False and mycoplasma-contaminated leukemia-lymphoma cell lines: time for a reappraisal.

Leukemia-lymphoma cell lines are important research tools in a variety of fields. To represent adequate model systems it is of utmost importance that cell lines faithfully model the primary tumor material and are not cross-contaminated with unrelated cell material (or contaminated with mycoplasma). As it has been previously reported that cross-contaminated cell lines represent a significant problem, it is of interest to know whether any improvement in the prevalence of such "false cell lines" had occurred since we called the alert in 1999. A retrospective review of our data archives covered 848 cell lines received from 1990 to 2014 from 290 laboratories in 23 countries spanning the spectrum of leukemia-lymphoma entities. Two variables were considered: authenticity and freedom from mycoplasma infection. Regarding provenance, we separately considered primary sources (original investigators having established the cell lines or reference repositories) and secondary sources. The percentages of mycoplasma-contaminated cell lines decreased significantly over the 25-year timespan. Among primary sourced material: mycoplasma-contamination fell from 23% to 0%; among secondary sourced: from 48% to 21%. The corresponding figures for cross-contamination declined from 15% to 6%, while among material obtained from secondary sources prevalence remained remarkably high, throughout the time periods at 14-18%. Taken together, our data indicate that using non-authenticated cell lines from secondary sources carries a risk of about 1:6 for obtaining a false cell line. The use of authentic leukemia-lymphoma cell lines holds important translational value for their model character and the reproducibility of the laboratory data in the clinical arena.

Shotgun Lipidomic Profiling of the NCI60 Cell Line Panel Using Rapid Evaporative Ionization Mass Spectrometry.

Rapid evaporative ionization mass spectrometry (REIMS) was used for the rapid mass spectrometric profiling of cancer cell lines. Spectral reproducibility was assessed for three different cell lines, and the extent of interclass differences and intraclass variance was found to allow the identification of these cell lines based on the REIMS data. Subsequently, the NCI60 cell line panel was subjected to REIMS analysis, and the resulting data set was investigated for its distinction of individual cell lines and different tissue types of origin. Information content of REIMS spectral profiles of cell lines were found to be similar to those obtained from mammalian tissues although pronounced differences in relative lipid intensity were observed. Ultimately, REIMS was shown to detect changes in lipid content of cell lines due to mycoplasma infection. The data show that REIMS is an attractive means to study cell lines involving minimal sample preparation and analysis times in the range of seconds.

Effects of minimal exposures to atmospheric pressure plasma on the activity of Salmonella Typhimurium: Deactivation of bacterial motility and suppression of host-cell invasion.

Atmospheric pressure plasma (APP) has been shown effective in sterilization by reducing the number of viable microbes during surface cleaning, food processing, or human tissue treatment. For safe conduct, the majority of previous research focused on complete abolition of microbes, which may require severe treatments. Our aim is to investigate the minimal treatment conditions necessary for effective inactivation of bacteria in such a manner that the APP treated bacteria would not be able to harm the host cells. For this, we ought to identify the objective criteria to make the bacteria dysfunctional. We choose the motile properties and the host-cell invasion capability as two measures to quantify the pathogenic state of bacteria. In this paper, we investigated how the APP treatment in a minimal dosage affects the activity of Salmonella Typhimurium. At 100 W and 15 kHz for 20 s, the APP treatment effectively suppressed active "run and tumble" type motility and induced formation of abnormally long structures. With 20 s exposure, the bacterial cells failed to cause pyroptosis in the host cells with >90% survival after 12 h of co-incubation. Our results suggest novel measures to evaluate the functional pathogenic state for identifying safe APP treatment conditions.

Engineered bacteria detect spatial profiles in glucose concentration within solid tumor cell masses.

Tumor heterogeneity makes cancer difficult to treat. Many small molecule cancer drugs target rapidly dividing cells on the periphery of tumors but have difficulty in penetrating deep into tumors and are ineffective at treating entire tumors. Targeting both rapidly dividing and slower growing regions of tumors is essential to effectively treat cancer. A cancer drug carrier that penetrates deep into tumors and identifies metabolically activity could supply treatment to those areas based on the local microenvironment. We hypothesized that glucose sensing bacteria could identify sugar gradients in solid tumors. To test this hypothesis, a genetic circuit was designed to trigger expression of a green fluorescent protein (GFP) reporter through the chemotaxis-osmoporin fusion protein, Trz1, a receptor for sensing glucose and ribose sugars. E. coli equipped with the Trz1-GFP expression system, were administered to an in vitro model of a continuously perfused tumor tissue that mimics systemic delivery and clearance of bacteria through a blood vessel adjacent to a solid tumor. The level of GFP expressed, per bacterium, was time independent and indicated the glucose concentration as a function of penetration depth within the microfluidic tumors. The measured glucose concentration, correlated (P-value = 2.6 × 10(-5) ) with tumor cell viability as a function of depth. Mathematical analysis predicted drug delivery by glucose-sensing bacteria would eliminate a higher percentage of the viable tumor cell population than a systemically administered drug. Glucose-sensing bacteria could deliver cancer therapies with increased drug penetration and nutrient-dependent dosing to continuously treat viable regions of cancer tissue that have a higher prevalence for metastatic dissemination. Biotechnol. Bioeng. 2016;113: 2474-2484. © 2016 Wiley Periodicals, Inc.

Metabolomics reveals mycoplasma contamination interferes with the metabolism of PANC-1 cells.

Mycoplasma contamination is a common problem in cell culture and can alter cellular functions. Since cell metabolism is either directly or indirectly involved in every aspect of cell function, it is important to detect changes to the cellular metabolome after mycoplasma infection. In this study, liquid chromatography mass spectrometry (LC/MS)-based metabolomics was used to investigate the effect of mycoplasma contamination on the cellular metabolism of human pancreatic carcinoma cells (PANC-1). Multivariate analysis demonstrated that mycoplasma contamination induced significant metabolic changes in PANC-1 cells. Twenty-three metabolites were identified and found to be involved in arginine and purine metabolism and energy supply. This study demonstrates that mycoplasma contamination significantly alters cellular metabolite levels, confirming the compelling need for routine checking of cell cultures for mycoplasma contamination, particularly when used for metabolomics studies. Graphical abstract Metabolomics reveals mycoplasma contamination changes the metabolome of PANC-1 cells.

Genome analysis of a Mycoplasma hyorhinis strain derived from a primary human melanoma cell line.

The complete genome of Mycoplasma hyorhinis strain MCLD has been sequenced and annotated. This genome differs by the inversion of a 14.4-kb and a 3.7-kb fragment and the deletion of a 9.9-kb fragment from M. hyorhinis strain HUB-1, isolated from swine respiratory tract. The genome revealed 778 coding sequences (CDSs), with a limited number of vlp genes encoding variable surface lipoproteins.

Effect of Helicobacter bilis infection on human bile duct cancer cells.

BACKGROUND: Helicobacter pylori infection is known to be associated with chronic atrophic gastritis, peptic ulcers, and gastric malignancies. However, the effects of other Helicobacter species have not been investigated extensively. In mice, a close relationship is observed between Helicobacter hepaticus and hepatocellular carcinoma, and Helicobacter species can be found in humans, most commonly in extragastric organs. There have also been reports that H. bilis may be associated with biliary malignancies in humans. The effect of H. bilis infection on a human bile duct cancer cell line was investigated in this study. METHODS: We prepared HuCCT-1, the human bile duct cancer cell line, which was cocultured with H. bilis and cultured alone as a control. HuCCT-1 with and without H. bilis were transfected with the NF-kappaB, E2 transcription factor (E2F), and cyclic AMP response element (CRE) luciferase vectors. The activity of NF-kappaB between H. bilis and the infected and noninfected HuCCT-1 cells was also measured by dual luciferase reporter assay. The concentration of vascular endothelial growth factor (VEGF) in the cocultured medium and control medium were measured by ELISA. To investigate the effect of H. bilis infection on HuCCT-1 with regard to human umbilical vein endothelial cell (HUVEC) tube formation, HUVECs and fibroblasts were cocultured in 24-well plates with and without the conditioned medium. RESULTS: NF-kappaB, E2F and CRE activity, production of VEGF, and angiogenesis in H. bilis-infected cell lines were enhanced compared with controls. CONCLUSIONS: H. bilis infection in a human bile duct cancer cell line activates transcript factors such as NF-kappaB that stimulate production of VEGF and lead to enhancement of angiogenesis. H. bilis infection may play an important role in malignancies in the biliary tract.

[Chinese herbal medicine Jianpi Jiedu Formula down-regulates the expression of vascular endothelial growth factor in human gastric cell line MKN45 induced by Helicobacter pylori by inhibiting cyclooxygenase-2].

OBJECTIVE: To investigate the effects of Jianpi Jiedu Formula (JPJDF), a compound Chinese herbal medicine, on vascular endothelial growth factor (VEGF) expression induced by Helicobacter pylori (Hp) in human gastric cancer cells, and to explore the possible mechanism. METHODS: Human gastric MKN45 cancer cells were infected with standard Hp NCTC11637 by coculture, and the cells were divided into 7 groups: normal control group, Hp group, NS398 inhibitor group, blank serum group, and 5%, 10% and 20% JPJDF groups. The expressions of VEGF and cyclooxygenase-2 (COX-2) mRNAs in human gastric cancer cells infected by Hp were evaluated by real-time fluorogenic quantitative polymerase chain reaction (PCR). After inhibiting the expression of COX-2 with COX-2 specific inhibitor NS398 (50 µmol/L), VEGF mRNA expression induced by Hp in human gastric cancer MKN45 cells was evaluated by real-time fluorogenic quantitative PCR. RESULTS: Real-time fluorogenic quantitative PCR results showed that COX-2 mRNA expression increased significantly after MKN45 cells were infected with Hp for 6 h (P<0.01), and VEGF mRNA expression also increased significantly after MKN45 cells were infected with Hp for 12 h as compared with the normal control group (P<0.01). After inhibiting COX-2 expression with COX-2 specific inhibitor NS398, Hp-induced VEGF mRNA expression significantly reduced (P<0.01). 5%, 10% and 20% JPJDF-containing sera all down-regulated VEGF and COX-2 mRNA expressions induced by Hp in a dose-dependent manner as compared with the Hp infection group. CONCLUSION: Hp could induce the expressions of COX-2 and VEGF in human gastric cancer cells, and JPJDF down-regulates Hp-induced expression of VEGF by inhibiting COX-2, which might be one of the important mechanisms of its prevention and treatment of gastric cancer.

Mycoplasma infection of cultured cells induces oxidative stress and attenuates cellular base excision repair activity.

Mycoplasma contamination is a major concern for in vitro cell culture models as its resistance to most antibiotics, which makes the prevention and treatment of infection challenging. Furthermore, numerous studies show that Mycoplasma infection alters a variety of cellular processes, in a wide range of cell lines. However, there is a lack of information pertaining to the effects of Mycoplasma infection on genomic stability. In this study, a dopaminergic neuronal cell line (BE-M17), a popular in vitro model for Parkinson's disease, was used to evaluate the effect of Mycoplasma infection on genomic instability, and base excision repair (BER) activity, using single cell gel electrophoresis (the comet assay). The results showed that Mycoplasma infection induced oxidative stress in the absence of an inflammatory response, with markedly increased levels of DNA damage [strand breaks/alkali-labile sites (SB/ALS), and oxidised purines], compared to uninfected cells. The source of the oxidative stress may have been increased ROS generation, or attenuation of cellular antioxidant capacity (or a combination of both). Uninfected cells initially repaired SB/ALS more rapidly than infected cells, although SB/ALS were fully repaired in both uninfected and infected cells 2 h after H2O2 challenge. However, while uninfected cells showed complete repair of oxidised purines within 24 h, for the infected cells, these were not fully repaired even after 30 h. In conclusion, this study showed that not only does Mycoplasma infection induce oxidative stress and DNA damage, but it also decreases the efficiency of the main pathway responsible for the repair of oxidatively damaged DNA i.e. BER. In this in vitro model, there is no mechanism for infection-induced inflammation, which could be a source of increased ROS production. Therefore, further studies are needed to evaluate how Mycoplasma infection causes oxidatively damaged DNA, and how it modulates cellular DNA repair.

Prostate-tumor-inducing gene-1 analysis in human prostate cancer cells and tissue in relation to Mycoplasma infection.

The potential role of PTI-1, in the natural story of prostate adenocarcinoma remains to be fully determined. PTI-1 expression was evaluated in human prostate cancer cell lines and in paraffin-embedded archive tissues. PTI-1 expression was found in Mycoplasma infected but not in non-infected cells. The lack of PTI-1 expression was also confirmed in fixed and paraffin-embedded human cancer prostate biopsies. The overall data indicate that, in prostate tumor cell lines, PTI-1 presence parallels Mycoplasma infection suggesting that PTI-1 might not necessarily play a major role in the onset of prostate tumorigenesis.

Biogenesis of Afipia-containing phagosomes in non-professional phagocytes.

Afipia felis is a Gram-negative alpha-proteobacterium, a rare cause of human cat scratch disease (CSD), and likely a pathogen of amoeba. Here, we show that various members of the genus Afipia attach to and are taken up by various non-professional phagocytic mammalian cells (epithelial CHO, endothelial EA.hy926, epithelial HeLa, epithelial INT407 cells, endothelial HMEC-1, endothelial HUVEC, and fibroblast L929 cells). However, only A. felis was able to do this efficiently. Invasion depended on a functional actin cytoskeleton and much less on microtubule dynamics. Bacteria were slowly taken up into HMEC-1 (and HUVEC) via pocket-like structures and they resided within membrane-surrounded phagosomes. While A. felis was found in a non-canonical endocytic compartment in macrophage cells, Afipia-containing phagosomes in HMEC-1 were transiently positive for early endosomal EEA1 and then became and remained positive for lysosome-associated membrane protein-1 (LAMP1) and the proton-pumping ATPase, suggesting undisturbed, albeit slowed, phagosome biogenesis in these cells. Similarly, at 24h of infection, most phagosomes in HeLa, INT407, HUVEC and in EA.hy926 cells were positive for LAMP1. In summary, A. felis enters various non-professional phagocytes and its compartmentation differs between macrophages and non-professional phagocytes.

Secretory IgA and mucin-mediated biofilm formation by environmental strains of Escherichia coli: role of type 1 pili.

Recent studies suggest the importance of secretory IgA (SIgA) and mucin in the mediation of biofilm formation by commensal bacteria within the mammalian gut. Studies using a variety of strains of Escherichia coli have indicated that the interaction between E. coli and SIgA is dependent on the type 1 pilus. In this study, the importance of the pilus in SIgA-mediated biofilm formation by a laboratory strain (MG1655) and environmental (fecal) strains of E. coli was evaluated. Transient expression of the type 1 pilus by the laboratory strain of E. coli failed to facilitate SIgA-mediated biofilm formation, whereas constitutive expression of the type 1 pilus by the laboratory strain was sufficient. In contrast, transient expression of the type 1 pilus was sufficient to facilitate SIgA-mediated biofilm formation by environmental isolates. The "threshold" for mucin-mediated biofilm formation appeared to be lower than that for SIgA-mediated biofilm formation, perhaps reflecting disparate roles of mucin and SIgA in mediating biofilm formation in the gut. These studies also provide the first procedures for the growth of bacterial biofilms on live epithelial cells in vitro, an important development that may facilitate future studies on the effects of bacterial biofilms on epithelial cells.

[Frequency, antimicrobial susceptibility and adherence patterns of Salmonella enterica isolated from chicken meat, beef and pork from Mexico City]. / Frecuencia, susceptibilidad antimicrobiana y patrón de adherencia de Salmonella enterica aislada de carne de pollo, res y cerdo de la Ciudad de México.

BACKGROUND: Food of animal origin is often involved in salmonellosis outbreaks. AIM: To evaluate the frequency of Salmonella enterica in chicken, beef and pork ground meat (a total of 2,592 samples) obtained from travelling markets and supermarkets at the Iztapalapa area of Mexico City, in order to determine the antimicrobial susceptibility and adherence capacity of isolated strains. METHODS: Isolation of S. enterica was carried out according to the BAM-FDA, the microbial susceptibility according with CLSI and adherence assay on HEp-2 cell line according with Baffone et al., 2001. RESULTS: S. enterica was isolated from 511 of all the analyzed samples (19.7%), from which 244 (47.7%), 152 (29.7%) and 115 (22.5%) corresponded to chicken, beef and pork ground meat, respectively. The highest frequency of resistance of S. enterica to antimicrobials was to ampicillin and chloramphenicol in chicken, perfloxacin and ampicillin in beef and carbenicillin, ampicillin, chloramphenicol, cefotaxime and perfloxacin in pork. Ninety percent of the strains showed an aggregative adherence pattern on HEp-2 cells. CONCLUSION: The frequency of S. enterica on meat products is high, which is the reason why a proper cooking of these ground meats is important in order to reduce the risk of acquiring salmonellosis.

[Comparative proteomics research on THP-1 cells infected with Brucella].

Brucella is a facultative intracellular pathogen that survives and multiplies inside host macrophages to cause brucellosis. The response of macrophage plays an essential role in the initiation of immune process following Brucella challenge. Nowadays, proteome approaches have been widely used in many different systems to investigate host-microbe interactions. The effect of pathogen-specific virulence mechanism can now be dissected using bacterial mutants and comparing different species. Attenuated vaccine strain 104M is defective in multification in host macrophage and is cleared relatively rapidly from tissues of the host, whereas virulent strains Brucella abortus 544A can produce chronic infection and cause brucellosis. In order to understand the underlying mechanisms of virulent Brucella intracellular survival, and detect the different-expressed proteins of THP-1 cells after infection with attenuated and virulent strains of Brucella abortus, a comparative proteomics research was conducted. Whole cellular protein profiling of THP-1 cells was presented by two dimensional (2D) electrophoresis and Coomassie Blue staining. After in-gel protein digestion, the different-expressed spots were detected by matrix assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF-MS). All the peptide mass fingerprints (PMFs) were searched by the program Mascot developed by Matrix Science Ltd. For identifying proteins, database of hemo sapiens was used. A total of 38 proteins with changed expression level were found. These proteins can be grouped into two familes: (1) the expression level increased after infection with 544A; (2) the expression level increased after infection with 104M. Out of the 38 proteins, 10 were mainly in the field of signal transduction, 6 were cytoskeletal proteins, 8 were substance metabolism related proteins and 3 were cell stress and defense associated proteins. Functions of the remaining proteins were unknown. These results provide insight into the changed global protein patterns of THP-1 cells after infection as well as a comprehensive foundation to further study of host-bacterial interaction.

C. trachomatis-infection accelerates metabolism of phosphatidylcholine derived from low density lipoprotein but does not affect phosphatidylcholine secretion from hepatocytes.

BACKGROUND: Chlamydia trachomatis is a prevalent sexually transmitted disease and the leading cause of infectious blindness in developing nations. It was not known if C. trachomatis-infection influenced metabolism of lipoprotein-derived phospholipids. Nor was it known if C. trachomatis-infection altered phosphatidylcholine (PC) secretion from hepatocytes. In the current study, low density lipoprotein (LDL)-derived [methyl-3H]PC metabolism was examined in L929 cells infected with C. trachomatis to determine if PC derived from LDL could serve as a potential source of PC trafficked to C. trachomatis. In addition, release of endogenously synthesized [methyl-3H]PC into the medium was examined in rat liver hepatocytes infected with C. trachomatis to determine if infection altered PC secretion. RESULTS: L929 cells 20 h post infection exhibited a 39% (p < 0.05) reduction in radioactivity in PC but total radioactivity incorporation was unaltered compared to controls. Lysophosphatidyl [methyl-3H]choline (LPC) and aqueous [methyl-3H]choline metabolites were elevated 3.6-fold (p < 0.05) and 16.5-fold (p < 0.05), respectively, in C. trachomatis-infected cells and this was due to a 51% increase (p < 0.05) in calcium-dependent phospholipase A2 activity. Hepatocytes 22 h post infection then incubated for 16 h with [methyl-3H]choline showed elevated [methyl-3H]PC biosynthesis but [methyl-3H]PC secreted into the medium was unaltered compared to controls. In contrast, both cellular and medium lyso [methyl-3H]PC were elevated in C. trachomatis-infected cells. CONCLUSION: This study is the first to show that metabolism of LDL-derived PC is accelerated in C. trachomatis infection and to support the notion that LDL-delivered PC may potentially serve as a source of PC trafficked to Chlamydia. In addition, C. trachomatis-infection does not inhibit PC secretion from hepatocytes indicating that the pool of newly synthesized PC destined for lipoprotein secretion may differ from the pool of PC used for C. trachomatis membrane biosynthesis.

Requirement of gene fadD33 for the growth of Mycobacterium tuberculosis in a hepatocyte cell line.

Gene fadD33 of Mycobacterium tuberculosis, one of the 36 homologues of gene fadD of Escherichia coli identified in the M. tuberculosis genome, predictively encodes an acyl-CoA synthase, an enzyme involved in fatty acids metabolism. The gene is underexpressed in the attenuated strain M. tuberculosis H37Ra relative to virulent H37Rv and plays a role in M. tuberculosis virulence in BALB/c mice by supporting mycobacterial replication in the liver. In the present paper, we investigated the role of fadD33 expression in bacterial growth within the hepatocyte cell line HepG2, as well as in human monocyte-derived THP-1 cells and peripheral blood mononuclear cells. M. tuberculosis H37Rv proved able to grow within HepG2 cells, while the intracellular replication of M. tuberculosis H37Ra was markedly impaired; complementation of strain H37Ra with gene fadD33 restored its replication to the levels of H37Rv. Moreover, disruption of gene fadD33 by allelic exchange mutagenesis reduced the intracellular growth of M. tuberculosis H37Rv, and complementation of the fadD33-disrupted mutant with gene fadD33 restored bacterial replication. Conversely, fadD33 expression proved unable to influence M. tuberculosis growth in human phagocytes, as fadD33-disrupted M. tuberculosis H37Rv mutant, as well as fadD33-complemented M. tuberculosis H37Ra, grew within THP-1 cells and peripheral monocytes basically at the same rates as parent H37Rv and H37Ra strains. The results of these experiments indicate that gene fadD33 expression confers growth advantage to M. tuberculosis in immortalized hepatocytes, but not in macrophages, thus emphasizing the importance of fadD33 in liver-specific replication of M. tuberculosis.

Frecuencia, susceptibilidad antimicrobiana y patrón de adherencia de Salmonella enterica aislada de carne de pollo, res y cerdo de la Ciudad de México / Frequency, antimicrobial susceptibility and adherence patterns of Salmonella enterica isolated from chicken meat, beef and pork from Mexico City

Resumen Introducción: Los alimentos de origen animal frecuentemente están implicados en brotes de salmonelosis. Objetivo: Evaluar la frecuencia de Salmonella enterica en carnes molidas de pollo, res y cerdo (un total de 2.592 muestras) obtenidas de mercados sobre ruedas y supermercados de la Delegación Iztapalapa en la Ciudad de México, determinar la susceptibilidad antimicrobiana y efectuar ensayos de adherencia en las cepas aisladas. Métodos: El aislamiento de S. enterica se hizo de acuerdo a la BAM-FDA, la susceptibilidad antimicrobiana de acuerdo con CLSI y el ensayo de adherencia en células HEp-2 conforme a Baffone y cols., 2001. Resultados: Salmonella enterica fue aislada en 511 del total de muestras analizadas (19,7%), de las cuales 244 (47,7%), 152 (29,7%) y 115 (22,5%) correspondieron a carne molida de pollo, res y cerdo, respectivamente. La mayor frecuencia de resistencia de S. enterica a antimicrobianos fue a ampicilina y cloranfenicol en pollo, perfloxacina y ampicilina en res y carbenicilina, ampicilina, cloranfenicol, cefotaxima y perfloxacina en cerdo. Noventa por ciento de las cepas mostraron un patrón de adherencia agregativo. Conclusión: La frecuencia de S. enterica en productos cárnicos es alta, por lo que es importante la adecuada cocción de la carne para disminuir el riesgo de una salmonelosis.

Background: Food of animal origin is often involved in salmonellosis outbreaks. Aim: To evaluate the frequency of Salmonella enterica in chicken, beef and pork ground meat (a total of 2,592 samples) obtained from travelling markets and supermarkets at the Iztapalapa area of Mexico City, in order to determine the antimicrobial susceptibility and adherence capacity of isolated strains. Methods: Isolation of S. enterica was carried out according to the BAM-FDA, the microbial susceptibility according with CLSI and adherence assay on HEp-2 cell line according with Baffone et al., 2001. Results: S. enterica was isolated from 511 of all the analyzed samples (19.7%), from which 244 (47.7%), 152 (29.7%) and 115 (22.5%) corresponded to chicken, beef and pork ground meat, respectively. The highest frequency of resistance of S. enterica to antimicrobials was to ampicillin and chloramphenicol in chicken, perfloxacin and ampicillin in beef and carbenicillin, ampicillin, chloramphenicol, cefotaxime and perfloxacin in pork. Ninety percent of the strains showed an aggregative adherence pattern on HEp-2 cells. Conclusion: The frequency of S. enterica on meat products is high, which is the reason why a proper cooking of these ground meats is important in order to reduce the risk of acquiring salmonellosis.

Characterization and authentication of cancer cell lines: an overview.

Studies of the same cell lines by different laboratories are common in the literature and often show different results with the same methodology. Use of best cell culture practices is essential to ensure consistent and reproducible results. Assay outcomes are easily influenced by many factors including changes in functionality, morphology, doubling time of cells, passage numbers, microbial contamination, and misidentification of cells. Simple observation, monitoring, and documentation of cell morphology and behavior, including growth rates, provide early warning and should be standard practice. Changes may indicate microbial contamination, genotypic drift due to high passage number, or cross-contamination with another cell line. Rapid molecular methods allow the identification of microbial and cross-contamination. Increasingly, authentication of cell lines is a prerequisite for scientific publication to avoid erroneous results entering the literature.

Isolation and culture of colon cancer cells and cell lines.

The preparation of cells from heavily contaminated tissue is challenging. It is usually best to avoid such specimens if possible, but for the study of colorectal and some other tumours, it is inevitable that this must be overcome. The best methods seem to use a combination of (1) debridement of necrotic or infected areas of the tumour, (2) enzymatic dissociation in the presence of antibiotics, (3) density centrifugation to remove debris, and (4) extensive washing of the tumour-derived cells prior to their use in cell culture methods. It is possible to obtain cells from up to 80-90% tumours with careful technique and cooperation from the clinical team.

Isolation and culture of ovarian cancer cells and cell lines.

Ovarian carcinomas show considerable heterogeneity of origin, both in terms of site and tissue. The most important and also most frequent of these tumors arise from the coelomic epithelium and are therefore characterized as epithelial ovarian carcinomas (EOC). EOC is often large and advanced at the time of presentation, so that cells are readily obtainable from surgical specimens or effusions. While the primary tumor may be chemosensitive, they often develop resistance and may do so rapidly. Due to the easy access to tumor cells and its biological behavior, EOC is considered to be an ideal model to investigate principal mechanisms of both antineoplastic drug sensitivity and resistance. Although studies on primary EOC cells are now preferred for many of these investigations, EOC cell line studies remain important too. This chapter gives an overview over major techniques required to establish and maintain primary EOC cell cultures and to initiate and cultivate permanently growing EOC cell lines.