Cardicola forsteri and C. orientalis (Digenea: Aporocotylidae) are significant pathogens affecting the Southern bluefin tuna (SBT), Thunnus maccoyii, aquaculture industry. The aims of this research were to identify a relationship between immune gene expression of SBT and C. forsteri and C. orientalis infection burdens; and to understand how immune gene expression changes over time during commercial ranching. SBT were collected during commercial operations in 2022 after 4, 8 and 10 weeks of ranching and the transcription of interleukin 1 beta (il1b), interleukin 8 (il8), tumour necrosis factor 2 (tnf2), immunoglobulin m (igm), major histocompatibility complex 2 (mhc2) and t cell receptor beta (tcrb) were measured from the anterior kidney and gills using reverse transcription quantitative polymerase chain reaction (RT-qPCR). Cardicola forsteri and C. orientalis infection was quantified as number of eggs/mm of SBT gill filament, number of adult flukes in SBT heart and C. forsteri and C. orientalis ITS-2 rDNA copy number/mg in SBT gills. Expression of il8 and igm in anterior kidney was positively correlated to C. forsteri and C. orientalis eggs/mm of SBT gill filament and to the copy number of C. forsteri ITS-2 rDNA in SBT gills. Expression of igm and mhc2 was higher in SBT positive for adult flukes and the expression of igm, mhc2 and tcrb in the anterior kidney was positively correlated to the number of adult flukes in SBT heart. Expression of tcrb in gills was negatively correlated to the number of adult flukes in SBT heart and negatively correlated to the copy number of C. orientalis ITS-2 rDNA. Expression of the adaptive immune genes igm, mhc2 and tcrb increased over the ranching duration in the kidney of SBT and the innate immune genes il1b and il8 decreased over the ranching duration in SBT gills. Despite evidence of an immune response to C. forsteri and C. orientalis, SBT were not able to clear infections during the standard industry ranching timeline.

Spotty Liver Disease (SLD), caused by Campylobacter hepaticus, greatly impacts the health and egg production of affected layer hens and is a disease of concern in the poultry industry. This study aimed to enhance the understanding of the immune response in chickens to C. hepaticus infection and their ability to resist reinfections. One hundred- and twenty-layer chickens were allocated to 10 groups and challenged, from one to three times, with C. hepaticus HV10T, with six weeks between each challenge. Blood and cloacal swabs were collected every three weeks to assess antibody responses and Campylobacter presence. Upon necropsy, bile, spleen, jejunum, and blood samples were collected for C. hepaticus detection and host gene expression analysis using qPCR and RNA sequencing. We found that most birds challenged with C. hepaticus for the second or third time did not develop liver lesions even with the presence of C. hepaticus in their bile, suggesting that birds were resistant to disease development following repeated exposure. Anti-C. hepaticus antibodies increased significantly six weeks after a single challenge but reduced from nine weeks. Quantitative PCR demonstrated that C. hepaticus could be recovered from the bile six weeks after a single challenge and increased significantly after a secondary challenge. RNA-seq and qPCR data demonstrate an elevation in pro-inflammatory cytokines, such as interleukin 6 (IL-6) and interleukin 1ß (IL-1ß), after a secondary challenge and down-regulation during a third challenge. Expression of many genes encoding barrier-supporting proteins genes were differentially expressed, with increased expression following a third challenge compared to expression after a single challenge. Comparison of gene expression in tissues of triple to a single challenged birds demonstrated that many genes involved in cytokine activity and the JAK-STAT cascade were down-regulated whereas other immune system pathways were up-regulated. Altogether, the results indicate that over time, immune memory, enhanced barrier function, and a balanced immune response developed, resulting in reduced impact of infection on the birds. These findings show that the impact of C. hepaticus infection can be ameliorated by immune responses and hence indicate that vaccines that induce appropriate protective immune responses should provide an effective tool to reduce SLD in poultry.

Previous studies of direct-fed microbial (DFM) supplements showed variable effects on the microbiota and physiology of dairy cows. The main aims of this study were to investigate the milk microbiota of cows supplemented with a lactobacilli-based DFM compared to untreated cows; describe the changes; and quantify the association between the taxa and cow productivity. The study followed seventy-five Holstein–Friesian dairy cows supplemented with a DFM over 16 months compared to seventy-five non-supplemented cows. Twenty-five cows from each group were sampled for microbiota analysis. The top taxa significantly associated with the variables were as follows: Age (Mammaliicoccus_319276, Turicibacter), milk production (Turicibacter, Bifidobacterium_388775), DIM (Stenotrophomonas_A_615274, Pedobacter_887417), milk fat percentage (Pseudomonas_E_647464, Lactobacillus), calendar month (Jeotgalicoccus_A_310962, Planococcus), milk protein percentage (Tistrella, Pseudomonas_E_650325), experimental group (Enterococcus_B, Aeromonas), SCC (Paenochrobactrum, Pseudochrobactrum), and trimester of pregnancy (Dyadobacter_906144, VFJN01 (Acidimicrobiales)). These were identified using multivariable analysis. Twenty-six genera were associated with the differences between experimental groups, including Pseudomonas, Lactococcus and Staphylococcus. Microbial taxa that changed in relative abundance over time included Atopostipes, Brevibacterium and Succinivibrio. Many of these genera were also part of the core microbiota. Supplementation with the DFM significantly altered the milk microbiota composition in the dairy cows, highlighting the impact of long-term DFM supplementation on microbial communities.

Parasitic diseases can be a significant constraint on aquaculture industries, which continue to develop in response to the rise in global demand for sustainable protein sources. Blood flukes, Cardicola forsteri and Cardicola orientalis, are economically significant parasites of Southern bluefin tuna (Australia), Pacific bluefin tuna (Japan), and Atlantic bluefin tuna (Mediterranean) as they are responsible for blood vessel obstruction in the gills leading to branchitis and mortalities when untreated. Here, we have defined the mitochondrial genomes for these species—the first for any aporocotylids. Oxford nanopore long-read sequencing was used to sequence C. orientalis from a single individual. The mitochondrial genome of C. forsteri was assembled and curated from available sequence data. Both Cardicola spp. mitogenomes contained 12 protein coding, 2 ribosomal and 22 tRNA genes, with the gene order matching that of Asian schistosomes. A control region was identified for each species which contained long and short repeats; the region for C. forsteri was longest, and the overall pattern differed between the two species. A surprisingly high nucleotide diversity was observed between the two species, generating interest into the mitochondrial genes of related species. This paper provides a useful resource for future genetics-based research of aporocotylids and other flatworm parasites of socioeconomic significance.

Necrotic enteritis (NE), caused by Clostridium perfringens, is a debilitating disease that results in significant production losses in the poultry industry. Traditionally, antibiotics have been used to control NE in flocks; however, due to concerns about the potential for selection of antibiotic resistance, antibiotic residues in meat, and restrictions on antibiotic use in some regions, alternative methods to control this disease are needed. In previous studies, proteins such as NetB, the key virulence factor, and alpha toxin have been used individually as subunit vaccines, but only partial protection was induced. It appears that a single subunit antigen is insufficient to produce high levels of protection. Here, an experimental vaccine, incorporating fragments from four antigens, was designed and tested. To simplify the production and delivery of multiple recombinant antigens, a novel quadrivalent (QV) fusion protein was designed. The QV-protein vaccine and each of the individual proteins were tested as vaccine candidates in a necrotic enteritis challenge model. The birds were vaccinated subcutaneously twice and then challenged with a pathogenic strain of C. perfringens. The development of necrotic lesions was scored, and serum IgY antibody responses were assessed. Compared with unvaccinated birds, birds vaccinated with QV-protein had significantly lower lesion scores (p < 0.05). Western blot and ELISA analyses demonstrated that vaccination with the QV-protein induced antibodies specific for all four target protein fragments within the QV-protein. In conclusion, the QV-protein provides the basis for ongoing NE vaccine development.

In this age of rapid technological advancement, next-generation sequencing (NGS) has emerged as a powerful tool for exploring the complexities of host-pathogen interactions. It enables the study of microorganism composition, functional potential, virulence factors, drug resistance mechanisms, host responses to infection, and many more facets of this intricate interplay. This protocol offers a cost-effective and scalable method for analysing the microbial composition of various samples. It accommodates up to 300 samples in a single Illumina MiSeq run, providing insights into bacterial, Archaeal, and fungal composition. This protocol primarily targets the V3-V4 region of the 16S rRNA gene for bacterial species-level identification and employs specific primers for Archaea and the internal transcribed spacer 1 gene (ITS-1) for fungal composition analysis. It has been successfully applied to diverse sample types.

The current study examined the effects of lactobacilli-based direct-fed microbial (DFM) supplementation on the microbiota composition and diversity in ruminal fluid samples collected from dairy cows. Over 18 months (September 2021 through January 2023), the rumen bacterial and archaeal communities of fifty cows, supplemented with the DFM (DFM; n = 25) or serving as un-supplemented controls (CON; n = 25), were examined using 16S rRNA gene amplification and sequence analysis of DNA extracted from ruminal samples. Microbial diversity was assessed through alpha- and beta-diversity metrics (p<0.05). Linear discriminant analysis effect size (LEfSe) analysis was performed to identify taxa driving the changes seen in the microbiota between experimental groups and temporally within each group (p<0.05). Bacillota and Bacteroidota were the major bacterial phyla, while Methanobacteriaceae was the predominant archaeal family. Bacterial genera such as Eubacterium_Q, Atopobium sp. UBA7741, and Sharpea were significantly more abundant in the DFM group, while Bacillus_P_294101 and SFMI01 had higher abundance in the CON group. The results also indicated significant temporal variations in ruminal microbial diversity, with specific taxa exhibiting different abundances between the DFM and CON groups. This study provides insights into how DFM feed additives can modulate the ruminal microbiota in dairy cows, revealing specific microbial shifts in response to supplementation.

The rumen microbiota of dairy cows plays a crucial role in fermenting fibrous material, essential for nutrient extraction and overall productivity, detoxification of anti-nutritional toxic compounds, synthesis of vital nutrients, and is essential for optimal animal health. This study investigated the impact of Lentilactobacillus-, Lactocaseibacillus-, and Lacticaseibacillus-based direct-fed microbial (DFM) supplementation on dairy cows’ faecal microbial composition and diversity. The study was carried out on a commercial dairy farm using 50 Holstein-Friesian cows randomly assigned into control (CON) and treatment (TRT) groups. Faecal samples were collected directly from the rectum every two months from September 2021 to January 2023. The bacterial 16S rRNA gene and fungal ITS-1 regions were amplified, sequenced, and analysed. Microbial diversity was assessed through alpha- and beta-diversity metrics. Linear discriminant analysis effect size (LEfSe) was performed to identify which taxa were driving the changes seen in the microbiota over time and treatment. Bacteroidaceae were the most prevalent bacterial family, followed by Lachnospiraceae and Muribaculaceae in both CON and TRT cows. Ascomycota, Basidiomycota, and Mucoromycota were the dominant three fungal phyla in the faeces of both CON and TRT cows. Bacterial genera Fructilactobacillus was abundant in the CON and Absicoccus in the TRT groups. Fungal taxa Chaetothryriales_incertae_sedis and Pseudomentella were absent in the faeces of TRT cows. Significant temporal and specific taxonomic differences were observed between the CON and TRT groups. The study’s findings underscore the dynamic nature of microbial communities and the importance of targeted dietary interventions. Further research is necessary to elucidate these microbial shifts, long-term impacts, and functional implications, aiming to optimise ruminant nutrition and enhance dairy cow performance.

Chickens in Australia have recently been identified with symptoms and morphological findings including spondylitis attributed to pathogenic Enterococcus cecorum. Notably, there is limited information on clinical E. cecorum strains in Australia. The cpsO gene, located downstream of the capsular polysaccharide (cps) locus, was recently reported to successfully differentiate between pathogenic and commensal E. cecorum strains, as this gene is highly conserved in the pathogenic strains. In this study, pathogenic E. cecorum, with a conserved cpsO gene, was detected on 1 of the 2 farms studied in Australia. E. cecorum strains isolated from clinical sites of the diseased birds from the second farm did not have the cpsO gene and were distant from the isolates of the first farm. A cpsO PCR of the caecal content of the birds on this farm was positive, while cpsO PCR of washed culture plates where the tissue extracts were spread onto and incubated for bacterial growth was negative. This suggests that pathogenic E. cecorum with the cpsO gene, as detected in Farm 1 and reported in other countries, was present in the second farm but could not grow on the selective agar plates during the initial step of E. cecorum isolation. Nevertheless, E. cecorum isolated from the clinical sites on the second farm might represent the pathogenic strain, but further animal studies are required to validate this possibility. Phylogenetic analysis showed that the pathogenic strains in Australia were most closely related to the clinical strains in North America.

Campylobacter concisus is an opportunistic bacterial pathogen linked with a range of human diseases. The objective of this study was to investigate the viable but nonculturable (VBNC) state of the bacterium. To induce the VBNC state, C. concisus cells were maintained in sterilized phosphate-buffered saline at 4°C for three weeks. The VBNC cells were monitored using quantitative analysis by propidium monoazide (PMAxx) coupled with quantitative real-time PCR (PMAxx-qPCR), targeting the DNA gyrase subunit B gene. The results demonstrated that C. concisus ATCC 51562 entered the VBNC state in 15 days, while ATCC 51561 entered the VBNC state in 9 days. The viable cell counts, assessed by PMAxx-qPCR, consistently remained close to the initial level of 107 CFU ml−1, indicating a substantial portion of the cell population had entered the VBNC state. Notably, morphological analysis revealed that the VBNC cells became coccoid and significantly smaller. The cells could be resuscitated through a temperature increase in the presence of a highly nutritious growth medium. In conclusion, under environmental stress, most C. concisus cells converted to the VBNC state. The VBNC state of C. concisus may be important for its environmental survival and spread, and the presence of VBNC forms should be considered in environmental and clinical monitoring.

The blood fluke Cardicola forsteri (Trematoda: Aporocotylidae) is a pathogen of ranched bluefin tuna in Japan and Australia. Genomics of Cardicola spp. have thus far been limited to molecular phylogenetics of select gene sequences. In this study, sequencing of the C. forsteri genome was performed using Illumina short-read and Oxford Nanopore long-read technologies. The sequences were assembled de novo using a hybrid of short and long reads, which produced a high-quality contig-level assembly (N50 > 430 kb and L50 = 138). The assembly was also relatively complete and unfragmented, comprising 66% and 7.2% complete and fragmented metazoan Benchmarking Universal Single-Copy Orthologs (BUSCOs), respectively. A large portion (> 55%) of the genome was made up of intergenic repetitive elements, primarily long interspersed nuclear elements (LINEs), while protein-coding regions cover > 6%. Gene prediction identified 8,564 hypothetical polypeptides, > 77% of which are homologous to published sequences of other species. The identification of select putative proteins, including cathepsins, calpains, tetraspanins, and glycosyltransferases is discussed. This is the first genome assembly of any aporocotylid, a major step toward understanding of the biology of this family of fish blood flukes and their interactions within hosts.

Vaccines are very effective in providing protection against many infectious diseases. However, it has proven difficult to develop highly efficacious vaccines against some pathogens and so there is a continuing need to improve vaccine technologies. The first successful and widely used vaccines were based on attenuated pathogens (e.g., laboratory passaged Pasteurella multocida to vaccinate against fowl cholera) or closely related non-pathogenic organisms (e.g., cowpox to vaccinate against smallpox). Subsequently, live vaccines, either attenuated pathogens or non-pathogenic microorganisms modified to deliver heterologous antigens, have been successfully used to induce protective immune responses against many pathogens. Unlike conventional killed and subunit vaccines, live vaccines can deliver antigens to mucosal surfaces in a similar manner and context as the natural infection and hence can often produce a more appropriate and protective immune response. Despite these advantages, there is still a need to improve the immunogenicity of some live vaccines. The efficacy of injectable killed and subunit vaccines is usually enhanced using adjuvants such mineral salts, oils, and saponin, but such adjuvants cannot be used with live vaccines. Instead, live vaccines can be engineered to produce immunomodulatory molecules that can stimulate the immune system to induce more robust and long-lasting adaptive immune responses. This review focuses on research that has been undertaken to engineer live vaccines to produce immunomodulatory molecules that act as adjuvants to increase immunogenicity. Adjuvant strategies with varying mechanisms of action (inflammatory, antibody-mediated, cell-mediated) and delivery modes (oral, intramuscular, intranasal) have been investigated, with varying degrees of success. The goal of such research is to define adjuvant strategies that can be adapted to enhance live vaccine efficacy by triggering strong innate and adaptive immune responses and produce vaccines against a wider range of pathogens.

The current in vitro study aimed to investigate the effects of a processed sugarcane extract on the viability of avian Eimeria sporozoites. Treatments were applied to hatched sporozoites: 1) without additives (no-treatment control); 2) with ethanol; 3) with salinomycin; 4) with Polygain™. All treatments were incubated in RPMI media containing live sporozoites at 37 °C for 14 h and then the number of viable sporozoites were counted. Compared to the no-treatment control, Polygain™ decreased (P < 0.001) the counts of E. maxima, E. acervulina, E. bruneti, and E. mitis sporozoites to a level similar to salinomycin (P > 0.05). In conclusion, Polygain™ could be a potential candidate as an anticoccidial agent.

Background Of increasing importance to the medical and veterinary communities is the zoonotic filarioid nematode Onchocerca lupi. Onchocercosis, thus far found in wolves, dogs, cats and humans, is diagnosed via skin snips to detect microfilariae and surgical removal of adults from the eye of the host. These methods are time-consuming, laborious and invasive, highlighting the need for new tools for the diagnosis of O. lupi in susceptible hosts. Symptoms related to the presence of the adults in the eye can range from none apparent to severe, including blindness. No reliable chemotherapeutic protocols are available, as yet, to eliminate the infection. Paramyosin, an invertebrate-specific protein, has been well-studied as an allergen, diagnostic marker and vaccine candidate. The aim of this study, therefore, was to isolate and characterise paramyosin from O. lupi to assess its suitability for the development of a serological diagnostic assay. Methods The adult and microfilarial stages of O. lupi were isolated from the eyes and skin of a 3-year-old male dog. Total RNA was extracted and reverse transcribed into single stranded cDNA. Reverse-transcription PCR was used to isolate a full-length paramyosin cDNA from adult worms and to investigate the temporal expression patterns of this gene. All amplicons were sequenced using dideoxy chain termination sequencing. Bioinformatics was used to predict the amino acid sequence of the gene, to compare the DNA and protein sequences with those available in public databases and to investigate the phylogenetic relationship of all molecules. Antibody binding sites were predicted using bioinformatics and mapped along with published antigenic epitopes against the O. lupi paramyosin protein. The native protein, and three smaller recombinantly expressed peptides, were subjected to western blot using serum from dogs both positive and negative for O. lupi . Results Paramyosin of O. lupi was herein molecularly characterized, encoded by a transcript of 2,643 bp and producing a protein of 881 amino acids (101.24 kDa). The paramyosin transcript was detected, by reverse transcription PCR, in adults and microfilariae, but not in eggs. Phylogenetic analysis indicates that this molecule clusters with paramyosins from other filarioids to the exclusion of those from other taxa. A total of 621 unique antibody binding epitopes were predicted for this protein and another 28 were conserved in other organisms. This information was used to design three peptides, for recombinant expression, to identify the antibody binding epitope(s) and reduce potential cross-reactivity with serum from dogs infected with other filarioid nematodes. Native paramyosin, purified from microfilariae and adults, was detected by antibodies present in serum from dogs with known O. lupi infections. Conclusions Data provided herein may assist in the development of a serological diagnostic test, based on antibodies to O. lupi paramyosin, for the diagnosis of this infection, in order to gain more information on the real distribution of this little known filarioid of zoonotic concern.

Cardiopulmonary infections by Angiostrongylus chabaudi affect domestic and wild felids but, due to limited information on the biology of this nematode, its pathogenicity remains unclear. This article describes the histopathological alterations associated with Angiostrongylus infection in a wildcat from Bulgaria, and reviews current literature on this feline angiostrongylid. Nematodes were isolated from lung lavage and faecal samples of a road killed wildcat in Southern Bulgaria. The morphological identification of parasite larvae as A. chabaudi was confirmed by molecular analysis of part of the 18S ribosomal RNA gene. Upon histopathological examination, severe granulomatous pneumonia, ranging from multifocal to coalescing, and pulmonary vascular lesions were observed. Extensive alveolar collapse, alveolar emphysematous changes, parenchymal haemorrhages and small artery wall hyperplasia were observed in the parenchyma adjacent to the granulomas. Histopathological examination revealed the presence of cross-sections of adult female parasites within the lumen of the pulmonary artery branches, the intima altered markedly by subendothelial proliferation and oedematous changes. This study compliments current knowledge of the pathogenesis of feline angiostrongylosis by A. chabaudi in wildcats, as well as of the distribution of this little-known parasite.

High-throughput molecular and computer technologies have become instrumental for systems biological explorations of pathogens, including parasites. For instance, investigations of the transcriptomes of different developmental stages of parasitic nematodes give insights into gene expression, regulation and function in a parasite, which is a significant step to understanding their biology, as well as interactions with their host(s) and disease. This chapter (1) gives a background on some key parasitic nematodes of socioeconomic importance, (2) describes sequencing and bioinformatic technologies for large-scale studies of the transcriptomes and genomes of these parasites, (3) provides some recent examples of applications and (4) emphasizes the prospects of fundamental biological explorations of parasites using these technologies for the development of new interventions to combat parasitic diseases.

Summary There is growing interest in the ‘farm effect’ on the spectrum of allergy. Evidence concerning the farm effect on asthma, eczema, and allergic rhinitis has been systematically synthesized, but without a specific focus on objective markers of sensitization. This focus is important, as farm exposures may be related to allergy but not to non‐allergic phenotypes of disease. We aimed to systematically review and meta‐analyse literature that has investigated associations between farm exposure at any age and objective measures of atopy, that is serum IgE or skin prick tests results. Using predefined inclusion and exclusion criteria, we identified 29 articles for review. IgE levels were measured in either childhood or adulthood by eighteen studies, while skin prick testing was performed in sixteen studies. Newcastle‐Ottawa Scale quality assessments indicated that the majority of these studies demonstrated a representative sample of selected participants. Due to significant heterogeneity in study measures and methodology between studies, only few were meta‐analysed. This meta‐analysis showed a significant protective effect of farm exposure before 1 year of life on allergic sensitization ( OR = 0.60 [0.52–0.70]). Farm exposure during childhood was also associated with a reduced risk of sensitization to cat or timothy ( OR = 0.60 [0.51–0.70]; OR =0.46 [0.41–0.51]). Studies investigating the effect of farm exposure in adult life could not be meta‐analysed, and their results were inconsistent. Insufficient studies investigated food sensitization as an outcome to allow synthesis. The majority of studies included in this review investigated childhood farm exposure, finding evidence to support a protective childhood ‘farm effect’ against subsequent atopy. There is inconsistent evidence on the association between farm exposure in adulthood and allergic sensitization. Further studies are needed to tease out the exact exposures and timing associated with farming environments that protect against allergic disease.

Natural enemies of ticks include the parasitoid wasp Ixodiphagus hookeri (Hymenoptera: Encyrtidae). The aim of this preliminary study was to investigate the occurrence of I. hookeri DNA in a community of ticks (Ixodes ricinus, Dermacentor marginatus, Hyalomma marginatum, Haemaphysalis inermis and Rhipicephalus turanicus). From May 2010 to March 2012, ticks were collected monthly by dragging and flagging, identified, and 481 adults and 305 nymphs screened molecularly for infection with I. hookeri. Of the samples tested (n = 786), 3.1% (n = 25) were positive for I. hookeri DNA, 7.2% (n = 22) in nymphs and 0.6% (n = 3) in adults. I. hookeri DNA was only detected in I. ricinus. This study shows that I. hookeri infests I. ricinus in southern Italy, with nymphs being the main developmental stage affected by this wasp.

Crenosoma vulpis is a metastrongyloid nematode primarily associated with respiratory tract infections of red foxes in North America and Europe. Sporadic cases have also been reported in domestic dogs. The present study aimed to provide morphological, molecular, and epidemiological data on the geographical distribution of this nematode throughout Italy. From 2012 to 2014, 12 of the 138 foxes examined, three dogs and one badger scored positive for C. vulpis. Forty adults were isolated from foxes and the badger, whereas first-stage larvae were detected in the three dogs. All specimens were morphologically identified as C. vulpis, and 28 nematodes were also molecularly characterized by sequencing mitochondrial (12S ribosomal DNA (rDNA)) and nuclear (18S rDNA) ribosomal genes. Four haplotypes were identified based on the 12S rDNA target gene, with the most representative (78.5 %) designated as haplotype I. No genetic variability was detected for the 18S rDNA gene. The molecular identification was consistent with the distinct separation of species-specific clades inferred by the phylogenetic analyses of both mitochondrial and ribosomal genes. Data herein reported indicates that C. vulpis has a wide distribution in foxes from southern Italy, and it also occurs in dogs from southern and northern regions of the country. Practitioners should consider the occurrence of this nematode in the differential diagnosis of canine respiratory disease, particularly in dogs living close to rural areas where foxes are present.

Onchocerca lupi is a neglected filarioid causing nodular lesions associated with acute or chronic ocular disease in dogs. Despite the recent appraisal of its zoonotic potential, human cases are increasingly reported in the Old and New Worlds. Therefore, the development of accurate tools for the rapid diagnosis of O. lupi infections in dogs is becoming a priority. In this study, we conducted a preliminary investigation aimed at evaluating the usefulness of a commercially available ELISA test for the detection of O. lupi antigens in canine sera. The potential use of this tool for larger epidemiological studies of canine onchocerciasis is discussed.