Ongoing outbreak of West Nile virus infection in humans, Greece, July to August 2011.

Between 16 July and 21 August 2011, 31 cases of West Nile neuroinvasive disease were reported from four regions in Greece. Of these, 17 occurred in districts that had not been affected in 2010. The reoccurrence of human cases in two consecutive years (following the large 2010 outbreak) and the spread of the virus in new areas suggest that West Nile virus is established in Greece, and its transmission may continue to occur in the future.

Leishmaniases in Northern Greece: seroprevalence of the infection and incidence of the disease during the period 2001-2006.

Increasing risk factors are making leishmaniases a growing public health concern for many countries around the world. The aim of this study was to assess the seroprevalence of Leishmania infantum infection in the general population and in HIV infected subjects of Northern Greece, bordering the Mediterranean basin where leishmaniasis is endemic. The clinical cases of the disease during the last 6 years (2001-2006) are also presented. A low frequency of L. infantum antibodies was found by IFA and ELISA in 1,525 healthy individuals (2.8%), aged 18-80 years, living in the 16 prefectures of Northern Greece (Macedonia and Thrace regions), and in 167 HIV positive subjects (0.6%). Fifty-seven clinical cases were diagnosed in the same area and an approximate annual incidence of 0.34/100,000 was estimated. No endemic foci were identified and the cases of the disease were sporadic. Most presented with the visceral form (VL), few with the cutaneous, and one with VL-HIV co-infection. A significant shift in the age of people at risk was observed, with children less affected than adults (children/adults ratio: 0.36). No relevant data from previous studies are available to demonstrate a possible change of the infection in Northern Greece. The results of this study could be used as a reference for leishmaniasis surveillance in the area.

Evaluation and comparison of fluorescence polarization assay with three of the currently used serological tests in diagnosis of human brucellosis.

Fluorescence polarization assay (FPA) is a method that has been used for the diagnosis of brucellosis in animals for many years. To test its possible usefulness for the diagnosis of human brucellosis, 230 sera from patients with clinical signs of brucellosis and positive serological tests (Rose Bengal, Standard Agglutination Test, iELISA), and 305 sera from a healthy population with no clinical/epidemiological/serological evidence were examined with FPA. By using ROC analysis, the cut-off value was estimated at 99 mP, with 93.5% sensitivity (95% CI 89.5-96.3) and 96.1% specificity (95% CI 93.2-97.9). The pairwise comparison of ROC curves between FPA and iELISA and between FPA and RBT revealed no significant statistic difference (P < 0.05). On the contrary it revealed a significant statistic difference between FPA and SAT (P > 0.05). SAT also had the lowest sensitivity (81.7%) among the three tests used in case definition while iELISA had a sensitivity of 90.8% and RBT a sensitivity of 88.7%. The Kappa analysis showed that FPA has a very good agreement (0.92) with the "status of the disease" and with iELISA (0.837). According to our results, FPA seems to be a valuable method for the diagnosis of brucellosis in humans. Taking into consideration the advantages of the method such as the speed of results obtaining, the objectivity of results interpretation, as well as the cost, FPA could be considered as a replacement for other established methods. However, further studies are needed to assess the reproducibility of FPA.

Nationwide surveillance of Streptococcus pneumoniae in Greece: patterns of resistance and serotype epidemiology.

This nationwide study assessed the antimicrobial susceptibility and seroprevalence of Streptococcus pneumoniae in paediatric carriage isolates and in clinical isolates from adult pneumococcal disease in Greece during the years 2004-2006. Among 780 isolates recovered from the nasopharynx of children <6 years old attending day-care centres, non-susceptibility rates to penicillin, cefuroxime, ceftriaxone, erythromycin, tetracycline and trimethoprim/sulfamethoxazole were 34.7%, 25.1%, 1.0%, 33.5%, 26.4% and 44.2%, respectively. Among 89 adult clinical isolates, the respective rates were 48.3%, 46.1%, 5.6%, 48.3%, 32.6% and 40.4%. High-level resistance to penicillin, cefuroxime and ceftriaxone was recorded for 14.4%, 23.3% and 0.1% of paediatric carriage isolates, whereas for clinical adult isolates the respective rates were 25.8%, 38.2% and 2.2%. No resistance to levofloxacin and moxifloxacin was recorded, although 3.5% of paediatric carriage isolates and 23.2% of adult clinical isolates had minimum inhibitory concentrations of ciprofloxacin >2mg/L. Serotypes 19F, 14, 23F and 6B were the most prevalent among carriage and clinical isolates. The 7-valent pneumococcal conjugate vaccine was estimated to provide coverage against 71.7% of paediatric carriage isolates and 51.3% of adult clinical isolates. Resistance rates among clinical isolates from adult sources were higher than those recorded among paediatric carriage S. pneumoniae isolates and displayed an increasingly resistant profile compared with previous reports from our country, warranting continuous vigilance.

Diarrheic shellfish poisoning due to toxic mussel consumption: the first recorded outbreak in Greece.

During the week of 14-20 January 2000, 120 people visited the Emergency Departments of hospitals in Thessaloniki, northern Greece, complaining of acute gastrointestinal illness after eating mussels (Mytilus galloprovincialis). The symptoms indicated diarrhoeic shellfish poisoning, and the toxicity of mussels harvested from Thermaikos Gulf in Thessaloniki during the outbreak was investigated using mouse bioassays. The bioassays revealed toxicity to mice by the mussel samples; while high numbers of toxic algae Dinophysis acuminata were identified in water samples from Thermaikos Gulf. The harvesting of mussels was immediately suspended and a monitoring programme for algal blooms was established from then onwards. During a follow-up of the mussels' toxicity from January 2000 to January 2005, two more mussel samples were found positive for diarrheic shellfish poisoning: one harvested in March 2001 from the area of the outbreak (Thermaikos Gulf) and the other harvested in January 2001 from Amvrakikos Gulf in north-western Greece. However, no sporadic cases or outbreaks were reported during this period.

Repeated occurrence of diverse extended-spectrum beta-lactamases in minor serotypes of food-borne Salmonella enterica subsp. enterica.

Screening of Greek nontyphoid salmonellae from 2000 to 2002 yielded three extended-spectrum beta-lactamase (ESBL)-producing human isolates. Salmonella enterica serotype Brandenburg harbored a multiresistant SHV-5 gene-carrying plasmid. S. enterica serotype Blockley and S. enterica serotype Hadar harbored a TEM-52 gene-carrying plasmid. An S. enterica serotype Virchow strain producing plasmid-mediated CTX-M-32 was isolated twice from poultry end products. All ESBL plasmids were self-transferable and carried by clones currently common in Greece.

Isolation of a rare Escherichia coli O157:H7 strain from farm animals in Greece.

A strain of Escherichia coli O157:H7 was isolated from goat faeces during a surveillance study on the prevalence of this serotype of E. coli in farm animals in Greece. Three hundred and fifty one faecal samples were collected from goat, sheep and cattle breeding farms in the area of Epirus, Northwestern Greece. The E. coli O157:H7 isolate was nonsorbitol-fermenter, produced only VT2 and showed a beta-glucuronidase positive activity, a rather unusual biochemical feature for the E. coli O157:H7 serotype. No other strain of E. coli O157:H7 was isolated from the faecal samples of the rest farm animals examined, thus the overall prevalence of animal carriage was found to be 0.2%. The findings also indicate that goats can be a reservoir of E. coli O157:H7 and goat milk, dairy products and meat may serve as a vehicle for the pathogen transmission to humans.

Isolation of Escherichia coli O157:H7 from foods in Greece.

The presence of Escherichia coli O157:H7 in various foods of animal origin was surveyed in northwestern Greece. Six hundred samples of unpasteurized cows', ewes' and goats' milk, raw minced meat, uncooked frozen beef hamburgers, sandwiches (containing ham or turkey, mixed vegetable salad with mayonnaise and lettuce), fresh traditional Greek pork sausages and swine intestines appropriate for traditional Greek kokoretsi were assayed for E. coli serogroup O157:H7 using the standard cultural method and the immunomagnetic separation technique. The pathogen was detected in 1 out of 100 (1.0%) samples of ewes' milk, 1 out of 75 (1.3%) fresh sausages and 1 out of 50 (2.0%) swine intestines prepared for kokoretsi. The isolated strains were nonsorbitol fermenters, MUG-negative, O157 agglutinating, verotoxin-producing and carried both VT1 and VT2 genes. The three isolated strains were tested for antibiotic resistance and were found to be susceptible to eight antimicrobial agents (ampicillin, chloramphenicol, kanamycin, nalidixic acid, norfloxacin, streptomycin, sulfamethoxazole-trimethoprim and tetracycline).

Serum is the preferred clinical specimen for diagnosis of human brucellosis by PCR.

Human brucellosis poses a significant public health problem in many developing countries and requires fast and accurate diagnosis. A PCR assay amplifying part of the 31-kDa Brucella abortus antigenic protein gene sequence was developed and applied to whole-blood and serum samples from 31 brucellosis patients and 45 healthy individuals. All patients except one had detectable Brucella DNA in either whole blood or serum (combined sensitivity, 97%), but the assay sensitivity was higher with serum samples (94%) than with whole-blood samples (61%). The assay specificity was excellent (100%). A confirmatory PCR assay targeting another Brucella gene region (omp-2) was also developed but lacked sensitivity. Serum is the optimal specimen for the diagnosis of brucellosis by PCR, a choice that leads to assay simplification and shortens turnaround time.

Prospective use of RFLP analysis on amplified Cryptococcus neoformans URA5 gene sequences for rapid identification of varieties and serotypes in clinical samples.

Clinical isolates of Cryptococcus neoformans, whole blood, cerebrospinal fluid, bronchoalveolar lavage fluid from patients with positive cryptococcal antigen latex-agglutination test, and spiked clinical material from healthy individuals, were tested by polymerase chain reaction (PCR) with primers amplifying C. neoformans URA5 gene sequences. To test compatibility of different DNA extraction protocols with the PCR-restriction fragment length polymorphism (RFLP) assay, a commercial DNA extraction kit (XTRAX; Gull Laboratories, UT, USA) was used alongside with the hexadecyltrimethylammonium bromide (CTAB) method on spiked biological fluids. Both methods extracted DNA from spiked clinical samples containing C. neoformans (8 +/- 2 cells ml(-1)) and generated amplification products suitable for restriction enzyme analysis. Alu I digestion differentiated the two varieties of C. neoformans. Three distinct RFLP patterns were obtained upon restriction with MspI corresponding to serotypes A, AD and B, C and D. URA5 PCR followed by RFLP analysis, coupled with a sensitive in-house or commercially available DNA extraction method from clinical samples, could be successfully incorporated into rapid routine diagnostic strategies. It could also provide an expeditious tool for epidemiology-based population genetics studies.

Haemorrhagic papular rash associated to Flavimonas oryzihabitans bacteraemia in a child.

Flavimonas oryzihabitans is a gram-negative rod that has rarely been implicated in human infections. The involvement of this organism has been documented in serious infections, the majority of which were cases of bacteraemia or peritonitis. We report the first isolation of the organism in Greece, from a case of bacteraemia, associated with haemorrhagic papular rash, in a paediatric patient and describe the phenotypic characteristics of the strain.

Shigellosis of childhood in northern Greece: epidemiological, clinical and laboratory data of hospitalized patients during the period 1971-96.

The aim of this study was to evaluate epidemiological, clinical and laboratory data of shigellosis in children from northern Greece, hospitalized in our department during the period 1971-96. In total, 422 cases of shigellosis, aged 1 month to 14 y (238M, 184F) were hospitalized during the study period. The annual distribution was approximately stable until 1990, the mean number of cases per year being about 20. During the last 4 y the incidence significantly decreased. Shigella was serotyped in 138/422 cases. Seventy six of the strains were S. flexneri (55%) and 56 S. sonnei (40%). In the majority of cases the clinical picture was mild. Severe dehydration was seen in only 6 patients. Ninety four patients (22%) had extra-intestinal manifestations. Most common of these were convulsions (16%) and, less frequently, disturbances of consciousness (n = 26), rash (n = 9), shock and disseminated intravascular coagulopathy (n = 2), nerve paralysis (n = 2), severe anaemia (n = 2) and haemolytic-uraemic syndrome (n = 1). Nine patients had acute encephalopathy of 12 h to 12 d duration. It is important to note that all these cases recovered completely with no residual neurological deficit, except for 1 girl who developed temporal epilepsy 8 y later. Spinal fluid was normal in all 42 examined patients. Antibiotics were given to 212 of 422 patients, mainly during the first half of the study period. Shigella resistance to antibiotic was significant for cotrimoxazole (24%) and ampicillin (16%). All patients were cured. Shigellosis is a mild disease in our area, with a decreasing prevalence.

Listeria meningitis in children: report of two cases.

We report two cases of meningitis caused by Listeria monocytogenes in children. The first patient was a healthy 14-month-old boy and the second patient a 3-year-old girl with Byler disease which, however, is not reported as a predisposing factor for listeriosis. We present these cases because Listeria infection, although common in neonates, is extremely infrequent during infancy and childhood.

Recent emergence of serogroup C meningococcal disease in Greece.

The number of cases of meningococcal disease reported to the Meningitis Reference Laboratory in Athens rose dramatically in 1996-1997. The aims were (1) to determine if the increase was due to introduction of new strains, (2) to assess the geographic and age distribution of the cases, (3) to compare antibiotic sensitivity patterns of the current isolates with strains from the early 1990s. In 1993-1994, 15/19 (74%) of the cases for which information on age was available were in children < or = 5 years; in 1995-1997, 80/179 (45%) of cases were in children < or = 5 years and 99 (55%) in the older age range (P < 0.02). From 593 cases in 1993 1997, 214 (36%) isolates were available for characterisation. Serogroup B was predominant in the early 1990s, but by 1997, serogroup C accounted for 46/72 (64%) of isolates and serogroup B for 25/72 (35%). Serogroup B was predominant in children < or = 5 years (44/78, 56%) but only 19/99 (18%) of older children and adults (P=0.0000005). Sulfonamide resistance decreased from 10/22 (45%) in 1993-1994 to 27/192 (14%) in 1995-1997 (P<0.01). Multilocus enzyme electrophoresis of 70 strains obtained during this period identified the epidemic ET-15 clone in 24 (34.3)%. The profiles of the Greek ET-15 isolates were identical to C:2a:P1.2(P1.5) strains responsible for the epidemic in the Czech Republic which began in 1993. This genotype was not found in Greek strains isolated prior to 1993. We conclude that the increase in meningococcal disease is due to introduction of the epidemic serogroup C:2a:P1.2(P1.5) strain responsible for disease in the Czech Republic and Canada.

Non-typhoid Salmonella meningitis.

The epidemiological, clinical and laboratory findings in 8 cases of meningitis due to non-typhoid salmonella were analyzed. Seven out of 8 patients were infants < 12 months old, 2 of whom died, 2 presented with recurrent meningitis. Three infants had reversible neurological sequelae on discharge, not found during follow-up in any. Conversely, one 2-month-old patient who was considered normal on discharge developed severe growth and mental retardation during follow-up.

Treatment of childhood bacterial meningitis with ceftriaxone once daily: open, prospective, randomized, comparative study of short-course versus standard-length therapy.

Fifty-two children were included in this study to evaluate and compare short- versus standard-length ceftriaxone therapy for bacterial meningitis. The duration of the short-course regimens was 4, 6 and 7 days for Neisseria meningitidis, Hemophilus influenzae and Streptococcus pneumoniae, respectively. The standard-length regimens were twice as long. On the basis of a computer-generated randomization list, 26 children were assigned either to the short- or to the standard-treatment regimen. Ceftriaxone was given intravenously once daily in a dose of 60 mg/kg after an initial loading dose of 100 mg/kg. The population characteristics, the severity of disease and the cerebrospinal fluid (CSF) findings were similar in the two study groups at admission. Bacteriological and clinical response were comparable. There were no significant differences in the incidence of neurological complications, prolonged fever (greater than or equal to 10 days), persistent pleocytosis and side effects between the two groups. Hearing loss occurred in 3 patients in the standard-length group and in no patients in the short-course group. Diarrhea was the only side effect and occurred in 14% of the patients. The results of the study indicate that the short-duration regimen was adequate for the treatment of meningitis caused by the three major meningeal pathogens. However, the small number of patients do not justify the adoption of the short-course regimen for all children with meningitis. At present, prolongation of ceftriaxone therapy or discontinuation of the drug under strict clinical observation of the patient should be considered in some cases.