Establishment of Full-Length cDNA Clones and an Efficient Oral Infection Model for Feline Coronavirus in Cats.

Feline infectious peritonitis virus (FIPV) is the etiologic agent of feline infectious peritonitis (FIP) and causes fatal disease in cats of almost all ages. Currently, there are no clinically approved drugs or effective vaccines for FIP. Furthermore, the pathogenesis of FIP is still not fully understood. There is an urgent need for an effective infection model of feline infectious peritonitis induced by FIPV. Here, we constructed a field type I FIPV full-length cDNA clone, pBAC-QS, corresponding to the isolated FIPV QS. By replacing the FIPV QS spike gene with the commercially available type II FIPV 79-1146 (79-1146_CA) spike gene, we established and rescued a recombinant virus, designated rQS-79. Moreover, we constructed 79-1146_CA infectious full-length cDNA pBAC-79-1146_CA, corresponding to recombinant feline coronavirus (FCoV) 79-1146_CA (r79-1146_CA). In animal experiments with 1- to 2-year-old adult cats orally infected with the recombinant virus, rQS-79 induced typical FIP signs and 100% mortality. In contrast to cats infected with rQS-79, cats infected with 79-1146_CA did not show obvious signs. Furthermore, by rechallenging rQS-79 in surviving cats previously infected with 79-1146_CA, we found that there was no protection against rQS-79 with different titers of neutralizing antibodies. However, high titers of neutralizing antibodies may help prolong the cat survival time. Overall, we report the first reverse genetics of virulent recombinant FCoV (causing 100% mortality in adult cats) and attenuated FCoV (causing no mortality in adult cats), which will be powerful tools to study pathogenesis, antiviral drugs, and vaccines for FCoV. IMPORTANCE Tissue- or cell culture-adapted feline infectious peritonitis virus (FIPV) usually loses pathogenicity. To develop a highly virulent FIPV, we constructed a field isolate type I FIPV full-length clone with the spike gene replaced by the 79-1146 spike gene, corresponding to a virus named rQS-79, which induces high mortality in adult cats. rQS-79 represents the first described reverse genetics system for highly pathogenic FCoV. By further constructing the cell culture-adapted FCoV 79-1146_CA, we obtained infectious clones of virulent and attenuated FCoV. By in vitro and in vivo experiments, we established a model that can serve to study the pathogenic mechanisms of FIPV. Importantly, the wild-type FIPV replicase skeleton of serotype I will greatly facilitate the screening of antiviral drugs, both in vivo and in vitro.

ADE and hyperinflammation in SARS-CoV2 infection- comparison with dengue hemorrhagic fever and feline infectious peritonitis.

The COVID-19 pandemic has rapidly spread around the world with significant morbidity and mortality in a subset of patients including the elderly. The poorer outcomes are associated with 'cytokine storm-like' immune responses, otherwise referred to as 'hyperinflammation'. While most of the infected individuals show minimal or no symptoms and recover spontaneously, a small proportion of the patients exhibit severe symptoms characterized by extreme dyspnea and low tissue oxygen levels, with extensive damage to the lungs referred to as acute respiratory distress symptom (ARDS). The consensus is that the hyperinflammatory response of the host is akin to the cytokine storm observed during sepsis and is the major cause of death. Uncertainties remain on the factors that lead to hyperinflammatory response in some but not all individuals. Hyperinflammation is a common feature in different viral infections such as dengue where existing low-titer antibodies to the virus enhances the infection in immune cells through a process called antibody-dependent enhancement or ADE. ADE has been reported following vaccination or secondary infections with other corona, Ebola and dengue virus. Detailed analysis has shown that antibodies to any viral epitope can induce ADE when present in sub-optimal titers or is of low affinity. In this review we will discuss ADE in the context of dengue and coronavirus infections including Covid-19.

Host Gene Expression of Macrophages in Response to Feline Coronavirus Infection.

Feline coronavirus is a highly contagious virus potentially resulting in feline infectious peritonitis (FIP), while the pathogenesis of FIP remains not well understood, particularly in the events leading to the disease. A predominant theory is that the pathogenic FIPV arises from a mutation, so that it could replicate not only in enterocytes of the intestines but also in monocytes, subsequently systemically transporting the virus. The immune status and genetics of affected cats certainly play an important role in the pathogenesis. Considering the importance of genetics and host immune responses in viral infections, the goal of this study was to elucidate host gene expression in macrophages using RNA sequencing. Macrophages from healthy male cats infected with FIPV 79-1146 ex vivo displayed a differential host gene expression. Despite the virus uptake, aligned viral reads did not increase from 2 to 17 h. The overlap of host gene expression among macrophages from different cats was limited, even though viral transcripts were detected in the cells. Interestingly, some of the downregulated genes in all macrophages were involved in immune signaling, while some upregulated genes common for all cats were found to be inhibiting immune activation. Our results highlight individual host responses playing an important role, consistent with the fact that few cats develop feline infectious peritonitis despite a common presence of enteric FCoV.

Diagnosis of Feline Infectious Peritonitis: A Review of the Current Literature.

Feline infectious peritonitis (FIP) is a fatal disease that poses several challenges for veterinarians: clinical signs and laboratory changes are non-specific, and there are two pathotypes of the etiologic agent feline coronavirus (FCoV), sometimes referred to as feline enteric coronavirus (FECV) and feline infectious peritonitis virus (FIPV) that vary fundamentally in their virulence, but are indistinguishable by a number of diagnostic methods. This review focuses on all important steps every veterinary practitioner has to deal with and new diagnostic tests that can be considered when encountering a cat with suspected FIP with the aim to establish a definitive diagnosis. It gives an overview on all available direct and indirect diagnostic tests and their sensitivity and specificity reported in the literature in different sample material. By providing summarized data for sensitivity and specificity of each diagnostic test and each sample material, which can easily be accessed in tables, this review can help to facilitate the interpretation of different diagnostic tests and raise awareness of their advantages and limitations. Additionally, diagnostic trees depict recommended diagnostic steps that should be performed in cats suspected of having FIP based on their clinical signs or clinicopathologic abnormalities. These steps can easily be followed in clinical practice.

Mucosal Immune Response to Feline Enteric Coronavirus Infection.

Feline infectious peritonitis is a devastating, fatal disease of domestic cats caused by a pathogenic mutant virus derived from the ubiquitous feline enteric coronavirus (FECV). Infection by FECV is generally subclinical, and little is known about the mucosal immune response that controls and eliminates the virus. We investigated the mucosal immune response against FECV in an endemically infected breeding colony over a seven-month period. Thirty-three cats were grouped according to FECV seropositivity and fecal virus shedding into naïve/immunologically quiescent, convalescent and actively infected groups. Blood, fecal samples and colon biopsies were collected to assess the mucosal and systemic immunologic and virologic profile. Results showed that cats with active FECV infections have strong systemic IgG and mucosal IgA responses that wane after virus clearance. Significant FECV-specific mucosal T cell IFNγ responses were not detected in any of the three groups. A shift toward an inflammatory state in the mucosa was suggested by increased IL17:FoxP3 expression. However, no histologic abnormalities were observed, and no shifts in lymphocyte subpopulation phenotype or proliferation were noted. Together, the results suggest that control of FECV is mediated by humoral mucosal and systemic responses and that perturbations in the primary reservoir organ (colon) are minimal.

Characterization of antiviral T cell responses during primary and secondary challenge of laboratory cats with feline infectious peritonitis virus (FIPV).

BACKGROUND: Feline infectious peritonitis (FIP) is considered highly fatal in its naturally occurring form, although up to 36% of cats resist disease after experimental infection, suggesting that cats in nature may also resist development of FIP in the face of infection with FIP virus (FIPV). Previous experimental FIPV infection studies suggested a role for cell-mediated immunity in resistance to development of FIP. This experimental FIPV infection study in specific pathogen free (SPF) kittens describes longitudinal antiviral T cell responses and clinical outcomes ranging from rapid progression, slow progression, and resistance to disease. RESULTS: Differences in disease outcome provided an opportunity to investigate the role of T cell immunity to FIP determined by T cell subset proliferation after stimulation with different viral antigens. Reduced total white blood cell (WBC), lymphocyte and T cell counts in blood were observed during primary acute infection for all experimental groups including cats that survived without clinical FIP. Antiviral T cell responses during early primary infection were also similar between cats that developed FIP and cats remaining healthy. Recovery of antiviral T cell responses during the later phase of acute infection was observed in a subset of cats that survived longer or resisted disease compared to cats showing rapid disease progression. More robust T cell responses at terminal time points were observed in lymph nodes compared to blood in cats that developed FIP. Cats that survived primary infection were challenged a second time to pathogenic FIPV and tested for antiviral T cell responses over a four week period. Nine of ten rechallenged cats did not develop FIP or T cell depletion and all cats demonstrated antiviral T cell responses at multiple time points after rechallenge. CONCLUSIONS: In summary, definitive adaptive T cell responses predictive of disease outcome were not detected during the early phase of primary FIPV infection. However emergence of antiviral T cell responses after a second exposure to FIPV, implicated cellular immunity in the control of FIPV infection and disease progression. Virus host interactions during very early stages of FIPV infection warrant further investigation to elucidate host resistance to FIP.

Differential recognition of peptides within feline coronavirus polyprotein 1 ab by sera from healthy cats and cats with feline infectious peritonitis.

The aim of the study was to identify peptides within the polyprotein (Pp) 1 ab that are differentially recognised by cats with either enteric or systemic disease following infection with feline coronavirus. Overlapping 12-mer peptides (n = 28,426) across the entire Pp1ab were arrayed on peptide chips and reacted with pooled sera from coronavirus seropositive cats and from one seronegative cat. Eleven peptides were further tested in ELISA with individual serum samples, and three were selected for further screening. Two peptides (16433 and 4934) in the nsp3 region encoding the papain 1 and 2 proteases were identified for final testing. Peptide 4934 reacted equally with positive sera from healthy cats and cats with feline infectious peritonitis (FIP), while peptide 16433 was recognized predominantly by FIP-affected cats. The value of antibody tests based on these peptides in differentiating between the enteric and FIP forms of feline coronavirus infection remains to be determined.

Pathogenesis of oral type I feline infectious peritonitis virus (FIPV) infection: Antibody-dependent enhancement infection of cats with type I FIPV via the oral route.

Feline infectious peritonitis virus (FIPV) causes a severe, immune-mediated disease called FIP in domestic and wild cats. It is unclear whether FIP transmits from cat to cat through the oral route of FIPV infection, and the reason for this includes that FIP is caused by oral inoculation with some FIPV strains (e.g., type II FIPV WSU 79-1146), but is not caused by other FIPV (e.g., type I FIPV KU-2 strain: FIPV-I KU-2). In this study, when cats passively immunized with anti-FIPV-I KU-2 antibodies were orally inoculated with FIPV-I KU-2, FIP was caused at a 50% probability, i.e., FIPV not causing FIP through oral infection caused FIP by inducing antibody-dependent enhancement. Many strains of type I FIPV do not cause FIP by inoculation through the oral route in cats. Based on the findings of this study, type I FIPV which orally infected cats may cause FIP depending on the condition.

Inflammatory Mediators in the Mesenteric Lymph Nodes, Site of a Possible Intermediate Phase in the Immune Response to Feline Coronavirus and the Pathogenesis of Feline Infectious Peritonitis?

Feline infectious peritonitis (FIP) is an almost invariably fatal feline coronavirus (FCoV)-induced disease thought to arise from a combination of viral mutations and an overexuberant immune response. Natural initial enteric FCoV infection may remain subclinical, or result in mild enteric signs or the development of FIP; cats may also carry the virus systemically with no adverse effect. This study screened mesenteric lymph nodes (MLNs), the presumed first site of FCoV spread from the intestine regardless of viraemia, for changes in the transcription of a panel of innate immune response mediators in response to systemic FCoV infection and with FIP, aiming to identify key pathways triggered by FCoV. Cats with and without FIP, the latter with and without FCoV infection in the MLN, were compared. Higher expression levels in FIP were found for toll-like receptors (TLRs) 2, 4 and 8. These are part of the first line of defence and suggest a response to both viral structural proteins and viral nucleic acid. Expression of genes encoding inflammatory cytokines and chemokines, including interleukin (IL)-1ß, IL-6, IL-15, tumour necrosis factor (TNF)-α, CXCL10, CCL8, interferon (IFN)-α, IFN-ß and IFN-γ, was higher in cats with FIP, consistent with inflammatory pathway activation. Expression of genes encoding transcription factors STAT1 and 2, regulating signalling pathways, particularly of the interferons, was also higher. Among cats without FIP, there were few differences between virus-positive and virus-negative MLNs; however, TLR9 and STAT2 expression were higher with infection, suggesting a direct viral effect. The study provides evidence for TLR involvement in the response to FCoV. This could open up new avenues for therapeutic approaches.

Discrepancies between feline coronavirus antibody and nucleic acid detection in effusions of cats with suspected feline infectious peritonitis.

Intra-vitam diagnosis of feline infectious peritonitis (FIP) is a challenge for veterinary diagnosticians, since there are no highly specific and sensitive assays currently available. With the aim to contribute to fill this diagnostic gap, a total of 61 effusions from cats with suspected effusive FIP were collected intra-vitam for detection of feline coronavirus (FCoV) antibodies and RNA by means of indirect immunofluorescence (IIF) assay and real-time RT-PCR (qRT-PCR), respectively. In 5 effusions there was no evidence for either FCoV RNA or antibodies, 51 and 52 specimens tested positive by IIF and qRT-PCR, respectively, although antibody titres≥1:1600, which are considered highly suggestive of FIP, were detected only in 37 effusions. Three samples with high antibody levels tested negative by qRT-PCR, whereas 18 qRT-PCR positive effusions contained no or low-titre antibodies. qRT-PCR positive samples with low antibody titres mostly contained low FCoV RNA loads, although the highest antibody titres were detected in effusions with CT values>30. In conclusion, combining the two methods, i.e., antibody and RNA detection would help improving the intra-vitam diagnosis of effusive FIP.

Immunohistochemical characterization of feline lymphoplasmacytic anterior uveitis.

OBJECTIVE: To characterize the immune cells present in different forms of feline anterior uveitis. SAMPLES: Eyes were obtained from 49 cats diagnosed with chronic idiopathic lymphoplasmacytic anterior uveitis, 7 cats with feline infectious peritonitis (FIP), and 9 cats euthanized for nonocular disease. METHODS: H&E sections were scored on the level of infiltrate in the anterior uvea. Immunohistochemistry was performed for FoxP3, CD3, and IL-17A, and positive cells were quantified in multiple images of each sample. A generalized estimating equation tested for an association between the level of inflammation and the prevalence of these cell types. RESULTS: Cells stained positive for IL-17A in idiopathic uveitis but not in FIP samples. We found significantly fewer FoxP3+ and CD3+ cells in low-grade compared with high-grade inflammation in idiopathic uveitis or FIP samples (P values all <.005), but no difference between FIP and high-grade samples. CONCLUSIONS: Idiopathic, but not FIP-associated, uveitis appears to have Th17 cell involvement. The numbers of FoxP3+ and CD3+ T-cells present appear directly correlated; thus, the severity of disease does not appear directly determined by the numbers of regulatory cells.

Diagnosis of non-effusive feline infectious peritonitis by reverse transcriptase quantitative PCR from mesenteric lymph node fine-needle aspirates.

OBJECTIVES: The aim of this study was to evaluate a feline coronavirus (FCoV) reverse transcriptase quantitative PCR (RT-qPCR) on fine-needle aspirates (FNAs) from mesenteric lymph nodes (MLNs) collected in sterile saline for the purpose of diagnosing non-effusive feline infectious peritonitis (FIP) in cats. METHODS: First, the ability of the assay to detect viral RNA in MLN FNA preparations compared with MLN biopsy preparations was assessed in matched samples from eight cats. Second, a panel of MLN FNA samples was collected from a series of cats representing non-effusive FIP cases (n = 20), FCoV-seropositive individuals (n = 8) and FCoV-seronegative individuals (n = 18). Disease status of the animals was determined using a combination of gross pathology, histopathology and/or 'FIP profile', consisting of serology, clinical pathology and clinical signs. RESULTS: Viral RNA was detected in 18/20 non-effusive FIP cases; it was not detected in two cases that presented with neurological FIP. Samples from 18 seronegative non-FIP control cats and 7/8 samples from seropositive non-FIP control cats contained no detectable viral RNA. Thus, as a method for diagnosing non-effusive FIP, MLN FNA RT-qPCR had an overall sensitivity of 90.0% and specificity of 96.1%. CONCLUSIONS AND RELEVANCE: In cases with a high index of suspicion of disease, RT-qPCR targeting FCoV in MLN FNA can provide important information to support the ante-mortem diagnosis of non-effusive FIP. Importantly, viral RNA can be reliably detected in MLN FNA samples in saline submitted via the national mail service. When applied in combination with biochemistry, haematology and serological tests in cases with a high index of suspicion of disease, the results of this assay may be used to support a diagnosis of non-effusive FIP.

Feline coronavirus antibody titer in cerebrospinal fluid from cats with neurological signs.

To investigate the utility of cerebrospinal fluid (CSF) anti-feline coronavirus (FCoV) antibody test for diagnosis of feline infectious peritonitis (FIP), the antibody titers were tested in CSF and sera from 271 FIP-suspected neurological cats. CSF antibody was detected in 28 cats, which were divided into 2 groups; 15 with CSF titer of 1:80 or lower and 13 with CSF titer of 1:640 or higher. In the latter group, reciprocal serum titer/reciprocal CSF titer was 8 or lower, which is extremely lower than normal range (256-2048), and FCoV RNA was detected in all of 11 CSF samples assayed by RT-PCR. Our findings indicate that CSF titer of 1:640 or higher may be served as a candidate for the index for diagnosing FIP.

Nanoparticulate vacuolar ATPase blocker exhibits potent host-targeted antiviral activity against feline coronavirus.

Feline infectious peritonitis (FIP), caused by a mutated feline coronavirus, is one of the most serious and fatal viral diseases in cats. The disease remains incurable, and there is no effective vaccine available. In light of the pathogenic mechanism of feline coronavirus that relies on endosomal acidification for cytoplasmic entry, a novel vacuolar ATPase blocker, diphyllin, and its nanoformulation are herein investigated for their antiviral activity against the type II feline infectious peritonitis virus (FIPV). Experimental results show that diphyllin dose-dependently inhibits endosomal acidification in fcwf-4 cells, alters the cellular susceptibility to FIPV, and inhibits the downstream virus replication. In addition, diphyllin delivered by polymeric nanoparticles consisting of poly(ethylene glycol)-block-poly(lactide-co-glycolide) (PEG-PLGA) further demonstrates an improved safety profile and enhanced inhibitory activity against FIPV. In an in vitro model of antibody-dependent enhancement of FIPV infection, diphyllin nanoparticles showed a prominent antiviral effect against the feline coronavirus. In addition, the diphyllin nanoparticles were well tolerated in mice following high-dose intravenous administration. This study highlights the therapeutic potential of diphyllin and its nanoformulation for the treatment of FIP.

Expression profiles of immune mediators in feline Coronavirus-infected cells and clinical samples of feline Coronavirus-positive cats.

BACKGROUND: There are two biotypes of feline coronavirus (FCoV): the self-limiting feline enteric coronavirus (FECV) and the feline infectious peritonitis virus (FIPV), which causes feline infectious peritonitis (FIP), a fatal disease associated with cats living in multi-cat environments. This study provides an insight on the various immune mediators detected in FCoV-positive cats which may be responsible for the development of FIP. RESULTS: In this study, using real-time PCR and multiplex bead-based immunoassay, the expression profiles of several immune mediators were examined in Crandell-Reese feline kidney (CRFK) cells infected with the feline coronavirus (FCoV) strain FIPV 79-1146 and in samples obtained from FCoV-positive cats. CRFK cells infected with FIPV 79-1146 showed an increase in the expression of interferon-related genes and pro-inflammatory cytokines such as MX1, viperin, CXCL10, CCL8, RANTES, KC, MCP1, and IL8. In addition, an increase in the expression of the above cytokines as well as GM-CSF and IFNγ was also detected in the PBMC, serum, and peritoneal effusions of FCoV-positive cats. Although the expression of MX1 and viperin genes was variable between cats, the expression of these two genes was relatively higher in cats having peritoneal effusion compared to cats without clinically obvious effusion. Higher viral load was also detected in the supernatant of peritoneal effusions compared to in the plasma of FCoV-positive cats. As expected, the secretion of IL1ß, IL6 and TNFα was readily detected in the supernatant of peritoneal effusions of the FCoV-positive cats. CONCLUSIONS: This study has identified various pro-inflammatory cytokines and interferon-related genes such as MX1, viperin, CXCL10, CCL8, RANTES, KC, MCP1, IL8, GM-CSF and IFNγ in FCoV-positive cats. With the exception of MX1 and viperin, no distinct pattern of immune mediators was observed that distinguished between FCoV-positive cats with and without peritoneal effusion. Further studies based on definitive diagnosis of FIP need to be performed to confirm the clinical importance of this study.

Frequency of electrophoretic changes consistent with feline infectious peritonitis in two different time periods (2004-2009 vs 2013-2014).

Objectives The aim of this study was to evaluate whether the frequency of electrophoretic changes in serum of cats with feline infectious peritonitis (FIP) changed in recent years vs past years. Methods Agarose gel electrophoresis (AGE) and capillary zone electrophoresis (CZE) from cats with FIP and healthy cats recorded in the periods 2004-2009 and 2013-2014 were retrospectively analysed. Relative and absolute values of each electrophoretic fraction were recorded and the number of cats showing single or combined electrophoretic changes consistent with FIP (hypoalbuminaemia, inverted albumin to globulin [A:G] ratio, increased total protein, total globulin, alpha [α]2-globulin and gamma [γ]-globulin concentration) were counted. Additionally, a visual analysis of electrophoretograms was also performed. Results for the two time periods were statistically compared. Results The details of 91 AGE procedures (41 from cats with FIP and 50 from healthy cats) and 45 CZE procedures (26 from cats with FIP and 19 from healthy cats) were obtained from the database. No significant differences between the two time periods were found both in FIP and in healthy cats analysed with CZE and in healthy cats analysed with AGE. Compared with 2004-2009, cats with FIP sampled in 2013-2014 with AGE showed a significantly lower concentration of total protein, γ-globulins and total globulins, and a significantly higher A:G ratio and percentage of albumin and α2-globulins. Using both AGE and CZE, in recent years the proportion of cats with high α2-globulins without gammopathy and the proportion of cats with gammopathy alone decreased. With a visual approach, the number of patterns considered as dubious increased in the second period with AGE (non-statistically significant). Conclusions and relevance The frequency of electrophoretic abnormalities in cats with FIP decreased in recent years, independently of the technique employed. Although the mechanism responsible for this change was not investigated in this study, this altered frequency may decrease the diagnostic accuracy of serum protein electrophoresis for FIP.

Upregulation of endothelial cell adhesion molecules characterizes veins close to granulomatous infiltrates in the renal cortex of cats with feline infectious peritonitis and is indirectly triggered by feline infectious peritonitis virus-infected monocytes in vitro.

One of the most characteristic pathological changes in cats that have succumbed to feline infectious peritonitis (FIP) is a multifocal granulomatous phlebitis. Although it is now well established that leukocyte extravasation elicits the inflammation typically associated with FIP lesions, relatively few studies have aimed at elucidating this key pathogenic event. The upregulation of adhesion molecules on the endothelium is a prerequisite for stable leukocyte-endothelial cell (EC) adhesion that necessarily precedes leukocyte diapedesis. Therefore, the present work focused on the expression of the EC adhesion molecules and possible triggers of EC activation during the development of FIP. Immunofluorescence analysis revealed that the endothelial expression of P-selectin, E-selectin, intercellular adhesion molecule 1 (ICAM-1) and vascular cell adhesion molecule 1 (VCAM-1) was elevated in veins close to granulomatous infiltrates in the renal cortex of FIP patients compared to non-infiltrated regions and specimens from healthy cats. Next, we showed that feline venous ECs become activated when exposed to supernatant from feline infectious peritonitis virus (FIPV)-infected monocytes, as indicated by increased adhesion molecule expression. Active viral replication seemed to be required to induce the EC-stimulating activity in monocytes. Finally, adhesion assays revealed an increased adhesion of naive monocytes to ECs treated with supernatant from FIPV-infected monocytes. Taken together, our results strongly indicate that FIPV activates ECs to increase monocyte adhesion by an indirect route, in which proinflammatory factors released from virus-infected monocytes act as key intermediates.

Natural resistance to experimental feline infectious peritonitis virus infection is decreased rather than increased by positive genetic selection.

A previous study demonstrated the existence of a natural resistance to feline infectious peritonitis virus (FIPV) among 36% of randomly bred laboratory cats. A genome wide association study (GWAS) on this population suggested that resistance was polygenic but failed to identify any strong specific associations. In order to enhance the power of GWAS or whole genome sequencing to identify strong genetic associations, a decision was made to positively select for resistance over three generations. The inbreeding experiment began with a genetically related parental (P) population consisting of three toms and four queens identified from among the survivors of the earlier study and belonging to a closely related subgroup (B). The subsequent effects of inbreeding were measured using 42 genome-wide STR markers. P generation cats produced 57 first filial (F1) kittens, only five of which (9.0%) demonstrated a natural resistance to FIPV infection. One of these five F1 survivors was then used to produce six F1/P-backcrosses kittens, only one of which proved resistant to FIP. Six of eight of the F1 and F1/P survivors succumbed to a secondary exposure 4-12 months later. Therefore, survival after both primary and secondary infection was decreased rather than increased by positive selection for resistance. The common genetic factor associated with this diminished resistance was a loss of heterozygosity.

Therapeutic effect of anti-feline TNF-alpha monoclonal antibody for feline infectious peritonitis.

Feline infectious peritonitis virus (FIPV) replication in macrophages/monocytes induced tumor necrosis factor (TNF)-alpha production, and that the TNF-alpha produced was involved in aggravating the pathology of FIP. We previously reported the preparation of a feline TNF-alpha (fTNF-alpha)-neutralizing mouse monoclonal antibody (anti-fTNF-alpha mAb). This anti-fTNF-alpha mAb 2-4 was confirmed to inhibit the following fTNF-alpha-induced conditions in vitro. In the present study, we investigated whether mAb 2-4 improved the FIP symptoms and survival rate of experimentally FIPV-inoculated SPF cats. Progression to FIP was prevented in 2 out of 3 cats treated with mAb 2-4, whereas all 3 cats developed FIP in the placebo control group. Plasma alpha1-glycoprotein and vascular endothelial growth factor levels were improved by the administration of mAb 2-4, and the peripheral lymphocyte count also recovered. These results strongly suggested that the anti-fTNF-alpha antibody is effective for the treatment of FIP.