MacConkey Agar Media Research Articles

Staphyloxanthin and Meropenem Effects on Some Capsular Genes Expressions of Klebsiella pneumoniae Isolated from UTIs

Klebsiella pneumoniae has become one of the most common etiologic agents in the context of recurrent UTI. Capsule is the earliest recognized factor determining bacterial pathogenicity. This study aimed to test the effects of staphyloxanthin, meropenem and a combination of them on capsule genes of Klebsiella pneumoniae. Eighty-eight gram negative bacterial isolates were collected from 150 urine specimens between September and November of 2022.after cultivation on MacConkey agar medium. Oxidase (negative) and catalase (positive) tests were performed. In addition, all isolates were found to be non-motile when their motility was further examined. The results presented that 77 (51.3%) could ferment lactose (lactose fermenter), while 30 isolates (20%) of them were confirmed as K. pneumonia. The antibiotics sensitivity test of (30) K. pneumoniae revealed that 28 (93%) of isolates were sensitive to meropenem, and about 20 (67%), 16 (53%), 8 (27%), 7 (23%), 5 (17%) and 4 (13%) were sensitive toward amikacin, ciprofloxacin, trimethoprim, cefepime, tetracycline and Augmentin respectively. S. aureus (producers of staphyloxanthin) isolates were taken; the staphyloxanthin pigment was extracted, purified by Millipore filter (0.22 mm). Conventional PCR results revealed the presence of galF gene (141 bp) in all eight tested isolates (100%). In addition, the wzi gene (177bp) was found in seven isolates (87.5%). According to the quantitative real-time PCR results, most treatments resulted in downregulation of galF and wzi genes expression, whereas some treatments resulted in upregulation.

Antimicrobial resistance pattern of Klebsiella species isolated from various clinical specimens

Objective: The present study was carried out to identify Klebsiella species and their antimicrobial resistance pattern from various clinical specimens. Methods: This cross study was conducted at a tertiary care hospital, Dhaka, Bangladesh from July2020 to June 2021. Written consent was taken from the concerned authority. A total 6739 clinical samples including urine, wound swab/ pus, sputum, blood and endotracheal aspirates were collected and processed for isolation of bacteria by following standard guidelines from adult patients. All the urine, wound swab, blood and endotracheal aspirates were inoculated in both blood agar and MacConkey agar media, for blood samples blood culture was done in BacT/ALERT 3D 60 aerobic bottles irrespective to antibiotics administration. After incubation at 370C aerobically for 24 hours incubated plates were examined. Phenotypic identification of the organisms were done by observing colony morphology, hemolytic properties, pigment production, Gram staining and biochemical tests (oxidase test, catalase test and biochemical reactions after inoculation in TSI, MIU, citrate agar media). The Kirby - Bauer disk diffusion method was used for antimicrobial susceptibility testing of the isolated organism using Mueller-Hinton agar and commercially available antibiotic discs (Oxoid Ltd, UK). The data were analyzed using SPSS software Version-20 (SPSS Inc., Chicago, IL, USA). Categorical variables were expressed as numbers (n) and percentages (%). Results: A total 6739 samples were included in the present study. Of which, 170 were urine, 162 were wound swabs, 71 were sputum, 52 were endotracheal aspirates and 45 were blood samples. Culture positivity among the samples is shown in table I. Conclusion: Most of the Klebsiella species are highly resistant to commonly used antibiotics. This higher resistance is the major cause morbidity and mortality related with infections in hospitalized patients. So, a regular surveillance of antibiotic susceptibility pattern is needed to prevent indiscriminate use of antibiotics. CME J 2024; 3(2); 12-15

Efflux pumps encoding genes (adeA and adeS) in relation to antibiotic resistance pattern in Acinetobacter baumannii strains isolated from Benha university hospital

Background Acinetobacter baumannii become a growing problem in hospitals as a predominant multidrug-resistant (MDR). This resistance capacity is generated by various mechanisms including efflux pumps that can direct antibiotics outwards and prevent antibiotics from affecting the bacteria. Methods Clinical samples were cultured on blood agar and MacConkey agar medium, identified by Gram stain and biochemical reactions, and then identified to the species level by Vitek2 automated system. The adeA and adeS genes among isolated strains were detected by conventional PCR. This study included 50 A. baumannii strains to assess the antibiotic resistance pattern of A. baumannii. Results A. baumannii strains were fully resistant to Piperacillin (100%) and lowest resistance to Imipenem (54%) was observed. Out of 50 isolates, 41 (82%) of A. baumanii isolates had adeA gene and 32 (64%) had adeS gene. The clinical strains that had adeA gene and adeS gene showed significantly higher resistance to Ciprofloxacin (92.7%) and (100%), respectively. Additionally, borderline significant differences were reported regarding Gentamycin (P = 0.065), Ceftazidime (P = 0.08), and Meropenem (P = 0.08), with the resistance being higher in the adeA positive group. Conclusion Efflux pumps genes (adeA, adeS) played a key role in antibiotic resistance of A. baumanii.

Design and Fabrication of Broad-Spectrum Antimicrobial Porous Metallo-Polymeric Microsphere for Water Disinfection.

An expedient and efficient approach is used to synthesize a new class of metallo-polymeric microspheres (MPMs) as antimicrobials to succumb the wide range of bacteria from water. Three types of MPMs, that is, poly[Silver (I)-methacrylate-co-methylmethacrylate] (pAgMA), poly[Copper (II)-methacrylate-co-methyl methacrylate] (pCuMA), and poly[Nickel (II)-methacrylate-co-methylmethacrylate] (pNiMA), are prepared via radical suspension polymerization technique in 3D shape with porous texture. The structural and morphological characterization of the prepared microspheres are examined by analytical techniques. The antimicrobial potentialities of prepared MPMs are investigated at the laboratory scale study, revealing that the MPMs exhibit strong antibacterial activity (≈99.9% killing) against Gram-negative and Gram-positive bacteria [Enterobacter hormaechei (EH), Bacillus megatarium (BM), and Bacillus bataviensis (BB)]. The MacConkey agar medium test reveals that MPMs have substantial biocidal efficacy against broad-spectrum Gram-negative bacteria present in tap water. The MPMs exhibit significant antimicrobial efficacy via contact killing owe to the presence of integrated biocidal metal moiety, which represents that the MPMs are safe for water disinfection.

Isolation and Identification of Vibrio Species from Different Types of Water Sources Along with Their Drug Susceptible Pattern

Abstract Background: Human activities are almost usually the cause of contaminated water supplies and places with large population densities and intensive land usage are particularly prone to pollution of drinking water sources. It is challenging to detoxify polluted water sources. Cholera is an acute form of diarrheal disease that plagued human civilization over the centuries. Vibrio spp., commonly found in aquatic environments, are the most common cause of bacterial gastroenteritis in the world, those may spread to humans through the ingestion of contaminated drinking water or exposure to seawater. Methods: In the present study, a total of 12 samples were collected from four types of water sources for the isolation of Vibrio spp. along with their drug-resistant pattern through conventional, biochemical, and antibiotic disc diffusion methods. Results: The samples were enriched into alkaline peptone water and then inoculated into culture media such as nutrient agar, MacConkey, and TCBs agar medium. After incubation for 24 h at 37°C on TCBS agar, yellow and green colonies were screened out for biochemical identification. Nonsucrose-fermenting Vibrio parahaemolyticus, Vibrio mimicus, and Vibrio vulnificus showed green colonies, and sucrose-fermenting Vibrio cholera and Vibrio alginolyticus showed yellow colonies on TCBS. The antibiotics ERY (15 μg), PCN (10 μg), CEX (30 μg), and VAN (30 μg) were completely resistant while the more effective antibiotics against Vibrio spp. were SXT (25 μg), CHL (30 μg), and GEN (30 μg). However, ciprofloxacin and nitrofurantoin showed moderate efficacy. Conclusion: To expand current knowledge on the occurrence, ecological niche, and persistence of potential human pathogenic Vibrio spp. in aquatic environments, the extensive laboratory experiment is required. To sum up, this kind of information is crucial for bringing attention to the existence of Vibrio spp. and enabling prompt implementation of preventive measures.

Design and Optimization of a yst-PCR to Detect Yersinia enterocolitica in Meat Food.

In this study, a polymerase chain reaction (PCR) directed to the yst chromosomal gene (yst-PCR) was used as a rapid, sensitive, and specific method to detect Yersinia enterocolitica strains belonging to different biotypes in foods; a competitive Internal Amplification Control (cIAC) is also developed. The cIAC had a molecular weight of 417 bp and was detected until a concentration of 0.85 ng/μL. No other strains of other Yersinia species, nor Enterobacteriales order were detected by this PCR. In pure culture, the detection limit (DL) of the yst-PCR was lower for ystA+ strain (10 colony-forming unit [CFU]/mL) than for ystB+ strain (1 × 102 CFU/mL); which was the concentration detected in Y. enterocolitica inoculated minced meat. The proposed protocol included an enrichment step in peptone sorbitol bile (PSB) broth at 25°C for 24 h followed by isolation on Mac Conkey agar and chromogenic medium. An aliquot of the PSB broth homogenate and a loopful from the confluent zone of solid media were collected to perform DNA extraction for yst-PCR, and typical colonies were characterized by biochemical assays. Among 30 non-contaminated food samples, 4 samples were yst-positive and no Y. enterocolitica isolates were obtained. It is suggested that this yst-PCR could be used in the investigation of Y. enterocolitica in foods.

Multidrug-Resistant ESBL-Producing Enterobacteriaceae Associated with Clinical Samples in a Tertiary Care Hospital, Sirajganj

Extended-spectrum β-lactamase (ESBL) microorganisms have been shown to increase multidrug resistance worldwide, which is a great concern. The prevalence of ESBL-producing clinical pathogens and their antimicrobial resistance patterns were identified in 66 isolates from patients in Khwaja Yunus Ali Hospital with several clinical infections cultured on blood agar and MacConkey agar media. The most prevalent bacteria were Escherichia coli (80.3%), which were identified by the BD-Phonex automated identifier machine, followed by Pseudomonas spp. (6%), Klebsiella spp. (12.1%), and others (4.4%). This research was conducted from January 2023 to June 2023. Generally, a large number of antibiotic resistance patterns and ESBL-producing common bacterial isolates were found in this study, where most of the resistant percentage was found in third-generation cephalosporin antibiotics, which increases the public health problem. In this study, the most prevalent bacteria E. coli isolates were mostly resistant to penicillin (100%), ceftazidime, cefotaxime, and cefuroxime (98.4%). Besides, 95.4% resistance was shown against ceftriaxone. The double-disc synergy test was done to determine the presence of ESBL-producing bacterial strains. The most widely ESBL-positive isolate was Escherichia coli (83%). Among the 66 sample strains produced, the ESBL maximum (53.03%) belonged to female patients, while 46.97% belonged to male patients. This study focuses on the prevalence and patterns of clinical pathogens and the antimicrobial susceptibility profiles of ESBL-producing bacterial infections in a tertiary-level health service center in Bangladesh. Generally, a large number of antibiotic resistance patterns and ESBL-producing common bacterial isolates were found in this study, which increases the public health risk. Therefore, to save human life, we ought to be taking appropriate action against the threat.

A Study on the Bacteriological Profile of Ascitic Fluids and Their Antibiotic Susceptibility Pattern in a Tertiary Care Hospital.

Background Ascitic fluid culture remains an essential step in the management of all patients with ascites, regardless of their presenting complaints. Diagnostic paracentesis should not be delayed or prevent timely administration of antibiotics, particularly in unstable patients. Hence, it is an essential part of the surveillance system of every hospital to perform ascitic fluid culture and assess the antibiotic susceptibility patterns of bacterial isolates. In view of this perspective, the present study was conducted at Chengalpattu Medical College Hospital, Tamil Nadu, India. Objective The aim of the study isto determine the bacterial isolatesof ascitic fluid samples and study their antibiotic susceptibility patterns. Materials and methods Ascitic fluids received in the central laboratory at the Department of Microbiology from various departments were included in this study. Preliminary identification of isolates wasperformed by direct Gram staining, acid-fast staining, and motility testing by the hanging drop method. Within one hour of receiving the samples, they were plated onto blood agar and MacConkey agar media and incubated for 18-24 hours at 37°C for isolation. Growth was checked, and species identification was done based on conventional methods. Antibiotic susceptibility testing was performed using the Kirby-Bauer disk diffusion method. Results In this study, a total of 100 ascitic fluid samples were collected, of which only eight (8%) showed growth. Among the eight isolates, six (75%) were Gram-negative bacilli (GNB). Four (66.66%) of the six GNB wereKlebsiella spp., while the remaining two (33.33%) were Escherichia coli. Both Gram-positive cocci wereStaphylococcus aureus. All the GNB isolates were susceptible to meropenem, piperacillin-tazobactam, and ceftriaxone, with varying susceptibilities to other drugs. Both Gram-positive isolates were found to be methicillin-sensitive Staphylococcus aureus. Conclusion GNB were the predominant organisms in cases of ascitic fluid infection, and they showed 100% susceptibility to carbapenem drugs (especially meropenem),piperacillin-tazobactam, and ceftriaxone.All these drugs can be kept in reserve for serious infections. Amikacin and gentamicin showed promising susceptibility. These drugs can be started empirically with patients on admission before performing culture. Drug adjustments may be later made based on culture reports.

Profile of multidrug-resistant bacteria causing urinary tract infections in inpatients and outpatients in Jakarta and Tangerang

Background Urinary tract infections (UTIs) caused by bacteria occupy the second highest rank of common infectious diseases in the world. Empirical use of antibiotics may give rise to multidrug-resistant (MDR) bacteria because of irrational prescription. Choice of antibiotics to treat UTIs is limited because of MDR bacteria. Thus, this study was conducted to investigate the bacterial antibiotic susceptibility patterns in inpatients and outpatients in Jakarta and Tangerang. Methods Bacterial isolates were obtained from midstream urine specimens from 43 inpatients and 43 outpatients with UTIs in Jakarta and Tangerang. Bacteria were isolated on blood and MacConkey agar media using colony count method. Isolate identification and their susceptibility patterns were performed using VITEK2 compact system according to manufacturers’ instructions. Data were analyzed using Chi-square test. Results A total of 89 bacterial isolates consisting of 15 bacterial species were successfully isolated from 86 specimens. Gram-negative bacteria were the most common etiology of UTIs in inpatients and outpatients. MDR bacteria were found in 52 of the 89 isolates. ESBL-producing E. coli was the most common MDR bacteria. ESBL-producing E. coli and other MDR bacteria showed good susceptibility to ertapenem, meropenem, amikacin, and tigecycline. There were no significant differences regarding the MDR bacterial count in inpatients and outpatients (p=0.521). Conclusion E. coli was found to be the most common MDR bacteria causing UTIs in inpatients and outpatients in Jakarta and Tangerang. Higher resistance to many antibiotics was found in MDR bacterial isolates in inpatients compared to outpatients. MDR bacteria in outpatient UTIs were highly resistant to commonly used antibiotics.

Prevalence of multidrug resistance and extended-spectrum beta-lactamase-producing Klebsiella pneumoniae from dairy cattle farm wastewater in East Java Province, Indonesia

Background and Aim: Antibiotic resistance in Klebsiella pneumoniae, especially extended-spectrum beta-lactamase (ESBL) producers, has become a global public health problem. This study aimed to estimate the prevalence of multidrug resistance (MDR) and ESBL-producing K. pneumoniae in wastewater from dairy farms. Materials and Methods: This study was conducted on dairy farms in East Java Province from June to October 2022. In total, 342 dairy farm wastewater samples were isolated on buffered peptone water media from six cities/regencies with the highest dairy cattle population in East Java. Samples were identified using MacConkey agar media, Gram-staining,eosin-methylene blue agar, and biochemical tests. In total, 14.32% (49/342) samples contained K. pneumoniae. Positive isolates were tested for antibiotic sensitivity. Klebsiella pneumoniae resistant to beta-lactam was confirmed using the double-disk synergy test to confirm the presence of ESBL-producing bacteria. Results: The percentage of antibiotic resistance in K. pneumoniae was 98% resistance to ampicillin, 67.3% to cefotaxime, 46.9% to tetracycline, 49% to ciprofloxacin, 98% to streptomycin, 14.3% to sulfamethoxazole–trimethoprim, and 83.7% to chloramphenicol. The prevalence of MDR in K. pneumoniae was 12.57% (43/342), with the highest prevalence in the five classes of antibiotics at 41.86% (18/43), and the prevalence of ESBL-producing K. pneumoniae was 5.55% (19/342), with the highest prevalence in the districts of Blitar and Pasuruan at 26.31% (5/19). Conclusion: Although the prevalence of ESBL-producing K. pneumoniae in wastewater samples from dairy farms was low, caution is recommended because they can be a reservoir for ESBL. Keywords: animal health, dairy cattle, extended-spectrum beta-lactamase, human health, Klebsiella pneumoniae, wastewater.

ASSESSMENT OF THE EFFECT OF AEROSOL DISINFECTION OF NANOSILVER AND HYDROPEROXIDE AG+ AT CAN THO UNIVERSITY OF MEDICINE AND PHARMACY

Background: One of the crucial methods for lowering hospital infections is air disinfection. Because the airborne contagion environment is difficult to control, the development of effective air disinfectants is critical to maintaining a safe air environment and preventing respiratory infections. At the Hospital of Can Tho University of Medicine and Pharmacy, nanosilver and hydroperoxide Ag+ are two different kinds of airborne infection control, so an analytical cross-sectional study was conducted with objectives: to compare the disinfection efficacy of two solutions, nanosilver and 5% Hydroperoxide - 0.005% Ag+, and to assess the efficacy of the silver nano solution and 5% Hydroperoxide - 0.005% Ag+ solution in disinfecting hospital air. Materials and methods: This study was conducted on 156 air microbiological samples in 13 wards at the Can Tho University of Medicine and Pharmacy Hospital using Koch's dust deposition method based on the principle of allowing dust to settle on the surface of nutrient agar boxes for a certain period. Data analysis using SPSS 26.0 software. Results: After disinfecting the rooms with nanosilver, the air quality was good in 69.23% of cases and pretty in 30.07%. With 5% Hydroperoxide - 0.005% Ag+, the number of rooms with good air quality was 15.39%, with pretty air was 69.23, and with poor air was 15.39%. Nanosilver can eradicate the fungus from Sabouraud Agar medium, and the density after disinfection was lower than before; the difference was not statistically significant (p>0.05). Moreover, on Macconkey Agar and Blood Agar medium, bacterial density after disinfecting by nanosilver was decreased; the difference was statistically significant (p < 0.05). Nanosilver reduced the bacterial density in 4 types of rooms better than 5% Hydroperoxide - 0.005% Ag+ solution. However, the difference was not statistically significant (p > 0.05). Conclusions: Nanosilver can be used to disinfect the hospital air as an alternative to 5% Hydroperoxide - 0.005% Ag+. Nanosilver disinfects well on gram-negative and gram-positive bacteria. Additional methods for improving the antifungal impact of nanosilver are required.

Bacterial Stomatitis in Wild Reticulated Pythons (Malayopython reticulatus) in Malaysia

Bacterial stomatitis is a common clinical form of upper alimentary tract disease in reptiles. The current study aimed to isolate and identify the common aerobes in the oral cavities of wild reticulated pythons and to profile their antimicrobial susceptibility. The need to conduct the current research was deemed in parallel with the increasing demand for snakes as pets and the growing emergence of multiple-drug-resistant organisms. A total of 40 fresh carcasses of the wild-caught reticulated pythons were assessed for the presence or absence of stomatitis. Oral swabs were obtained and cultured on blood and MacConkey agar media. The colony and cellular morphologies of the isolates were evaluated, followed by Gram-positive and Gram-negative bacterial identification. Antimicrobial susceptibility testing was performed using Kirby-Bauer disk diffusion method against selected antibiotics, namely gentamicin (GEN), amoxicillin (AMX), cephalexin (LEX), azithromycin (AZM), tetracycline (TET), and ciprofloxacin (CIP), commonly used to treat bacterial infection in reptiles. Results indicated that the prevalence of stomatitis was 77.5%. Among 153 isolates identified, 76.47% of bacteria were identified from pythons with stomatitis lesions, while 23.53% of bacteria were identified from pythons without stomatitis. Of 153 isolates, Gram-negative bacteria were shown to be predominant (94.77%). The three most isolated bacterial species were Aeromonas spp. (14.38%), Klebsiella pneumoniae (11.76%), and Alcaligenes faecalis (8.5%). Meanwhile, coagulase-negative Staphylococcus spp. (4.58%) and Corynebacterium spp. (0.66%) were the only isolated Gram-positive aerobes. Most isolates were observed to be equally susceptible to GEN and CIP (at 95.8%) but highly resistant to AMX (83.3%) and LEX (75.0%). In conclusion, bacterial stomatitis in wild-caught reticulated pythons was highly prevalent and often seen as a mixed bacterial infection (96.8%). The isolated bacteria consistently show susceptibility towards GEN and CIP and thus could be considered the primary line of antibiotics in treating this disease.

Enhancing Contact Tracing for Serratia marcescens Biofilm on High-Usage Body Towels in Rivers State Bathrooms

The placement of body towels before and after use in bathrooms where Serratia marcescens proliferates calls for concern, as Serratia marcescens, an airborne opportunistic pathogen has been reported keratinolytic. Serratia marcescens, a bacterium commonly noticed with a pink or red slimy appearance in toilet sinks, bowls, and tiles presents an aesthetically, unappealing and disgusting appearance in the toilet surroundings. The study aimed to trace the presence of Serratia marcescens on frequently used body towels hung in the bathroom doors. Swabs from forty (40) differently used towels were collected from twenty (20) volunteered homes and analyzed using standard microbiological procedures. Microbiological procedures involved inoculating the swab sample on a prepared peptone broth and plating on MacConkey agar media, followed by identification and streak of the recovered isolate onto Congo red agar media for biofilm formation. Results showed the recovery of Serratia marcescens isolates. The three homes showed a Serratia marcescens count of 3 X 10, 1 X 102, and 7 X 10 CFU per swab for house units C, I, and P respectively. Serratia marcescens could form a biofilm, a basic feature that allowed it to strive on a body towel. The results derived strongly identified the presence of Serratia marcescens biofilm on body towels hung in the bathrooms. This could have health implications for towel users due to the bacteria's keratolytic properties. Hence, the need for constant surveillance to support effective measures of hygiene, aimed at preventing the spread of Serratia marcescens is recommended.

Enhancing Contact Tracing for Serratia marcescens Biofilm on High-Usage Body Towels in Rivers State Bathrooms


 The placement of body towels before and after use in bathrooms where Serratia marcescens proliferates calls for concern, as Serratia marcescens, an airborne opportunistic pathogen has been reported keratinolytic. Serratia marcescens, a bacterium commonly noticed with a pink or red slimy appearance in toilet sinks, bowls, and tiles presents an aesthetically, unappealing and disgusting appearance in the toilet surroundings. The study aimed to trace the presence of Serratia marcescens on frequently used body towels hung in the bathroom doors. Swabs from forty (40) differently used towels were collected from twenty (20) volunteered homes and analyzed using standard microbiological procedures. Microbiological procedures involved inoculating the swab sample on a prepared peptone broth and plating on MacConkey agar media, followed by identification and streak of the recovered isolate onto Congo red agar media for biofilm formation. Results showed the recovery of Serratia marcescens isolates. The three homes showed a Serratia marcescens count of 3 X 10, 1 X 102, and 7 X 10 CFU per swab for house units C, I, and P respectively. Serratia marcescens could form a biofilm, a basic feature that allowed it to strive on a body towel. The results derived strongly identified the presence of Serratia marcescens biofilm on body towels hung in the bathrooms. This could have health implications for towel users due to the bacteria's keratolytic properties. Hence, the need for constant surveillance to support effective measures of hygiene, aimed at preventing the spread of Serratia marcescens is recommended.

Crown gall development on cannabis (Cannabis sativa L., marijuana) plants caused by Agrobacterium tumefaciens species-complex

Crown gall, caused by Agrobacterium tumefaciens (syn. Rhizobium radiobacter), is characterized by gall formation on crowns, stems or roots on many plant species. We describe the occurrence of this disease for the first time on cannabis (Cannabis sativa L. marijuana) plants in two licenced production facilities in British Columbia. Galls were observed on crowns, stems and roots of several genotypes, including ‘White Rhino’, ‘Pink Kush’ and ‘Sour Kush’. The affected plants displayed no other visible symptoms. The incidence of crown gall was extremely low, estimated at 0.01% of total plants. Isolations made from root and stem galls on MacConkey medium (selective for Gram -ve bacteria) and on D1 medium (selective for Agrobacterium spp.) yielded a range of bacterial species, which did not include Agrobacterium sp. However, the presence of Agrobacterium in gall tissues was confirmed following PCR amplification with primers for the indole-acetic acid (iaa) H gene on the T-DNA region of Agrobacterium spp. and showed > 99% similarity to A. tumefaciens. Artificial inoculations were conducted using an A. tumefaciens strain from ATCC (Rhizobium radiobacter, strain designation TT134 [6-1-2, CIP 104 336], cat. 15955) which resulted in gall formation on five cannabis genotypes tested. These galls contained the iaaH gene as determined by PCR and the pathogen was successfully re-isolated on MacConkey agar medium at 4 and 10 weeks post-inoculation. Our findings confirm the occurrence of A. tumefaciens on naturally infected and artificially infected cannabis plants grown under greenhouse conditions and demonstrate reproducible gall symptoms on several cannabis genotypes following A. tumefaciens inoculation.

Evaluation of Plasmid Mediated Drugs Resistance among Wound Bacterial Infection Obtained from Patients Attending some Hospitals in Kano State, Nigeria

This study was carried out to evaluate the plasmid mediated drugs resistance among wound bacteria isolates obtained from (352 swabs specimens) patients attending some hospitals in Kano State (National Orthopaedic Hospital Dala, Aminu Kano Teaching Hospital, Infectious Diseases Hospital, Bichi General Hospital and Wudil General Hospital). Swab specimens were obtained aseptically using the randomized sampling method and cultured on Blood and MacConkey agar media and incubated aerobically and anaerobically for 24 hours. Result showed that two hundred and twelve (212, 60.23%) out of 352 harbors some bacteria, which consist of six different species (Pseudomonas aeruginosa, Escherichia coli, Proteus mirabilis, Klebsiella pneumoniae, Staphylococcus aureus and Streptocoocus pyogenes) as isolated from wound swabs specimens. S. aureus (39%) and S. pyogenes (7%) showed the highest and the lowest prevalences respectively. Antibiotic susceptibility testing was done using the Kirby-Bauer disk diffusion technique on Mueller-Hinton agar, where 140 (66%) isolates were susceptible while 72 (34%) isolates were resistant. Bacteria isolates exhibited the highest resistance to Cefixime, Azithromycin and Erythromycin and least resistance to Ciprofloxacin (most effective/active) among the antibiotics used. The resistant isolates were subjected to curing experiment. Plasmid detection and isolation showed that out of 72 resistant isolates, 49 contain plasmids which accounts for 68%. After plasmid curing and second sensitivity test, 38 (78%) isolates that were initially resistant becomesusceptible to the same antibiotics used. Hence, post plasmid-curing sensitivity test revealed increased susceptibility pattern of isolates to the previously used antibiotics. This indicates that plasmid-borne multidrug resistant gene had been denatured by the sodium dodecyl sulfate used as the curing agent. Therefore, this study justifies the need to encourage infection control, and proper treatment to prevent the spread and re-emergence of plasmid-mediated drugs resistant bacteria.

First report of potato (Solanum tuberosum L.) blackleg disease caused by Dickeya solani in Mayabeque, Cuba.

Blackleg, caused by Pectobacterium spp. and Dickeya spp., is an important disease of potatoes. During the period from November 20 and March 2021, stems of potato plants showing necrosis and rot symptoms, and chlorotic leaves, were collected from commercial production areas of the Mayabeque province of Cuba (Fig. 1A). After disinfestation of affected stems, small fragments of the stem were cut and macerated in a sterile 0.85% NaCl solution. Serial dilutions of bacterial suspension were prepared and streaked onto nutrient agar in Petri plates. Two colonies per sample showing the characteristic of "fried egg" were selected for further investigation, and an isolated was selected and named D7. The isolated bacterium was rod shaped, gram-negative, motile, oxidase and indole production negative, with anaerobic growth, and able to use lactose as carbon source in Mac Conkey Agar medium. One colony of the isolate D7 was selected and multiplied. Total DNA of the bacteria cells was extracted and used to amplify the genes pelADE (Nassar et al., 1996) and gapA (Cigna et al., 2017), to differentiate Dickeya from Pectobacterium. The sequence obtained showed 99.75% and 99.88% nucleotide identity with Dickeya solani for pelADE (Genbank accession number ON644347) and gapA (Genbank accession number ON644346), respectively. To confirm the pathogenicity of the isolate D7, four 15-day-old potato plants, including two plants of each 'Otolia' and 'Naima' potatoes were inoculated with a bacterial suspension of the isolate D7 (108 CFU/ml) in sterile water by stabbing. Control plants were stabbed with sterile water. Inoculated plants were maintained at 28°C, relative humidity of about 90%, and at 12 h light/12 h dark, as described by (Chen et al. 2014). After 3 to 5 days, typical blackleg disease symptoms (water-soaked lesions and necrosis) developed at the inoculated areas of plants (Fig. 1B). No symptom was observed in the control plants. Bacterium was re-isolated from symptomatic plants and the isolates had the same cultural, physiological, and biochemical characteristics to the isolate D7. To our knowledge, this is the first report of D. solani causing blackleg in potato fields in Cuba. Further studies to determine the spread of this pathogen in potato producing areas in Cuba is underway.

Detection of Uropathogens and their Antimicrobial Susceptibility Pattern by VITEK2 Automated System in the Intensive Care Unit

Background: Indwelling urinary catheters are frequently used in critically ill patients. Approximately 66% of patients in intensive care have a urinary catheter in place. Catheter-associated urinary tract infection (CAUTI) is a significant cause of morbidity and mortality, affecting all ages. Antibiotic resistance is one of the biggest public health challenges of our time. Infection with antibiotic-resistant bacteria has become an increasingly complex problem in ICU. Objective: To identify catheter-associated uropathogens and their antimicrobial susceptibility pattern by VITEK2 automated system in intensive care unit patients in Rajshahi Medical College Hospital.
 Material and Method: A cross-sectional descriptive study was conducted in the Microbiology department of Rajshahi Medical College, the Intensive Care Unit of Rajshahi Medical College Hospital, and a commercial laboratory in Rajshahi during the period of January 2021 to December 2021. The specimen (urine) was inoculated in blood agar, nutrient agar, and MacConkey's agar media and incubated aerobically at 37˚C for 24 hours. Bacterial isolations and susceptibility tests were done by VITEK2 automated system.
 Result: Out of the total 96 samples, culture yielded growth was 36 (37.50%), and culture-negative cases were 60(62.50%). In all age groups, males were predominant, totaling 63 (66.20%). A maximum of 33 cases were found in the age group of 18-30 years where males 23(69.70%) and females 10(30.30%). Among the culture-positive isolates, gram-negative organisms were higher (87.17%) than gram-positive (12.82%). Among 39 isolates, Escherichia coli was the predominant organism of 12(30.76%). Other isolates were Klebsiella spp. 10(25.64%), Pseudomonas spp. 5(12.82%), Acinetobacter spp. 4(10.25%), Enterobacter spp. 2(05.12%), CoNS 3(07.69%) respectively. Regarding the antimicrobial susceptibility test, gram-negative organisms were highly susceptible to Colistin, followed by Ertapenem, Imipenem, Meropenem, and Amikacin. The most effective drugs for Gram-positive were Linezolid. Other susceptible drugs against Gram-positive bacteria were Vancomycin and Tigecycline.
 Conclusion: Most of the isolated bacteria are multidrug resistant. Determining antimicrobial susceptibility by VITEK2 will aid the physician in choosing appropriate antibiotics and treating multidrug-resistant cases.
 TAJ 2022; 35: No-2: 43-49

Morphology, Biochemical, and Molecular Characterization of Pasteurella multocida Causing Hemorrhagic Septicemia in Indonesia.

Pasteurella multocida is a Gram-negative bacterium that causes hemorrhagic septicemia (HS) in buffaloes and cattle. The disease causes serious problems in Indonesian livestock and is classified as a serious transmissible animal disease. Previous research has determined the diversity of P. multocida using a serotyping method based on the antigenic properties of capsule polysaccharides. An alternative method for analysis utilizes sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and random amplified polymorphic DNA (RAPD). This study aimed to characterize and determine P. multocida diversity in several regions of Indonesia based on phenotypic character, protein profile, and the band pattern of RAPD results. Bacterial identification was performed using traditional biochemical techniques and API® 20NE systems and then confirmed molecularly using polymerase chain reaction (PCR). The freeze-thawing technique was performed to obtain the bacterial protein extract, and DNA extraction was executed using DNAzol. The extracted protein and RAPD product were then electrophoresed on 12% polyacrylamide gel and 1.5% agarose gel, respectively. The results indicate that the molecular weight range of the protein bands is 12-209 kDa, and the band pattern of the RAPD results ranged from 307-3,100 bp. Based on phenotypical analysis, P. multocida from South Sulawesi Province exhibited a variety of growth characteristics in MacConkey agar media. Using the hierarchical clustering analysis of the band patterns of RAPD and the whole-cell protein profiles, four and five clusters were formed, respectively. These results indicate molecular diversity among P. multocida from several regions of Indonesia.

Multidrug resistant and extended spectrum β-lactamase producing gram negative bacterial uropathogens among females in a tertiary hospital, Pokhara

Introduction: Extended spectrum β-lactamase producing gram negative bacilli has emerged and spread worldwide as an important cause of urinary tract infections and indicates a major threat. The study aimed to determine the prevalence of multidrug resistant and extended spectrum beta-lactamase producing gram negative bacterial uropathogens among females in a tertiary level hospital. Methods: A hospital-based cross-sectional study was conducted in Pokhara Academy of Health Sciences, Pokhara from February to November, 2019. A total number of 301 midstream urine samples were collected and examined using MacConkey agar and blood agar medium. Antimicrobial susceptibility testing was done by Kirby Bauer disk diffusion method on Mueller Hinton agar using Clinical and Laboratory Standards Institute guidelines. Results: Out of 301 mid-stream urine samples, 99(33%) sample showed significant bacterial growth. Among them, 78(79%) were gram negative bacteria. Escherichia coli were the predominant organism. Multidrug resistant gram negative isolates were 65.4%. Among 78 Gram negative isolates, 31(39.7%) were extended spectrum β-lactamase producers. Among extended spectrum β-lactamase producers, 27(87.1%) were MDR. Highest frequency of extended spectrum β-lactamases production was seen in E. coli, 23(74.2%). Majority of gram negative bacteria showed susceptibility toward colistin and nitrofurantoin. Ampicillin was found to be highly resistant towards gram negative uropathogen. Conclusions: This study found that higher proportion of multi-drug resistants were among gram negative isolates and further more among extended spectrum β-lactamase producing gram negative isolates. Thus, there is urgent need to address the issue of antimicrobial resistant and promote rational use of the antibiotics in our region.