Abstract A scarcity of information on the occurrence of zoonotic vector‐borne pathogens (VBPs), alongside a lack of human and animal health authorities’ awareness of pre‐existing data, augment the risk of VBP infection for local people and limit our ability to establish control programs. This holds especially true in low‐middle income countries such as Bosnia and Herzegovina (BiH). This dearth of information on zoonotic VBPs is bolstered by the inability of previously used diagnostic tests, including conventional molecular diagnostic methods, to detect the full spectrum of relevant pathogens. Considering this, we set out to apply a microfluidic qPCR assay capable of detecting 43 bacterial and protozoan pathogens from blood to accrue critical baseline data for VBPs occurrence in BiH. A total of 408 dogs were tested of which half were infected with at least one VBP of zoonotic or veterinary importance. Leishmania infantum was found in 18% of dogs, reaching a prevalence as high as 38% in urbanized areas of Sarajevo. These data highlight substantially higher levels of L. infantum prevalence when compared to that previously reported using conventional methods using the same samples. Additionally, this high‐throughput microfluidic qPCR assay was able to detect pathogens rarely or never reported in canines in BiH, including Anaplasma phagocytophilum (3%), Anaplasma platys (0.2%), haemotropic Mycoplasma (1%) and Hepatozoon canis (26%). Our report of the endemicity of important zoonotic pathogens and those of clinical significance to dogs emphasizes the need for urgent implementation of surveillance and control for VBPs in BiH, targeting both animal and human infections within the country.

Abstract Background Crimean–Congo haemorrhagic fever (CCHF) is a widespread tick‐borne zoonosis with reported detection of virus and/or virus‐specific antibodies from over 57 countries across Africa, Asia, Europe and the Middle East and is endemic in the Balkans. Detection of Crimean–Congo Haemorrhagic Fever Virus (CCHFV) antibodies in domestic ruminants has been important in providing initial evidence of virus circulation and in localising CCHFV high‐risk spots for human infection. Objectives The present study investigated the possible exposure of sheep to CCHFV in Bosnia and Herzegovina (B&H). Methods To investigate the presence of anti‐CCHFV antibodies in sheep, all sera (n = 176) were tested using multi‐species double antigen enzyme‐linked immunosorbent assay (ELISA). Reactive sera were further complementary tested by adapted commercial indirect immunofluorescence assay (IFA) using FITC‐conjugated protein G instead of anti‐human immunoglobulins. Results CCHFV specific antibodies were detected in 17 (9.66%) animals using ELISA test. All negative sera were determined as negative by both tests, while 13 out of 17 ELISA‐positive reactors were also determined as unambiguously positive by IFA test. The age group with the highest proportion of seropositive rectors were the oldest animals. Conclusions This is the first report of anti‐CCHFV antibodies in sheep from B&H providing the evidence of CCHFV circulation in the country's sheep population. So far, these findings indicate the circulation of the virus in the westernmost region of the Balkans and point to the potential CCHFV spread further out of this endemic area.

The order Accipitriformes comprises the largest group of birds of prey with 260 species in four families. So far, 21 haemosporidian parasite species have been described from or reported to occur in accipitriform birds. Only five of these parasite species have been characterized molecular genetically. The first part of this study involved molecular genetic screening of accipitriform raptors from Austria and Bosnia-Herzegovina and the first chromogenic in situ hybridization approach targeting parasites in this host group. The aim of the second part of this study was to summarize the CytB sequence data of haemosporidian parasites from accipitriform raptors and to visualize the geographic and host distribution of the lineages. Blood and tissue samples of 183 accipitriform raptors from Austria and Bosnia-Herzegovina were screened for Plasmodium, Haemoproteus and Leucocytozoon parasites by nested PCR, and tissue samples of 23 PCR-positive birds were subjected to chromogenic in situ hybridization using genus-specific probes targeting the parasites’ 18S rRNAs. All published CytB sequence data from accipitriform raptors were analysed, phylogenetic trees were calculated, and DNA haplotype network analyses were performed with sequences from clades featuring multiple lineages detected in this host group. Of the 183 raptors from Austria and Bosnia-Herzegovina screened by PCR and sequencing, 80 individuals (44%) were infected with haemosporidian parasites. Among the 39 CytB lineages detected, 18 were found for the first time in the present study. The chromogenic in situ hybridization revealed exo-erythrocytic tissue stages of Leucocytozoon parasites belonging to the Leucocytozoon toddi species group in the kidneys of 14 infected birds. The total number of CytB lineages recorded in accipitriform birds worldwide was 57 for Leucocytozoon, 25 for Plasmodium, and 21 for Haemoproteus. The analysis of the DNA haplotype networks allowed identifying numerous distinct groups of lineages, which have not yet been linked to morphospecies, and many of them likely belong to yet undescribed parasite species. Tissue stages of Leucocytozoon parasites developing in accipitriform raptors were discovered and described. The majority of Leucocytozoon and Haemoproteus lineages are specific to this host group, but most Plasmodium lineages were found in birds of other orders. This might indicate local transmission from birds kept at the same facilities (raptor rescue centres and zoos), likely resulting in abortive infections. To clarify the taxonomic and systematic problems, combined morphological and molecular genetic analyses on a wider range of accipitriform host species are needed.

Background The order Accipitriformes comprises the largest group of birds of prey with 260 species in four families. So far, 21 haemosporidian parasite species have been described from or reported to occur in accipitriform birds. Only five of these parasite species have been characterized molecular genetically. The first part of this study involved molecular genetic screening of accipitriform raptors from Austria and Bosnia-Herzegovina and the first chromogenic in situ hybridization approach targeting parasites in this host group. The aim of the second part of this study was to summarize the CytB sequence data of haemosporidian parasites from accipitriform raptors and to visualize the geographic and host distribution of the lineages. Methods Blood and tissue samples of 183 accipitriform raptors from Austria and Bosnia-Herzegovina were screened for Plasmodium , Haemoproteus and Leucocytozoon parasites by nested PCR, and tissue samples of 23 PCR-positive birds were subjected to chromogenic in situ hybridization using genus-specific probes targeting the parasites’ 18S rRNAs. All published CytB sequence data from accipitriform raptors were analysed, phylogenetic trees were calculated, and DNA haplotype network analyses were performed with sequences from clades featuring multiple lineages detected in this host group. Results Of the 183 raptors from Austria and Bosnia-Herzegovina screened by PCR and sequencing, 80 individuals (44%) were infected with haemosporidian parasites. Among the 39 CytB lineages detected, 18 were found for the first time in the present study. The chromogenic in situ hybridization revealed exo-erythrocytic tissue stages of Leucocytozoon parasites belonging to the Leucocytozoon toddi species group in the kidneys of 14 infected birds. The total number of CytB lineages recorded in accipitriform birds worldwide was 57 for Leucocytozoon , 25 for Plasmodium , and 21 for Haemoproteus . Conclusion The analysis of the DNA haplotype networks allowed identifying numerous distinct groups of lineages, which have not yet been linked to morphospecies, and many of them likely belong to yet undescribed parasite species. Tissue stages of Leucocytozoon parasites developing in accipitriform raptors were discovered and described. The majority of Leucocytozoon and Haemoproteus lineages are specific to this host group, but most Plasmodium lineages were found in birds of other orders. This might indicate local transmission from birds kept at the same facilities (raptor rescue centres and zoos), likely resulting in abortive infections. To clarify the taxonomic and systematic problems, combined morphological and molecular genetic analyses on a wider range of accipitriform host species are needed.

Red foxes are the most abundant wild carnivore species in Europe commonly exposed to pathogenic Leptospira and Hepatozoon canis. Despite high seroprevalence, the clinical disease caused by these pathogens in red foxes has never been reported. Herein, we report the first-ever case of a fatal Leptospira spp. and H. canis coinfection in a two-month-old red fox cub with acute haemolytic anaemia, mild bronchopneumonia, intraalveolar haemorrhage, and tubulonephrosis. The presence of pathogenic Leptospira spp. DNA was detected in the kidney and lung tissues of the infected animal. In contrast to our previous knowledge, we believe that such fatal cases due to concomitant infection by Leptospira spp. and H. canis, especially in young animals, may commonly occur in nature. However, further studies are required to identify other factors that possibly contribute to the severity and the pathogenic effect of Leptospira spp. and H. canis infections in red foxes.

Abstract Infections with various bacteria, especially gram-negative aerobes, are a well-recognized problem in captive cold-blooded animals with immunocompromised health status, or in those kept under poor conditions. Pseudomonas is one of the most represented genera. Here, we present a case of fatal disseminated infection caused by Pseudomonas aeruginosa in a captive green iguana kept at the “Pionirska dolina” Zoo in Sarajevo, Bosnia and Herzegovina. At necropsy, severe stomatitis, pneumonia, hepatitis and nephritis, accompanied with focally extensive dermatitis were observed. Histopathology revealed multifocal necrosis in various visceral organs. Culture and subsequent MALDI-TOF MS analysis were conducted to identify the isolate as P. aeruginosa. Antimicrobial susceptibility testing revealed a wide susceptibility of the isolate, however applied therapy was instilled too late in the presented case. This case demonstrates the significance of timely and accurate identification, and antimicrobial susceptibility testing of bacterial isolates implicated in the pathology of captive reptiles. The importance of monitoring the adequate environmental conditions (enclosure temperature, humidity and conformation), health status and possible clinical signs of illness are highlighted.

Abstract The study has aimed to investigate and determine the anatomical position, shape, size, and histological features of the ductus venosus, and its role as a shunt in the fetal circulatory system in domestic ruminants. The research was conducted on 19 bovine, 11 sheep and 5 goat fetuses, aborted at the late stage of pregnancy or deceased just after delivery. The general anatomy of the ductus venosus was investigated by in-situ dissection of the corrosive cast obtained by injection of 25% solution of Vinylite mass through the umbilical vein. For histological examination, the fetal tissue samples were stained with Hematoxylin and Eosin, Masson’s trichrome, Verhoeff-Van Gieson and Gomoriꞌs silver stain. The results showed that ruminant fetal ductus venosus is a curved, trumpet-shaped vessel, situated in the central part of the liver, above the porta hepatis. Its ventral part is constricted in the form of an isthmus, having a prominent lip-like thickening at the junction with the portal sinus. Histological examination showed the dominant presence of collagen and elastic fibers in its tunica media, with thin bands of smooth muscle fibers oriented in a longitudinal and circular direction indicating ability for vasoconstriction and vasodilatation.

Melanomacrophages of fish are commonly explored as biomarkers of water pollution and are considered to be sensitive albeit non-specific health indicators in water ecosystems. Sharks as long living marine species are good sentinel species. This study presents morphometric data for splenic and hepatic melanomacrophages (MMC), and observed histopathology in ten lesser-spotted catsharks, Scyliorhinus canicula (L.), one of the most abundant shark species in the eastern Adriatic Sea. At necropsy, we collected random tissue samples from liver, brain, gallblader, pancreas, spleen, kidney, gills, entire digestive system, thyroid gland, rectal gland, entire urogenital (male samples) and genital system (female samples). Collected tissue samples were routinely processed and stained with hematoxylin-eosin, Periodic Acid-Schiff, and Masson Trichrome for microscopic examinations and morphometry. There was a minimal number of histopathological lesions in the examined sharks, but morphometric values reported herein were three folds higher than in previous studies in free-ranging sharks. Studies on larger numbers of sharks are needed to elucidate the biological significance of our finding in the context of population decline of the lesser-spotted catshark.