Evidence and antibiotic resistance profiles of clinical Acinetobacter calcoaceticus-Acinetobacter baumannii (ACB) and non-ACB complex members in companion animals: A 2020-2022 retrospective study.

To evaluate the frequency of Acinetobacter spp., belonging to both Acinetobacter calcoaceticus-baumannii (ACB) and non-ACB complex, and their antibiotic resistance profiles in veterinary medicine, a three-year (2020-2022) retrospective study was carried out on sick companion animals. Epidemiological data from different clinical canine, feline, and equine samples, were acquired. For each strain, MALDI-TOF MS identification and susceptibility to a panel of 11 antibiotics, by Kirby-Bauer and E-test methods, were performed. Out of 628 bacteriological examinations, 2.5% resulted positive for strains belonging to Acinetobacter genus. Frequencies of 2.3%, 1.9%, and 3% were obtained from both in-visiting and hospitalized dogs, cats, and horses, respectively. Members of ACB-complex accounted for 50% of isolates. Since all strains resulted susceptible to aminoglycosides and polymyxins, no pandrug-resistant (PDR) species were recorded. While 12.5% A. baumannii resulted extensively-drug resistant (XDR), a higher percentage of multidrug-resistant strains was recorded among non-ACB strains (35.5%) than ACB strains (25%). Susceptibility was observed in the same percentage in both groups (62.5%). All ACB strains confirmed their intrinsic resistances. Non-ACB species showed lower resistances against antipseudomonal penicillins plus beta-lactamase inhibitors (P=0.1306), III generation cephalosporins (P=0.0547), and tetracyclines (P=0.0209) than ACB species. Carbapenem-resistance was observed for XDR A. baumannii (12.5%) and, in particular for MDR non-ACB complex members (25%). To our knowledge, A. lactucae represents the first description in two sick dogs in Italy. Furthermore, our results emphasize the role of non-ACB-complex species as important zoonotic pathogens, which could be reservoirs of clinically relevant resistance profiles.

Fungal metabolite 6-pentyl-α-pyrone reduces canine coronavirus infection.

Canine coronavirus (CCoV) can produce a self-limited enteric disease in dogs but, because of notable biological plasticity of coronaviruses (CoVs), numerous mutations as well as recombination events happen leading to the emergence of variants often more dangerous for both animals and humans. Indeed, the emergence of new canine-feline recombinant alphacoronaviruses, recently isolated from humans, highlight the cross-species transmission potential of CoVs. Consequently, new effective antiviral agents are required to treat CoV infections. Among the candidates for the development of drugs against CoVs infection, fungal secondary metabolites (SMs) represent an important source to investigate. Herein, antiviral ability of 6-pentyl-α-pyrone (6 PP), a SM obtained by Trichoderma atroviride, was assessed against CCoV. During in vitro infection, nontoxic concentration of 6 PP significantly increased cell viability, reduced morphological signs of cell death, and inhibited viral replication of CCoV. In addition, we found a noticeable lessening in the expression of aryl hydrocarbon receptor (AhR), a strategic modulator of CoVs infection. Overall, due to the variety of their chemical and biological properties, fungal SMs can decrease the replication of CoVs, thus identifying a suitable in vitro model to screen for potential drugs against CoVs, using a reference strain of CCoV (S/378), non-pathogenic for humans.

Antiviral activity of Taurisolo® during bovine alphaherpesvirus 1 infection.

Bovine alphaherpesvirus 1 (BoAHV-1), the pathogen causing Infectious Bovine Rhinotracheitis (IBR) and predisposing to polymicrobial infections in cattle, provokes farm economic losses and trading restrictions in the world. However, nontoxic antiviral agents for BoAHV-1 infection are still unavailable, but plant extracts, such as flavonoid derivatives possess activity against BoAHV-1. Taurisolo®, a nutraceutical produced by Aglianico grape pomace, has recently shown promising antiviral activity. Herein, the potential activity of Taurisolo® during BoAHV-1 infection in Madin Darby bovine kidney (MDBK) cells was tested. Taurisolo® enhanced cell viability and reduced morphological death signs in BoAHV-1-infected cells. Moreover, Taurisolo® influenced the expression of bICP0, the key regulatory protein of BoAHV-1, and it strongly diminished virus yield. These effects were associated with an up-regulation of aryl hydrocarbon receptor (AhR), a transcription factor involved in microbial metabolism and immune response. In conclusion, our findings indicate that Taurisolo® may represent a potential antiviral agent against BoAHV-1 infection. Noteworthy, AhR could be involved in the observed effects and become a new target in antiviral therapy.

In Vitro Evaluation of Antiviral Activities of Funicone-like Compounds Vermistatin and Penisimplicissin against Canine Coronavirus Infection.

Recent studies have demonstrated that 3-O-methylfunicone (OMF), a fungal secondary metabolite from Talaromyces pinophilus belonging to the class of funicone-like compounds, has antiviral activity against canine coronaviruses (CCoV), which causes enteritis in dogs. Herein, we selected two additional funicone-like compounds named vermistatin (VER) and penisimplicissin (PS) and investigated their inhibitory activity towards CCoV infection. Thus, both compounds have been tested for their cytotoxicity and for antiviral activity against CCoV in A72 cells, a fibrosarcoma cell line suitable for investigating CCoV. Our findings showed an increase in cell viability, with an improvement of morphological features in CCoV-infected cells at the non-toxic doses of 1 µM for VER and 0.5 µM for PS. In addition, we observed that these compounds caused a strong inhibition in the expression of the aryl hydrocarbon receptor (AhR), a ligand-activated transcription factor which is activated during CCoV infection. Our results also showed the alkalinization of lysosomes in the presence of VER or PS, which may be involved in the observed antiviral activities.

Effectiveness of the Fungal Metabolite 3-O-Methylfunicone towards Canine Coronavirus in a Canine Fibrosarcoma Cell Line (A72).

Canine coronavirus (CCoV), an alphacoronavirus, may cause self-limiting enteric disease in dogs, especially in puppies. The noteworthy plasticity of coronaviruses (CoVs) occurs through mutation and recombination processes, which sometimes generate new dangerous variants. The ongoing SARS-CoV-2 pandemic and the isolation of a novel canine-feline recombinant alphacoronavirus from humans emphasizes the cross-species transmission ability of CoVs. In this context, exploring antiviral compounds is essential to find new tools for fighting against CoVs infections. Fungi produce secondary metabolites, which are often developed as antibiotics, fungicides, hormones, and plant growth regulators. Previous examinations of benzo-γ-pyrone 3-O-methylfunicone (OMF), obtained from Talaromyces pinophilus, showed that it reduces the infectivity of hepatitis C virus and bovine herpesvirus 1. Based on this evidence, this study evaluated the antiviral ability of OMF against CCoV infection in a canine fibrosarcoma (A72) cell line. During CCoV infection, a non-toxic dose of OMF markedly increased features of cell viability. Moreover, OMF induced a significant reduction in virus yield in the presence of an intense downregulation of the viral nucleocapsid protein (NP). These findings occurred in the presence of a marked reduction in the aryl hydrocarbon receptor (AhR) expression. Taken together, preliminary findings suggest that OMF inhibiting AhR shows promising activity against CCoV infection.

Canine Coronavirus Activates Aryl Hydrocarbon Receptor during In Vitro Infection.

The aryl hydrocarbon receptor (AhR) is a ligand-activated transcription factor that interacts with substrates, including microbial metabolites. Recent advances reveal that AhR is involved in the host response to coronaviruses (CoVs) infection. Particularly, AhR antagonists decrease the expression of angiotensin-converting enzyme 2 (ACE2) via AhR up-regulation, resulting in suppression of severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) infection in mammalian cells. Herein, we report that AhR is expressed in canine fibrosarcoma (A72) cells, where it is considerably activated by infection with genotype II of canine coronavirus (CCoV-II). The pharmacological inhibition of AhR, by CH223191, suppressed cell death signs and increased cell viability. Furthermore, the AhR antagonist induced a meaningful decline in virus yield, accompanied by the inhibition of the expression of viral nuclear protein (NP). Fascinatingly, during CCoV infection, a novel co-expression of NP and AhR expression was found. Taken together, our preliminary findings show that infection with CCoV activates AhR, and pharmacologic AhR inhibition reduces CCoV replication, identifying AhR as a possible candidate target for CCoV antiviral therapy.

First Description of Serological Evidence for SARS-CoV-2 in Lactating Cows.

Following the COVID-19 epidemic outbreak in Ariano Irpino, Campania region (Italy), we tested lactating cows for the presence of SARS-CoV-2 on a cattle farm at which, prior to the investigation, 13 of the 20 farmworkers showed COVID-19-like symptoms, and one of them died. Twenty-four lactating cows were sampled to detect SARS-CoV-2. All nasal and rectal swabs and milk samples were negative for SARS-CoV-2 RNA. Of the 24 collected serum samples, 11 showed antibodies against SARS-CoV-2 nucleocapsid protein, 14 showed antibodies against SARS-CoV-2 spike protein, and 13 developed neutralising antibodies for SARS-COV-2; all samples were negative for Bovine Coronavirus (BCoV), another betacoronavirus. To our knowledge, this is the first report of natural serological evidence of SARS-CoV-2 infection in lactating cows. We hypothesise that this may be a case of reverse zoonosis. However, the role of cattle in SARS-CoV-2 infection and transmission seems to be negligible.

Antiviral Property of the Fungal Metabolite 3-O-Methylfunicone in Bovine Herpesvirus 1 Infection.

Bovine herpesvirus type-1 (BoHV-1) is a widespread pathogen that provokes infectious rhinotracheitis and polymicrobial infections in cattle, resulting in serious economic losses to the farm animal industry and trade restrictions. To date, non-toxic active drugs against BoHV-1 are not available. The exploitation of bioactive properties of microbial products is of great pharmaceutical interest. In fact, fungi are a promising source of novel drugs with a broad spectrum of activities and functions, including antiviral properties. Hence, the potential antiviral properties of 3-O-methylfunicone (OMF), a secondary metabolite produced by Talaromyces pinophilus, were evaluated on BoHV-1. In this study, during BoHV-1 infection in bovine cells (MDBK), the non-toxic concentration of 5 µM OMF considerably reduced signs of cell death and increased cell proliferation. Furthermore, OMF significantly decreased the virus titer as well as the cytopathic effect and strongly inhibited the expression of bICP0, the major regulatory protein in the BoHV-1 lytic cycle. These findings were accompanied by a considerable up-regulation in the expression of the aryl hydrocarbon receptor (AhR), a multifunctional transcription factor also linked to the host's response to a herpesvirus infection. Overall, our results suggest that by involving AhR, OMF shows potential against a BoHV-1 infection.

Involvement of herpesviruses in cases of abortion among water buffaloes in southern Italy.

A six-year study on water buffaloes from the Campania Region (Southern Italy) was conducted to evaluate the presence of bovine/bubaline herpesviruses in cases of abortion. A total of 244 buffalo foetuses were analysed by real-time PCR to detect the presence of: bovine alphaherpesvirus 1(BoHV-1), bubaline alphaherpesvirus 1 (BuHV-1), bovine alphaherpesvirus 2 (BuHV-2), and bovine gammaherpesvirus 4 (BoHV-4). The foetuses of 14 water buffaloes that showed abortions were positive for BuHV-1 (4 animals) and/or BoHV-4 (11 animals), with one of these cases showing co-infection with BuHV-1 and BoHV-4. This study reports the first identification of BoHV-4 in water buffaloes. Cases of abortion were analysed using both molecular and cultural assays for the presence of other pathogens. In nearly all the abortion cases positive for BoHV-4, the virus was identified as a co-infecting agent together with other microorganisms, whereas in two abortion cases, it was the only pathogen found.