Klebsiella Type Research Articles

Genetic analysis of capsular polysaccharide synthesis gene clusters in 79 capsular types of Klebsiella spp.

A total of 79 capsular types have been reported in Klebsiella spp., whereas capsular polysaccharide synthesis (cps) regions were available in only 22 types. Due to the limitations of serotyping, complete repertoire of cps will be helpful for capsular genotyping. We therefore resolved the rest 57 cps and conducted comparative analysis. Clustering results of 1,515 predicted proteins from cps loci categorized proteins which share similarity into homology groups (HGs) revealing that 77 Wzy polymerases were classified into 56 HGs, which indicate the high specificity of wzy between different types. Accordingly, wzy-based capsular genotyping could differentiate capsule types except for those lacking wzy (K29 and K50), those sharing identical wzy (K22 vs. K37); and should be carefully applied in those exhibited high similarity (K12 vs. K41, K2 vs. K13, K74 vs. K80, K79 vs. KN1 and K30 vs. K69). Comparison of CPS structures in several capsular types that shared similarity in their gene contents implies possible functions of glycosyltransferases. Therefore, our results provide complete set of cps in various types of Klebsiella spp., which enable the understandings of relationship between genes and CPS structures and are useful for identification of documented or new capsular types.

Antibiogram Typing and Biochemical Characterization of Klebsiella pneumoniae after Biofield Treatment

Antibiogram Typing and Biochemical Characterization of Klebsiella pneumoniae after Biofield Treatment

Antibiogram Typing and Biochemical Characterization of Klebsiella pneumoniae after Biofield Treatment

Klebsiella pneumoniae (K. pneumoniae) is a common nosocomial pathogen causing respiratory tract (pneumoniae) and blood stream infections. Multidrug-resistant (MDR) isolates of K. pneumoniae infections are difficult to treat in patients in health care settings. Aim of the present study was to determine the impact of Mr. Trivedi’s biofield treatment on four MDR clinical lab isolates (LS) of K. pneumoniae (LS 2, LS 6, LS 7, and LS 14). Samples were divided into two groups i.e. control and biofield treated. Control and treated groups were analyzed for antimicrobial susceptibility pattern, minimum inhibitory concentration (MIC), biochemical study and biotype number using MicroScan Walk-Away® system. The analysis was done on day 10 after biofield treatment as compared with control group. Antimicrobial sensitivity assay showed that there was 46.42% alteration in sensitivity of tested antimicrobials in treated group of MDR K. pneumonia isolates. MIC results showed an alteration in 30% of tested antimicrobials out of thirty after biofield treatment in clinical isolates of K. pneumoniae. An increase in antimicrobial sensitivity and decrease in MIC value was reported (in LS 6) in case of piperacillin/tazobactam and piperacillin. Biochemical study showed a 15.15% change in biochemical reactions as compared to control. A significant change in biotype numbers were reported in all four clinical isolates of MDR K. pneumoniae after biofield treatment as compared to control group. On the basis of changed biotype number after biofield treatment, new organism was identified as Enterobacter aerogenes in LS 2 and LS 14. These results suggest that biofield treatment has a significant effect on altering the antimicrobial sensitivity, MIC values, biochemical reactions and biotype number of multidrug-resistant isolates of K. pneumoniae.

Capsular types of Klebsiella pneumoniae revisited by wzc sequencing.

Capsule is an important virulence factor in bacteria. A total of 78 capsular types have been identified in Klebsiella pneumoniae. However, there are limitations in current typing methods. We report here the development of a new genotyping method based on amplification of the variable regions of the wzc gene. Fragments corresponding to the variable region of wzc were amplified and sequenced from 76 documented capsular types of reference or clinical strains. The remaining two capsular types (reference strains K15 and K50) lacked amplifiable wzc genes and were proven to be acapsular. Strains with the same capsular type exhibited ≧94% DNA sequence identity across the variable region (CD1-VR2-CD2) of wzc. Strains with distinct K types exhibited <80% DNA sequence identity across this region, with the exception of three pairs of strains: K22/K37, K9/K45, and K52/K79. Strains K22 and K37 shared identical capsular polysaccharide synthesis (cps) genes except for one gene with a difference at a single base which resulted in frameshift mutation. The wzc sequences of K9 and K45 exhibited high DNA sequence similarity but possessed different genes in their cps clusters. K52 and K79 exhibited 89% wzc DNA sequence identity but were readily distinguished from each other at the DNA level; in contrast, strains with the same capsular type as K52 exhibited 100% wzc sequence identity. A total of 29 strains from patients with bacteremia were typed by the wzc system. wzc DNA sequences confirmed the documented capsular type for twenty-eight of these clinical isolates; the remaining strain likely represents a new capsular type. Thus, the wzc genotyping system is a simple and useful method for capsular typing of K. pneumoniae.

Wzi Gene Sequencing, a Rapid Method for Determination of Capsular Type for Klebsiella Strains

Pathogens of the genus Klebsiella have been classified into distinct capsular (K) types for nearly a century. K typing of Klebsiella species still has important applications in epidemiology and clinical microbiology, but the serological method has strong practical limitations. Our objective was to evaluate the sequencing of wzi, a gene conserved in all capsular types of Klebsiella pneumoniae that codes for an outer membrane protein involved in capsule attachment to the cell surface, as a simple and rapid method for the prediction of K type. The sequencing of a 447-nucleotide region of wzi distinguished the K-type reference strains with only nine exceptions. A reference wzi sequence database was created by the inclusion of multiple strains representing K types associated with high virulence and multidrug resistance. A collection of 119 prospective clinical isolates of K. pneumoniae were then analyzed in parallel by wzi sequencing and classical K typing. Whereas K typing achieved typeability for 81% and discrimination for 94.4% of the isolates, these figures were 98.1% and 98.3%, respectively, for wzi sequencing. The prediction of K type once the wzi allele was known was 94%. wzi sequencing is a rapid and simple method for the determination of the K types of most K. pneumoniae clinical isolates.

Capsular serotypes and multilocus sequence types of bacteremic Klebsiella pneumoniae isolates associated with different types of infections

We investigated the epidemiology of different serotypes of Klebsiella pneumoniae isolates causing bacteremic liver abscess (LA) using multilocus sequence typing (MLST). MLST and molecular typing were performed for 41 K1 (19 LA), 37 K2 (5 LA), and 33 non-K1/K2 (6 LA) isolates that were derived from a previous one-year K. pneumoniae bacteremia cohort. Capsular serotypes and rmpA of these isolates were determined by polymerase chain reaction (PCR) methods. Among the 41 K1 isolates, 39 were ST23 and the remaining two isolates were ST23 single-locus variant. There were 11 STs among K2 isolates. ST65 was the most common (n = 10), followed by ST86, ST373, and ST375. Only ST65 (n = 3), ST373 (n = 1), and ST375 (n = 1) caused LA, and ST65 was a three-locus variant of ST23. For non-K1/K2 isolates, the ST types varied widely. ST218 (K57) was the most common type (n = 6, 18 %), and it was a single-locus variant of ST23 and caused two cases of LA. The existences of rmpA among serotypes varied (100 % for K1, 89 % for K2, and 55 % for non-K1/K2). For isolates causing LA, all of them were positive for rmpA. For non-K1/K2 isolates causing infections other than LA, the positivity of rmpA ranged from 0 % (biliary tree infection) to 67 % (pneumonia). In this one-year cohort, all K1 isolates were ST23 or its single-locus variants, but the composition of ST types among K2 isolates was quite variable. ST23 and its one- (ST1005 and ST218) and three-locus (ST65) variants comprised 80 % of isolates causing LA.

Susceptibility of gram-negative aerobic bacilli from intra-abdominal pathogens to antimicrobial agents collected in the United States during 2011

During 2011, a total of 1442 gram-negative pathogens from intra-abdominal infections were collected as part of the Study for Monitoring Antimicrobial Resistance Trends (SMART) from 19 hospital sites within the United States. Susceptibility to ertapenem and comparators and molecular analysis of ertapenem resistant isolates was performed. Extended-spectrum beta-lactamase ESBL (ESBL) isolates were determined using the Clinical and Laboratory Standards Institute's recommended phenotypic test. Isolates were identified to the species level, and tested for antimicrobial susceptibility using custom MicroScan dehydrated broth microdilution panels ESBLs and carbapenemases were characterized using the Check-Points microarray. Strain typing of Klebsiella pneumoniae was performed by rep-PCR on the DiversiLab System. The majority of isolates were Escherichia coli (36%), K. pneumoniae (18.6%), Pseudomonas aeruginosa (12.1%) and Enterobacter cloacae (8.4%). Incidence of ESBL-positive isolates was 12.7%, 9.7%, 3.6% and 3.1% for K. pneumoniae, E. coli, Proteus mirabilis and Klebsiella oxytoca, respectively. Against the majority of isolates and species tested, the most active antibiotics were amikacin, ertapenem, and imipenem, with the carbapenems being the most active invitro, including against ESBL-positive isolates of E. coli. All other antibiotics exhibited diminished activity. Against K. pneumoniae, the carbapenems were notably less active against ESBL-positive isolates though their activity against this sub-population was still the highest of all antibiotics tested; however, 41.1% (14 of 34) of the phenotypically ESBL-positive K. pneumoniae co-produced a carbapenemase (KPC2 or KPC3), and >90% of the isolates producing only an ESBL remained susceptible to ertapenem. Further monitoring of susceptibility of intra-abdominal isolates is warranted due to limited therapeutic options available to physicians.

Surface modification of PLGA nanofibrous biocomposites using flavonoids for biomedical applications

Rutin-containing poly(L-lactide-co-glycolide) (PLGA) nanofibrous biocomposites were prepared by electrospinning technique and antibacterial properties of the biocomposites were examined. Two types of bacterial strains, Staphylococcus aureus and Klebsiella pneumoniae were used in this test. The results showed that the rutin-containing PLGA nanofibrous biocomposites exhibited significant antibacterial effects against both the bacteria. Human nasopharyngeal epidermoid carcinoma (KB) epithelial cells were used to evaluate cytotoxicity. It was found that there was no cytotoxicity for the rutin-containing PLGA nanofibrous biocomposites.

Phenotypic and Molecular Characteristics of Carbapenem-Non-Susceptible Enterobacteriaceae from a Teaching Hospital in Wenzhou, Southern China

Carbapenem resistance in Enterobacteriaceae is increasing and has become a matter of great concern. The aim of this study was to characterize carbapenem-non-susceptible Enterobacteriaceae from a teaching hospital. A total of 49 carbapenem-non-susceptible Enterobacteriaceae clinical isolates recovered in 2007-2010 from the First Affiliated Hospital of Wenzhou Medical College were analyzed by antimicrobial susceptibility testing. The carbapenemase phenotype, outer membrane protein profiles, and clonal relatedness were investigated using the modified Hodge test, sodium dodecyl sulfate-polyacrylamide gel electrophoresis, and pulsed-field gel electrophoresis (PFGE). Multilocus sequence typing (MLST) of Klebsiella pneumoniae was also performed. β-Lactamase genes were examined by PCR and sequencing, and the transferability of carbapenemase genes was determined by a conjugation experiment. The rates of imipenem, meropenem, and ertapenem resistance were 59.2%, 40.8%, and 96.0%, respectively. Thirty isolates exhibited carbapenemase activity, and 32 isolates carried carbapenemase genes. Furthermore, 10 and 9 clinical isolates posessed AmpC β-lactamase and extended-spectrum β-lactamase (ESBL) genes, respectively. Eight of 32 carbapenemase-producing isolates were proved to be carried by conjugative plasmids, and there was porin loss in 34.7% (17/49) of the isolates. PFGE analysis demonstrated that 9 KPC-2-producing Serratia marcescens belonged to a clonal strain, suggesting the clonal dissemination of these KPC-2-bearing isolates among different wards. The MLST of K. pneumoniae revealed that two KPC-2 producers were ST11. This study suggests that KPC-2-type carbapenemase is the main contributor to carbapenems resistance in carbapenemase-producing Enterobacteriaceae, and that ESBL, AmpC β-lactamase overproduction, and porin loss contribute to the resistance level among these isolates; in carbapenemase-non-producing Enterobacteriaceae, ESBL, AmpC enzyme, and porin loss contribute to the carbapenems resistance of Enterobacteriaceae, especially the ertapenem resistance of Enterobacter cloacae.

ISOLATION OF KLEBSIELLA SPP. FROM SOME MEATS

Meats often contains micro-organism which may cause disease, Including Klebsiella spp. Which cause wide range of disease states, notably pneumonia, urinary tract infections (UTIs), and bacteremia. The aim of this study was collected 100 samples of meats (Calf, sheep, chicken) from the meats shops present in the different area in Damascus and countryside in a sterile container, for isolated Klebsiella and growing it on Selective Media to identify and characterize its morphology, biochemical and Molecular Characteristics by using Polymerase chain reaction PCR. Colonies of the microorganisms were counted on difference media then isolated from a selective media. The colonies of Klebsiella isolated are circular, dome-shaped. 3-4 mm diameter with mucoid aspect, stickiness and surrounded by capsule. Our results showed that 46% of examined specimen contained Klebsiella spp.The results showed that it can investigate of Klebsiella in meats samples by biochemical reactions and polymerase chain reaction. In next study we will be produced primers To differentiate between two types of Klebsiella (K. pneumoniae and K. oxytoca).

In Vitro Assessment of Antibacterial Activity and Cytocompatibility of Quercetin‐Containing PLGA Nanofibrous Scaffolds for Tissue Engineering

Flavonoids, such as quercetin, have been reported to exhibit a wide range of biological activities related to their antioxidant capacity. The aim of this study was to investigate the protective effect of quercetin on cell adhesion, and the viability and proliferation of KB epithelial cells. Quercetin‐ (1, 5 wt%)‐containing poly (l‐lactide‐co‐glycolide) (PLGA) nanofibrous scaffolds (PLGA/Q 1, PLGA/Q 5) were prepared by electrospinning technique and their antibacterial properties were examined. Two types of bacteria strains, Staphylococcus aureus (SA) and Klebsiella pneumoniae (KP), were used to evaluate the antibacterial properties of the scaffolds. The results showed that the quercetin‐containing PLGA nanofibrous scaffolds exhibited significant antibacterial effects against the two bacterial strains. KB epithelial cells were also used to evaluate the cytocompatibility of the scaffolds. From the results, it was found that the PLGA nanofibrous scaffolds with 1 wt% of quercetin had good cell compatibility. It is considered that the PLGA nanofibrous scaffolds with 1 wt% quercetin have potential to be used in tissue engineering.

Molecular Typing and Virulence Analysis of Serotype K1 Klebsiella pneumoniae Strains Isolated from Liver Abscess Patients and Stool Samples from Noninfectious Subjects in Hong Kong, Singapore, and Taiwan

Serotype K1 Klebsiella pneumoniae with multilocus sequence type 23 (ST23) has been strongly associated with liver abscess in Taiwan. Few data regarding the strain types and virulence of this serotype from other Asian countries are available. Serotype K1 K. pneumoniae strains isolated from liver abscess and stool samples from subjects hospitalized in Hong Kong, Singapore, and Taiwan hospitals were examined. Forty-seven serotype K1 isolates were identified: 26 from liver abscess samples and 21 from stool samples. MLST revealed 7 sequence types: 85.1% (40 of 47 isolates) belonged to ST23, 1 isolate belonged to ST163 (a single-locus variant of ST23), and 2 isolates were ST249 (a 3-locus variant of ST23). New STs, namely, ST367, ST425, and ST426, were allocated to 3 of 4 isolates from stool samples. The virulence of these strains was determined by neutrophil phagocytosis and mouse infection models. Except for two ST23 isolates, all Klebsiella pneumoniae isolates were resistant to phagocytosis. Resistance to serum killing varied in isolates of ST23, while all non-ST23 strains were susceptible to serum killing except one with ST249 from a liver abscess. All hypervirulent isolates with a 50% lethal dose of <10(2) CFU were from ST23, were resistant to phagocytosis and serum killing, and also carried both virulence-associated genes, rmpA and aerobactin. Multilocus sequence typing genotype 23 was the most prevalent sequence type among serotype K1 K. pneumoniae isolates from both liver abscess and stool samples in the Asia Pacific region. Serotype K1 K. pneumoniae isolates with capsule expression leading to phagocytic resistance and with the aerobactin gene were associated with hypervirulence.

Molecular epidemiological analysis of KPC-2 and IMP-4 carbapenemase possessing Klebsiella pneumonia isolated from children

Objective To investigate the distribution of acquired carbapenemases in carbapenemresistant strains of Klebsiella pneumoniae, and explore its role in epidemiology of nosocomial infection. Methods From November 2008 to March 2009, twenty clinical isolates of carbapenem-resistant Klebsiella pneumoniae were collected from children hospitalized in Wuhan children's hospital. MICs of antibiotics were tested by DNA of Klebsiella pneumoniae. Modified Hodge test was used to screen strains producing carbapenemases,combined imipenem(IPM)-EDTA , meropenem(MEM)- EDTA and ceftazidime(CAZ) - EDTA double-disk synergy test (DDST) were used to detect metallo-β-lactamase-producing. PCR amplification of the carbapenemase and integrase genes, and sequencing were performed. Plasmid conjugation transfer experiments and Southern hybridization were applied to study the mode of drug resistance transmission. Results Four types of Klebsiella pneumoniae were detected by PFGE, type A consisted of 5 strains, including 3 strains of type Al and 2 strains of type A2), type B (2 strains), type C (12strains) and type D (1 strain). Type A and C were the main drug resistant clones. Eight strains of Klebsiella pneumoniae carried both KPC-2 and IMP-4 genes, 10 strains carried IMP-4 gene, 2 strains carried KPC-2 gene. None of NDM-1 ,GIM, SPM, SIM, OXA-23, and VIM carbapenemase genes was detected in 20 isolates. All of 20 isolates carried lntl which were found to be located on bacterial chromosome by Southern blot. Conclusions KPC-2and IMP-4 genes are the major carbapenemase genes in Klebsiella pneumoniae isolated in Wuhan.Transmission of drug resistance is mainly through vertical transmission of type C resistant clone and horizontal transmission of Intl on bacteria chromosome. Key words: Klebsiella pneumoniae; Carbapenems; Beta-Lactamases; Integrons

Comparison of (GTG) 5-oligonucleotide and ribosomal intergenic transcribed spacer (ITS)-PCR for molecular typing of Klebsiella isolates

Molecular typing of Klebsiella species has become important for monitoring dissemination of β-lactamase-producers in hospital environments. The present study was designed to evaluate poly-trinucleotide (GTG) 5- and rDNA intergenic transcribed spacer (ITS)-PCR fingerprint analysis for typing of Klebsiella pneumoniae and Klebsiella oxytoca isolates. Multiple displacement amplified DNA derived from 19 K. pneumoniae (some with an ESBL-phenotype), 35 K. oxytoca isolates, five K. pneumoniae, two K. oxytoca, three Raoultella, and one Enterobacter aerogenes type and reference strains underwent (GTG) 5 and ITS-PCR analysis. Dendrograms were constructed using cosine coefficient and the Neighbour joining method. (GTG) 5 and ITS-PCR analysis revealed that K. pneumoniae and K. oxytoca isolates, reference and type strains formed distinct cluster groups, and tentative subclusters could be established. We conclude that (GTG) 5 and ITS-PCR analysis combined with automated capillary electrophoresis provides promising tools for molecular typing of Klebsiella isolates.

Nosocomial blood stream infection in intensive care units at Assiut University Hospitals (Upper Egypt) with special reference to extended spectrum β-lactamase producing organisms

AimThis study investigated the nosocomial blood stream infection (BSI) in the adult ICUs in Assiut university hospitals to evaluate the rate of infection in different ICUs, causative microorganisms, antimicrobial resistance, outcome of infection, risk factors, prevalence of extended spectrum B-lactamase producing organisms and molecular typing of Klebsiella pneumoniae strains to highlight the role of environment as a potential source of nosocomial BSI.MethodsThis study was conducted over a period of 12 months from January 2006 to December 2006. All Patients admitted to the different adult ICUs were monitored daily by attending physicians for subsequent development of nosocomial BSI. Blood cultures were collected from suspected patients to detect the causative organisms. After antimicrobial susceptibility testing, detection of ESBLs was conducted among gram negative isolates. Klebsiella pneumoniae isolates were tested by PCR to determine the most common group of B-lactamase genes responsible for resistance. Klebsiella pneumoniae isolates from infected patients and those isolated from the environment were typed by RAPD technique to investigate the role of environment in transmission of infection.ResultsThe study included 2095 patients who were admitted to different ICUs at Assiut University Hospitals from January 2006 to December 2006. Blood samples were collected from infected patients for blood cultures. The colonies were identified and antibiotic sensitivities were performed. This study showed that the rate of nosocomial BSI was 75 per 1000 ICU admissions with the highest percentages in Trauma ICU (17%). Out of 159 patients with primary bloodstream infection, 61 patients died representing a crude mortality rate of 38%. Analysis of the organisms causing BSI showed that Gram positive organisms were reported in 69.1% (n = 121); MRSA was the most prevalent (18.9%), followed by methicillin resistant coagulase negative Staphylococci (16%). Gram negative bacilli were reported in 29.1% (n = 51). In this case, Klebsiella pneumoniae was the most common (10.3%) followed E coli (8.6%). Candida spp. was reported only in (1.7%) of isolates. Antibiotics sensitivities of Gram positive organisms showed that these organisms were mostly sensitive to vancomycin (90.1%), while Gram negative organisms were mostly sensitive to imipenem (90.2%). In this study we tested Gram negative isolates for the production of the ESBL enzyme and concluded that 64.7% (33/51) of patients' isolates and 20/135 (14.8%) environmental isolates were confirmed to be ESBL producers. The type of β-lactamase gene was determined by polymerase chain reaction which showed that SHV was the main type. Molecular typing was done for 18 Klebsiella pneumoniae strains that caused nosocomial BSI and for the 36 Klebsiella pneumoniae strains which were isolated from the environmental samples by the RAPD method. The two environmental strains were identical, with one isolated from a patient, which confirms the serious role of the hospital environment in the spread of infections.ConclusionNosocomial BSI represents a current problem in Assiut University Hospitals, Egypt. Problems associated with BSI include infection with multidrug resistant pathogens (especially ESBLs) which are difficult to treat and are associated with increased mortality. Of all available anti-microbial agents, carbapenems are the most active and reliable treatment options for infections caused by ESBL isolates. However, overuse of carbapenems may lead to resistance of other Gram-negative organisms.

Molecular typing of Klebsiella pneumoniae isolates from public hospitals in Recife, Brazil

Thirty nosocomial isolates of Klebsiella pneumoniae, collected from three public hospitals in Recife, Brazil, between 1999 and 2000, were analysed in order to determine their epidemiological relatedness and genetic characteristics. The isolates were characterised by biotyping, antibiotyping, protein analysis, plasmid profile and random amplified polymorphic DNA (RAPD). The majority of the clinical isolates were resistant to multiple antibiotics, in particular to beta-lactams, and 30% were found to be ESBLs producers. RAPD proved to be the most effective technique in discriminating unrelated K. pneumoniae isolates. It was confirmed by the highly genetic similarity found among related isolates from an hospital outbreak. We conclude that K. pneumoniae infections in Recife has been caused by a variety of bacterial genotypes. This is the first report that revealed the subtypes of K. pneumoniae in Brazil by plasmid analysis and RAPD.

NOSOCOMIAL KLEBSIELLA INFECTION IN NEONATES IN A TERTIARY CARE HOSPITAL: PROTEIN PROFILE BY SDS-PAGE AND KLEBOCIN TYPING AS EPIDEMIOLOGICAL MARKERS

Purpose: To find out the prevalence of Klebsiella in hospital acquired neonatal infections in a tertiary care set up and to evaluate the role of klebocin typing and protein profile by SDS-PAGE in epidemiological typing of the isolates. Methods: Hospital born neonates transferred to the neonatal unit after birth and available in the unit 48 hours later comprised the study group. Two hundred and three neonates were found eligible for inclusion in the study. Repeated blood cultures, other relevant clinical specimens and environmental samples were collected and identified according to the standard techniques. Isolated clinical and environmental Klebsiella pneumoniae strains were subjected to klebocin typing and protein profiling by SDS-PAGE at regular intervals. Results: Multi drug resistant K. pneumoniae were the commonest organism isolated in 30 neonates leading to the incidence of Klebsiella nosocomial infection to be 14.7%. Klebocin typing of the K. pneumoniae isolates showed four patterns with type 312 being the commonest (43.4%). Whole cell protein analysis by SDS-PAGE of K. pneumoniae isolates revealed four types of banding pattern. Analysis of the typing method showed that the typeability and reproducibility of klebocin was 83.3% and 73.3% respectively whereas typeability and reproducibility of SDS-PAGE was 100%. Conclusions: Based on the present study it is concluded that SDS-PAGE typing method is better than klebocin typing in neonatal nosocomial infection. It is also suggested that protein profile by SDS-PAGE may be used as a tool for epidemiological typing of Klebsiella pneumoniae isolates in laboratories where genomic based molecular typing technique is not available.

Capsule impedes adhesion to and invasion of epithelial cells by Klebsiella pneumoniae.

The adhesion of K21a, K26, K36, and K50 capsulated Klebsiella strains to ileocecal (HCT-8) and bladder (T24) epithelial cell lines was significantly lower than that of their corresponding spontaneous noncapsulated variants K21a/3, K26/1, K36/3, and K50/3, respectively. Internalization of the bacteria by both epithelial cell lines was also significantly reduced. Similarly, a capsule-switched derivative, K2(K36), that exhibited a morphologically larger K36 capsule and formed more capsular material invaded the ileocecal epithelial cell line poorly compared to the corresponding K2 parent strain. None of the capsulated strains exhibited significant mannose-sensitive type 1 fimbriae, whereas two of the noncapsulated variants K21a/3 and K50/3 exhibited potent mannose-sensitive hemagglutinating activity. Although hemagglutinating activity that could be attributed to mannose-resistant Klebsiella type 3 fimbriae was weak in all strains, in several cases the encapsulated parent strains exhibited lower titers than their corresponding noncapsulated variants. Although the level of adhesion to the ileocecal cells is not different from adhesion to bladder cells, bacterial internalization by bladder cells was significantly lower than internalization by ileocecal cells, suggesting that bladder cells lack components required for the internalization of Klebsiella.

Molecular typing of multiresistant Klebsiella pneumoniae isolated from children from northern Jordan.

Twenty-nine clinical isolates of community acquired Klebsiella pneumoniae obtained from 17 children with malnutrition were characterized by antibiotic susceptibility, plasmid analysis, and random amplified polymorphic DNA (RAPD) techniques. Disc diffusion methodology was used to test the susceptibility of the isolates to 13 antibiotics: amoxycillin, cephapirin, ceftazidime, cefoxitin, cefotaxime, aztreonam, gentamicin, ciprofloxacin, chloramphenicol, erythromycin, nalidixic acid, trimethoprim and amoxycillin-clavulanic acid. All the isolates showed multiresistance patterns (15 patterns) ranging from resistance to two antibiotics to resistance to 10 antibiotics. All isolates were resistant to amoxycillin and erythromycin. Ten K. pneumoniae isolates producing extended-spectrum beta-lactamases (ESBLs) as evidenced by the double-disc diffusion synergy test were isolated sporadically from six patients. Six of these 10 isolates were hyperproducers of ESBL, which resulted in increased resistance to the beta-lactam-beta-lactamase inhibitor combination amoxycillin-clavulanic acid. Plasmid analysis showed plasmid ranging in size from 48 kilobases (kb) to 1.4 kb. All the 29 isolates shared the same plasmid 26 kb. There was a consistent relationship between antibiotype and plasmid profiles for each pair of isolates obtained from five individual patients. RAPD analysis using a single (10-mer) primer demonstrated that the isolates that have the same antibiotype and the same plasmid profile had different RAPD fingerprint patterns. These results demonstrate that the RAPD technique is better than antibiotype characterization and a plasmid analysis profile for typing K. pneumoniae as well as for revealing strain differences.

RAPD typing of Klebsiella pneumoniae, Klebsiella oxytoca, Serratia marcescens and Pseudomonas aeruginosa isolates using standardized reagents.

OBJECTIVE: To perform quality assessment of standardized random amplified polymorphic DNA (RAPD) analysis for epidemiologic typing of Klebsiella pneumoniae, K. oxytoca, Serratia marcescens and Pseudomonas aeruginosa. METHODS: Thirty K. pneumoniae, 15 K. oxytoca, 30 S. marcescens and 33 P. aeruginosa epidemiologically unrelated isolates and four collections of clinically related isolates of each species were included in the study. RAPD analysis was performed using Ready-To-Go RAPD Analysis beads with primer ERIC-1R and Ready-To-Go primer 2 for K. pneumoniae and K. oxytoca, primer set ERIC-2/1026 and Ready-To-Go primer 2 for S. marcescens, and primers D-10514 and D-14306 for P. aeruginosa. RESULTS: All epidemiologically unrelated K. pneumoniae and K. oxytoca isolates were distinguished. Twenty-nine types were distinguished among the 30 unrelated S. marcescens isolates and 32 types among the 33 unrelated P. aeruginosa isolates. Indistinguishable banding patterns were obtained in repeated analyses of two isolates and from 11 serial subcultures of three isolates of each species included in the study. The RAPD data from the clinically related isolates correlated with the epidemiologic origin of the isolates. CONCLUSIONS: The use of Ready-To-Go RAPD Analysis beads resulted in reproducible and stable banding patterns with a high discriminatory capacity, and the RAPD typing results corresponded with the epidemiologic origin of the isolates.