The immune response in canine and human leishmaniasis and how this influences the diagnosis- a review and assessment of recent research.

Leishmaniasis is a widespread but still underdiagnosed parasitic disease that affects both humans and animals. There are at least 20 pathogenic species of Leishmania, most of them being zoonotic. The diagnosis of leishmaniasis remains a major challenge, with an important role being played by the species of parasites involved, the genetic background, the immunocompetence of the host. This paper brings to the fore the sensitivity of the balance in canine and human leishmaniasis and addresses the importance of the host's immune response in establishing a correct diagnosis, especially in certain cases of asymptomatic leishmaniasis, or in the situation the host is immunosuppressed or acquired leishmaniasis through vertical transmission. The methods considered as a reference in the diagnosis of leishmaniasis no longer present certainty, the diagnosis being influenced mostly by the immune response of the host, which differs according to the presence of other associated diseases or even according to the breed in dogs. Consequently, the diagnosis and surveillance of leishmaniasis cases remains an open topic, requiring new diagnostic methods adapted to the immunological state of the host.

Septicemic Outbreak in A Rainbow Trout Intensive Aquaculture System: Clinical Finds, Etiological Agents, and Predisposing Factors.

On the 23rd of September 2022, a small intensive aquaculture unit populated with rainbow trout (Oncorhynchus mykiss) reported increased mortality in adults and juvenile fish. The unit comprised 12 enclosed concrete basins with a capacity of ten cubic meters of water, populated with 150 kg of fish each. Fish were subjected to a clinical examination on the site, after which whole fish were harvested for a bacteriological and histopathological examination. Water quality parameters were examined using classic biochemical methods and Fourier Transform Infrared Spectroscopy in order to find out whether the environment in which the fish live is also a predisposing factor that could facilitate different pathogens and induce a state of disease in the fish. Real-time PCR was performed on strains of Aeromonas spp. sampled from the fish to accurately identify the pathogen species. The goal was to accurately identify the problems and predisposing factors that lead to disease outbreaks.

Short Review on the Biological Activity of Cyclodextrin-Drug Inclusion Complexes Applicable in Veterinary Therapy.

Cyclodextrins (CDs) are a family of carrier molecules used to improve the pharmacokinetic parameters of therapeutic molecules. These cyclic oligosaccharides have medical and pharmaceutical applications by being able to form inclusion complexes with molecules that are poorly soluble in water. The benefits of these complexes are directed towards improving the chemical and biological properties-i.e., solubility, bioavailability, stability, non-toxicity and shelf life of drug molecules. Since the 1960s, the first inclusion complexes used in therapeutics were those with α-, ß- and γ-CD, which proved their usefulness, but had certain degrees of particularly renal toxicity. Currently, to correct these deficiencies, ß-CD derivatives are most frequently used, such as sulfobutylether-ß-CD, hydroxypropyl-ß-CD, etc. Therefore, it is of interest to bring to the attention of those interested the diversity of current and potential future clinical applications of inclusion complexes in veterinary medicine and to present the contribution of these inclusion complexes in improving drug efficacy. The most important biological activities of ß-CD complexed molecules in the veterinary field are summarized in this short review.

Comparative Study of Flesh Quality, Blood Profile, Antioxidant Status, and Intestinal Microbiota of European Catfish (Silurus glanis) Cultivated in a Recirculating Aquaculture System (RAS) and Earthen Pond System.

With the increasing demand for European catfish, traditional extensive growth methods in polyculture are no longer sufficient to meet market needs. Therefore, this study aimed to identify indicators for improving recirculating aquaculture system (RAS) technology by determining and comparing growth performance, flesh quality, blood profile, oxidative status, and intestinal microbiota parameters between fish cultivated in a RAS and an earthen pond. Results revealed that RAS-grown fish had a higher fat content compared to pond-grown fish, while no significant differences were found for growth parameters. Sensory analysis showed no significant difference in taste between the two groups. Blood composition analysis showed small differences. Oxidative status analyses showed higher catalase and glutathione peroxidase activities in RAS-grown fish and slightly higher superoxide dismutase activity in pond-grown fish. Microbial analysis showed differences in the intestinal microflora, with a higher total number of aerobic germs and anaerobic germs and a lower total number of sulfite-reducing clostridia in RAS-grown fish. This study provides valuable insights into the comparative performance of a RAS and a pond rearing system in European catfish production, potentially informing future growth technologies.

The Risk of Emerging of Dengue Fever in Romania, in the Context of Global Warming.

(1) Background: Few studies to date have assessed the influences induced by climate change on the spatial distribution and population abundance of Aedes albopictus using the latest climate scenarios. In this study, we updated the current distribution of Ae. albopictus mosquitoes and evaluated the changes in their distribution under future climate conditions, as well as the risk of dengue virus emergence in Romania. (2) Methods: Under the two scenarios: High scenario (HS) when no drastic measures to reduce the effects of global warming will be taken, or they are not effective and low scenario (LS) when very stringent greenhouse control measures will be implemented. (3) Results: The results estimate an increase in temperatures in Romania of up to 2.6 °C in HS and up to 0.4 °C in LS, with an increase in the period of virus replication within the vector from June to October in HS and from May to September in LS. Moreover, in 2022, Ae. albopictus was reported in a new county, where it was not identified at the last monitoring in 2020. (4) Conclusions: The rapid spread of this invasive species and the need to implement monitoring and control programs for the Aedes population in Romania are emphasized.

Strategies for the Diagnosis of Granulocytic Anaplasmosis in Two Naturally Infected Dogs.

This study describes granulocytic anaplasmosis in two dogs naturally infected with Anaplasma phagocytophilum. The 3-year-old dogs (male and female) came from the same household and were referred to the Faculty of Veterinary Medicine in Iasi for blood donation. They were subject to standard routine tests: haematology blood test, blood smear, and serological tests (VETSCAN® FLEX4 and IDEXX SNAP 4Dx Plus). The female dog had no medical problems, while the male dog experienced joint pain. The blood smear was negative for tick-borne pathogens, and the haematology findings indicated thrombocytopenia in both dogs, with the male dog also displaying eosinophilia. The two dogs were mildly positive in the ELISA tests for the detection of Anaplasma spp. antibodies; therefore, the blood samples were tested using the qRT-PCR method for Anaplasma platys and Anaplasma phagocytophilum. The qRT-PCR result was negative for A. platys, but it was positive for A. phagocytophilum. The treatment consisted of the administration of doxycycline for 28 days. In conclusion, the high number of cases with non-specific clinical signs, the different sensitivity and specificity of the immunochromatographic serological tests, as well as the possibility of confusing the morula during the cytological examination, make the molecular test mandatory for precise diagnosis.

An Overview on Leishmaniasis in Romania: Diagnosis and Therapeutics.

Leishmaniasis, a vector-borne disease considered to be one of the twenty neglected diseases by the World Health Organization, represents one of the public health concerns in endemic countries. In humans, as well as in animal counterparts, the infection can evolve with different clinical localizations, such as those that are cutaneous, mucocutaneous and visceral. Romania has been traditionally considered a nonendemic country for Leishmania species infection and has had sporadic positive human cases; however, the climate change recorded in recent decades has created potentially optimal conditions for the preponderant vectors of Phlebotomus spp., which has lately been identified in various parts of country. Moreover, with people and dogs (the prevailing hosts) traveling in endemic countries, the disease was imported and diagnosed in both species, and became a medical concern. In this review, we focused on the: (1) epidemiological data of leishmaniasis cases, both in humans and animals, reported by Romania; (2) diagnostic tools available for confirmation since there is a lack of gold-standard laboratory methods for human and dog patients; and (3) conventional antileishmanial therapy.

Dirofilaria repens predominates in shelter dogs from South Romania.

Canine filarioids are zoonotic vector-borne parasitic nematodes distributed mostly in the tropics and subtropics. The aim of this study was to determine the prevalence, distribution, etiology and genetic variation of canine filarioid infections by three different PCR techniques in four Southern Romanian counties. Blood samples of 300 shelter dogs were screened for infections with canine filarioids by real-time PCR. To determine filarioid species and coinfections, samples positive in the initial screening were further tested by conventional PCR and sequenced. Results indicated that 17% of the tested dogs were positive for at least one filarioid species. The prevalence of D. repens infection was 11.7%, significantly higher than that of D. immitis (4.7%) and A. reconditum (1.3%) (p ≤ 0.003). The high prevalence of canine filarioid infections represents a challenge to animal and human health in the South of Romania, and they should be constantly monitored.

Occurrence of Babesia Species and Co-Infection with Hepatozoon canis in Symptomatic Dogs and in Their Ticks in Eastern Romania.

Although the distribution of Babesia spp. and Hepatozoon canis is well known in Romania, there is still a marked lack of information in many places of the country. This study aimed to investigate the occurrence of these haemoparasites in symptomatic dogs and in their ticks in Iasi, eastern Romania. Ninety owned dogs were subjected to clinical examination at the Faculty of Veterinary Medicine of Iasi and all detectable ticks (58 ticks from 15 dogs) were collected. Additionally, 124 ticks collected from the coat of other dogs (no. = 23) were included. Three Babesia species were found in dogs: Babesia canis (94.4%), Babesia vogeli (3.3%), and Babesia rossi (2.2%). All the dogs resulted negative for H. canis. The ticks were identified as follows: Ixodes ricinus (64%), Dermacentor reticulatus (33%), and Rhipicephalus sanguineus group (3%). B. canis (Minimum Infection Rate; MIR = 81%), B. vogeli (MIR = 3%), and Babesia microti-like piroplasm (MIR = 1%) were found in ticks. Moreover, 15 ticks were positive for H. canis, 6 were co-infected with B. canis, and 1 with B. microti-like piroplasm. This is the first molecular identification of B. rossi in two symptomatic dogs from Romania, although further studies are needed to investigate the vector competence of other ticks from Europe.

First report of Dirofilaria repens infection in a microfilaraemic cat from Romania.

The present study describes the first report of Dirofilaria repens infection with the presence of both microfilariae and adult nematodes in a cat from Northeastern Romania. Briefly, a 5-year-old male mixed breed cat was presented to a veterinary clinic in Iasi (Romania), for neutering, in early February 2020. During the surgery, two whitish worms were removed from the internal part of the scrotum. Two adult nematodes, one female and one male, were identified, on the basis of morphological features, as D. repens with whitish, cylindrical bodies, measuring 12.5 cm and 6.5 cm in length, respectively. At histology, the female nematode showed two cavitated structures containing myriads of variably arranged microfilariae. The male had a transversal diameter of 350 µm, a 10 µm thick cuticle and a ridge-period of 10 Μm. multiplex PCR confirmed the diagnosis of D. repens from both nematodes. The Knott's test revealed the presence of microfilariae of D. repens. Routine biochemistry panel was within range with one exception, urea serum level slightly increased. The haematology results revealed an increased number of neutrophils, lymphocytes and eosinophils. The cat had an infection with Otodectes cynotis as well. The cat was discharged with the following therapy recommended: oral doxycycline (10 mg/kg) for 30 days and topical moxidectin, monthly doses. After six months, the Knott's test gave negative results. Further studies should include new insights of D. repens infection in cats concerning its epidemiology, diagnosis and control.

Epidemiological Study of Canine Babesiosis and Hepatozoonosis in the South of Romania.

PURPOSE: The aim of this study was to evaluate the prevalence and distribution of canine babesiosis and hepatozoonosis in the south of Romania as well as to assess if there are differences in the prevalence between regions in that area. METHODS: Blood samples from 300 dogs from the south of Romania were screened for the presence of piroplasmids by PCR. Positive samples were further tested for the presence of Babesia spp. and Hepatozoon canis. Pearson's chi-square with Yates correction was used to determine if there is statistical difference between infected populations. RESULTS: Fifty-four percent (163/300) of the dogs were positive for Hepatozoon spp. or piroplasmids. Forty-eight percent (143/300) were infected with H. canis and 9.6% (29/300) were positive to B. canis. Three percent (9/300) of the dogs were co-infected with Babesia spp. and H. canis. The prevalence of H. canis in the south-west of Romania was significantly higher (p < 0.05) than that of three other regions while the prevalence of Babesia spp. infection in the south-east of the country was significantly higher (p < 0.05) compared to its south-west. CONCLUSIONS: A high rate of H. canis infection was found in the stray dog population surveyed with the highest infection rate detected in south-western Romania. Babesia canis was found to pose the highest threat in south-eastern Romania, whereas B. vogeli infection was focused in the same region of high H. canis infection in south-western Romania. These infections should be of important concern to veterinarians and dog owners in Romania.

Brainstem Auditory Evoked Potentials in Raccoon Dogs (Nyctereutes procynoides).

Raccoon dogs (Nyctereutes procynoides) are canids indigenous to Eastern Asia being one of the most invasive non-native animals in Europe and potential vectors for several hazardous parasitic and viral diseases. To present, there is a lack of studies regarding objective techniques used to appreciate the integrity and functionality of the nervous system in this species. Brainstem auditory evoked potentials (BAER) is a cost-effective, quick and noninvasive technique to assess the functionality of nervous system. The aim of the study is to provide reference values and evaluates the reliability of BAER recording with needle electrodes in clinically healthy raccoon dogs. Nine individuals were investigated for BAER under xylazine and ketamine general anesthesia. Four waves (I, II, III and V) were identified and evaluated for latencies, morphologically similarity to those previously reported for other species (dogs, horses, cats, and ferret). Our data can be used in further studies to asses objectively the auditory system function in raccoon dogs.

Seroepidemiological survey of human exposure to Dirofilaria spp. in Romania and Moldova.

The present study aimed to evaluate the extent of Dirofilaria immitis and D. repens exposure in humans from eastern and southern areas of Romania and central Moldova by serological methods. The serological screening was performed on a total of 450 serum samples (187 from Romania and 263 from Moldova). The sera were collected using a convenience sampling with the help of physicians from the hospitals of the study areas. All samples were analysed by a non-commercial ELISA test for the detection of IgG antibodies against adult somatic antigens of D. immitis and D. repens. The results showed a total of 49 (10.9%; 95% CI = 8.3-14.1%) individuals from Romania and Moldova with a positive response to IgG antibodies against both adult somatic antigens of D. immitis and D. repens. Specifically, 48 (10.7%; 95% CI = 8.0-14.0%) patients were positive for IgG-antibodies against adult somatic antigens of D. immitis, one (0.2%; 95% CI = 0.4-1.2%) against D. repens antigens, and four (0.9%; 95% CI = 0.4-3.3%). were positive for antigens of both parasites. At country level, out of 187 samples from Romania, 13 (6.9%; 95% CI = 4.1-11.5%) were positive for anti-D. immitis IgG with high exposure in the southern part of the country (Bucharest). Of the 263 people from Moldova, 36 (13.7%; 95% CI = 10.0-18.4%) were positive for D. immitis antigens from which three (1.1%, 95% CI = 0.4-3.3%) were positive for the antibodies against antigens of both parasites. Only one sample was found positive for anti-D. repens IgG. Positive IgG-ELISA results were confirmed by Western blot analysis. In addition, for further confirmation, a complementary ELISA was performed for anti-WSP IgG antibodies against Wolbachia endosymbionts. Our findings showed a noticeable exposure of humans from Romania and Moldova to Dirofilaria parasites. Serology can be useful for indicating exposure to Dirofilaria spp. in a healthy population in order to obtain useful data on the epidemiological scenario of human dirofilariosis in Eastern Europe.

Geographic distribution of canine heartworm (Dirofilaria immitis) infection in stray dogs of eastern Romania.

A survey was conducted in the eastern part of Romania to assess the prevalence and geographical distribution of Dirofilaria immitis in dogs. Plasma samples were collected from 458 stray dogs hosted in shelters in 8 counties and tested serologically for the presence of heartworm. In addition, 45 blood samples from dogs of a shelter in Galati city were examined by the modified Knott and multiplex polymerase chain reaction (PCR) techniques. The immmunological assay showed a heartworm infection prevalence of 8.9% in the dogs. Optical density results for positive samples ranged between 0.217 and 2.683. Geographical information systems (GIS) were used to produce overlays of distribution maps of D. immitis prevalence and predictive maps based on temperature suitability. High prevalence of D. immitis was found in the central East up to the northern border of the country, i.e. Galati county (60%), followed by the counties of Vaslui (12.0%) and Iasi (7.7%). Out of 45 samples examined using the Knott test, 23 were positive for circulating microfilariae (51.1%), while 19 dogs were positive for D. immitis and 4 for both D. immitis and D. repens with the multiplex PCR test. The high prevalence for D. immitis shown in dogs in the Southeast (Galati, 42.2%) also by multiplex PCR gave strong support to the results achieved by the serological tests. The present study confirms the ability of GIS to predict the distribution and epidemiology of dirofilariosis in different geographical territories as has been already demonstrated by the empirical epidemiological data obtained at the continental, national and intraregional levels.

Climate Change Is Increasing the Risk of the Reemergence of Malaria in Romania.

The climatic modifications lead to global warming; favouring the risk of the appearance and development of diseases are considered until now tropical diseases. Another important factor is the workers' immigration, the economic crisis favouring the passive transmission of new species of culicidae from different areas. Malaria is the disease with the widest distribution in the globe. Millions of people are infected every year in Africa, India, South-East Asia, Middle East, and Central and South America, with more than 41% of the global population under the risk of infestation with malaria. The increase of the number of local cases reported in 2007-2011 indicates that the conditions can favour the high local transmission in the affected areas. In the situation presented, the establishment of the level of risk concerning the reemergence of malaria in Romania becomes a priority.

PCR identification of five species from the Anopheles maculipennis complex (Diptera: Culicidae) in North-Eastern Romania.

The members of the Anopheles maculipennis complex have been incriminated for the transmission of the malaria in Europe, which was endemic until the middle of the century. The global warming and the intensification of the intercontinental travel constitute a risk of the re-emergence of the malaria in Europe, given the presence of the Anopheles vectors. The study has attempted the identification by using the PCR (Polymerase Chain Reaction) of the members of the Anopheles maculipennis complex from the North-eastern area of Romania from the city of Iasi. In total there have been identified by using the PCR amplifying the ITS2 sequence of the ribosomal DNA, 217 specimens belonging to the complex of A. maculipennis among which: 58 A. atroparvus, 18 A. melanoon, 2 A. labranchiae, 52 A. maculipennis and 87 A. messeae. The ITS2 sequences of the ribosomal DNA have been compared to those of the species belonging to the A. maculipennis available in GenBank. The Species A. labranchiae is reported for the first time in Romania, being identified in the larval stage IV. The adaptation of a new species to the climatic conditions present in the North-eastern Romania, confirms the phenomenon of global warming and also the intensification of the travelling. As a result of the analysis of the A. labranchiae sequence, this one corresponds to the extent of 96% to the species from Italy, registered in GenBank, given the fact that a high number of the inhabitants of the municipality of Iasi are working in this country.

Comparative assessment of effectiveness of ketoprofen and ketoprofen/beta-cyclodextrin complex in two experimental models of inflammation in rats.

Oral administration of non-steroidal anti-inflammatory drugs (NSAIDs) can lead to adverse effects such as gastrointestinal distress. The complexation of different groups of active substances with ß-cyclodextrin (ß-CD) has drawn considerable interest over recent years. The purpose of this study was to analyze the ketoprofen/ß-cyclodextrin (K/ß-CD) conjugate complex as well as to assess its anti-inflammatory effect after oral administration (doses of 30 mg/m(2) and 15 mg/m(2) of body surface), compared with ketoprofen. The studies were done on two models of experimentally-induced acute inflammation in rats (n = 48, 6/group), by means of intraplantar administration of a 10% aqueous kaolin suspension and intraperitoneal administration of a 1% sodium thioglycolate solution. The dynamics of the acute inflammatory process and the anti-inflammatory effects were monitored using plethysmometric determinations after 3, 6, 9, 12, 24 and 48 h (plantar inflammation), and the absorbance of the exudates (spectrophotometrically read) and nucleated cell counts after 24 h (peritoneal inflammation). The coupling of ketoprofen with ß-CD resulted in increased solubility (100% in 60 min) of the newly-formed product, which further resulted in a higher bioavailability compared with ketoprofen (<40% in 120 min). In both models of experimentally-induced inflammation, the K/ß-CD complex had a higher anti-inflammatory activity than ketoprofen.