Real-time genomic investigation underlying the public health response to a Shiga toxin-producing Escherichia coli O26:H11 outbreak in a nursery.

Shiga toxin-producing Escherichia coli (STEC) is a significant cause of gastrointestinal infection and the haemolytic-uremic syndrome (HUS). STEC outbreaks are commonly associated with food but animal contact is increasingly being implicated in its transmission. We report an outbreak of STEC affecting young infants at a nursery in a rural community (three HUS cases, one definite case, one probable case, three possible cases and five carriers, based on the combination of clinical, epidemiological and laboratory data) identified using culture-based and molecular techniques. The investigation identified repeated animal contact (animal farming and petting) as a likely source of STEC introduction followed by horizontal transmission. Whole genome sequencing (WGS) was used for real-time investigation of the incident and revealed a unique strain of STEC O26:H11 carrying stx2a and intimin. Following a public health intervention, no additional cases have occurred. This is the first STEC outbreak reported from Israel. WGS proved as a useful tool for rapid laboratory characterization and typing of the outbreak strain and informed the public health response at an early stage of this unusual outbreak.

Contribution of real-time PCR to Plasmodium species identification and to clinical decisions: a nationwide study in a non-endemic setting.

Treatment choice for patients with malaria in Israeli hospitals is based on microscopy and rapid diagnostic tests (RDTs). Here, we demonstrate the cumulative value of real-time polymerase chain reaction (PCR) in optimizing the treatment of malaria. Between January 2009 and December 2015, 451 samples from 357 patients were tested in our laboratory using a real-time PCR assay. Hospital laboratory results (without real-time PCR) were compared to those obtained in our laboratory. A total of 307 patients had a malaria-positive laboratory finding in the hospital. Out of those, 288 were confirmed positive and 19 negative using real-time PCR. Two negative hospital results were found to be positive by real-time PCR. More specifically, of 153 cases positive for Plasmodium falciparum by real-time PCR, only 138 (90%) had been correctly identified at the hospitals. Similarly, 66 (67%) of 99 cases positive for P. vivax, 2 (11%) of 18 cases positive for P. ovale, and 3 (30%) of 10 cases positive for P. malariae had been correctly identified. Of 10 cases of mixed infection, only one had been identified as such at the hospital. Thus, real-time PCR was required for correct identification in 81 (28%) out of 290 positive cases. In 52 (18%) of those, there was an erroneous categorization of relapsing versus non-relapsing parasites. In a nationwide study, we found that the use of real-time PCR is definitely beneficial and may change the decision regarding the choice of treatment.

Molecular epidemiology of Campylobacter jejuni infection in Israel-a nationwide study.

The incidence of Campylobacter infection in Israel, particularly among children <2 years of age, has risen over the last decade and became one of the highest among industrialized countries. This study explored the molecular epidemiology of Campylobacter jejuni in Israel over a decade (2003-2012) using multilocus sequence typing (MLST) combined with demographic metadata. Representative clinical isolates (438) from a large national repository together with selected veterinary isolates (74) were subject to MLST. The distribution of age groups, ethnicity and clinical source across various genotypes was evaluated using Poisson modelling. The 512 studied isolates were assigned 126 distinct sequence types (STs) (18.8% novel STs) grouped into 21 clonal complexes (CCs). Most human, poultry and bovine STs clustered together in the leading CCs. Three dominant STs (ST21, ST6608, ST4766) were detected only since 2006. Patients infected with the leading CCs were similarly distributed along densely populated areas. The frequency of blood isolates was higher in patients infected with CC353 (relative rate (RR)=2.0, 95% CI 1.03-3.9, adjusted p value (adj.p) 0.047) and CC42 (RR=4.4, 95% CI 1.7-11.6, adj.p 0.018) and lower with CC257 (RR=0.3, 95% CI 0.1-0.9, adj. p 0.047). The distribution of age groups and ethnicity also varied across the leading CCs. In conclusion, C. jejuni isolates in a national sample appeared highly diverse with a high proportion of new STs. Phylogenic analysis was compatible with poultry and cattle as possible food sources of clinical infection. Demographic characteristics of the infected patients coupled with strain invasiveness across different genotypes revealed a complex epidemiology of C. jejuni transmission in Israel.

Design and application of a core genome multilocus sequence typing scheme for investigation of Legionnaires' disease incidents.

Sequence-based typing (SBT) for Legionella pneumophila (Lp) has dramatically improved Legionnaires' disease (LD) cluster investigation. Microbial whole genome sequencing (WGS) is a promising modality for investigation but sequence analysis methods are neither standardised, nor agreed. We sought to develop a WGS-based typing scheme for Lp using de novo assembly and a genome-wide gene-by-gene approach (core genome multilocus sequence typing, cgMLST). We analysed 17 publicly available Lp genomes covering the whole species variation to define a core genome (1,521 gene targets) which was validated using 21 additional published genomes. The genomes of 12 Lp strains implicated in three independent cases of paediatric humidifier-associated LD were subject to cgMLST together with three 'outgroup' strains. cgMLST was able to resolve clustered strains and clearly identify related and unrelated strains. Thus, a cgMLST scheme was readily achievable and provided high-resolution analysis of Lp strains. cgMLST appears to have satisfactory discriminatory power for LD cluster analysis and is advantageous over mapping followed by single nucleotide polymorphism (SNP) calling as it is portable and easier to standardise. cgMLST thus has the potential for becoming a gold standard tool for LD investigation. Humidifiers pose an ongoing risk as vehicles for LD and should be considered in cluster investigation and control efforts.

Recent trends in the epidemiology of shigellosis in Israel.

We provide an update on the epidemiology of shigellosis in Israel using data generated by a sentinel laboratory-based surveillance network for the period 1998-2012. The average annual incidence of culture-proven shigellosis was 97/100 000. We estimated that each case of shigellosis accounted for 25 cases in the community indicating the high burden of disease. Orthodox Jewish communities, living in highly crowded conditions and with a high number of children aged <5 years were the epicentre of country-wide biennial propagated epidemics of S. sonnei shigellosis. S. flexneri was the leading Shigella serogroup in Israeli Arabs. S. flexneri 2a and S. flexneri 6 alternated as the most common serotypes. Both S. sonnei and S. flexneri isolates showed high rates of resistance to ampicillin and trimethoprim/sulfamethoxazole and very low rates of resistance to quinolones and third-generation cephalosporins. Shigellosis due to S. sonnei conferred 81% (95% confidence interval 69-89) protection against the homologous Shigella serotype when epidemic exposure re-occurred 2 years later. These data are of value in the process of Shigella vaccine development.

Risk factors for sporadic infection with Salmonella Infantis: a matched case-control study.

This matched case-control study investigated the risk factors for sporadic Salmonella Infantis infection in 263 affected children and 263 age-, gender- and neighbourhood-matched controls. Information about exposure to potential risk factors was obtained via telephone interview and evaluated by conditional logistic regression analysis. Age groups ≤ 1 year (n=77) and >1 year (n = 186) were analysed separately. Of those aged ≤ 1 year, breastfeeding was a significant protective factor against infection [matched odds ratio (mOR) 0.24, 95% confidence interval (CI) 0.10-0.59, P < 0.01]. In the older group, consumption of eggs (mOR 1.87, 95% CI 1.00-3.49, P = 0.05) was a significant risk factor and thawing chicken in water (mOR 2.55, 95% CI 0.94-6.91, P = 0.07) was borderline risk factor, while consumption of carrots (mOR 0.46, 95% CI 0.26-0.83, P < 0.01), drinking tap water (mOR 0.44, 95% CI 0.22-0.85, P = 0.02), religious lifestyle (mOR 0.40, 95% CI 0.21-0.74, P < 0.01) and having a high number of children in the household (mOR 0.72, 95% CI 0.58-0.88, P < 0.01) were significant protective factors. Consumers should avoid eating undercooked eggs and food handlers should be educated regarding proper handling and cooking of eggs. Breastfeeding should be strongly encouraged by public health authorities. The public must be educated on stringent hygiene practices, especially proper cooking of eggs to reduce infection rates.

Geographical variations in Salmonella incidence in Israel 1997-2006: the effect of rural residency.

The overall incidence and serotype distribution of non-typhoid Salmonella (NTS) may vary between different geographical localities. To investigate possible regional differences and the effect of demographic factors, we studied 15 865 episodes of laboratory-confirmed NTS infection in Israel. Using Poisson models we found significant variation in the average annual incidence rate of NTS in 15 administrative sub-districts, which was inversely associated with the percent of rural residency (incidence rate ratio 0.75, 95% confidence interval 0.65­0.86, P<0.001). Variation was also found in the relative incidence of the most prominent serotypes (Enteritidis, Virchow, Typhimurium, Hadar, Infantis), which was affected by rural residency, the percent of non-Jewish population in the sub-district, and the percent of population aged o55 years in the sub-district.Rural residency had a major effect on the epidemiology of salmonellosis in Israel. Future research is required to understand whether decreased incidence in rural areas is an under-detection bias or reflects true differences in NTS illnesses.

Recent trends in the epidemiology of non-typhoidal Salmonella in Israel, 1999-2009.

The aim of the present study was to assess the recent trends in the epidemiology of non-typhoid Salmonella in Israel using a sentinel laboratory-based surveillance network. Between 1999 and 2009, 8758 Salmonella stool isolates were reported by five sentinel laboratories. There was a significant decrease in the incidence rate of Salmonella isolates from 70·5/100,000 in 1999 to 21·6/100,000 in 2005 followed by a slight increase to 30·3/100,000 in 2009. Of all Salmonella, 64·3% were isolated from children in the 0-4 years age group. Up to 2008, S. Enteritidis was the most prevalent serotype and in 2009 S. Infantis emerged as the most common Salmonella serotype. The decrease in the incidence of S. Enteritidis and S. Typhimurium and increase in S. Infantis among humans were associated with a similar trend among breeding flocks, which followed significant preventive interventions conducted against S. Enteritidis and S. Typhimurium infections in poultry. Tight surveillance and education of food handlers and consumers should be enhanced to reduce the foodborne transmission of Salmonella in Israel.

Curtailed short-term and long-term survival following infection with non-typhoid Salmonella in Israel.

Among bacterial foodborne pathogens, non-typhoid Salmonella enterica (NTS) is a leading cause of death worldwide. This study assessed short-term and long-term mortality following NTS infection in Israel, and evaluated the effects of age, sex, source of isolation and different serotypes on mortality. The source of data was a national registry of NTS isolates submitted to the Salmonella Reference Center, Government Central Laboratories, in Jerusalem, Israel, during 1997-2006. Vital status was derived from the registry of the Israeli Ministry of the Interior. The survival of a cohort of 15,919 patients infected with the top five NTS serotypes was evaluated by calculating age-standardized mortality ratios (SMRs) and by Cox proportional hazards multivariate regressions at three follow-up time intervals: 30 days, 1 year and end of follow-up. The median follow-up time was 6.44 years (mean, 6.21 years; range, 1 day to 10.80 years). The cumulative crude mortality rates at the three time intervals were 0.68%, 1.86% and 4.40%, respectively, corresponding to increased SMRs of 16.95 (95% CI 13.9-20.46), 4.25 (95% CI 3.78-4.76), and 1.83 (95% CI 1.70-1.97), respectively. Cox regressions revealed that increasing age, extraintestinal source of isolation and NTS serotype had significant effects on mortality within all three follow-up intervals. The risk of mortality was increased for serotypes Infantis and Typhimurium, and decreased for serotypes Virchow and Hadar, as compared with serotype Enteritidis. The study revealed curtailed short-term and long-term survival following NTS infection that persisted for many years following detection by culture.

The epidemiology of extraintestinal non-typhoid Salmonella in Israel: the effects of patients' age and sex.

Extraintestinal disease occurs in 5-8% of non-typhoid Salmonella enterica (NTS) infections and is more likely to be associated with hospitalization and death. The study examined the epidemiology of extraintestinal NTS infections in Israel and the possible effects of patients' age and sex. NTS isolates passively submitted to the National Salmonella Reference Center during 1996-2006 were the source for the study cohort. Poisson regression models were used to assess incidence trends over the study years and to evaluate the effects of patients' age and sex on the incidence of extraintestinal NTS manifestations. A total of 36,822 stool and 1,415 (3.7%) patient-unique NTS isolates from blood (74.1%), urine (18.3%), and other sources (3.7%) were studied. Serotypes Enteritidis, Virchow, and Typhimurium accounted for 66.3% of the isolates. Analysis showed a highly significant quadratic (U-shaped) relationship between patients' age and the incidence of extraintestinal isolation (p < 0.001), with increasing risk in the two extremes of age. Differences between the incidence of blood and urine sources were significant in patients <10 and >or=60 years old (relative risk [RR] = 5.88, 95% confidence interval [CI] 3.36-10.30, p < 0.001 and RR = 1.66, 95% CI 1.09-2.53, p = 0.017, respectively). Males >or=60 years of age were more likely than females of the same age to have bacteremia (RR = 1.90, 95% CI 1.39-2.61, p > 0.001) and less likely to have urinary NTS isolation (RR = 0.50, 95% CI 0.28-0.89, p = 0.018). Serotype Virchow had the highest incidence in patients <10 years of age, while serotype Enteritidis had the highest incidence in patients >or=60 years old. The study revealed a complex effect of patients' age and sex on the epidemiology of extraintestinal NTS manifestations.

The changing panorama of bacterial enteric infections.

We studied the age-specific population-based incidence of bacterial enteric infections caused by Shigella, Salmonella and Campylobacter, in Jerusalem. During 1990-2008, 32,408 cases were reported (incidence rate 232.1/100,000 per annum). The patterns of Shigella (47.4% of cases), Salmonella (34.4%) and Campylobacter (18.2%) infections evolved noticeably. Campylobacter rates increased from 15.0 to 110.8/100,000 per annum. Salmonella rates increased from 74.2 to 199.6/100,000 in 1995 then decreased to 39.4/100,000. Shigella showed an endemic/epidemic pattern ranging between 19.7 and 252.8/100,000. Most patients (75%) were aged <15 years; children aged <5 years comprised 56.4% of cases, despite accounting for only 12.9% of the population. Campylobacter was the predominant organism in infants aged <1 year and Shigella in the 1-4 years group. The hospitalization rates were: Shigella, 1.8%; Campylobacter, 2.3%; Salmonella, 6.9%. Infants were 2.2 times more likely to be hospitalized than children aged 1-14 years (P=0.001). Household transmission occurred in 21.2% of Shigella cases compared with 5% in the other bacteria.

A survey of laboratory-confirmed isolates of invasive listeriosis in Israel, 1997-2007.

During the 11-year period from 1997 to 2007, 321 isolates of Listeria monocytogenes from sporadic cases of invasive listeriosis were reported to the national reference laboratory in Israel. Of these isolates, 113 (35%) were identified from perinatal cases, and 208 (65%) from non-perinatal cases. The prevalent serovars were 4b, 1/2b, 1/2a and 4c. Serovar 4b was identified in 80.5% of the perinatal isolates (P=0.0162), while the number of 1/2b and 1/2a strains increased in the > or = 60 years old group (P=0.0285). Resistance to tetracycline was found in eight 4b isolates. The seasonal distribution showed that 206 isolates (64.2%) were submitted during the hot season (May-October). The estimated morbidity for the study period was 4.4 per million. The incidence of invasive listeriosis was higher in the perinatal group (5.6/100 000), than in individuals aged > or = 60 years (1.5/100 000).

Prevalence and correlates of diphtheria toxoid antibodies in children and adults in Israel.

A seroepidemiological study was performed to evaluate immunity to diphtheria and to determine the correlates of diphtheria toxoid antibody levels among children and adults in Israel. In total, 3,185 sera from an age-stratified sample of children and adults, obtained in 2000-2001, were tested for diphtheria toxoid antibodies by an in-house double-antigen ELISA. A level of or=0.1 IU/mL (full protection or seropositivity). Seronegativity increased significantly in subjects aged >50 years, reaching levels of 9.7%, 12.6% and 18.9% in the groups aged 50-54, 55-59 and >60 years, respectively (p 0.001), with rates of basic immunity following a similar pattern. Subjects born abroad had higher seronegativity rates than those born in Israel (7.7% vs. 4.9%; p 0.019). No difference in diphtheria toxoid antibody levels was found according to other demographical variables, such as gender, Jewish or Arab ethnicity, urban or rural settlements, and the subjects' place of residence. The level of immunity to diphtheria among children and adults in Israel was satisfactory, with the exception of individuals aged >50 years. The risk of diphtheria outbreaks is low, but sporadic cases may occur among individuals lacking basic immunity against the disease.

Salmonella enterica serotype Virchow: epidemiology, resistance patterns and molecular characterisation of an invasive Salmonella serotype in Israel.

This study outlines the unique epidemiology of Salmonella enterica serotype Virchow in Israel. Between 1997 and 2002, the overall incidence of non-typhoid Salmonella enterica (NTS) decreased from 69.3 to 53.3 infections/100,000 population, but the incidence of S. Virchow increased (from 7.2 to 9.1 infections/100,000). Since 2000, S. Virchow has become the second-ranking NTS isolate, accounting for 17% and 27% of all stool and blood NTS isolates, respectively. Infants aged < 1 year had the highest incidence of isolation from stools (92.8/100,000). The incidence of isolation from blood was highest for infants aged <1 year (4.4/100,000). Only 6% of isolates were susceptible to all ten antibiotic agents tested; 34% were resistant to one agent, 54% to one to three agents, and 40% to four to six agents. A high proportion of the tested isolates were resistant to nalidixic acid (89%), streptomycin (56%), tetracycline (43%), trimethoprim-sulphamethoxazole (38%) and chloramphenicol (28%), but none to ciprofloxacin or ceftriaxone. Pulsed-field gel electrophoresis revealed two closely related clusters, each containing a predominant pulsotype. Coupled with its invasive propensity, the increasing incidence of highly resistant S. Virchow in Israel is of real concern. Future research should focus on the sources of S. Virchow in the food chain in order to institute effective control measures.

Variability of Shigella flexneri serotypes in Israel during a period of two years: 2000 and 2001.

During a period of 2 years (2000 and 2001) 996 Shigella flexneri strains from sporadic cases in Israel were sent to the National Shigella Reference Centre (NSRC) by hospital and outpatient clinics. The most common serotypes were 2a, 6 and 1b, accounting for 88.4% of all isolates. They were investigated according to the monthly distribution of the strains, and the age and sex of the patients. The severity of the disease was assessed by a hospital/outpatient distribution (H/Od) of the isolates, based on the location of the sending laboratory. The most affected age groups were 0-11 months and 1-4 years, and the prevalent serotype was 2a, while serotype 6 was dominant in the 5-14 years age group. More cases were registered during the hot season, and there were some serotype-related variations. Overall, 62.1% of the samples were from male patients. Serotype 1b was dominant in the male/female ratio, although it was third in general prevalence. According to the H/Od serotype 2a was more common in hospitalized males and serotype 6 in outpatients, both male and female. These variations, as well as changes in serotype prevalence in the past, underscore the importance of serotype monitoring as part of the public health strategies for reducing the burden of Shigella flexneri infections.

Blood invasiveness of Salmonella enterica as a function of age and serotype.

We explored the dual influence of the patient's age and the infecting serotype on the blood invasiveness patterns of non-Typhi Salmonella enterica (NTS). Blood invasiveness ratio (BIR) was calculated as the ratio between the number of blood and blood + stool isolates. Analysis of 14,951 NTS isolates showed that the BIR increased drastically above the age of 60 years, reaching levels 3.5-7 times higher compared to age group < 2 years. Different patterns of age-related invasiveness were observed for the five most common NTS serotypes (Enteritidis, Typhimurium, Virchow, Hadar, Infantis). Among children < 2 years, the BIR was highest for serotype Virchow and lowest for serotype Hadar, while in persons > or = 60 years it was highest for serotypes Enteritidis and lowest for serotype Infantis. The tendency of NTS serotypes to invade the bloodstream was significantly influenced by the patient's age, however the impact of age differed for various NTS serotypes.

Microspectrofluorometry and fluorescence imaging in the study of human cytopathology.

The study of energy pools and dynamics of specific pathways in living cells by microspectrofluorometry and fluorescence imaging produces spectral and topographic images characterizing structural and functional changes associated with cytopathology. Microspectro-fluorometry and fluorescence imaging have been applied, together with organelle morphometry to a number of cells mimicking certain cytopathologies, including melanoma cells, long-term malignant cells, and gene-defective cells. These investigations of cellular pathology indicate that there is a convergence of various physiopathological processes. Cellular states that have similarities include senescence, detoxification, and transformation. While the NAD(P)H metabolic transients have been studied before, our emphasis in this article is on very rapidly scanned fluorescence images related to organelle integration and photoinduced cellular senescence.

Clinical, epidemiological, and microbiological features of Vibrio vulnificus biogroup 3 causing outbreaks of wound infection and bacteraemia in Israel. Israel Vibrio Study Group.

BACKGROUND: Vibrio vulnificus is a gram-negative bacterium that causes septicaemia and wound infection. Cases occur sporadically, and no previous outbreaks due to a common source or a clonal strain have been reported. In the summer and autumn of 1996 and 1997, an outbreak of invasive V. vulnificus infection occurred in Israel in people who had recently handled fresh, whole fish purchased from artificial fish-ponds. METHODS: We reviewed clinical and epidemiological information, and undertook an environmental investigation to assess disease characteristics, modes of transmission, phenotypic characteristics of the bacterium, and fish-marketing policy. The clonal nature of 19 isolates was studied by biotyping, pulsed-field gel electrophoresis, and restriction-fragment length polymorphism (RFLP) analysis of a PCR fragment. FINDINGS: During 1996-97, 62 cases of wound infection and bacteraemia occurred. 57 patients developed cellulitis, four had necrotising fasciitis, and one developed osteomyelitis. In all cases, the fish were cultivated in inland fish-ponds. In the summer of 1996, fish-pond managers initiated a new marketing policy, in which fish were sold alive instead of being packed in ice. Phenotypically, the isolates had five atypical biochemical test results. The isolates were non-typeable by pulsed-field gel electrophoresis, and all had the same PCR-RFLP pattern which had not been seen previously. INTERPRETATION: The cause of the outbreak was a new strain of V. vulnificus, classified as biogroup 3. A new fish-marketing policy that began in 1996 may have exposed susceptible people to the organism.

Multiprobe fluorescence imaging and microspectrofluorimetry of cell transformation and differentiation: implications in terms of applied biochemistry and biotechnology.

The dichotomy of cellular transformation versus differentiation does not preclude the hypothesis of a unified underlying mechanism that can switch either way as a result of growth factors, cell-membrane receptors, secondary messengers, integrating switch kinases and/or nuclear receptors. Its study for biopharmaceutical and biotechnological applications requires a methodology capable of dealing with such pleiotropy. In the multiprobe-multiparameter approach, one must remain wary of cumulative toxic effects and misinterpretations. 'Smart' instrumentation does not mean 'smart' probes. It turns out that the cell's own endogenous probes, the fluorescent coenzymes, may be akin to 'smart' probes, open to study in situ of many-fold interrelated pathways in cell energetics and dynamics. Resolution at the micro- and even nano-compartment levels is not altogether impossible. Thus an innovative search in terms of what may be called 'intracellular reconnaissance with fluorescent probes and biopharmaceuticals' necessitates recourse to multiple tentative probings along the pleiotropic mechanisms as far in resolution as one can go. Among the characteristic findings using this approach are: (i) morphometric alterations in the mitochondria and melanosomes of melanoma cells treated with azelaic acid; (ii) deregulation of mitochondrial control and extramitochondrial metabolism in similarly treated cells; (iii) considerable acceleration of NAD(P) transient kinetics in atractylate-treated L sarcoma cells; (iv) alterations of mitochondria and Golgi in fusion-deficient myoblasts; (v) tentative recognition of beta-glucosidase deficiency in Gaucher disease cells by the use of fluorescent and fluorogenic lysosomal probes; and (vi) UVA-induced accumulation of Schiff bases (a kind of accelerated photo-aging) in yeast and kidney epithelial cells. Because these studies utilize probing at whatever points along the concerned pathways become accessible, at first glance they may look disconnected. What and where is the connecting thread, for instance, between studying melanoma metabolism, melanosome morphometry, hepatocyte organelle morphogenesis and transformation, myotube organelle morphogenesis and fusion-non-fusion, and lysosomal activity in gene-deficient cells? In the mapping of the regulatory and deregulatory mechanisms involved in the switching of differentiation or transformation, each of the above topics carries an information content towards resolution of the pleiotropic puzzle. The integration of such information with increasing resolution and access to intracellular microdomains may ultimately allow focus on the precise target, the switch from differentiation to transformation or vice versa.