Virus isolation in cell culture for confirmatory diagnostic of rabies in bovine specimens

Kanitz, Fábio Adriano; Cargnelutti, Juliana Felipetto; Weiblen, Rudi; Batista, Helena Beatriz de Carvalho Ruthner; Flores, Eduardo Furtado

doi:10.1590/0103-8478cr20141690

Cited by 4 publications

(4 citation statements)

References 5 publications

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“…and high sensitivity to this virus [20][21][22][23]. The optimal culture conditions found in this study differ slightly from the previous reports in relation to FBS supplementation [8,19,[22][23][24] and cell concentration [19,22,23,25,26], nonetheless were proven to obtain the best viral recovery rates.…”

Section: Plos Neglected Tropical Diseasescontrasting

confidence: 69%

Inactivated rabies vaccines: Standardization of an in vitro assay for residual viable virus detection

et al. 2020

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Human rabies, a neglected viral zoonosis, is preventable through domestic animals vaccination and post-exposure prophylaxis using inactivated rabies vaccines. During vaccine production, several mandatory in vivo quality control trials, such as potency, live virus, and safety, are responsible for the use of large numbers of laboratory animals. Over the years, global organizations encouraged the development of alternative methods to reduce, replace and refine the use of animals in the pharmaceutical industry. In this study we standardized an in vitro assay for determination of residual live virus combining viral isolation techniques with direct immunofluorescence detection and viral quantification by a molecular method. Standardization of viral recovery steps and quantification by RT-qPCR were performed and the combined method was shown to be 3 fold more sensitive than the in vivo assay. It was possible to identify viral suspensions cultures, which still had residual viable rabies virus particles, evidencing the importance to implement this method in quality control schemes of rabies vaccine production. In addition, this developed assay is more practical, inexpensive and less time consuming, producing results in just 4 days, which may allow greater agility in the internal quality control of the vaccine. The in vitro method may reduce 2/3 rd of laboratory animals numbers used for this purpose, since it can be applied in the intermediate quality control of inactivated rabies vaccine production.

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Section: Plos Neglected Tropical Diseasescontrasting

confidence: 69%

Inactivated rabies vaccines: Standardization of an in vitro assay for residual viable virus detection

et al. 2020

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“…We also compared the sensitivity of virus inoculation in cell culture as its use for rabies diagnosis has been reported as a reliable, cost‐effective, ethically preferable alternative to MIT (Fuoco et al., 2018; Kanitz et al., 2015; Madhusudana et al., 2010). RTCIT detected six positive samples (15%) that were negative or inconclusive in DFAT/MIT.…”

Section: Discussionmentioning

confidence: 99%

“…As such, MIT can be replaced by in vitro assays to confirm rabies diagnosis, which must be validated in the laboratory to ensure its accuracy. Studies have shown that inoculation of specific cell lines provides comparable results to MIT (Castilho, 2007;Corona et al, 2018;Kanitz et al, 2015). Such assays, collectively named Rabies Tissue Culture Infection Tests (RTCIT), require cell culture facilities (Corona et al, 2018).…”

Section: Introductionmentioning

confidence: 99%

Laboratory validation of confirmatory tests for rabies diagnosis: Approaches to reduce animal use and facilitate sample collection

Cappelari

Godinho

Silva

et al. 2022

Transbounding Emerging Dis

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Rabies is an encephalitis caused by rabies virus, whose transmission occurs upon contact with infected animals' saliva. The diagnosis is usually performed post-mortem through a direct fluorescent antibody test (DFAT). If the DFAT results are negative, they must be confirmed with an isolation test, usually the mouse inoculation test (MIT), which implies the suffering and death of the animals, high costs and most importantly, up to 28 days to confirm a negative result. Another issue related to rabies diagnosis is the sample collection and storage, which is critical for the rabies virus' RNA genome.Thus, this study aimed to evaluate (i) reverse transcriptase polymerase chain reaction (RT-PCR) and Rabies Tissue Culture Infection Tests (RTCIT) in comparison to DFAT and MIT and (ii) FTA ® cards as an alternative sample collection and preservation method.Eighty animal samples were evaluated through DFAT, RTCIT and RT-PCR; MIT was performed only in DFAT-negative samples. FTA ® cards were evaluated with a subset of 64

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“…In susceptible hosts, RABV is a primary neurotrophic virus that infects neurons in both the central and peripheral nervous systems [ 30 ]. In cell cultures, however, RABV can infect many types of cells, including those of non-neural origins, such as BHK-21 cells [ 1 , 12 , 21 ] and Vero cells [ 33 ]. A local RABV isolated from a rabid dog in Bali also replicated very well in BHK-21 cells.…”

Section: Discussionmentioning

confidence: 99%

Protective antibody response of Balb/c mice to Bali rabies virus isolate propagated in BHK-21 cells

Astawa

Agustini²,

Tenaya³

et al. 2018

The Journal of Veterinary Medical Science

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The protective antibody response of Balb/c mice to Bali rabies virus (RABV) in BHK-21 cells was studied. The virus was isolated from a rabid dog and was adapted to replicate in BHK-21 cell culture for seven passages. The BHK-21-adapted Bali RABV (BHK-Bali RABV) was inactivated with binary ethylenimine and 24 mice were immunized twice at 21-days intervals with the inactivated virus and Rabisin® vaccine. Virus replication was detected using indirect immunofluorescence, immunocytochemistry, and western blotting assays. Enzyme-linked immunosorbent assay examination 2 weeks after the first immunization revealed RABV antibody titers that were mostly below the minimum protective level (<0.5 equivalent unit, EU). Antibody titers increased sharply after the second immunization. Antibody titers in serum of mice induced by inactivated BHK-Bali RABV one week after the second immunization were slightly lower (0.8–3.8 EU) than those induced by Rabisin vaccine (0.9–6.3 EU). RABV antibody titers were stable for at least 6 weeks after the second immunization. Both Rabisin vaccine and inactivated BHK-Bali RABV induced neutralizing antibodies with neutralization titers (50% protective dose per ml) of 29.84 for 0.1 ml Rabisin, 211.41 for 0.2 ml Rabisin, 27.41 for 0.1 ml BHK-Bali RABV, and 28.25 for 0.2 ml BHK-Bali RABV. Thus, inactivated BHK-Bali RABV induces a protective immune response in Balb/c mice, but at lower levels compared to induction by Rabisin vaccine.

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Virus isolation in cell culture for confirmatory diagnostic of rabies in bovine specimens

Cited by 4 publications

References 5 publications

Inactivated rabies vaccines: Standardization of an in vitro assay for residual viable virus detection

Inactivated rabies vaccines: Standardization of an in vitro assay for residual viable virus detection

Laboratory validation of confirmatory tests for rabies diagnosis: Approaches to reduce animal use and facilitate sample collection

Protective antibody response of Balb/c mice to Bali rabies virus isolate propagated in BHK-21 cells

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