, following a brief period of vancomycin treatment, strains C6, C8, and C9, which were obtained from blood and other body parts, exhibited a significant reduction in heterogeneity as determined by population analysis profile-area under the curve (PAP-AUC) detection. Genotyping analysis revealed that these bacterial strains belonged to the same SCCmecIVa-ST59-t437-agrI genotype and shared the same virulome and resistome. In this study, a comparative genomics analysis was conducted between strain C1 and strain N315 to identify potential hVISA-associated mutations. Ultimately, a total of 205 mutation sites in 19 candidate genes, likely associated with the hVISA phenotype, were identified.

Molecular basis and evolutionary cost of a novel macrolides/lincosamides resistance phenotype in Staphylococcus haemolyticus.

Staphylococcus haemolyticus (S. haemolyticus) is a coagulase-negative Staphylococcus that has become one of the primary causes of nosocomial infection. After a long period of antibiotic use, S. haemolyticus has developed multiple resistance phenotypes for macrolides and lincosamides. Herein, we evaluated four S. haemolyticus clinical isolates, of which three had antibiotic resistance patterns reported previously. The fourth isolate was resistant to both erythromycin and clindamycin in the absence of erythromycin induction. This novel phenotype, known as constitutive macrolides-lincosamides-streptogramins resistance, has been reported in other bacteria but has not been previously reported in S. haemolyticus. Investigation of the isolate demonstrated a deletion in the methyltransferase gene ermC, upstream leader peptide. This deletion resulted in constitutive MLS resistance based on whole-genome sequencing and experimental verification. Continuous expression of ermC was shown to inhibit the growth of S. haemolyticus, which turned out to be the fitness cost with no MLS pressure. In summary, this study is the first to report constitutive MLS resistance in S. haemolyticus, which provides a better understanding of MLS resistance in clinical medicine. IMPORTANCE This study identified a novel phenotype of macrolides/lincosamides resistance in Staphylococcus haemolyticus which improved a better guidance for clinical treatment. It also clarified the mechanistic basis for this form of antibiotic resistance that supplemented the drug resistance mechanism of Staphylococcus. In addition, this study elaborated on a possibility that continuous expression of some resistance genes was shown to inhibit the growth of bacteria themselves, which turned out to be the fitness cost in the absence of antibiotic pressure.

Molecular Evolutionary Mechanisms of CYP6ER1vA-Type Variant Associated with Resistance to Neonicotinoid Insecticides in Field Populations of Nilaparvata lugens.

The evolution of insecticide resistance has threatened the control of Nilaparvata lugens. Research on mechanisms behind neonicotinoid resistance in N. lugens remains incomplete. This study examined P450-mediated resistance to neonicotinoids in a resistant N. lugens strain (XA-2017-3G). The overexpression of CYP6ER1 in the XA-2017-3G strain plays a role in neonicotinoid resistance, as confirmed by RNA interference. Phenotypic analyses of CYP6ER1-mediated resistance in strains, including laboratory-susceptible, field-collected, and imidacloprid-laboratory further-selected strains, revealed that the vA-type/vL-type genotype exhibited greater resistance to neonicotinoids compared to the vA-type/vA-type genotype. The mRNA expression levels of CYP6ER1vA-type were closely correlated with the levels of neonicotinoid resistance in N. lugens strains, in which CYP6ER1vA-type overexpression is in part attributed to increased copy numbers of CYP6ER1. CYP6ER1vA-type-mediated neonicotinoid resistance was further confirmed by a CYP6ER1vA-type transgenic Drosophila melanogaster line. Taken together, our findings strongly suggest that the overexpression of CYP6ER1vA-type, which can be partially attributed to copy number variations, plays a crucial role in N. lugens resistance to neonicotinoids.

Bittersweet Sugars: How Unusual Glycan Structures May Connect Epithelial-to-Mesenchymal Transition and Multidrug Resistance in Cancer.

Cancer cells are characterized by metabolic reprogramming, which enables their survival in of-ten inhospitable conditions. A very well-documented example that has gained attraction in re-cent years and is already considered a hallmark of transformed cells is the reprogramming of carbohydrate metabolism. Such a feature, in association with the differential expression of en-zymes involved in the biosynthesis of glycoconjugates, generically known as glycosyltransfer-ases, contributes to the expression of structurally atypical glycans when compared to those ex-pressed in healthy tissues. The latest studies have demonstrated that glycophenotypic alterations are capable of modulating multifactorial events essential for the development and/or progres-sion of the disease. Herein, we will address the importance of glycobiology in modern medi-cine, focusing on the ability of unusual/truncated O-linked glycans to modulate two complex and essential phenomena for cancer progression: the acquisition of the multidrug resistance (MDR) phenotype and the activation of molecular pathways associated with the epithelial-mesenchymal transition (EMT) process, an event deeply linked with cancer metastasis.

Increased expression of the pathological O-glycosylated form of oncofetal fibronectin in the multidrug resistance phenotype of cancer cells.

Changes in protein glycosylation are a hallmark of transformed cells and modulate numerous phenomena associated with cancer progression, such as the acquisition of multidrug resistance (MDR) phenotype. Different families of glycosyltransferases and their products have already been described as possible modulators of the MDR phenotype. Among the glycosyltransferases intensively studied in cancer research, UDP-N-acetyl-d-galactosamine:polypeptide N-acetylgalactosaminyltransferase-6 (pp-GalNAc-T6), which is widely expressed in many organs and tissues, stands out. Its influence in several events associated with kidney, oral, pancreatic, renal, lung, gastric and breast cancer progression has already been described. However, its participation in the MDR phenotype has never been studied. Here, we demonstrate that human breast adenocarcinoma MCF-7 MDR cell lines, generated by chronic exposure to doxorubicin, in addition to exhibiting increased expression of proteins belonging to the ABC superfamily (ABCC1 and ABCG2), and anti-apoptotic proteins (Blcl-2 and Bcl-xL), also present high expression of pp-GalNAc-T6, the enzyme currently proposed as the main responsible for the biosynthesis of oncofetal fibronectin (onf-FN), a major extracellular matrix component expressed by cancer cells and embryonic tissues, but absent in healthy cells. Our results show that onf-FN, which is generated by the addition of a GalNAc unit at a specific threonine residue inside the type III homology connective segment (IIICS) domain of FN, is strongly upregulated during the acquisition of the MDR phenotype. Also, the silencing of pp-GalNAc-T6, not only compromises the expression of the oncofetal glycoprotein, but also made the MDR cells more sensitive to all anticancer drugs tested, partially reversing the MDR phenotype. Taken together, our results demonstrate for the first time the upregulation of the O-glycosylated oncofetal fibronectin, as well as the direct participation of pp-GalNAc-T6 during the acquisition of a MDR phenotype in a breast cancer model, giving credence to the hypothesis that in transformed cells, glycosyltransferases and/or their products, such as unusual extracellular matrix glycoproteins can be used as potential therapeutic targets for the treatment of cancer.

One Health Surveillance of Antimicrobial Resistance Phenotypes in Selected Communities in Thailand.

Integrated surveillance of antimicrobial resistance (AMR) using the One Health approach that includes humans, animals, food, and the environment has been recommended by responsible international organizations. The objective of this study was to determine the prevalence of AMR phenotypes in Escherichia coli and Klebsiella species isolated from humans, pigs, chickens, and wild rodents in five communities in northern Thailand. Rectal swabs from 269 pigs and 318 chickens; intestinal contents of 196 wild rodents; and stool samples from 69 pig farmers, 155 chicken farmers, and 61 non-farmers were cultured for E. coli and Klebsiella species, which were then tested for resistance to ceftriaxone, colistin, and meropenem. The prevalence of ceftriaxone-resistant E. coli and Klebsiella species in pigs, chickens, rodents, pig farmers, chicken farmers, and non-farmers was 64.3%, 12.9%, 4.1%, 55.1%, 38.7%, and 36.1%, respectively. Colistin resistance in pigs, chickens, rodents, pig farmers, chicken farmers, and non-farmers was 41.3%, 9.8%, 4.6%, 34.8%, 31.6%, and 24.6%, respectively. Meropenem resistance was not detected. The observed high prevalence of AMR, especially colistin resistance, in study food animals/humans is worrisome. Further studies to identify factors that contribute to AMR, strengthened reinforcement of existing regulations on antimicrobial use, and more appropriate interventions to minimize AMR in communities are urgently needed.

The Relationship between Ciprofloxacin Resistance and Genotypic Changes in S. aureus Ocular Isolates.

Staphylococcus aureus (S. aureus) is a frequent cause of eye infections with some isolates exhibiting increased antimicrobial resistance to commonly prescribed antibiotics. The increasing resistance of ocular S. aureus to ciprofloxacin is a serious concern as it is a commonly used as a first line antibiotic to treat S. aureus keratitis. This study aimed to analyse genetic mutations in the genomes of 25 S. aureus isolates from infections or non-infectious ocular conditions from the USA and Australia and their relationship to ciprofloxacin resistance. Overall, 14/25 isolates were phenotypically resistant to ciprofloxacin. All isolates were analyzed for mutations in their quinolone resistance-determining regions (QRDRs) and efflux pump genes. Of the fourteen resistant isolates, 9/14 had ciprofloxacin resistance mutations within their QRDRs, at codons 80 or 84 within the parC subunit and codon 84 within the gyrA subunit of DNA gyrase. The highest resistance (MIC = 2560 µg/mL) was associated with two SNPs in both gyrA and parC. Other resistant isolates (3/14) had mutations within norB. Mutations in genes of other efflux pumps and their regulator (norA, norC, mepA, mdeA, sepA, sdrM, mepR, arlR, and arlS) or the DNA mismatch repair (MMR) system (mutL and mutS) were not associated with increased resistance to ciprofloxacin. The functional mutations associated with ciprofloxacin resistance in QRDRs (gyrA and parC) and norB suggests that these are the most common reasons for ciprofloxacin resistance in ocular isolates. Novel SNPs of gyrA Glu-88-Leu, Asn-860-Thr and Thr-845-Ala and IIe-855-Met, identified in this study, need further gene knock out/in studies to better understand their effect on ciprofloxacin resistance.

Beyond Circulating Tumor Cell Enumeration: Cell-Based Liquid Biopsy to Assess Protein Biomarkers and Cancer Genomics Using the RareCyte® Platform.

The practice of medicine has steadily employed less invasive methods to obtain information derived from the tumor to guide clinical management of patients. Liquid biopsy-the sampling of blood-is a non-invasive method for generating information previously only available from tissue biopsies of the tumor mass. Analysis of fragmented circulating tumor DNA in the plasma is clinically used to identify actionable mutations and detect residual or recurrent disease. Plasma analysis cannot, however, assess cancer phenotypes, including the expression of drug targets and protein biomarkers. Circulating tumor cells (CTCs) are intact cancer cells that have entered the blood that have the potential for distant metastasis. While enumeration of CTCs is prognostic of outcome, recently developed technology allows for the interrogation of protein biomarkers on CTCs that could be predictive of response. Furthermore, since CTCs contain intact whole cancer genomes, isolating viable CTCs detected during therapy could provide a rational approach to assessing mutational profiles of resistance. Identification, characterization and molecular analysis of CTCs together will advance the capacity of liquid biopsy to meet the requirements of twenty-first century medicine.

Sexual maturity, lack of partner choice and sperm precedence in the promiscuous ladybird beetle Eriopis connexa (Germar): Who is my father?

Ladybird beetles present as key predator of many agricultural pests. Among them, Eriopis connexa stands out due some important traits for pest management programs such as common occurrence, population selected for resistance to pyrethroid insecticides, and possibility to be commercially available by rearing using alternative prey. Despite that, little is known about the role of polyandry and its effect on paternity in this species. Ladybird beetles engage in multiple matings, raising questions about the benefits of polyandry and paternity. We studied the selection of mating pair, sperm precedence, and offspring paternity by performing experiments on (1) the age of sexual maturity, (2) the frequency and refractory mating behavior within a photophase period, (3) the preference of insecticide-susceptible females to mate with either susceptible or resistant male phenotypes; and (4) the effect of multiple matings, on progeny paternity. Sexual maturity in E. connexa was found to begin within 3 days of adult emergence for both sexes, but fully developed after 5 days. The highest frequency of mating exhibited by sexual mature pairs occurred within first hour of pairing and the phenotype for insecticide resistance did not affect the choice of a mating partner. The resistance trait marker in the resistant beetle phenotype indicates that progeny paternity results from a mixture of available sperm and do not depend on mating order. These data suggest that released beetles would have a simple prospect for progeny production for both insecticide resistant and susceptible beetles.