Gram-negative Bacterial Isolates Research Articles

Spectrum and Antibiotic Sensitivity Pattern of Bloodstream Bacterial Isolates from Septicemic Neonates

Aim: To evaluate the etiology of neonatal sepsis. We aimed to explore the antimicrobial sensitivity patterns among neonates. Methodology: This observational-based study was conducted in POF hospital Wah Cantt from September 2020 to March 2021. We selected 300 sepsis patients based on the WHO neonatal sepsis screening tool. Blood samples were collected by trained laboratory technicians using the aseptic technique. Laboratory technicians prepared a 5cm (in diameter) patch of skin over the proposed site of venipuncture. We drew one ml sample of blood from a fresh venipuncture site. This blood was added to a bottle containing 5–10 mL of blood Agar culture media. Results: We observed that babies with less than 5 Apgar scores reported a high risk of neonatal sepsis. We observed gram-positive bacteria in 49 (55.7%) cases while 39 (44.3%) were reported gram-negative bacteria. At the time of diagnosis, we observed that 67.3% of cases of gram-positive bacteria were highly reported in the LONS group. Around 23 (63.3%) gram-negative bacterial isolates were from EONS. Regarding bacterial pathogen, we observed 22 (25%) cases of Coagulase-negative staphylococcus aureus and 18 (20%) were of E. coli which results in neonatal sepsis. Conclusion: Our results concluded that third-line antimicrobials can be a beneficial treatment for managing neonatal sepsis. However, failure of irst two antimicrobials was observed due to high utilization before an accurate diagnosis of neonatal sepsis. Keywords: Antimicrobial, Sensitivity Patterns,. Neonatal sepsis

Chemical Composition of Kickxia aegyptiaca Essential Oil and Its Potential Antioxidant and Antimicrobial Activities.

The exploration of new bioactive compounds from natural resources as alternatives to synthetic chemicals has recently attracted the attention of scientists and researchers. To our knowledge, the essential oil (EO) of Kickxia aegyptiaca has not yet been explored. Thus, the present study was designed to explore the EO chemical profile of K. aegyptiaca for the first time, as well as evaluate its antioxidant and antibacterial activities, particularly the extracts of this plant that have been reported to possess various biological activities. The EO was extracted from the aerial parts via hydrodistillation and then characterized by gas chromatography-mass spectrometry (GC-MS). The extracted EO was tested for its antioxidant activity via the reduction in the free radicals, 2,2-diphenyl-1-picrylhydrazyl (DPPH) and 2,2′-azinobis(3-ethylbenzothiazoline-6-sulfonic acid) (ABTS). In addition, the EO was tested as an antibacterial mediator against eight Gram-negative and Gram-positive bacterial isolates. Forty-three compounds were identified in the EO of K. aegyptiaca, with a predominance of terpenoids (75.46%). Oxygenated compounds were the main class, with oxygenated sesquiterpenes attaining 40.42% of the EO total mass, while the oxygenated monoterpenes comprised 29.82%. The major compounds were cuminic aldehyde (21.99%), caryophyllene oxide (17.34%), hexahydrofarnesyl acetone (11.74%), ar-turmerone (8.51%), aromadendrene oxide (3.74%), and humulene epoxide (2.70%). According to the IC50 data, the K. aegyptiaca EO revealed considerable antioxidant activity, with IC50 values of 30.48 mg L−1 and 35.01 mg L−1 for DPPH and ABTS, respectively. In addition, the EO of K. aegyptiaca showed more substantial antibacterial activity against Gram-positive bacterial isolates compared to Gram-negative. Based on the minimum inhibitory concentration (MIC), the EO showed the highest activity against Escherichia coli and Bacillus cereus, with an MIC value of 0.031 mg mL−1. The present study showed, for the first time, that the EO of K. aegyptiaca has more oxygenated compounds with substantial antioxidant and antibacterial activities. This activity could be attributed to the effect of the main compounds, either singular or synergistic. Thus, further studies are recommended to characterize the major compounds, either alone or in combination as antioxidants or antimicrobial agents, and evaluate their biosafety.

In-vitro activity of ceftolozane/tazobactam against recent clinical bacterial isolates from two Saudi Arabian hospitals

BackgroundAntimicrobial resistance is continuously increasing. Adding tazobactam to ceftolozane improves the latter’s activity spectrum against resistant strains. We aimed to determine the susceptibility of recently collected bacterial isolates to ceftolozane/tazobactam (C/T) and other antibiotics. MethodsThis was a prospective cohort study conducted between March 2017 and March 2018. The in-vitro activities of C/T and 14 other antibiotics were assessed against 192 gram-negative bacterial (GNB) isolates (P. aeruginosa, K. pneumonia, E. coli, and other Enterobacterales) prospectively collected from two hospitals in Saudi arabia; in the laboratories of the International Health Management Associates Inc. Samples were obtained from intensive care units (ICUs) and non-ICU locations. The minimum inhibitory concentrations (MICs) of the antibiotics were determined by broth microdilution. Isolates were obtained from different infection sites [urine (31.8%), urinary bladder samples (15.1%), abscess/pus (20.3%), endotracheal aspirates (18.8%)]. ResultsOur sample showed substantial drug resistance; 66.1% of the collected isolates showed either multiple or extensive drug resistance. Susceptibility rates of P. aeruginosa (n = 50), E.coli (n = 40), K. pneumoniae (n = 64) and other Enterobacterales (n = 38) to C/T were 74%, 87.5%, 48.4% and 71.1%, respectively. According to MIC50 values (1 µg/mL for both P. aeruginosa and other Enterobacterales, 0.5 µg/mL for E.coli, and 4 µg/mL for K. pneumoniae), C/T was among the most potent antibiotics against these isolates. ConclusionsC/T displayed high potency against all examined bacterial isolates. It was mainly active against E.coli followed by P.aeruginosa and other Enterobacterales and its lowest susceptibility rate was reported against K. pneumoniae.

Molecular Characterization of Antibiotic Resistant Genes among Gram Negative Clinical Isolates

Increasing antibiotic resistance is a threat worldwide. Deep studies have to be done for correct and appropriate usage of antibiotics for treating infections efficiently and to curtail the spread of resistance. Hence, this study is aimed at understanding the prevalence of antibiotic resistance genes among 35 gram-negative bacterial isolates. By performing biochemical tests, the isolates were identified to be Escherichia coli (n = 15), Klebsiella oxytoca (n = 3), Klebsiella pneumoniae (n = 5), Proteus mirabilis (n = 4), Proteus vulgaris (n = 1) and Pseudomonas aeruginosa (n = 7). Antibiotic susceptibility tests showed that most E. coli isolates were resistant to ampicillin, levofloxacin and cefazolin. Ampicillin resistance was found in all of Klebsiella oxytoca, Klebsiella pneumoniae and Proteus mirabilis isolates. All Proteus mirabilis isolates were resistant to ciprofloxacin and co-trimoxazole. Pseudomonas aeruginosa showed maximum resistance to gentamycin and tobramycin. All 35 isolates were sensitive to amikacin, imipenem and meropenem. Polymerase Chain Reaction showed that AmpC and SHV genes were predominant in E. coli and Klebsiella species respectively. Among Proteus mirabilis and Pseudomonas aeruginosa, IntI gene was more prevailing. NDM1 gene was not detected in any of the isolates. This study also showed the prevalence of isolates resistant to multiple drugs due to the co-existence of resistant genes. Therefore, control of spread of infections, routine study in this field and proper guidelines for prescribing antibiotics is very necessary.
 HIGHLIGHTS
 
 This study is aimed at understanding the prevalence of antibiotic resistance genes among 35 gram-negative bacterial isolates
 A considerable number of isolates had antibiotic resistance genes and some isolates showed resistance to multiple drugs due to the co-existence of resistant genes
 Infections caused by bacteria that are resistant to antibiotics are difficult to treat and may sometimes lead to death
 Regular study and proper use of antibiotics is essential to curtail the spread of antibiotic resistant bacteria. Regular
 
 GRAPHICAL ABSTRACT

Plasmid-mediated quinolone resistance determinants in clinical bacterial pathogens isolated from the Western Region of Ghana: a cross-sectional study.

quinolones are critically important antibiotics that are reserved for treating very severe infections caused by multidrug-resistant bacterial pathogens. However, their indiscriminate uses have resulted in an increased number of resistant strains in many parts of the world including Ghana. We determined the quinolone resistance profile of Gram-negative bacterial pathogens and characterized the underlying molecular determinants of resistance. Gram-negative pathogens obtained from clinical specimens at three hospital laboratories were tested for resistance to quinolones and other commonly used antibiotics. ESBL production among the Enterobacterial isolates was confirmed using the combined disc diffusion method. We then used PCR to determine seven types of plasmid-mediated quinolone resistance genes present in the isolates resistant to nalidixic acid and ciprofloxacin. in this study, 29.5% of the isolates were resistant to ciprofloxacin, with the highest of 50% among E. coli resistance to the other quinolones was levofloxacin (24.4%), norfloxacin (24.9%), and nalidixic acid (38.9%). Significant proportions of the quinolone-resistant isolates were ESBL producers (P-values < 0.001). The aac(6´)-Ib-cr, qnrS, oqxA, and qepA genes were present in 43 (89.6%), 27 (56.3%), 23 (47.9%), and one (2.1%) of the isolates, respectively. None of the isolates tested positive to qnrA, qnrB, and oqxB genes. The presence of the aac(6´)-Ib-cr gene positively correlated with resistance to ceftriaxone, cefotaxime, and gentamicin (P-values < 0.05). high proportions of Gram-negative bacterial isolates were resistant to quinolones and most of these isolates possessed multiple PMQR genes. There is a need to implement measures to limit the spread of these organisms.

Prevalence and Antibiotic Sensitivity pattern of Bacterial Isolates from Catheterized UTI Patients: A Local Study

Prevalence and multidrug resistance among bacteria in catheter-associated urinary tract infections (CAUTIs) has been on the rise in recent times. Hence, the prevalence rate and antibacterial susceptibility of bacteria in CAUTIs in ICU patients was evaluated. A total of 120 patients admitted to the ICU of Nishtar Hospital, Multan, were recruited for this study. Both gram-positive and gram-negative bacterial isolates were characterized based on biochemical tests including catalase test, oxidase test, indole test, TSI test, citrate test, coagulase test and growth on 6.5% NaCl agar. The prevalence of bacterial species was Escherichia coli (32%), Staphylococcus aureus (26%), Pseudomonas spp. (18%), Proteus spp. (14%) and Enterococcus spp. (2%). A considerable degree of resistance against commonly prescribed antibiotics was observed. Gram negative bacteria showed resistance to ciprofloxacin, piperacillin-tazobactam and amikacin as well as susceptibility to imipenem, tigecycline and polymixin. Gram positive bacteria showed resistance to antibiotics such as piperacillin-tazobactam, ampicillin, gentamicin, oxacillin and ceftazidime suggesting the ineffectiveness of these antibiotics for treating bacterial infections among CAUTI patients and demonstrating the latest trends in antimicrobial drug resistance profile in local population.

Development of a Monte Carlo simulation model to predict pasteurized fluid milk spoilage due to post-pasteurization contamination with gram-negative bacteria

Psychrotolerant gram-negative bacteria introduced as post-pasteurization contamination (PPC) are a major cause of spoilage and reduced shelf life of high-temperature, short-time pasteurized fluid milk. To provide improved tools to (1) predict pasteurized fluid milk shelf life as influenced by PPC and (2) assess the effectiveness of different potential interventions that could reduce spoilage due to PPC, we developed a Monte Carlo simulation model that predicts fluid milk spoilage due to psychrotolerant gram-negative bacteria introduced as PPC. As a first step, 17 gram-negative bacterial isolates frequently associated with fluid milk spoilage were selected and used to generate growth data in skim milk broth at 6°C. The resulting growth parameters, frequency of isolation for the 17 different isolates, and initial concentration of bacteria in milk with PPC, were used to develop a Monte Carlo model to predict bacterial number at different days of shelf life based on storage temperature of milk. This model was then validated with data from d 7 and 10 of shelf life, collected from commercial operations. The validated model predicted that the parameters (1) maximum growth rate and (2) storage temperature had the greatest influence on the percentage of containers exceeding 20,000 cfu/mL standard plate count on d 7 and 10 (i.e., spoiling due to PPC), which indicates that accurate data on maximum growth rate and storage temperature are important for accurate predictions. In addition to allowing for prediction of fluid milk shelf life, the model allows for simulation of "what-if" scenarios, which allowed us to predict the effectiveness of different interventions to reduce overall fluid milk spoilage due to PPC through a set of proof-of-concept scenario (e.g., frequency of PPC in containers reduced from 100% to 10%; limiting distribution temperature to a maximum of 6°C). Combined with other models, such as previous models on fluid milk spoilage due to psychrotolerant spore-forming bacteria, the data and tools developed here will allow for rational, digitally enabled, fluid milk shelf life prediction and quality enhancement.

Drug Susceptibility Pattern and Associated Factors of Bacteria Isolated from Urinary Tract Infection among Pregnant Women Attending Antenatal Care in Teaching Referral Hospital, Southeast Ethiopia

Background: Urinary tract infection (UTI) is caused by bacteria from the digestive tract which climbs the opening of the urethra and begins to multiply to cause infection. UTI in pregnancy is associated with significant morbidity for both mother and baby. In most developing countries including Ethiopia, screening for UTI in pregnancy is not considered an essential part of Antenatal Care. This study aimed to assess the bacterial profile and antibiotic susceptibility pattern and associated risk factors among pregnant women in Madda Walabu University Goba Referral Hospital, Southeast Ethiopia. Methods: An institution-based cross-sectional study was conducted from June-August, 2020 . A total of 234 pregnant women were enrolled; data were collected using a structured questionnaire by a trained interviewer. Urine samples were collected from all pregnant women and culture on cysteine lysine electrolytes deficiency medium. Data were analyzed using SPSS 20.0. Descriptive statistics were used to explain the study participants with relevant variables. Logistic regression was used for data comparison. P-value &lt;0.05 was accepted as statistically significant. Results: The overall prevalence of UTI was 23.9% among both asymptomatic and symptomatic groups. Of this screened midstream urine samples showed that 16% and 32.2%, had significant bacteriuria in the asymptomatic and symptomatic groups respectively. The prevalence of UTI was significantly associated with previous history of catheterization and urinary tract infection (p&lt;0.05). E. coli was the most frequently isolated organism (42.9%) followed by coagulase-negative Staphylococcus (26.8%), and S. aureus (12.5%). Gram-negative and Gram-positive bacteria accounted for (59%) and (41%) respectively. Gram-negative isolates showed resistance to ampicillin, naldixic acid and trimethoprim/sulfamethoxazole. Also, all Gram-negative bacterial isolates revealed a high level of resistance against trimethoprim/sulfamethoxazole. Conclusion: Significant bacteriuria has been observed from both symptomatic and asymptomatic pregnant women. The majority of the isolates were resistant to commonly prescribed antibiotics. This calls for an early screening of all pregnant women for UTI and those found to be infected need treatment with an appropriate drug to avoid complications.

Phenotypic characterization of carbapenem non-susceptible gram-negative bacilli isolated from clinical specimens.

BackgroundMultidrug resistant, extremely drug-resistant, pan-drug resistant, carbapenem-resistant, and carbapenemase-producing gram-negative bacteria are becoming more common in health care settings and are posing a growing threat to public health.ObjectiveThe study was aimed to detect and phenotypically characterize carbapenem no- susceptible gram-negative bacilli at the Ethiopian Public Health Institute.Materials and methodsA prospective cross-sectional study was conducted from June 30, 2019, to May 30, 2020, at the national reference laboratory of the Ethiopian Public Health Institute. Clinical samples were collected, inoculated, and incubated for each sample in accordance with standard protocol. Antimicrobial susceptibility testing was conducted using Kirby-Bauer disk diffusion method. Identification was done using the traditional biochemical method. Multidrug-resistant and extensively drug-resistant isolates were classified using a standardized definition established by the European Centre for Disease Prevention and Control and the United States Centers for Disease Prevention and Control. Gram-negative organisms with reduced susceptibility to carbapenem antibiotics were considered candidate carbapenemase producers and subjected to modified carbapenem inactivation and simplified carbapenem inactivation methods. Meropenem with EDTA was used to differentiate metallo-β-lactamase (MBL) from serine carbapenemase. Meropenem (MRP)/meropenem + phenylboronic acid (MBO) were used to differentiate Klebsiella pneumoniae carbapenemase (KPC) from other serine carbapenemase producing gram-negative organisms.ResultsA total of 1,337 clinical specimens were analyzed, of which 429 gram-negative bacterial isolates were recovered. Out of 429 isolates, 319, 74, and 36 were Enterobacterales, Acinetobacter species, and Pseudomonas aeruginosa respectively. In our study, the prevalence of multidrug-resistant, extensively drug-resistant, carbapenemase-producing, and carbapenem nonsusceptible gram-negative bacilli were 45.2%, 7.7%, 5.4%, and 15.4% respectively. Out of 429 isolates, 66 demonstrated reduced susceptibility to the antibiotics meropenem and imipenem. These isolates were tested for carbapenemase production of which 34.8% (23/66) were carbapenemase producers. Out of 23 carbapenemase positive gram-negative bacteria, ten (10) and thirteen (13) were metallo-beta-lactamase and serine carbapenemase respectively. Three of 13 serine carbapenemase positive organisms were Klebsiella pneumoniae carbapenemase.ConclusionThis study revealed an alarming level of antimicrobial resistance (AMR), with a high prevalence of multidrug-resistant (MDR) and extremely drug-resistant, carbapenemase-producing gram-negative bacteria, particularly among intensive care unit patients at the health facility level. These findings point to a scenario in which clinical management of infected patients becomes increasingly difficult and necessitates the use of “last-resort” antimicrobials likely exacerbating the magnitude of the global AMR crisis. This mandates robust AMR monitoring and an infection prevention and control program.

Prevalence of antimicrobial resistance in Gram-negative clinical isolates from a major secondary hospital in Kuwait: a retrospective descriptive study.

Building an antimicrobial resistance (AMR) surveillance system in a country requires analysis of available data on AMR in clinical isolates. This study's objective was to determine the AMR prevalence of Gram-negative bacterial (GNB) isolates cultured from clinical specimens at a major general hospital in Kuwait. A retrospective descriptive study was conducted on AMR profiles of GNB clinical isolates (n=5290) between January and December 2018. Data were extracted from the laboratory information system in the hospital. The GNB organisms (i.e., Escherichia coli, Klebsiella pneumoniae, Pseudomonas aeruginosa, and Acinetobacter baumannii) were isolated from five main locations at the hospital (i.e., intensive care units [ICUs], surgical wards, medical wards, pediatric wards, and outpatient polyclinics). Overall, high AMR prevalence (>50%) against ampicillin, cefuroxime, cefotaxime, ceftazidime, ciprofloxacin, and trimethoprim/sulfamethoxazole, were observed across the GNB organisms. However, low resistance prevalence (<10%) were observed against amikacin, ertapenem, meropenem, and piperacillin/tazobactam. In general, AMR prevalence in E. coli isolates from ICU, medical and surgical wards was significantly (p<0.05) higher compared to other locations, whereas, AMR prevalence in P. aeruginosa isolates from pediatric ward was higher (p<0.05) compared to other locations. The overall multidrug resistance (MDR) prevalence was 38.7% (95% CI: 37.4-40.0). The highest MDR prevalence was among E. coli isolates from respiratory specimens (48%); wounds, bones, or other tissues (47.7%); and body fluids (47.1%). Similarly, MDR prevalence in K. pneumoniae, P. aeruginosa, and A. baumannii isolated from respiratory specimens was significantly (p<0.05) higher compared to other specimen types. The most frequent MDR phenotypes in the four GNB organisms and across the different specimen types included three antimicrobial drug classes: penicillins, cephalosporins, and fluroquinolones. Our findings demonstrate high AMR prevalence among common Gram-negative bacteria at this major hospital. Monitoring data on antimicrobial susceptibility of common bacterial organisms is critical for assessing trends in AMR at hospitals and for informing policy decisions.

Antimicrobial susceptibility patterns of bacterial isolates from routine clinical specimens of a tertiary hospital in Bangladesh

Background and objectives: To prevent the emergence and spreading of antimicrobial resistance, especially multidrug resistance in pathogenic bacteria, the selection of appropriate antibiotics is a prerequisite for the effective treatment of infection.This study aimed to analyze the prevalence and antimicrobial resistance patterns of bacterial isolates from various clinical samples in a tertiary care hospital. Methods: This study was conducted at a teaching hospital of Dhaka city, Bangladesh from January 2020 to March 2021. The results of culture and antimicrobial susceptibility of bacterial isolates from various clinical samples were collected and analysed. Identification of bacteria and antimicrobial susceptibility test were performed according to the standard methods. Results: A total of 1277 bacterial isolates was analyzed. Of them, 1072 (83.95%) were Gram-negative, and 205 (16.05%) were Gram-positive bacteria. Among the isolates, Escherichia coli (n=576), Enterobacter spp. (n=150), Klebsiella spp. (n=140), and Staphylococcus aureus (n=117) were predominant.The Enterobacteriaceae showed higher resistance to cephradine (94.3%) and cefuroxime (76.7%), whereas least resistant to imipenem (10.1%) and meropenem (14.8%). Pseudomonas spp. was highly resistant to ceftriaxone (80.2%), and colistin (70.8%), whereas least resistant to piperacillin-tazobactam (15.1%). Colistin was the most effective agent (resistance 6.7%) against Acinetobacter spp. Linezolid (resistance 1%) and vancomycin (resistance 2%) were highly effective against Gram-positive bacteria. Among the Staphylococcus aureus, 95.7% were methicillin-resistant (MRSA). A total of 889 (69.6%) bacterial isolates were identified as multidrug resistant. Multidrug resistance was more prevalent in Gram-positive isolates (79.5%) than that of Gram-negative bacteria (67.7%). Furthermore, 7.5% of Gram-negative bacterial isolates were resistant to all seven classes of antibiotics tested. Conclusions: This study revealed presence of high rate of resistance to several antimicrobial agents in bacteria isolated from various clinical samples.The findings would help healthcare professionals to select appropriate antibiotics for the effective treatment of infections and to develop antibiotic stewardship protocol. IMC J Med Sci 2022; 16(1): 005 *Correspondence: M. Mahboob Hossain, Microbiology Program, Department of Mathematics and Natural Sciences (MNS), BRAC University, 66 Mohakhali, Dhaka 1212. E-mail: mmhossain@bracu.ac.bd

Occurrence of extended-spectrum β-lactamase, AmpC, and carbapenemase-producing genes in gram-negative bacterial isolates from human immunodeficiency virus infected patients

BackgroundProgressive decline of immune response in HIV patients makes them susceptible to frequent bacterial infections. High usage of antibiotics influences the emergence of multidrug-resistant bacteria and worsens the clinical outcomes. In this study, the occurrence of drug-resistant genes in Gram-negative bacterial isolates from HIV patients in South India was analyzed. MethodsA total of 173 Gram-negative bacterial (GNB) isolates from HIV patients were screened for antibiotic susceptibility profile using the Kirby-Bauer diskdiffusion method. Positivity of drug-resistant genes was analyzed using polymerase chain reaction method. ResultsIn this study, 72.8% of bacterial isolates were obtained from urine specimens, and Escherichia coli (47.4%) was the predominantly isolated bacterium. Overall, 87.3% and 83.2% of GNB were resistant to 3rd generation cephalosporin antibiotics such as cefotaxime and ceftazidime, respectively, 56.6% were resistant to cephamycin (cefoxitin) and 43% to carbapenem (imipenem) antibiotics. Extended-spectrum β-lactamases (ESBL) production was noted among 79.5% of GNB isolates, followed by AmpC (57.1%) and Metallo β-lactamases (37.3%). Molecular analysis revealed that ESBL genes such as blaTEM (94.1%), blaCTX-M (89.2%), and blaSHV (24.2%) were detected at higher levels among GNB isolates. Carbapenemase-producing genes such as blaOXA-48 (20%), blaOXA-23 (2.6%), and both blaOXA-23 and blaOXA-51 like genes (2.6%) and AmpC producing genes such as blaCIT (26.7%), blaDHA (3.6%), and blaACC (1.8%) were detected at low-level. ConclusionsThis study concludes that ESBL producing genes are detected at high level among gram-negative bacterial isolates from HIV patients in South India.

Phenotypic Methods for the Detection of Metallo-Beta-Lactamase Production by Gram-negative Bacterial Isolates from Hospitalized Patients in A Tertiary Care Hospital in India

Drug-resistant bacteria are a global health concern owing to the high morbidity and mortality they can cause, especially in countries such as India. Gram-negative bacteria, including Enterobacteriaceae, Pseudomonas, and Acinetobacter, are primarily responsible for expanding the scope of drug resistance. These antibiotic-resistant pathogens are particularly associated with serious infections in hospitals. The production of carbapenemase by gram-negative bacteria appears to be the major reason for their resistance to carbapenems. The study was a prospective study done from March 2018 to December 2020. All the carbapenem-resistant isolates from various clinical samples were further tested for the production of carbapenemases/metallo-beta-lactamases production by various phenotypic tests like carbaNp, Imipenem–EDTA combined disc synergy test, Double-disc synergy test and E-test methods. Of all carbapenem-resistant gram-negative bacteria isolated from patients in a hospital in India, 237 (88.1%) carbapenemase producers were identified, among which 217 (91.5%) were metallo-beta-lactamase (MBL) producers. Therefore, the detection of MBL producers is important for preventing their infectious spread. The present study revealed that most MBL producers were isolated from patients of 0–9 to years of age (63.9%). The double-disc synergy test (DDST) and E-test MBL strips were more sensitive than the combined disc test in detecting MBLs. Because the DDST was the simplest and most effective method, it can be used for the routine laboratory screening of MBL producers in hospitals.

Bacterial Assessment of Buffalo Meat in Kathmandu Valley

Meat is highly nutritious and mostly consumed food. Usually, meat gets contaminated because of improper handling, open transportation practice, unhygienic cutting tools, and can cause various food–borne illnesses. This study was focused to determine bacterial load and occurrences of potential pathogenic bacteria in the raw buffalo meat sold in Kathmandu valley. Altogether 40 raw buffalo meat samples were collected and assessed. In addition, antibiotic susceptibility of gram-negative bacterial isolates was tested. The total plate count (TPC), and coliform count (CC) in the samples were found comparatively highest from the Asan area of Kathmandu with 9.6±0.02 log CFU/gm and 8.9±0.00 log CFU/gm respectively. Altogether 10 genera of gram-negative bacteria were identified based on the various biochemical tests. The isolated gram-negative bacteria included Proteus spp (39%, 21/54), Pseudomonas (19%, 10/54) Citrobacter spp. (9%, 5/54), E. coli (7%, 4/54), Serratia marcescens (5%, 3/54), Salmonella spp. (9%, 5/54), Enterobacter spp. (2%,1/54), Morganella morganeii (2%,1/54), Klebsiella (2%, 1/54), Yersinia enterocolitica (6%,3/54). The antibiotic susceptibility tests for the isolates was carried against six different antibiotics including-Ofloxacin (OF), Ciprofloxacin (CIP), Nalidixic Acid (NA), Nitrofurantoin (NIT 300), Amoxycillin (AMX) and Chloramphenicol (C). Out of total 54 isolates, 36 were found to be multidrug- resistant. The results of this study clearly revealed buffalo meat sold in Kathmandu valley was contaminated with potential bacterial pathogens which may cause various food- borne illnesses. The occurrence of multi - drug resistant bacteria in the meat samples is alarming threat to public health. Adequate measures to protect bacterial contamination in buffalo meat is highly recommended.

Black-and-White Ruffed Lemur (Varecia variegata) in Captivity: Analysis of the Oral Microbiota in a One Health Perspective.

Simple SummaryThe emergence of multidrug-resistant bacteria is a serious public health problem. Wild animals are known to be sources of multidrug-resistant bacteria as well as infectious diseases, some of them transmissible to humans. Due to many factors, such as the destruction of natural habitats or climate change, contact between wild animal species and humans is increasing. Thus, it is particularly important that studies be carried out in wildlife species to assess the possible existence of multidrug-resistant bacteria. In this case, the chosen species was the black-and-white ruffed lemur belonging to a zoo. Through a sample of the oral cavity, it was possible to know that half of the bacteria isolated in this group of animals were resistant to, at least, one antibiotic and one of them is resistant to an antibiotic for exclusive use in a hospital environment. Captive wild mammals can be a source of multidrug-resistant bacteria and studies such as this could contribute to the development of strategies to prevent the spread of this public health program.This study aimed to characterize the susceptibility profile to antibiotics and biofilm formation of Gram-negative bacterial isolates obtained from the oral cavity of the black-and-white ruffed lemur (Varecia variegata). From eight individuals from a zoo located in Portugal, samples of the oral microbiota were collected with sterile swabs and then placed in closed tubes with a transport medium. Culture was carried out for media of Gram-negative bacteria. Twenty-two isolates were obtained and subjected to susceptibility tests to twenty-five antimicrobial agents belonging to seven different classes. All tested isolates demonstrated resistance to, at least, one antibiotic, and it was possible to observe multidrug resistance in 11 of the 22 isolates (50%). It should be noted that an isolate showed phenotypic resistance to imipenem, an antibiotic for exclusive use in a hospital environment. All the isolates showed an increasing ability of biofilm formation over time. The obtained results show that wild mammals in captivity could be reservoirs and potential sources of multi-resistant pathogens. In view of this fact and considering the One Health concept, it will be advisable to establish local monitoring programs worldwide that benefit and protect human, animal and environmental health.

Risk factors and mortality associated with multi-drug-resistant Gram-negative bacterial infection in adult patients following abdominal surgery

Multi-drug-resistant (MDR) Gram-negative bacterial (GNB) infection remains a significant cause of morbidity and mortality among surgical patients. The objective of this study was to recognize the risk factors for MDR GNB infection in patients following abdominal surgery, and determine the predictors independently associated with death. From 2010 to 2017, a retrospective cohort study was conducted among patients with abdominal surgery admitted to the surgical intensive care unit (ICU). Patients with GNB infection were included for analyses. In total, 364 patients experienced GNB infection following abdominal surgery. Of these, 117 (32.1%) were MDR GNB infection. Of 133 MDR GNB isolates, the most common isolate was Escherichia coli (45.1%). Patients with MDR GNB infection had significantly longer ventilator-days and hospital stay, as well as higher 30-day and in-hospital mortality compared with non-MDR GNB patients. Multi-variable analysis showed that longer length of pre-ICU stay, surgical re-exploration, receipt of group 2 carbapenems (e.g. imipenem, meropenem and doripenem) and fluoroquinolones, and higher total bilirubin were independent risk factors for the acquisition of MDR GNB infection. Predictors for 30-day mortality among patients with MDR GNB infection were chronic kidney disease, receipt of group 2 carbapenems and inappropriate empirical antimicrobial therapy. This study provides important information about the risk factors for MDR GNB infection and 30-day mortality among patients following abdominal surgery.

COVID-19 and Antimicrobial Resistance: Data from the Greek Electronic System for the Surveillance of Antimicrobial Resistance-WHONET-Greece (January 2018-March 2021).

Changes in hospitals’ daily practice due to COVID-19 pandemic may have an impact on antimicrobial resistance (AMR). We aimed to assess this possible impact as captured by the Greek Electronic System for the Surveillance of Antimicrobial Resistance (WHONET-Greece). Routine susceptibility data of 17,837 Gram-negative and Gram-positive bacterial isolates from blood and respiratory specimens of hospitalized patients in nine COVID-19 tertiary hospitals were used in order to identify potential differences in AMR trends in the last three years, divided into two periods, January 2018–March 2020 and April 2020–March 2021. Interrupted time-series analysis was used to evaluate differences in the trends of non-susceptibility before and after the changes due to COVID-19. We found significant differences in the slope of non-susceptibility trends of Acinetobacter baumannii blood and respiratory isolates to amikacin, tigecycline and colistin; of Klebsiella pneumoniae blood and respiratory isolates to meropenem and tigecycline; and of Pseudomonas aeruginosa respiratory isolates to imipenem, meropenem and levofloxacin. Additionally, we found significant differences in the slope of non-susceptibility trends of Staphylococcus aureus isolates to oxacillin and of Enterococcus faecium isolates to glycopeptides. Assessing in this early stage, through surveillance of routine laboratory data, the way a new global threat like COVID-19 could affect an already ongoing pandemic like AMR provides useful information for prompt action.

Subinhibitory concentration stress of colistin enhanced PhoPQ expression in Escherichia coli harboring mcr‐1

The increased and inappropriate use of colistin led to the emergence of its resistance among Gram-negative bacterial isolates and the most common mechanism of colistin resistance in Gram-negative bacteria is the modification of the lipopolysaccharide mediated by two-component regulatory systems, PhoPQ and PmrAB. The aim of the present study was to investigate the transcriptional expression of the PhoPQ system against colistin stress in clinical isolates of Escherichia coli with colistin-resistant phenotype. Six colistin-resistant E. coli isolates were obtained from Silchar Medical College and Hospital, Silchar that were of clinical origin and received for routine culture and sensitivity testing. Screening for colistin resistance was done by broth microdilution method and further screened for the presence of the different types of plasmid-mediated colistin resistance mcr genes namely, mcr-1 to mcr-10 by polymerase chain reaction (PCR). The screened positive isolates were subjected to PCR assay targeting phoP and phoQ genes and their expression was measured by quantitative real-time PCR. The results of this study revealed that two E. coli isolates (TS2 and TS4) were found to carry the mcr-1 gene. PhoP and PhoQ gene amplification was observed in all the isolates. Transcriptional analysis showed that the isolates harboring the mcr-1 gene showed an enhanced level of expression in the PhoP, PhoQ genes in the presence of a subinhibitory concentration of colistin whereas no significant expression was observed for the isolates which were devoid of the mcr gene. This study demonstrates the involvement of mcr-1 in the PhoPQ system in clinical isolates of colistin-resistant E. coli which will help in designing a molecular marker for detecting colistin-resistant E. coli and contribute to the assessment of resistance burden and infection control strategy.

OXA-48 Carbapenemase-Encoding Transferable Plasmids of Klebsiella pneumoniae Recovered from Egyptian Patients Suffering from Complicated Urinary Tract Infections.

Simple SummaryGram-negative bacteria are common causes of urinary tract infections (UTIs), some of which can resist treatment by antibiotics, including carbapenems, which are last resort treatment options. This study aimed to report the resistance of some Gram-negative bacteria causing complicated UTIs to carbapenems at two important hospitals in Cairo, Egypt, and to determine the possible transfer of this resistance to other bacterial species. The collected bacteria were tested for antibiotic resistance and detection of the genes responsible for this resistance. A total of 256 Gram-negative bacterial clinical isolates were collected, 65 (25.4%) of which showed carbapenem resistance. The detected carbapenem resistance genes were blaOXA-48, blaVIM, blaKPC, and blaNDM genes. The blaOXA-48, among other genes, was successfully transferred to a previously susceptible bacteria, making it resistant. The study concluded that the rate of carbapenem resistance among Gram-negative bacteria causing UTIs in Cairo, Egypt is relatively high and can be transferred among bacterial hosts.Gram-negative bacteria are common causes of urinary tract infections (UTIs). Such pathogens can acquire genes encoding multiple mechanisms of antimicrobial resistance, including carbapenem resistance. The aim of this study was to detect the carbapenemase-producing ability of some Gram-negative bacterial isolates from urine specimens of patients suffering from complicated UTIs at two vital tertiary care hospitals in Cairo, Egypt; to determine the prevalence of carbapenemase genes among plasmid-bearing isolates; and explore the possibility of horizontal gene transfer to other bacterial species. The collected isolates were subjected to antimicrobial susceptibility testing, phenotypic analysis of carbapenemase production, and molecular detection of plasmid-borne carbapenemase genes, then the extracted plasmids were transformed into competent E. coli DH5α. A total of 256 Gram-negative bacterial clinical isolates were collected, 65 (25.4%) isolates showed carbapenem resistance of which 36 (55.4%) were carbapenemase-producers, and of these 31 (47.7%) harbored plasmids. The extracted plasmids were used as templates for PCR amplification of blaKPC, blaNDM, blaVIM, blaOXA-48, and blaIMP carbapenemase genes. The blaOXA-48 gene was detected in 24 (77.4%) of the tested isolates while blaVIM gene was detected in 8 (25.8%), both blaKPC and blaNDM genes were co-present in 1 (3.2%) isolate. Plasmids carrying the blaOXA-48 gene from 4 K. pneumoniae clinical isolates were successfully transformed into competent E. coli DH5α. The transformants were carbapenemase-producers and acquired resistance to some of the tested antimicrobial agents as compared to untransformed E. coli DH5α. The study concluded that the rate of carbapenem resistance among Gram-negative bacterial uropathogens in Cairo, Egypt is relatively high and can be transferred horizontally to other bacterial host(s).

Antimicrobial susceptibility patterns of respiratory Gram-negative bacterial isolates from COVID-19 patients in Switzerland

BackgroundBacterial superinfections associated with COVID-19 are common in ventilated ICU patients and impact morbidity and lethality. However, the contribution of antimicrobial resistance to the manifestation of bacterial infections in these patients has yet to be elucidated.MethodsWe collected 70 Gram-negative bacterial strains, isolated from the lower respiratory tract of ventilated COVID-19 patients in Zurich, Switzerland between March and May 2020. Species identification was performed using MALDI-TOF; antibiotic susceptibility profiles were determined by EUCAST disk diffusion and CLSI broth microdilution assays. Selected Pseudomonas aeruginosa isolates were analyzed by whole-genome sequencing.ResultsPseudomonas aeruginosa (46%) and Enterobacterales (36%) comprised the two largest etiologic groups. Drug resistance in P. aeruginosa isolates was high for piperacillin/tazobactam (65.6%), cefepime (56.3%), ceftazidime (46.9%) and meropenem (50.0%). Enterobacterales isolates showed slightly lower levels of resistance to piperacillin/tazobactam (32%), ceftriaxone (32%), and ceftazidime (36%). All P. aeruginosa isolates and 96% of Enterobacterales isolates were susceptible to aminoglycosides, with apramycin found to provide best-in-class coverage. Genotypic analysis of consecutive P. aeruginosa isolates in one patient revealed a frameshift mutation in the transcriptional regulator nalC that coincided with a phenotypic shift in susceptibility to β-lactams and quinolones.ConclusionsConsiderable levels of antimicrobial resistance may have contributed to the manifestation of bacterial superinfections in ventilated COVID-19 patients, and may in some cases mandate consecutive adaptation of antibiotic therapy. High susceptibility to amikacin and apramycin suggests that aminoglycosides may remain an effective second-line treatment of ventilator-associated bacterial pneumonia, provided efficacious drug exposure in lungs can be achieved.