Gr900

Virus-free Mastomys coucha were obtained from Janvier Labs (Le Genest-Saint-Isle, France). At the DKFZ Mastomys coucha were housed under specified pathogen-free (SPF) conditions in individually ventilated cages (Tecniplast GR900) at 22+/-2˚C and 55+/-10% relative humidity in a light/dark cycle of 14/10 h. Mastomys were fed with mouse breeding diet and allowed access to water ad libitum. According to the three R rules of animal experimentation, the animals used in this exploratory study were subgroups (n=6 animals per group; groups: PBS, MnPV L1-VLPs, HPV16 RG1-VLPs, CUT-PANHPVAX) of a larger exploratory study also including testing the protective efficacy of an alternative L1 isoform which is published elsewhere (48 (link)).

In this study, we used male 12-week-old C57BL/6N mice (24–27 g) that were purchased from Charles River Laboratories (Sulzfeld, Germany). Mice were randomized to a Naive, sham-operated group and an fMCAo group (n = 10 in each). Altogether, 30 mice were accommodated in groups of 5–6 in individually ventilated cages (GR900, Tecniplast, Buguggiate, Italy). Environmental enrichment was provided in each cage, namely autoclaved aspen wood chips (1031004, LBS-Biotech, Crawley, UK) in addition to enrichment items including a polycarbonate tunnel (K3487), crawl ball (K3329), aspen blocks (1023005) and aspen wood wool (1034005) obtained from LBS-Biotech (Reigate, UK). Each cage had preset parameters—a temperature of 25 ± 1 °C, humidity between 50 and 60%, and a 12-hour day/night cycle with lights on from 07:00 to 19:00. All mice had free access to filtered tap water and a standard pelleted chow (19.2% protein, 4.1% fat, 6.1% fiber and 5.9% ash) (1324, Altromin, Mucedola, Settimo Milanese, Italy). The experimental design of the study is provided in Figure 1.

Mastomys coucha from the DKFZ colonies were maintained under SFP conditions in individually ventilated cages (Tecniplast GR900) or in type 3 cages in positive pressure isolators on aspen bedding with curled wood wool as environmental enrichment. Mastomys were constantly kept in a light/dark cycle of 14/10h, an average temperature of 22+/-2°C and a humidity of 55+/-10% according to Directive 2010/63/EU, appendix III and the German legislation. Mastomys were fed ad libitum (Mouse and Rat Maintenance No. 3437, KLIBA NAFAG, Kaiseraugst, Switzerland) and had unlimited access to autoclaved water.
Virus-free animals were obtained by hysterectomies of pregnant Mastomys coucha under sterile conditions and kept in a specific pathogen free isolator unit with positive air pressure at the DKFZ [21 (link)]. To confirm the virus-free status of the animals, sera and skin samples are regularly tested by ELISA (E2 and L1) and PCR, respectively.