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Symptoms and sources of Yersinia enterocolitica -infection: a case-control study. BMC Infectious Diseases 2010, 10 (1) : 122–131.PubMedCrossRef 32. Selleckchem DZNeP Anonymous: Infectious Diseases in Finland 2005. Publications of National Public Health Institute Series B 2006., 17/2006: 33. Capilla S, Ruiz J, Goni P, Castillo J, Rubio MC, Jimenez de Anta MT, Gomez-Lus R, Vila J: Characterization of the molecular mechanisms of quinolone resistance in Yersinia enterocolitica O:3 clinical isolates. J Antimicrob Chemother 2004, 53 (6) : 1068–1071.PubMedCrossRef 34. Partridge SR, Tsafnat G, selleck compound Coiera E, Iredell JR: Gene cassettes and cassette arrays in mobile resistance integrons.

FEMS Microbiol Rev 2009, 33 (4) : 757–784.PubMedCrossRef 35. Karami N, Nowrouzian F, Adlerberth I, Wold AE: Tetracycline resistance in Escherichia coli and persistence in the infantile colonic microbiota. Antimicrob Agents Chemother 2006, 50 (1) : 156–161.PubMedCrossRef 36. Sihvonen LM, Haukka K, Kuusi M, Virtanen MJ, Siitonen A, YE Study Group: Yersinia enterocolitica and Y. enterocolitica -like species in clinical stool specimens of humans: identification and prevalence of bio/serotypes in Finland. Eur J Clin Microbiol Infect Dis 2009, 28 (7)

: 757–765.PubMedCrossRef 37. Lukinmaa S, Nakari UM, Liimatainen A, Siitonen A: Genomic diversity within phage types of Salmonella enterica ssp. enterica serotypes Enteritidis and Typhimurium. Foodborne Pathog Dis 2006, 3 (1) : 97–105.PubMedCrossRef 38. Hunter PR, Gaston MA: Numerical index of the discriminatory ability of typing systems: an application of Simpson’s index of diversity. J Clin Microbiol 1988, 26 (11) : 2465–2466.PubMed 39. CLSI: Performance standards for antimicrobial susceptibility MRIP testing: M100-S16. Clinical and Laboratory Standards Institute; 2006. 40. Cheasty T, Day M, Threlfall E: Increasing incidence of resistance to nalidixic acid in shigellas from humans in England and Wales: implications for therapy. Clinical Microbiology and Infection 2004, 10: 1033–1035.PubMedCrossRef 41. Gripenberg-Lerche C, Zhang L, Ahtonen P, Toivanen P, Skurnik M: Construction of urease-negative mutants of Yersinia enterocolitica serotypes O:3 and O:8: role of urease in virulence and arthritogenicity. Infect Immun 2000, 68 (2) : 942–947.PubMedCrossRef Authors’ contributions LMS participated in the design of the study, did or supervised the MLVA, PFGE, DNA sequencing, and antimicrobial susceptibility testing, carried out the data analysis, and drafted the manuscript. ST performed the conjugation experiment.

For cardiotoxicity of anticancer drugs are typical low levels of cTnT. The majorities of these troponins are bound to actin and are released slowly. This characteristic, along with the slow clearance of troponins from the body permits the detection of

not only acute damage but also of ongoing injury [19]. Baseline plasma hs-cTnT levels were elevated in 5 (13,5%) patients which might be associated with previous anthracycline exposure. Persistent low-level elevations at selleck chemicals least 30 days after HSCT have been observed in our study in almost one third of patients, suggesting prolonged effects of chemotherapy or radiotherapy on the myofibrillar system of cardiomyocytes. Only some authors have demonstrated the role of cardiac

troponins in detection of cardiotoxicity after allogeneic HSCT [13, 20–22]. In fact, several studies have already shown continuous damage of chemotherapy or radiotherapy to the cardiac myofibrillar system [23–25]. In our study, levels of NT-proBNP showed positive correlation with hs-cTnT, which might indicate a relationship Autophagy Compound Library screening between the degree of structural myocyte damage and functional myocardial impairment. These observations were underscored by the echocardiographic studies which did reveal significant changes in systolic and diastolic parameters. Conclusions Persistent elevations in cardiac biomarkers may reflect the presence of an underlying reduced functional myocardial reserve or reduced cardiac tolerance to cardiac stressors. Serial measurements of plasma NT-proBNP Meloxicam and hs-cTnT concentrations might be a useful tool for identification of patients at risk of developing cardiotoxicity after allogeneic HSCT and requiring cardiological follow up. Further studies are needed to clarify whether combination of both biomarkers improve detection of cardiotoxicity. Continued follow up is required to

bring more insight into the predictive value of these biomarkers. Acknowledgements The authors thank Marek Polomsky, M.D. from the University of Rochester, New York, USA for revision of the manuscript. This work was supported by a grant from the Scientific Grant Agency of the Ministry of Health, Slovak Republic 2007/42-UK-18. References 1. Lodi D, Iannitti T, Palmieri B: Stem cells in clinical practice: applications and warnings. J Exp Clin Cancer Res 2011, 30:9.PubMedCrossRef 2. Bhatia S, Francisco L, Carter A, Sun CL, Baker KS, Gurney JG, McGlave PB, Nademanee A, O’Donnell M, Ramsay NK, Robison LL, Snyder D, Stein A, Forman SJ, Weisdorf DJ: Late mortality after hematopietic stem cell transplantation and functional status of long-term survivors: report from the Bone Marrow Transplant Survivor Study. Blood 2007, 110:3784–3791.PubMedCrossRef 3.

Among isolates with complete patterns, 72/162 (44.4%) were clustered. Despite potential fitness costs associated with resistance-conferring mutations [25], the proportion of clustered

Tariquidar datasheet strains was not significantly different among drug-sensitive (60/137, 43.8%) and drug-resistant (12/25, 48.0%) isolates of M. tuberculosis. To distinguish between primary resistance and acquired resistance, clustered isolates sharing identical drug resistance-conferring mutations were considered. Five of the 12 (41.7%) drug-resistant isolates involved in molecular clusters shared their drug resistance-conferring mutations with other isolates in the same cluster, thus strongly suggesting patient-to-patient transmission. Conclusions This study provides so far missing data about drug resistance-conferring mutations in M. tuberculosis isolates

from Madang in PNG. Monitoring drug resistance is essential to prevent the spread of resistant bacteria, especially in diseases requiring lengthy treatments such as TB. Our data suggests that not all present Akt activator drug resistance associated mutations may be detected by molecular tests, which mainly focus on a subset of polymorphisms only. However, given the complex implementation of culture-based DST in resource-constrained settings, PNG may be well suited for an accelerated roll-out of molecular drug resistance testing in order to better tackle the emergence and the transmission of drug-resistant M. tuberculosis strains. Methods Study site and patient characteristics In 2005-2007, a pilot study was conducted in Madang (PNG) at the Modillion Hospital, which is the main point of care in Madang province. In April 2009, a cohort study was initiated in the same hospital and two smaller health centers in close vicinity to Madang town. Patients above 14 years were included if having microscopically confirmed pulmonary TB or other clinical evidence suggesting smear-negative TB. Treatment and

follow-up were planed according to the directly observed treatment, short-course (DOTS) program. Demographic and clinical data were available for all Fossariinae patients, except those recruited during the 2005-2007 pilot study. Sample processing Sputum samples were examined by light microscopy after Ziehl-Neelsen staining. Decontamination was conducted according to Petroff’s method [26]. DST was performed by proportion method [27] at the Queensland Mycobacterial Reference Laboratory in Australia using BACTEC™ MGIT™ 960 (Beckton Dickinson, USA) and the following drug concentrations: RIF (1.0 μg/mL), INH (0.1 and 0.4 μg/mL), Ethambutol (5.0 μg/mL), Pyrazinamide (100 μg/mL), Streptomycin (1.0 μg/mL), Amikacin (1.0 μg/mL), Kanamycin (5.0 μg/mL), Ofloxacin (2.0 μg/mL), Capreomycin (2.5 μg/mL), ETH (5.0 μg/mL), p-Aminosalicylic acid (4.0 μg/mL), and Cycloserine (50.0 μg/mL).

pneumoniae strains that infect

otherwise healthy individuals have emerged from initial endemic foci in Taiwan and China, and are now spreading into North America and Europe [4–6]. This highlights the increasing threat that K. pneumoniae poses to public health and the importance of elucidating its mechanisms of pathogenesis. Most K. pneumoniae strains possess a thick polysaccharide capsule which is involved in protection from opsonisation and phagocytosis and is a well recognized in vivo virulence factor [7]. HDAC activation Various studies have also highlighted roles for surface-exposed lipopolysaccharides, multiple iron acquisition systems and adhesins in K. pneumoniae infection [1, 7, 8]. Several strain-specific virulence determinants of the pyogenic liver abscess-associated

this website isolate K. pneumoniae NTUH-K2044 have been well characterised [9–11]. However, the functions of strain-specific genomic regions in K. pneumoniae strains associated with other types of infection remain poorly studied. Comparative analyses using computational and in vitro experimental techniques have shown that K. pneumoniae strains possess an extremely plastic genome that consists of a conserved core genome interspersed by strain-specific accessory components [12–15]. This was further highlighted in a recent study which calculated that only 54.7% of known K. pneumoniae genes were shared by three sequenced isolates (Kp342, MGH78578, NTUH-K2044) [15]. Genomic islands (GI), typically ranging from 10 kb to 200 kb in size and frequently inserted

within tRNA gene (tRNA) hotspots, comprise a substantial proportion of the accessory genome. GI acquisition offers an efficient ‘quantum leap’ based route to gaining virulence factors, antibiotic resistance determinants and/or metabolic pathways pre-tailored for the exploitation of new environments [16, 17]. Epidemiological studies have suggested that K. pneumoniae infections are preceded by Phosphoglycerate kinase colonization of the gastrointestinal tract [18]. Adhesion and colonization are essential steps in the infection process and are often mediated by fimbriae, which are small hair-like extensions on the bacterial cell surface that can interact with other surfaces via tip-located adhesin proteins [19]. The majority of environmental and clinical K. pneumoniae isolates are known to express type 1 fimbriae and type 3 fimbriae, which have recently been classified into the γ1 and γ4-fimbrial subgroups using the Nuccio and Bäumler fimbrial classification system, which was created from a large scale phylogenetic analysis of fimbrial usher proteins [20–23]. Recent in vivo experiments have demonstrated a role for K. pneumoniae type 1 fimbriae in urinary tract infections [22].

In addition, the women in this case C188-9 chemical structure report presented with current amenorrhea of varying duration, i.e., short-term amenorrhea defined as the cessation of menses for <100 days and long-term amenorrhea defined as the absence of menses for >100 days [13]. Screening procedures Participants signed an informed consent approved by the Institutional Review Board at the University of Toronto or Pennsylvania State University. Height and weight were measured, and participants completed questionnaires to assess medical

history, exercise and menstrual history, eating behaviors, and psychological health. A physical exam and blood sample was performed to determine overall health. A semi-structured psychological interview

was conducted selleck chemicals to ensure that the women were not experiencing major psychiatric disorders, and a registered dietitian assessed eating patterns and food preferences. Dual-energy x-ray absorptiometry (DXA) scans were performed to assess BMD and body composition. Baseline procedures During a 4-week baseline period, menstrual calendars and daily urine samples for the assessment of menstrual function were collected. Body weight was measured weekly. At week 3 of baseline, energetic markers (leptin, ghrelin, total triiodothyronine (TT3)), markers of bone formation and resorption, body composition, resting energy expenditure (REE), and dietary intake were assessed. Participants also completed a test of aerobic fitness. Classification of baseline menstrual status Upon study entry, classification of menstrual status was based on self-reported menstrual history, which was confirmed by a 28-day urinary profile of E1G, PdG, and luteinizing hormone (LH) profiles during a 4-week baseline period. FHA was assessed by confirming a negative pregnancy test, normal endocrine panel, no menses in the past 90 days, and

not documentation of chronically suppressed E1G and PdG profiles observed during the baseline period. Intervention procedures for energy calculations Both participants were asked to increase their caloric intake 20-30% above baseline TEE while maintaining their usual exercise training regimen. For the purpose of this report, baseline TEE was operationally defined as the sum of REE and purposeful EEE. Energy bars that contained approximately 250–300 kilocalories (1,046-1,255 kJ) were provided by the research staff to increase caloric intake. The target increase in caloric intake was gradually achieved by a slow increase in calories during the first several weeks of the intervention to encourage compliance. A registered dietitian met with the participants regularly to provide strategies to meet the target caloric intake.

CB: Professor

at the Department of Genetics Selleck ARN-509 and Biotechnology, Faculty of Agricultural Sciences, University of Aarhus, Denmark. AR: Professor at the Laboratory of Surgical Research, Institute of Clinical Medicine, University of Tromsø, Norway. Acknowledgements The assistance of veterinarians Hege Hasvold and Siri Knudsen, and technicians Ragnhils Osnes and Hege Hagerup is highly acknowledged. Peter Sørensen is acknowledged for his support to the analysis of the microarray data. This study was supported by a grant from the Northern Norway Regional Health Authority (Helse Nord RHF). References 1. Arai M, Yokosuka O, Chiba T, Imazeki F, Kato M, Hashida J, Ueda Y, Sugano S, Hashimoto K, Saisho H, Takiguchi M, Seki N: Gene expression profiling reveals the mechanism and pathophysiology of

mouse liver CRT0066101 supplier regeneration. J Biol Chem 2003, 278:29813–29818.PubMedCrossRef 2. Fukuhara Y, Hirasawa A, Li XK, Kawasaki M, Fujino M, Funeshima N, Katsuma S, Shiojima S, Yamada M, Okuyama T, Suzuki S, Tsujimoto G: Gene expression profile in the regenerating rat liver after partial hepatectomy. J Hepatol 2003, 38:784–792.PubMedCrossRef 3. Locker J, Tian JM, Carver R, Concas D, Cossu C, Ledda-Columbano GM, Columbano A: A common set of immediate-early response genes in liver regeneration and hyperplasia. Hepatology 2003, 38:314–325.PubMedCrossRef 4. Su AI, Guidotti LG, Pezacki JP, Chisari FV, Schultz PG: Gene expression during the priming phase of liver regeneration after partial hepatectomy in mice. Proc Natl Acad Sci USA 2002, 99:11181–11186.PubMedCrossRef 5. White P, Brestelli JE, Kaestner KH, Greenbaum LE: Identification of transcriptional networks during liver regeneration. J Biol Chem 2005, 280:3715–3722.PubMedCrossRef 6. Taub R: Liver regeneration: From myth to mechanism. Nat Rev Mol Cell Biol 2004, 5:836–847.PubMedCrossRef Resveratrol 7. Fujiyoshi M, Ozaki M: Molecular mechanisms of liver regeneration

and protection for treatment of liver dysfunction and diseases. J Hepatobiliary Pancreat Sci 2011, 18:13–22.PubMedCrossRef 8. Koniaris LG, McKillop IH, Schwartz SI, Zimmers TA: Liver regeneration. J Am Coll Surg 2003, 197:634–659.PubMedCrossRef 9. Campbell JS, Prichard L, Schaper F, Schmitz J, Stephenson-Famy A, Rosenfeld ME, Argast GM, Heinrich PC, Fausto N: Expression of suppressors of cytokine signaling during liver regeneration. J Clin Invest 2001, 107:1285–1292.PubMedCrossRef 10. Aldeguer X, Debonera F, Shaked A, Krasinkas AM, Gelman AE, Que XG, Zamir GA, Hiroyasu S, Kovalovich KK, Taub R, Olthoff KM: Interleukin-6 from intrahepatic cells of bone marrow origin is required for normal murine liver regeneration. Hepatology 2002, 35:40–48.PubMedCrossRef 11. Debonera F, Aldeguer X, Shen XD, Gelman AE, Gao F, Que XY, Greenbaum LE, Furth EE, Taub R, Olthoff KM: Activation of interleukin-6/STAT3 and liver regeneration following transplantation. J Surg Res 2001, 96:289–295.

In the Netherlands the creation of sown field margins, known as ‘fauna margins’, is a common form of subsidised AES.

It is assumed that these margins provide habitat for animals in the broad sense, i.e., for birds, small mammals and invertebrates. Due to the manner in which the scheme is regulated, they are commonly installed for a period of 6 years only. As AES may not always be effective in promoting biodiversity (Kleijn et al. 2001, 2006; Kohler et al. 2007; Blomqvist et al. 2009) and often cost a considerable amount of money, it is of great importance to assess the contribution of these margins to biodiversity. Invertebrates, being a species-rich and diverse group of small animals, seem to be especially fit to use as focus group for studying the biodiversity of small landscape elements like fauna margins. The age of such margins might be expected to be a leading factor in invertebrate occurrence, with older margins selleckchem having a greater chance of invertebrate colonisation (Corbet 1995). However, only a limited number of papers have been published on the Cediranib order development of invertebrate communities

in field margins after initial establishment (more papers have been published on plant succession, e.g., Kleijn et al. 1998; Critchley et al. 2006; Manhoudt et al. 2007; Musters et al. 2009). Most of them found in increase with age of the margins (Denys and Tscharntke 2002; Olson and Wäckers 2007; Frank and Reichhart 2004; Woodcock et al. 2008; Musters et al. 2009), although Woodcock et al. (2008) found predatory beetles to peak in the second year after establishment Isotretinoin and to decrease in 2 year thereafter. However, none of these studies deal with a broad range of invertebrate groups and only Musters et al. (2009) and Denys and Tscharntke (2002) discuss patterns over a considerable period of time. To gain more insight into the development of invertebrate groups in field margins, and especially the patterns for distinct functional groups, we performed an inventory on their diversity and abundance in a large

number of these margins in the province of Zeeland, the Netherlands. We formulated two research objectives: (1) How does the number of invertebrate taxa in these strips relate to the age of the margin? (2) How is the abundance of three functional feeding groups—predators, herbivores and detritivores—related to the age of the margin? From the literature cited above, we expected that the field margins would become more species rich with age and that invertebrates would become more abundant. The second question is of major importance, as two of these functional groups may have a direct impact on farming practice: predators that function as enemies of pest organisms and herbivores that might be damaging to crops. It is however possible that the two groups affect each other, resulting in unexpected changes in abundance (Corbet 1995).

Figure 6 Reconstruction of the wound with the free rectus abdominis muscle flap: Line drawing illustrating the free rectus abdominis muscle transfer for thoracotomy wound reconstruction: The right internal mammary artery and vein were anastomosed in an end to end fashion to the right deep inferior epigastric artery and vein, respectively. IMA/V: The check details internal mammary artery and vein, DIEA/V: The deep inferior epigastric artery and

vein, EIA/V: The external iliac artery and vein, R: The rectus abdominis muscle, S: The sternum, F: Fascial closure. Figure 7 The free rectus muscle transferred to the wound: The free rectus abdominis muscle flap transferred to the wound. The right internal mammary vessels extending from the third to fourth intercostal space were prepared for microvascular anastomoses after removal of the third cartilaginous rib. Figure 8 The inset of the rectus muscle: The right chest incision in the recipient site was closed and the free rectus

muscle flap was inset. Figure 9 Postoperative picture: Two months after the reconstruction. Discussion Wound complications associated with emergency thoracotomy have not been reported in the literature. In light of the almost non-existent infection rate, surgical debridement and the reconstruction of EDT wounds is rarely necessitated. The management of the complicated EDT wound was initiated Rolziracetam by adequate surgical debridement and appropriate antibiotic treatment prior to definitive reconstruction. In addition, coverage especially with a muscle flap was planned to overcome Ipatasertib nmr the infection and to supplement the healing in such a wound with exposed heart. The pectoralis major, the latissimus dorsi, the rectus abdominis,

and omental flap are most frequently employed flaps in the chest and sternal region wound reconstruction [3, 4]. However, in our case, reconstruction of the thoracotomy wound presented several reconstructive challenges. The pectoralis major or latissimus dorsi muscle flaps were not suitable with regards to the location of the EDT wound. The omental flap was not employed to avoid laparotomy and associated risks. On the other hand, the rectus abdominis muscle could not be utilized since the superior epigastric vessels, the pedicle of a superiorly based flap, were found to be unreliable. The superior epigastric artery originates from the internal mammary artery at the level of the seventh rib. Then, it descends between the costal and xiphoid slips of the diaphragm, anterior to the lower fibers of the transversus thoracis and transversus abdominis. Entering the rectus sheath, at first behind the rectus abdominis muscle and then perforating and supplying it, it anastomoses with the deep inferior epigastric branch of the external iliac [5] (Figure 4).

The mecA gene, the structural determinant that encodes PBP2a, is therefore considered as a useful molecular marker of putative methicillin resistance in S. aureus and CoNS [9, 10]. Clinical laboratory tests

for methicillin resistance are highly dependent on growing conditions such as temperature, pH and salt concentration [11]. Thus, these factors emphasize the need to develop a rapid, accurate and sensitive method for detection of methicillin-resistant staphylococci, which does not depend on growth conditions. Nucleic-acid-based tests using PCR are increasingly being used in laboratories to replace time-consuming, labor intensive and less sensitive conventional diagnostic methods, such as biochemical identification and Kirby-Bauer antimicrobial susceptibility tests. Various PCR methods have been developed to identify: (i) Staphylococcus genus [12]; (ii) methicillin-resistance selleck screening library [13]; and (iii) Panton-Valentine leukocidin (PVL)-producing Staphylococcus genus [14]. These methods do not detect all of the above-mentioned targets simultaneously. Hence, the present study focused on the design of a pentaplex PCR for methicillin-resistant staphylococci with an internal control for the detection of Staphylococcus genus (16S rRNA gene), methicillin-resistant staphylococci (mecA gene), community-acquired

MRSA (lukS gene), and discrimination between S. aureus and CoNS BCKDHB (femA gene). Results In the present study, the pentaplex PCR was optimized successfully to identify the Staphylococcus genus (16S check details rRNA), S. aureus species (femA), methicillin resistance (mecA) and PVL toxin (lukS) genes simultaneously. Stepwise optimization of primer concentration, annealing temperature, MgCl2, dNTP and Taq polymerase was carried out. The pentaplex PCR gave the best results

when 3.13 mM MgCl2, 200 μM dNTP, 0.75 U Taq polymerase and 60°C annealing temperature were used. The analytical sensitivity of the pentaplex PCR at the DNA level was found to be 10 ng DNA (data not shown), whereas, at the bacterial level, it was found to be 104 CFU/mL (data not shown). The analytical specificity of the pentaplex PCR assay at the genus level was determined using 10 staphylococcal reference strains and found to be positive for the Staphylococcus genus specific 16S rRNA gene. A representative gel picture of methicillin resistance with reference strains is shown in Figure 1, while the other 10 Gram-positive non-staphylococcal and 13 Gram-negative strains were negative. All the reference strains of S. aureus were positive for femA gene by pentaplex PCR, while other CoNS species were negative (Table 1). Hence, all methicillin-resistant reference strains were positive for mecA gene by pentaplex PCR. However, the methicillin-sensitive reference strains were negative for mecA gene by pentaplex PCR (Table 1).

Horizontal reading of the graph indicates the percentage of unigenes shared by several libraries. D. GO annotation results for this website High Scoring Pairs (HSP) coverage of 0%. GO annotation was first conducted using the Score Function (SF) of the BLAST2GO software. The GO terms selected by the annotation step were then merged with InterProScan predictions (SF + IPR). Finally, the Annex annotation was run (SF + IPR + ANNEX). E. Annotation distribution of GO terms. Two

non-normalized libraries were constructed from asymbiotic and symbiotic ovaries (AO and SO) starting with 1 µg of polyA RNAs. They were prepared using Creator SMART cDNA Library Construction kit (Clontech/BD Biosciences), following the manufacturer’s instructions. cDNA was digested by SfiI, purified (BD Chroma Spin – 400 column) and ligated into pDNRlib vector for Escherichia coli transformation. Amplified double strand cDNA (ds cDNA) was prepared using a SMART approach [28]. SMART Oligo II oligonucleotide (Clontech/BD Biosciences) and CDS primer were used for first-strand cDNA synthesis. SMART-amplified cDNA samples were further digested by RsaI endonuclease. The SSH libraries from asymbiotic and symbiotic ovaries (SSH-A and SSH-S) were constructed

starting with 20 µg of total RNA. SSH libraries from specimens challenged and not challenged by S. typhimurium (SSH-C and SSH-NC) were performed on 20.4 µg of a total RNA equally pooled from different tissues (i.e., ovaries, gut, cæca, fat tissues, hemocytes, hematopoietic organ, nerve chain, and brain) harvested at each PI3K Inhibitor Library molecular weight time point. The pooled total RNA was obtained by mixing equal amounts of total RNA

extracted separately for each tissue and for each time point. Subtractive hybridizations were performed BCKDHB using SSH method in both directions (Asymbiotic vs. Symbiotic A/S and vice-versa S/A; Not Challenged vs. Challenged NC/C and vice-versa C/NC) as described in [29, 30] using the PCR-Select cDNA Subtraction Kit (Clontech/BD Biosciences). SSH libraries were prepared by Evrogen (Moscow, Russia). The Mirror Orientation Selection (MOS) procedure was used for SSH-A/S and SSH-C/NC as described in [31] in order to reduce the number of false-positive clones in the SSH-generated libraries. Purified cDNAs from SSH-A/S and SSH-C/NC were cloned into the pAL16 vector (Evrogen) and used for E. coli transformation. Finally, the normalized library (N) was prepared with 75 µg of a pooled total RNA from an equimolar proportion of asymbiotic and symbiotic ovaries, and 6h, 9h, and 15h challenged asymbiotic females. As for the libraries of challenged specimens, total RNA was extracted separately from the same tissues. This N library was prepared by Evrogen (Moscow, Russia). Total RNA sample was used for ds cDNA synthesis using SMART approach [28]. SMART prepared amplified cDNA was then normalized using Duplex Specific Nuclease (DSN) normalization method [32].