pseudomonas and Klebsiella species. These bacteria are resistant to antibiotic but medicinal plants are most effective against different type of infection. In medicinal plants present the bioactive compounds which showed the antibacterial activity against Klebsiella, Pseudomonas, Staphylococci, Proteus. Multi drug resistant bacteria to one or more therapeutic classes but it showed inhibition zone against some medicinal plants. It detect the
Population surveillance was conducted in the western interior region of British Columbia, Canada (2016 popu- lation 182,422) as previously described . The regional microbiology laboratory at the Royal Inland Hospital in Kamloops identified all residents in the selected area with Klebsiella species BSI. A senior infectious disease consultant (KBL) then performed a case-by-case chart review to abstract clinical information. The Charlson Comorbidity Index (CCI) was used to classify comorbid illnesses . This study was granted a waiver of individ- ual informed consent by the Interior Health Research Ethics Board (file 201,314,052-I).
Background: Resistance to variety of common antimicrobials has made the proliferation of extended spectrum β-lactmase (ESBL) producing strains a serious global health concern that complicated treatment strategies. Antimicrobial resistance to cephalosporin, penicillin and aztreonam are bmediated by Extended-Spectrum Beta-Lactamases (ESBL) via hydrolysis of antibiotics. The most common bacteria associated with ESBL among the Enterobacteriaceae are Klebsiella species . This study was focus on detection of TEM gene associated ESBL Klebsiella species in Khartoum stat.
Klebsiella pneumoniae and Klebsiella oxytoca are the two most frequently encountered Klebsiella species giving rise to infections in humans, but other Klebsiella species can also be found in clinical specimens: Klebsiella ozae- nae, Klebsiella rhinoscleromatis, Klebsiella terrigena, Klebsiella planticola, Klebsiella ornithinolytica, and Enterobac- ter aerogenes (Klebsiella mobilis). However, many of these species are indistinguishable by the conventional meth- ods employed routinely in the clinical microbiological laboratory. Several investigators have suggested various additional tests, but as yet there is no standardized test panel for identifying all Klebsiella species and sub- species. In the present study, performed in three national Klebsiella reference laboratories, we have evaluated a test panel consisting of 18 biochemical tests on 242 strains comprising all Klebsiella species and subspecies. The test panel was designed to identify organisms preliminarily identified as belonging to the genus Klebsiella on the basis of conventional methods or automated identification systems. With the described test panel it is possible to find one or more positive test results differentiating any Klebsiella species, except Klebsiella rhino- scleromatis, from its closest relative.
Amplification of cps genomic region. Among the ORFs identified in the cps region (1), the gnd gene, which codes for 6-phosphogluconate dehydrogenase, was identified downstream of the longest transcriptional unit. We designed two primers flanking the cps cluster, based on (i) the sequence of the cps genomic region of a K2 serotype (1), (ii) the sequence of the K. pneumoniae genome project K52 strain MGH78578 of the Genome Sequencing Center at Washington University Medical School (http://genome.wustl.edu/), and (iii) the sequence diversity of the gnd gene among Klebsiella species (S. Brisse, unpublished data). Primer CPS-1 (5⬘-GCT GGT AGC TGT TAA GCC AGG GGC GGT AGC G) was complementary to the JUMPstart sequence (15) that is located just upstream of ORF3 on the sequence from strain K2 Chedid (1, 33). This JUMPstart sequence was found to be conserved in genome project strain MGH78578. Reverse primer rCPS (5⬘-TAT TCA TCA GAA GCA GCA CGC AGC TGG GAG AAG CC) was complementary to a conserved region of the gnd gene sequence (gene positions 1042 to 1007). A second reverse primer, which was named rCPS2 (5⬘-GCG CTC TGG CTG GTC CAT TTA CCG GTC CCT TTG) and whose sequence was specific for a region located 232 nucleotides upstream of primer rCPS (which was thus 232 bp closer to the forward primer), was designed in order to amplify the cps regions of strains for which the amplification with primer rCPS failed. The molecular sizes of the products amplified from strain MGH78578 are expected to be 18,766 bp in PCRs with primers CPS-1 and rCPS and 18,534 bp in PCRs with primers CPS-1 and rCPS2.
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nal transcribed spacer (ITS) sequence, which is not subject to the same selective pressure as the rRNA genes and conse- quently has a 10-times-greater evolution rate, appears to be able to overcome the apparent limitation of rRNA genes (2, 14). Sequence and length polymorphisms found in the ITS are increasingly being used as tools for bacterial species and sub- species identification (17, 27, 29, 37), typing (25, 37), and evolutionary studies (1, 12, 33). Gu ¨rtler et al. reported that ITS sequence analysis is complementary to the 16S rRNA gene for phylogenetic analysis (13). A PCR method based on ITS sequences has been developed for the detection and iden- tification of K. pneumoniae subsp. pneumoniae (22, 24). In- formation related to ITS regions in Klebsiella is still lacking, with only 1 ITS sequence from K. oxytoca and 21 ITS se-
Antimicrobial resistance (AMR) within a wide range of infectious agents is a public health threat of broad concern to countries (1) . Increasingly, governments around the world are beginning to pay attention to a problem so serious, that it threatens the achievements of modern medicine. AMR is a complex global public health challenge, and no single or simple strategy will suffice to fully contain the emergence and spread of infectious organisms that become resistant to the available antimicrobial drugs. The development of AMR is a natural phenomenon in microorganisms and is accelerated by the selective pressure exerted by use and misuse of antimicrobial agents in humans and animals (2) . The current lack of new antimicrobials on the horizon to replace those that become ineffective brings added urgency to the need to protect the efficacy of existing drugs.Hospitals, and particularly intensive care units, are an important breeding ground for the development and spread of antibiotic resistant bacteria. . An important cause of increasing antibiotic resistance is the selection of resistant bacterial strains by mutation and transfer of mobile resistance genes.Out breaks with a common source of multiple resistant bacteria ,often caused by organisms such as Pseudomonas spp,Klebsiella spp,and Acinetobacter spp are another hazard (2,3) .
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In the present study, the isolated gram negative bacteria from skin wound were E. coli (46.4%), Klebsiella species (10.7%), Proteus species (27.4%), Pseudomonas species (7.1%) and others (8.3%) which correlate with the study done by Haque et al.  in the same hospital. Klebsiella species (88.9%) was the leading ESBL pro- ducers from skin wound followed by Proteus species (78.3%), Enterobacter species (71.4%), E. coli (61.5%) and Pseudomonas species (100.0%). In another study in Bangladesh Haque and Salam21 have reported that ESBL production for Klebsiella species was 57.9% followed by Proteus species (50.0%), E. coli (47.8%) and Pseudomonas species (31.3%) . The high frequency of ESBLs in Klebsiella species is of great concern since infections caused by this bacterium were very common. In addition to that resistance of the organism may be due to the presence of some virulence factor like hyper viscosity, polysaccharide capsule and production of en- dotoxin, carbapenemase, which make it more resistant . Furthermore, they also spread easily with pathogenic and efficient at acquiring and disseminating resistance plasmid  .
strain KPN1705 against At-22 (K. variicola), NJST258_2 (K. pneumoniae), and 700603 (K. quasipneumoniae) reference genomes. The SNP distribution was uniform across the three reference genomes, which made the possibility of KPN1705 being the result of a single large recombination event less likely (data not shown). To further evaluate its phylogeny, we determined that KPN1705 was a single locus variant of the K. pneu- moniae ST1155. The only other ST1155 strain reported in the literature is designated 10982 and is proposed to represent a novel species (22). Strain 10982 was isolated from a perianal swab from an intensive care unit (ICU) patient in Maryland in 2005. Thus, our strain KPN1705 appears most closely related to a novel Klebsiella species that colonizes the gastrointestinal (GI) tract of humans and may be capable of causing human disease. SHV-LEN-OKP core chromosomal beta-lactamases are not restricted by Kleb- siella species. The SHV-LEN-OKP beta-lactamases are core chromosomal genes of Klebsiella that have been suggested to be differentiators of Klebsiella species: K. pneu- moniae (SHV restricted), K. quasipneumoniae (OKP restricted), and K. variicola (LEN FIG 2 Phylogenetic tree of K. quasipneumoniae from humans. Polymorphisms in the K. quasipneumoniae strains were called against the 700603 reference genome (black cross). The clade KpIIA is at the top left, while the KpIIB clade is at the bottom right. ESBL-producing strains sequenced for this study are represented by circles. Non-ESBL-producing strains of both clades identiﬁed in our collection are represented by diamonds. Strains previously sequenced by Holt et al. (8) are represented by squares. Strains associated with human invasive infection sites are indicated in red, and human colonization is indicated in blue.
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Susceptibilities. A total of 2,268 drug-organism combina- tions were compared, with the organism results as shown in Table 4. Many of the errors (two major and eight minor) occurred with Klebsiella species and nitrofurantoin. Discrepan- cies also seemed to be more common with P. aeruginosa iso- lates, although there were no specific antibiotics that showed more problems than others. The other errors were distributed randomly among species and drugs. Among disparities, testing from routine media (reference method) yielded 14 resistant results (25% of all discordant results) that showed intermedi- ate results for the same organism from the CO plate. Most of the minor errors were interpretive differences that involved only one dilution difference in MIC, probably within the ex- perimental error of the systems.
In Uganda, antibiotics are readily available over the counter in community pharmacies , which portends significant rates of resistance among flora circulating within the community. In addition, inappropriate use of antimicrobials by pastoralists for animal diseases and the carriage of drug resistant potentially nosocomial bacteria in the livestock gut has been documented [11, 12]. Cur- rently, most available data focuses on susceptibility of isolates from established infections without much atten- tion on commensal isolates, particularly the Enterobac- teriaceae [11, 12]. Furthermore, drug resistance is also prevalent in hospital settings in Uganda . Moreover, the health care seeking behaviors in the community may vary depending on socioeconomic status of individu- als involved . Yet, information on resistance surveil- lance particularly for isolates from community settings is scarce as surveillance mainly focuses on susceptibility of isolates collected from clinical specimens in hospitals. The aim of this study was to characterize the antimicro- bial susceptibility profiles of E. coli and Klebsiella species isolated as flora from faeces and urine of clients attending outpatient clinics in Uganda, and to investigate the fac- tors associated with carriage of drug resistant isolates.
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(iii) Clone analysis and species identification. To identify the species ampli- fied by PCR and to ensure that the PCR amplicons were indeed bacterial rRNA genes rather than artifacts, a systematic clone analysis was employed. The PCR products were cloned into the pCR8 vector, and at least 20 random transfor- mants were selected for plasmid purification and restriction enzyme digestion. Following electrophoresis, a total of 10 clones with positive 16S rRNA gene inserts were selected for species identification. Plasmids from these 10 clones were extracted by using a WizardPlus SV minipreps DNA purification system (Promega), and their inserts were sequenced (Genomics Core Facility, Case Western Reserve University, Cleveland, OH) by using primers GW1 and GW2 (Table 1), which provided the sequence of the 16S rRNA gene insert from opposite ends. When the DNA sequences obtained with primers GW1 and GW2 were combined, the entire 16S rRNA gene was identified. The sequences were assembled and aligned by using the VectorNTI program (Invitrogen, Carlsbad, CA). The NCBI BLAST nucleotide sequence database (http://www.ncbi.nlm.nih .gov/blast/blast.cgi) was searched for preliminary species identification, regard- less of whether the species was cultivated or uncultivated. Phylogenetic analysis was then performed to identify the most closely related species.
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Our findings highlight the need for improved surveillance, in- fection control, and prevention practices. We argue that CPO control should be a joint collaborative effort by all hospitals and health authorities. It is only through collaboration and coopera- tion that we can advance the common cause of curbing and re- versing the spread of CPOs. In BC, a province-wide CPO surveil- lance program mandated by the Ministry of Health was started in July 2014, with collaboration between the BCCDC PHL and the Provincial Infection Control Network of British Columbia (https: //www.picnet.ca/wp-content/uploads/Surveillance-Protocol-for -CPO-2014-08-11.pdf). This program links laboratory strain characteristics to patient clinical and epidemiological informa- tion, facilitating the implementation of infection control mea- sures. The establishment of effective surveillance programs re- quires knowledge of locally prevalent resistance mechanisms, which would help devise appropriate phenotypic and genotypic protocols (24, 45, 46). NDM was the most prevalent carbapen- emase detected in BC during our study period, followed by OXA- 48-like and VIM carbapenemases (Table 2). Of note, the majority of the OXA-48-like positive K. pneumoniae isolates were isolated in 2012, with their numbers decreasing in 2013 and the first quar- ter of 2014. In contrast to our local epidemiological findings, a recently published paper on the epidemiology of CPOs in Quebec by Lefebvre et al. (47) reported that their landscape is dominated by KPC-producing K. pneumoniae. K. pneumoniae was also the most commonly isolated species in BC in our study, followed by E. cloacae and E. coli (Fig. 2). Surveillance protocols need to be established, bearing in mind the variation in carbapenem MICs in different CPO species (45), and while not every center has the resources necessary for genotypic confirmation of CPOs, those that do would benefit from knowing which carbapen- emases are prevalent in the population and should be targeted, at a minimum.
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Clinical strains. Between 5 and 20 October 2000, 305 single clinical isolates were studied by using the Phoenix system, and findings were compared to the results of the manual methods of ID and AST in use in our laboratories. These 305 strains included 130 Enterobacteriaceae strains, 57 non-lactose-fermenting gram-negative bacillus (NFGNB) strains, 76 Staphylococcus isolates, and 42 Streptococcus and Enterococcus isolates. The following species were included: Escherichia coli (69 isolates), Klebsiella pneumoniae (14 isolates), Klebsiella oxy- toca (6 isolates), Enterobacter cloacae (12 isolates), Enterobacter aerogenes (7 isolates), Citrobacter freundii (6 isolates), Proteus mirabilis (7 isolates), Proteus vulgaris (1 isolate), Proteus penneri (1 isolate), Serratia marcescens (1 isolate), Serratia liquefaciens (1 isolate), Hafnia alvei (1 isolate), Morganella morganii (1 isolate), Providencia stuartii (1 isolate), Salmonella enterica (2 isolates), Pseudo- monas aeruginosa (37 isolates), Pseudomonas fluorescens (2 isolates), Pseudomo- nas putida (1 isolate), Stenotrophomonas maltophilia (7 isolates), Alcaligenes xylosoxidans (2 isolates), Acinetobacter baumanii (5 isolates), Acinetobacter lwoffi (2 isolates), Aeromonas hydrophila (1 isolate), Staphylococcus aureus (42 iso- lates), Staphylococcus epidermidis (17 isolates), Staphylococcus hominis (7 iso- lates), Staphylococcus haemolyticus (7 isolates), Staphylococcus warneri (3 iso- lates), Enterococcus faecalis (23 isolates), Enterococcus faecium (5 isolates), Enterococcus gallinarum (1 isolate), Streptococcus pneumoniae (4 isolates), beta- hemolytic Streptococcus strains A, B, C, and G (7 isolates), and Streptococcus mitis (2 isolates) (the last three species were used only for ID).
Since the 1980s, the incidence of infections due to extended-spectrum β-lactamase-producing Escherichia coli and Klebsiella species (ESBL-EK) has increased. Several studies of clinical outcomes in patients with infections due to ESBL-producing organisms have shown higher mortality and reduced rates of clinical and microbiologic response compared to infections due to non-ESBL-pro- ducing organisms [8-12]. However the impact of ESBL- producing organisms on clinical outcome has not been well described in patients with SBP and advanced liver cir- rhosis. We conducted the current study to evaluate the outcomes of SBP due to ESBL-EK, based on their isolation from ascites, compared with those of SBP due to non- ESBL-EK. We also investigated the impact of ineffective initial antimicrobial therapy on outcome in patients with SBP due to ESBL-EK, and the risk factors for infection by ESBL-producing microorganisms.
The impact of cassava mill effluent on total aerobic bioload of soil samples in Aba was determined between the months of August and October, 2013. The soil samples designated A, B, C, D, E were collected from cassava processing mill effluent dump sites with sterile containers and subjected to standard microbiological analysis. The results revealed a decrease in microbial load of soil as distance approaches the dump sites. The total bacterial and fungal counts of soil samples collected 50m away from the dump sites ranged from 3.0 ×10 6 to 3.4 × 10 6 cfu/g and 4.1 × 10 6 to 4.6 × 10 6 cfu/g respectively, while those at the dump sites ranged from 1.2 × 10 6 to 1.5 × 10 6 cfu/g for total aerobic bacterial count and 2.3 × 10 6 to 2.9 × 10 6 cfu/g for the total aerobic fungal count. The various bacteria isolated from the dump site include Bacillus species, Pseudomonas species, Klebsiella species, Escherichia coli, Micrococcus species and Chromobacterium species while the fungal isolates include Aspergillus species, Penicillum species, Mucor species, Rhizopus species and yeast. The above results indicated an adverse effect of cassava mill effluent on soil microorganisms and calls for regulations on the disposal of the effluent to avoid environmental degradation.
for hydrocarbon remediation is Carbon: Nitrogen: phosphorous equal 100:10:4. The nitrogen, phosphorous and potassium of the collected polluted soil sample ranged from the nitrogen of 73kg/acre, Phosphorous of 8kg/acre, and 104kg/acre. They were also differing in the micronutrient in the soil. The 5 different isolated, identified from the diesel polluted soil; the isolates were Pseudomonas, Bacillus, Citrobacter, Enterobacter and E. coli. Highly degrading potential organism of diesel was identified has Bacillus cereus by 16s rNA sequencing. Bacillus cereus was able to degrade. The seven (Micrococcus, Pseudomonas, Flavobacterium, Serratia, Moraxella, Bacillus and Klibesella) different species (Bacillus species, Klebsiella species, Citrobacter and Pseudomonas species) of potential hydrocarbon degrading organism which utilizes hydrocarbon has a sole carbon source for their growth was identified from hydrocarbon contaminated soil collected in Mexico (Santhini et al., 2009). Some of the researchers have reported that degradation of soil bacteria ranges from 0.13 (Jones et al., 1970) to 50% (Pinholt et al., 1979), and marine bacteria ranges from (0.003% (Hollaway, et al., 1980) to 100% (Mulkins and Phillips 1974). Bacillus Sp was effective hydrocarbon degradation (Amund and Adebiyi, 1991; Atlas, 1992; Nwachuku and Ugoji, 1995; Nwachuku, 2001; Benkacaker and Ekundato, 1997; Diaz et al., 2000). Bacillus Sp identified from hydrocarbon contaminated soils has a potential to degrade benzene, crude, decanol, ethyl-benzene, n- tetradecanol and xylene (Ghazali et al., 2004). The hydrocarbons from the environment has the following bacteria such as Bacillus megaterium, Bacillus cereus, Micrococcus luteus, Staphylococcus aureus, Lactobacillus acidophilus, Neisseria fluorescence and Corynebacterium xerosis were the potent degraders of hydrocarbons (gasoline and diesel) (Jyothi et al., 2012).
Extended-spectrum ␤ -lactamases, plasmid-mediated AmpC ␤ -lactamases (PABLs), and plasmid-mediated metallo- ␤ -lactamases confer resistance to many ␤ -lactams. In Japan, although several reports exist on the prevalence of extended-spectrum ␤ -lactamases and metallo- ␤ -lactamases, the prevalence and characteristics of PABLs remain unknown. To investigate the production of PABLs, a total of 22,869 strains of 4 enterobac- terial species, Escherichia coli, Klebsiella pneumoniae, Klebsiella oxytoca, and Proteus mirabilis, were collected during six 6-month periods from 17 clinical laboratories in the Kinki region of Japan. PABLs were detected in 29 (0.13%) of 22,869 isolates by the 3-dimensional test, PCR analysis, and DNA sequencing analysis. PABL-positive isolates were detected among isolates from 13 laboratories. Seventeen of 13,995 (0.12%) E. coli isolates, 8 of 5,970 (0.13%) K. pneumoniae isolates, 3 of 1,722 (0.17%) K. oxytoca isolates, and 1 of 1,182 (0.08%) P. mirabilis isolates were positive for PABLs. Of these 29 PABL-positive strains, 20 (69.0%), 6 (20.7%), 2 (6.9%), and 1 (3.4%) carried the genes for CMY-2, DHA-1, CMY-8, and MOX-1 PABLs, respectively. Pattern analysis of randomly amplified polymorphic DNA and pulsed-field gel electrophoretic analysis revealed that the prevalence of CMY-2-producing E. coli strains was not due to epidemic strains and that 3 DHA-1-producing K. pneumoniae strains were identical, suggesting their clonal relatedness. In conclusion, the DHA-1 PABLs were predominantly present in K. pneumoniae strains, but CMY-2 PABLs were predominantly present in E. coli strains. The present findings will provide significant information to assist in preventing the emergence and further spread of PABL-producing bacteria.
aeruginosa. Biofilms are microbial communities encased within polysaccharide rich extracellular matrix on surfaces of wounds. They are associated with drastically enhanced resistance against most antimicrobial agents leading to treatment failures. In our study 42 isolates showed biofil m production by Tissue Culture Plate method. Most common organism producing biofilm was Pseudomonas species followed by Staphylococcus aureus; this was in correlation with Seth et al whose study evaluated the effect of clinical strategies against biofilm infected wounds in quantitative in vivo models and showed that Pseudomonal biofilm markedly impairs wound healing. Biofilm producing organisms were associated with therapeutic failure and infection was resolved only on surgical debridement. In TCP method, from the total number of 95 isolates tested for biofilm formation, strong biofilm producers were 36 (37.8 %), 6 (6.3%) were moderate and 53 (55. 7%) isolates were considered as non or weak biofilm producers. In tube method, from the total number of 95 isolates tested for biofilm formation, strong biofilm producers were 30 (31.5 %), 10 (10.5%) were moderate and 60 (63.1%) isolates were considered as non or weak biofilm producers. This was in concordance with the tissue culture plate method of biofilm detection. In congo red agar method, from the total number of 95 isolat es tested for biofilm formation, 20 displayed black colonies but no dry crystalline morphology and 22 displayed black dry crystalline morphology and the rest 53 (55. 7%) isolates were considered as non biofilm producers as they did not displayed black, dry and crystalline colonies.
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The antimicrobial resistance profile of K. pneumoniae, E. coli and Acinetobacter species isolates in our study revealed a generally higher resistance rate than reported in African studies   . The previous studies     show the efficiency of Gentamicin against Gram Negative rods, but our data show a lower effec- tiveness of this antibiotic for treatment of K. pneumoniae and Acinetobacter isolates with their rate of resistance 87.2% and 64.7% respectively. For E. coli, Acinetobacter and K. pneumoniae, we reported high resistance (about 60%) to fluoroquinolones (ciprofloxacin) which limit the available oral treatment.