Chryssoula Nicolaou

Comparative antimicrobial susceptibility of biofilm versus planktonic forms of Salmonella enterica strains isolated from children with gastroenteritis

In the present study, 194 Salmonella enterica strains, isolated from infected children and belonging to various serotypes, were investigated for their ability to form biofilms and the biofilm forms of the isolated strains were compared to their corresponding planktonic forms with respect to the antimicrobial susceptibility. For the biofilm-forming strains, the minimum inhibitory concentration for bacterial regrowth (MICBR) from the biofilm of nine clinically applicable antimicrobial agents was determined, and the results were compared to the respective MIC values of the planktonic forms. One hundred and nine S. enterica strains out of 194 (56%) belonging to 13 serotypes were biofilm-forming. The biofilm forms showed increased antimicrobial resistance compared to the planktonic bacteria. The highest resistance rates of the biofilm bacteria were observed with respect to gentamicin (89.9%) and ampicillin (84.4%), and the lowest rates with respect to ciprofloxacin and moxifloxacin (2.8% for both). A remarkable shift of the MICBR50 and MICBR90 toward resistance was observed in the biofilm forms as compared to the respective planktonic forms. The development of new consensus methods for the determination of the antimicrobial susceptibility of biofilm forms seems to be a major research challenge. Further studies are required in order to elucidate the biofilm antimicrobial resistance mechanisms of the bacterial biofilms and their contribution to therapeutic failure in infections with in vitro susceptible bacteria.

Identification and antimicrobial resistance of campylobacter species isolated from animal sources.

Background: Campylobacter spp. are together with Salmonella spp. the leading causes of human bacterial gastroenteritis worldwide. The most commonly isolated species in humans are Campylobacter jejuni and C. coli. The isolation, identification, and antimicrobial resistance of Campylobacter spp. from poultry and raw meat from slaughterhouses, has been investigated for the first time in Greece. During the period from August 2005 to November 2008 a total of 1080 samples were collected: (a) 830 fecal samples from five poultry farms, (b) 150 cecal samples from chicken carcasses in a slaughterhouse, and (c) 100 fecal samples from one pig farm near the region of Attica. The identification of the isolates was performed with conventional (sodium hippurate hydrolysis and commercial identification system (Api CAMPY system, bioMerieux, France), as well as with and molecular methods based on 16S rRNA species specific gene amplification by PCR and subsequent sequence analysis of the PCR products. Results: Sixteen Campylobacter strains were isolated, all collected from the poultry farms. None of the strains was identified as C. jejuni. Antimicrobial susceptibility to six antimicrobials was performed and all the strains were susceptible to ciprofloxacin, amoxicillin–clavulanic acid, and gentamicin. Thirteen out of 14 C. coli were resistant to erythromycin and all C. coli strains were resistant to ampicillin. Conclusion: Our results emphasize the need for a surveillance and monitoring system with respect to the prevalence and antimicrobial resistance of Campylobacter in poultry, as well as for the use of antimicrobials in veterinary medicine in Greece.

An unusual case of insomnia associated with Whipple encephalopathy : first case reported from Greece

AbstractWhipple disease is a relapsing systemic illness caused by Tropheryma whippelii. Central nervous system involvement occurs in 5%–40% of all patients. Hypothalamic manifestations occur in 31% of Whipple encephalopathy, including polydipsia, hyperphagia, change in libido and insomnia. We report a case of a 48–year–old man with severe insomnia, depression, dementia, dysarthria, myoclonic movements of the limbs and ophthalmoplegia. The diagnosis of Whipple encephalopathy was confirmed by PCR analysis of blood and faeces. He received a full dose of antibiotic treatment. Despite clinical improvement, resolution of the lesions detected in MRI scan of the brain and negative results of the PCR in blood, faeces and cerebrospinal fluid six months later, insomnia persisted and finally subsided after the administration of carbamazepine (600 mg/day). Our case supports the finding that carbamazepine might be useful in the treatment of insomnia associated with Whipple encephalopathy.

Cerebral Whipple's disease diagnosed using PCR: the first case reported from Greece.

AbstractBackground: Whipple’s disease (WD) is a rare multisystemic bacterial infection, with variable clinical manifestations occasionally involving the central nervous system. As the cultivation of the etiologic agent, Tropheryma whippelii, is difficult, a laboratory diagnosis is usually based on histological methods. In the last few years, molecular detection of the bacterial 16SrRNA genes by PCR, with 2 primer sets, has greatly contributed to the ability of clinicians to diagnose this disease. We present a cerebral case of WD in a 48-year-old male, successfully diagnosed using PCR with T. whippelii in the blood and feces. As far as we know this is the first case reported from Greece. Methods: For the diagnosis of WD, histological examination of duodenum biopsy for diastase-resistant, non-acid fast, periodic acid Schiff (PAS)-positive inclusions in macrophages, and molecular detection of the 16SrRNA genes of T. whippelii by PCR in cerebrospinal fluid, blood, and feces, were performed. Results: The histological detection was negative. PCR results were positive in the blood and feces of the patient and negative in the cerebrospinal fluid. Seven months after the onset of antimicrobial therapy, PCR was negative in all three clinical specimens. Conclusions: The application of PCR proved to be an invaluable tool for the recognition, differential diagnosis and early initiation of antimicrobial therapy for the patient diagnosed with WD, a disease which is generally fatal if it remains untreated.

Multilocus sequence typing (and phylogenetic analysis) of Campylobacter jejuni and Campylobacter coli strains isolated from clinical cases in Greece

BackgroundThe molecular epidemiology of C. jejuni and C. coli clinical strains isolated from children with gastroenteritis, was investigated using the multilocus sequence typing method (MLST). This analysis establishes for the first time in Greece and constitutes an important tool for the epidemiological surveillance and control of Campylobacter infection in our country.MethodsThe MLST genotypes were compared with those gained by other typing methods (HS-typing, PFGE and FlaA typing) and were also phylogenetically analyzed, in order to uncover genetic relationships.ResultsAmong 68 C. jejuni strains, 41 different MLST-Sequence Types (MLST-STs) were found. Fifty six strains or 34 MLST-STs could be sorted into 15 different MLST-Sequence Type Complexes (MLST-STCs), while twelve strains or seven MLST-STs did not match any of the MLST-STCs of the database. Twenty C. coli strains belonged to 14 different MLST-STs. Eleven MLST-STs were classified in the same MLST-STC (828), and three were unclassifiable. There was no significant association between the MLST-STs and the results of the other typing methods.Phylogenetic analysis revealed that some strains, classified to the species of C. jejuni, formed a separate, phylogenetically distinct group. In eight strains some alleles belonging to the taxonomic cluster of C. jejuni, were also detected in C. coli and vice versa, a phenomenon caused by the genetic mosaic encountered inside the genus Campylobacter.ConclusionsThe MLST-ST determination proved to be a very useful tool for the typing as well as the identification of Campylobacter on the species level.

Detection of Chlamydia Pneumoniae in the Cerebrospinal Fluid of Patients with Multiple Sclerosis by Combination of Cell Culture and PCR

AbstractBackground: During the course of multiple sclerosis (MS) intrathecal oligoclonal IgGs are present in the cerebrospinal fluid (CSF). The intracellular human pathogen Chlamydia pneumoniae may play a role either as a causative pathogenetic agent in the disease, or C. pneumoniaeinfccted MS patients could be immunologically less able to clear the agent from the central nervous system (CNS). Methods: CSF samples were studied in 100 individuals — 70 MS patients and 30 age-matched controls with other neurological diseases. CSF was taken by lumbal puncture; cell cultures were performed by the cell vial technique, followed by a 4-day incubation at 37°C. A nested PCR was performed. Results:C. pneumoniae was detectable in the CSF of only 2.9% of the MS patients and none of control patients (with no significant difference between the MS patients and controls). IgG antibodies were positive in only 1.43% of the MS patients and 3.33% of the controls. IgA antibodies were positive in 6.66% of the control patients and none of the patients were positive for IgM antibodies. There was no statistically significant difference between the two groups of patients with respect to the three antibody classes. Conclusions: The results confirm the high leave of controversy surrounding a possible link between C. pneumoniae and MS, and the matter requires further thorough investigation.

Prevalence of Four Virulence Genes in Campylobacter jejuni Determined by PCR and Sequence Analysis

AbstractIntroduction: The presence of four virulence genes (racR, wlaN, cgtB, virB11) in 356 Campylobacter jejuni strains isolated from confirmed clinical cases was examined by PCR and sequence analysis. The investigated genes were chosen on the basis of their variation in prevalence. Methods: The virulence genes were detected by PCR and the amplified products were submitted for sequence analysis. Results: The gene with the highest prevalence was racR (87.08%). virB was present in only 1.69% of the C. jejuni strains, and wlaN and cgtB were detected in 16.01% and 24.44%, respectively. Five strains associated with Guillain-Barré syndrome and Miller-Fischer syndrome out of the total of 356 (1.40%) were positive for cgtB.Conclusion: Our findings suggest that racR may encode factors necessary for bacterial pathogenicity in humans, while the roles of the other three genes remain ambiguous.

Detection of Chlamydia Pneumoniae in the Cerebrospinal Fluid of Patients with Multiple Sclerosis by Combination of Cell Culture and PCR No Evidence for Possible Association

cerebrospinal fluid (CSF). The intracellular human pathogen Chlamydia pneumoniae may play a role either as a causative pathogenetic agent in the disease, or C. pneumoniae-infected MS patients could be immunologically less able to clear the agent from the central nervous system (CNS). Methods: CSF samples were studied in 100 individuals – 70 MS patients and 30 age-matched controls with other neurological diseases. CSF was taken by lumbal puncture; cell cultures were performed by the cell vial technique, followed by a 4-day incubation at 37°C. A nested PCR was performed. Results: C. pneumoniae was detectable in the CSF of only 2.9% of the MS patients and none of control patients (with no significant difference between the MS patients and controls). IgG antibodies were positive in only 1.43% of the MS patients and 3.33% of the controls. IgA antibodies were positive in 6.66% of the control patients and none of the patients were positive for IgM antibodies. There was no statistically significant difference between the two groups of patients with respect to the three antibody classes. Conclusions: The results confirm the high leave of controversy surrounding a possible link between C. pneumoniae and MS, and the matter requires further thorough investigation.

Heat-stable antigen serotyping of Campylobacter jejuni strains isolated from hospitalized children in Athens, Greece

A hundred and twentynine Campylobacter jejuni strains isolated from hospitalized children with gastroenteritis were serotyped by the heat-stable antigen scheme (HS, Penners method). Isolates belonged to two different periods. Group A contained strains isolated in 1987–1988 and group B contained strains which were isolated in 1998–2000. A variety of serotypes was found. Serotype HS:2 was predominant, followed by the HS:4 complex and HS:1,44. Many clinically important Guillain-Barré Syndrome associated serotypes – like HS:19 – were identified. There were no significant differences in the distribution of serotypes between the two periods. The present report provides reference data, as this is the first C. jejuni serotyping study ever made in Greece.

Serologic markers for HBV, HCV and HIV in immigrants visiting the Athens' polyclinic of 'Doctors of the World - Greece'.

G. Papadakis , N.A. Okoba , C. Nicolaou , F. Boufidou , A. Ioannidis , S. Bersimis , Doctors of the World e Greece, S. Chatzipanagiotou * Greek Delegation of Doctors of the World, Athens, Greece Department of Medical Biopathology, Eginition Hospital, Medical School, University of Athens, Athens, Greece Department of Nursing, Faculty of Human Movement and Quality of Life Sciences, University of Peloponnese, Greece Department of Statistics and Insurance Science, University of Piraeus, Piraeus, Greece