Ongoing Activity of Toscana Virus Genotype A and West Nile Virus Lineage 1 Strains in Turkey: A Clinical and Field Survey

Toscana virus (TOSV), West Nile virus (WNV) and tickborne encephalitis virus (TBEV) are among major viral pathogens causing febrile disease and meningitis/encephalitis. The impact of these viruses was investigated at a referral centre in Ankara Province, Central Anatolia in 2012, where previous reports suggested virus circulation but with scarce information on clinical cases and vector activity. Serum and/or cerebrospinal fluid samples from 94 individuals were evaluated, in addition to field‐collected arthropod specimens that included 767 sandflies and 239 mosquitoes. Viral nucleic acids in clinical samples and arthropods were sought via specific and generic nested/real‐time PCRs, and antibody responses in clinical samples were investigated via commercial indirect immunofluorescence tests (IIFTs) and virus neutralization. A WNV antigen assay was also employed for mosquitoes. WNV neuroinvasive disease has been identified in a 63‐year‐old male via RNA detection, and the WNV strain was characterized as lineage 1. TOSV infections were diagnosed in six individuals (6.3%) via RNA or IgM detection. Partial sequences in a 23‐year‐old female, presented with fever and transient pancytopenia, were characterized as TOSV genotype A. Febrile disease with arthralgia and/or peripheral cranial nerve involvement was noted in cases with TOSV infections. Previous WNV and TOSV exposures have been observed in 5.3% and 2.1% of the subjects, respectively. No confirmed TBEV exposure could be identified. Morphological identification of the field‐collected mosquitoes revealed Culex pipiens sensu lato (74.4%), Anopheles maculipennis (20.9%), An. claviger (2.1%) and others. Sandfly species were determined as Phlebotomus papatasi (36.2%), P. halepensis (27.3%), P. major s. l. (19.3%), P. sergenti (8.9%), P. perfiliewi (4.4%), P. simici (2.6%) and others. Viral infections in arthropods could not be demonstrated. TOSV genotype A and WNV lineage 1 activity have been demonstrated as well as serologically proven exposure in patients. Presence of sandfly and mosquito species capable of virus transmission has also been revealed.     

Aedes vexans and Culex pipiens as the potential vectors of Dirofilaria immitis in Central Turkey

This study was carried out to investigate the potential vectors and relative mosquito infection rates of Dirofilaria immitis throughout two mosquito seasons (2008-2009) in Kayseri province where is located in Central Anatolian part of Turkey. For this aim, totally 1198 genomic DNA pools, extracted and grouped according to the species and collection site (1-17 specimens/pool) from 6153 mosquito specimens, were examined by PCR using species-specific primers for D. immitis. The captured mosquitoes from 46 focuses were survived under in vitro conditions for 7 days to allow the development of larval stages of D. immitis. DNA extraction was performed individually to both thorax-head and abdomens in order to determine infective and infected mosquito specimens, respectively. The most abundant mosquito species in the study area was determined as Ae. vexans (51.7%) and this was followed by Cx. pipiens (42.1%), Cx. theileri (3.1%), Cs. annulata (1.5%), An. maculipennis (1.0%) and Cx. hortensis (0.6%). The PCR results indicated that 9/312 and 12/312 pools from Ae. vexans abdomens and thorax-heads were positive for filarial DNAs, respectively where as 3/241 pools of each abdomens and thorax-heads from Cx. pipiens were positive for D. immitis DNAs. The minimum infection rates (MIRs) for Ae. vexans and Cx. pipiens were calculated as 0.41 and 0.12, respectively. Although D. immitis DNA's were found in both pools from Ae. vexans and Cx. pipiens, the calculated MIRs provide evidence that Ae. vexans could be the main potential vector of D. immitis in Kayseri.     

West Nile virus outbreak in captive and wild raptors,Czech Republic, 2018

West Nile virus lineage 2 (WNV‐2) was detected in the brain of 17 goshawks (Accipiter gentilis) that succumbed to neuroinvasive disease in the Czech Republic during 2018: twelve birds were captive and five wild. Furthermore, two wild sparrowhawks (Accipiter nisus) and three other captive birds of prey (golden eagle Aquila chrysaetos, hybrid saker falcon Falco cherrug × F. rusticolus and Harris's hawk Parabuteo unicinctus) also died due to WNV encephalitis. The 2018 outbreak in Czech raptors clearly reflects a new epidemiological situation and indicates an increasing risk of both raptor and human infection with WNV‐2 in the country.     

West Nile Virus in North‐Eastern Italy, 2011: Entomological and Equine IgM‐Based Surveillance to Detect Active Virus Circulation

Since 2008, West Nile Virus (WNV) has expanded its range in several Italian regions, and its yearly recurrence suggests the virus may have become endemic in some areas. In 2011, a new plan based also on the detection of IgM antibodies was implemented in the north‐eastern Italian regions of Veneto and Friuli Venezia Giulia, aiming to early detect WNV infections in areas where the virus had already circulated during the previous summers, and in adjacent zones. From July to November 2011, 1880 sera from 521 equine premises were screened by a commercial IgM capture ELISA. Mosquitoes were captured by CDC‐CO₂ traps at 61 locations in the two regions. Collected mosquitoes were identified, pooled by species/date/location and examined by real‐time RT‐PCR and sequencing. Passive surveillance was carried out on clinically affected horses and non‐migratory wild birds found dead. IgM sero‐positive equines were detected in 19 holdings, five in the area with WNV circulation (AWC) and 14 in the surveillance area (SA); 10 more horse premises tested positive to further serological controls within 4 km of the positive holdings. A total of 85 398 mosquitoes of 15 species were collected and 2732 pools examined. Five Culex pipiens pools tested positive for the presence of WNV. Passive surveillance on non‐migratory wild birds allowed detection of the virus only in one found dead collared dove (Streptopelia decaocto), of 82 birds sampled. The WNV belonged to the lineage 2, which had been isolated for the first time in Italy earlier in 2011. By the first week of October, nine human cases had been confirmed in the same area. The implementation of a protocol combining IgM screening of horses with surveillance on mosquito vectors proved to be valuable for early detecting WNV circulation.     

Mosquito collection in endemic areas of Japanese encephalitis in Hokkaido, Japan

A study was conducted during 1985 and 1986 to evaluate the roles of mosquito species as possible vectors of Japanese encephalitis (JE) virus in Hokkaido. The number of Culex tritaeniorhynchus was very low among the four pig farms where outbreaks of abortion caused by JE virus were observed in swine populations. At one farm near Sapporo, only one Cx. tritaeniorhynchus was found among a total of 510 mosquitoes collected during the survey period from July to October 1985, even when JE virus activity among sentinel pigs was revealed by seroconversion. At another farm in the south, no individuals of this mosquito species were found among 987 mosquitoes collected at the time of the outbreaks of abortion. Cx. pipiens pallens, Anopheles species, Aedes vexans nipponii, and Ae. japonicus were predominant over Cx. tritaeniorhynchus. Cx. tritaeniorhynchus, almost a solve vector species of JE virus in the southern part of Japan, is probably not a vector of the virus in Hokkaido. The collected mosquitoes (2,332 from 1985 and 1,403 from 1986) were processed for virus isolation but no JE virus was isolated. More extensive field studies are necessary to provide further information on the role of mosquito species other than Cx. tritaeniorhynchus in the transmission of JE virus in the northern limits of its range including Hokkaido.     

Detection of Salmonella enterica Serovar Montevideo and Newport in Free‐ranging Sea Turtles and Beach Sand in the Caribbean and Persistence in Sand and Seawater Microcosms

Salmonellae are Gram‐negative zoonotic bacteria that are frequently part of the normal reptilian gastrointestinal flora. The main objective of this project was to estimate the prevalence of non‐typhoidal Salmonella enterica in the nesting and foraging populations of sea turtles on St. Kitts and in sand from known nesting beaches. Results suggest a higher prevalence of Salmonella in nesting leatherback sea turtles compared with foraging green and hawksbill sea turtles. Salmonella was cultured from 2/9 and identified by molecular diagnostic methods in 3/9 leatherback sea turtle samples. Salmonella DNA was detected in one hawksbill turtle, but viable isolates were not recovered from any hawksbill sea turtles. No Salmonella was detected in green sea turtles. In samples collected from nesting beaches, Salmonella was only recovered from a single dry sand sample. All recovered isolates were positive for the wzx gene, consistent with the O:7 serogroup. Further serotyping characterized serovars Montevideo and Newport present in cloacal and sand samples. Repetitive‐element palindromic PCR (rep‐PCR) fingerprint analysis and pulsed‐field gel electrophoresis of the 2014 isolates from turtles and sand as well as archived Salmonella isolates recovered from leatherback sea turtles in 2012 and 2013, identified two distinct genotypes and four different pulsotypes, respectively. The genotyping and serotyping were directly correlated. To determine the persistence of representative strains of each serotype/genotype in these environments, laboratory‐controlled microcosm studies were performed in water and sand (dry and wet) incubated at 25 or 35°C. Isolates persisted for at least 32 days in most microcosms, although there were significant decreases in culturable bacteria in several microcosms, with the greatest reduction in dry sand incubated at 35°C. This information provides a better understanding of the epizootiology of Salmonella in free‐ranging marine reptiles and the potential public health risks associated with human interactions with these animals in the Caribbean.     

The incrimination of Aedes (stegomyia) aegypti as the vector of Dirofilaria repens in Nigeria

Six local species of culicides were identified as the common mosquitoes in Zaria, out of 15 species captured using various adult and larval collection methods. These common culicides are Culex pipiens fatigans, Anopheles gambiae grp., Mansonia africana, Culex pipiens pipiens, Aedes (stegomyia) aegypti and Aedes vittatus. They were each fed directly on a local dog naturally infected with Dirofilaria repens to evaluate their refractoriness/susceptibility to dirofilarial infection. In a number of donor-feeding trials, 39. 4% Culex pipiens fatigans; 58.9% An gambiae grp.; 60.5% Mansonia africana; 1.8% of Culex pipiens pipiens; 23.4% Ae aegypti and 3.3% of Ae vittatus successfully fed on the microfilaraemic host. Only Aedes aegypti was susceptible to the infection as all 40 (100%) Ae aegypti reaching 10-14 day post-blood meal had infective (L(3)) larvae of D. repens. The remaining five species were refractory. The microfilariae in the five non-susceptible mosquitoes were always found trapped in the blood meal in the insects midgut (stomach). These trapped microfilaria were dead by the 2nd day in the insect's midgut. However, in the susceptible Ae aegypti, the microfilariae were set free from the blood meal in the midgut and within 24h migrated to the malpighian tubules (MT) of the mosquitoes. All Ae aegypti dissected 5-7 day post-infective blood meal showed the typical quiescent sausage stage (L(2)) larvae in the malpighian tubules. At day-10 post-blood meal, relatively active infective (L(3)) larvae of D. repens were found in the MT; and by day 12-14, highly motile infective larvae had reached the insect's head and proboscis, with infective larvae occasionally oozing out during dissection through the tip of the proboscis. The rate of development of D. repens to infective larvae was faster in mosquitoes infected in July when the environmental temperature was 24.5 degrees C than those infected in November when the temperature was 22.5 degrees C. The latter were delayed for 4 days. The breeding sources of Ae aegypti, the local vector implicated were also identified. As no particular vector of this zoonotic filaria has been identified previously in Nigeria, these findings could make any control programme more focussed and easier.     

Comparison of florfenicol pharmacokinetics in Exopalaemon carinicauda at different temperatures and administration routes

Florfenicol is a broad‐spectrum antibacterial drug. Exopalaemon carinicauda is important in the prawn aquaculture industry in China. Florfenicol pharmacokinetics in E. carinicauda were studied at different temperatures and via different routes of administration to provide a scientific basis for the rational use of drugs for E. carinicauda production. At water temperatures of 22 ± 0.4°C and 28 ± 0.3°C, after intramuscular (IM) injection and oral (per ora (PO)) administration of florfenicol at 10 mg/kg body weight (BW) and 30 mg/kg BW, respectively, the florfenicol concentration in the plasma, hepatopancreas, gills, muscles, and carapace of E. carinicauda was determined by high‐performance liquid chromatography. After IM injection at different temperatures, the metabolism of florfenicol in E. carinicauda conformed to a two‐compartment open model with zero‐order absorption. After PO administration, the metabolism of florfenicol in E. carinicauda was consistent with a two‐compartment open model with first‐order absorption. Using an identical administration route but different water temperatures, the metabolism of florfenicol in E. carinicauda was quite different. Overall, florfenicol was absorbed rapidly and distributed widely in E. carinicauda, but elimination was slow and the bioavailability was not high. A low temperature and PO administration resulted in a low elimination rate.     

Characterization of the Campylobacter jejuni Population in the Barnacle Geese Reservoir

Campylobacter spp. are the most common cause of bacterial gastroenteritis worldwide and have been isolated from a wide number of different hosts and environmental sources. Waterfowl is considered a natural reservoir for this zoonotic bacterium and may act as a potential infection source for human campylobacteriosis. In this study, faecal samples from 924 barnacle geese were tested for the presence of C. jejuni and C. coli. The resulting C. jejuni and C. coli populations were characterized by multilocus sequence typing (MLST), structure analysis by BAPS and phylogenetic analysis based on full genome sequences. The prevalences of C. jejuni in barnacle geese faeces were 11.5% and 23.1% in 2011 and 2012, respectively, and only 0.2% of the samples were positive for C. coli in both years. Furthermore, a possible adaption of the clonal complexes (CCs) ST‐702 and ST‐1034 to the barnacle geese reservoir was found, as these two CCs represented the majority of the typed isolates and were repeatedly isolated from different flocks at several time‐points. Further core genome phylogenetic analysis using ClonalFrame revealed a formation of a distinct monophyletic lineage by these two CCs, suggesting a certain degree of clonality of the C. jejuni population adapted to barnacle geese. Therefore, although STs also commonly found in humans patients (e.g. ST‐45) were among the barnacle geese C. jejuni isolates, this reservoir is probably an infrequent source for human campylobacteriosis.     

Ovine sperm DNA oxidation quantification using an 8‐OHdG immunodetection assay

Our aim was to optimize 8‐hydroxy‐2′‐deoxyguanosine (8‐OHdG) immunodetection in order to detect DNA damage caused by oxidative stress that may not be detected by other DNA integrity analysis techniques, especially due to the high compaction of DNA in ruminants. Semen samples from 6 rams were cryopreserved. After thawing, samples were subjected to the DNA oxidation quantification using an 8‐OHdG immunodetection assay by flow cytometry. We have evaluated two different incubation times (30 min vs. overnight) at 4°C of the primary antibody (monoclonal anti‐8‐OHdG antibody). We have also compared the results of this technique with the sperm chromatin structure assay (SCSA^®). The analysis revealed that there were no significant differences (p > .05) between different incubation times. However, overnight incubation seems to cause more non‐specific binding of the secondary antibody. Significant differences (p < .05) between subjects and oxidation controls (8 M H₂O₂/800 μM FeSO₄?7H₂O) were evident. We can conclude that the 8‐OHdG immunodetection assay for DNA oxidation quantification of ram sperm can be performed subjecting sperm samples to a very high oxidative treatment.     

Giardia duodenalis in small animals and their owners in Germany: A pilot study

Giardia duodenalis is a relevant gastrointestinal protozoan pathogen of humans and animals. This species complex consists of eight genetically different assemblages. Assemblages A and B are pathogenic to humans and pets, thus confer zoonotic potential. The risk of zoonotic transmission has been controversially discussed. The aim of this monocentric cross‐sectional pilot study was to investigate G. duodenalis assemblages in humans and pets living in common households in Berlin/Brandenburg (Germany). Samples from dogs, cats and humans sharing the same households were screened for Giardia infection by antigen‐detecting assays. All human samples were additionally analysed by a Giardia‐specific qPCR. Cyst quantification and sequences of different gene loci (triosephosphate isomerase (tpi), glutamate dehydrogenase (gdh), β‐giardin (bg) and for dogs SSUrDNA) were analysed. A total of 38 households (31 households with dogs and seven with cats) with 69 human individuals participated in the study. Initial antigen‐detecting assays revealed Giardia‐positive results for 13 (39%) canine, one (14%) feline and one human sample. Reanalysis of the human samples by qPCR revealed two more positive specimens (4%). Two of these three samples were identified as assemblage B at all tested loci. Success rate of assemblage typing for pet samples was generally low and comprised mainly the SSUrDNA locus only. Overall, six of 13 Giardia‐positive canine samples were typable (2× A, 1× co‐infection: A and B, 1× C; 2× D). One pair of samples (dog and human) from the same household had a similar but not identical assemblage B sequence at tpi locus. Assemblage A was also detected in the dog specimen, which hampered sequence analysis. In conclusion, although exhibiting limitations due to the sample size, our study highlights the need for better and standardized typing tools to distinguish G. duodenalis strains with higher resolution in order to perform proper case–control studies for a realistic estimation of zoonotic risk.     

Two Cases of Cutaneous Diphtheria Associated with Occupational Pig Contact in Germany

In 2010, two independent cases of cutaneous diphtheria caused by toxigenic C. ulcerans were identified in Germany. Both patients had intense occupational contact with pigs. Diagnostic work‐up comprising biochemical differentiation, rpoB sequencing, matrix‐assisted laser desorption/ionization time‐of‐flight mass spectrometry (MALDI‐TOF) analysis, real‐time tox PCR and Elek test as well as public health measures including an intensified source tracing involving 83 asymptomatic pigs of an associated pig farm are presented.     

Detection of hepatitis E virus RNA in rats caught in pig farms from Northern Italy

Hepatitis E virus (HEV) strains belonging to the Orthohepevirus genus are divided into four species (A–D). HEV strains included in the Orthohepevirus A species infect humans and several other mammals. Among them, the HEV‐3 and HEV‐4 genotypes are zoonotic and infect both humans and animals, of which, pigs and wild boar are the main reservoirs. Viruses belonging to the Orthohepevirus C species (HEV‐C) have been considered to infect rats of different species and carnivores. Recently, two studies reported the detection of HEV‐C1 (rat HEV) RNA in immunocompromised and immunocompetent patients, suggesting a possible transmission of rat HEV to humans. The role of rats and mice as reservoir of HEV and the potential zoonotic transmission is still poorly known and deserves further investigation. To this purpose, in this study, the presence of HEV RNA was investigated in the intestinal contents and liver samples from 47 Black rats (Rattus rattus) and 21 House mice (Mus musculus) captured in four pig farms in Northern Italy. The presence of both Orthohepevirus A and C was investigated by the real‐rime RT‐PCR specific for HEV‐1 to HEV‐4 genotypes of Orthohepevirus A species and by a broad spectrum hemi‐nested RT‐PCR capable of detecting different HEV species including rat HEV. The intestinal content from two Black rats resulted positive for HEV‐C1 RNA and for HEV‐3 RNA, respectively. None of the House mice was HEV RNA positive. Sequence analyses confirmed the detection of HEV‐C1, genotype G1 and HEV‐3 subtype e. The viral strain HEV‐3e detected in the rat was identical to swine HEV strains detected in the same farm. Liver samples were negative for the detection of either rat HEV or HEV‐3.     

Influences of fermentation and ripening temperatures on the enzymatic activity and physicochemical and sensory properties of salted egg white sufu

This study developed a novel form of sufu. Salted egg white sufu (SEWS) was produced by fermenting steamed salted egg white with douchi koji and ripening at 25°C, 35°C, or 45°C for 19 days. The results show that the protease activity of the koji reduced pronouncedly during fermentation, whereas the one in the SEWS initially increased but subsequently decreased. The total nitrogen and amino nitrogen contents and degree of protein hydrolysis increased during fermentation and decreased during preripening. SEWS, particularly processed at 35°C, contained more free amino acids, notably glutamic acid and leucine, than steamed salted egg white did. After processing, SEWS had darker colors, particularly when manufactured at higher temperatures, and hardness and springiness decreased. The 35°C and 45°C SEWS had higher sensory acceptability. In conclusion, ripening at 35°C for 19 days is recommended for producing this novel animal protein‐based sufu.     

Comparison of Different Sampling Strategies and Laboratory Methods for the Detection of C. jejuni and C. coli from Broiler Flocks at Primary Production

Summary The objective of the study was to evaluate the performance of different combinations of sample type, transport medium and culture methods for the recovery of Campylobacter jejuni and C. coli from broiler flocks at primary production. Boot swabs moistened with one of four different transport media [maximum recovery diluent (n = 120), Exeter broth (EX) (n = 120), buffered peptone water (n = 120) and modified semi‐solid Cary‐Blair (n = 120)], caecal samples (n = 40) and faecal samples (n = 120) from 40 broiler flocks were compared and sensitivity estimates obtained using a Bayesian model. Samples were cultured onto mCCDA before and after enrichment in EX and incubated microaerobically at 41.5°C. Campylobacter suspect colonies were identified to the species level by multiplex PCR. Results from the Bayesian model indicated that boot swabs after enrichment had higher sensitivity (90–94%) than caecal contents before or after enrichment (84% and 89%, respectively) and faecal samples after enrichment (82%) for the detection of Campylobacter spp., although these differences were not statistically significant. Enrichment significantly increased the sensitivity of boot swab and caecal samples for detection of Campylobacter spp. and C. jejuni, respectively. However, the enrichment of caecal samples resulted in a significant decrease in the sensitivity of these samples for detection of C. coli. There was much greater variation in the sensitivity estimates of the methods for detecting C. coli than for C. jejuni, and the ranking of methods was different between the two species. Boot swabs gave the best sensitivity values for detection of C. jejuni, and enrichment culture of faecal samples was the most sensitive method for detection of C. coli.     

Epidemiology of Campylobacter jejuni in raccoons (Procyon lotor) on swine farms and in conservation areas in southern Ontario

Campylobacter is a leading cause of foodborne illness in humans worldwide. Sources of infection are often difficult to identify, and are, generally, poorly understood. Recent work suggests that wildlife may represent a source of Campylobacter for human infections. Using a repeated cross‐sectional study design, raccoons were trapped on five swine farms and five conservation areas in southern Ontario from 2011 to 2013. Our objectives were to: (a) assess the impact of seasonal, climatic, location, annual and raccoon demographic factors on the occurrence of Campylobacter jejuni in these animals; and (b) identify clusters of C. jejuni in space, time and space‐time using spatial scan statistics. Multi‐level multivariable logistic regression was used to examine the odds of isolating C. jejuni, with site and animal modelled as random intercepts. The following independent variables were examined: raccoon age and sex, year, location type, season, temperature and rainfall. A total of 1,096 samples were obtained from 627 raccoons; 46.3% were positive for C. jejuni. The following interactions and their main effects were significant (p < .05) and retained in the final model: season × temperature, year × rainfall, year × temperature. Based on the results from our multivariable model and spatial scan statistics, climatic variables (i.e. rainfall, temperature and season) were associated with the carriage of C. jejuni by raccoons, but the effects were not consistent, and varied by location and year. Although raccoons may pose a zoonotic risk due to their carriage of Campylobacter, further work is required to characterize the transmission and movement of this microorganism within the ecosystem.     

Distribution of Neoehrlichia mikurensis in Ixodes ricinus ticks along the coast of Norway: The western seaboard is a low‐prevalence region

Neoehrlichia mikurensis is a tick‐borne pathogen widespread among ticks and rodents in Europe and Asia. A previous study on Ixodes ricinus ticks in Norway suggested that N. mikurensis was scarce or absent on the south‐west coast of Norway, but abundant elsewhere. The aim of this study was to further investigate the prevalence and distribution of N. mikurensis along the western seaboard of Norway in comparison with more eastern and northern areas. The second aim of the study was to examine seasonal variation of the bacterium in one specific location in the south‐eastern part of Norway. Questing I. ricinus were collected from 13 locations along the coast of Norway, from Brønnøysund in Nordland County to Spjærøy in Østfold County. In total, 11,113 nymphs in 1,113 pools and 718 individual adult ticks were analysed for N. mikurensis by real‐time PCR. The mean prevalence of N. mikurensis in adult ticks was 7.9% while the estimated pooled prevalence in nymphs was 3.5%. The prevalence ranged from 0% to 25.5%, with the highest prevalence in the southernmost and the northernmost locations. The pathogen was absent, or present only at low prevalence (<5%), at eight locations, all located in the west, from 58.9°N to 64.9°N. The prevalence of N. mikurensis was significantly different between counties (p < .0001). No significant seasonal variation of N. mikurensis prevalence was observed in the period May to October 2015. Our results confirm earlier findings of a low prevalence of N. mikurensis in the western seaboard of Norway.     

Comparatively examining of the apelin‐13 levels in the Capoeta trutta (Heckel, 1843) and Cyprinus carpio (Linnaeus, 1758)

Apelin is a recently discovered peptide produced by several tissues in the various vertebrates and fish. Apelin has been suggested to have role in regulation of many diverse physiological functions including food intake, energy homoeostasis, immunity, osmoregulation and reproduction. In this study, apelin‐13 levels in the blood serum of Cyprinus carpio and Capoetta trutta were determined. Then the results were compared between two species and sexes of each species. Apelin‐13 level was analysed using the enzyme‐linked immunoassay (ELISA) kit (Rat apelin‐13 ELISA kit, catalog no: CSB‐E14367r). Apelin‐13 level in the blood serum of C. trutta was significantly higher than those of the C. carpio (p < 0.05). However, its levels were observed to be no significant difference (p > 0.05) that compared to between sexes of each species. There was a significant negative correlation (r = ?0.829, p = 0.0001) between the apelin‐13 level and body weight of C. carpio. However, no significant correlation (r = ?0.022, p = 0.924) between the apelin‐13 level and weight of C. trutta observed.     

The occurrence of Salmonella,extended‐spectrum β‐lactamase producing Escherichia coli and carbapenem resistant non‐fermenting Gram‐negative bacteria in a backyard poultry flock environment

Increase in the number of small‐scale backyard poultry flocks in the USA has substantially increased human‐to‐live poultry contact, leading to increased public health risks of the transmission of multi‐drug resistant (MDR) zoonotic and food‐borne bacteria. The objective of this study was to detect the occurrence of Salmonella and MDR Gram‐negative bacteria (GNB) in the backyard poultry flock environment. A total of 34 backyard poultry flocks in Washington State (WA) were sampled. From each flock, one composite coop sample and three drag swabs from nest floor, waterer‐feeder, and a random site with visible faecal smearing, respectively, were collected. The samples were processed for isolation of Salmonella and other fermenting and non‐fermenting GNB under ceftiofur selection. Each isolate was identified to species level using MALDI‐TOFF and tested for resistance against 16 antibiotics belonging to eight antibiotic classes. Salmonella serovar 1,4,[5],12:i:‐ was isolated from one (3%) out of 34 flocks. Additionally, a total of 133 ceftiofur resistant (Cef^R) GNB including Escherichia coli (53), Acinetobacter spp. (45), Pseudomonas spp. (22), Achromobacter spp. (8), Bordetella trematum (1), Hafnia alvei (1), Ochrobactrum intermedium (1), Raoultella ornithinolytica (1), and Stenotrophomonas maltophilia (1) were isolated. Of these, 110 (82%) isolates displayed MDR. Each flock was found positive for the presence of one or more Cef^R GNB. Several MDR E. coli (n = 15) were identified as extended‐spectrum β‐lactamase (ESBL) positive. Carbapenem resistance was detected in non‐fermenting GNB including Acinetobacter spp. (n = 20), Pseudomonas spp. (n = 11) and Stenotrophomonas maltophila (n = 1). ESBL positive E. coli and carbapenem resistant non‐fermenting GNB are widespread in the backyard poultry flock environment in WA State. These GNB are known to cause opportunistic infections, especially in immunocompromised hosts. Better understanding of the ecology and epidemiology of these GNB in the backyard poultry flock settings is needed to identify potential risks of transmission to people in proximity.