Detection of pathogenic Vibrio species and antibiogram activity in Asian Seabass (Lates calcarifer) in Tumpat, Kelantan

@#Some of Vibrio species is well known as pathogenic bacteria in aquaculture and the marine industry. Its infection is able to generate a massive outbreak and affect the fish population, especially for net caged fish such as seabass. This study was conducted to investigate the prevalence of Vibrio spp. isolated from seabass (Lates calcarifer) in Sri Tujuh Lagoon, Tumpat, Kelantan. Then, to determine the antibiotic resistance in Vibrio isolates. Polymerase chain reaction (PCR) was used to detect Vibrio species using specific primer VR169 and VR744 with estimation base pair size band, 597 bp and further identified by sequencing. On the other hand, antibiotic susceptibility tests were continued by using 13 types of antibiotics; kanamycin (K30), chloramphenicol (C30), neomycin (N10), ampicillin (AMP10), nitrofurantoin (F300), tetracycline (TE30), streptomycin (S10), norfloxacin (NOR10), ciprofloxacin (CIP5), nalidixic acid (NA30), gentamicin (CN10), doxycycline (DO30) and sulfamethoxazole (SXT100). As a result, 14 Vibrio isolates were identified, including Vibrio fluvialis (n=6), Vibrio parahaemolyticus (n=3), Vibrio harveyi (n=2) and each isolate for Vibrio vulnificus, Vibrio alginolyticus and Vibrio spp. The results showed that all isolates were sensitive to most antibiotics except ampicillin, neomycin and streptomycin. The MAR index value was ranging from 0 to 0.31. This study demonstrates the prevalence of Vibrio spp. in seabass and the report on multidrug resistance strains that could be of concern to the fish farmers. In addition, data from this study can be further used in fish disease management plans.

Antibiotic resistance profile and RAPD analysis of Campylobacter jejuni isolated from vegetables farms and retail markets

Objective: To investigate antibiotic resistance profile and characterize Campylobacter jejuni (C. jejuni) isolates using random amplified polymorphic DNA (RAPD) analysis. Methods: Ninety eight C. jejuni isolates from farms and retail outlets were screened against 10 antibiotics commonly used clinically and agriculturally by using disk diffusion method. RAPD analysis was done to characterize 98 C. jejuni isolates. Results: Fifty-one percent of the isolates had multiple antibiotic resistance index 0.2 and below. This indicated that the isolates in the vegetables were not from the high risk environment or extensive farming practices. C. jejuni isolates found resistant towards penicillin G (93%), vancomycin (86%), ampicillin (35%), erythromycin (28%), genta-mycin (4%), amikacin (3%), enrofloxacin (1%), norfloxacin (1%) and no resistance to-wards ciprofloxacin. RAPD clustering analysis showed that the contamination of C. jejuni in vegetables was likely due to cross contamination at retail markets. Conclusions: C. jejuni contamination in vegetables at retail markets was due to cross contamination. Current finding proved that C. jejuni in small scale vegetables production was less expose towards antibiotic abuse.

Antibiotic resistance profile and RAPD analysis of Campylobacter jejuni isolated from vegetables farms and retail markets

Objective: To investigate antibiotic resistance profile and characterize Campylobacter jejuni (C. jejuni) isolates using random amplified polymorphic DNA (RAPD) analysis. Methods: Ninety eight C. jejuni isolates from farms and retail outlets were screened against 10 antibiotics commonly used clinically and agriculturally by using disk diffusion method. RAPD analysis was done to characterize 98 C. jejuni isolates. Results: Fifty-one percent of the isolates had multiple antibiotic resistance index 0.2 and below. This indicated that the isolates in the vegetables were not from the high risk environment or extensive farming practices. C. jejuni isolates found resistant towards penicillin G (93%), vancomycin (86%), ampicillin (35%), erythromycin (28%), gentamycin (4%), amikacin (3%), enrofloxacin (1%), norfloxacin (1%) and no resistance towards ciprofloxacin. RAPD clustering analysis showed that the contamination of C. jejuni in vegetables was likely due to cross contamination at retail markets. Conclusions: C. jejuni contamination in vegetables at retail markets was due to cross contamination. Current finding proved that C. jejuni in small scale vegetables production was less expose towards antibiotic abuse.

Analysis of pathogenic bacterial distribution and drug resistance of hospital infection during 2011-2013 / 国际检验医学杂志

Objective To statistically analyze the bacterial flora change and drug resistance situation in the patients with bacteri‐al infection to provide reference for clinical rational drug use and the management of nosocomial infection .Methods The clinical samples were conventionally isolated and cultured .The bacteria identification and drug sensitivity test were performed by using the bioMerieux company VITEK2 automatic microorganism analyzer .The confirmation test of drug susceptibility adopted the disk dif‐fusion method recommended by the American Clinical and Laboratory Standardization Committee (CLSI) .Results The sample sources in the hospital infection during 2011-2013 were main sputum ,secretions and midstream urine ;the main infectious bacteria showed the increasing trend ,the top 5 of bacteria were :Escherichia coli (ECO) ,Klebsiella pneumoniae (KPN) ,Acinetobacter bau‐manii (ABA) ,Pseudomonas aeruginosa (PAE) ,Staphylococcus aureus (SAU);the top three of common multi‐drug resistant bacte‐ria were ABA ,PAE and ECO ,their constituent ratio during these three years had a small amplitude increase ;the antibiotics for Gram negative bacilli (G -) resistance rate of more than 70% during these 3 years were :ampicillin ,cefuroxime sodium and cefu‐roxime axetil;the antibiotics for Gram positive cocci (G+ ) resistance rate of more than 70% during these 3 years were penicillin and erythromycin ..Except for 4 cases of Enterococcus faecalis ,no other vancomycin‐resistant strains were found .Conclusion The sam‐ple source of bacterial infection is dominated by sputum ,the gram negative bacteria are the main force of hospital infection ,showing a increasing trend every year ,multi- drug resistant strains are also continuously rising ,penicillin is unsuitable to the clinical treat‐ment of bacterial infection ;imipenem has very high sensitivity to ECO and KPN .The hospital should attach great importance to the infection management ,strengthen the application and management of antibiotics ,decrease the outbreak and prevalence of bacterial infection and reduce the increase of drug resistant strains .

Integron mediated multidrug resistance in extended spectrum beta-lactamase producing clinical isolates of Klebsiella pneumoniae

The present study describes integron mediated multiple antibiotic resistance in extended-spectrum β-lactamase producing clinical isolates of Klebsiella pneumoniae. One hundred and four clinical isolates of K. pneumoniae from two Iranian hospitals were screened for extended-spectrum β-lactamase production and susceptibility of the extended-spectrum β-lactamase producing isolates was determined to 17 antibiotics by disc diffusion. Presence of integron classes 1, 2 and 3 was detected by PCR and integrase specific primers. Isolates harboring class 1 integron were then screened for variable regions using PCR. Fifty isolates (48%) produced extended-spectrum β-lactamases among which, 22 (44%) harbored class 1, 3 (6%) carried class 2 and none contained class 3 integons. Integron carriage was significantly associated with higher rates of multiple antibiotic resistance in extended-spectrum β-lactamase producing clinical isolates of K. pneumoniae. Integron harboring isolates were more resistant to aztreonam (51.3%), ceftazidime (42.6%), cefotaxime (43.3%), cefepime (24.6%), kanamycin (43.2%), tobramycin (30.7%), norfloxcacin (32%) and spectinomycin (25.6%) compared to the organisms without integrons. On the other hand, resistance to nitrofurantoin and streptomycin was significantly higher among the integron negative isolates. PCR amplification of class1 integron variable regions revealed 9 different sized DNA fragments and isolates with similar profiles for class 1 integron variable regions showed the same antibiotic resistance phenotypes.

Antibiotic susceptibility pattern of isolates of Pseudomonas aeruginosa in a Saudi Arabian Hospital.

Pseudomonas aeruginosa is one of the most common gram-negative bacteria. identified in the clinical specimens of hospital admitted patients. A major problem in P. aeruginosa infection may be that this pathogen exhibits a high degree of resistance to a broad spectrum of antibiotics. The study aimed to isolate and determine the antimicrobial susceptibility patterns of the P. aeruginosa. This prospective study was done over a period of six months. Forty one clinical isolates of Pseudomonas aeruginosa (P. aeruginosa) were isolated from sputum specimens of the patients suspected of having respiratory tract infection. The antibiotic susceptibility profiles of all the isolates were determined using disk diffusion method as recommended by Clinical Laboratory Standards Institute. Ciprofloxacin was found to be the most effective antimicrobial agent with 85.4% susceptibility followed by imipenem (75.6%), aminoglycosides (amikacin, 95.1% and gentamicin, 90.3%), and the beta-lactams (cefepime 65.8%, ceftazidime, 51.2%). Piperacillin showed the maximum resistance (46.3%) followed by Aztreonam (36.6%). Regular antimicrobial susceptibility surveillance is essential for area-wise monitoring of the resistance patterns. An effective national and state level antibiotic policy and draft guidelines should be introduced to preserve the effectiveness of antibiotics and for better patient management.

Antibacterial effect of Allium sativum cloves and Zingiber officinale rhizomes against multiple-drug resistant clinical pathogens

Objective: To evaluate the antibacterial properties of Allium sativum (garlic) cloves and Zingiberofficinale (ginger) rhizomes against multi-drug resistant clinical pathogens causing nosocomial infection. Methods: The cloves of garlic and rhizomes of ginger were extracted with 95% (v/v) ethanol. The ethanolic extracts were subjected to antibacterial sensitivity test against clinical pathogens. Results: Anti-bacterial potentials of the extracts of two crude garlic cloves and ginger rhizomes were tested against five gram negative and two gram positive multi-drug resistant bacteria isolates. All the bacterial isolates were susceptible to crude extracts of both plants extracts. Except Enterobacter sp. and Klebsiella sp., all other isolates were susceptible when subjected to ethanolic extracts of garlic and ginger. The highest inhibition zone was observed with garlic (19.45 mm) against Pseudomonas aeruginosa (P. aeruginosa). The minimal inhibitory concentration was as low as 67.00 μg/mL against P. aeruginosa. Conclusions: Natural spices of garlic and ginger possess effective anti-bacterial activity against multi-drug clinical pathogens and can be used for prevention of drug resistant microbial diseases and further evaluation is necessary.

Antibiotic resistance of Vibrio parahaemolyticus isolated from pond-reared Litopenaeus vannamei marketed in Natal, Brazil

Ten out of fifty fresh and refrigerated samples of shrimp (Litopenaeus vannamei) collected from retailers in Natal (Rio Grande do Norte, Northeastern Brazil) tested positive for Vibrio parahaemolyticus. The Kanagawa test and multiplex PCR assays were used to detect TDH and TRH hemolysins and the tdh, trh and tlh genes, respectively. All strains were Kanagawa-negative and tlh-positive. Antibiotic susceptibility testing was done for seven antibiotics by the agar diffusion technique. Five strains (50 percent) presented multiple antibiotic resistance to ampicillin (90 percent) and amikacin (60 percent), while two strains (20 percent) displayed intermediate-level resistance to amikacin. All strains were sensitive to chloramphenicol. Intermediate-level susceptibility and/or resistance to other antibiotics ranged from 10 to 90 percent, with emphasis on the observed growing intermediate-level resistance to ciprofloxacin. Half our isolates yielded a multiple antibiotic resistance index above 0.2 (range: 0.14-0.29), indicating a considerable risk of propagation of antibiotic resistance throughout the food chain.

Occurrence of heavy metals and antibiotic resistance in bacteria from internal organs of american bullfrog (Rana catesbeiana) raised in Malaysia

A total of 40 bacteria have been successfully isolated from internal organs of the American bullfrog (Rana catesbeiana) raised in Malaysia, namely, eight isolates of Aeromonas spp., 21 of Edwardsiella spp., six of Flavobacterium spp. and five of Vibrio spp. In terms of antibiotic susceptibility testing, each isolate was tested against 21 antibiotics, resulting in 482 (57.3 percent) cases of sensitivity and 61 (7.3 percent) cases of partial sensitivity. Meanwhile, 297 (35.4 percent) bacterial isolates were registered as resistant. The multiple antibiotic resistance (MAR) index of each bacterial species indicated that bacteria from raised bullfrogs have been exposed to tested antibiotics with results ranging from 0.27 to 0.39. Additionally, high percentages of heavy metal resistance among these isolates were observed, with values ranging from 85.0 to 100.0 percent. The current results provided us information on bacterial levels of locally farmed bullfrogs exposed to copper, cadmium, chromium as well as 21 types of antibiotics.(AU)

Characterization of Shigella spp. by antimicrobial resistance and PCR detection of ipa genes in an infantile population from Porto Velho (Western Amazon region), Brazil

The incidence of Shigella spp. was assessed in 877 infants from the public hospital in Rondônia (Western Amazon region, Brazil) where Shigella represents the fourth cause of diarrhea. Twenty-five isolates were identified: 18 were Shigella flexneri, three Shigella sonnei, three Shigella boydii and one Shigella dysenteriae. With the exception of S. dysenteriae, all Shigella spp. isolated from children with diarrhea acquired multiple antibiotic resistances. PCR detection of ipa virulence genes and invasion assays of bloody diarrhea and fever (colitis) were compared among 25 patients testing positive for Shigella. The ipaH and ipaBCD genes were detected in almost all isolates and, unsurprisingly, all Shigella isolates associated with colitis were able to invade HeLa cells. This work alerts for multiple antibiotic resistant Shigella in the region and characterizes presence of ipa virulence genes and invasion phenotypesin dysenteric shigellosis.

Sequence analysis of multiple-antibiotic-resistant active efflux systemgene acrAB of Salmonella typhi / 重庆医科大学学报

Objective:To identify the sequence of the multiple-antibiotic-resistant efflux acrAB gene in Salmonella typhi 275(a clinically isolated strain) and analyze its structure,amino acid sequence.Methods:The whole acrAB gene of Salmonella typhi 275 were amplified by PCR with the primers designed from Genebank,and the sequence of products and the amino acid sequence were detected.Results:The whole sequence of acrAB gene of Salmonella typhi 275 contained 5156 bases and 99.38% identity were found in comparison with the reference sequence of Salmonella typhi (Genebank No. AL627267);84.69% identity were detected in comparison with that of Escherichia coli (Genebank No.ECUO0734).The acrR,actA,acrB of Salmonella typhi 275 coded AcrR,AcrA,AcrB proteins with 217,397 and 1049 amino acids,respectively.1 amino acid alteration existed in both AcrA (Ser270Thr)and AcrB (Met964Thr) in comparison with the reference Genebank amino acids sequence.Compared with Escherichia coli,28,33 and 56 amino acids alteration and 86.98%,91.69% and 94.66% identity in the AcrR,Acta and AcrB were found respectively.2 additional amino acids existed in AcrR of Salmonella typhi 275.The promoter region of Acta and acrR was located in 4367-4507 bases and the SD sequence was in 4373-4377 bases.Conclusion:The multiple-antibiotic-resistant active efflux system gene acrAB of Salmonella typhi shows high homology in bases,amino acids and protein structure with that of Escherichia coli and it is the possible reason that Salmonella typhi is resistant to multi-antibiotics without similar chemical structure.

The mutation of mar gene in clinical isolated Escherichia coli resistant to fluoroquinolones / 中华传染病杂志

Objective To study the antibiotic resistance mediated by the mar mechanism in clinical isolated Escherichia coli. Methods To analyse the antibiotic resistance pattern and the out membrane protein spectrum of clinical isolated E. coli . The plasmid pNJR3 2 carrying the wild type gyr A gene was transferred into clinical multiple resistant strain LC 1. The resistant pattern and plasmid pattern of LC 1 before and after plasmid transferring were compared. The mar OR of clinical strain LC 1 was cloned and sequenced through PCR. The nucleotides of mar gene were compared with the wild type issued by GenBank. Results Of all the 49 clinical strains , six strains express Mrp5, a mar specific out membrane protein. No changes were detected on susceptibility to fluoroquinolone of clinical strain LC 1 after pNJR3 2 transferred suggesting that gyr A was not the main reason contributing to fluoroquinolone resistance. Sequence analysis showed three mutant spots in mar OR of clinical strain LC 1. Conclusion There does exist mar mechanism in clinical isolated Escherichia coli resistant to fluoroquinolone. The mutation in mar OR may contribute to the mar phenotype of LC 1.

Antibiotic Resistance and Genotyping of Pseudomonas aeruginosa Isolated from Burn Unit by Pulsed-field Gel Electrophoresis / 中华医院感染学杂志

OBJECTIVE To analyze antibiotic resistance pattern and use pulsed-field gel electrophoresis(PFGE) to study the molecular epidemiology of Pseudomonas aeruginosa isolated from burn unit. METHODS P.aeruginosa had been isolated and tested by K-B method from clinical samples and antibiotic resistance was analyzed and studied retrospectively. RESULTS The drug resistance of P.aeruginosa to nine antibiotics was high,the multiple drug resistance rate was 30%. CONCLUSIONS The resistance rates to commonly used antibacterials in P.aeruginosa are high and the resistance pattern is wide.PEGE is a better genotyping method to study molecular epidemiology and analytic homology.