K1 Types Research Articles

Synergy of bacteriophage depolymerase with host immunity rescues sepsis mice infected with hypervirulent Klebsiella pneumoniae of capsule type K2.

The hypervirulent Klebsiella pneumoniae (hvKp) with K1 and K2 capsular types causes liver abscess, pneumonia, sepsis, and invasive infections with high lethality. The presence of capsular polysaccharide (CPS) resists phagocytic engulfment and contributes to excessive inflammatory responses. Bacteriophage depolymerases can specifically target bacterial CPS, neutralizing its defense. Based on our previous research, we expressed and purified a bacteriophage depolymerase (Dep1979) targeting hvKp with capsule type K2. Interestingly, although Dep1979 lacked direct bactericidal activity in vitro, it exhibited potent antibacterial activity in vivo. Low-dose Dep1979 (0.1 mg/kg) improved the 7-day survival of immunocompetent mice to 100%. Even at 0.01 mg/kg, mice achieved 100% survival at 5 days, although efficacy sharply declined at doses as low as 0.001 mg/kg. Following Dep1979 treatment, reduced expression of inflammatory factors and no apparent tissue damage were observed. However, therapeutic efficacy significantly diminished in immunosuppressed mice. These findings underscore the critical role of Dep1979 in disarming CPS, which synergizes with host immunity to enhance antibacterial activity against hvKp.

Analysis of virulence profiles in clinical isolates of Klebsiella pneumoniae from renal abscesses: clinical significance of hypervirulent isolates.

Klebsiella pneumoniae can cause a wide range of infections. Hypervirulent K. pneumoniae (hvKp), particularly associated with the K1 and K2 capsular types, is an increasingly significant microorganism with the potential to cause invasive infections, including renal abscesses. Despite the rising prevalence of hvKp infections, information on renal abscesses caused by K. pneumoniae is limited, and the clinical significance of hvKp associated with specific virulence genes remains elusive. This study performed at a 1200-bed tertiary hospital sought to identify the clinical and microbiological characteristics of renal abscesses caused by K. pneumoniae, focusing on various virulence genes, including capsular serotypes and multilocus sequence typing (MLST). Over an 8-year period, 64 patients with suspected renal abscesses were reviewed. Ten patients diagnosed with K. pneumoniae-related renal abscesses were ultimately enrolled in the study. Among the isolates from the 10 patients, capsular serotype K2 was predominant (40.0%), followed by K1 (30.0%). The most common sequence type by MLST was 23 (40.0%). In particular, six patients (60.0%) harbored specific genes indicative of hvKp: iucA, peg-344, rmpA, and rmpA2. Our findings highlight the importance of hvKp as a pathogen in renal abscesses. Although the nature of hvKp is relatively unknown, it is widely recognized as a highly virulent pathogen that can infect relatively healthy individuals of various ages and simultaneously cause infections at multiple anatomical sites. Therefore, when treating patients with K. pneumoniae-related renal abscesses, caution is necessary when considering the characteristics of hvKp, such as potential bacteremia, multi-organ abscess formation, and metastatic spread.

Clinical and microbiological characteristics of bloodstream infection caused by Klebsiella pneumoniae harboring rmpA in Japanese adults

We investigated the clinical features of bloodstream infections (BSIs) caused by Klebsiella pneumoniae harboring rmpA and molecular characteristics of the bacteria. We retrospectively investigated adult patients with K. pneumoniae BSI from January 2010 to March 2021 at Nagasaki University Hospital. A matched case–control study in a 1:3 ratio was conducted to clarify the clinical and bacterial characteristics of BSI caused by rmpA-positive K. pneumoniae compared with those caused by rmpA-negative isolates. Antimicrobial susceptibility testing and multilocus sequence typing (MLST) were performed for rmpA-positive isolates. The rmpA was detected in 36 (13.4%) of the 268 isolates. Of these 36 isolates, 31 (86.1%) harbored iucA and 35 (97.2%) each possessed peg-344 and iroB; capsular types were identified as K1 in 9 (25.0%) and K2 in 10 isolates (27.8%). Contrarily, of the 108 rmpA-negative isolates, which were matched for case–control studies, 5 isolates (4.6%) harbored iucA and 1 (0.9%) each possessed peg-344 and iroB; 2 (1.9%) and 3 isolates (2.8%) had K1 and K2 capsular types, respectively. Among the rmpA-positive isolates, ST23/K1 (eight isolates) was the most frequent, followed by ST412/non-K1/K2 (seven isolates), ST86/K2 (five isolates), and ST268/non-K1/K2 (four isolates). In a multivariate analysis using clinical factors, liver abscess positively correlated with rmpA-positive isolates, whereas biliary tract infection and use of anticancer drugs negatively correlated with rmpA-positive isolates in patients with K. pneumoniae BSI. Considering the correlation between rmpA-positive isolates and clinical features, rmpA can be used as a marker for understanding the pathophysiology of K. pneumoniae BSI.

Urban wastewater contributes to the emergence of carbapenem-resistant Klebsiella pneumoniae (CRKP) in an urban receiving river in eastern India.

The present study revealed the emergence of carbapenem-resistant Klebsiella pneumoniae (CRKP) and the associated driving factors in an urban river system surrounding Cuttack city, Odisha. The high contamination factor and contamination degree indicate poor water quality. The CRKP isolates showed 100% resistance against piperacillin, amoxicillin-clavulanic acid, piperacillin-tazobactam, ceftriaxone, ceftazidime, meropenem, and imipenem but less resistance to colistin (12.85%). Among the CRKP isolates, carbapenemase genes blaNDM, blaOXA-48-like, and blaKPC were detected in 94.28%, 35%, and 10% of isolates, respectively. The resistance genes (blaNDM, blaTEM, and blaCTX-M) were found to be significantly correlated with toxic metals (As, Cd, Co, Cu, Fe, Mn, Pb) (P<0.05). Detection of virulence factors (yersiniabactin and aerobactin) and capsular serotypes (K1, K2, and K54 types) explain the pathogenicity of CRKP isolates. Enterobacterial repetitive intergenic consensus-PCR based molecular typing separated the CRKP strains into 13 clusters, of which VI and XI clusters showed similar resistance and virulence determinants, indicating the dissemination of clones from wastewater to the river system. Our results provide first-hand information on assessing risks to public health posed by the CRKP isolates and toxic metals in the Kathajodi River. Molecular surveillance of nearby hospitals for the prevalence of CRKP will help trace their transmission route.

Capsule-Targeting Depolymerases Derived from Acinetobacter baumannii Prophage Regions.

In this study, several different depolymerases encoded in the prophage regions of Acinetobacter baumannii genomes have been bioinformatically predicted and recombinantly produced. The identified depolymerases possessed multi-domain structures and were identical or closely homologous to various proteins encoded in other A. baumannii genomes. This means that prophage-derived depolymerases are widespread, and different bacterial genomes can be the source of proteins with polysaccharide-degrading activities. For two depolymerases, the specificity to capsular polysaccharides (CPSs) of A. baumannii belonging to K1 and K92 capsular types (K types) was determined. The data obtained showed that the prophage-derived depolymerases were glycosidases that cleaved the A. baumannii CPSs by the hydrolytic mechanism to yield monomers and oligomers of the K units. The recombinant proteins with established enzymatic activity significantly reduced the mortality of Galleria mellonella larvae infected with A. baumannii of K1 and K92 capsular types. Therefore, these enzymes can be considered as suitable candidates for the development of new antibacterials against corresponding A. baumannii K types.

Prostatic Abscess Caused by Klebsiella pneumoniae: A 6-Year Single-Center Study

Hypervirulent Klebsiella pneumoniae (hvKp) is an important strain that can cause multiple organ infections. Although hvKp infection cases are increasing, there is limited information on the prostatic abscesses caused by K. pneumoniae. Furthermore, the clinical significance of hvKp associated with K1 or K2 capsular types or virulence genes in prostatic abscesses remains unclear. Therefore, we aimed to elucidate the clinical and microbiological characteristics of prostatic abscesses caused by K. pneumoniae in relation to various virulence genes. A retrospective study was performed at a 1200-bed tertiary hospital between January 2014 and December 2019. Patients diagnosed with prostatic abscesses with K. pneumoniae isolated from blood, urine, pus, or tissue cultures were enrolled in this study. Our results demonstrate that 30.3% (10/33) of the prostatic abscesses were caused by K. pneumoniae. All strains isolated from patients with prostatic abscesses due to K. pneumoniae were the K1 capsular type, and eight patients (80.0%) carried rmpA and iutA genes that identified hvKp. These findings suggest that hvKp is an important pathogen in prostatic abscesses. Therefore, when treating patients with K. pneumoniae prostatic abscesses, attention should be paid to the characteristics of hvKp, such as bacteremia, multiorgan abscess formation, and metastatic spread.

Hypervirulent Klebsiella pneumoniae Strains Modulate Human Dendritic Cell Functions and Affect TH1/TH17 Response.

Hypervirulent Klebsiella pneumoniae (Hv-Kp) strains have emerged as pathogens causing life-threatening, invasive disease even in immunocompetent hosts. Systemic dissemination usually occurs following perturbations of the gut microbiota and is facilitated by Hv-Kp resistance to phagocytosis and complement activity. Hv-Kp are usually associated with K1 or K2 capsular types, produce several iron uptake systems (e.g., aerobactin and salmochelin) and are often but not invariably, capsular material hyper-producers (hypermucoviscous phenotype: HMV). Whether Hv-Kp escape the immune response at mucosal site is unknown. In this work, we studied the effects of Hv-Kp on human dendritic cells (DCs), central players of the IL-23/IL-17 and IL-12/IFN-γ axis at mucosal sites, essential for pathogen clearance. Four Hv-Kp and HMV strains were selected and their activity on DC maturation and cytokine production was compared to that of non-virulent Kp strains with classic or HMV phenotypes. While the maturation process was equally induced by all Kp strains, significant differences between virulent and non-virulent strains were found in the expression of genes for cytokines involved in T-cell activation and differentiation. The non-virulent KP04C62 and the classic Kp, KPC157 induced high expression of TH1 (IL-12p70 and TNFα) and TH17 cytokines (IL-23, IL-1β and IL-6), while Hv-Kp poorly activated these cytokine genes. Moreover, conditioned media from DCs cultured with non-virulent Kp, either classical or hypercapsulated, induced the activation of IL-17 and IFN-γ genes in preactivated CD4+-cells suggesting their TH17/TH1 differentiation. Conditioned media from Hv-Kp poorly activated IL-17 and IFN-γ genes. In summary, our data indicate that Hv-Kp interfere with DC functions and T-cell differentiation and suggest that the escape from the IL-23/IL-17 and IL-12/IFN-γ axes may contribute to pathogen dissemination in immunocompetent hosts.

Hypervirulent Klebsiella pneumoniae - clinical and molecular perspectives.

Hypervirulent Klebsiella pneumoniae (hvKp) has emerged as a concerning global pathogen. hvKp is more virulent than classical K.pneumoniae (cKp) and capable of causing community-acquired infections, often in healthy individuals. hvKp is carried in the gastrointestinal tract, which contributes to its spread in the community and healthcare settings. First recognized in Asia, hvKp arose as a leading cause of pyogenic liver abscesses. In the decades since, hvKp has spread globally and causes a variety of infections. In addition to liver abscesses, hvKp is distinct from cKp in its ability to metastasize to distant sites, including most commonly the eye, lung and central nervous system (CNS). hvKp has also been implicated in primary extrahepatic infections including bacteremia, pneumonia and soft tissue infections. The genetic determinants of hypervirulence are often found on large virulence plasmids as well as chromosomal mobile genetic elements which can be used as biomarkers to distinguish hvKp from cKp clinical isolates. These distinct virulence determinants of hvKp include up to four siderophore systems for iron acquisition, increased capsule production, K1 and K2 capsule types, and the colibactin toxin. Additionally, hvKp strains demonstrate hypermucoviscosity, a phenotypic description of hvKp in laboratory conditions that has become a distinguishing feature of many hypervirulent isolates. Alarmingly, multidrug-resistant hypervirulent strains have emerged, creating a new challenge in combating this already dangerous pathogen.

Prevalence of pks gene cluster and characteristics of Klebsiella pneumoniae-induced bloodstream infections.

BackgroundThe emerging pks‐positive (pks+) strains have aroused great public concern recently. Colibactin, encoded by pks gene cluster, has been reported to be involved in DNA damage and increased virulence. Little is known about its prevalence among Klebsiella pneumoniae‐induced bloodstream infections (BSIs). Therefore, the aim of this study was to investigate the prevalence of pks gene cluster, and molecular and clinical characteristics of K pneumoniae‐induced BSIs.MethodsA total of 190 non‐duplicate K pneumoniae bloodstream isolates were collected at a university hospital in China from March 2016 to March 2018. Molecular characteristics including capsular types, virulence, and pks genes were detected by polymerase chain reaction (PCR). Clinical characteristics and antimicrobial susceptibility were also investigated.ResultsOverall, 21.6% (41/190) of K pneumoniae bloodstream isolates were hypervirulent K pneumoniae(hvKP). The prevalence of pks gene cluster was 26.8% (51/190). The positive rates of K1, K57, and genes associated with hypervirulence, that is, rmpA, wcaG, mrkD, allS, ybtS, kfu,and iucA, were significantly higher in the pks+isolates than the pks‐negative (pks −) isolates (P < 0.05), while the pks+ isolates were significantly less resistant to 11 antimicrobial agents than the pks − isolates. Multivariate analysis showed diabetes mellitus, and K1 and K20 capsular types as independent risk factors for pks + K pneumoniaebloodstream infections.ConclusionsThe pks + K pneumoniae was prevalent in individuals with bloodstream infections in mainland China. The high rates of hypervirulent determinants among pks + K pneumoniaerevealed the potential pathogenicity of this emerging gene cluster. Diabetes mellitus, and K1 and K20 capsular types were identified as independent risk factors associated with pks+ K pneumoniaebloodstream infections. This study highlights the significance of clinical awareness and epidemic surveillance of pks+strains.

Baseline Parasite Profile in Developing Irradiation Malaria Vaccine in Indonesia: Molecular Analysis of Papua Samples

Malaria remains a major public health problem in Indonesia, therefore the development of its vaccine that can be created with radiation is urgently needed. Aim of this research was to determine the genotype of blood specimens obtained from 4-22 years old malaria outpatients in Jayapura General Hospital, Papua. The information obtained will be a great value in additional knowledge for vaccine development of gamma ray irradiated parasites. Microscopic examination on thin blood smear was done, followed by Polymerase Chain Reaction (PCR) diagnosis to further confirm the parasite using 18S rRNA gene and mitochondrial primers on deoxyribonucleic acid extracted using chelex-100 ion exchanger and the presence of each mutation in the genes was examined using restriction fragment length polymorphism method. Our result showed that 9 samples infected with Plasmodium falciparum and 1 with P. vivax. Beside that 9 samples carried mutations in pfcrt and 22% in pfmdr1. Mutations involving N1042D at pfmdr1 gene was also detected in 3 samples (33%). Mutations in dhfr gene at codon S108N were detected in all 9 samples. Mutations in dhps gene at codon K540E were also detected in 3 (33%) samples. Analysis of the parasite genotype indicated that infection occurred with multiple parasite strains. The majority of the samples carried the 3D7 type of MSP2 gene while others carried the FC27 type. The MSP1 genes observed carried K1, MAD20 and RO33 types but some other types could not be determined.

The genetic polymorphism of merozoite surface protein-1 in Plasmodium falciparum isolates from Aceh province, Indonesia

An estimated of 3.3 million Indonesian population were infected with malaria. However, extensive genetic polymorphism of the field isolates msp-1 of P. falciparum represents a major obstacle for the development of malaria treatment. The aim of this study was to investigate the genetic diversity of msp-1 genotype in field isolates of P. falciparum collected in Aceh Province. A total of 90 patients with malaria (+) were selected from eleven district hospitals in Aceh from 2013-2015. Data were collected by anamnesis, complete physical examination and laboratory tests for msp-1. All protocols to diagnose malaria followed the WHO 2010 guideline. All samples were stored in Eijkman Biology Molecular Institute, Jakarta. Among 90 samples, 57.7% were male, and 42.3% were female with the most cases found between 21-30 years old. From the allele typing analysis of P. falciparum from Aceh; K1, MAD20, and RO33 allele types were identified. MAD20 type was the highest allele found in this study (57.9%). It was found in single and mixed infection. A moderate level of the mixed allele was also observed.

Comparative genome analysis of novel Podoviruses lytic for hypermucoviscous Klebsiella pneumoniae of K1, K2, and K57 capsular types

Hypermucoviscous (HV) strains of capsular types K1, K2 and K57 are the most virulent representatives of the Klebsiella pneumoniae species. Eight novel bacteriophages lytic for HV K. pneumoniae were isolated and characterized. Three bacteriophages, KpV41, KpV475, and KpV71 were found to have a lytic activity against mainly K. pneumoniae of capsular type K1. Two phages, KpV74, and KpV763 were lytic for K2 capsular type K. pneumoniae, and the phage KpV767 was specific to K57-type K. pneumoniae only. Two more phages, KpV766, and KpV48 had no capsular specificity. The phage genomes consist of a linear double-stranded DNA of 40,395–44,623bp including direct terminal repeats of 180–246 bp. The G + C contents are 52.3–54.2 % that is slightly lower than that of genomes of K. pneumoniae strains being used for phage propagation. According to the genome structures, sequence similarity and phylogenetic data, the phages are classified within the genus Kp32virus and Kp34virus of subfamily Autographivirinae, family Podoviridae. In the phage genomes, genes encoding proteins with putative motifs of polysaccharide depolymerase were identified. Depolymerase genes of phages KpV71 and KpV74 lytic for hypermucoviscous K. pneumoniae of K1 and K2 capsular type, respectively, were cloned and expressed in Escherichia coli, and the recombinant gene products were purified. The specificity and polysaccharide-degrading activity of the recombinant depolymerases were demonstrated.

Epidemiological typing of multidrug-resistant Klebsiella pneumoniae, which causes paediatric ventilator-associated pneumonia in Egypt.

Multidrug-resistant Klebsiella pneumoniae is a common nosocomial pathogen that plays an important role in ventilator-associated pneumonia (VAP). This study aimed to define the clonal relatedness of K. pneumoniae strains isolated from paediatric VAP in addition to those isolated from environmental samples. This study included 19 clinical and 4 environmental K. pneumoniae isolates recovered from the paediatric intensive care unit (PICU) in Assiut University Children's Hospital. The K. pneumoniae isolates were confirmed by biotyping using API strips and subjected to antimicrobial susceptibility testing. The genes coding K1 and K2 capsular types were detected by PCR. The clonal relationships between the K. pneumoniae isolates were determined by pulsed-field gel electrophoresis (PFGE). Ten resistotypes were detected among all the K. pneumoniae isolates, while PFGE identified seventeen K. pneumoniae pulsotypes. Similar PFGE patterns were found between environmental and clinical isolates and between isolates recovered from different patients, suggesting the circulation of K. pneumoniae pathogens in the PICU and the role of the environment in the spread of infection. No correlation was found between the resistotypes and pulsotypes of the K. pneumoniae isolates. PFGE showed higher discriminatory power for the typing of nosocomial K. pneumoniae [Simpson's diversity index (DI)=0.96] than resistotyping (DI=0.72). As far as we know, this is the first report of the isolation of the same multidrug-resistant (MDR) K. pneumoniae pulsotype from patients and environmental samples in the same hospital ward in Egypt. This study provides a step on the way to understanding the genotyping and epidemiology of MDR K. pneumoniae for enhanced prevention of bacterial transmission.

Prevalence and characteristics of pks genotoxin gene cluster-positive clinical Klebsiella pneumoniae isolates in Taiwan

The pks gene cluster encodes enzymes responsible for the synthesis of colibactin, a genotoxin that has been shown to induce DNA damage and contribute to increased virulence. The present study investigated the prevalence of pks in clinical K. pneumoniae isolates from a national surveillance program in Taiwan, and identified microbiological and molecular factors associated with pks-carriage. The pks gene cluster was detected in 67 (16.7%) of 400 isolates from various specimen types. Multivariate analysis revealed that isolates of K1, K2, K20, and K62 capsular types (p < 0.001), and those more susceptible to antimicrobial agents (p = 0.001) were independent factors strongly associated with pks-carriage. Phylogenetic studies on the sequence type (ST) and pulsed-field gel electrophoresis patterns indicated that the pks-positive isolates belong to a clonal group of ST23 in K1, a locally expanding ST65 clone in K2, a ST268-related K20 group, and a highly clonal ST36:K62 group. Carriage of rmpA, iutC, and ybtA, the genes associated with hypervirulence, was significantly higher in the pks-positive isolates than the pks-negative isolates (95.5% vs. 13.2%, p < 0.001). Further studies to determine the presence of hypervirulent pks-bearing bacterial populations in the flora of community residents and their association with different disease entities may be warranted.

Genetic Diversity of Plasmodium falciparum Populations in Malaria Declining Areas of Sabah, East Malaysia.

Malaysia has a national goal to eliminate malaria by 2020. Understanding the genetic diversity of malaria parasites in residual transmission foci can provide invaluable information which may inform the intervention strategies used to reach elimination targets. This study was conducted to determine the genetic diversity level of P. falciparum isolates in malaria residual foci areas of Sabah. Malaria active case detection was conducted in Kalabakan and Kota Marudu. All individuals in the study sites were screened for malaria infection by rapid diagnostic test. Blood from P. falciparum-infected individuals were collected on filter paper prior to DNA extraction. Genotyping was performed using merozoite surface protein-1 (MSP-1), merozoite surface protein-2 (MSP-2), glutamate rich protein (GLURP) and 10 neutral microsatellite loci markers. The size of alleles, multiplicity of infection (MOI), mean number of alleles (Na), expected heterozygosity (He), linkage disequilibrium (LD) and genetic differentiation (FST) were determined. In Kalabakan, the MSP-1 and MSP-2 alleles were predominantly K1 and FC27 family types, respectively. The GLURP genotype VI (751–800 bp) was predominant. The MOI for MSP-1 and MSP-2 were 1.65 and 1.20, respectively. The Na per microsatellite locus was 1.70. The He values for MSP-1, MSP-2, GLURP and neutral microsatellites were 0.17, 0.37, 0.70 and 0.33, respectively. In Kota Marudu, the MSP-1 and MSP-2 alleles were predominantly MAD20 and 3D7 family types, respectively. The GLURP genotype IV (651–700 bp) was predominant. The MOI for both MSP-1 and MSP-2 was 1.05. The Na per microsatellite locus was 3.60. The He values for MSP-1, MSP-2, GLURP and neutral microsatellites were 0.24, 0.25, 0.69 and 0.30, respectively. A significant LD was observed in Kalabakan (0.495, p<0.01) and Kota Marudu P. falciparum populations (0.601, p<0.01). High genetic differentiation between Kalabakan and Kota Marudu P. falciparum populations was observed (FST = 0.532). The genetic data from the present study highlighted the limited diversity and contrasting genetic pattern of P. falciparum populations in the malaria declining areas of Sabah.

Allelic Forms of Merozoite Surface Protein-3 in Plasmodium falciparum Isolates From Southeast of Iran.

Background:Genetic diversity has provided Plasmodium falciparum with the potential capacity of avoiding the immune response, and possibly supported the natural selection of drug or vaccine-resistant parasites. Merozoite surface protein-3 (MSP-3) has been used to develop vaccines and investigate the genetic diversity regarding P. falciparum malaria in Iran.Objectives:The main goal of this study was to analyze the polymorphic antigen MSP-3 genes across southeast of Iran among four different districts, to identify the differences in the allele frequency and genetic diversity.Materials and Methods:Nested polymerase chain reaction amplification was used to determine polymorphisms of N-terminal region of the MSP-3 gene. A total of 85 microscopically positive P. falciparum infected individuals from southeast of Iran were included in this study.Results:Of the 85 confirmed P. falciparum samples obtained from four different districts, 72 were successfully scored for MSP-3.The MSP-3 allele classes (K1 and 3D7 types) showed comparable prevalence in all districts. Overall frequencies of K1 and 3D7 allele classes were 94.5 % for both.Conclusions:Since no study has yet looked at the extent of P. falciparum MSP-3 in this geographic region, these data can be helpful to support development of a vaccine based on MSP-3 against malaria. There should be a comparative analysis in different seasonal peaks to indicate the allelic polymorphism of MSP-3 over a period.

Genetic Diversity of Variable Region Block 2 in the Merozoite Surface Protein-1 (MSP1) in Plasmodium falciparum Field Isolates from South-East of Iran

Merozoite surface protein 1 of (PfMSP-1) is a leading malaria vaccine candidate. However, extensive genetic diversity of this gene in field isolates of represents a major obstacle for the development of an effective vaccine against malaria. The present study was aimed at analysing genetic polymorphisms of K1, MAD20 and RO33 allelic types of MSP-1 block 2 among isolates from Sistan and Baluchestan, Iran. In this study a total of 94 infected persons from Sistan and Baluchistan Province of Iran, were included. Blood samples were collected from March 2011 to September 2012. Block 2 of the MSP-1 gene was genotyped by allele-specific nested polymerase chain reaction (PCR) after DNA extraction. Eighty-nine (94.7%) of the 94 samples were successfully amplified; 7 distinct MSP-1 genotypes were identified by size differences on agarose gels. MAD20 was the predominant MSP-1 allelic family identified in 46.1% (41/89) of the samples while RO33 family had the least frequency (7.9%). A total of 9/89 (10.1%) samples exhibited multiple infections with two alleles at PfMSP-1. The present study shows that the level of genetic diversity is relatively low in south-east of Iran and most of infections are composed of one clone, which is consistent with an area of low malaria transmission. These data are useful for malaria prevention and control in Iran.

Identification of Klebsiella pneumoniae K1 and K2 Capsular Types by PCR and Quellung Test

Background:: Klebseilla pneumoniae causes urinary tract infections, nosocomial pneumonia and intra-abdominal infections. Capsular antigens are considered to be the ultimate virulence determinants. Among 77 capsular serotypes of K. pneumoniae, serotypes K1 and K2 are the most virulent ones in humans. Objectives:: We designed a PCR method for detection of capsular serotypes K1 and K2 of K. pneumoniae using genes cps cluster wzc and orf10 which are required for biosynthesis of capsular polysaccharides in K1 and K2 types, respectively. Materials and Methods:: We collected 89 K. pneumoniae clinical isolates from the patients of Labafinejad Hospital located in Tehran. Clinical isolates were mostly chosen from urine. We used PCR technique to detect isolates possessing K1 and K2 capsular polysaccharides based on the wzc and orf10 genes which are required for biosynthesis of capsular polysaccharide in K1 and K2 types, respectively. The results of PCR were compared with those obtained by capsular serotyping using Quellung test. Results:: Of 89 isolates of K. pneumoniae tested by both techniques, 10 (11.2%) belonged to K1 and 13 (14.6%) belonged to K2 serotypes, respectively. Conclusions:: We found that these serotypes are probably important in clinical specimens. PCR was a simple and inexpensive tool for identification of K. pneumoniae clinical isolates of K1 and K2 geno-serotypes.

Genetic diversity of Plasmodium falciparum field isolates in central Sudan inferred by PCR genotyping of Merozoite surface protein 1 and 2

Background:Characterization of Plasmodium falciparum diversity is commonly achieved by amplification of the polymorphic regions of the merozoite surface proteins 1 (MSP1) and 2 (MSP2) genes.Aims:The present study aimed to determine the allelic variants distribution of MSP1 and MSP2 and multiplicity of infection in P. falciparum field isolates from Kosti, central Sudan, an area characterized by seasonal malaria transmission.Materials and Methods:Total 121 samples (N = 121) were collected during a cross-sectional survey between March and April 2003. DNA was extracted and MSP1 and MSP2 polymorphic loci were genotyped.Results:The total number of alleles identified in MSP1 block 2 was 11, while 16 alleles were observed in MSP2 block 3. In MSP1, RO33 was found to be the predominant allelic type, carried alone or in combination with MAD20 and K1 types, whereas FC27 family was the most prevalent in MSP2. Sixty two percent of isolates had multiple genotypes and the overall mean multiplicity of infection was 1.93 (CI 95% 1.66-2.20). Age correlated with parasite density (P = 0.017). In addition, a positive correlation was observed between parasite densities and the number of alleles (P = 0.022).Conclusion:Genetic diversity in P. falciparum field isolates in central Sudan was high and consisted of multiple clones.

Genetic analysis of the merozoite surface protein-1 block 2 allelic types in Plasmodium falciparum clinical isolates from Lao PDR

BackgroundMSP-1 is one of the potential malarial vaccine candidate antigens. However, extensive genetic polymorphism of this antigen in the field isolates of Plasmodium falciparum represents a major hindrance for the development of an effective vaccine. Therefore, this study aimed to establish the prevalence and genetic polymorphisms of K1, MAD20 and RO33 allelic types of msp-1 block 2 among P. falciparum clinical isolates from Lao PDR.MethodsPlasmodium falciparum isolates were collected from 230 P. falciparum-infected blood samples from three regions of Lao PDR. K1, MAD20 and RO33 were detected by nested PCR; SSCP was used for polymorphism screening. The nested PCR products of each K1, MAD20 and RO33 allelic types that had different banding patterns by SSCP, were sequenced.ResultsThe overall prevalence of K1, MAD20 and RO33 allelic types in P. falciparum isolates from Lao PDR were 66.95%, 46.52% and 31.30%, respectively, of samples under study. Single infections with K1, MAD20 and RO33 allelic types were 27.83%, 11.74% and 5.22%, respectively; the remainders were multiple clonal infections. Neither parasite density nor age was related to MOI. Sequence analysis revealed that there were 11 different types of K1, eight different types of MAD20, and 7 different types of RO33. Most of them were regional specific, except type 1 of each allelic type was common found in 3 regions under study.ConclusionsGenetic polymorphism with diverse allele types was identified in msp-1 block 2 among P. falciparum clinical isolates in Lao PDR. A rather high level of multiple clonal infections was also observed but the multiplicity of infection was rather low as not exceed 2.0. This basic data are useful for treatment and malaria control program in Lao PDR.