Causative Strain Research Articles

Clinical detection and characterization of bacterial pathogens in the genomics era.

The availability of genome sequences obtained using next-generation sequencing (NGS) has revolutionized the field of infectious diseases. Indeed, more than 38,000 bacterial and 5,000 viral genomes have been sequenced to date, including representatives of all significant human pathogens. These tremendous amounts of data have not only enabled advances in fundamental biology, helping to understand the pathogenesis of microorganisms and their genomic evolution, but have also had implications for clinical microbiology. Here, we first review the current achievements of genomics in the development of improved diagnostic tools, including those that are now available in the clinic, such as the design of PCR assays for the detection of microbial pathogens, virulence factors or antibiotic-resistance determinants, or the design of optimized culture media for ‘unculturable’ pathogens. We then review the applications of genomics to the investigation of outbreaks, either through the design of genotyping assays or the direct sequencing of the causative strains. Finally, we discuss how genomics might change clinical microbiology in the future.Electronic supplementary materialThe online version of this article (doi:10.1186/s13073-014-0114-2) contains supplementary material, which is available to authorized users.

Getting personal perspectives on individualized treatment duration in multidrug-resistant and extensively drug-resistant tuberculosis.

Tuberculosis (TB) differs from most other bacterial infectious diseases by a very long duration of combination antibiotic therapy required to achieve relapse-free cure. Although the standard recommended "short-course" treatment length for TB is 6 months, the World Health Organization recommends a duration of 20 months for the treatment of patients with multidrug-resistant and extensively drug-resistant TB (M/XDR-TB). Apart from the long duration of anti-TB therapy, treatment of M/XDR-TB is very expensive and often associated with adverse drug events. The optimal duration for treatment of TB likely differs between individuals and depends on a variety of variables, such as the extent of the disease, the immune status of the host, and the virulence and the drug resistance of the causative strain of Mycobacterium tuberculosis. Some patients with M/XDR-TB may have to be treated with currently available antituberculosis drug regimens for more than 20 months, whereas much shorter treatment durations may be possible to achieve cure for the majority of patients with M/XDR-TB. Personalization of the duration of treatment for TB, especially for patients with M/XDR-TB, would be highly desired. Until recently there has been little interest in the identification of biosignatures that could eventually lead to individual recommendations for the duration of anti-TB therapy. This pulmonary perspective reviews the knowledge on clinical and radiological scores, host- and pathogen disease-related profiles, molecules, and signatures that are currently explored as biomarkers to personalize the duration of therapy in TB.

Genomic characterization of a large outbreak of Legionella pneumophila serogroup 1 strains in Quebec City, 2012.

During the summer of 2012, a major Legionella pneumophila serogroup 1 outbreak occurred in Quebec City, Canada, which caused 182 declared cases of Legionnaire's disease and included 13 fatalities. Legionella pneumophila serogroup 1 isolates from 23 patients as well as from 32 cooling towers located in the vicinity of the outbreak were recovered for analysis. In addition, 6 isolates from the 1996 Quebec City outbreak and 4 isolates from patients unrelated to both outbreaks were added to allow comparison. We characterized the isolates using pulsed-field gel electrophoresis, sequence-based typing, and whole genome sequencing. The comparison of patients-isolated strains to cooling tower isolates allowed the identification of the tower that was the source of the outbreak. Legionella pneumophila strain Quebec 2012 was identified as a ST-62 by sequence-based typing methodology. Two new Legionellaceae plasmids were found only in the epidemic strain. The LVH type IV secretion system was found in the 2012 outbreak isolates but not in the ones from the 1996 outbreak and only in half of the contemporary human isolates. The epidemic strains replicated more efficiently and were more cytotoxic to human macrophages than the environmental strains tested. At least four Icm/Dot effectors in the epidemic strains were absent in the environmental strains suggesting that some effectors could impact the intracellular replication in human macrophages. Sequence-based typing and pulsed-field gel electrophoresis combined with whole genome sequencing allowed the identification and the analysis of the causative strain including its likely environmental source.

Development of tailored real-time RT-PCR assays for the detection and differentiation of serotype O, A and Asia-1 foot-and-mouth disease virus lineages circulating in the Middle East

Rapid and accurate diagnosis is essential for effective control of foot-and-mouth disease (FMD). In countries where FMD is endemic, identification of the serotypes of the causative virus strains is important for vaccine selection and tracing the source of outbreaks. In this study, real-time reverse transcription polymerase chain reaction (rRT-PCR) assays using primer/probe sets designed from the VP1 coding region of the virus genomes were developed for the specific detection of serotype O, A and Asia-1 FMD viruses (FMDVs) circulating in the Middle East. These assays were evaluated using representative field samples of serotype O strains belonging exclusively to the PanAsia-2 lineage, serotype A strains of the Iran-05 lineage and serotype Asia-1 viruses from three relevant sub-groups. When RNA extracted from archival and contemporary field strains was tested using one- or two-step rRT-PCR assays, all three primer/probe sets detected the RNA from homotypic viruses and no cross-reactivity was observed with heterotypic viruses. Similar results were obtained using both single- and multiplex assay formats. Using plasmid standards, the minimum detection level of these tests was found to be lower than two copies. The results illustrate the potential of tailored rRT-PCR tools for the detection and categorization of viruses circulating in the Middle East belonging to distinct subgroups of serotypes O, A and Asia-1. These assays can also overcome the problem of serotyping samples which are found positive by the generic rRT-PCR diagnostic assays but negative by virus isolation and antigen-detection ELISA which would otherwise have to be serotyped by nucleotide sequencing. A similar approach could be used to develop serotyping assays for FMDV strains circulating in other regions of the world.

Correlation Analysis of EV71 Detection and Case Severity in Hand, Foot, and Mouth Disease in the Hunan Province of China

An increase in the incidence of hand, foot and mouth disease (HFMD) cases has been observed in the Hunan province of mainland China since 2009 with a particularly higher level of severe cases in 2010–2012. Intestinal viruses of the picornaviridae family are responsible for the human syndrome associated with HFMD with enterovirus 71 (EV71) and Coxsackievirus A16 (Cox A16) being the most common causative strains. HFMD cases associated with EV71 are generally more severe with an increased association of morbidity and mortality. In this study, the etiology surveillance data of HFMD cases in Hunan province from March 2010 to October 2012 were analyzed to determine if there is a statistically relevant linear correlation exists between the detection rate of EV71 in mild cases and the proportion of severe cases among all HFMD patients. As the cases progressed from mild to severe to fatal, the likelihood of EV71 detection increased (25.78%, 52.20% and 84.18%, respectively). For all cases in the timeframe evaluated in this study, the presence of virus was detected in 63.21% of cases; among cases showing positivity for virus, EV71 infection accounted for 50.14%. These results provide evidence to support the observed higher morbidity and mortality associated with this outbreak and emphasizes the importance of early detection in order to implement necessary prevention measures to mitigate disease progression.

First report of an Acanthamoeba genotype T13 isolate as etiological agent of a keratitis in humans.

Several strains of free-living amoebae (FLA) belonging to the genus Acanthamoeba are able to cause a painful sight-threatening disease of the cornea designated as Acanthamoeba keratitis (AK). In this case report, a 22-year-old woman, wearer of soft contact lenses, was treated after the initial examination, and follow-up laboratory results led to the diagnosis of Acanthamoeba keratitis. The patient recovered under the targeted therapy, demonstrating that the acanthamoebae were the etiological agents of the keratitis in this case. The acanthamoebae belonged morphologically to group II. Genotyping of the causative Acanthamoeba strain based on sequences of the PCR amplimer ASA.S1 amplified from 18S ribosomal DNA by using the genus-specific primers JDP1 and JDP2 followed. The phylogenetic comparison of ASA.S1 confirmed that the isolated Acanthamoeba strain is closely related to genotype T13 supported by pairwise sequence identities of 97.1-98.0% and bootstrap support of 980 replicates with reference sequences of genotype T13. These results regarding the Acanthamoeba keratitis-causing isolate KaBo expands the number of known pathogenic genotypes to 12. To our knowledge, this is the first report of a T13 Acanthamoeba genotype being associated with keratitis in humans.

Mass mortality of Eurasian Tree Sparrows (Passer montanus) from Salmonella Typhimurium dt40 in Japan, winter 2008-09.

An outbreak of salmonellosis in wild passerines caused mass mortality of Eurasian Tree Sparrows (Passer montanus) in Hokkaido, Japan, 2005-06; however, the etiology was poorly understood. In winter 2008-09, sparrow mortality again occurred in Hokkaido, and 202 deaths in 100 incidents at 94 sites were reported. We conducted a comprehensive investigation to evaluate the cause and impact on sparrow populations. We collected 26 carcasses at 13 sites, including a zoological park. In addition, Salmonella screening of zoo animals was conducted as a biosecurity measure. Salmonella Typhimurium was isolated from multiple organs in all examined sparrows; they were diagnosed with septicemic salmonellosis. Eleven sites (85%) were related to wild bird feeding and six of eight sparrow fecal samples, including from the zoo, were S. Typhimurium-positive. No infection was detected in zoo animals. Isolates belonged to three phage types: DT40 (88%), DT110 (8%), and DT120 (4%). Pulsed-field gel electrophoresis patterns were the same in all isolates, regardless of phage type. Biochemical characteristics and antibiotic-resistance profiles of DT40 were similar in all isolates, indicating a single origin. The mortality was likely associated with that in 2005-06 because the isolates had the same profiles. Tissue levels of sodium, calcium, and magnesium (the main components of chemical deicer suspected to be the major cause of poisoning deaths in 2005-06 mortality) were not higher in the affected sparrows. We conclude that an emerging epidemic infection with S. Typhimurium DT40 related to bird feeding was the cause of sparrow mortality in 2008-09 and suggest that this causative strain is host-adapted to sparrows in Japan. The mortality might have had some impact on the local population, but its influence was limited.

Hospital Clostridium difficile Cases Nearly Doubled During 2000s

DENVER – The incidence of Clostridium difficile infections in United States hospitals nearly doubled between 2001 and 2010, with little evidence of recent decline, an analysis of national data showed. “These data underscore the importance of directing resources to the prevention of [C. difficile infection, CDI], as well as developing public policy for reducing the incidence of these infections in U.S. hospitals,” Kelly R. Daniels, PharmD, said in an interview prior to the annual Interscience Conference on Antimicrobial Agents and Chemotherapy, where the study was presented. “Judicious use of antibiotics is essential to reducing these infections, as antibiotics are the main risk factor for the development of CDI,” she noted. Dr. Daniels, a graduate student in the translational science PhD program at the University of Texas, Austin, and her associates retrospectively reviewed U.S. National Hospital Discharge Surveys from 2001 to 2010. They included patients aged 18 years and older who were discharged from the hospital with an ICD-9-CM diagnosis code for CDI (008.45) and used data weights to determine national estimates. They presented incidence rates as CDI cases per 1,000 hospitalizations, and they used multivariable logistic and linear regression models to compare mortality and hospital length of stay between primary and secondary CDI. Over the 10-year study period, 2.2 million patients were discharged from the hospital with diagnoses of CDI. Their median age was 75 years, most (86%) were white, and more than half (59%) were female. One-third of cases (33%) were primary CDI, while the remainder were secondary CDI. The three most common concomitant infectious diagnoses were urinary tract infection (21%), pneumonia (14%), and sepsis/septicemia (12%). Dr. Daniels reported that the incidence of CDI increased from 4.5 cases/1,000 hospitalizations in 2001 to 8.2 cases/1,000 hospitalizations in 2010. Similar trends were observed in patients with primary and secondary CDI. The overall mortality rate was 7.1% for the study period and was significantly higher among patients with secondary CDI, compared with those who had primary CDI (8.8% vs. 3.3%, respectively; relative risk, 1.8). The median hospital length of stay was 8 days and was significantly higher among patients with secondary CDI, compared with those who had primary CDI (9 days vs. 5 days, respectively; RR, 13.3). “We found that in-hospital -mortality among patients with primary CDI is decreasing, while mortality among patients with secondary CDI is increasing,” Dr. Daniels said. “This trend is different from prior studies, which demonstrated increases in CDI-related mortality from the 1990s to the early 2000s.” The decline in in-hospital -mortality among patients with primary CDI might reflect improvements in care in recent years, she explained. In contrast, the increase in mortality among those with secondary CDI may be caused by changes in the frequency or severity of other comorbid illnesses. However, “this was not specifically examined as part of our study,” Dr. Daniels added. She acknowledged certain limitations of the analysis, including the fact that it relied on administrative codes to identify cases of CDI. “Although CDI codes have high accuracy for identifying CDI, they cannot be considered equivalent to medical chart reviews,” Dr. Daniels explained. Also, “the use of administrative codes precludes our ability to confirm the CDI diagnosis using laboratory methods or to identify the causative strain of C. difficile.” In addition, the National Hospital Discharge Surveys don't include long-term care hospitals or federal hospitals. “Therefore, our estimates may not be generalizable to those settings and may underestimate the true burden of CDI in the United States,” Dr. Daniels explained. No outside funding was obtained for the conduct of this study. Dr. Daniels disclosed that she is supported by the National Institutes of Health National Center for Advancing Translational Sciences Loan Repayment Program.

Molecular detection and isolation of West Nile virus from a human case in northern Greece, 2013.

In order to laboratory confirm the first suspected West Nile fever case in 2013 in northern Greece, a combination of serological molecular and culture methods were applied. It was shown that the causative West Nile virus strain belonged to lineage 2, and possessed the amino acid substitution H249P in the NS3 protein, as in previous years. The significance of this specific strain in Europe remains to be elucidated.

Influenza vaccine strains: Licensing perspectives

Influenza vaccine strains: Licensing perspectives

Genotype diversity of Mycobacterium isolates from children in Jimma, Ethiopia.

BackgroundPaediatric tuberculosis (TB) is poorly addressed in Ethiopia and information about its magnitude and the genotype distribution of the causative Mycobacterium tuberculosis strains responsible for its spread are scanty.MethodsGastric lavage or sputum samples were collected from consecutively enrolled TB suspect children visiting Jimma University Hospital in 2011 and cultured on Middlebrook 7H11 and Löwenstein-Jensen media. Acid fast bacterial (AFB) isolates were subjected to molecular typing targeting regions of difference (RDs), 16S rDNA gene and the direct repeat (DR) region using multiplex polymerase chain reaction (mPCR), gene sequencing and spoligotyping, respectively. Molecular drug susceptibility testing of M. tuberculosis isolates was performed by Genotype®MTBDRplus line probe assay (LPA) (Hain Life Sciences, Germany).ResultsGastric lavage (n = 43) or sputum (n = 58) samples were collected from 101 children and 31.7% (32/101) of the samples were positive for AFB by microscopy, culture and/or PCR. Out of 25 AFB isolates, 60% (15/25) were identified as M. tuberculosis by PCR, and 40% isolates (10/25) were confirmed to be non-tuberculous mycobacteria (NTM) by genus typing and 16S rDNA gene sequencing. Lineage classification assigned the M. tuberculosis strains into Euro-American (EUA, 66.7%; 10/15), East-African-Indian (EAI; 2/15), East-Asian (EA; 1/15) and Indio-Oceanic (IO; 1/15) lineages. Seven M. tuberculosis strains were new to the SpolDB4 database. All of the M. tuberculosis isolates were susceptible to isoniazid (INH) and rifampicin (RIF), except for one strain (of spoligotype SIT-149 or T3_ETH family) which had a mutation at the inhA locus which often confers resistance to INH (low level) and ethionamide.ConclusionsAnalysis of the genetic population structure of paediatric M. tuberculosis strains suggested similarity with that of adults, indicating an on-going and active transmission of M. tuberculosis from adults to children in Ethiopia. There were no multidrug-resistant TB (MDR-TB) strains among the isolates.

Meningococcemia due to the 2000 Hajj-Associated Outbreak Strain (Serogroup W-135 ST-11) with Immunoreactive Complications

We present the first reported case of systemic infection with Neisseria meningitidis serogroup W-135 sequence type (ST)-11 in Japan. A 44-year-old woman presented with high fever, sore throat, and fatigue and was diagnosed with N. meningitidis bacteremia. The causative strain was identified as serogroup W-135 ST-11 by polymerase chain reaction and multilocus sequence typing. Approximately 1 month after treatment, she developed high fever, dyspnea, chest pain, and shoulder pain due to pericarditis, polyarthritis, and tenosynovitis, which are all relatively common immunoreactive complications of W-135 ST-11 meningococcal infections. This causative strain was the same as that responsible for an outbreak of meningitis among Hajj pilgrims in 2000. The strain is now found worldwide because it can attain a high carriage rate and has a long duration of carriage. We suspect that our patient's infection was acquired from an imported chronic carrier.

Sacroiliac joint pain as an important element of psoriatic arthritis diagnosis

Psoriatic arthritis (PsA) is a chronic inflammatory disease characterized by the coexistence of arthritis with psoriasis of the skin and nails. The sacroiliac joints were observed in 34-78% of patients with psoriatic arthritis. Due to such a high prevalence of SIJ dysfunction, understanding pathophysiology of pain and the associated pain pattern becomes a very important aspect of PsA diagnosis. As far as the etiology of SI joint dysfunction is concerned, it has not been disambiguated yet. Among the main causative factors, injuries and strains of the structures surrounding the joint are noted. Joint pathology usually manifests itself by pain occurring within the area of the joint. The causes of pain may be divided into two categories: intra-articular and extra-articular. Pain caused by the SI joint may be nociceptive or neural in nature, whereas the pain pattern characteristic of the joint correlates with its innervation and is consistent with S2 dorsal rami.

Special Issue on Infectious Disease Control of Natural Disasters

Large natural disasters such as earthquakes, tsunamis and typhoons often produce many refugees, forcing them to live inconvenient and unsanitary lives in temporary places of refuge. Even if they can remain in their homes, hygienic conditions may be worsened by interrupted electricity, water, fuel and other lifelines. Winter disasters bring more concerns, such as influenza and cold weather, while those in summer raise problems of diarrhea-related disease. Two of Japan’s largest recent earthquakes, the 1995 Great Hanshin-Awaji Earthquake and the 2011 Great East Japan Earthquake, did not cause large infectious disease outbreaks thanks to proper countermeasures. Even so, such outbreaks frequently occur worldwide. One recent example is the 2010 cholera outbreak following an earthquake in Haiti. In an added complication, it is thought that the outbreak was from a foreign source – an Asian epidemic strain of cholera. Many refugees have resulted from ethnic and tribal conflicts in Africa. In a visit to Kenya as a short-term expert for the Japan International Cooperation Agency (JICA), I observed a cholera outbreak in a Somali refugee camp. Health facilities at the camp were extremely bad, with the occurrence of drought adding to the suffering in these regions. Many developing countries in tropical and subtropical regions have problems of insufficient food supplies due to recent population growth. Disasters breaking out in these areas leave an unwelcome legacy of undernourishment and malnutrition, especially among young children. In this situation, the number of fatalities due to diarrhea is very high. Such fatalities are fewer in developed countries, but diarrhea in children under 5 years of age in developing countries remains a serious problem. World Health Organization (WHO) statistics published this year show that the global number of deaths in 2010 of children under 5 was some 76 million, 10% of which suffered from diarrhea-related disease. Malaria, pneumonia, premature birth, birth asphyxia and neonatal sepsis are additional causes of these deaths. This special issue details the countermeasures taken against infectious diseases in recent large disasters. As stated above, no serious outbreaks of infection were observed in the Great East Japan Earthquake, but damage to a local atomic power plant was extensive, as reported by the mass media. In addition to refugees from the earthquake and tsunami, many residents near the atomic power plant were forced to move out of their homes and towns because of the possible release of radiation, even though their homes had not been destroyed or even damaged. The lack of serious infectious disease outbreaks were more than made up for, however, by many problems with infectious disease. The subject of infectious disease risk and public health recovery is described by Dr. Hitoshi Oshitani of Tohoku University. Another article covers communicable diseases following the Great Earthquake described by Dr. Kentaro Iwata of Kobe University. The Great Earthquake and resulting tsunami in Sumatra, Indonesia, in December 2005 left more than 2,300,000 victims. Because this happened in a tropical region, public health control, especially food sanitation, was the worst problem. Dr. Nasronudin of Airlangga University in Indonesia communicates his experiences in this situation. The 2011 Haiti earthquake also involved a tropical region. As a Central American country, Haiti has had no experience with cholera in nearly a century and faces a cholera outbreak after the earthquake. Dr. G. B. Nair a cholera specialist at the Translational Health Science and Technology Institute of India, investigated the situation in Haiti and found that the causative strain was an Asian epidemic cholera. We therefore asked Dr. Nair to write about the Haiti cholera epidemic. Global microbial culture collection facilities have many microbial stocks that, if somehow released by a disaster, would cause at least two serious problems – one of environmental pollution by pathogenic organisms triggering infectious disease and another of the loss of valuable microbial resources. This makes it vital to maintain safe, secure culture collections against disasters. Dr. Takayuki Ezaki, Gifu University, describes this subject. Finally, we thank the authors for their contributions and the reviewers for their invaluable comments.

Comparative study of the commonly used virulence tests for laboratory diagnosis of ovine footrot caused by Dichelobacter nodosus in Australia

Footrot in sheep and goats is expressed as a spectrum of clinical entities ranging from benign, which is a self limiting interdigital dermatitis to highly virulent, in which severe under running of the horn of the hoof occurs. Interactions between the host, the virulence of the causative strain of Dichelobacter nodosus and environmental conditions determine the severity of the disease. Clinical diagnosis of virulent footrot, which a notifiable disease in some states of Australia, is not always straightforward. Therefore, the gelatin gel and elastase tests for protease activity, and the intA PCR test for an inserted genetic element in D. nodosus are commonly used to support or to confirm a clinical diagnosis. A comparative study of these laboratory tests with a large number of samples collected from 12 flocks of sheep with clinically virulent footrot was conducted. Based on the elastase test, 64% of the isolates tested were classified as virulent compared to 91% on the gelatin gel test and 41% according to the intA test. The agreement between the elastase and the gelatin gel test was low (kappa=0.12) as were the agreements between other tests. Only about 21% of the isolates were virulent in all 3 tests. Therefore these tests on their own may not provide standard and reliable results and are likely to remain as supplementary tests for clinical diagnosis of the disease.

Feline toxoplasmosis in Finland

Three subgroups of the Finnish cat population underwent investigation for different aspects of feline toxoplasmosis. Blood samples of 445 purebred pet cats and 45 shelter cats were screened for Toxoplasma gondii-specific immunoglobulin G antibodies with a direct agglutination test. The overall seroprevalence was 48.4%; older cats and cats receiving raw meat in their diet were more often seropositive. Fecal samples were obtained from 131 shelters cats; 2 of the cats were found shedding T. gondii-like oocysts, and the oocysts shed by 1 of the 2 were confirmed as T. gondii with polymerase chain reaction. Among 193 cats submitted for necropsy during a 3.5-year period, 6 (3.1%) had been diagnosed with generalized toxoplasmosis and were retrospectively further investigated. The main pathological lesions included acute interstitial pneumonia, acute necrotizing hepatitis, and nonsuppurative meningoencephalitis with glial granulomas. Immunohistochemical staining demonstrated a mild to massive parasite burden in tissues with pathological lesions as well as in unaffected tissues. The results of the direct multilocus genotyping of T. gondii parasites detected were consistent with endemic genotype II, and the causative parasite strains were isolated from 2 of the generalized toxoplasmosis cases. The results indicate that cats in Finland commonly encounter T. gondii and contribute to the environmental oocyst burden, while the endemic genotype II can also prove fatal to the parasite's definitive host. Preventing feline T. gondii infections is not only of public health importance but also a welfare issue for the cats themselves.

Negative Impact of Linezolid on Human Neutrophil Functions in vitro

Background/Aims: In a recent phase III clinical trial on linezolid, more patients in the linezolid treatment arm acquired Gram-negative catheter-related bloodstream infections despite the adequate therapy of infections caused by Gram-negative bacteria. We tested our hypothesis that linezolid impairs phagocytosis and the killing of Gram-negative bacteria by polymorphonuclear leukocytes (PMN). Methods: The influence of clinically relevant concentrations (5, 20 and 50 mg/l) of linezolid on granulocyte function in vitro was tested. Phagocytosis was determined by flow cytometry, and killing of bacteria was evaluated by plate counting. Chemotaxis was examined by an under-agarose cell migration assay. Gram-positive and Gram-negative bacteria were used. Results: Linezolid significantly impaired phagocytosis of a specific Escherichia coli strain in a concentration-dependent manner, whereas the effect on Pseudomonas aeruginosa was less prominent. No such effects were observed with a different E. coli strain or Staphylococcus aureus. Neither killing nor the chemotactic behaviour of PMN was significantly affected by linezolid. Conclusions: The observed concentration-dependent impairment of the phagocytic function might contribute to the higher frequency of catheter-related Gram-negative bloodstream infections in patients treated with linezolid. Individual patient risk may also depend on the causative Gram-negative strain.

Complete genome sequencing and evolutionary analysis of dengue virus serotype 1 isolates from an outbreak in Kerala, South India

In this study, dengue virus (DENV) isolates from a localized, small-scale, non-seasonal dengue outbreak were genetically characterized. The outbreak occurred during the pre-monsoon months (April-May) in a medical college campus in Kerala, South India in 2009 affecting 76 people. Analysis of 39 viral RNA positive serum samples by a serotype specific reverse-transcription polymerase chain reaction identified dengue virus serotype 1 (DENV1) as the causative strain. Formation of a distinct genetic clade was revealed in the initial phylogenetic analysis using nucleotide sequences of a partial (303 bp) Capsid-Pre-membrane protein (C-PrM) coding region of 37 outbreak strains. The sequences of these strains clustered with that of the Genotype III DENV-1 strains from India, and 32 among them formed a single major sub-clade. Whole-genome sequencing (10,693 bp) of two strains (RGCB585/2009 and RGCB592/2009) selected from this major sub-clade, and subsequent phylogenetic analysis using the full-length coding region sequence showed that the sequences grouped with that of the isolates from Thailand (1980), Comoros (1993), Singapore (1993), and Brunei (2005) among the Indo-Pacific isolates. The sequences of the two strains had a nucleotide identity of 97-98 % and an amino acid identity of 98-99 % with these closely related strains. Maximum amino acid similarity was shown with the Singapore 8114/93 isolate (99.6 %). Four mutations-L46M in the capsid, D278N in the NS1, L123I, and L879S in the NS5 protein coding regions-were seen as signature substitutions uniformly in RGCB585/2009 and RGCB592/2009; in another isolate from Kerala (RGCB419/2008) and in the Brunei isolate (DS06-210505). These four isolates also had in common a 21-nucleotide deletion in the hyper-variable region of the 3'-non-translated region. This first report on the complete genome characterization of DENV-1 isolates from India reveals a dengue outbreak caused by a genetically different viral strain. The results point to the possibility of exotic introduction of these circulating viral strains in the region.

Response of Different Antibiotic Resistant Group of Streptococcus pyogenes to Environmental Stresses

Streptococcus species is considered as an important pathogen for human and animals. The antibiotic resistance mechanism in this species is continuously increased. On the other side, the tolerance of environmental stresses play an effective role in the severity of many streptococcal causative disease. In this study we assayed survey on the causative agents of pharyngitis and tonsillitis patients. The predominant causative strain was Streptococcus pyogenes with 93% isolating ratio frequency. The other pathogenic species were S. agalactia 5.3% and S. pneumonia 1.7%. According to the antibiotic resistant test the S. pyogenes isolates were classified into six different groups. A selected strain from each antibiotic resistant group was tested for tolerance of a restrictive environmental factors. The variations of the environmental niches of isolates were in consistence with their antibiotic resistant variation.

Cardiovascular implantable electronic device endocarditis treated with daptomycin with or without transvenous removal

Background and MethodsNine patients with cardiovascular implantable electronic device (CIED) endocarditis were treated with daptomycin after the failure of previous treatment. The blood and CIED lead cultures of 1 patient were negative. In the other 8 patients, we observed 6 monomicrobic infections and 2 polymicrobic infections. Overall, 10 strains were isolated in these patients: 4 methicillin-sensitive Staphylococcus aureus, 2 methicillin-sensitive Staphylococcus epidermidis, 1 methicillin-resistant Staphylococcus aureus, 1 methicillin-resistant Staphylococcus epidermidis, 1 methicillin-sensitive Staphylococcus hominis, and 1 Propionibacterium acnes. The CIED was removed transvenously in 7 patients. Two patients were too sick for the removal of their CIED, and were cured with 6 mg/kg of daptomycin for 60 and 110 days, respectively, without adverse events. ResultsOne patient died 4 days after the removal of his CIED because of a complicated abdominal aortic aneurysm. The other 8 patients were cured, with a mean follow-up of 17 ± 8 months. The removed leads were negative, after daptomycin therapy, in 4 cases out of 7. The mean ratio between peak daptomycin concentration and minimal inhibitory concentration (MIC) of the causative strains was 38.3 ± 18.5. For patients whose data were available, the ratio between peak daptomycin concentration and minimal bactericidal concentration (MBC) was 13.2 ± 3.2. ConclusionDaptomycin monotherapy may be a useful therapeutic tool in difficult-to-treat CIED endocarditis, resulting in a high rate of cures and sterilized leads removed. The ratio between peak daptomycin concentration and MIC or MBC may be useful as predictive tool for treatment success.