Genus Klebsiella Research Articles

Phenotypic and Molecular Detection of BlaNDM Gene Among Drug-Resistant Klebsiella Isolates.

In the past few centuries, a widespread increase in antimicrobial resistance has been observed among Klebsiella species. The antibiotic- resistant strains of the genus Klebsiella are becoming a serious threat in clinical settings due to their involvement in severe invasive and non-invasive infections. The emergence of resistance among these strains is associated with their strong enzymatic activity against several broad-spectrum antibiotics. These enzymes include beta-lactamases, extended-spectrum beta-lactamases (ESBL), AmpC beta-lactamases, and carbapenemases. These resistance enzymes are capable of hydrolyzing various broad-spectrum drugs like extended-spectrum cephalosporin and carbapenems. The present study was conducted to determine the emerging resistance among Klebsiella strains by identifying the production of carbapenemase enzyme phenotypically and the frequency of the NDM resistance gene by a polymerase chain reaction. In this study, 236 Gram-negative isolates from different clinical laboratories were identified. Out of which, 125 isolates were found as Klebsiella species by using standard microbiological techniques. Minimum inhibitory concentrations (MIC) were determined using eight representative antibiotics by the Macro broth dilution method. Phenotypic detection of carbapenemase producing Klebsiella species was performed by Modified Hodge Test. Phenotypic findings were then checked and compared with genotypic results obtained by using the Polymerase chain reaction (PCR) for the detection of resistance genes responsible for the production of carbapenemase. In this study, carbapenemase production was found only in 6 (5%) Klebsiella isolates by using the phenotypic method; however, 3 isolates out of 125 were screened positive for the gene NDM-1. Since we are considering carbapenems as the last therapeutic option for treating infections, mainly caused by Gram-negative isolates, the prevailing resistance against this drug is widely disseminating. It is better to evaluate the antibiotic susceptibility, phenotypic screening as well genotypic screening (where possible) for implementing strict antibiotic control policies in health care settings, hospitals, laboratories, etc.

Gut Microbiota Composition of Biliary Atresia Patients Before Kasai Portoenterostomy Associates With Long-term Outcome.

ABSTRACTBackground and Aims:Biliary atresia (BA) is a cholestatic, fibro-obliterative cholangiopathy of unknown etiology. BA is primarily treated by a surgical approach, that is, the Kasai portoenterostomy (KPE), to obtain clearance of jaundice (COJ). The gut microbiota (GM) composition has been associated with the course of several cholestatic liver diseases. It is largely unknown, however, whether GM composition associates with the outcome of KPE. We compared the GM composition of BA patients and controls and assessed if GM composition before KPE was related to COJ after KPE.Methods:We compared feces of term-born BA patients before KPE and controls (patients undergoing inguinal hernia repair) by 16S rRNA sequencing. Composition and alpha diversity of the GM were compared between BA and controls before KPE and after KPE, between patients with COJ versus without COJ (total serum bilirubin < or ≥20 μmol/L <6 months post-KPE).Results:Alpha diversity was comparable between BA (n = 12, age 1.6 [1.3–1.8] months) and controls (n = 6, age 2.0 [1.4–2.1] months; P = 0.22). Compared with controls, BA patients had lower abundances of Bifidobacteriaceae (β = −1.98, P < 0.001) and Lachnospiraceae (β = −1.84, P = 0.007), and higher abundances of Streptococcus (β = −1.13, P = 0.003). The alpha diversity before KPE correlated negatively with COJ (R = −0.63, P = 0.03). Lower alpha diversity pre-KPE was associated with COJ [+] (βlogit = −0.64, P = 0.04). We observed greater abundances of genus Acinetobacter (β = 1.27, P = 0.03) and family Clostridiaceae (β = 1.45, P = 0.03) and lower abundances of the family Enterobacteriaceae (genera Klebsiella (β = −1.21, P = 0.01), Salmonella (β = −1.57, P = 0.02)) in COJ [+] versus COJ [−].Conclusions:The GM of BA patients before Kasai portoenterostomy associates with outcome, clearance of jaundice, suggestive of predictive, and mechanistic roles of the gut microbiota composition in bile homeostasis.

Halotolerant and metalotolerant bacteria strains with heavy metals biorestoration possibilities isolated from Uburu Salt Lake, Southeastern, Nigeria

Environmental contaminations by heavy metals are currently an increasing public health concern globally. One key challenge of these toxic metals is the extremely difficulties involved in their detoxification from the environment and effluents because of their non-degradability. An efficient biologic agent with potentials of remediating these toxic metals may ease these ever-increasing problems. We reported toxic metals tolerance and bioremediation potentials of novel bacteria sp. Strains USL2S, USL4W and USL5W isolated from Uburu salt lake, Ebonyi State, Nigeria. The phenotypic characteristics and the 16S rRNA gene analyses revealed that USL2S strain belongs to the genus Klebsiella, whereas USL4W and USL5W strains belong to the genus Pseudomonas. The bacteria isolates grew well in media containing 5–15 % of sodium chloride. The bacteria isolate showed capacity to tolerate 50.0 mM Hg+2 and Pb+2, 17.0, 12.50 and 4.0mM Ni+2, Cd+2, and Zn+2 respectively in solid media. Pseudomonas putida A4W Strain also tolerated 16.0 mM Cu+2, while Klebsiella sp. Strain USL2S, Pseudomonas putida USL5W Strain tolerated 4.0 mM each. AAS analyses showed 85, 95, and 95 % Hg; 97.13, 98.89, and 97.55 % Pb; 73.33, 77.42 and 69.72 Cd; 88.06, 99.54, and 97.91 % Ni; 100, 100 and 83.62% Cu; 42.30, 84.52 and 98.80 % Zn removal from media broth incorporated with the tested metals by Klebsiella sp. USL2S, Pseudomonas sp. Strain USL5W and Pseudomonas sp. USL4W respectively. We therefore recommend these novel moderately halophilic and metal tolerant isolates as possible biologic agents for effective bioremediation of mercury, lead, cadmium, nickel, copper and zinc in contaminated environments and effluents.

Exploring the Global Spread of Klebsiella grimontii Isolates Possessing blaVIM-1 and mcr-9.

The spread of plasmid-mediated carbapenemases within Klebsiella oxytoca is well-documented. In contrast, data concerning the closely related species Klebsiella grimontii are scarce. In fact, despite the recent report of the first blaKPC-2-producing K. grimontii, nothing is known about its clonality and antibiotic resistance patterns. In a retrospective search in our collection, we identified 2 blaVIM-positive K. oxytoca strains. Whole-genome sequencing with both Illumina and Nanopore indicated that our strains actually belonged to K. grimontii and were of sequence type 172 (ST172) and ST189. Moreover, the two strains were associated with 297-kb IncHI2/HI2A-pST1 and 90.6-kb IncFII(Yp) plasmids carrying blaVIM-1 together with mcr-9 and blaVIM-1, respectively. In the IncHI2/HI2A plasmid, blaVIM-1 was located in a class 1 integron (In110), while mcr-9 was associated with the qseC-qseB-like regulatory elements. Overall, this plasmid was shown to be very similar to those carried by other Enterobacterales isolated from food and animal sources (e.g., Salmonella and Enterobacter spp. detected in Germany and Egypt). The IncFII(Yp) plasmid was unique, and its blaVIM-1 region was associated with a rare integron (In1373). Mapping of In1373 indicated a possible origin in Austria from an Enterobacter hormaechei carrying a highly similar plasmid. Core-genome phylogenies indicated that the ST172 K. grimontii belonged to a clone of identical Swedish and Swiss strains (≤15 single nucleotide variants [SNVs] to each other), whereas the ST189 strain was sporadic. Surveillance of carbapenemase-producing K. oxytoca strains should be reinforced to detect and prevent the dissemination of new species belonging to the Klebsiella genus.

Proposal for reunification of the genus Raoultella with the genus Klebsiella and reclassification of Raoultella electrica as Klebsiella electrica comb. nov.

The order Enterobacterales was divided into seven families including the family Enterobacteriaceae in 2016. The genus Klebsiella within the family Enterobacteriaceae was divided into two genera Klebsiella and Raoultella in 2001. Here, our phylogenomic analysis shows that the genus Raoultella is nested within the genus Klebsiella. Klebsiella and Raoultella together are monophyletic and share average amino acid identities (AAIs) of 86.9–89.6% above the AAI threshold (86%) for genus delimitation within the family Enterobacteriaceae. Klebsiella and Raoultella share AAIs of 79.9%–85.0% with the other genera within the subfamily “Klebsiella clade”, which are in the range of inter-genus AAIs (74–85%) within the family Enterobacteriaceae. Klebsiella and Raoultella also share six known conserved signature indels. Therefore, we propose to reunify Klebsiella and Raoultella to the single genus Klebsiella and reclassify Raoultella electrica as Klebsiella electrica comb. nov. Our genome-based taxonomic analyses also identified seven potential novel species within the unified genus Klebsiella.

Clinical characteristics and outcome of bacteraemia caused by Enterobacter cloacae and Klebsiella aerogenes: more similarities than differences

The genus Enterobacter is a common cause of nosocomial infections. Historically, the most frequent Enterobacter species were those of Enterobacter cloacae complex and Enterobacter aerogenes. In 2019, E. aerogenes was re-classified as Klebsiella aerogenes owing to its higher genotypic similarity with the genus Klebsiella. Our objective was to characterise and compare the clinical profiles of bacteraemia caused by E. cloacae and K. aerogenes. This 3-year multicentre, prospective cohort study enrolled consecutive patients with bacteraemia by E. cloacae or K. aerogenes. Baseline characteristics, bacteraemia features (source, severity, treatment), antibiotic susceptibility, resistance mechanisms and mortality were analysed. The study included 285 patients with bacteraemia [196 (68.8%) E. cloacae and 89 (31.2%) K. aerogenes]. The groups showed no differences in age, sex, previous use of invasive devices, place of acquisition, sources or severity at onset. The Charlson score was higher among patients with E. cloacae bacteraemia [2 (1-4) vs. 1 (0.5-3); P=0.018], and previous antibiotic therapy was more common in patients with K. aerogenes bacteraemia (57.3% vs. 41.3%; P=0.01). Mortality was 19.4% for E. cloacae and 20.2% for K. aerogenes (P = 0.869). Antibiotic susceptibility was similar for both species, and the incidence of multidrug resistance or ESBL production was low (6% and 5.3%, respectively), with no differences between species. Bacteraemias caused by E. cloacae and K. aerogenes share similar patient profiles, presentation and prognosis. Patients with E. cloacae bacteraemia had more co-morbidities and those with K. aerogenes bacteraemia had received more antibiotics.

Evaluation of cellulose degrading bacteria isolated from the gut-system of cotton bollworm, Helicoverpa armigera and their potential values in biomass conversion

Background Cotton bollworm, Helicoverpa armigera is a widely distributed, devastating pest of over 200 crop plants that mainly consist of some cellulosic materials. Despite its economic importance as a pest, little is known about the diversity and community structure of gut symbiotic bacteria potentially functioned in cellulose digestion in different gut-sections of H. armigera. In view of this lacuna, we attempted to evaluate and characterize cellulose-degrading bacteria (CDB) from foregut, midgut, and hindgut -regions of H. armigera by using a culture-dependent approach. Methodology The symbiotic bacteria were isolated from different gut-systems of H. armigera by enrichment techniques using Carboxymethyl cellulose sodium salt (CMC) as carbon source. The isolated bacteria were purified and subsequently screened for cellulose-degradation by plate-based method to display the zones of CMC clearance around the colonies. The identification and phylogeny of the gut-bacteria were reconstructed by using 16S rRNA gene sequencing. Different enzymes such as endoglucanase, exoglucanase, β-glucosidase, and xylanase were assayed to determine the cellulolytic repertoire of the isolated bacteria. Results The enrichment of CDB and subsequent plate based screening methods resulted in isolation of 71 bacteria among which 54% of the bacteria were obtained from foregut. Among the isolated bacteria, 25 isolates showed discernible cellulose-degradation potential on CMC-agar plates. The phylogenetic analysis based on 16S rRNA gene amplification and sequencing affiliated these cellulolytic bacteria to two major phyla viz., Firmicutes and Proteobacteria. The members of the genus Klebsiella accounted for 39.43% of the total isolated bacteria while 31% of the Bacillus strains were enriched from hindgut region. The principal component analysis (PCA) further suggested that the members of Bacillus and Klebsiella together dominated the foregut and hindgut regions as they accounted for 68% of the total CDB. The four potential isolates selected on the basis of plate-based activities were further evaluated for their lignocellulases production by using various agricultural wastes as substrates. The PCA of the enzyme activities demonstrated that potential isolates majorly secreted endoglucanase and xylanase enzymes. Among the agro-wastes, multivariate analysis validated wheat husk (WH) and sugarcane bagasse (SCB) as most favorable substrates for xylanase and endoglucanase productions respectively. The overall findings suggest that H. armigera harbors diverse bacterial communities in different gut-sections that could assist the host in digestion processes, which may potentially serve as a valuable reservoir of some unique symbionts applied for biomass conversion in biofuel industry.

A Comparative Analysis of Virulence Factors in Klebsiella Species

Klebsiella is a genus of bacteria frequently associated with nosocomial infections due to their multi-drug resistant properties. Klebsiella pneumoniae accounts for a large majority of hospital-acquired pneumonia, urinary tract infections, and septicemia. The pathogenicity of K. pneumoniae clinical isolates has been greatly established to better understand and treat these infections. Klebsiella oxytoca is also primarily healthcare associated and is considered to be an emerging nosocomial pathogen, as it causes the second most frequent infections from the Klebsiella genus. Although it shares many characteristics with K. pneumoniae, its virulence factors have not been well studied due to its lesser degree of incidence. As an emerging pathogen, more information must be gathered in order to learn how to best treat this infectious, antibiotic-resistant bacteria. This study aims to first determine if the pathogenic factors found in clinical Klebsiella pneumoniae isolates are also present in Klebsiella oxytoca and then compare the production of these virulence factors under normal and inducing conditions. This work compares the expression of pathogenic factors in five isolates of K. oxytoca, three isolates of K. pneumoniae, and one strain of Escherichia coli. The pathogenic factors chosen for this study include antibiotics resistance, siderophore production, biofilm formation abilities, and reactive oxygen species (ROS) detoxification. To determine antibiotic resistance, the minimum inhibitory concentration of nine different antibiotics was calculated for each isolate. Siderophore production was measured using a Chrome Azurol Sulfonate colorimetric test in both the absence and presence of 2,2’-dipyridyl, an iron chelator. Biofilm formation was detected with Crystal Violet dye in both the absence and presence of acetaminophen and hydrochlorothiazide, both of which are commonly used therapeutic pharmaceuticals in nosocomial settings, as well as 2,2’-dipyridyl. ROS detoxification via superoxide dismutase enzyme activity was measured through native gel activity assays. Our findings indicate that K. pneumoniae has resistance to more antibiotics than K. oxytoca, but the two species have comparable siderophore production, biofilm formation capabilities, and ROS detoxification capabilities. In conclusion, Klebsiella pneumoniae and Klebsiella oxytoca share several mechanisms of pathogenicity under normal and inducing conditions and drug resistance is the most notable difference among the strains tested in this work.

Dissemination and characteristics of Klebsiella spp. at the processed cheese plant

The genus Klebsiella is not generally considered as a major foodborne pathogen and with regard to food production it represents also a hygiene indicator. This study was focused on the monitoring of Klebsiella spp. dissemination at the processed cheese plant and determination of virulence determinants and antibiotic resistance of obtained strains. Klebsiella spp. were detected in 43% (37/87) of samples: swabs from the cheese processing environment (34/69), personnel (2/3) and raw material (milk powder) from an opened packaging (1/7). All tested samples of final processed cheeses were negative for the presence of Klebsiella spp. Obtained strains were identified as K. oxytoca (n = 32) and K. pneumoniae (n = 23). Typing results enabled to reveal the presence of two predominant PFGE clusters of K. pneumoniae and K. oxytoca suggesting the occurrence of suspect persistent strains, and spread of Klebsiella spp. between the production environment and the personnel. The melting process of processed cheese production was able to eliminate Klebsiella spp. in the final products, although some K. oxytoca and K. pneumoniae strains carried genes of the locus of heat resistance 1 (LHR1), which may lead to their increased heat resistance. K. pneumoniae and K. oxytoca isolated from the processed cheese plant did not represent any hypervirulent or multidrug-resistant strains which could be a potential threat to public health.

Carboniferous Blattinopsidae: revision of Klebsiella and new genus and species from Avion (Insecta, Paoliida)

ABSTRACT The genus Klebsiella Meunier, 1908 from the latest Carboniferous of Commentry, is revised, confirming its attribution to the Blattinopsidae. The family name Klebsiellidae should have priority on its junior synonym Blattinopsidae, but the common usage over time could allow maintaining the later. The first Blattinopsidae from the Moscovian Konservat–lagerstätte of Avion is described as a new genus and species Avionblattinopsis oudardi gen. et n. sp. on the basis of a single forewing. It differs from the other genera of this family in the vein ScP distally fused to the vein RA. It increases our knowledge about this family, known between the Late Carboniferous and the Middle Permian. An emended family diagnosis is proposed. Protoblattiniella minutissima Meunier, 1912, based on a mature nymph would better fit with the Paoliidae than with the Blattinopsidae.

Heavy Metals in Soils and the Remediation Potential of Bacteria Associated With the Plant Microbiome

High concentrations of non-essential heavy metals/metalloids (arsenic, cadmium, and lead) in soils and irrigation water represent a threat to the environment, food safety, and human and animal health. Microbial bioremediation has emerged as a promising strategy to reduce the concentration of heavy metals in the environment due to the demonstrated ability of microorganisms, especially bacteria, to sequester and transform these compounds. Although several bacterial strains have been reported to be capable of remediation of soils affected by heavy metals, published information has not been comprehensively analyzed to date to recommend the most efficient microbial resources for application in bioremediation or bacterial-assisted phytoremediation strategies that may help improve plant growth and yield in contaminated soils. In this study, we critically analyzed eighty-five research articles published over the past 15 years, focusing on bacteria-assisted remediation strategies for the non-essential heavy metals, arsenic, cadmium, and lead, and selected based on four criteria: i) The bacterial species studied are part of a plant microbiome, i.e., they interact closely with a plant species ii) these same bacterial species exhibit plant growth-promoting characteristics, iii) bacterial resistance to the metal(s) is expressed in terms of the Minimum Inhibitory Concentration (MIC), and iv) metal resistance is related to biochemical or molecular mechanisms. A total of sixty-two bacterial genera, comprising 424 bacterial species/strains associated with fifty plant species were included in our analysis. Our results showed a close relationship between the tolerance level exhibited by the bacteria and metal identity, with lower MIC values found for cadmium and lead, while resistance to arsenic was widespread and significantly higher. In-depth analysis of the most commonly evaluated genera, Agrobacterium, Bacillus, Klebsiella, Enterobacter, Microbacterium, Pseudomonas, Rhodococcus, and Mesorhizobium showed significantly different tolerance levels among them and highlighted the deployment of different biochemical and molecular mechanisms associated with plant growth promotion or with the presence of resistance genes located in the cad and ars operons. In particular, the genera Klebsiella and Enterobacter exhibited the highest levels of cadmium and lead tolerance, clearly supported by molecular and biochemical mechanisms; they were also able to mitigate plant growth inhibition under phytotoxic metal concentrations. These results position Klebsiella and Enterobacter as the best potential candidates for bioremediation and bacteria-assisted phytoremediation strategies in soils contaminated with arsenic, cadmium, and lead.

Granuloma Inguinale

Granuloma ingunale (GI) or donovanosis is a genital ulcer disease caused by theCalymmatobacterium granulomatis. It is a Gram-negative, facultative, obligateintracellular and pleomorphic bacterium. This bacterium has phylogeneticallyclosed to and placed within the Klebsiella genus. Clinically, the disease is com-monly characterized as painless, slowly progressive ulcerative lesions on thegenitals or perineum without regional lymphadenopathy. The lesions are highlyvascular and bleed easily on contact Extragenital lesions may occur but are rareand more common in newborns from mothers with GI genital lesions. Thisdisease is often neglected, therefore it is often misdiagnosed and inaccuratetherapy. Treatment time is 3 weeks or until clinical cure has been achieved forall proposed regimens. It often occurs both in men and women of reproductiveage (20-40 years). This article consists of several theoretical references that havebeen viewed to have a better understanding of GI.

Granuloma Inguinale

Granuloma ingunale (GI) or donovanosis is a genital ulcer disease caused by theCalymmatobacterium granulomatis. It is a Gram-negative, facultative, obligateintracellular and pleomorphic bacterium. This bacterium has phylogeneticallyclosed to and placed within the Klebsiella genus. Clinically, the disease is com-monly characterized as painless, slowly progressive ulcerative lesions on thegenitals or perineum without regional lymphadenopathy. The lesions are highlyvascular and bleed easily on contact Extragenital lesions may occur but are rareand more common in newborns from mothers with GI genital lesions. Thisdisease is often neglected, therefore it is often misdiagnosed and inaccuratetherapy. Treatment time is 3 weeks or until clinical cure has been achieved forall proposed regimens. It often occurs both in men and women of reproductiveage (20-40 years). This article consists of several theoretical references that havebeen viewed to have a better understanding of GI.

In vitro colonic fermentation of a plant sterol-enriched beverage in a dynamic-colonic gastrointestinal digester

The impact of a plant sterol-enriched beverage on the sterol metabolism, organic acid production and microbiota composition was evaluated by means of a dynamic gastrointestinal and colonic fermentation model. After one week of fermentation, an absence of sterol metabolites was reported, in accordance with the lack of microbiota related to their metabolism. Although total organic acid content was lower in the ascending colon (AC) compared to the transversal (TC) and descending colon (DC) (28–57 mmol/L vs. 55-87 and 44–64 mmol/L, respectively), its increments, with respect to the initial value, were higher (2-fold vs. 1.6- and 1.5-fold). Increments of acetate, butyrate and propionate were observed in all colon vessels, whereas lactate production was only observed in the AC at the first hours of fermentation. Results showed a progressive increase in microbial species richness and evenness from the AC to the DC. Beverage fermentation also modulated certain members of the microbiota: Bifidobacterium (within 24 h) and Megasphaera genera were stimulated, and Mitsuokella was reduced in AC, while the growth of Klebsiella genus was promoted and Bacteroidetes phylum decreased in TC and DC. These results show that the beverage modulates the microbial community and activity, which could be physiologically relevant to the host.

Distribution of Beta-Lactamase Producing Gram-Negative Bacterial Isolates in Isabela River of Santo Domingo, Dominican Republic.

Bacteria carrying antibiotic resistance genes (ARGs) are naturally prevalent in lotic ecosystems such as rivers. Their ability to spread in anthropogenic waters could lead to the emergence of multidrug-resistant bacteria of clinical importance. For this study, three regions of the Isabela river, an important urban river in the city of Santo Domingo, were evaluated for the presence of ARGs. The Isabela river is surrounded by communities that do not have access to proper sewage systems; furthermore, water from this river is consumed daily for many activities, including recreation and sanitation. To assess the state of antibiotic resistance dissemination in the Isabela river, nine samples were collected from these three bluedistinct sites in June 2019 and isolates obtained from these sites were selected based on resistance to beta-lactams. Physico-chemical and microbiological parameters were in accordance with the Dominican legislation. Matrix-assisted laser desorption ionization-time of flight mass spectrometry analyses of ribosomal protein composition revealed a total of 8 different genera. Most common genera were as follows: Acinetobacter (44.6%) and Escherichia (18%). Twenty clinically important bacterial isolates were identified from urban regions of the river; these belonged to genera Escherichia (n = 9), Acinetobacter (n = 8), Enterobacter (n = 2), and Klebsiella (n = 1). Clinically important multi-resistant isolates were not obtained from rural areas. Fifteen isolates were selected for genome sequencing and analysis. Most isolates were resistant to at least three different families of antibiotics. Among beta-lactamase genes encountered, we found the presence of blaTEM, blaOXA, blaSHV, and blaKPC through both deep sequencing and PCR amplification. Bacteria found from genus Klebsiella and Enterobacter demonstrated ample repertoire of antibiotic resistance genes, including resistance from a family of last resort antibiotics reserved for dire infections: carbapenems. Some of the alleles found were KPC-3, OXA-1, OXA-72, OXA-132, CTX-M-55, CTX-M-15, and TEM-1.

Characterization of four virulent Klebsiella pneumoniae bacteriophages, and evaluation of their potential use in complex phage preparation

BackgroundNowadays, hundreds of thousands of deaths per year are caused by antibiotic resistant nosocomial infections and the prognosis for future years is much worse, as evidenced by modern research. Bacteria of the Klebsiella genus are one of the main pathogens that cause nosocomial infections. Among the many antimicrobials offered to replace or supplement traditional antibiotics, bacteriophages are promising candidates.MethodsThis article presents microbiological, physicochemical and genomic characterization of 4 virulent bacteriophages belonging to Siphoviridae, Myoviridae and Podoviridae families. Phages were studied by electron microscopy; their host range, lytic activity, adsorption rate, burst size, latent period, frequency of phage-resistant forms generation, lysis dynamics and sensitivity of phage particles to temperature and pH were identified; genomes of all 4 bacteriophages were studied by restriction digestion and complete genome sequence.ResultsStudied phages showed wide host range and high stability at different temperature and pH values. In contrast with single phages, a cocktail of bacteriophages lysed all studied bacterial strains, moreover, no cases of the emergence of phage-resistant bacterial colonies were detected. Genomic data proved that isolated viruses do not carry antibiotic resistance, virulence or lysogenic genes. Three out of four bacteriophages encode polysaccharide depolymerases, which are involved in the degradation of biofilms and capsules.ConclusionsThe bacteriophages studied in this work are promising for further in vivo studies and might be used in phage therapy as part of a complex therapeutic and prophylactic phage preparation. The conducted studies showed that the complex preparation is more effective than individual phages. The use of the complex phage cocktail allows to extend the lytic spectrum, and significantly reduces the possibility of phage-resistant forms generation.

Isolation, Characterization and Activity Test of Soil Origin Bacteria Amilage

This study aimed to obtain amylase-producing potent bacteria from soil and test the amylase activity produced. Soil samples were taken from the Biological Education and Research Forest, Andalas University. The isolation was done by using the stratified dilution technique on agar media. The screening of amylase activity employed the qualitative and quantitative tests on agar starch. From 8 isolated amylolytic bacteria, there were three isolates with amylolytic potential. The results of characterization and identification based on Bergey's Manual of Determinative Bacteriology show that isolates A4, A1, and A6 belonged to the genus Bacillus, Corynebacterium, and Klebsiella. The bacteria obtained can then be produced and optimized for the needs of industrial enzymes.

Multidrug-resistant Klebsiella quasipneumoniae subsp. similipneumoniae carrying blaNDM-blaCTX-M15 isolated from flies in Rio de Janeiro, Brazil

ObjectivesFlies have been implicated in the dispersal of medically important bacteria including members of the genus Klebsiella between different environmental compartments. The aim of this study was to retrieve and characterize antibiotic-resistant bacteria from flies collected near to hospitals. MethodsFlies were collected in the vicinity of medical facilities and examined for bacteria demonstrating phenotypic resistance to ceftriaxone, followed by determination of phenotypic and genotypic resistance profiles. In addition, whole genome sequencing followed by phylogenetic analysis and resistance genotyping were performed with the multidrug-resistant (MDR) strain Lemef23, identified as Klebsiella quasipneumoniae subsp. similipneumoniae. ResultsThe strain Lemef23, classified by multiple locus sequence typing as novel ST 3397, harboured numerous resistance genes. The blaNDM was located on a Tn3000 element, a common genetic platform for the carriage of this gene in Brazil. Inference of phylogenetic orthology of strain Lemef23 and other clinical isolates suggested an anthropogenic origin. ConclusionsThe findings of this study support the role of flies as vectors of MDR bacteria of clinical importance and provide the first record of blaNDM-1 and blaCTXM-15 in a Brazilian isolate of K. quasipneumoniae subsp. similipneumoniae, demonstrating the value of surveying insects as reservoirs of antibiotic resistance.

Description of the ovarian microbiota of Aedes aegypti (L) Rockefeller strain

Aedes aegypti is one of the vectors responsible for transmitting the viruses that cause dengue, Zika and chikungunya in the human population. Mosquitoes have bacterial communities in different organs, mainly in the midgut, but to a lesser extent in their reproductive organs, such as the ovaries, where replication and vertical transmission is decisive for dengue virus. These bacteria also influence metabolic and physiological processes such as ingestion and digestion of blood. In this study, aerobic bacterial communities associated with ovaries of A. aegypti Rockefeller strain were determined, describing their potential function during ovocitary development. The groups of mosquitoes were separated into three treatments: diet with 10% sugar solution, diet with blood supply, and blood feeding combined with tetracycline. The ovaries were extracted from the mosquitoes, and then put in enriched culture media (blood and nutritive agar) by direct inoculation, for subsequent isolation and macroscopic and microscopic characterization of the colonies. The taxonomic determination of bacterial isolates was achieved by sequence analysis of the 16S rRNA gene. A higher bacterial load was observed in the sugar feeding group (6 × 10³ CFU/ml) in contrast to the group fed only with blood, with and without an antibiotic (4.03-4.04 × 10³CFU/ml; 4.85-5.04 × 10³CFU/ml). As a result, a total of 35 colonies were isolated, of which 80% were gram-negative and 20% gram-positive; 72% were lactose negative and 8% lactose positive. Of the total bacteria, 83% had gamma hemolysis, 17% alpha hemolysis, and none presented beta hemolysis. After phenotypic and biochemical characterization, 17 isolates were selected for molecular identification. Only phyla Actinobacteria and Proteobacteria were found. Bacteria associated with ovaries of A. aegypti were mainly identified as belonging to the Serratia and Klebsiella genera. Some bacteria (Serratia marcescens, Pantoea dispersa and Klebsiella oxytoca) have wide biotechnological potential due to their entomopathogenic power and their bioactivity against different pathogens.

Application of biochemical typing in veterinary medicine in bee enterobacterioses to determine Klebsiella Pneumoniae

The article presents laboratory diagnostics (in vitro), namely, identification of pure culture of pathogenic bacteria of Klebsiella Pneumoniae species in case of enterobacteriosis in bees in winter-spring and summer-autumn times. The purpose of the study was the biochemical typification of bacteria of the species Klebsiella Pneumoniae with humane medicine methods, that isolated in the case of dysbacteriosis of bees which have a characteristic symptomatic complex of a decrease in the strength of bee families, which leads to a decrease in the resistance of the bee family, their diarrhea, crawling, and then swarming or death of bees. Contamination of beehive frames and walls of beehive by fecation leads to sharp deterioration of the apiary's veterinary and sanitary condition and significant economic damage for beekeepers. Pure culture of pathogenic bacteria served as an object for experiment. The Family of the bacteria was established earlier – Enterobacteriaceae, and was confirmed by “Zhytomyr Regional State Laboratory of the State Service of Ukraine for Food Safety and Consumer Protection”. Laboratory diagnostics of dysbiosis in bees caused by enterobacteria was performed in such a sequence: 1. Sowing of pathological material taken from sick bees on selective nutrient media for enterobacteria and extraction of pure culture; 2. Microscopy of typical colonies; 3. Determination of bacteria genus; 4. Determination of bacteria motor activity: 5. Urease test; 6. Indole test; 7. Phenyalaalanine test; 8. Study of basic enzymatic properties of bacteria. In a series of laboratory biochemical studies of pure culture microorganisms isolated from mixed culture from diseased bees the isolated bacterial strain belongs to the Family Enterobacteriaceae, Genus Klebsiella, Species Klebsiella pneumoniae. The novelty of the application of the algorithm for determining the species of Klebsiella pneumoniae enterobacteriae allows to diagnose dysbacterioses in winter-spring and summer-autumn times clearly and economically. The isolated Klebsiella pneumoniae bacteria serve as experimental cultures for testing drugs of various directions and actions in laboratory conditions and are kept at the Department of Microbiology, Pharmacology and Epizootology, Faculty of Veterinary Medicine of Polissya National University (formerly Zhytomyr National Agroecological University). Further application of complex diagnostics of enterobacteriosis of bees, including methods of biochemical typification, will allow to expand etiological factors of bee family collapse.