Optimizing and purifying extracellular amylase from soil bacteria to inhibit clinical biofilm-forming bacteria

Background Bacterial biofilms have become a major threat to human health. The objective of this study was to isolate amylase-producing bacteria from soil to determine the overall inhibition of certain pathogenic bacterial biofilms. Methods We used serial dilution and the streaking method to obtain a total of 75 positive amylase isolates. The starch-agar plate method was used to screen the amylolytic activities of these isolates, and we used morphological and biochemical methods to characterize the isolates. Optimal conditions for amylase production and purification using Sephadex G-200 and SDS-PAGE were monitored. We screened these isolates’ antagonistic activities and the purified amylase against pathogenic and multi-drug-resistant human bacteria using the agar disk diffusion method. Some standard antibiotics were controlled according to their degree of sensitivity. Finally, we used spectrophotometric methods to screen the antibiofilm 24 and 48 h after application of filtering and purifying enzymes in order to determine its efficacy at human pathogenic bacteria. Results The isolated Bacillus species were Bacillus megaterium (26.7%), Bacillus subtilis (16%), Bacillus cereus (13.3%), Bacillus thuringiesis (10.7%), Bacillus lentus (10.7%), Bacillus mycoides (5.3%), Bacillus alvei (5.3%), Bacillus polymyxa (4%), Bacillus circulans (4%), and Micrococcus roseus (4%). Interestingly, all isolates showed a high antagonism to target pathogens. B. alevi had the highest recorded activity (48 mm) and B. polymyxa had the lowest recorded activity (12 mm) against Staphylococcus aureus (MRSA) and Escherichia coli, respectively. On the other hand, we detected no antibacterial activity for purified amylase. The supernatant of the isolated amylase-producing bacteria and its purified amylase showed significant inhibition for biofilm: 93.7% and 78.8%, respectively. This suggests that supernatant and purified amylase may be effective for clinical and environmental biofilm control. Discussion Our results showed that soil bacterial isolates such as Bacillus sp. supernatant and its purified amylase are good antibiofilm tools that can inhibit multidrug-resistant former strains. They could be beneficial for pharmaceutical use. While purified amylase was effective as an antibiofilm, the isolated supernatant showed better results.

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Multidrug resistant Salmonella spp. and Escherichia coli from a popular Ready-to-Eat local food (Fufu) from commercial food vendors in Ghana

Journal of Food Safety and Hygiene ◽

10.18502/jfsh.v5i4.5702 ◽

2021 ◽

Author(s):

Courage Kosi Setsoafia Saba ◽

Akosua Bonsu Karikar ◽

Enoch Yeleliere ◽

Patrick Takyi ◽

Stephen Wilson Kpordze

Keyword(s):

Foodborne Pathogens ◽

Pathogenic Bacteria ◽

Multidrug Resistant ◽

Diffusion Method ◽

Microbial Count ◽

Salmonella Spp ◽

Public Health Importance ◽

Disk Diffusion Method ◽

E Coli ◽

Susceptibility Profiles

Microbial contamination of vended foods are of public health importance due to the potential of becoming a reservoir of foodborne pathogens and resistant strains of bacteria. This study looked at the presence of pathogenic bacteria in a popular Ready-To-Eat (RTE) traditional food, Fufu in Ghana. Sixty (60) Fufu samples were obtained from various food joints categorized as Opened, Semi-closed and Closed or Restaurants. Samples were processed and analyzed using standard bacteriological methods. The susceptibility profiles of the isolates were obtained by using the Kirby-Bauer disk diffusion method with the EUCAST guidelines with the five antibiotics. Prevalence of E. coli was 85% and Salmonella species was 68%. Microbial count of isolated E. coli ranged from 0 to 3×106 cfu/ml. There were no significant differences (p>0.05) among the different modes of operations. Fufu samples from Opened, Semi-closed and Closed joints were respectively contaminated with E. coli and Salmonella species as follows: 92%, 76%; 80%, 60% and 80%, 65%. The Salmonella species showed highest resistance to erythromycin (58.5%) and E. coli species were commonly resistant to Ceftazidime (88.2%) and Ceftriaxone (94.1%). All isolates were susceptible to nitrofurantoin. Multidrug resistance was detected among 27.5% of E. coli strains and 14.6% of Salmonella species. Fufu from the different eating joints in the Tamale Metropolis were substantially contaminated with multidrug resistant pathogens. The study recommends surveillance studies of resistant pathogens in foods, increased education and training of food vendors on sanitation, food handling and safety practices in the region.

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Microbial Growth as Determinant of Antibiotic Production with Biotic Elicitors Stimulation

Jurnal Kimia Sains dan Aplikasi ◽

10.14710/jksa.23.3.89-95 ◽

2020 ◽

Vol 23 (3) ◽

pp. 89-95

Author(s):

Citra Hardiyanti ◽

Khairullinas Khairullinas ◽

Jeky Sasemar Lumban ◽

Titania Tjandrawati Nugroho ◽

Yuana Nurulita

Keyword(s):

Staphylococcus Aureus ◽

Pathogenic Bacteria ◽

Antibiotic Production ◽

Multidrug Resistant ◽

Positive Control ◽

Diffusion Method ◽

Disk Diffusion Method ◽

Antibiotic Resistant ◽

Penicillium Sp ◽

Biotic Elicitor

An antibiotic-resistant and multidrug-resistant (MDR) issue open the role of researchers to continue to search for natural potential as a source of new antimicrobials. One of the potential fungi isolates that can produce antimicrobial active compounds from Indonesian tropical peat soils is Penicillium sp. LBKURCC34. In this study, the production of antimicrobial compounds from local isolates was carried out by batch fermentation method in liquid media with the addition of biotic elicitors to increase the extraction activity and yield. This study aims to optimize the results based on the time the elicitor is added. Staphylococcus aureus was used as a biotic elicitor, which was added on days 2, 3, and 4 in the production of antibiotics by fermentation incubation of 6-14 days. The antibiotic production media was extracted with ethyl acetate and evaporated. The antimicrobial test was carried out by the disk diffusion method against pathogenic bacteria Escherichia coli, Staphylococcus aureus, Bacillus subtilis, and Staphylococcus epidermidis using three crude extract contents (19; 38; and 57 µg/disc). Amoxicillin® was used as a positive control (10 µL/disc). The results showed that the addition of S. aureus biotic elicitor extended the log phase growth of the fungus Penicillium sp. LBKURCC34. The optimum condition of production was obtained by adding initiator treatment on the 3rd day for 14 days incubation with the highest yield and could inhibit the growth of all pathogenic microbes.

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The Role of Purified α-amylase from Soil Bacteria Versus Crude Cell Supernatant from Medium against Clinical Biofilm-forming Bacteria

10.20944/preprints201912.0344.v1 ◽

2019 ◽

Author(s):

Rokaya Elamary ◽

Fawziah Albarakaty ◽

Wesam Salem

Keyword(s):

Human Life ◽

Agar Plate ◽

Antagonistic Activity ◽

Significant Inhibition ◽

Bacterial Biofilm ◽

Diffusion Method ◽

Bacillus Species ◽

Good Tool ◽

Terrestrial Environment ◽

Disk Diffusion Method

Bacterial biofilms have become a significant and growing threat to human life, nature, and environmental health. The aim of this study is to isolate amylase-producing bacteria from the terrestrial environment (soil) for investigating their general inhibition of some pathogenic human bacterial biofilm. A total of 75 amylase producing isolates were obtained by serial dilution and streaking method. Amylotic activity of these isolates was screened by a starch agar plate method. Isolates were characterized by morphological and biochemical methods. The isolated Bacillus species were B. megaterium (26.7%), B. subtilis (16%), B. cereus (13.3%), B. thuringiesis (10.7%), B. lentus (10.7%), B. mycoides (5.3%), B. alvei (5.3%), B. polymyxa (4%), B. circulans (4%) and Micrococcus roseus (4%). The optimum conditions for amylase production were monitored. Antagonistic activity of these isolates and purified amylase against multidrug-resistant pathogenic human bacteria by agar disk diffusion method. The sensitivity level of some standard antibiotics served as control. Interestingly, it was found that all isolates exhibited great antagonistic activity against the target pathogens. The greatest activity recoded by B. alevi (48 mm) against Staphylococcus aureus (MRSA) and the lowest activity recorded by B. polymyxa (12 mm) against E. coli while low inhibition recorded for amylase. Antibiofilm efficacy of isolates supernatant and purified enzyme also monitored by spectrophotometric methods. The results revealed highly significant inhibition with percentages of 93.6 and 78.8% respectively. So, they represent a good tool for biofilm control in clinical and environmental applications.

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Contamination of the Surfaces of a Health Care Environment by Multidrug-Resistant (MDR) Bacteria

International Journal of Microbiology ◽

10.1155/2019/3236526 ◽

2019 ◽

Vol 2019 ◽

pp. 1-7 ◽

Cited By ~ 2

Author(s):

Laila Chaoui ◽

RajaaAit Mhand ◽

Fouad Mellouki ◽

Naima Rhallabi

Keyword(s):

Staphylococcus Aureus ◽

Pathogenic Bacteria ◽

Multidrug Resistant ◽

Cross Sectional Study ◽

Hospital Environment ◽

Diffusion Method ◽

Health Care Environment ◽

Disk Diffusion Method ◽

Cross Sectional ◽

Gram Negative

Nosocomial infections (NIs) are known worldwide and remain a major problem despite scientific and technical advances in the field of health. The severity of the infection depends on the characteristics of the microorganisms involved and the high frequency of resistant pathogens in the hospital environment. The aim of this study is to determine the distribution of pathogenic bacteria (and their resistance to antibiotics) that spread on hospital surfaces, more specifically, on those of various departments in the Provincial Hospital Center (PHC) of Mohammedia, Morocco. A cross-sectional study was conducted from March 2017 to April 2018. Samples were collected by swabbing the hospital surfaces, and the isolated bacteria were checked for their susceptibility to antibiotics by the Kirby–Bauer disk diffusion method following the standards of the Clinical and Laboratory Standards Institute (CLSI). Among 200 swab samples, 176 (88%) showed bacterial growth. Gram-negative isolates were predominant at 51.5% (101/196), while the Gram-positives were at 48.5% (95/196). The main isolates are Enterobacteria weighted at 31.6% (62/196), Staphylococcus aureus reaching 24% (47/196), Pseudomonas aeruginosa at 9.2% (18/196), and Acinetobacter spp. with 3.3% (6/196). Moreover, the antimicrobial susceptibility profile of the isolates showed that about 31.7% (32/101) of the Gram-negative isolates were found to be MDR. This resistance is also high among isolates of S. aureus of which 44.7% (20/47) were methicillin-resistant Staphylococcus aureus (MRSA). Contamination of hospital surfaces by MDR bacteria is a real danger to public health. The concept of environmental bacterial reservoir is a reality that requires strict compliance with current guidelines and recommendations for hand hygiene, cleaning, and disinfection of surfaces in hospitals.

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Antibiotic Resistance of Uropathogens Isolated from Patients Hospitalized in District Hospital in Central Poland in 2020

Antibiotics ◽

10.3390/antibiotics10040447 ◽

2021 ◽

Vol 10 (4) ◽

pp. 447

Author(s):

Barbara Kot ◽

Agata Grużewska ◽

Piotr Szweda ◽

Jolanta Wicha ◽

Urszula Parulska

Keyword(s):

Antibiotic Resistance ◽

Multidrug Resistant ◽

Diffusion Method ◽

Disk Diffusion Method ◽

Beta Lactamases ◽

E Coli ◽

Central Poland ◽

Resistance Patterns ◽

Extended Spectrum Beta Lactamases ◽

Tract Infections

The aim of this study was to determine antibiotic resistance patterns and the prevalence of uropathogenes causing urinary tract infections (UTIs) in patients hospitalized in January–June 2020 in central Poland. Antimicrobial susceptibility testing was performed using the disk-diffusion method. Escherichia coli (52.2%), Klebsiella pneumoniae (13.7%), Enterococcus faecalis (9.3%), E. faecium (6.2%), and Proteus mirabilis (4,3%) were most commonly isolated from urine samples. E. coli was significantly more frequent in women (58.6%) (p = 0.0089) and in the age group 0–18, while K. pneumoniae was more frequent in men (24.4%) (p = 0.0119) and in individuals aged 40–60 and >60. Gram-negative species showed resistance to ampicillin. K. pneumoniae were resistant to amoxicillin plus clavulanic acid (75.0%), piperacillin plus tazobactam (76.2%), cefotaxime (76.2%), cefuroxime (81.0%), ciprofloxacin (81.0%), and trimethoprim plus sulphamethoxazole (81.0%). Carbapenems were effective against all E. coli and P. mirabilis. Some K. pneumoniae (13.6%) produced metallo-β-lactamases (MBLs). E. coli (22.6%), K. pneumoniae (81.8%), and all E. faecium were multidrug-resistant (MDR). Some E. coli (26.2%), K. pneumoniae (63.6%), and P. mirabilis (14.3%) isolates produced extended-spectrum beta-lactamases (ESBL). Vancomycin-resistant E. faecium was also found. This study showed that the possibilities of UTIs therapy using available antibiotics become limited due to the increasing number of antibiotic-resistant uropathogens.

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Detection of pathogenic bacteria associated with earphones used by students of Stamford University Bangladesh

Stamford Journal of Microbiology ◽

10.3329/sjm.v10i1.50723 ◽

2020 ◽

Vol 10 (1) ◽

pp. 1-4

Author(s):

Omor Ahmed Chowdhury ◽

Md Raihan Ahmed ◽

Md Raihan Dipu ◽

Md Aftab Uddin

Keyword(s):

Pathogenic Bacteria ◽

Diffusion Method ◽

Resistant Bacteria ◽

Bacterial Isolates ◽

Biochemical Tests ◽

Disk Diffusion Method ◽

Antibiotic Resistant ◽

Potential Source ◽

Different Types ◽

Staphylococcus Spp

The use of earphones has increased in recent times throughout the world especially among the different level of students such as school, college or university who have a higher tendency of sharing these among them. Unlike airline headsets, headphones and stethoscope ear-pieces, ear phones are often shared by multiple users and can be a potential medium for transmission of pathogens, which can give rise to various ear related infections. The objective of this study was to detect the pathogenic bacteria from the ear-phones used by the students of Stamford University Bangladesh. A total of 16 ear-phone swabs were collected by sterile cotton swabs. The swabs were inoculated onto blood agar and incubated aerobically overnight at 37oC. Microscopic observation and standard biochemical tests were performed to confirm the identification of all the bacterial isolates. Six presumptively identified Staphylococcus spp. (38%) were tested against six different types of antibiotics following Kirby-Bauer disk diffusion method. Isolates were found to be 84% resistant against Cotrimoxazole and demonstrated 100% sensitivity to Vancomycin and Ciprorofloxacin. The findings of this study suggest the users to disinfect their respective ear phones and not to exchange them as they may act as a potential source to transfer pathogenic and antibiotic resistant bacteria among the ear phone users. Stamford Journal of Microbiology, Vol.10 (1) 2020: 1-4

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The activity of different extracts from Panax quinquefolium L. cultures against pathogenic Staphylococcus aureus with respect to ginsenoside content

Archives of Biological Sciences ◽

10.2298/abs150330104s ◽

2015 ◽

Vol 67 (4) ◽

pp. 1277-1284 ◽

Cited By ~ 2

Author(s):

Monika Sienkiewicz ◽

Anna Głowacka ◽

Edward Kowalczyk ◽

Ewa Kochan

Keyword(s):

Staphylococcus Aureus ◽

Hairy Root ◽

Multidrug Resistant ◽

Clinical Isolates ◽

Diffusion Method ◽

Hairy Root Cultures ◽

Root Cultures ◽

Panax Quinquefolium ◽

Disk Diffusion Method ◽

Ginsenoside Content

Ginsenosides can be isolated from various cultures of Panax quinquefolium L., American ginseng. The aim of the study was to determine the antibacterial activity of extracts from leaves, stalks, hairy root cultures and field roots of P. quinquefolium L. containing ginsenosides against Staphylococcus aureus isolates obtained from various clinical materials. The agar well diffusion assay was used to evaluate microbial growth inhibition at various concentrations of extracts. The susceptibility of the clinical isolates to recommended antibiotics was determined with the disk-diffusion method. The results showed that the tested extracts inhibited the growth of all S. aureus clinical isolates, including MRSA (methicillin-resistant S. aureus) with MIC values ranging from 0.5 mg/mL to 1.7 mg/mL. The level of antimicrobial activity of extracts depends on the ginsenoside content. Both field roots and hairy root cultures represent excellent sources of these metabolites. Extracts with ginsenosides were found to inhibit multidrug-resistant staphylococci and can be a valuable complement to antistaphylococcal therapy.

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Prevalence, Risk Factors, and Antibiogram of Nontyphoidal Salmonella from Beef in Ambo and Holeta Towns, Oromia Region, Ethiopia

International Journal of Microbiology ◽

10.1155/2021/6626373 ◽

2021 ◽

Vol 2021 ◽

pp. 1-13

Author(s):

Endrias Zewdu Gebremedhin ◽

Gezahegn Tafesse Soboka ◽

Bizunesh Mideksa Borana ◽

Lencho Megersa Marami ◽

Edilu Jorga Sarba ◽

...

Keyword(s):

Risk Factors ◽

Food Safety ◽

Antimicrobial Susceptibility ◽

Multidrug Resistant ◽

Animal Origin ◽

Diffusion Method ◽

Isolation And Identification ◽

Disk Diffusion Method ◽

Cross Sectional ◽

Food Of Animal Origin

Background. Salmonella has been recognized as a major cause of food-borne illness associated with the consumption of food of animal origin. The present cross-sectional study was conducted from December 2017 to May 2018 in Ambo and Holeta towns to assess the prevalence, risk factors, and antimicrobial susceptibility patterns of nontyphoidal Salmonella isolates from raw beef samples from abattoirs, butchers, and restaurants in Ambo and Holeta towns, Oromia region, Ethiopia. Methods. A total of 354 beef samples were collected from abattoirs, butchers, and restaurants. Salmonella isolation and identification were carried out using standard bacteriological methods recommended by the International Organization for Standardization. Antimicrobial susceptibility testing was performed using the disk diffusion method. Besides, a structured questionnaire was used to collect sociodemographic data and potential risk factors for contamination of meat. Chi-square tests and logistic regression were used for data analyses. Results. Of the total 354 meat samples examined, 20 (5.7%, 95% confidence interval (CI): 3.5–8.6) were positive for Salmonella. Two serotypes belonging to S. typhimurium (11 isolates) and I:4,5,12: i:- (9 isolates) were identified. The Salmonella detection rate in abattoirs, butchers, and restaurants was 4.2% (5/118), 8.5% (10/118), and 4.2% (5/118), respectively. The antimicrobial susceptibility test showed that 40%, 30%, and 20% of the Salmonella isolates were resistant to azithromycin, amoxicillin, and ceftriaxone, respectively. The odds of Salmonella isolation when meat handlers are illiterate were 7.8 times higher than those when they are educated to the level of secondary and above ( P = 0.032 ). Similarly, the likelihood of Salmonella isolation was 6.3 and 7.6 times higher among workers of butcher and restaurants, respectively, who had no training ( P = 0.003 ) and no knowledge ( P = 0.010 ) on food safety and hygiene. Conclusions. The study showed widespread multidrug-resistant Salmonella isolates in the study areas. Therefore, raw meat consumption and indiscriminate use of antimicrobial drugs should be discouraged. Provision of food safety education for meat handlers and further surveillance of antimicrobial-resistant isolates are suggested.

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Synergistic effects of Persicaria odorata (Daun Kesom) leaf extracts with standard antibiotics on pathogenic bacteria

IIUM Medical Journal Malaysia ◽

10.31436/imjm.v16i2.321 ◽

2020 ◽

Vol 16 (2) ◽

Author(s):

P. M. Ridzuan ◽

Hairul Aini Hamzah ◽

Anis Shah ◽

Norazian Mohd Hassan ◽

Baharudin Roesnita

Keyword(s):

Pathogenic Bacteria ◽

Synergistic Effects ◽

Gas Chromatography Mass Spectrometry ◽

Diffusion Method ◽

Synergistic Activity ◽

Leaf Extracts ◽

Gram Positive ◽

Disk Diffusion Method ◽

Bacteria Growth ◽

Gram Positive Bacteria

Antibacterial activity of different types of P. odorata leaf extracts was evaluated in combination with standard antibiotics. Persicaria. odorata leaves were extracted with n-hexane (n-hex), dichloromethane (DCM) and methanol (MeOH). Each extract was applied on vancomycin (30µg), erythromycin (15µg) and gentamicin (10µg) discs, respectively. Disk diffusion method was used to evaluate the synergistic activity of each combination on Staphylococcus aureus, Staphylococcus epidermidis, Streptococcus pyogenes, Streptococcus pneumoniae, Pseudomonas aeruginosa, Salmonella typhi, and Escherichia coli. Minimum inhibitory concentration (MIC) and gas chromatography mass spectrometry (GCMS) analysis was performed on the active extract. Synergistic effects seen were mainly from the n-hex+antibiotics combinations, mainly on the Gram-positive bacteria (7 additive, 5 antagonistic), with MIC range from 50 µg/ml to 100 µg/ml, as well as Gram-negative bacteria (2 additive, 2 indifferent, 5 antagonistic). In particular, synergism showed by the combination of n-hex+van were all additive against the susceptible bacteria. DCM extract combination showed synergistic effects on three Gram-positive species (S. aureus, S. epidermidis, S. pyogenes). Meanwhile, MeOH+antibiotics combination showed significant additive synergistic effects (p<0.05) on S. aureus and S. epidermidis. The major compounds of leaves extract were decanal and β-citral. n-Hex extract superiorly inhibited Gram-positive bacteria growth as compared to DCM and MeOH extracts. The additive synergistic property of the n-hex P. odorata extract could be further studied for possible use as an antibacterial agent.

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Molecular Detection and Antibiotyping of Multidrug-Resistant Salmonella Isolated from Houseflies in a Fish Market

Pathogens ◽

10.3390/pathogens8040191 ◽

2019 ◽

Vol 8 (4) ◽

pp. 191 ◽

Cited By ~ 5

Author(s):

Sobur ◽

Hasan ◽

Haque ◽

Mridul ◽

Noreddin ◽

...

Keyword(s):

Molecular Detection ◽

Multidrug Resistant ◽

Diffusion Method ◽

Resistant Bacteria ◽

Salmonella Spp ◽

Disk Diffusion Method ◽

Antibiotic Resistant ◽

Fish Market ◽

Pcr Method ◽

High Level

Houseflies (Musca domestica) are well-known mechanical vectors for spreading multidrug-resistant bacteria. Fish sold in open markets are exposed to houseflies. The present study investigated the prevalence and antibiotypes of multidrug-resistant (MDR) Salmonella spp. in houseflies captured from a fish market. Direct interviews with fish vendors and consumers were also performed to draw their perceptions about the role of flies in spreading antibiotic-resistant bacteria. A total of 60 houseflies were captured from a local fish market in Bangladesh. The presence of Salmonella spp. was confirmed using PCR method. Antibiogram was determined by the disk diffusion method, followed by the detection of tetA, tetB, and qnrA resistance genes by PCR. From the interview, it was found that most of the consumers and vendors were not aware of antibiotic resistance, but reported that flies can carry pathogens. Salmonella spp. were identified from the surface of 34 (56.7%) houseflies, of which 31 (91.2%) were found to be MDR. This study revealed 25 antibiotypes among the isolated Salmonella spp. All tested isolates were found to be resistant to tetracycline. tetA and tetB were detected in 100% and 47.1% of the isolates, respectively. Among the 10 isolates phenotypically found resistant to ciprofloxacin, six (60%) were found to be positive for qnrA gene. As far as we know, this is the first study from Bangladesh to report and describe the molecular detection of multidrug-resistant Salmonella spp. in houseflies in a fish market facility. The occurrence of a high level of MDR Salmonella in houseflies in the fish market is of great public health concerns.

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