API 20E Research Articles

Antimicrobial susceptibility pattern of ceftazidime-avibactam against Eschericia Coli.

Objectives: To identify the susceptibility of Ceftazidime-avibactam against Escherichia coli and their frequency in different clinical specimens. To correlate the susceptibility pattern of ceftazidime-avibactam against Escherichia coli in different age groups and gender. To evaluate susceptibility of drugs among Escherichia coli as compared to traditional antibiotics. Study Design: Cross-sectional Study. Setting: PNS Shifa Hospital Karachi. Period: September 2019 to May 2020. Material & Methods: The sample size was 150. The age group 10-50 years. Ethical permission was taken. Informed consent was taken. The specimens were inoculated on Blood and MacConkey’s agar culture plates. The culture plate was inoculated at 37℃ in incubator for 24 to 48 hours. Identification of Enterobacteriaceae was done by colony morphology, gram staining, biochemical tests, and API 20E. After identification, the susceptibility profile of conventional antibiotics was identified. Mueller Hinton agar was used to check the antibiotic susceptibility of Ceftazidime-avibactam by disk diffusion method. Results: Ceftazidime-avibactam shows sensitivity against 82.7% of the isolates while 17.3 % isolates were resistant. The minimum inhibitory concentration of microorganisms is measured by E-test method that revealed most of the isolate show sensitivity less than 1 µg/mL concentration whereas few of them showed susceptibility on 2-8 µg/mL concentration. We compared our results with other classes of antibiotics used commonly This gives an insight for improved treatment methodologies for future prospects particularly diseases caused by members of Enterobacteriaceae. Conclusion: It was concluded that Ceftazidime-avibactam is a novel drug combination that shows high sensitivity against Escherichia coli.

Isolation and determination of Vibrio spp. pathogen from Sciaenops ocellatus suffering from hemorrhagic disease under cage culture in Vietnam

This study was carried out to isolate and determine the Vibrio spp. from the Red drum fish (Sciaenops ocellatus) suffering from the hemorrhagic disease in Vietnam. In this study, 18 strains of Vibrio bacteria were identified from 27 samples of Red drum fish. The isolated bacterial strains were identified with the 16S rRNA sequencing method and checked for morphological, physiological, and biochemical characteristics by using the API 20E KIT. Results of the study revealed the presence of twelve strains of V. alginolyticus, three strains of V. fluvialis, and three strains of V. orientalis. All Vibrio strains have gene similarities with those on the Genbank ranging from 98.05 to 100%. The biochemical characteristics of these 18 isolates were similar and these are susceptible to tetracycline and doxycycline and entirely resistant to ampicillin, amoxicillin, and erythromycin.

PM 7/20 (3) Erwinia amylovora

PM 7/20 (3) <i>Erwinia amylovora</i>

Multidrug-Resistant and Extended-Spectrum β-Lactamase (ESBL) - Producing Enterobacterales Isolated from Carriage Samples among HIV Infected Women in Yaoundé, Cameroon.

The exacerbation of antimicrobial resistance (AMR) is a major public health threat worldwide. In sub-Saharan Africa, there is a scarcity of data regarding multidrug-resistant (resistance to at least one antibiotic of three or more families of antibiotics) as well as extended spectrum β-lactamase-producing Enterobacterales (ESBL-PE), isolated among clinical and asymptomatically healthy patients, especially in women living with HIV (WLHIV) despite their immunocompromised status. The overarching aim of this study was set to determine the prevalence and characterize genotypically multi-drug resistant Enterobacterales (MDR-E) and ESBL- PE isolated from vaginal swabs of WLHIV attending the Yaoundé Central Hospital, Yaoundé, Cameroon. A cross-sectional study was conducted among WLHIV during a four-month periods from 1 February to 31 May 2021. A total of 175 WLHIV, of childbearing age and under antiretroviral treatment were contacted. One hundred and twenty participants (120) were recruited and vaginal swabs were collected from them. After culture on Eosine-Methylen Blue (EMB) agar, the identification of Enterobacterales was performed using API 20E kit. A double-screening of ESBL-PE was performed using a combined disc diffusion method and ROSCO Diagnostica kits. An antibiotic susceptibility test was carried out by disc diffusion as per the Kirby-Bauer method and the β-lactamase resistance genes, blaCTX-M, blaCTX-M-group1-2-9, blaTEM were molecularly characterized using a conventional Polymerase Chain Reaction (PCR). Overall, 30.83% (37/120) of the included WLHIV were colonized with Enterobacterales and the prevalence of vaginal carriage of MDR Enterobacterales among them was 62.16% (23/37). Among MDR-E isolates, the most prevalent species were E. coli (56.0%; 14/25) and K. pneumoniae (20.0%; 5/25). High rates of resistance to trimethoprim-sulfamethoxazole (96.0%; 24/25), amoxicillin-clavulanic acid (88.0%; 22/25) and gentamicin (72%; 18/25) were observed. The resistance mechanisms detected among these isolates were ESBL (48.0%; 12/25), ESBL+ porin loss (8.0%; 2/25), ESBL+AmpC (24%; 6/25), with blaCTX-M, blaCTX-M-group-1,2,9 being identified at 48.0% (12/25) for each of them and blaTEM at 72.0% (18/25). Our findings confirm the high-prevalence of MDR as well as ESBL-PE isolated in WLHIV, and suggest that a real time monitoring system of antimicrobial resistant bacteria coupled with the reinforcement of infection prevention control (IPC) strategies are needed to sustainably contain these life-threatening pathogens especially in the most vulnerable populations.

The phenotypic, genotypic and pathogenicity comparison ofEdwardsiella ictaluriIndonesian local isolates causing enteric septicemia of catfish

This study aimed to compare the phenotypic and genotypic characteristics, and pathogenicity of Edwardsiella ictaluri derived from several catfish farming in Indonesia. The isolated bacteria were from West Java, East Java, South Sumatra, Jambi, Lampung, Riau and South Kalimantan. The phenotypic analysis based on conventional biochemical characteristics, API 20E and enzyme activity by APIzym showed that the E. ictaluri from various locations in Indonesia had the different growth patterns, ability to hydrolyze sugar (saccharose and amylase), hydrolyzed gelatin, lysed blood agar media and leucine and valine arylamidase enzyme activity. The sensitivity antibiotic testing showed most of the tested isolates were multi-drug resistance to five antimicrobial classes (macrolides, lincosamides, rifamycin, penicillins and quinolones). The phylogenetic analysis with sequencing 16S rRNA had 99.56%–100% similarity to E. ictaluri. The genotypic analysis of the similarity strains with Genetic Fingerprinting Repetitive sequence-mediated PCR (Rep-PCR) denoted different genotype profiles of each primer ERIC 1 ERIC II, BOX and GTG5. The protein profiles were analysed using the SDS-PAGE method. The whole-cell isolates of E. ictaluri coded PJbH, P and PBm1G had outer membrane protein (OMP) and riboflavin biosynthesis protein (RibD). The pathogenicity test led to high mortality for 97.78%–100% in the whole-cell treatment, while the low mortality was 6.67% in the ECP treatment. These results concluded that E. ictaluri isolates from various farming in Indonesia had different phenotypic and genotypic variations and protein components of E. ictaluri cells proved to be virulent and played a role in the pathogenesis of E. ictaluri infection in catfish.

Antimicrobial Resistance of Tannin Extract against E. coli Isolates from Sheep.

Plants have been long valuable sources of natural materials that have served to preserve human and animal health; as a result, pharmacological purposes have arisen from the use of plant compounds in most countries, according to a World Health Organization report. The present study aimed to assess the antimicrobial resistance of tannin extract against Escherichia coli (E. coli) isolates in sheep. A total of 100 samples from sheep were used to isolate E. coli and treated with tannin extract (90% purity) to investigate the in vitro effect, as compared to some antibiotics (Clindamycin, Cephalexin, Kanamycin, Tetracycline, and Vancomycin). The bacterial samples were cultured in a selective and differential medium, and Gram staining was used to examine them. The biochemical assays were performed to purify and expose these cultures; moreover, the API 20E system and RapidTM ONE kits were utilized to confirm the bacterial strain. Based on the findings, 50% of the samples showed a positive result for the presence of E. coli. The well diffusion technique was used to investigate the antibacterial activity to confirm the antibacterial action of tannin extract (from pomegranate peel) in different concentrations against E. coli. The highest zone of inhibition for the bacteria ranged from 12±0.5 to 30.3±0.2 at 50% concentrations, proving that tannins extract was significantly effective against E. coli. The presence of E. coli was detected in 50 % of the samples. The well-diffusion technique was used to evaluate the antimicrobial property of tannin extract through various concentrations with the highest zone of inhibition for the bacteria ranging from 12.5 to 30.30.2 at 50%, demonstrating that tannin extract was significantly effective on E. coli.

Antibacterial Activity of Boswellia carterii Aqueous Extract and Its Effect on Phagocytosis in vitro.

Boswellia serrate has been traditionally used for the treatment of several inflammatory diseases, and bacterial resistance to antibiotics has recently increased the use of bioproducts. The present study aimed to assess the antibacterial effect and phagocytic ability of the aqueous extract of the Boswellia serrata in bacteria isolated in nosocomial infections. Boswellia carterii plant was collected and prepared from the aqueous extract in different concentrations. A total of 125 samples were collected from various clinical sources, including urine, sputum, wounds, otitis, and blood, from patients of both genders in different age groups. The results demonstrated that out of the 59 infected samples, urine samples had the highest infection (68%), followed by wounds, sputum, and otitis reported as (60%), (44%), and (40%), respectively. On the other hand, blood samples had the lowest percentage of infection (28%). Microscopic diagnostic results, biochemical tests, API Staph System, API 20E System, and Vitek 2 Compact pointed out that the highest infection rates were related to Staphylococcus aureus (32.20%), Pseudomonas aeruginosa (25.33%), and Escherichia coli (22.03%), while the lowest infection rate was detected in Klebsiella pneumonia (20.33%). The results indicated that aqueous extract of Boswellia carterii had an antibacterial activity for all bacterial isolates, 25 mg/ml of extract gave an inhibition zone of 10.8 mm,10.4 mm, 7mm, and 10mm for S. aureus, E. coli, P aeruginosa, and K. pneumonia, respectively, while 200 mg/ml of extract gave 24 mm, 22 mm, 18.4 mm, and 20 mm, respectively. The results pointed to a significant increase in the phagocytosis rate, with the phagocytosis of blood samples treated with Boswellia carterii extract (79.7%), as compared to control samples (57.75%). As evidenced by the results of this study, the aqueous extract of the Boswellia carterii plant showed antibacterial effects and a positive impact on the phagocytic ratio; nonetheless, it is recommended that further studies be conducted to characterize the compounds of this herb.

Pathogenicity of Aeromonas hydrophila in Cultured African Catfish (Clarias gariepinus)

Aeromonas infections are becoming a serious risk issue in commercial aquaculture, and a wide range of fish and shellfish species has been documented as being vulnerable. Five isolates of Aeromonas hydrophila were identified from African catfish (Clarias gariepinus) cultured in Selangor, West Malaysia in this study. A conventional rapid identification approach (API 20E strip) was used for preliminary identification based on the biochemical properties of the isolated bacteria. Polymerase chain reaction (PCR) with the specific primer 16 rDNA, on the other hand, was used as an accurate and confirmed identification. A pathogenicity test via intramuscular (IM) injection was used to investigate the virulence of A. hydrophila. With a high degree of similarity (98%) to the NCBI or Genbank databases, the isolates were identified as A. hydrophila. The LD50 was calculated using pathogenicity test findings and was found to be 2.1 ´ 106.33 CFU mL−1, while 1 ´ 108 CFU mL−1 in the experimentally injected fish, resulted in 100% mortality. Several organs, including the kidney, liver, and spleen, showed histopathological abnormalities. Those changes mainly include increase in the presence of hemosiderin deposits, congested portal vessels, vacuolated hepatocytes, generalised loss of tubular cells, and oedematous degeneration in the infected organs.

Antimicrobial Resistance, Pathogenic, and Molecular Characterization of Escherichia coli from Diarrheal Patients in South Korea.

Diarrheal diseases due to foodborne Escherichia coli are the leading cause of illness in humans. Here, we performed pathogenic typing, molecular typing, and antimicrobial susceptibility tests on seventy-five isolates of E. coli isolated from stool samples of patients suffering from foodborne diseases in Busan, South Korea. All the isolates were identified as E. coli by both biochemical analysis (API 20E system) and matrix-assisted laser desorption/ionization-time of flight mass spectrometry (MALDI-TOF MS). The bacteria displayed entero-pathogenic E. coli (EPEC) (47.0%), entero-aggregative E. coli (EAEC) (33.3%), entero-toxigenic E. coli (ETEC) (6.6%), ETEC and EPEC (6.6%), EPEC and EAEC (4%), and ETEC and EAEC (2.7%) characteristics. The E. coli isolates were highly resistant to nalidixic acid (44.0%), tetracycline (41.3%), ampicillin (40%), ticarcillin (38.7%), and trimethoprim/sulfamethoxazole (34.7%); however, they were highly susceptible to imipenem (98.6%), cefotetan (98.6%), cefepime (94.6%), and chloramphenicol (94.6%). Although 52 strains (69.3%) showed resistance against at least 1 of the 16 antibiotics tested, 23 strains (30.7%) were susceptible to all the antibiotics. Nine different serotypes (O166, O8, O20, O25, O119, O159, O28ac, O127a, and O18), five genotypes (I to V, random-amplified polymorphic DNA), and four phenotypes (A to D, MALDI-TOF MS) were identified, showing the high level of heterogeneity between the E. coli isolates recovered from diarrheal patients in South Korea.

Effect of cultivation conditions on hemolysin production from clinical isolates of Serratia marcescens

Sixty-four isolates were collected from clinical sources in Baghdad and Mosul city. The identification of S. marcescens confirmed by using API 20E and VITEK-2 systems. Twelve isolates of S. marcescens produced hemolysin, which were detected by two ways, qualitative hemolytic assay and quantitative hemolytic assay. The optical density of hemolysin producing isolates at standard condition (37oC, pH=7, 24h) showed that Serratia marcescens (SmU9) isolate gave the highest absorbance of hemolysin production (0.7) but at (25°C, pH=7,24h) the absorbance of hemolysin production turn into (3.0), compared with the absorbance at 571nm for Control without hemolysin (0.060) and completely hemolysis by Triton x-100 (6.0). However, the optimum optical density of the best hemolysin producing isolate (SmU9) at the optimum cultivation conditions (25°C, pH=9, 24h, 150 rpm, 1% inoculum size in Nutrient broth) was (6.0).

ETIOLOGY OF BACTERIAL WETWOOD OF QUERCUS ROBUR L.

Phytopathogenic bacteria in the plant organism form an integral part of the accompanying microflora, as well as pathogens of pathological processes that do not just weaken the plant, but shortly (with acute pathogenesis) lead to degradation and complete dieback. Notably, bacteriosis is described by typical macroscopic signs of the course of the disease, but the exact aetiology of the pathological process can be reliably established only based on bacteriological analysis with the identification of morphological, cultural, and biochemical properties of isolates. The purpose of this study is to experimentally confirm the direct causes of oak degradation caused by bacterial wetwood in the tree stands under study, as well as to investigate the morphological and biochemical properties of the pathogen. This study employed classical microbiological, phytopathological, and biochemical methods that establish the aetiology of the disease, analyse typical symptoms, include microscopy of the affected parts of the oak, isolation, and identification of the pathogen. The properties of bacterial isolates were figured out according to generally accepted methods and using the API 20E test system and the NEFERMtest24 MikroLaTEST®, ErbaLachema a test system. It was experimentally confirmed that by all macroscopic signs (crown openness, exudate discharge from bark cracks, presence of depressed (sunken) necrotic wet wounds in certain areas of cracks, development of a wet pathological core, presence of epicormic sprouts, etc.) the identified disease is a systemic, vascular-parenchymal bacteriosis, known as bacterial wetwood of common oak. The isolated bacterial isolates were identified by morphological, physiological, and biochemical properties as Lelliottia nimipressuralis – the causative agent of bacterial wetwood of common oak. This suggests that the aetiology of degradation of common oak in Ukraine is closely related to bacteriosis, and the results of this study allow for early phytosanitary diagnostics of the state of common oak in natural conditions based on typical symptomatic signs

ChromagarTM Col-Apse for Detection of Colistin Resistance in Clinical Isolates of Multidrug Resistant Gram Negative Bacilli.

To determine the diagnostic accuracy of CHROMagarTM COL-APSE, for detection of colistin resistance in clinical isolates of multi drug resistant (MDR) gram negative bacilli (GNB). Cross-sectional validation study. Department of Microbiology, Armed Forces Institute of Pathology (AFIP), Rawalpindi, Pakistan from February 2019 to August 2019. Total 96 MDR-GNB in clinical isolates were included. Isolates were identified using gram stain, catalase, oxidase, API 20E and API 20NE. After taking approval from Institutional Ethical Review Committee, colistin susceptibility was determined simultaneously by CHROMagarTM COL-APSE (using 1x105 CFU/ml inoculum) and Broth Micro Dilution (BMD) Minimum Inhibitory Concentration (MIC) method as per CLSI. EUCAST guidelines were followed for interpretation of susceptibility profile. Results were validated with gold standard test, i.e. BMD. Out of 96 MDR clinical isolates, the distribution was K. pneumoniae n=63, E. coli n=18, A. baumannii n=11, C. freundii n=3, and E. cloacae n=1. The sensitivity, specificity, positive predictive value, negative predictive value and diagnostic accuracy of CHROMagarTM COL-APSE for detection of colistin resistance, keeping BMD-MIC method as gold standard, was 97.96%, 97.87%, 97.96%, 97.87% and 97.92%, respectively. CHROMagarTM COL-APSE can be used as screening agar by direct streaking of specimen as well as diagnostic for detection of colistin resistance by the use of standardised inoculum. It can be used as a critical time saving method for colistin resistance detection in absence of expertise or manpower required for BMD as well as the cost required for genetic sequencing to detect MCR genes. Key Words: Multi drug resistant (MDR), Gram negative bacilli (GNB), Colistin resistance, CHROMagarTM COL-APSE.

First Report of Bacterial Canker on Blueberry (Vaccinium corymbosum) Caused by Pseudomonas syringae pv. syringae in Serbia.

During May 2021, necrosis of young twigs and flower buds were observed on two-year-old highbush blueberry plants (Vaccinium corymbosum) cv. Draper, in a 1 hectare orchard in the municipality of Šabac, Serbia. Disease symptoms included reddish-brown to black irregularly shaped cankers developing on the shoot tips that extended downwards along the branches. In some plants, cankers surrounded the stem, causing shoot-tip dieback and necrosis of the buds. Beneath the bark, a distinct margin between diseased and healthy tissue was visible. A few weeks before symptoms development, seven freezing events with temperature from -3°C to -1°C, and five near-freezing temperatures were recorded in this area, leading to the hypothesis that symptoms were associated to the presence of ice nucleating bacteria belonging to Pseudomonas syringae. The observed disease incidence was 80%, while 10% of the plants died. Bacteria were isolated from symptomatic tissue on King's medium B (KB). After 2 to 3 days of incubation at 27°C, predominantly grey-whitish, shiny, round, convex bacterial colonies were observed on agar plates. Ten isolates producing a fluorescent pigment on KB were selected for further characterization by biochemical and molecular tests. The isolates were Gram, oxidase and arginine-dihydrolase negative, levan positive, induced hypersensitive response on tobacco leaves and showed no pectinolytic activity on potato slices. Based on the results of API 20E and API 20NE tests (BioMerieux, France), and the fact that isolates did not utilize tartrate nor had tyrosinase activity, they were preliminarly identified as Pseudomonas syrinage pv. syringae (Braun-Kiewnick and Sands 2001). Additionally, all tested isolates had ice-nucleation activity at -5°C. The syrB gene responsible for syringomycin synthesis, was amplified in all isoaltes with the specific primer pair B1/B2 (Sorensen et al. 1998). The 16S rRNA gene sequences of five selected isolates (GenBank MZ410287 to 91) showed 100% identity to P. s. pv. syringae isolated from Prunus avium in United Kingdom (GenBank CP026568) and France (GenBank LT962480). Sequences of gyrB gene (Sarkar and Guttman 2004) of two selected isolates (GenBank MZ420633 and MZ420634) showed 98,44% identity to the P. s. pv. syrinage strain isolated in France (GenBank LT962480). Pathogenicity of the isolates was confirmed on 2-year-old blueberry plants cv. Draper, by inoculating two plants per isolate. One-cm long wounds were made on branches using a scalpel and 20 µl of bacterial suspension (106 CFU/ml) was infiltrated into the tissue. The cuts were then covered with moist sterile cotton pads and wraped in parafilm for 3 days. Inoculation was also performed on two leaves per plant by needleless syringe infiltration (106 CFU/ml). Sterile distilled water was used as a negative control. Plants were maintained in greenhouse at 27°C day and 15°C night temperature. Three weeks after inoculation, the inoculated branches and leaves developed necrosis, leaves spots and cankers respectively, resembling the natural infection. Symptoms were not observed on the control plants. Bacteria were reisolated from symptomatic tissue and their identity was confirmed by amplifying the syrB gene sequence and additional biochemical tests. This is the first report of bacterial canker of highbush blueberry caused by P. s. pv. syringae in Serbia. In Europe, there was only one report on Pseudomonas spp. causing disease on blueberry leaves in Poland (Kaluzna et al. 2013). Due to market demands and export potential, blueberry production in Serbia has been rapidly increasing. In 2015, highbush blueberry was cultivated on 220 ha, while in 2020 the area increased to 1899 ha. However, under favourable environmental conditions, blueberry production might be severely affected by bacterial canker.

High faecal carriage of extended-spectrum beta-lactamase producing Enterobacteriaceae (ESBL-PE) among hospitalized patients at Sylvanus Olympio Teaching Hospital, Lomé, Togo in 2019

Background: Extended-spectrum beta-lactamase producing Enterobacteriaceae (ESBL-PE) are a global health concern, associated with increased morbidity and mortality. Even in the absence of infections, colonization by these pathogens is still a great threat because of the risk of cross transfer among hospitalized patients. Faecal carriage of ESBL-PE remained poorly documented in Africa. This study aimed to determine faecal carriage rate of ESBL-PE, factors associated with carriage, and antimicrobial susceptibility of the strains among hospitalized patients at Sylvanus Olympio Teaching Hospitals (CHU SO) in Lomé, Togo.&#x0D; Methodology: This was a cross-sectional study of 105 randomly selected hospitalized patients between September and November 2019. Socio-demographic and clinical data as well as rectal swabs were collected after obtaining the consent of the selected participants. Rectal swabs were cultured on selective bromocresol purple (BCP) lactose agar containing 6µg/l ceftazidime, for isolation of Enterobacteriaceae. Identification of each isolate was performed using Uriselect 4 medium and API 20E. Antibiotic susceptibility of the bacterial isolates was performed by the Bauer-Kirby agar disc diffusion test and interpreted according to CASFM-EUCAST recommendations.&#x0D; Results: The faecal carriage rate of ESBL-PE among selected hospitalized patients was 80.9% (85/105). Escherichia coli was the most frequent bacteria 69.5% (73/105), followed by Klebsiella pneumoniae 22.8% (24/105). The antibiotic profile of ESBL producing Escherichia coli showed resistance to amoxycillin+clavulanic acid (72.6%), ticarcillin+clavulanic acid (82.2%), piperacillin+tazobactam (30.1%), cefoxitin (30.1%) ciprofloxacin (84.9%), levofloxacin (76.7%), nalidixic acid (83.6%), chloramphenicol (26.0%), gentamicin (49.3%), sulfamethoxazole-trimethoprim (86.3%), imipenem (5.5%), and ertapenem (21.9%). All (100%) isolates were sensitive to amikacin and fosfomycin. None of the characteristics or risk factors assessed was significantly associated with faecal carriage of ESBL-PE.&#x0D; Conclusion: Faecal carriage rate of ESBL-PE in these hospitalized patients was very high, but no factor was associated with carriage of ESBL-PE among the study population. Implementation of infection control measures, and surveillance are needed to limit the spread of these resistant pathogens within CHU SO healthcare facilities.

Occurrence and antibiotic resistance of Vibrio parahaemolyticus isolated from the Tunisian coastal seawater.

Vibrio parahaemolyticus is a gram-negative bacterium ubiquitous in seawater or estuarine water throughout the world. It is a major cause of seafood gastroenteritis complications. In this study, the presence of V. parahaemolyticus was investigated in 66 seawater samples collected during 2018 from 15 stations spread along the Tunisian coast using selective media including CHROMagar Vibrio media. The results show that only eight samples contained V. parahaemolyticus. However, while Vibrio alginolyticus was detected in all samples; both Vibrio cholerae and Vibrio vulnificus were not found. Nine of the presumed V. parahaemolyticus colonies were purified on tryptic soy agar from eight positive samples then identified by the API 20E biochemical test and confirmed by the presence of a specific target toxR gene. The detection of virulence genes, thermostable direct haemolysin (tdh) and thermostable-related haemolysin (trh), by the polymerase chain reaction (PCR) showed the presence of only two trh-positive isolates. The assessment of antibiotic susceptibility of the V. parahaemolyticus isolated revealed a complete resistance to colistin, amikacin, penicillin and cefotaxime and a total sensitivity to chloramphenicol, nitrofurantoin and sulfamethoxazole-trimethoprim with a multiple antibiotic resistance index (MAR) ranging from 0.4 to 0.5.

The Identification of Multidrug-Resistant Microorganisms including Bergeyella zoohelcum Acquired from the Skin/Prosthetic Interface of Amputees and Their Susceptibility to Medihoney™ and Garlic Extract (Allicin).

Users of prosthetic devices face the accumulation of potentially drug-resistant pathogenic bacteria on the skin/prosthesis interface. In this study, we took surface swabs of the skin/prosthesis interface of eleven disabled athletes to identify microorganisms present. In addition to determining their antimicrobial resistance profile, we assessed their sensitivity to Manuka honey and Garlic extract (allicin). Eleven volunteers were directed to swab the skin at the skin/prosthesis interface. After initial isolation of microorganisms, we employed the following general microbiological methods: Gram stain, Catalase test, Oxidase test, lactose fermenting capability, haemolytic capability, Staphaurex, mannitol fermenting capability, Streptex; API Staph, 20E, Candida, and BBL crystal identification system tests. Once identified, isolates were analysed for their sensitivity to penicillin, erythromycin, ampicillin, vancomycin, ceftazidime, ciprofloxacin, gentamicin, and colistin-sulphate. Isolates were also analysed for their sensitivity to allicin (Garlic Extract (GE)) and Manuka honey (Medihoney™) (MH). Eleven isolates were identified: Bacillus cereus, Staphylococcus haemolyticus, Staphylococcus aureus, Micrococcus luteus, Pseudomonas oryzihabitans, Micrococcus spp., Bacillus subtilis, Group D Streptococcus, Pantoea spp., Enterobacter cloacae, and Bergeyella zoohelcum. All isolates were resistant to 1 unit of penicillin and 10 μg of ampicillin. Bergeyella zoohelcum was observed to have the widest range of resistance with observed resistance against five of the eight antimicrobials employed in this study. This study highlights the prevalence of uncommon drug-resistant microorganisms on the skin within a vulnerable population, highlighting the potential for MH or GE intervention.

COLISTIN AGAR; EVALUATION OF A NOVEL DIAGNOSTIC APPROACH TO DETECTION OF COLISTIN RESISTANCE

Objective: To determine the diagnostic accuracy of Colistin agar for detection of Colistin resistance in clinical isolates of MultiDrug Resistant Gram-Negative Bacilli.&#x0D; Study Design: Cross-sectional validation study.&#x0D; Place and Duration of Study: Department of Microbiology, Armed Forces Institute of Pathology, Rawalpindi Pakistan, from Feb to Aug 2019.&#x0D; Methodology: A total of 100 Multi-Drug Resistant Gram-Negative Bacilli in clinical isolates were included. Isolates were identified using Gram stain, Catalase, Oxidase, API 20E, and API 20NE. After approval from the institutional ethical review committee, Colistin susceptibility was determined simultaneously by Colistin agar and Broth Micro Dilution Minimum Inhibitory Concentration method as per CLSI. For susceptibility criteria, EUCAST guidelines were followed. Results were validated with the gold standard test, i.e., Broth Micro Dilution.&#x0D; Results: Out of 100 Multi-Drug Resistant clinical isolates, the distribution was K. pneumoniae n=60, E.coli n=16, A. baumannii n=11, C. freundii n=8, and E. cloacae n=5. The sensitivity, specificity, positive predictive value, negative predictive value, and diagnostic accuracy of Colistin agar for detection of Colistin resistance, keeping Broth Micro Dilution Minimum Inhibitory Concentration method as the gold standard was 96.67%, 97.14%, 93.55%, 98.55%, and 97%, respectively.&#x0D; Conclusion: Colistin agar has excellent diagnostic accuracy for the detection of colistin resistance with standardized inoculum density. Due to its ease of use, cost-effectiveness, and accurate results, it can be used in lab setups deficient in manpower and advanced equipment for Broth Micro Dilution or genetic sequencing.

Carbapenemase Producing Klebsiella species from Hospitalised Patients at Three Referral Hospitals in Yaounde; Cameroon

Background: Carbapenemases are the most potent beta-lactamases capable of hydrolysing all beta-lactam drugs. Data scarcity on laboratory evidence of carbapenemase producers obstructs infection prevention and effective control measures. This study aimed to detect and phenotypically characterize Klebsiella species resistant to carbapenems. Method: A total of eighty-seven (87) Klebsiella spp. isolates were collected from three Yaounde-based tertiary hospitals over a period of twelve months (February 2019 to January 2020). The isolates were identified on API 20E and antimicrobial susceptibility testing was done according to the Kirby Bauer disc diffusion method. Carbapenem resistance was detected based on reduction in inhibition zone diameter of meropenem less than 28mm and resistance phenotypes determined by the combination disc synergy test. Results: Four Klebsiella species were identified namely: Klebsiella pneumoniae 62/87 (71.3%), K. oxytoca 11/87 (12.6%), K. ozaenae 7/87 (8%), K. rhinoscleromatis 7/87 (8%). Klebsiella pneumoniae was most resistant to carbapenems with a considerably high number of multidrug resistant isolates 39/56 (69.6%). Patients from the 0-9 years age group were most represented 29/87 (33.3%). Carbapenemases belonging to Ambler classes A, B and D were detected as follows: KPC 2/87 (2.3%), MBL 2/87(2.3%), OXA-48 4/87 (4.6%). All four Klebsiella species were producers of KPC carbapenemase 12/87 (13.8%) and Extended Spectrum β-lactamase 68/87 (78.2%) enzymes. Conclusion: These results revealed that more than half of Klebsiella species analysed were multidrug resistant and about a quarter in hospitalized patients at the referral hospitals were mostly KPC carbapenemase producers. This raises concerns over the need to control and confine the spread of these isolates of clinical and epidemiological importance for effective infection prevention and control.

Evidence of the Transmission of Resistance of Enterobacteria to Betalactamines Between Mothers and Children in Brazzaville

Beta-lactams have always been used in many cases as first-line antibiotics in human medicine. The emergence of beta-lactamase-producing strains of Enterobacteriaceae is nowadays a public health problem. In order to demonstrate the transplacental transmission of enterobacteria resistance to beta-lactams between mothers and their children. Seventy-two (72) stool samples were collected from mothers and two-day-old newborns at the Talangaï referral hospital. Isolation of enterobacteria was done on selective media. Identification of beta-lactamase producing strains was performed by biochemical characteristics using API 20E strips. The resistance profile to carbapenems, monobactam and cephalosporins was performed by antibiogram. A total of 50 strains were purified and identified, of which 30 (60%) were isolated from the mother-newborn couple and 20 (40%) from the newborns. Among these strains, we identified in the mother-newborn group: 10 (33%) <i>Enterobacter </i>sp, 10 (33%) <i>Escherichia coli</i>, 6 (20%) <i>Klebsiella</i> oxytoca; 2 (7%) <i>Shigella </i>sp and 2 (7%) <i>Salmonella</i> sp. High resistance to carbapenems and cephalosporins was observed with <i>Shigella</i> sp. The other strains of enterobacteria isolated from newborns only, among which were identified: 7 (35%) <i>Escherichia coli</i>; 5 (25%) Klebsiella oxytoca; 5 (25%) <i>Enterobacter </i>sp, 3 (15%) <i>Shigella</i> sp. newborns without their mothers showed a variable resistance profile to the antibiotics tested. Three resistance phenotypes were observed in the mother-newborn group including: FEP PRL CL, FEP AMP MA CR ATM IMP ETP and FEP AMP CAZ MA CL ETP. The resistance phenotypes observed in the mothers were identical to those found in their respective offspring. These results show that each newborn is born with a rate of resistance acquired at birth, which testifies to a transplacental transmission between mother and child, consequence of the emergence of the resistance of enterobacteria to betalactamines in the Congolese population.

Surveillance of Resistance to Imipenem and Meropenem in Broad-spectrum Beta-lactamase-producing &amp;lt;i&amp;gt;E. coli&amp;lt;/i&amp;gt; Strains Isolated from Urine Samples in Njombe, Cameroon

The emergence and spread of carbapenems resistance <i>Enterobacteriaceae</i> remain a major public health, a real threat as well as a silent tsunami. This phenomenon leading to reduce the therapeutic option and increase the additional cost. The general objective of this study was to determine the frequency of <i>Escherichia coli</i> strains producing broad-spectrum beta lactamases and resistant to carbapenems. We carried out a descriptive, cross sectional and prospective study between August and November 2020 at the Hospital Saint Jean de Malte in Njombe, on a consecutive sample of 249 patients received at the bacteriology unit. Our study population consisted of all patients who came for inpatient or outpatient consultations, were prescribed an cytobacterioligical urine exam, and in whom an <i>E. coli</i> strain was isolated. The identification of <i>E. coli</i> strains was confirmed using the Api 20E mini gallery. Resistance to carbapenems (Meropenem and Imipenem) was defined by determining the Minimal Inhibitory Concentration (MIC) by the microdilution plate method. Of the 249 cytobacterioligical urine exam samples received during our study period, 131 presented a pathogenic germ and <i>E. coli</i> strains were identified in 85 of them. The age of the patients in whom the <i>E. coli</i> strains were identified ranged from 70 to 85 years and the male sex dominated with a frequency of 38.2%. Of these identified <i>E. coli</i> strains, we detected 08 (9.41%) BLSE-producing strains. The resistance rate of the isolated <i>E. coli</i> strains producing BLSE was 75% and 50% for Meropenem and Imipenem respectively. The results of this study underline the urgent need for a regular surveillance system for broad-spectrum antibiotics in our context.