Polyketide Synthase-Mediated O-Methyloxime Formation in the Biosynthesis of the Oximidine Anticancer Agents.

Bacterial trans-acyltransferase polyketide synthases (trans-AT PKSs) are modular megaenzymes that employ unusual catalytic domains to assemble diverse bioactive natural products. One such PKS is responsible for the biosynthesis of the oximidine anticancer agents, oxime-substituted benzolactone enamides that inhibit vacuolar H+ -ATPases. Here, we describe the identification of the oximidine gene cluster in Pseudomonas baetica and the characterization of four novel oximidine variants, including a structurally simpler intermediate that retains potent anticancer activity. Using a combination of in vivo, in vitro and computational approaches, we experimentally elucidate the oximidine biosynthetic pathway and reveal an unprecedented mechanism for O-methyloxime formation. We show that this process involves a specialized monooxygenase and methyltransferase domain and provide insight into their activity, mechanism and specificity. Our findings expand the catalytic capabilities of trans-AT PKSs and identify potential strategies for the production of novel oximidine analogues.

Programmed Iteration Controls the Assembly of the Nonanoic Acid Side Chain of the Antibiotic Mupirocin.

Mupirocin is a clinically important antibiotic produced by Pseudomonas fluorescens NCIMB 10586 that is assembled by a complex trans-AT polyketide synthase. The polyketide fragment, monic acid, is esterified by a 9-hydroxynonanoic acid (9HN) side chain which is essential for biological activity. The ester side chain assembly is initialised from a 3-hydroxypropionate (3HP) starter unit attached to the acyl carrier protein (ACP) MacpD, but the fate of this species is unknown. Herein we report the application of NMR spectroscopy, mass spectrometry, chemical probes and in vitro assays to establish the remaining steps of 9HN biosynthesis. These investigations reveal a complex interplay between a novel iterative or "stuttering" KS-AT didomain (MmpF), the multidomain module MmpB and multiple ACPs. This work has important implications for understanding the late-stage biosynthetic steps of mupirocin and will be important for future engineering of related trans-AT biosynthetic pathways (e.g. thiomarinol).

The Kalimantacin Polyketide Antibiotics Inhibit Fatty Acid Biosynthesis in Staphylococcus aureus by Targeting the Enoyl-Acyl Carrier Protein Binding Site of FabI.

The enoyl-acyl carrier protein reductase enzyme FabI is essential for fatty acid biosynthesis in Staphylococcus aureus and represents a promising target for the development of novel, urgently needed anti-staphylococcal agents. Here, we elucidate the mode of action of the kalimantacin antibiotics, a novel class of FabI inhibitors with clinically-relevant activity against multidrug-resistant S. aureus. By combining X-ray crystallography with molecular dynamics simulations, in vitro kinetic studies and chemical derivatization experiments, we characterize the interaction between the antibiotics and their target, and we demonstrate that the kalimantacins bind in a unique conformation that differs significantly from the binding mode of other known FabI inhibitors. We also investigate mechanisms of acquired resistance in S. aureus and identify key residues in FabI that stabilize the binding of the antibiotics. Our findings provide intriguing insights into the mode of action of a novel class of FabI inhibitors that will inspire future anti-staphylococcal drug development.

Control of ß-Branching in Kalimantacin Biosynthesis: Application of 13 C†NMR to Polyketide Programming.

The presence of ß-branches in the structure of polyketides that possess potent biological activity underpins the widespread importance of this structural feature. Kalimantacin is a polyketide antibiotic with selective activity against staphylococci, and its biosynthesis involves the unprecedented incorporation of three different and sequential ß-branching modifications. We use purified single and multi-domain enzyme components of the kalimantacin biosynthetic machinery to address in vitro how the pattern of ß-branching in kalimantacin is controlled. Robust discrimination of enzyme products required the development of a generalisable assay that takes advantage of 13 C NMR of a single 13 C label incorporated into key biosynthetic mimics combined with favourable dynamic properties of an acyl carrier protein. We report a previously unassigned modular enoyl-CoA hydratase (mECH) domain and the assembly of enzyme constructs and cascades that are able to generate each specific ß-branch.

A Decade in the MIST: Learnings from Investigations of Drug Metabolites in Drug Development under the "Metabolites in Safety Testing" Regulatory Guidance.

Since the introduction of metabolites in safety testing (MIST) guidance by the Food and Drug Administration in 2008, major changes have occurred in the experimental methods for the identification and quantification of metabolites, ways to evaluate coverage of metabolites, and the timing of critical clinical and nonclinical studies to generate this information. In this cross-industry review, we discuss how the increased focus on human drug metabolites and their potential contribution to safety and drug-drug interactions has influenced the approaches taken by industry for the identification and quantitation of human drug metabolites. Before the MIST guidance was issued, the method of choice for generating comprehensive metabolite profile was radio chromatography. The MIST guidance increased the focus on human drug metabolites and their potential contribution to safety and drug-drug interactions and led to changes in the practices of drug metabolism scientists. In addition, the guidance suggested that human metabolism studies should also be accelerated, which has led to more frequent determination of human metabolite profiles from multiple ascending-dose clinical studies. Generating a comprehensive and quantitative profile of human metabolites has become a more urgent task. Together with technological advances, these events have led to a general shift of focus toward earlier human metabolism studies using high-resolution mass spectrometry and to a reduction in animal radiolabel absorption/distribution/metabolism/excretion studies. The changes induced by the MIST guidance are highlighted by six case studies included herein, reflecting different stages of implementation of the MIST guidance within the pharmaceutical industry.

Growth Recovery of Lemna gibba and Lemna minor Following a 7-Day Exposure to the Herbicide Diuron.

In agricultural catchments, aquatic ecosystems can experience a pulse exposure to pesticides. Following such exposure, non-target organisms that are not extirpated may recover. This paper investigates the potential of two duckweed species (Lemna minor and Lemna gibba) to recover from a 7-day exposure to different concentrations (0.4-208 µg L(-1)) of the herbicide diuron. There was significant inhibition in the growth and biomass after the initial 7-day exposure (e.g. frond number EC50=59.2 and 52.2 µg L(-1) for L. minor and L. gibba, respectively). Following transfer to clean media, recovery (the highest concentration yielding no significant difference in the effect endpoint from the control) was observed for all effects endpoints at concentrations ranging 60-111 µg L(-1) for L. minor and 60-208 µg L(-1) for L. gibba. These results suggest that recovery is possible for primary producers at environmentally relevant concentrations considered significant in ecological risk assessment.

Identifying D-positive donors using a second automated testing platform.

Because of the variability of D expression, one method may be inadequate to correctly classify donors with variant RHD alleles. We evaluated the use of a solid -phase automated platform (ImmucorGamma Galileo) to confirm D- test results obtained on first-time donors on the Beckman Coulter PK7300 automated microplate test system. Samples with discordant results were analyzed by serologic tube methods, RHD genotyping using the BLOODchip platform (Progenika) and, if necessary, sequencing. We estimated the number of cases of alloimmunization in women younger than 50 years likelyto be prevented by the addition of Galileo testing. From May 2011 to May 2012, 910,220 donor samples were tested; 15,441 were first-time donors with concordant D- results. Five donors tested D- on the PK7300 and weak D+ on the Galileo; one was found to be a false positive on further testing. On manual testing, the other four donors had positive indirect antiglobulin test results with one to three of the antisera used and were C+. On BLOODchip testing, two donors were classified as D+, and two were assigned a "no call". D variants included weak D type 67, weak D type 9, and two novel variants. Approximately 10 percent of D- units are transfused to women younger that 50 years. Assuming an alloimmunization rate of 30 percent, use of the Galileo would prevent approximately one alloimmunization every 5 to 6 years in this patient group. We conclude that the yield of preventing alloimmunization in this population by adding a second automated seologic testing platform is very low.

Capitalizing on deliberate, accidental, and GM-driven environmental change caused by crop modification.

The transgenic traits associated with the majority of commercial genetically modified crops are focused on improving herbicide and insecticide management practices. The use of the transgenic technology in these crops and the associated chemistry has been the basis of studies that provide evidence for occasional improvement in environmental benefits due to the use of less residual herbicides, more targeted pesticides, and reduced field traffic. This is nicely exemplified through studies using Environmental Impact Quotient (EIQ) assessments. Whilst EIQ evaluations may sometimes illustrate environmental benefits they have their limitations. EIQ evaluations are not a surrogate for Environmental Risk Assessments and may not reflect real environmental interactions between crops and the environment. Addressing the impact cultivated plants have on the environment generally attracts little public attention and research funding, but the introduction of GM has facilitated an expansion of research to address potential environmental concerns from government, NGOs, industry, consumers, and growers. In this commentary, some evidence from our own research and several key papers that highlight EIQ assessments of the impact crops are having on the environment are presented. This information may be useful as an education tool on the potential benefits of GM and conventional farming. In addition, other deliberate, accidental, and GM-driven benefits derived from the examination of GM cropping systems is briefly discussed.

Radiolabelled mass-balance excretion and metabolism studies in laboratory animals: are they still necessary?

The conduct of excretion and metabolism studies using radiolabelled drugs in multiple laboratory animal species has been a mainstay of the suite of support activities provided by drug metabolism groups within pharmaceutical research and development organizations for decades. Drug metabolism scientists carry out exhaustive analyses of plasma and excretory matrices to comprehensively determine the profiles of metabolites in these species. While these analyses have taught us considerably regarding principles of drug metabolism and excretion, it is our contention that the routine conduct of such studies for every new drug development compound in every laboratory animal species used in toxicology studies is no longer necessary. The recently released regulatory guidance regarding metabolites and safety testing have better defined what we need to know regarding metabolite profiles in humans relative to animals. In this commentary, we propose a strategy wherein a radiolabel metabolism study is conducted only in humans, and that these data are utilized as a springboard to direct the exploration of steady-state human versus animal metabolite exposures. Such a strategy better serves the purpose of what is needed to support our understanding of the safety of a new drug candidate. Valuable expertise in drug metabolism and biotransformation can be redeployed to meet the burgeoning needs in drug design efforts to optimize structures with regard to metabolic clearance properties, understanding pharmacologically active metabolites, and reducing generation of chemically reactive metabolites.

Classic and Current Opinions in Human Organ and Tissue Transplantation.

Graft tolerance is a pathophysiological condition heavily reliant on the dynamic interaction of the innate and adaptive immune systems. Genetic polymorphism determines immune responses to tissue/organ transplantation, and intricate humoral and cell-mediated mechanisms control these responses. In transplantation, the clinician's goal is to achieve a delicate equilibrium between the allogeneic immune response, undesired effects of the immunosuppressive drugs, and the existing morbidities that are potentially life-threatening. Transplant immunopathology involves sensitization, effector, and apoptosis phases which recruit and engages immunological cells like natural killer cells, lymphocytes, neutrophils, and monocytes. Similarly, these cells are involved in the transfer of normal or genetically engineered T cells. Advances in tissue transplantation would involve a profound knowledge of the molecular mechanisms that underpin the respective immunopathology involved and the design of precision medicines that are safe and effective.

Programmed Iteration Controls the Assembly of the Nonanoic Acid Side Chain of the Antibiotic Mupirocin.

Mupirocin is a clinically important antibiotic produced by Pseudomonas fluorescens NCIMB 10586 that is assembled by a complex trans-AT polyketide synthase. The polyketide fragment, monic acid, is esterified by a 9-hydroxynonanoic acid (9HN) side chain which is essential for biological activity. The ester side chain assembly is initialised from a 3-hydroxypropionate (3HP) starter unit attached to the acyl carrier protein (ACP) MacpD, but the fate of this species is unknown. Herein we report the application of NMR spectroscopy, mass spectrometry, chemical probes and in vitro assays to establish the remaining steps of 9HN biosynthesis. These investigations reveal a complex interplay between a novel iterative or "stuttering" KS-AT didomain (MmpF), the multidomain module MmpB and multiple ACPs. This work has important implications for understanding the late-stage biosynthetic steps of mupirocin and will be important for future engineering of related trans-AT biosynthetic pathways (e.g. thiomarinol).

From definition to implementation: a cross-industry perspective of past, current and future MIST strategies.

The article describes and discusses the evolution of strategies to characterize metabolites in support of safety studies over the last 40 years, as well as future trends. Approaches to derive qualitative and quantitative information on metabolites are described, with a particular focus on the comparison of options to quantify metabolites in the absence of authentic standards. Current strategies to assess metabolite profiles are summarized into four general approaches and compared against a number of key criteria. Potential future strategies are discussed, including the use of clinical samples as the starting point for metabolite investigations, minimizing the need for animal radiolabelled studies and establishing metabolite safety without radiolabelled studies in animals or human.

Excretion and metabolism of lersivirine (5-{[3,5-diethyl-1-(2-hydroxyethyl)(3,5-14C2)-1H-pyrazol-4-yl]oxy}benzene-1,3-dicarbonitrile), a next-generation non-nucleoside reverse transcriptase inhibitor, after administration of [14C]Lersivirine to healthy volunteers.

Lersivirine [UK-453,061, 5-((3,5-diethyl-1-(2-hydroxyethyl)(3,5-14C2)-1H-pyrazol-4-yl)oxy)benzene-1,3-dicarbonitrile] is a next-generation non-nucleoside reverse transcriptase inhibitor, with a unique binding interaction within the reverse transcriptase binding pocket. Lersivirine has shown antiviral activity and is well tolerated in HIV-infected and healthy subjects. This open-label, Phase I study investigated the absorption, metabolism, and excretion of a single oral 500-mg dose of [14C]lersivirine (parent drug) and characterized the plasma, fecal, and urinary radioactivity of lersivirine and its metabolites in four healthy male volunteers. Plasma C(max) for total radioactivity and unchanged lersivirine typically occurred between 0.5 and 3 h postdose. The majority of radioactivity was excreted in urine (approximately 80%) with the remainder excreted in the feces (approximately 20%). The blood/plasma ratio of total drug-derived radioactivity [area under the plasma concentration-time profile from time zero extrapolated to infinite time (AUC(inf))] was 0.48, indicating that radioactive material was distributed predominantly into plasma. Lersivirine was extensively metabolized, primarily by UDP glucuronosyltransferase- and cytochrome P450-dependent pathways, with 22 metabolites being identified in this study. Analysis of precipitated plasma revealed that the lersivirine-glucuronide conjugate was the major circulating component (45% of total radioactivity), whereas unchanged lersivirine represented 13% of total plasma radioactivity. In vitro studies showed that UGT2B7 and CYP3A4 are responsible for the majority of lersivirine metabolism in humans.

Total Synthesis of Kalimantacin A.

The kalimantacins make up a family of hybrid polyketide-nonribosomal peptide-derived natural products that display potent and selective antibiotic activity against multidrug resistant strains of Staphylococcus aureus. Herein, we report the first total synthesis of kalimantacin A, in which three fragments are prepared and then united via Sonogashira and amide couplings. The enantioselective synthetic approach is convergent, unlocking routes to further kalimantacins and analogues for structure-activity relationship studies and clinical evaluation.

A Priming Cassette Generates Hydroxylated Acyl Starter Units in Mupirocin and Thiomarinol Biosynthesis.

Mupirocin, a commercially available antibiotic produced by Pseudomonas fluorescens NCIMB 10586, and thiomarinol, isolated from the marine bacterium Pseudoalteromonas sp. SANK 73390, both consist of a polyketide-derived monic acid homologue esterified with either 9-hydroxynonanoic acid (mupirocin, 9HN) or 8-hydroxyoctanoic acid (thiomarinol, 8HO). The mechanisms of formation of these deceptively simple 9HN and 8HO fatty acid moieties in mup and tml, respectively, remain unresolved. To define starter unit generation, the purified mupirocin proteins MupQ, MupS, and MacpD and their thiomarinol equivalents (TmlQ, TmlS and TacpD) have been expressed and shown to convert malonyl coenzyme A (CoA) and succinyl CoA to 3-hydroxypropionoyl (3-HP) or 4-hydroxybutyryl (4-HB) fatty acid starter units, respectively, via the MupQ/TmlQ catalyzed generation of an unusual bis-CoA/acyl carrier protein (ACP) thioester, followed by MupS/TmlS catalyzed reduction. Mix and match experiments show MupQ/TmlQ to be highly selective for the correct CoA. MacpD/TacpD were interchangeable but alternate trans-acting ACPs from the mupirocin pathway (MacpA/TacpA) or a heterologous ACP (BatA) were nonfunctional. MupS and TmlS selectivity was more varied, and these reductases differed in their substrate and ACP selectivity. The solution structure of MacpD determined by NMR revealed a C-terminal extension with partial helical character that has been shown to be important for maintaining high titers of mupirocin. We generated a truncated MacpD construct, MacpD_T, which lacks this C-terminal extension but retains an ability to generate 3-HP with MupS and MupQ, suggesting further downstream roles in protein-protein interactions for this region of the ACP.

Metabolites in safety testing: metabolite identification strategies in discovery and development.

The publication of the FDA MIST guidelines in 2008, together with the acknowledged importance of metabolism data for the progression of novel compounds through drug discovery and drug development, has resulted in a renewed focus on the metabolite identification strategies utilised throughout the pharmaceutical industry. With the plethora of existing and emerging technologies available to the metabolite identification scientist, it is argued that increased diligence should be applied to metabolism studies in the early stages of both drug discovery and drug development, in order to more routinely impact chemical design and to comply with the concepts of the MIST guidance without re-positioning the definitive radiolabelled studies from there typical place in late development.Furthermore, these strategic elements should be augmented by a broad and thorough understanding of the impact of the derived metabolism data, most notably considerations of absolute abundance, structure and pharmacological activity, such that they can be put into proper context as part of a holistic safety strategy.The combination of these approaches should ensure a metabolite identification strategy that successfully applies the principles of the MIST guidance throughout the discovery/development continuum and thereby provides appropriate confidence in support of human safety.

Multi-Antibiotic Resistance and Factors Affecting Carriage of Extended Spectrum ß-Lactamase-Producing Enterobacteriaceae in Pediatric Population of Enugu Metropolis, Nigeria.

Extended-spectrum ß-lactamase (ESBL)-producing organisms have become a serious challenge in healthcare delivery globally. The prevalence of ESBL carriage in healthy and sick children in Enugu, Nigeria, was bacteriologically investigated in this study. Four hundred and twenty-two biological samples (mid-stream urine and feces) were bacteriologically analyzed. The isolates were screened for ESBL production using Clinical and Laboratory Standards Institute (CLSI) breakpoints. The suspected ESBL producers were confirmed using double disc synergy test method. Out of the 162 isolates screened, 32 (19.8%) were confirmed as ESBL positive, with a prevalence of 25.32% among sick children in Enugu State University Teaching Hospital (ESUTH), Parklane, Enugu and 13.89% in apparently healthy children in a community setting. Klebsiella spp. and Escherichia coli had the highest prevalence of 34.6% and 28.6%, respectively; Citrobacter spp. and Enterobacter spp. were 18.2% and 16.7%, respectively. The ESBL positive isolates were resistant to sulfamethoxazole/trimethoprim (100%), tetracycline (100%), kanamycin (96.9%), nitrofurantoin (84.4%), ciprofloxacin (68.6%), and chloramphenicol (62.5%) but susceptible to meropenem (100%), colistin (56.3%), and gentamicin (50%). Klebsiella spp. had the highest ESBL occurrence among sick children while E. coli had the highest ESBL occurrence among healthy children in Enugu. All ESBL-positive isolates were multiply resistant to conventional antibiotics. The emergence and spread of ß-lactamase-producing Enterobacteriaceae in hospital and community environments highlight the possibility for an infection outbreak if not checked.

Assessment of the absorption, metabolism and absolute bioavailability of maraviroc in healthy male subjects.

AIMS: Two studies were conducted to: (i) quantify the amount of drug-related radioactivity in blood, plasma, urine and faeces following a (14)C-labelled dose of maraviroc; and (ii) investigate the pharmacokinetics, safety and tolerability of intravenous (i.v.) maraviroc and determine the absolute bioavailability of oral maraviroc. Metabolite profiling was also conducted. Data from both of these studies were used to construct a mass-balance model for maraviroc. METHODS: Study 1 was an open-label study in three healthy male subjects. All subjects received a single 300-mg oral solution dose of (14)C-labelled maraviroc. Study 2 included two cohorts of subjects. Cohort 1 involved a double-blind (third party open), four-way crossover study where eight subjects received escalating i.v. doses of maraviroc (3, 10 and 30 mg) with placebo insertion. Cohort 2 involved an open, two-way crossover study where 12 subjects received 30 mg maraviroc by i.v. infusion and 100 mg maraviroc orally in random order. In study 1, blood samples and all urine and faeces were collected up to at least 120 h postdose. In study 2, blood samples were taken at intervals up to 48 h postdose. Urine was also collected up to 24 h postdose in cohort 1 only. RESULTS: After oral administration in study 1, maraviroc was rapidly absorbed with a plasma T(max) reached by 2 h postdose for all three subjects. The maximum concentrations of radioactivity also occurred within 2 h for all subjects. There was a higher amount of radioactivity in plasma than in blood (blood/plasma ratio approximately 0.6 for AUC(t) and C(max)). Unchanged maraviroc was the major circulating component in plasma, accounting for approximately 42% of the circulating radioactivity. Following a 300-mg (14)C-labelled maraviroc dose, means of 76.4% and 19.6% of radioactivity were recovered in the faeces and urine, respectively. The mean total recovery of dosed radioactivity was 96%, with the majority of radioactivity being recovered within 96 h postdose. Profiling of the urine and faeces showed similar and extensive metabolism in all subjects. Unchanged maraviroc was the major excreted component (33%). The major metabolic pathways were determined and involved oxidation and N-dealkylation. Intravenous doses of maraviroc (3-30 mg) were well tolerated in study 2, and drug exposure was approximately proportional to dose within the studied range. Approximately 23% of total clearance (44 l h(-1)) was accounted for by renal clearance (10.2 l h(-1)). Mean volume of distribution at steady state was 194 l. Absolute bioavailability of a 100-mg oral tablet dose, by comparison with a 30-mg i.v. dose, was calculated to be 23.1%. CONCLUSIONS: Maraviroc is rapidly absorbed and extensively metabolized, although unchanged maraviroc is the major circulating component in plasma and is the major excreted component after oral dosing. The pharmacokinetics of maraviroc after i.v. administration is approximately proportional over the dose range studied. Renal clearance contributes 23% of total clearance. The absolute bioavailability of 100 mg oral maraviroc is 23%.

The effect of vegetation on pesticide dissipation from ponded treatment wetlands: quantification using a simple model.

Field data shows that plants accelerate pesticide dissipation from aquatic systems by increasing sedimentation, biofilm contact and photolysis. In this study, a graphical model was constructed and calibrated with site-specific and supplementary data to describe the loss of two pesticides, endosulfan and fluometuron, from a vegetated and a non-vegetated pond. In the model, the major processes responsible for endosulfan dissipation were alkaline hydrolysis and sedimentation, with the former process being reduced by vegetation and the latter enhanced. Fluometuron dissipation resulted primarily from biofilm reaction and photolysis, both of which were increased by vegetation. Here, greater photolysis under vegetation arose from faster sedimentation and increased light penetration, despite shading. Management options for employing constructed wetlands to polish pesticide-contaminated agricultural runoff are discussed. The lack of easily fulfilled sub-models and data describing the effect of aquatic vegetation on water chemistry and sedimentation is also highlighted.

Bacteriology and Antibiogram of Urinary Tract Infection Among Female Patients in a Tertiary Health Facility in South Eastern Nigeria.

BACKGROUND: Urinary Tract Infection (UTI) is a common contagion among men and women with the incidence relatively higher among women due to their differing anatomy. An understanding of the kind of pathogens implicated in urinary tract infections as well as antibiotic susceptibility profiling may help the clinician make rationally correct empirical choice in their treatment. OBJECTIVE: This study is aimed at determining the type and antibiotic susceptibility pattern of bacterial uropathogens isolated from female patients attending Chukwuemeka Odumegwu Ojukwu University Teaching Hospital (COOUTH), Awka, Nigeria. METHOD: Two hundred and forty patients with clinically diagnosed UTI and who were on at least 5 days' antibiotic holiday were recruited into the study. Their demographic characteristics were captured using pre-tested questionnaire. Their clean catch mid-stream urine samples were collected using sterile universal container and sent to the Microbiology Department for processing. Within 30 minutes of samples collection, the specimens were cultured and the isolates were identified, after 24 h of incubation, using standard microbiological techniques. Antibiotic susceptibility tests were done with standard antibiotic discs using the Kirby-bauer disc diffusion method. RESULTS: Out of the 240 urine samples, 89.17% yielded significant bacteriuria. The pathogens implicated were Escherichia coli (28.5%), Staphylococcus aureus (28.0%), Salmonella spp (22.8%) and Pseudomonas aeruginosa (20.5%). HIV status, patients age, pregnancy status and marital status all significantly affected bacteriuria rate (p value < 0.05), while patients' location (sub-urban/rural dwelling), and level of education did not (p value > 0.05). The pattern of microbial resistance to antibiotics suggests that ceftazidime, fosfomycin and cefoxitin may not be used as first-line agents in the empirical treatment of UTIs rather; levofloxacin, meropenem or aztreonam should be considered. Levofloxacin was significantly effective against all the isolates and may be administered empirically while waiting for the culture result (Mean % susceptibility was 79.85). CONCLUSION: E. coli and S. aureus were the predominant pathogens in the study and many were resistant to the commonly prescribed antibiotics and so leave the clinicians with only few alternative drugs for UTIs treatment. Routine surveillance and monitoring studies need to be constantly conducted to update clinicians on the prevalent pathogens and the rational and empirical treatment of UTIs. Aggressive and consistent health education using every possible media is also recommended to combat the menace of drug resistance occasioned by inappropriate antibiotic use.