Antibiotic Resistance Profile of CTX-M-type Extended-Spectrum Beta-Lactamases in Escherichia coli and Klebsiella pneumoniae in Accra, Ghana

Extended-spectrum beta-lactamases (ESBLs) are plasmid-mediated beta lactamases that are capable of hydrolysing beta-lactams except carbapenems and cephamycins. The most common ESBL types include CTX-M, TEM and SHV. This genetic diversity in the various ESBL-producing organisms may reflect characteristic differences in relation to pathogenesis, antibiotic resistance expression, response to therapy, transmission and infection control. This work sought to determine the characteristic antibiotic minimum inhibition concentrations (MICs) and antimicrobial sensitivity profile of CTX-M-type ESBLs in Accra. Hundred (100) DNA templates were extracted from ESBL-producing K. pneumoniae and E. coli isolates. The specific ESBL types were determined by polymerase chain reaction with specific primers and reaction conditions. The MICs of the antibiotics were determined using Vitek 2 Compact System (bioMérieux, Marcy I’Etoile, France). The results showed that CTX-M-type ESBL have cefotaxime MIC in the resistant range of >64 µg/ml. The CTX-M-type ß-lactamases showed co-resistances to gentamicin (88.6%), tetracycline (71.4%), trimethoprim-sulphamethoxazole (98.6%).The resistance of CTX-M-type ESBL producing organisms to fluoroquinolones have been well established in this work with resistances in ciprofloxacin (71.4%) and norfloxacin (71.4%) with MIC90 being >4 µg/ml and >16 µg/ml respectively. The beta-lactam-beta-lactamase inhibitor combination of piperacillin-tazobactam was more susceptible to CTX-M-type ESBL than amoxicillin-clavulanate. Imipenem and amikacin has been established as the in vitro drug of choice for the management of organisms producing CTX-M-type ESBL in this present work. Efforts should be made to control the increasing prevalence of CTX-M-type producing organisms in the communities and hospital settings in Accra with their adverse multiple-drug resistance. Keywords: Extended spectrum beta-lactamase, CTX-M-type ESBL, Resistance, Antibiotics

Antibiotic Resistance Profile of Non-Extended Spectrum Beta-Lactamase-producing Escherichia coli and Klebsiella pneumoniae in Accra, Ghana

One of the major challenges facing health professionals is the prevalence of antibiotic resistance. Most Gram-negative bacteria produce beta-lactamases which are enzymes that in-activate ?-lactams. Recent publications suggested that extended spectrum beta-lactamase production in E. coli and K. pneumoniae is one of the main causes of antimicrobial resistance in penicillins, cephalosporins and some non-beta-lactam antibiotics in Accra. This present work sought to determine the resistance profile of antimicrobials to non-ESBL-producing isolates in Accra. The 400 K. pneumoniae and E. coli isolates were screened for non-ESBL-producing strains using the combined disk method. The minimum inhibition concentration for 17 antibiotics was determined using Vitek 2 Compact System (bioMérieux, Marcy I’Etoile, France).  Among the 400 total bacterial isolates, 198 (49.5%) were non-ESBL producers. Co-resistances to ampicillin (66.7%), piperacillin (59.1%), tetracycline (77.8%) and trimethoprim/sulphamethoxazole (68.2%) have been collaborated in this work. The increasing rise in resistance to the beta-lactam/beta-lactamase inhibitor combination antibiotics such as amoxicillin/clavulanic acid (13.6%) and piperacillin/tazobactam (18.7%) is problematic since they have become the empirical drug of choice for treating most infections. The steady increase in resistance to gentamicin (17.2%) as well as the floroquinolones such as ciprofloxacin (39.4%) and norfloxacin (34.9%) is alarming. In the absence of ESBLs, cephalosporins generally have been effective in treating infections caused by enterobacteria. Nitrofurantoin remains reliable for managing non-life threatening urinary tract infections. Amikacin and imipenem continue to be effective third-line treatment options for Gram-negative bacteria infections.  As antibiotic resistance increases and the development of new antimicrobials declines, it is imperative that we use antimicrobials that are still effective rationally. Evidence based antibiotic prescriptions and usage as well as regular evaluation of antibiotic resistance will help to control the spread of antibiotic resistance in Accra, Ghana. Keywords: Extended spectrum beta-lactamase, Resistance, Antibioti

Comparative Analysis of Antimicrobial Resistance of Extended-Spectrum Beta-Lactamase Producers and Non-Extended-Spectrum Beta-Lactamase Producers among Bacterial Isolates in Accra, Ghana

Antibiotic resistance may occur naturally but misuse of antibiotics in humans and animals is accelerating the process. One of the modes of resistant mechanism is the production of extended-spectrum beta-lactamases (ESBLs) by the bacteria. ESBLs are plasmid-mediated beta-lactamases that are capable of hydrolysing penicillins, cephalosporin and several non-beta-lactam antibiotics. This laboratory-based study sought to compare the rate of antimicrobial resistance between ESBL and non-ESBL-producers in Accra. Four hundred K. pneumoniae and E. coli isolates were collected at the Korle Bu Teaching Hospital and screened for ESBL and non-ESBL-producers using the combined disk method and Vitek 2 system. The minimal inhibition concentrations (MICs) for 17 commonly used antibiotics were determined using Vitek 2 System. The results indicated significant difference (P<0.05) between the antimicrobial resistance of ESBL-producers and non-ESBL producers except for amikacin and imipenem. The 198 non-ESBL phenotypes recorded relatively low antimicrobial resistance to cefotaxime 4(2%), ceftazidime 4(2%), nitrofurantoin 6(3%), amoxicillin/clavulanic acid 27(13.6%), gentamicin 34(17.2%) and ciprofloxacin 78(39.4%). In contrast, the 202 ESBL producers registered high antibiotic resistance to cefotaxime 197(97.5%), ceftazidime 175(86.6%), nitrofurantoin 94(46.5%), amoxicillin/clavulanic acid 64(31.7%), gentamicin 166(82.2%) and ciprofloxacin 161(79.7%). Cephalosporins and nitrofurantoin are suitable for the treatment of non-ESBL producers while imipenem and amikacin is the drug of choice for treating ESBL-producing infections. Evidence based antibiotic usage will help to control the spread of resistance by ESBL producers in Accra, Ghana. Also, there is the need to intensify research in the use of natural products to treat ESBL infections. Keywords: Extended spectrum beta-lactamase, Resistance, Bacteria, Antibiotic

Emergence of Carbapenem-resistant Enterobacteriaceae among Extended-spectrum Beta-lactamase Producers in Accra, Ghana

Previous studies in Accra had established that extended-spectrum beta-lactamase (ESBL) producers were increasingly becoming a public health nuisance. Since ESBL producers resulted in multi-drug resistance among most beta-lactams and non-beta-lactams, the antibiotic of choice for the treatment of these ESBL infections was carbapenems such as imipenem, meropenem and ertapenems. Hence the emergence and spread of carbapenem resistant bacteria may lead to therapeutically dead end for life-threatening infections. This study therefore focussed on the occurrence of carbapenem resistant Enterobacteriaceae among ESBL producers from clinical specimens analysed at MDS-Lancet Laboratories, Accra, Ghana. One thousand (1000) clinical isolates were identified and analysed for ESBL producers using the combined disk synergy method. In order to determine the carbapenemase producing ESBL bacteria and the antibiotic of choice for treating carbapenem resistant infections, antimicrobial susceptibility testing was performed to determine the minimum inhibition concentration of the antibiotics used against the ESBL producers. The antibiotics used included imipenem, meropenem ertapenem and other antibiotics. The results indicated that 600 (75%) of the clinical isolates were ESBL producers. Among the 600 ESBL producers, 43 (7.2) were carbapenem resistant bacteria including 7 different Gram negative bacterial species. Among the carbapenemase producers, Escherichia coli (34.9%) and Klebsiella pneumoniae (25.6%) were the dominant bacterial species. The carbapenem resistant bacteria indicated multi-drug resistance to penicillins (100%), cephalosporins (100%), amoxicillin-clavulanic acid (100%), piperacillin-tazobactam (100%), ciprofloxacin (83.7%), gentamycin (79.7%), amikacin (27.9%), colistin (18.6%) and fosfomycin (11.6%).  Colistin seems to be is the drug of choice for treating carbapenem resistant strains. Although fosfomycin showed a higher activity, it is only recommended for urinary tract infections. Evidence based antibiotic usage and nosocomial infection control will help to control the emergence of carbapenem resistant strains in Accra, Ghana. Also, there is the need to intensify research in the use of natural products to treat resistant bacterial infections. Keywords: Carbapenem, ESBLs, Antibiotics, Colisti

Molecular Characterization of Extended-Spectrum Beta-Lactamases in Escherichia coli and Klebsiella pneumoniae in Accra, Ghana

Extended-spectrum beta-lactamases (ESBLs) are plasmid-mediated beta lactamases that are capable of hydrolysing beta-lactams except carbapenems and cephamycins. The ESBL types include SHV, TEM and CTX-M, OXA, PER and VEB-1. The most common ones isolated from clinical specimen are the CTX-M, SHV and TEM.  The specific ESBL-producing organisms have different genetic characteristics which mark their identification at the molecular level. This work sought to determine the genetic characterization of ESBL-producing K. pneumoniae and E. coli in Accra. The molecular investigations of the ESBL-coding genes included extraction of 100 DNA templates of phenotypic ESBL-producing isolates by boiling method, preparation of the PCR reaction mixture using appropriate primers, standard PCR reaction in a thermocycler,   agarose gel electrophoresis, bands visualization by ultraviolet trans-illumination and bands photography using a Kodak EDAS 290 gel documentation system. The results significantly (p<0.05) indicated that of the 100 ESBL producers, 90(90%) possess CTX-M genes and 25(25%) had TEM genes. None of the ESBL producers possesses SHV genes. Seventy (70%) of the ESBL producers possess only CTX-M genes and 5(5%) had only TEM genes. Twenty (20%) of the isolates had both CTX-M and TEM genes. Of the 100 ESBL phenotypes, 78(78%) and 2(2%) were positive for CTX-M-1group and CTX-M-9group ESBL genes respectively. Organisms producing CTX-M-type ESBL are more prevalent in Accra than other ESBL types. CTX-M-1group producing isolates dominated the ESBL phenotypes with CTX-M-15 likely to be the dominate CTX-M-type ESBL. There is the need for further studies into the characteristic transmission, pathogenesis, antibiotic resistance expression, and infection control of CTX-M-type ESBL and TEM-type ESBL in Accra. Keywords: Extended spectrum beta-lactamase, CTX-M genes, TEM genes, SHV genes, Molecula

Possible Health Risk due to the Environmental Exposure of High Levels of Lead in Exhaust Soot of Automobiles in Parts of Accra, Ghana

Internal combustion engines produce soot as a result of incomplete gasoline and diesel combustion.  Leaded exhaust soot emitted into the atmosphere has serious health and environmental concerns. Lead has been outlawed as an automotive gasoline additive in most countries including Ghana because of its cumulative toxicity in humans especially children and damaging effect on catalytic converters in automobiles. Nevertheless, leaded fuels are apparently being produced, imported and used illegally in some countries as octane rating booster because of its profitability. Refined gasoline and diesel are imported into Ghana through bulk oil distribution firms. This preliminary study assessed the level of lead in automotive exhaust soot from randomly selected automobiles in parts of Accra. Exhaust soot samples obtained from ten diesel and ten gasoline automobiles were collected for analysis of its lead concentration using atomic absorption spectrophotometry. The results showed the presence of lead in 4(40%) and 10(100%) of the randomly selected diesel and gasoline vehicular exhaust soot respectively. The concentration of lead in the exhaust soot of diesel-powered automobiles ranged from 0.060mg/kg to 0.435mg/kg and that of the gasoline-powered vehicles recorded values ranging from 0.195mg/kg to 2.055mg/kg. With this rather high level of lead in the vehicular soot, it could be concluded that the exhaust soot can be a significant source of lead in the atmosphere in parts of Accra. Lead exposure is known to cause debilitating developmental and neurological effects in children and cardiovascular effects in adults. The high levels of lead in the exhaust soot may be attributed to the possibility of lead additives in the gasoline and diesel used by those automobiles. Regulators of the petroleum downstream industry such as the National Petroleum Authority must routinely test for lead in imported refined petroleum products and enforce the ban on the importation, sale and usage of the outlawed leaded fuel in Ghana. Further studies should be conducted on the levels of lead in air and blood lead levels in fuel dispensers, fuel tanker drivers and fuel loading workers of bulk oil distribution firms. Keywords: Lead, Exhaust, Soot, Gasoline, Automobil

Antibacterial Activity of Alchornea cordifolia (Christmas bush) Leaves Extract on Carbapenem Resistant Enterobacteriaceae Causing Multi-drug Resistant Systemic Infections

Carbapenems have been used for the treatment of systemic infections caused by Enterobacteriaceae. However, recent studies suggested that some Enterobacteriaceae are producing carbapenemases, which has limited the treatment options for carbapenem-resistant Enterobacteriaceae. Some of the emerging carbapenemase resistant Enterobacteriaeae that are causing multi-drug resistant systemic infections include Escherichia coli, Klebsiellae pneumoniae, Providencia rettgeri, Proteus mirabili, Pantoea species, Citrobacter koseri and Acinetobacter baumanii. There is therefore the need for alternative treatment regimens for carbapenem-resistant Enterobacteriaceae. This study determined the in vitro efficacy of Alchornea cordifolia on carbapenem-resistant Enterobacteriaceae using agar well diffusion and well microplate dilution method. Serial dilutions of the ethanolic crude extract of the leaves were prepared and used against the carbapenem-resistant Enterobacteriaceae. The phytochemical screening was also performed to determine the antibacterial compounds. The christmas bush leaves extracts concentrations ranging from 50 mg/ml – 200 mg/ml showed active diameter zone of inhibition. The ethanolic extract of Christmas bush leaves had minimum inhibition concentration and minimum bactericidal concentration of 3.13mg/ml indicating significant antibiotic activity against the carbapenem-resistant Enterobacteriaceae. The phytochemical screening of the Christmas bush leaves showed the presence of antimicrobial phytochemicals such as flavonoids. This offers the possibility of developing effective antimicrobial agent to treat multi-drug resistant systemic infections. Therefore, there is the need to determine the toxicological effect and perform clinical trials of the active antimicrobial compounds isolated in the leave extracts of Christmas bush shrub. Keywords: Alchornea cordifolia, Flavonoids, Antibacterial, Carbapenem-resistant Enterobacteriaceae DOI: 10.7176/JNSR/10-10-03 Publication date:May 31st 202

Alchornea Cordifolia (Christmas Bush) Leaves Extract Shows Activity Against Extended-Spectrum Beta-Lactamase-Producing Klebsiella Pneumoniae That Cause Multi-Drug Resistant Urinary Tract Infections

Klebsiella pneumoniae are one of the most common cause of multi-drug resistant urinary tract infections such as cystitis and pyelonephritis. Extended-spectrum beta-lactamases are plasmid-mediated enzymes commonly found in the Klebsiella pneumoniae and other Gram negative bacteria. They are capable of hydrolysing beta-lactams except carbapenems and cephamycins. Extended-spectrum beta-lactamases also confer resistance to several non-beta-lactam antibiotics, limiting treatment options for urinary tract infections caused by extended-spectrum beta-lactamases -producing K. pneumoniae. This study determined the in vitro efficacy of Alchornea cordifolia on extended-spectrum beta-lactamases -producing K. pneumoniae. Serial dilutions of the ethanolic extract of the leaves were prepared and tested against the extended-spectrum-beta-lactamases-producing K. pneumoniae. The phytochemical screening was performed to determine the antibacterial compounds. The Christmas bush leaves extracts concentrations ranging from 50 mg/ml – 200 mg/ml showed significant active diameter zone of inhibition. The ethanolic extract of A. cordifolia leaves had minimum inhibition concentration and minimum bactericidal concentration of 3.13 mg/ml indicating significant antibiotic activity against the ESBL isolates. The phytochemical screening of the Christmas bush leaves showed the presence of antimicrobial phytochemicals such as flavonoids. Ethanolic extracts of Christmas bush leaves is proving to be efficacious against multi-drug resistant urinary tract infections. This offers hope for the development of effective antibiotics due to the presence of flavonoids in the A. cordifolia leaf extracts. Therefore, there is the need to determine the toxicological effect and clinical trials of the active antimicrobial compounds isolated in the leaf extracts of A. cordifolia shrub. Keywords: Alchornea. cordifolia, Flavonoids, Multi-drug resistant, Extended-spectrum beta-lactamase, Klebsiella pneumoniae DOI: 10.7176/JNSR/11-24-03 Publication date: December 31st 202

Antimicrobial Activity of Psidium Guajava (Guava) Leaves Extract on Extended-Spectrum Beta-Lactamase-Producing Klebsiella Pneumoniae that Cause Multi-Drug Resistant Urinary Tract Infections

Extended-spectrum beta-lactamases (ESBLs) are plasmid-mediated enzymes commonly found in the Enterobacteriaceae that are capable of hydrolysing ß-lactams except carbapenems and cephamycins. ESBLs also confer resistance to several non-ß-lactam antibiotics. ESBL producing isolates are predominantly Klebsiella pneumoniae and Eschericia coli. Klebsiella pneumoniae are one of the most common causes of urinary tract infections such as cystitis and pyelonephritis developed by about 150 million people in every given year. ESBL-producing Klebsiella pneumoniae appear susceptible to cephalosporins in vitro using conventional breakpoints but ineffective in vivo. This has limited treatment options for urinary tract infections caused by extended-spectrum beta-lactamases -producing K. pneumoniae. This study determined the in vitro efficacy of Psidium guajava on extended-spectrum beta-lactamases -producing K. pneumoniae using agar well diffusion and well microplate dilution method. Serial dilutions of the ethanolic extract of the leaves were prepared and tested against the extended-spectrum beta-lactamases-producing K. pneumoniae. The phytochemical screening was performed to determine the antibacterial compounds. The guava leaves extracts concentrations ranging from 50 mg/ml – 200 mg/ml showed active diameter zone of inhibition. The ethanolic extract of guava leaves had minimum inhibition concentration and minimum bactericidal concentration of 6.25mg/ml indicating significant antibiotic activity against the ESBL isolates. The phytochemical screening of the guava leaves showed the presence of antimicrobial phytochemicals such as flavonoids. Ethanolic extracts of guava leaves is proving to be efficacious against multi-drug resistant ESBL-producing K. pneumoniae, which a major cause urinary tract infections. This offers hope for the development of effective antibiotics against multi-drug resistant urinary tract infections due to the presence of flavonoids in the guava leaf extracts. Therefore, there is the need to determine the toxicological effect of the active antimicrobial compounds isolated in the leaf extracts of guava plant. Keywords: Psidium guajava, Flavonoids, Multi-drug resistant, Extended-spectrum beta-lactamase, Klebsiella pneumoniae DOI: 10.7176/JNSR/11-22-02 Publication date: November 30th 202

Phenotypic Characterization of AmpC beta-lactamase among Cefoxitin Resistant Escherichia coli and Klebsiella pneumoniae Isolates in Accra, Ghana

AmpC ?-lactamases hydrolyze penicillins, cephalosporins and cephamycins and resist inhibition by clavulanate, sulbactam, and tazobactam. Strains with AmpC genes are inherently resistant to multiple agents, making the selection of an effective antibiotic difficult. This present work sought to investigate the occurrence of AmpC beta-lactamases-producing phenotypes in E. coli and K. pneumoniae and their antimicrobial sensitivity profile. Four hundred K. pneumoniae and E. coli non-duplicate isolates were collected and their antibiotic sensitivity testing for cefoxitin and other 16 antibiotics were determined using Vitek 2 Compact System (bioMérieux, Marcy I’Etoile, France).  The isolates resistant to cefoxitin were confirmed as AmpC beta-lactamases-producing phenotypes with disk synergy testing (DST) using cefotaxime or ceftazidime with or without boronic acid. An increase in zone diameter of ?5mm in the presence of boronic acid indicates the presence of AmpC beta-lactamases in the test organism. The results showed that of the 50 cefoxitin resistant isolates screened from 400 bacterial isolates, 5(10%) were AmpC beta-lactamase-producers with 60%, 60%, 60%, 80% and 100% multiply antibiotic resistance in gentamicin, ciprofloxacin, norfloxacin, trimethoprim/sulfamethoxazole and tetracycline respectively. Nitrofurantoin which indicated 100% susceptibility with MIC90 of 32µg/ml may be a therapeutic option especially for non-life-threatening urinary tract infection. Imipenem was the antibiotic of choice with 100% susceptibility rates (MIC90 of ?1µg/ml). Though the insignificant (p>0.05) levels of AmpC beta-lactamase phenotypes may not require routine detection in health facilities, there is the need to implement evolutionary antibiotic administration policies and pragmatic infection control measures in the hospitals.      Keywords: AmpC beta-lactamase, Cefoxitin, ?-lactams, E. coli, K. pneumonia