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Journal of Clinical Pathology Dec 1971Mycoplasmas were sought in the salivary secretions and minor salivary gland tissue of 26 patients with Sjögren's syndrome or the allied sicca complex. A mycoplasma (M....
Mycoplasmas were sought in the salivary secretions and minor salivary gland tissue of 26 patients with Sjögren's syndrome or the allied sicca complex. A mycoplasma (M. orale type 1) was recovered from the stimulated parotid saliva of only one case. Possible mechanisms of mycoplasmal cell damage in this and allied disorders are considered and some future lines of investigation are suggested.
Topics: Adult; Aged; Culture Media; Female; Humans; Male; Middle Aged; Mycoplasma; Parotid Gland; Saliva; Salivary Glands; Sjogren's Syndrome
PubMed: 5139987
DOI: 10.1136/jcp.24.9.810 -
In Vitro Cellular & Developmental... May 1994Mycoplasmal contamination remains a significant impediment to the culture of eukaryotic cells. For certain cultures, attempts to eliminate the infection are feasible...
Mycoplasmal contamination remains a significant impediment to the culture of eukaryotic cells. For certain cultures, attempts to eliminate the infection are feasible alternatives to the normally recommended disposal of the contaminated culture. Here, three antibiotic regimens for mycoplasmal decontamination were compared in a large panel of naturally infected cultures: a 1-wk treatment with the fluoroquinolone mycoplasma removal agent (MRA), a 2-wk treatment with the fluoroquinolone ciprofloxacin, and three rounds of a sequential 1-wk treatment with BM-Cyclin containing tiamulin and minocyclin. These antibiotic treatments had a high efficiency of permanent cure: MRA 69%, ciprofloxacin 75%, BM-Cyclin 87%. Resistance to mycoplasma eradication was observed in some cell cultures: BM-Cyclin 0%, MRA 20%, ciprofloxacin 20%. Nearly all resistant contaminants that could be identified belonged to the species Mycoplasma arginini and M. orale. Detrimental effects of the antibiotics were seen in the form of culture death caused by cytotoxicity (in 5 to 13% of the cultures). Alterations of the cellular phenotypic features or selective clonal outgrowth might represent further untoward side effects of exposure to these antibiotics. Overall, antibiotic decontamination of mycoplasmas is an efficient, inexpensive, reliable, and simple method: 150/200 (75%) chronically and heavily contaminated cultures were cured and 50/200 (25%) cultures could not be cleansed and were either lost or remained infected. It is concluded that eukaryotic cell cultures containing mycoplasmas are amenable to antibiotic treatment and that a cure rate of three-quarters is a reasonable expectation.
Topics: Cells, Cultured; Ciprofloxacin; Diterpenes; Drug Resistance, Microbial; Minocycline; Mycoplasma; Quinolones
PubMed: 8069460
DOI: 10.1007/BF02631456 -
Indian Journal of Medical Microbiology Oct 2007A two-stage nested polymerase chain reaction (PCR) assay system was described that amplifies the 16S-23S rRNA spacer region sequences of Mycoplasma and Acholeplasma...
PURPOSE
A two-stage nested polymerase chain reaction (PCR) assay system was described that amplifies the 16S-23S rRNA spacer region sequences of Mycoplasma and Acholeplasma infections in cell cultures and virus stocks.
METHODS
Established cell lines and virus stocks were screened for the presence of Mycoplasma by using nested PCR using two sets of outer and inner primers, amplifies 16S-23S rRNA. PCR and restriction fragment length polymorphism (RFLP) assay was used to detect and identify most of the species-specific Mycoplasmas involved in cell cultures and virus stock contaminants. Infected cultures detected by PCR-RFLP were further treated with BM-cyclin (5 microg/mL) and passaged for three times and tested for Mycoplasma infections by PCR-RFLP.
RESULTS
Mycoplasma pirum and Mycoplasma orale infections were detected by nested PCR. Species specificity was identified by using RFLP of Vsp I, Cla I and Hin dIII restriction enzymes. Mycoplasma infections were cured by treatment with BM-cyclin. This was further confirmed by non-amplification of PCR amplimers in BM-cyclin treated vs. non-treated cultures.
CONCLUSIONS
Regular monitoring of cell cultures for Mycoplasma infections and identification of species-specific Mollicutes will identify the source of contaminations. This approach can be used for quality control of the biological reagents used in cell culture and virology laboratories.
Topics: Anti-Bacterial Agents; Cell Culture Techniques; DNA, Bacterial; DNA, Ribosomal Spacer; Diterpenes; Minocycline; Mycoplasma; Polymerase Chain Reaction; Polymorphism, Restriction Fragment Length; Quality Control; Virology
PubMed: 18087086
DOI: 10.4103/0255-0857.37340 -
FEMS Microbiology Letters Nov 1992A fast and simple method to detect bacterial and especially mycoplasma contamination in tissue culture by means of polymerase chain reaction (PCR) amplification is...
A fast and simple method to detect bacterial and especially mycoplasma contamination in tissue culture by means of polymerase chain reaction (PCR) amplification is described. In a first step the universal primer pairs P1/P2 (190-bp fragment) and P3/P4 (120-bp fragment) directed to different conserved parts of the prokaryotic 16S rRNA gene are used. A positive signal after amplification on cell culture DNA with these primers provides an indication of bacterial infection. Using the internal primers IP1, IP3 and IP'3 complementary to a part of the V4 and V8 variable regions of the 16S rRNA gene, in combination with a universal primer, cultures contaminated with mycoplasma could be identified. Six mycoplasma species, typical contaminants in tissue cultures, were investigated: Mycoplasma orale, M. fermentans, M. arginini, M. hyorhinis, M. hominis and Aeromonas laidlawii. This mycoplasma test is an easy, specific and sensitive assay which should be extremely useful in any tissue culture setting.
Topics: Bacteria; Base Sequence; Cells, Cultured; DNA, Bacterial; DNA, Ribosomal; Molecular Sequence Data; Mycoplasma; Polymerase Chain Reaction; RNA, Ribosomal
PubMed: 1468621
DOI: 10.1016/0378-1097(92)90293-w -
The American Review of Respiratory... May 1978Mycoplasma strain 215-M, closely related to Mycoplasma orale type 1, was grown in human macrophage monolayers in vitro. Sonicated homogenates of macrophages diluted in...
Mycoplasma strain 215-M, closely related to Mycoplasma orale type 1, was grown in human macrophage monolayers in vitro. Sonicated homogenates of macrophages diluted in saline solution were used for intradermal skin tests in 19 patients with sarcoidosis and in 25 patients with a variety of other diseases. Positive skin-test reactions to this newly developed Mycoplasma test material were observed significantly more often in the patients with sarcoidosis than in the other patients.
Topics: Adult; Aged; Antibodies, Bacterial; Female; Humans; Intradermal Tests; Male; Middle Aged; Mycoplasma; Sarcoidosis; Skin Tests
PubMed: 655497
DOI: 10.1164/arrd.1978.117.5.975 -
Journal of Thoracic Disease Feb 2022The current COVID-19 pandemic is posing a major challenge to public health on a global scale. While it is generally believed that severe COVID-19 results from...
BACKGROUND
The current COVID-19 pandemic is posing a major challenge to public health on a global scale. While it is generally believed that severe COVID-19 results from over-expression of inflammatory mediators (i.e., a "cytokine storm"), it is still unclear whether and how co-infecting pathogens contribute to disease pathogenesis. To address this, we followed the entire course of the disease in cases with severe or critical COVID-19 to determine the presence and abundance of all potential pathogens present-the total "infectome"-and how they interact with the host immune system in the context of severe COVID-19.
METHODS
We examined one severe and three critical cases of COVID-19, as well as a set of healthy controls, with longitudinal samples (throat swab, whole blood, and serum) collected from each case. Total RNA sequencing (meta-transcriptomics) was performed to simultaneously investigate pathogen diversity and abundance, as well as host immune responses, in each sample. A Bio-Plex method was used to measure serum cytokine and chemokine levels.
RESULTS
Eight pathogens, SARS-CoV-2, (), (), (), (), , herpes simplex virus (HSV) and human cytomegalovirus (CMV), identified in patients with COVID-19 appeared at different stages of the disease. The dynamics of inflammatory mediators in serum and the respiratory tract were more strongly associated with the dynamics of the infectome compared with SARS-CoV-2 alone. Correlation analysis revealed that pulmonary injury was directly associated with cytokine levels, which in turn were associated with the proliferation of SARS-CoV-2 and co-infecting pathogens.
CONCLUSIONS
For each patient, the cytokine storm that resulted in acute lung injury and death involved a dynamic and highly complex infectome, of which SARS-CoV-2 was a component. These results indicate the need for a precision medicine approach to investigate both the infection and host response as a standard means of infectious disease characterization.
PubMed: 35280492
DOI: 10.21037/jtd-21-1284 -
Journal of Clinical Pathology Oct 1972Using a modified cell-free culture medium, a mycoplasma was isolated from sarcoid lymph nodes in two cases and from sarcoid skin lesions in four out of seven cases of...
Using a modified cell-free culture medium, a mycoplasma was isolated from sarcoid lymph nodes in two cases and from sarcoid skin lesions in four out of seven cases of chronic sarcoidosis. Growth inhibition tests showed that the isolates were related to Mycoplasma orale type 1. By the indirect haemagglutination method, 244 cases of definite or probable sarcoidosis, 160 patients with other diseases, and 355 blood donors were tested for antibodies against an isolated mycoplasma (strain 215-M). Titres [unk] 16 were found in 14% of the patients with sarcoidosis and in 8% of the patients with other diseases but only in 0.6% of the blood donors. The proportion of patients with high antibody titres among those with sarcoidosis and erythema nodosum was smaller (8%) than among those with other forms of sarcoidosis (17%). The role of the mycoplasmas isolated from sarcoid tissues remains obscure, but it is possible that these organisms are only an expression of altered immunity in sarcoidosis.
Topics: Adult; Antibodies, Bacterial; Biopsy; Blood Donors; Cell-Free System; Erythema Nodosum; Female; Hemagglutination Tests; Humans; Lymph Nodes; Male; Microscopy, Electron, Scanning; Middle Aged; Mycoplasma; Sarcoidosis; Skin
PubMed: 4646295
DOI: 10.1136/jcp.25.10.837 -
Applied and Environmental Microbiology Sep 2008In this study, we describe our results on the evaluation of the ability of different permissive mammalian cell lines to support the biological enrichment of mycoplasma...
In this study, we describe our results on the evaluation of the ability of different permissive mammalian cell lines to support the biological enrichment of mycoplasma species known to be bacterial contaminants of cell substrates. The study showed that this approach is able to significantly improve the efficiency of mycoplasma detection based on nucleic acid testing or biochemical technologies (e.g., MycoAlert mycoplasma detection). Of 10 different cell lines (Vero, MDBK, HEK-293, Hep-G2, CV-1, EBTr, WI-38, R9ab, MDCK, and High Five) used in the study, only MDCK cell culture was found to support the efficient growth of all the tested mycoplasmas (Mycoplasma arginini, M. bovis, M. fermentans, M. gallinaceum, M. gallisepticum, M. synoviae, M. hominis, M. hyorhinis, M. orale, M. salivarium, and Acholeplasma laidlawii) known to be most frequently associated with contamination of cell substrates and cell lines in research laboratories or manufacturing facilities. The infection of MDCK cells with serial dilutions of each mycoplasma species demonstrated that these common cell line contaminants can be detected reliably after 7-day enrichment in MDCK cell culture at contamination levels of 0.05 to 0.25 CFU/ml. The High Five insect cell line was also found to be able to support the efficient growth of most mycoplasma species tested, except for M. hyorhinis strain DBS1050. However, mycoplasma growth in insect cell culture was demonstrated to be temperature dependent, and the most efficient growth was observed when the incubation temperature was increased from 28 degrees C to between 35 and 37 degrees C. We believe that this type of mycoplasma enrichment is one of the most promising approaches for improving the purity and safety testing of cell substrates and other cell-derived biologics and pharmaceuticals.
Topics: Animals; Bacteriological Techniques; Cell Culture Techniques; Cell Line; Chlorocebus aethiops; Coculture Techniques; DNA, Bacterial; Dogs; Insecta; Mycoplasma; Mycoplasma Infections; Polymerase Chain Reaction; Sensitivity and Specificity; Vero Cells
PubMed: 18606798
DOI: 10.1128/AEM.00720-08 -
In Vitro Feb 1979A total of 6432 cell cultures was assayed for mycoplasmas over a 6-year period by aerobic and anaerobic incubation of agar and broth media. Mycoplasmas were detected in...
A total of 6432 cell cultures was assayed for mycoplasmas over a 6-year period by aerobic and anaerobic incubation of agar and broth media. Mycoplasmas were detected in 375 cultures (5.8%). M. orale and A. laidlawii accounted for 61.3% of the isolates. Anaerobic incubation detected 98.1% of the isolates; aeorbic incubation detected 45.8%. Of factors studied to determine their effect on mycoplasma assay, only two, anaerobic incubation and presence of mycoplasmacidal/static antibiotics, were significant. In separate studies, 86 of 2656 cell cultures (3.2%) were infected with strains of M. hyorhinis that did not grow on cell-free media. Recommendations are given for microbiological assay of cell-culture mycoplasmas.
Topics: Aerobiosis; Anaerobiosis; Cells, Cultured; Culture Media; Mycoplasma; Species Specificity
PubMed: 457180
DOI: 10.1007/BF02618100 -
Brazilian Journal of Medical and... Jul 2006A total of 301 cell cultures from 15 laboratories were monitored for mycoplasma (Mollicutes) using PCR and culture methodology. The infection was detected in the cell...
A total of 301 cell cultures from 15 laboratories were monitored for mycoplasma (Mollicutes) using PCR and culture methodology. The infection was detected in the cell culture collection of 12 laboratories. PCR for Mollicutes detected these bacteria in 93 (30.9%) samples. Although the infection was confirmed by culture for 69 (22.9%) samples, PCR with generic primers did not detect the infection in five (5.4%). Mycoplasma species were identified with specific primers in 91 (30.2%) of the 98 samples (32.6%) considered to be infected. Mycoplasma hyorhinis was detected in 63.3% of the infected samples, M. arginini in 59.2%, Acholeplasma laidlawii in 20.4%, M. fermentans in 14.3%, M. orale in 11.2%, and M. salivarium in 8.2%. Sixty (61.2%) samples were co-infected with more than one mycoplasma species. M. hyorhinis and M. arginini were the microorganisms most frequently found in combination, having been detected in 30 (30.6%) samples and other associations including up to four species were detected in 30 other samples. Failure of the treatments used to eliminate mycoplasmas from cell cultures might be explained by the occurrence of these multiple infections. The present results indicate that the sharing of non-certified cells among laboratories may disseminate mycoplasma in cell cultures.
Topics: Base Sequence; Cells, Cultured; DNA, Bacterial; Electrophoresis, Agar Gel; Humans; Molecular Sequence Data; Polymerase Chain Reaction; Tenericutes
PubMed: 16862282
DOI: 10.1590/s0100-879x2006000700009