Identification Of Culturable Bacteria Research Articles

Distribution characteristics of antibiotic resistance in direct-eating food and analysis of Citrobacter freundii genome and pathogenicity

Purpose This study aims to study the distribution characteristics of antibiotic resistance in direct-eating food and analysis of Citrobacter freundii genome and pathogenicity. Residual antibiotics and antibiotic resistance genes (ARGs) in the environment severely threaten human health and the ecological environment. The diseases caused by foodborne pathogenic bacteria are increasing daily, and the enhancement of antibiotic resistance of pathogenic bacteria poses many difficulties in the treatment of disease. Design/methodology/approach In this study, six fresh fruits and vegetable samples were selected for isolation and identification of culturable bacteria and analysis of antibiotic resistance. The whole genome of Citrobacter freundii isolated from cucumber was sequenced and analyzed by Oxford Nanopore sequencing. Findings The results show that 270 strains of bacteria were identified in 6 samples. From 12 samples of direct food, 2 kinds of probiotics and 10 kinds of opportunistic pathogens were screened. The proportion of Citrobacter freundii screened from cucumber was significantly higher than that from other samples, and it showed resistance to a variety of antibiotics. Whole genome sequencing showed that Citrobacter freundii was composed of a circular chromosome containing signal peptides, transmembrane proteins and transporters that could induce antibiotic efflux, indicating that Citrobacter freundii had strong adaptability to the environment. The detection of genes encoding carbohydrate active enzymes is more beneficial to the growth and reproduction of Citrobacter freundii in crops. A total of 29 kinds of ARGs were detected in Citrobacter freundii, mainly conferring resistance to fluoroquinolones, aminoglycosides, carbapenem, cephalosporins and macrolides. The main mechanisms are the change in antibiotic targets and efflux pumps, the change in cell permeability and the inactivation of antibiotics and the detection of virulence factors and ARGs, further indicating the serious risk to human health. Originality/value The detection of genomic islands and prophages increases the risk of horizontal transfer of virulence factors and ARGs, which spreads the drug resistance of bacteria and pathogenic bacteria more widely.

Isolation and Identification of Culturable Bacteria from South China Seawater and Preliminary Screening of Marine Biocontrol Bacteria.

Marine microorganisms have evolved special metabolic pathways to produce numerous bioactive substances with novel structures and unique functions. This study analyzed the diversity of culturable bacteria in marine water samples from the South China Sea and screened the isolated bacteria with pathogenic fungi. A total of 200 culturable strains of 72 different bacteria were obtained from 56 water samples from the South China Sea. They belonged to three phyla and four classes, namely Gammaproteobacteria, Alphaproteobacteria, Bacilli and Actinomycetia. Bacilli was the dominant class, comprising up to 59.72%, followed by Gammaproteobacteria (20.83%). Bacillus, Pseudomonas, Paenibacillus and Rhizobium were the most dominant genera. Among these strains, HY-88 and HY-91 encoding BamC, FenB and PKSI genes were selected and identified as Bacillus subtilis. The respective inhibition rates of the HY-88 caused by plate confrontation against Magnaporthe grisea, Fusarium oxysporum, Botrytis cinerea, anthrax and Botrytis cinerea were 90.91%, 54.29%, 52.17% and 51.72%, in comparison with HY-91 86.36%, 48.57%, 47.83% and 34.48%. In addition, the supernatant of HY-88 showed a lesion inhibition rate of 74.5%, which was significantly higher than HY-91 (60.55%). In addition, HY-88 and HY-91 showed strong antifungal activity to Colletotrichum viniferum on detached Shine Muscat grapes. Tolerance tests showed that the HY-88 and HY-91 grew at 10-40 °C, 7-10% NaCl and pH 3-11. HY-88 and HY-91 could inhibit various fungal plant diseases, which lays a foundation for the development of new biopesticides.

An Efficient Micropropagation Protocol for the Endangered European Shrub February Daphne (Daphne mezereum L.) and Identification of Bacteria in Culture

Daphne mezereum of the Thymelaeaceae family is a medicinal shrub occurring naturally in Europe and under legal protection in Poland. In the present study, a protocol developed for mass propagation of February daphne from nodal explants is presented. Micropropagation is one of the in vitro techniques that allow the preservation of rare and valuable plants by developing efficient methods for their propagation. In the proliferation stage, explants were cultured in the Woody Plant Medium (WPM) with different cytokinins, and in the rooting stage on the semi-solid WPM medium with perlite, indole-3-butyric acid (IBA) in various concentrations was used. The maximum proliferation rate (five shoots per explant) was observed on the medium containing 4.14 μM of meta-Topolin (mT). When the regenerated shoots were rooted in vitro in the presence of IBA in various concentrations, that of 19.68 μM induced the highest number of roots per shoot (6.63) and the maximal root length (2.15 cm). It is also worth remembering that plants are often colonized by different groups of microorganisms, which also affect the diversity of the ecosystem. The endophytic bacteria inhabiting the D. mezereum shoots are Mycobacterium.

Bordetella pertussis, a reemerging pathogen in pediatric respiratory infections. A study in Quito, Ecuador

Bordetella pertussis is the causative agent of pertussis, which mainly affects unvaccinated children, while Bordetella parapertussis causes a disease presenting clinical characteristics that are indistinguishable from whooping cough. Despite high vaccination coverage, pertussis remains a public health concern worldwide, with approximately 140000 cases reported annually. Here we determined the prevalence of B. pertussis and B. parapertussis infection among infants under one year of age by polymerase chain reaction (PCR); our aim being to identify whether the data obtained relates to the relevant sociodemographic and clinical data. The study included 86 samples of nasopharyngeal swabs from infants aged between 0–12 months, who were reported as probable cases of whooping cough by the health centers around the Ecuadorian highlands, from August 2016 to July 2017. The nasopharyngeal swabs were cultured and microbiological and molecular analyses were performed. B. pertussis was identified by PCR in 41% of the samples (30/86), more than half of which corresponded to infants aged between 0–3 months. Moreover, a statistically significant correlation (p<0.05) between the identification of bacteria in culture and the catarrhal stage of the disease was observed. The results obtained from the study highlighted the need for an active national surveillance of pertussis, in particular for laboratory testing, to provide a highly sensitive and more specific diagnosis of Bordetella infection.

A retrospective analysis of cases of Spontaneous Bacterial Peritonitis in cirrhosis patients.

Spontaneous Bacterial Peritonitis (SBP) is an infection in patients with cirrhosis and carries significant mortality. The management of SBP is evolving with the rise of multidrug resistant organisms. Our aim was to perform a retrospective analysis to determine if identification of bacteria in culture could aid in prognosis and provide information regarding optimal treatment. We analyzed our 10-year experience of SBP in a single academic center (Northwestern Memorial Hospital). We obtained information regarding SBP prophylaxis, culture data and resistance patterns of bacteria, choice/duration of inpatient antibiotics, and key laboratory measurements and determined outcomes including mortality, hospital duration, and ICU stay. Patients with SBP had a 17.8% mortality and had culture positive SBP 34.4% of the time. Antimicrobial resistance was seen in 21.3% of cases and trended towards worsening mortality, with worsened mortality associated with first line use of piperacillin-tazobactam (p = 0.0001). Patients on SBP prophylaxis who developed SBP had improved mortality (p<0.0001) unless there was a positive culture, in which case patients had worsened mortality (p = 0.019). Patient with a higher PMN counts after repeat paracentesis had higher mortality (p = 0.02). Our results show that SBP continues to be a morbid and deadly condition and identification of an organism is key in treatment. The standard initial antibiotic for SBP may need to be modified to reflect emerging resistant pathogens and gram-positive organisms. Further, antibiotic prophylaxis should be utilized only in select cases to prevent development of resistance.

Isolation and Identification of Culturable Bacteria and Fungi from Mixed Dipterocarp and Mangrove Forests of Brunei Darussalam

Unique forests comprise of culturable microbes with unique properties that are of potential biotechnological uses. Considering that the pristine mixed dipterocarp forest and mangrove forest in Brunei Darussalam located in northwest Borneo in Southeast Asia are not well explored concerning their microbial diversity, this study aims to isolate and identify the culturable bacteria and fungi from these forest ecosystems. Bacteria and fungi were isolated using nutrient agar and potato dextrose agar, respectively. Microbial identification was done by DNA barcoding using 16S rRNA for bacteria and rRNA-ITS marker for fungi and supplemented by Biolog. A total of 16 bacterial and 19 fungal isolates were obtained which were identified to six bacterial and four fungal genera. Mixed dipterocarp forest harbored six genera of bacteria (Agrobacterium, Bacillus, Ralstonia, Staphylococcus, Burkholderia, and Cedecea), whereas mangrove forest showed two genera (Bacillus and Staphylococcus). Both types of forests showed three genera of fungi which were Trichoderma, Penicillium, and Aspergillus, with Saksenaea isolated only from mangrove forests. This study not only enhances the understanding of the diversity of culturable microbes in Brunei’s forest ecosystems, which has been less investigated till date but also of their ecological and biotechnological value.

Microbial life beyond the grave: 16S rRNA gene-based metagenomic analysis of bacteria diversity and their functional profiles in cemetery environments

Recent studies have identified cemeteries as potential environmental reservoirs of multi-drug resistant pathogenic bacteria that could contaminate groundwater sources posing public health threats. However, these findings were based on the identification of culturable bacteria and at times not below burial grounds. Investigation on the bacterial diversity and functional profiles of bacterial communities above and below burial grounds in human cemeteries are few. The current study used high-throughput sequencing techniques to determine the bacterial composition and their associated functional profiles in cemetery soil samples collected at the surface and below burial ground in two South African cemeteries (Maitland Cemetery in Cape Town and Fontein Street Cemetery in Middelburg) to evaluate the potential health threat to surrounding populations through contamination of groundwater. Significant differences were observed between sample depths with the clustering of the surface (0 m) and the 2 m samples into separate groups. Pseudomonas and Corynebacterium were the most abundant genera across all samples. Pseudomonas and Rhodococcus were the dominant genera in the 2 m samples while Prauserella and Staphylococcus were dominant in the surface samples. The 2 m samples showed a lower alpha diversity but recorded higher proportions of human diseases functional classes compared to the surface samples. Human disease functional profiles revealed involvement, in infectious (cholera), neurodegenerative (Alzheimer's disease) cardiovascular (hypertrophic cardiomyopathy) immune system (Systemic lupus erythematosus) metabolic (Type I & II diabetes) diseases and cancer. Antibiotic resistance and antibiotics synthesis signatures were also identified. Thus, cemeteries could be potential sources of microbial and antibiotic pollution in groundwater, especially in areas with shallow water tables such as Maitland. Selection of sites for use as cemeteries should, therefore, require a proper understanding of the hydrogeological characteristics of the selected site. However, further studies are required to trace the actual movement of these pollutants into groundwater resources.

Isolation and identification of culturable bacteria, capable of heterotrophic growth, from rapid sand filters of drinking water treatment plants

The microbial community in sand filters (SFs) of drinking water treatment plants (DWTPs) likely contributes to SF functionalities, such as organic carbon removal through heterotrophic metabolism. However, the dynamics and functionality of the SF microbiome and microbial communities in oligotrophic freshwater environments in general, are poorly understood. Therefore, the availability of bacterial strains from these oligotrophic environments is of great interest, but such organisms are currently underrepresented in culture collections. Focusing on heterotrophic carbon metabolism, bacteria were isolated from SFs using conventional media and media that contained SF extracts to mimic the SF environment. The majority of isolates belonged to Betaproteobacteria, more specifically to the families Comamonadaceae (genera Acidovorax, Curvibacter, Hydrogenophaga, Simplicispira, Paucibacter, Pelomonas, Piscinibacter and Rhodoferax) and Oxalobacteraceae (Undibacterium). Additionally, members of Alphaproteobacteria (Mesorhizobium), Gammaproteobacteria (Aeromonas and Perlucidibaca) and Actinobacteria (Rhodococcus and Brachybacterium) were isolated. Several of those genera have only rarely been described, but appear typical inhabitants of oligotrophic freshwater environments. In this regard, the Comamonadaceae isolates are of particular interest. Our study shows that bacteria representative of oligotrophic environments can be isolated using simple isolation procedures. The isolates provide a microbial framework for extending our knowledge of the taxonomy, physiology and functionality of oligotrophic freshwater microbiomes and their interactions with possible invaders.

16S rDNA based identification of culturable bacteria from highly polluted Varthur and Bellandur lakes of Bangalore, India

16S rDNA based identification of culturable bacteria from highly polluted Varthur and Bellandur lakes of Bangalore, India

Microbiological Analysis of Hemodialysis Water in a Developing Country.

Microbiological control of hemodialysis fluid is important for the prevention of hemodialysis-associated illness. Bacterial populations inhabiting a distribution system for hemodialysis water were studied over a 4 month period in five hospitals (one in Tehran, and the others at Alborz). All the samples from the four hospitals at Alborz had colony counts of ≥100 CFU/ml, which at different points of sampling were higher than the maximum recommended values. A total of 80 samples taken at different points in each hospital's hemodialysis distribution system were collected, and 229 planktonic bacteria isolated on R2A medium. No growth was detected by culturing the samples on Blood agar or Mueller-Hinton agar, according to routine procedures currently used in the five hospitals. A representative of isolates from each of 45 different morphotypes were identified using 16S RNA sequencing. A diverse bacterial community, containing predominantly gram-positive members of Kocuria, Arthrobacter and Staphylococcus and Mycobacterium, was detected. Bacteria from the genera Acinetobacter, Burkholderia, Halomonas, Herbaspirillum, Pseudomonas, and Sphingomonas were identified, which has been described in the build-up of biofilms. Some of the species reported here may represent a health risk to patients receiving hemodialysis treatment. In conclusion, it is recommended that standard protocols for evaluation of microbial contamination be used for regular monitoring and identification of culturable bacteria.

Isolation and identification of culturable bacteria from wild Anopheles culicifacies, a first step in a paratransgenesis approach.

BackgroundDue to the effect of midgut bacteria on proliferation of parasites and their potential as paratransgenesis tools, their identification in malaria vector mosquitoes is important. Anopheles culicifacies s.l. is one of the main malaria vectors in Asia; however, its midgut microbiota remains un-studied. This work was primarily designed to isolate potential candidates for use in a paratransgenesis approach, but also to give a picture of the midgut microbiota of wild-caught An. culicifacies larvae and adults from the southeast corner of Iran, which has the highest malaria endemicity in the country.MethodsA total of 68 larvae and 34 adult females (newly eclosed and older) from three different biotopes in Iran were analyzed for their midgut microflora. The mosquitoes had their midgut bacterial contents plated on three different culture media (brain heart agar, nutrient agar and blood agar) yielding 57 bacterial isolates. The 16S rRNA genes of the isolates were sequence analyzed for species designation, which then was confirmed by biochemical analysis.ResultsA total of twelve bacterial genera were identified: Acinetobacter, Aeromonas, Bacillus, Chryseobacterium, Delftia, Exiguobacterium, Kurthia, Microbacterium, Pseudomonas, Staphylococcus, Thorsellia and Variovorax. In older females, only Gram-negative bacteria were found, whereas larvae and newly-eclosed adults also harbored Gram-positive bacteria. The diversity of isolates also varied between sampling sites and mosquito stages, with the largest number of genera found in the Anguri district and in larvae, respectively. Pseudomonas was the most common genus retrieved from all sampling sites, and in both larvae and adults, suggesting a potential transstadial passage of these bacteria. Interestingly, identical 16S sequences of Pseudomonas were found in mosquitoes originating from different habitats at least 45 km apart, which could suggest that these bacteria have been adapted to the mosquitoes.ConclusionsThe study of vector mosquito microbiota has recently gathered increased interest because of the potential influence on vector competence. By adding data from a hitherto uncharacterized malaria mosquito, a better picture of gut flora in vector mosquitoes was obtained. Furthermore, some species of the predominant genus Pseudomonas will be evaluated for the selection of a paratransgenesis candidate.

Mobilization of manganese by basalt associated Mn(II)-oxidizing bacteria from the Indian Ridge System

The Indian Ridge System basalt bearing Mn-oxide coatings had todorokite as the major and birnesite as the minor mineral. We posit that microorganisms associated with these basalts participate in the oxidation of Mn and contribute to mineral deposition. We also hypothesized that, the Mn-oxidizing microbes may respond reversibly to pulses of fresh organic carbon introduced into the water column by mobilizing the Mn in Mn-oxides. To test these two hypotheses, we enumerated the number of Mn-oxidizers and -reducers and carried out studies on the mobilization of Mn by microbial communities associated with basalt. In medium containing 100μM Mn2+, 103 colony forming units (CFU) were recovered with undetectable number of reducers on Mn-oxide amended medium, suggesting that the community was more oxidative. Experiments were then conducted with basalt fragments at 4±2°C in the presence ‘G+’ and absence ‘G−’ of glucose (0.1%). Controls included set-ups, some of which were poisoned with 15mM azide and the others of which were heat-killed. The mobilization of Mn in the presence of glucose was 1.76μgg−1d−1 and in the absence, it was 0.17μgg−1d−1 after 150d. Mn mobilization with and without added glucose was 13 and 4 times greater than the corresponding azide treated controls. However, rates in ‘G+’ were 16 times and ‘G−’ 24 times more than the respective heat killed controls. The corresponding total counts in the presence of added glucose increased from 1.63×106 to 6.71×107 cellsg−1 and from 1.41×107 to 3.52×107cellsg−1 in its absence. Thus, the addition of glucose as a proxy for organic carbon changed the community’s response from Mn(II)-oxidizing to Mn(IV)-reducing activity. The results confirm the participation of Mn oxidizing bacteria in the mobilization of Mn. Identification of culturable bacteria by 16S rRNA gene analysis showed taxonomic affiliations to Bacillus, Exiguobacterium, Staphylococcus, Brevibacterium and Alcanivorax sp.

Decolorization of synthetic melanoidins-containing wastewater by a bacterial consortium

The presence of melanoidins in molasses wastewater leads to water pollution both due to its dark brown color and its COD contents. In this study, a bacterial consortium isolated from waterfall sediment was tested for its decolorization. The identification of culturable bacteria by 16S rDNA based approach showed that the consortium composed of Klebsiella oxytoca, Serratia mercescens, Citrobacter sp. and unknown bacterium. In the context of academic study, prevention on the difficulties of providing effluent as well as its variations in compositions, several synthetic media prepared with respect to color and COD contents based on analysis of molasses wastewater, i.e., Viandox sauce (13.5% v/v), caramel (30% w/v), beet molasses wastewater (41.5% v/v) and sugarcane molasses wastewater (20% v/v) were used for decolorization using consortium with color removal 9.5, 1.13, 8.02 and 17.5%, respectively, within 2 days. However, Viandox sauce was retained for further study. The effect of initial pH and Viandox concentration on decolorization and growth of bacterial consortium were further determined. The highest decolorization of 18.3% was achieved at pH 4 after 2 day of incubation. Experiments on fresh or used medium and used or fresh bacterial cells, led to conclusion that the limitation of decolorization was due to nutritional deficiency. The effect of aeration on decolorization was also carried out in 2 L laboratory-scale suspended cell bioreactor. The maximum decolorization was 19.3% with aeration at KLa=2.5836 h(-1) (0.1 vvm).

Soil microbial counts and identification of culturable bacteria in an extreme by arid zone

Sixteen samples of two soil cores (about 550 and 180 cm in depth) were drilled at intervals in the lower reach of Heihe river basin (northwest of China) in order to illustrate soil microbial characteristics and diversity of culturable bacteria in an extreme by arid environment. Soil water content, organic matter, total nitrogen, pH, direct cell counts, and culturable microorganism counts were evaluated. The total cell concentration was 19-1120/microg (i.e. 0.19-11.2 x 10(8) per g) soil, the culturable bacteria count being 0.2-10.9 per microg (i.e. 2 x 10(5)-10.9 x 10(6) CFU/g) soil. The number of direct cell counts obtained by 4',6-diamidino-2-phenylindole-staining or the cound of culturable microbes after enrichment with different media were statistically significantly correlated with soil organic matters, total nitrogen content, soil water content and surface vegetation; this partly explained the larger number in the deeper first core than in the shallower one. As part of identification of 228 colonies isolated from the two cores, thirty-two were selected for 16S rDNA amplification, sequencing and molecular identification. These 32 isolates were affiliated to 5 major groups of bacteria: alpha-Proteobacteria, 5-Proteobacteria, gamma-Proteobacteria, the high-G+C G+-bacteria, the low-G+C G- -bacteria, and the Cytophaga-Flexibacter-Bacteroides group. Twenty-eight were rod- or short-rod shaped, which accounted for >87.5% of all species; only 4 of 32 species were cocci (<12.5%).