Spread Of Drug Resistance Research Articles

Antimalarial drug resistance profiling of Plasmodium falciparum infections in India using Ion Torrent deep sequencing

IntroductionTracking the emergence and spread of antimalarial drug resistance is critical for supporting progress towards the control and eventual elimination of malaria in South Asia, especially India. Plasmodium falciparum has evolved resistance to virtually every antimalarial drug, and significant progress has been made to identify the molecular genetic mechanisms involved in the most common types of resistance.MethodsAn amplicon sequencing protocol was used for molecular surveillance of antimalarial drug resistance in a total of 158 patient isolates collected from December 2012 to September 2015 from three sites in south, west and east India: Tamil Nadu, Gujarat, and Odisha respectively. Five full length Plasmodium falciparum genes whose mutant proteins are implicated in antimalarial drug resistance were investigated: Pfcrt for chloroquine, Pfdhfr for pyrimethamine, Pfdhps for sulfadoxine, Pfk13 for artemisinin and Pfmdr1 for resistance to multiple antimalarial drugs.ResultsWe observed a high proportion of wild-type Pfcrt and Pfdhfr haplotypes from the P. falciparum-dominant site Rourkela, while mutant Pfcrt and Pfdhfr haplotypes were fixed at the P. vivax dominant sites Chennai and Nadiad. The wild-type Pfdhps haplotype was predominant across all study sites. We identified mutations in the propeller domain of Pfk13, although they are not associated with resistance to artemisinin. Finally, using samples taken from the same patient on day 2, day 7, and day 14 after artemisinin combination treatment, we were able to observe changes in allele frequency of drug resistance genes during the course of an infection.DiscussionThis is the first high-throughput deep sequencing study of five full-length P. falciparum genes in clinical isolates from three different study sites in India with varying transmission ecologies. Amplicon sequencing based on ion-torrent has the potential to track changes in the frequency of drug resistant alleles as a patient is undergoing drug therapy, and to identify new resistance mutations as they increase in frequency in the patient. This study showed possibility of whole gene sequencing, leads to in-depth molecular surveillance of multiple antimalarial resistant candidates and furthermore suggests investigations on reversal of resistant genotypes due to usage of artemisinin combination therapy in P. falciparum prevalent areas of the country.

Genotyping USA laboratory-maintained isolates and European clinical isolates of Dirofilaria immitis to assess macrocyclic lactone susceptibility or resistance at predictive SNP sites using droplet digital PCR

Dirofilaria immitis is a parasitic nematode that causes cardiovascular dirofilariosis ("heartworm disease”) primarily in canids. The principal approach for mitigating heartworm infection involves the use of macrocyclic lactone (ML) for prophylaxis. Recent research has substantiated the emergence of D. immitis displaying resistance to MLs in the USA. Numerous factors, such as the mobility of companion animals and competent vectors could impact the spread of drug resistance. Genomic analysis has unveiled that isolates resistant to ML exhibit unique genetic profiles when compared to their wild-type (susceptible) counterparts. Out of the ten single nucleotide polymorphism (SNP) markers validated in clinical samples of D. immitis from the USA, four have demonstrated their effectiveness in distinguishing between isolates with varying ML efficacy phenotypes. This study explores the potential of these confirmed SNPs for conducting surveillance studies. Genotypic analysis using SNP markers emerges as a valuable tool for carrying out surveys and evaluating individual clinical isolates. Two USA laboratory-maintained isolates (Berkeley, WildCat) and twenty-five random European clinical samples of either adult worms or microfilariae (mf) pools isolated from domestic dogs, were tested by droplet digital PCR (ddPCR)-based duplex assay. This approach elucidates genetic evidence pertaining to the development of drug resistance and provides baseline data on resistance related genotypes in Europe. The data on these clinical samples suggests genotypes consistent with the continued efficacy of ML treatment regimens in Europe. In addition, this assay can be significant in discriminating cases of drug-resistance from those possibly due to non-compliance to the recommended preventive protocols.

Antimalarial Efficacy of Ethanol Extract of Bridelia micrantha Stem Bark against Plasmodium berghei-Infected Mice.

The spread of drug resistance is a significant issue, particularly in endemic countries with limited resources. The aim of this study was to evaluate antimalarial and antioxidant activity of B. micrantha in order to justify its use in traditional medicine. Evaluation of the in vivo antimalarial activity of B. micrantha was carried out according to the model of the suppressive and curative test of Peters' over 4 days in infected Swiss albino mice. Antioxidant parameters and stress were measured after intraperitoneal administration of 1 × 107 infected red blood cells. At doses of 150 mg/kg, 300 mg/kg, and 600 mg/kg, administration of B. micrantha substantially produced suppression of P. berghei infection by 67.75%, 73.46%, and 78.99%, respectively, while 84.64% of the untreated group (1% DMSO) had suppression from chloroquine. The curative test significantly decreased the levels of parasitaemia and death in the treated groups. Furthermore, after B. micrantha extract was given to infected mice, a noteworthy increase in total protein, aspartate aminotransferase (AST), and alanine aminotransferase (ALT) was observed. On the other hand, hepatic catalase (CAT) and superoxide dismutase (SOD) productions were considerably greater than that of the healthy control. Mice had considerably lower levels of nonenzymatic antioxidant markers such as glutathione, NO, and MDA showing that the liver was protected. The infected groups responded favorably to the ethanol extract of B. micrantha. This result justifies investigation for its use in Cameroon.

Abstract 2780: Induced pluripotent stem cell (iPSC) based vaccines potentializes anti-CTLA4 immune response in a murine lung carcinoma model

Abstract Despite the development of new immunotherapy strategies, lung cancer remains the leading cause of cancer-related deaths. One of the reasons is attributed to the persistence of cancer stem cells (CSCs) within the tumor, contributing to drug resistance, dormancy and metastatic spread. To eradicate CSCs, iPSC-based vaccination represents a novel approach based on the paradigm that CSCs and iPSCs share common tumor associated antigens (TAAs) with the ability to elicit a strong anti-tumor immune response.We previously showed that induced allogeneic pluripotent stem cells (miPSCs), combined with a histone deacetylase inhibitor (HDACi), induced a major reduction of tumor growth and metastatic spread to lungs in an aggressive triple negative breast cancer model (4T1) in syngeneic mice. We report here the use of the same protocol in a lung carcinoma model using LLC1 cell line in combination with an anti-CTLA4 antibody. We compared the effectiveness of allogenic miPSCs treatment alone or with an anti-CTLA4 mouse antibody on mice-bearing LLC1-Luc tumors (mean+se: 17,5+0,6 mm3). Five mice received 5 subcutaneous (SC) injection of 2 × 106 miPSCs, 5 mice received 3 intraperitoneal injections of anti-CTLA4 antibody at 5mg/kg and 5 mice were treated with the combinatory regimen (miPSCs and anti- CTLA4) with the same doses. In addition, all treated mice received a HDACi in the drink water throughout the experience.After 24 days, all tumor volumes from the 3 groups were significantly reduced compared to the control mice receiving only PBS. We obtained a reduction of 72% (p=0.007) and 74% (p=0.009) respectively for the mice treated solely with miPSCs or anti-CTLA4. An important synergic effect was obtained for the mice treated with miPSCs and anti- CTLA4 with a reduction of the tumor size by 92% (p=0.005) compared to the control mice. We also found a significant inhibition of metastatic spread to lungs for the 3 groups that was more pronounced in mice receiving the combination treatment with a significant reduction of 91 % compared to the control group. All these therapeutic effects were correlated with a significant increase of CD4+ and CD8+ cells that was more pronounced in mice receiving the combinatory regimen.We also have performed single-cell RNA sequencing analysis (10x Genomics) of LLC1 tumors treated with miPSCs and anti- CTLA4 compared to mice treated with monotherapy and untreated mice. T-distributed stochastic neighbor embedding plot reveals distinct clusters of stromal and immune cells across tumors with a significant upregulation of CD8+ and CD4+ T cells in mice receiving the combinatory regimen.Overall, these results demonstrate for the first time the clinical relevance of using an off-the-shelf allogeneic iPSC-based vaccine combined with an anti-CTLA4 antibody as a new anti-cancer immunotherapy strategy against aggressive non-small cell lung cancer (NSCLC) with metastatic potential. Citation Format: Frank Griscelli, Diana Chaker, Noufissa Oudrhiri, Christophe Desterke, Ali G. Turhan, Annelise Bennaceur Griscelli. Induced pluripotent stem cell (iPSC) based vaccines potentializes anti-CTLA4 immune response in a murine lung carcinoma model [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 2780.

Availability of drugs and resistance testing for bedaquiline, pretomanid, linezolid, and moxifloxacin (BPaL(M)) regimen for rifampicin-resistant tuberculosis in Europe

ObjectivesMultidrug-resistant/rifampicin-resistant tuberculosis is a major obstacle to successful tuberculosis control. The recommendation by the WHO to use bedaquiline, pretomanid, linezolid, and moxifloxacin (BPaL(M)) for 6 months, based on results of two trials with high efficacy and low toxicity, has revolutionized treatment options. MethodsIn this study, representatives of the Tuberculosis Network European Trials group in 44 of 54 countries of the WHO Europe region documented the availability of the medicines and drug susceptibility testing (DST) of the BPaL(M) regimen through a structured questionnaire between September and November 2023. ResultsIn total, 24 of 44 (54.5%), 42 of 44 (95.5%), 43 of 44 (97.7%), and 43 of 44 (97.7%) countries had access to pretomanid, bedaquiline, linezolid, and moxifloxacin, respectively. Overall, 23 of 44 (52.3%) countries had access to all the drugs composing the BPaL(M) regimen. In total, 21 of 44 (47.7%), 37 of 44 (84.1%), 40 of 44 (90.9%), and 41 of 44 (93.2%) countries had access to DST for pretomanid, bedaquiline, linezolid, and moxifloxacin, respectively. Overall, DST was available for all medicines composing the BPaL(M) regimen in 21 of 44 (47.7%) countries, including countries where pretomanid DST was available at specialized laboratories. The availability of DST for the drugs the countries had access to, varied from 87.5% to 95.3% (pretomanid 21 of 24 (87.5%), bedaquiline 37 of 42 (88.1%), linezolid 40 of 43 (93.1%) and moxifloxacin 41 of 43 (95.3%)). DiscussionIn only about half of the countries participating in the survey, clinicians had access to all the BPaL(M) regimen drugs. A complete DST for the BPaL(M) medicines was possible in less than half of the countries, because of the low accessibility of DST for pretomanid. Equal access to new regimens is urgently needed in Europe and a rapid scale up of DST, especially for pretomanid, is important to prevent unnoticed spread of drug resistance.

Chemoproteomics-based profiling reveals potential antimalarial mechanism of Celastrol by disrupting spermidine and protein synthesis

BackgroundMalaria remains a global health burden, and the emergence and increasing spread of drug resistance to current antimalarials poses a major challenge to malaria control. There is an urgent need to find new drugs or strategies to alleviate this predicament. Celastrol (Cel) is an extensively studied natural bioactive compound that has shown potentially promising antimalarial activity, but its antimalarial mechanism remains largely elusive.MethodsWe first established the Plasmodium berghei ANKA-infected C57BL/6 mouse model and systematically evaluated the antimalarial effects of Cel in conjunction with in vitro culture of Plasmodium falciparum. The potential antimalarial targets of Cel were then identified using a Cel activity probe based on the activity-based protein profiling (ABPP) technology. Subsequently, the antimalarial mechanism was analyzed by integrating with proteomics and transcriptomics. The binding of Cel to the identified key target proteins was verified by a series of biochemical experiments and functional assays.ResultsThe results of the pharmacodynamic assay showed that Cel has favorable antimalarial activity both in vivo and in vitro. The ABPP-based target profiling showed that Cel can bind to a number of proteins in the parasite. Among the 31 identified potential target proteins of Cel, PfSpdsyn and PfEGF1-α were verified to be two critical target proteins, suggesting the role of Cel in interfering with the de novo synthesis of spermidine and proteins of the parasite, thus exerting its antimalarial effects.ConclusionsIn conclusion, this study reports for the first time the potential antimalarial targets and mechanism of action of Cel using the ABPP strategy. Our work not only support the expansion of Cel as a potential antimalarial agent or adjuvant, but also establishes the necessary theoretical basis for the development of potential antimalarial drugs with pentacyclic triterpenoid structures, as represented by Cel.5LH6oqsCtKBdVDis1L_PeKVideo

Antigenic strain diversity predicts different biogeographic patterns of maintenance and decline of antimalarial drug resistance.

The establishment and spread of antimalarial drug resistance vary drastically across different biogeographic regions. Though most infections occur in sub-Saharan Africa, resistant strains often emerge in low-transmission regions. Existing models on resistance evolution lack consensus on the relationship between transmission intensity and drug resistance, possibly due to overlooking the feedback between antigenic diversity, host immunity, and selection for resistance. To address this, we developed a novel compartmental model that tracks sensitive and resistant parasite strains, as well as the host dynamics of generalized and antigen-specific immunity. Our results show a negative correlation between parasite prevalence and resistance frequency, regardless of resistance cost or efficacy. Validation using chloroquine-resistant marker data supports this trend. Post discontinuation of drugs, resistance remains high in low-diversity, low-transmission regions, while it steadily decreases in high-diversity, high-transmission regions. Our study underscores the critical role of malaria strain diversity in the biogeographic patterns of resistance evolution.

Antigenic strain diversity predicts different biogeographic patterns of maintenance and decline of antimalarial drug resistance

The establishment and spread of antimalarial drug resistance vary drastically across different biogeographic regions. Though most infections occur in sub-Saharan Africa, resistant strains often emerge in low-transmission regions. Existing models on resistance evolution lack consensus on the relationship between transmission intensity and drug resistance, possibly due to overlooking the feedback between antigenic diversity, host immunity, and selection for resistance. To address this, we developed a novel compartmental model that tracks sensitive and resistant parasite strains, as well as the host dynamics of generalized and antigen-specific immunity. Our results show a negative correlation between parasite prevalence and resistance frequency, regardless of resistance cost or efficacy. Validation using chloroquine-resistant marker data supports this trend. Post discontinuation of drugs, resistance remains high in low-diversity, low-transmission regions, while it steadily decreases in high-diversity, high-transmission regions. Our study underscores the critical role of malaria strain diversity in the biogeographic patterns of resistance evolution.

Antimicrobial Susceptibility and Characterization of Extended-Spectrum β-Lactamase-Producing Escherichia coli Isolated from Stools of Primary Healthcare Patients in Ethiopia.

Antimicrobial resistance of Escherichia coli is a growing problem in both developed and developing countries. This study aimed to investigate the phenotypic antimicrobial resistance of E. coli isolates (n = 260) isolated from the stool specimen of patients attending public health facilities in Addis Ababa and Hossana. This study also aimed to characterize phenotypically confirmed extended-spectrum beta-lactamase (ESBL)-producing E. coli isolates (n = 22) using whole-genome sequencing. Resistance to 18 different antimicrobials was assessed using the disc diffusion method according to the European Committee on Antimicrobial Susceptibility Testing (EUCAST) guidelines. The highest resistance rate among the E. coli isolates was found for ampicillin (52.7%), followed by trimethoprim-sulfamethoxazole (29.6%). Of all isolates, 50 (19.2%) were multidrug-resistant and 22 (8.5%) were ESBL producers. ESBL genes were detected in 94.7% of the sequenced E. coli isolates, and multiple β-lactamase genes were detected in 57.9% of the isolates. The predominant ESBL gene identified was blaCTX-M-15 (78.9%). The blaTEM-1B gene was detected in combination with other ESBL genes in 57.9% of the isolates, while only one of the sequenced isolates contained the blaTEM-1B gene alone. The blaCTX-M-3 gene was detected in three isolates. The genes blaCTX-M-15 and blaTEM-1B as well as blaCTX-M-15 and blaTEM-169 were confirmed to coexist in 52.6% and 10.5% of the sequenced E. coli isolates, respectively. In addition, blaOXA-1 was identified together with blaCTX-M-15 and blaTEM-1B in one isolate, and in one isolate, blaTEM-169 together with blaCTX-M-15 and blaTEM-1B was found. The results obtained show that measures need to be taken to reduce the spread of drug resistance and ensure the long-term use of available antimicrobials.

Assessment of antimalarial activity of crude extract of Chan-Ta-Lee-La and Pra-Sa-Chan-Dang formulations and their plant ingredients for new drug candidates of malaria treatment: In vitro and in vivo experiments.

The emergence and spread of antimalarial drug resistance have become a significant problem worldwide. The search for natural products to develop novel antimalarial drugs is challenging. Therefore, this study aimed to assess the antimalarial and toxicological effects of Chan-Ta-Lee-La (CTLL) and Pra-Sa-Chan-Dang (PSCD) formulations and their plant ingredients. The crude extracts of CTLL and PSCD formulations and their plant ingredients were evaluated for in vitro antimalarial activity using Plasmodium lactate dehydrogenase enzyme and toxicity to Vero and HepG2 cells using the tetrazolium salt method. An extract from the CTLL and PSCD formulations exhibiting the highest selectivity index value was selected for further investigation using Peter's 4-day suppressive test, curative test, prophylactic test, and acute oral toxicity in mice. The phytochemical constituents were characterized using gas chromatography-mass spectrometry (GC-MS). Results showed that ethanolic extracts of CTLL and PSCD formulations possessed high antimalarial activity (half maximal inhibitory concentration = 4.88, and 4.19 g/mL, respectively) with low cytotoxicity. Ethanolic extracts of the CTLL and PSCD formulations demonstrated a significant dose-dependent decrease in parasitemia in mice. The ethanolic CTLL extract showed the greatest suppressive effect after 4 days of suppressive (89.80%) and curative (35.94%) testing at a dose of 600 mg/kg. Moreover, ethanolic PSCD extract showed the highest suppressive effect in the prophylactic test (65.82%) at a dose of 600 mg/kg. There was no acute toxicity in mice treated with ethanolic CTLL and PSCD extracts at 2,000 mg/kg bodyweight. GC-MS analysis revealed that the most abundant compounds in the ethanolic CTLL extract were linderol, isoborneol, eudesmol, linoleic acid, and oleic acid, whereas ethyl 4-methoxycinnamate was the most commonly found compound in the ethanolic PSCD extract, followed by 3-hydroxy-2-(4-hydroxy-3-methoxyphenyl)-4H-chromen-4-one, flamenol, oleic acid amide, linoleic acid, and oleic acid. In conclusions, ethanolic CTLL and PSCD extracts exhibited high antimalarial efficacy in vitro. The ethanolic CTLL extract at a dose of 600 mg/kg exhibited the highest antimalarial activity in the 4-day suppressive and curative tests, whereas the ethanolic PSCD extract at a dose of 600 mg/kg showed the highest antimalarial activity in the prophylactic test.

A spatio-temporal model of multi-marker antimalarial resistance.

The emergence and spread of drug-resistant Plasmodium falciparum parasites have hindered efforts to eliminate malaria. Monitoring the spread of drug resistance is vital, as drug resistance can lead to widespread treatment failure. We develop a Bayesian model to produce spatio-temporal maps that depict the spread of drug resistance, and apply our methods for the antimalarial sulfadoxine-pyrimethamine. We infer from genetic count data the prevalences over space and time of various malaria parasite haplotypes associated with drug resistance. Previous work has focused on inferring the prevalence of individual molecular markers. In reality, combinations of mutations at multiple markers confer varying degrees of drug resistance to the parasite, indicating that multiple markers should be modelled together. However, the reporting of genetic count data is often inconsistent as some studies report haplotype counts, whereas some studies report mutation counts of individual markers separately. In response, we introduce a latent multinomial Gaussian process model to handle partially reported spatio-temporal count data. As drug-resistant mutations are often used as a proxy for treatment efficacy, point estimates from our spatio-temporal maps can help inform antimalarial drug policies, whereas the uncertainties from our maps can help with optimizing sampling strategies for future monitoring of drug resistance.

Drug resistance markers in Plasmodium vivax isolates from a Kanchanaburi province, Thailand between January to May 2023.

Plasmodium vivax has become the predominant species in the border regions of Thailand. The emergence and spread of antimalarial drug resistance in P. vivax is one of the significant challenges for malaria control. Continuous surveillance of drug resistance is therefore necessary for monitoring the development of drug resistance in the region. This study aims to investigate the prevalence of the mutation in the P. vivax multidrug resistant 1 (Pvmdr1), dihydrofolate reductase (Pvdhfr), and dihydropteroate synthetase (Pvdhps) genes conferred resistance to chloroquine (CQ), pyrimethamine (P) and sulfadoxine (S), respectively. 100 P. vivax isolates were obtained between January to May 2023 from a Kanchanaburi province, western Thailand. Nucleotide sequences of Pvmdr1, Pvdhfr, and Pvdhps genes were amplified and sequenced. The frequency of single nucleotide polymorphisms (SNPs)-haplotypes of drug-resistant alleles was assessed. The linkage disequilibrium (LD) tests were also analyzed. In Pvmdr1, T958M, Y976F, and F1076L, mutations were detected in 100%, 21%, and 23% of the isolates, respectively. In Pvdhfr, the quadruple mutant allele (I57R58M61T117) prevailed in 84% of the samples, followed by (L57R58M61T117) in 11%. For Pvdhps, the double mutant allele (G383G553) was detected (48%), followed by the triple mutant allele (G383M512G553) (47%) of the isolates. The most prevalent combination of Pvdhfr (I57R58M61T117) and Pvdhps (G383G553) alleles was sextuple mutated haplotypes (48%). For LD analysis, the association in the SNPs pairs was found between the intragenic and intergenic regions of the Pvdhfr and Pvdhps genes. The study has recently updated the high prevalence of three gene mutations associated with CQ and SP resistance. Genetic monitoring is therefore important to intensify in the regions to further assess the spread of drug resistant. Our data also provide evidence on the distribution of drug resistance for the early warning system, thereby threatening P. vivax malaria treatment policy decisions at the national level.

Antibiotic susceptibility trends in bacterial isolates from wound infections

Pyogenic infections leading to pus formation are associated with various inflammatory mediators and the proliferation of microbial flora. Wound infections can be caused by both aerobic and anaerobic bacteria, viruses, fungi, or even protozoans. Timely treatment of infected patients is critical since untreated or poorly treated pyogenic infections can lead to mortality or severe complications. Successful treatment is defined by choosing the right antibiotic, which is hindered by the spreading of bacterial drug resistance. It is well documented that overuse of antimicrobials before and especially during the COVID-19 pandemic has resulted in the emergence of drug resistance among various microorganisms. Our study aimed to identify common etiological agents associated with wound infections and to evaluate their antibiotic resistance patterns. A cross-sectional study was conducted by analyzing the antibiograms of different aerobic bacteria isolated from wound samples of patients from the Palakkad District Hospital in Kerala, India, from 2017 through 2019. All variables, including age, sex, bacterial isolate, and antibiogram, were tabulated and the chi-square test was used for analysis. Out of the 1,269 bacterial isolates analyzed, the most common gram-negative strain was Escherichia coli (23%) followed by Pseudomonas aeruginosa (20%). Among the gram-positive isolates, Staphylococcus aureus was the most common (19%). We found only 15% of Staphylococcus aureus isolates were methicillin-resistant, against the national average of 41%. The most resistant strain in our study was Acinetobacter baumannii. Statistically significant changes in antibiotic resistance patterns were observed over the three years of the study period.

Antimicrobial Sensitivity Pattern Analysis of Bacterial Species Isolated from Urine Samples

Background: Antimicrobial resistance is a significant global health issue. In order to apply efficient control measures to stop the rapid spread of drug resistance, it is vital to comprehend the sensitivity pattern of antibiotics in various types of therapeutically relevant samples.

Multi-drug-resistant tuberculosis and its associated factors among pulmonary tuberculosis patients linked to first-line anti-tuberculosis drugs in north-west Ethiopia

Introduction. Multi-drug-resistant tuberculosis (MDR-TB) is an emerging global challenge. Ethiopia is one of the 20 top countries with the highest estimated numbers of incidents of MDR-TB. Recently, the World Health Organization warned that drug-resistant TB is escalating and called for concerted action to reduce the spread of drug resistance. Hypothesis. The current study investigated MDR-TB in patients receiving first-line anti-TB drug treatment and associated factors. Aim. The study aimed to determine the prevalence of MDR-TB and its associated factors among smear-positive pulmonary TB patients receiving first-line anti-TB drug treatment. Methodology. An institution-based cross-sectional study was employed. All data were collected from laboratory result log books and information via a questionnaire. Samples from 205 smear-positive pulmonary TB patients were selected among first-line drug treatment by a systematic sampling method. Specimens were transported to Felege Hiwot referral hospital laboratory for GeneXpert testing. Factors associated with an outcome variable in binary multi-variable logistic regression analysis at P<0.05 were considered statistically significant variables. An ethical approval letter was taken to the respective health facility and written consent was obtained from each participant. Results. The overall prevalence of MDR-TB was 9.3 % (95 % CI, 5.4 13.7 %). Sign and symptom experience of anti-TB drug side effects [adjusted odds ratio (AOR)=0.18, 95 % CI=0.03–0.99, P=0.049] and co-morbidity (AOR=0.03, 95 % CI=0.01–0.55, P=0.02) were statistically associated with the development of MDR-TB infection Conclusion. The prevalence of MDR-TB was high (9.3 %) and contributed highly to new cases (8.3 %). Factors associated with MDR-TB were previous treatment, co-morbidity and laboratory diagnosis method prior to TB treatment. Therefore, this finding aims to maximize early detection and treatment, strengthening TB infection control, and proper implementation of directly observed therapy short course recommendations to reduce the burden of MDR-TB.

Removal of selected sulfonamides and sulfonamide resistance genes from wastewater in full-scale constructed wetlands

Sulfonamides are high-consumption antibiotics that reach the aquatic environment. The threat related to their presence in wastewater and the environment is not only associated with their antibacterial properties, but also with risk of the spread of drug resistance in bacteria. Therefore, the aim of this work was to evaluate the occurrence of eight commonly used sulfonamides, sulfonamide resistance genes (sul1–3) and integrase genes intI1–3 in five full-scale constructed wetlands (CWs) differing in design (including hybrid systems) and in the source of wastewater (agricultural drainage, domestic sewage/surface runoff, and animal runs runoff in a zoo). The CWs were located in low-urbanized areas in Poland and in Czechia. No sulfonamides were detected in the CW treating agricultural tile drainage water. In the other four systems, four sulfonamide compounds were detected. Sulfamethoxazole exhibited the highest concentration in those four CWs and its highest was 12,603.23 ± 1000.66 ng/L in a CW treating a mixture of domestic sewage and surface runoff. Despite the high removal efficiencies of sulfamethoxazole in the tested CWs (86 %–99 %), it was still detected in the treated wastewater. The sul1 genes occurred in all samples of raw and treated wastewater and their abundance did not change significantly after the treatment process and it was, predominantly, at the level 105 gene copies numbers/mL. Noteworthy, sul2 genes were only found in the influents, and sul3 were not detected. The sulfonamides can be removed in CWs, but their elimination is not complete. However, hybrid CWs treating sewage were superior in decreasing the relative abundance of genes and the concentration of SMX. CWs may play a role in the dissemination of sulfonamide resistance genes of the sul1 type and other determinants of drug resistance, such as the intI1 gene, in the environment, however, the magnitude of this phenomenon is a matter of further research.

Examining the Frequency of bla CTX-M and bla TEM Genes in Escherichia Coli Isolates with Multiple Drug Resistance Isolated from Patients Hospitalized in Ahvaz University Hospitals

Background and Objectives In the last few decades, Escherichia coli strains producing extended-spectrum beta-lactamases (ESBLs) have increased and are now considered a major problem in hospitalized patients. The aim of this study was to investigate the prevalence of blaCTX-M and blaTEM genes in Escherichia coli with multiple drug resistance. Subjects and Methods This study was conducted from October 2021 to May 2022. The research site included the microbiology laboratories of Golestan, Imam Khomeini, and Shahid Beqai hospitals where isolates suspected of Escherichia coli were isolated from urine culture samples of patients with urinary tract infection. All Escherichia coli samples were identified by conventional biochemical and molecular methods. The antibiotic sensitivity of the isolates was determined by disk diffusion method. Then, in order to finally confirm the presence of blaCTX-M and blaTE genes, PCR method was used. Results Out of the 160 bacterial isolates examined, 138 (86.3%) showed multidrug resistance (MDR). The highest and lowest percentage of resistance was related to Amoxicillin-Clavonic acid (84.1%) and Meropenem (5.8%), respectively. Based on the combined disk method, 96 isolates (69.6%) were positive for Beta-lactamase enzymes (ESBLs). Among the 96 ESBL-positive isolates, 85 (88.5%) and 38 (39.6%) carried blaCTX-M and blaTEM genes, respectively. Conclusion Given the significant prevalence of Escherichia coli strains that are resistant to multiple drugs and carrying ESBLs beta-lactamase genes in hospital, there is a need for effective surveillance measures to prevent the further spread of drug resistance in these isolates.

Evolution and transmission of antibiotic resistance is driven by Beijing lineage Mycobacterium tuberculosis in Vietnam.

Drug-resistant tuberculosis (TB) infection is a growing and potent concern, and combating it will be necessary to achieve the WHO's goal of a 95% reduction in TB deaths by 2035. While prior studies have explored the evolution and spread of drug resistance, we still lack a clear understanding of the fitness costs (if any) imposed by resistance-conferring mutations and the role that Mtb genetic lineage plays in determining the likelihood of resistance evolution. This study offers insight into these questions by assessing the dynamics of resistance evolution in a high-burden Southeast Asian setting with a diverse lineage composition. It demonstrates that there are clear lineage-specific differences in the dynamics of resistance acquisition and transmission and shows that different lineages evolve resistance via characteristic mutational pathways.

Genomics of Plasmodium vivax in Colombia reveals evidence of local bottle-necking and inter-country connectivity in the Americas.

Colombia aims to eliminate malaria by 2030 but remains one of the highest burden countries in the Americas. Plasmodium vivax contributes half of all malaria cases, with its control challenged by relapsing parasitaemia, drug resistance and cross-border spread. Using 64 Colombian P. vivax genomes collected between 2013 and 2017, we explored diversity and selection in two major foci of transmission: Chocó and Córdoba. Open-access data from other countries were used for comparative assessment of drug resistance candidates and to assess cross-border spread. Across Colombia, polyclonal infections were infrequent (12%), and infection connectivity was relatively high (median IBD = 5%), consistent with low endemicity. Chocó exhibited a higher frequency of polyclonal infections (23%) than Córdoba (7%), although the difference was not significant (P = 0.300). Most Colombian infections carried double pvdhfr (95%) and single pvdhps (71%) mutants, but other drug resistance mutations were less prevalent (< 10%). There was no evidence of selection at the pvaat1 gene, whose P. falciparum orthologue has recently been implicated in chloroquine resistance. Global population comparisons identified other putative adaptations. Within the Americas, low-level connectivity was observed between Colombia and Peru, highlighting potential for cross-border spread. Our findings demonstrate the potential of molecular data to inform on infection spread and adaptation.

THU549 Targeting The Growth Hormone Receptor Attenuates Tumor Exosome Mediated Propagation Of Chemoresistance In Melanoma

Abstract Disclosure: P. Kulkarni: None. R. Basu: None. T. Bonn: None. J.J. Kopchick: None. Elevated ATP-binding cassette (ABC) transporters in tumor cells resulting in decreased intracellular drug accumulation is a major reason for chemotherapy resistance in cancer. Mounting evidence indicates that extracellular vesicles (EVs), particularly exosomes, the smallest subset of EVs, play a crucial role in the spread of drug resistance. We have previously shown that growth hormone (GH) decreases therapeutic efficacy in melanoma via upregulating the levels of ABC transporters, whereas when GHR is knocked down or GH/GHR interaction is inhibited by GHR antagonist, Pegvisomant, these ABC transporters are significantly downregulated. No study has yet directly assessed the effect of GH or GHR antagonism on exosomal cargoes in terms of promoting drug resistance. We observe that human melanoma cells (Malme3M, SK-Mel28, and MDA-MB-435) when treated with GH or doxorubicin alone or in combination increase the exosomal protein levels of multiple multi-drug efflux pumps, namely ABCC1, ABCC2, ABCB1, and ABCG2. On adding Pegvisomant to these treatments, we found a significant decrease in the levels of these exosomal drug efflux pumps induced by both autocrine and endogenously added GH. Furthermore, we assessed the role of GH in exosome-mediated dissemination of chemoresistance from drug-resistant to drug-sensitive cells. When GH and doxorubicin-treated melanoma-derived exosomes were administered to treatment-naïve melanoma cells, a significant increase in drug efflux was observed in the latter, compared to the control. On the other hand, when Pegvisomant co-treated melanoma-derived exosomes were administered to treatment-naïve melanoma cells, the drug efflux was attenuated, indicating chemo-sensitization. Collectively, these observations indicate that Pegvisomant suppresses various exosomal cargoes modulating drug resistance in melanoma. Acknowledgment: This work was supported in part by the State of Ohio’s Eminent Scholar Program includes a gift from Milton and Lawrence Goll, by the AMVETS, Ohio University’s Student Enhancement Award, and Edison Biotechnology Institute. Presentation: Thursday, June 15, 2023