scholarly journals Plaque assay and cloning of scrub typhus rickettsiae in irradiated L-929 cells

1977 ◽  
Vol 6 (1) ◽  
pp. 76-80
Author(s):  
S C Oaks ◽  
J V Osterman ◽  
F M Hetrick
Keyword(s):  
Scrub Typhus ◽  
Spotted Fever ◽  
Plaque Assay ◽  
Plaque Formation ◽  
Plaque Size ◽  
Representative Species ◽  
Plaque Purification ◽  
Gamma Irradiated ◽  
Intraperitoneal Inoculation

It was demonstrated that gamma-irradiated L-929 cells support plaque formation by three strains of Rickettsia tsutsugamushi and representative species of the spotted fever and typhus group rickettsiae. Sensitivity of the plaque assay for detection of viable scrub typhus rickettsiae was similar to that achieved with intraperitoneal inoculation of random-bred mice. The concentration of irradiated cells and the temperature and length of incubation were all found to affect plaque size. A technique combining terminal dilution and plaque purification was used to obtain clones of three strains of scrub typhus rickettsiae.

Plaque Formation by Chlamydia in L Cells

1970 ◽  
Vol 1 (3) ◽  
pp. 259-262
Author(s):  
Joyce Banks ◽  
B. Eddie ◽  
Julius Schachter ◽  
K. F. Meyer
Keyword(s):  
Cell Lines ◽  
Plaque Assay ◽  
Mouse Fibroblast ◽  
Plaque Formation ◽  
Fibroblast Cells ◽  
Human Origin ◽  
L Cells ◽  
Highly Sensitive ◽  
Slow Growing ◽  
Potential Tool

Chlamydiae were found capable of producing plaques in several cell lines. Mouse fibroblast cells, L-929, proved the most sensitive to infection and yielded plaques of the highest clarity. Assay of chlamydial infectivity by plaque titration was at least as sensitive as egg ld 50 determination. Among chlamydial isolates of avian, mammalian, and human origin, only slow-growing trachoma-inclusion-conjunctivitis agents did not produce plaques. The plaque assay is highly sensitive, reproducible, and offers a potential tool for investigations requiring accurate measurement of small changes in chlamydial infectivity.

scholarly journals A Brief History of the Major Rickettsioses in the Asia–Australia–Pacific Region: A Capstone Review for the Special Issue of TMID

2020 ◽  
Vol 5 (4) ◽  
pp. 165
Author(s):  
Daniel H. Paris ◽  
Daryl J. Kelly ◽  
Paul A. Fuerst ◽  
Nicholas P. J. Day ◽  
Allen L. Richards
Keyword(s):  
Scrub Typhus ◽  
Spotted Fever ◽  
Specific Antigen ◽  
Far East ◽  
Pacific Region ◽  
Special Issue ◽  
Rickettsial Disease ◽  
History Of ◽  
Rickettsial Diseases ◽  
The Impact

The rickettsioses of the “Far East” or Asia–Australia–Pacific region include but are not limited to endemic typhus, scrub typhus, and more recently, tick typhus or spotted fever. These diseases embody the diversity of rickettsial disease worldwide and allow us to interconnect the various contributions to this special issue of Tropical Medicine and Infectious Disease. The impact of rickettsial diseases—particularly of scrub typhus—was substantial during the wars and “police actions” of the last 80 years. However, the post-World War II arrival of effective antibiotics reduced their impact, when recognized and adequately treated (chloramphenicol and tetracyclines). Presently, however, scrub typhus appears to be emerging and spreading into regions not previously reported. Better diagnostics, or higher population mobility, change in antimicrobial policies, even global warming, have been proposed as possible culprits of this phenomenon. Further, sporadic reports of possible antibiotic resistance have received the attention of clinicians and epidemiologists, raising interest in developing and testing novel diagnostics to facilitate medical diagnosis. We present a brief history of rickettsial diseases, their relative importance within the region, focusing on the so-called “tsutsugamushi triangle”, the past and present impact of these diseases within the region, and indicate how historically, these often-confused diseases were ingeniously distinguished from each another. Moreover, we will discuss the importance of DNA-sequencing efforts for Orientia tsutsugamushi, obtained from patient blood, vector chiggers, and rodent reservoirs, particularly for the dominant 56-kD type-specific antigen gene (tsa56), and whole-genome sequences, which are increasing our knowledge of the diversity of this unique agent. We explore and discuss the potential of sequencing and other effective tools to geographically trace rickettsial disease agents, and develop control strategies to better mitigate the rickettsioses.

scholarly journals Gamma-Irradiated Scrub Typhus Immunogens: Development and Duration of Immunity

1978 ◽  
Vol 22 (1) ◽  
pp. 80-86 ◽  
Author(s):  
George H. G. Eisenberg ◽  
Joseph V. Osterman
Keyword(s):  
Scrub Typhus ◽  
Gamma Irradiated

scholarly journals Plaque Assay for Q Fever and Scrub Typhus Rickettsiae

1970 ◽  
Vol 19 (6) ◽  
pp. 963-965
Author(s):  
Joseph E. McDade ◽  
Peter J. Gerone
Keyword(s):  
Scrub Typhus ◽  
Q Fever ◽  
Plaque Assay

Maternal high-fat diet promotes calcified atherosclerotic plaque formation in adult offspring by enhancing transformation of vascular smooth muscle cells to osteochondrocytic-like phenotype

2020 ◽  
Vol 41 (Supplement_2) ◽  
Author(s):  
D Miyawaki ◽  
H Yamada ◽  
H Kubota ◽  
T Sugimoto ◽  
M Saburi ◽  
...  
Keyword(s):  
Smooth Muscle ◽  
Smooth Muscle Cells ◽  
Vascular Smooth Muscle Cells ◽  
High Fat Diet ◽  
Fold Increase ◽  
Plaque Formation ◽  
Adult Offspring ◽  
Plaque Size ◽  
High Fat ◽  
Osteogenic Activity

Abstract Background and objective Maternal high-fat diet (HFD) has been shown to modulate vascular function and remodeling in adult offspring. Atherosclerotic vascular calcification is closely associated with the onset of cardiovascular event. We therefore investigated the impact of maternal HFD on calcification of atherogenic plaques. Methods and results Eight-week-old female apo-E−/− mice (C57BL/6) were fed an HFD or a normal diet (ND) one week prior to mating, and the diet was continued throughout gestation and lactation. Offspring of both groups were fed a high-cholesterol diet (HCD) from 8 weeks of age. Ex vivo osteogenic activity of aortic root and aortic arch was analyzed using in vivo imaging system (IVIS) with OsteoSense 680. Sixteen-week-old male offspring of HFD-fed dams (O-HFD) showed a 1.4-fold increase in fluorescent intensity compared with those of ND-fed dams (O-ND) (p<0.05). Likewise, female O-HFD showed a significantly increased osteogenic activity in aortic arch (154%, p<0.05). Percentages of plaque area and oil red O-positive area were comparable between O-ND and O-HFD of both genders, suggesting that augmented osteogenic activity in O-HFD is not dependent on the plaque size. To investigate the underlying mechanism of augmented calcified plaque formation in O-HFD, vascular smooth muscle cells (VSMCs) of thoracic aorta form 8-week-old male offspring were primarily cultured and VSMCs calcification was induced by treatment with calcification media supplemented with phosphate (2.6 mM). Alizarin-red-positive area upon 10 days stimulation showed a 3.4-fold increase in VSMCs from O-HFD compared with that from O-ND (p<0.01). Consistently, western blotting analysis revealed that expression level of osteocalcin was significantly higher in O-HFD than O-ND, suggesting that osteochondrocytic transformation of VSMCs is augmented in O-HFD. Conclusion Our findings demonstrate that maternal HFD accelerates the development of atherogenic calcification independent of plaque size. In vitro transformation to osteochondrocytic-like cells is enhanced in VSMCs from offspring of HFD-fed dams. Inhibition of VSMCs skewing toward osteochondrocytic-like cells could be a potential therapeutic target for preventing the development of atherosclerotic vascular calcification. Funding Acknowledgement Type of funding source: None

scholarly journals Optimization and Evaluation of a Multiplex Quantitative PCR Assay for Detection of Nucleic Acids in Human Blood Samples from Patients with Spotted Fever Rickettsiosis, Typhus Rickettsiosis, Scrub Typhus, Monocytic Ehrlichiosis, and Granulocytic Anaplasmosis

2020 ◽  
Vol 58 (9) ◽  
Author(s):  
Megan E. Reller ◽  
J. Stephen Dumler
Keyword(s):  
Quantitative Pcr ◽  
Scrub Typhus ◽  
Spotted Fever ◽  
Febrile Illness ◽  
Epidemiologic Studies ◽  
Qpcr Assay ◽  
Spotted Fever Group ◽  
Analytical Sensitivity ◽  
Content Type ◽  
Real Time Quantitative Pcr

ABSTRACT Spotted fever group rickettsioses (SFGR), typhus group rickettsioses (TGR), scrub typhus (caused by Orientia tsutsugamushi), ehrlichiosis, and anaplasmosis often present as undifferentiated fever but are not treated by agents (penicillins and cephalosporins) typically used for acute febrile illness. Inability to diagnose these infections when the patient is acutely ill leads to excess morbidity and mortality. Failure to confirm these infections retrospectively if a convalescent blood sample is not obtained also impairs epidemiologic and clinical research. We designed a multiplex real-time quantitative PCR (qPCR) assay to detect SFGR, TGR, O. tsutsugamushi, and infections caused by Anaplasma phagocytophilum and Ehrlichia chaffeensis with the ompA, 17-kDa surface antigen gene, tsa56, msp2 (p44), and vlpt gene targets, respectively. Analytical sensitivity was ≥2 copies/μl (linear range, 2 to 2 × 105) and specificity was 100%. Clinical sensitivities for SFGR, TGR, and O. tsutsugamushi were 25%, 20%, and 27%, respectively, and specificities were 98%, 99%, and 100%, respectively. Clinical sensitivities for A. phagocytophilum and E. chaffeensis were 93% and 84%, respectively, and specificities were 99% and 98%, respectively. This multiplex qPCR assay could support early clinical diagnosis and treatment, confirm acute infections in the absence of a convalescent-phase serum sample, and provide the high-throughput testing required to support large clinical and epidemiologic studies. Because replication of SFGR and TGR in endothelial cells results in very low bacteremia, optimal sensitivity of qPCR for these rickettsioses will require use of larger volumes of input DNA, which could be achieved by improved extraction of DNA from blood and/or extraction of DNA from a larger initial volume of blood.

scholarly journals Searching and Finding the Hidden Treasure: A Retrospective Analysis of Rickettsial Disease Among Dutch International Travelers

2020 ◽  
Author(s):  
Sophia G de Vries ◽  
Louise E van Eekeren ◽  
Hans van der Linden ◽  
Benjamin J Visser ◽  
Martin P Grobusch ◽  
...  
Keyword(s):  
Retrospective Analysis ◽  
Medical Records ◽  
Scrub Typhus ◽  
Spotted Fever ◽  
Febrile Illness ◽  
Empiric Antibiotic Therapy ◽  
Spotted Fever Group ◽  
Orientia Tsutsugamushi ◽  
Rickettsial Disease ◽  
Empiric Antibiotic

Abstract Background Rickettsial disease (RD) is a prevalent and underestimated cause of febrile illness worldwide, especially in the absence of an inoculation eschar. We attempted to quantify this underestimation at our clinic, by investigating past cases of febrile illness in travelers who had tested negative for leptospirosis, a disease that can initially present similarly to non-eschar RD, and which we routinely consider when other important causes of unspecified febrile illness have tested negative. Methods We performed a retrospective analysis in febrile returned travelers from Asia, Africa, or the Americas between 2010 and 2017, who had tested negative for leptospirosis. Serologic immunofluorescence assays were performed for Orientia tsutsugamushi (scrub typhus), typhus group, and spotted fever group RD. We performed a medical records review of all patients who tested positive. In case of a fitting medical history, cases were deemed either confirmed (based on convalescent serology) or suspected (based on single serology). Results Among 97 patients, convalescent serology was available in 16 (16.5%) patients, and a single serology in 81 (83.5%) patients. RD was the likely diagnosis in 8 of 16 (50.0%) patients with convalescent serology, and in 8 of 81 (9.9%) with single serology. Of the 16 confirmed/suspected cases, 11 (69%) had been missed and 7 (44%) had not received adequate empiric antibiotic therapy. Conclusions This study shows that non-eschar RD is an important and poorly recognized cause of illness in travelers, even in a specialized travel clinic. A lower threshold to test and treat for RD is warranted in returning travelers with febrile illness.

Clinical Features of Rickettsial Infection in Children in Tropical Australia—A Report of 15 Cases

2020 ◽  
Vol 66 (6) ◽  
pp. 655-660 ◽  
Author(s):  
Alexandra G A Stewart ◽  
Simon Smith ◽  
Enzo Binotto ◽  
Josh Hanson
Keyword(s):  
Scrub Typhus ◽  
Spotted Fever ◽  
Case Fatality ◽  
Spotted Fever Group ◽  
Tertiary Referral Hospital ◽  
Admission Rates ◽  
Rickettsial Infections ◽  
Retrospective Audit ◽  
Tropical Australia ◽  
The Tropics

Abstract Rickettsial infections are an under-recognized cause of acute, undifferentiated fever in the tropics. In Asia, intensive care unit (ICU) admission rates as high as 21% and case-fatality rates of up to 5% have been reported. This 20-year retrospective audit of children and adults with serologically confirmed scrub typhus or spotted fever group (SFG) infection was performed at a tertiary-referral hospital in tropical Australia. There were 15 paediatric cases during the study period (11 scrub typhus, 3 SFG and 1 undifferentiated). Hypotension [5/15 (33%)], tachycardia [6/15 (40%)] and tachypnoea [6/15 (40%)] were common at presentation. Children were more likely to be hypotensive at admission than adults [5/15 (33%) vs. 5/118 (4%), p = 0.002]. However, no child died or was admitted to ICU, compared with 18/120 (15%) adults who required ICU support during the study period, one of whom died. Paediatric rickettsial infections have a relatively benign clinical course in tropical Australia with serious complications appearing far less frequently than have been reported in the Asian literature.

PLAQUE FORMATION BY POLIO AND MEASLES VIRUSES IN BS-C-1 (HOPPS) CELLS DERIVED FROM THE AFRICAN GREEN MONKEY

1965 ◽  
Vol 11 (3) ◽  
pp. 435-439 ◽  
Author(s):  
John Furesz ◽  
Pierre Moreau
Keyword(s):  
Cell Line ◽  
Measles Virus ◽  
Plaque Assay ◽  
Plaque Formation ◽  
African Green Monkey ◽  
High Passage ◽  
Green Monkey ◽  
Simple Medium ◽  
Low Passage ◽  
Passage Cells

BS-C-1 cells, cultivated in a simple medium designated M-E, were found to be suitable for a plaque assay of polio and measles viruses. In the plaque assay both low- and high-passage levels of the BS-C-1 cell line were highly susceptible to measles virus but titers of polioviruses were lower in the high-passage cells. Low-passage cells, preserved and stored at −196 °C for prolonged periods, are recommended for the titration of polio and measles viruses.

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