Ultrastructure of Felis catus whole fetus (Fcwf-4) cell culture following infection with feline coronavirus

Objectives Feline infectious peritonitis (FIP) is caused by infection with feline coronavirus (FCoV). FCoV is incredibly contagious and transmission is via the faecal–oral route. FCoV infection, and therefore FIP, is most common in breeder and rescue catteries, where many cats are kept indoors, using litter trays. Whether it is possible to break the cycle of FCoV infection and reinfection using cat litters has never been investigated. The aim of the study was to examine the effect of cat litters on FCoV infectivity and virus load in multi-cat households, and transmission frequency. Methods Fifteen cat litters were mixed and incubated with FCoV, centrifuged and the supernatants tested in vitro for the ability to prevent virus infection of cell culture. To test applicability of in vitro results to real life, virus load was measured in two households in a double crossover study of four Fuller’s earth-based cat litters by testing rectal swabs using FCoV reverse transcriptase quantitative PCR. Results Four litters abrogated FCoV infection of cell culture, nine reduced it to a greater or lesser extent and two had no effect. One brand had different virus inhibitory properties depending on where it was manufactured. Fuller’s earth-based litters performed best, presumably by adsorbing virus. In the field study, there appeared to be less virus shedding on one Fuller’s earth-based cat litter. Conclusions and relevance The in vitro study successfully identified cat litters that inactivate FCoV; such litters exist so do not need to be developed. Fuller’s earth-based litters best prevented infection of cell culture, but did not completely abrogate FCoV transmission in two multi-cat households. A dust-free clumping Fuller’s earth litter appeared to fare best, but virus shedding also varied on the control litters, complicating interpretation. Sawdust-based cat litters are not useful in FCoV-endemic households because they track badly and have a poor effect on virus infection.

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Development of Feline Ileum- and Colon-Derived Organoids and Their Potential Use to Support Feline Coronavirus Infection

Cells ◽

10.3390/cells9092085 ◽

2020 ◽

Vol 9 (9) ◽

pp. 2085 ◽

Cited By ~ 2

Author(s):

Gergely Tekes ◽

Rosina Ehmann ◽

Steeve Boulant ◽

Megan L. Stanifer

Keyword(s):

Cell Culture ◽

Molecular Mechanisms ◽

Domestic Cat ◽

Contributing Factors ◽

Feline Infectious Peritonitis ◽

Culture Models ◽

Coronavirus Infection ◽

Cell Culture Models ◽

Feline Coronavirus ◽

Potential Use

Feline coronaviruses (FCoVs) infect both wild and domestic cat populations world-wide. FCoVs present as two main biotypes: the mild feline enteric coronavirus (FECV) and the fatal feline infectious peritonitis virus (FIPV). FIPV develops through mutations from FECV during a persistence infection. So far, the molecular mechanism of FECV-persistence and contributing factors for FIPV development may not be studied, since field FECV isolates do not grow in available cell culture models. In this work, we aimed at establishing feline ileum and colon organoids that allow the propagation of field FECVs. We have determined the best methods to isolate, culture and passage feline ileum and colon organoids. Importantly, we have demonstrated using GFP-expressing recombinant field FECV that colon organoids are able to support infection of FECV, which were unable to infect traditional feline cell culture models. These organoids in combination with recombinant FECVs can now open the door to unravel the molecular mechanisms by which FECV can persist in the gut for a longer period of time and how transition to FIPV is achieved.

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Type I feline coronavirus spike glycoprotein fails to recognize aminopeptidase N as a functional receptor on feline cell lines

Journal of General Virology ◽

10.1099/vir.0.82666-0 ◽

2007 ◽

Vol 88 (6) ◽

pp. 1753-1760 ◽

Cited By ~ 31

Author(s):

Charlotte Dye ◽

Nigel Temperton ◽

Stuart G. Siddell

Keyword(s):

Cell Culture ◽

Cell Lines ◽

Aminopeptidase N ◽

Surface Glycoprotein ◽

Cellular Receptor ◽

Type I ◽

Type Ii ◽

Virus Surface ◽

Functional Receptor ◽

Feline Coronavirus

There are two types of feline coronaviruses that can be distinguished by serology and sequence analysis. Type I viruses, which are prevalent in the field but are difficult to isolate and propagate in cell culture, and type II viruses, which are less prevalent but replicate well in cell culture. An important determinant of coronavirus infection, in vivo and in cell culture, is the interaction of the virus surface glycoprotein with a cellular receptor. It is generally accepted that feline aminopeptidase N can act as a receptor for the attachment and entry of type II strains, and it has been proposed that the same molecule acts as a receptor for type I viruses. However, the experimental data are inconclusive. The aim of the studies reported here was to provide evidence for or against the involvement of feline aminopeptidase N as a receptor for type I feline coronaviruses. Our approach was to produce retroviral pseudotypes that bear the type I or type II feline coronavirus surface glycoprotein and to screen a range of feline cell lines for the expression of a functional receptor for attachment and entry. Our results show that type I feline coronavirus surface glycoprotein fails to recognize feline aminopeptidase N as a functional receptor on three continuous feline cell lines. This suggests that feline aminopeptidase N is not a receptor for type I feline coronaviruses. Our results also indicate that it should be possible to use retroviral pseudotypes to identify and characterize the cellular receptor for type I feline coronaviruses.

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A T.E.M. study of mouse mammary epithelial cell secretory differentiation cultured on collagen and Matrigel

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s0424820100085241 ◽

1991 ◽

Vol 49 ◽

pp. 188-189

Author(s):

W.N. Bentham ◽

V. Rocha

Keyword(s):

Cell Culture ◽

Mammary Epithelial ◽

Secretory Process ◽

Cell Culture System ◽

Protein Maturation ◽

Cell Culture Techniques ◽

Culture Techniques ◽

Mouse Mammary Epithelial Cell ◽

Secretory Differentiation

It has been an interest of our lab to develop a mammary epethelial cell culture system that faithfully duplicates the in vivo condition of the lactating gland. Since the introduction of collagen as a matrix on which cells are cultivated other E.C.M. type matrices have been made available and are used in many cell culture techniques. We have previously demonstrated that cells cultured on collagen and Matrigel do not differentiate as they do in vivo. It seems that these cultures often produce cells that show a disruption in the secretory process. The appearance of large ribosomal studded vesicles, that specifically label with antibody to casein, suggest an interruption of both protein maturation and secretion at the E.R. to golgi transition. In this report we have examined cultures on collagen and Matrigel at relative high and low seeding densities and compared them to cells from the in vivo condition.

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Solid-phase immunoelectron microscopy for rapid diagnosis of enteroviruses

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s0424820100111240 ◽

1984 ◽

Vol 42 ◽

pp. 226-227 ◽

Cited By ~ 1

Author(s):

K. Pegg-Feige ◽

F. W. Doane

Keyword(s):

Electron Microscopy ◽

Cell Culture ◽

Immunoelectron Microscopy ◽

Detection Method ◽

Solid Phase ◽

Protein A ◽

Plant Viruses ◽

Rapid Diagnosis ◽

Virus Diagnosis ◽

Antiviral Antibody

Immunoelectron microscopy (IEM) applied to rapid virus diagnosis offers a more sensitive detection method than direct electron microscopy (DEM), and can also be used to serotype viruses. One of several IEM techniques is that introduced by Derrick in 1972, in which antiviral antibody is attached to the support film of an EM specimen grid. Originally developed for plant viruses, it has recently been applied to several animal viruses, especially rotaviruses. We have investigated the use of this solid phase IEM technique (SPIEM) in detecting and identifying enteroviruses (in the form of crude cell culture isolates), and have compared it with a modified “SPIEM-SPA” method in which grids are coated with protein A from Staphylococcus aureus prior to exposure to antiserum.

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Automated cell counting of astrocytes on patterned substrates containing aliphatic and charged properties

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s042482010014124x ◽

1995 ◽

Vol 53 ◽

pp. 974-975

Author(s):

W. Shain ◽

H. Ancin ◽

H.C. Craighead ◽

M. Isaacson ◽

L. Kam ◽

...

Keyword(s):

Cell Culture ◽

Cell Attachment ◽

Culture Method ◽

Cell Counting ◽

Data Sets ◽

Nuclear Staining ◽

Double Positive ◽

A Cell ◽

Wafer Test ◽

Cell Densities

Neural protheses have potential to restore nervous system functions lost by trauma or disease. Nanofabrication extends this approach to implants for stimulating and recording from single or small groups of neurons in the spinal cord and brain; however, tissue compatibility is a major limitation to their practical application. We are using a cell culture method for quantitatively measuring cell attachment to surfaces designed for nanofabricated neural prostheses.Silicon wafer test surfaces composed of 50-μm bars separated by aliphatic regions were fabricated using methods similar to a procedure described by Kleinfeld et al. Test surfaces contained either a single or double positive charge/residue. Cyanine dyes (diIC18(3)) stained the background and cell membranes (Fig 1); however, identification of individual cells at higher densities was difficult (Fig 2). Nuclear staining with acriflavine allowed discrimination of individual cells and permitted automated counting of nuclei using 3-D data sets from the confocal microscope (Fig 3). For cell attachment assays, LRM5 5 astroglial cells and astrocytes in primary cell culture were plated at increasing cell densities on test substrates, incubated for 24 hr, fixed, stained, mounted on coverslips, and imaged with a 10x objective.

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