Prevalence of Salmonella in broiler chickens in Kagoshima, Japan in 2009 to 2012 and the relationship between serovars changing and antimicrobial resistance.

BACKGROUND: This study aimed to examine the prevalence, serovars, and antimicrobial resistance of Salmonella isolates from broiler chickens in Kagoshima, Japan. A total of 192 flocks and 3071 samples were collected from broiler chickens at local farms in Kagoshima, Japan from 2009 to 2012. RESULT: Among the tested farms, 49.0% of flocks were positive for Salmonella, and 243 isolates were obtained from 3071 cecal samples (7.9%). All the Salmonella isolates were one of three serovars: S. Infantis (57.6%); (140/243), S. Manhattan (40.3%; 98/243 and S. Schwarzengrund (2.1%; 5/243). The proportion of S. Infantis isolates decreased from 66.0% in 2009 to 50.0% in 2011 but increased to 57.6% in 2012, while the proportion of S. Manhattan isolates significantly increased from 26.4 to 50% from 2009 to 2011, and decreased moderately to 40.9% in 2012. Most of the recovered Salmonella isolates were resistant to three antimicrobials, i.e., streptomycin (95.1%), sulfamethoxazole (91.0%) and oxytetracycline (91.4%). In contrast, all Salmonella strains were susceptible to chloramphenicol. Comparison of this study to previous studies of the antimicrobial susceptibility of Salmonella isolates showed that: the percentage of antibiotic-resistance isolates increased dramatically for two antibiotics, ampicillin (from 22.4 to 55.1%) and cefotaxime (from 9.1 to 52.7%). In contrast, the percentage of ofloxacin-resistant isolates decreased across the three survey periods, from 20.8% in 2004-2006 to 1.6% in the present study period (2009-2012). In addition, S. Infantis exhibited a variety of resistance to antimicrobials examined from sensitive to resistance to eight antimicrobials. Multidrug resistance to more than 6 six antimicrobials was detected in 113 (46.5%) of the isolates, and most of them were S. Manhattan. CONCLUSIONS: There was a marked change in the serovars and antimicrobial resistance profiles of the Salmonella isolates in this study compared to those in previous studies. The percentage of S. Manhattan isolates increased as did the percentages of ampicillin- and cefotaxime-resistant isolates.

Effect of dietary fermented mushroom bed on egg production in laying hens.

Egg productivity is declined by stress. It has been reported that some food supplements can improve the egg productivity due to a reduction of environmental stress. We evaluated the effect of fermented waste mushroom bed (FWMB) as a dietary additive on egg productivity. Hens were fed control food (control group, n = 100) or 3% FWMB-added food (FWMB group, n = 100) for 16 months. The number of eggs, soft-shelled eggs, and broken eggs were recorded for 15 months. We also evaluated stress-related markers (ovotransferrin, lipid peroxide, and the heterophil-to-lymphocyte ratio). The FWMB group had slightly increased egg production compared with control hens. The FWMB group produced significantly less broken and soft-shelled eggs than the control group. All stress-related markers were significantly lower in the FWMB group than in the control group. Gut flora was also affected by FWMB feeding. The increased egg production and decreased proportion of broken and soft-shelled eggs might be related to the prevention of stressful conditions by FWMB.

Effects of dietary feed supplementation with heat-treated Lactobacillus sakei HS-1 on the health status, blood parameters, and fecal microbes of Japanese Black calves before weaning.

Background and Aim: Our previous research suggested that heat-killed Lactobacillus sakei HS-1 (HK-LS HS-1) is potentially beneficial for improving intestinal microbes and reducing the number of medical treatments. This study aimed to investigate the effect of HK-LS HS-1 as a supplement in milk replacers (MRs) on clinical health during the 1-month preweaning period. Materials and Methods: Eighteen female calves were randomly assigned to either a group receiving the HK-LS HS-1 supplement (n = 9) or a control group without it (n = 9). We then investigated the effect of including supplementary HK-LS HS-1; 0.2% in MRs twice daily at 09:00 and 16:00 on the health, serum biochemical parameters (measured using an automated biochemical analyzer), and fecal bacteriological changes of preweaning Japanese Black calves at the day of the start of supplementation (before HK-LS HS-1 supplementation; day 0), at weaning (day 30), and at 2 weeks (day 45) and 4 weeks (day 60) after weaning. Results: During the supplementation period (0-30 days), (1) an increase (p = 0.023) was observed in albumin, and there was a tendency of increase in total cholesterol level in the HK-LS HS-1 group but not in the control group; (2) substantial differences were obtained after the weaning period (30-60 days), although no differences were observed from 0-30 days in both groups. The anti-Müllerian hormone (AMH) level was substantially increased after weaning in the control group. No differences were observed in the amounts of Coliform spp. and Staphylococcaceae spp. between the two groups; thus, HK-LS HS-1 supplementation had similar antibacterial effects. A significant reduction was observed in the time to weaning of the HK-LS HS-1 group in the field trial. Conclusion: Supplementation with HK-LS HS-1 from an early stage after birth to weaning is a cost-effective treatment to improve the growth rate of preweaning calves. However, supplementation during only preweaning periods appears to have no beneficial effects on preventing weaning stress, especially in terms of AMH levels.

The incidence of Campylobacter contamination levels through Chicken-Sashimi Processing steps in A Small-scale Poultry processing plant applying the External stripping method.

This study aimed to analyze the incidence of Campylobacter in a small-scale chicken meat processing plant producing "chicken-sashimi", and determine the effectiveness of surface burning as a treatment during processing. The most probable number (MPN) method was used to analyze the load of Campylobacter in 48 samples from four different processing steps (de-feathering, chilling, surface burning, and final-products; 12 samples each). We found the highest load of isolated bacteria in chicken skin after de-feathering. Campylobacter was not detected after the surface burning step despite a large load of bacteria present in the cecum content. Campylobacter was absent in the final products. Adequate surface burning can avoid Campylobacter contamination of chicken sashimi in the processing plant by applying the external stripping method.

Decreasing trend of ß-lactam resistance in Salmonella isolates from broiler chickens due to the cessation of ceftiofur in ovo administration.

Ceftiofur, a third-generation cephalosporin antimicrobial, was used in Japanese hatcheries for many years before 2012. We continue to study Salmonella in broilers and their antimicrobial resistance. The current study aimed to express how the cessation of ceftiofur affects the antimicrobial resistance of Salmonella isolated from broiler chickens in Kagoshima Prefecture, Japan in 2017 and 2018. A total of 274 Salmonella isolates were recovered from 1535 cecal content samples obtained from 96 broiler flocks over years 2017 and 2018. Among the S. enterica isolates, the predominant serovars were S. Manhattan (128/274, 46.7%), S. Schwarzengrund (120/274, 43.8%), and S. Infantis (26/274, 9.5%). The isolates showed a high proportion of antimicrobial resistance for oxytetracycline, sulfamethoxazole, and streptomycin. However, the ß-lactam resistance rates were significantly decreased (p < 0.01) in 2017, while no ß-lactam resistant isolates detected in 2018. The highlight of this study was the complete disappearance of ß-lactam resistance in Salmonella isolates from broiler chicken in Kagoshima, Japan.

Footpad dermatitis in broiler chickens in Japan.

The prevalence of footpad dermatitis (FPD) in broiler chickens in Japan was investigated. In the first examination at slaughterhouses, lesions were commonly observed on the footpads of a total of 8,985 broiler chickens from 45 flocks on 36 farms. In 3 flocks, all the birds examined had lesions. In the other 42 flocks, the incidence of FPD ranged from 31.9% (81/254) to 99.5% (1/222). The footpad lesions were classified into 4 categories according to the severity of dermatitis as follows; score 0, 1,181 birds (13.1%); score 1, 2,992 birds (33.3%); score 2, 3,000 birds (33.4%); and score 3, 1,812 birds (20.2%). The mean scores of the flocks varied widely from 0.31 to 2.69. Males had higher mean scores than females. No significant differences were found between the mean scores of birds reared in windowless houses and those in open-sided houses. On 4 farms, the investigation was carried out in different seasons, and 3 of them showed higher mean scores in winter than in summer. Next, observation of 2,255 birds from 15 flocks on 5 farms during a 4-week rearing period revealed that FPD was already produced at 7 days of age and worsened with age.

Toxoplasma gondii antibody prevalence and isolation in free-ranging cats in Okinawa, Japan.

Cats are an important host of Toxoplasma gondii from an epidemiological perspective because they are the only definitive hosts that excrete oocysts in their feces. In this study, 201 free-ranging cats in Okinawa were examined for T. gondii infection. Using the latex agglutination test, we detected antibodies against T. gondii in 26.9% (54/201) of the cats. Oocysts of T. gondii were not detected upon microscopic examination of the feces of 128 cats. T. gondii was isolated from the tissues of 9 out of 24 seropositive or pseudo-seropositive cats with a bioassay using laboratory mice. Genotyping for the GRA6 gene revealed that five and four of the isolates were type I and II, respectively.

Vasomotor effects of noradrenaline, 5-hydroxytryptamine, angiotensin II, bradykinin, histamine, and acetylcholine on the bat (Rhinolophus ferrumequinum) basilar artery.

The responsiveness of the basilar artery to intrinsic vasoactive substances is species-specific and can be a unique characteristic. We investigated the responsiveness of the bat (Rhinolophus ferrumequinum) basilar artery to noradrenaline (NA), 5-hydroxytryptamine (5-HT), angiotensin (Ang) II, bradykinin (BK), histamine (His), and acetylcholine (ACh). NA, 5-HT, Ang II, and BK induced contraction, whereas His and ACh induced relaxation, in a concentration-dependent manner. The NA cumulative concentration-response curve was shifted to the right in parallel with phentolamine (an α-antagonist). However, propranolol, a ß-antagonist, had no significant effect. The 5-HT curve was shifted to the right in parallel by ketanserin (a 5-HT2 antagonist) and methiothepin (a 5-HT1 and 5-HT2 antagonist). Losartan (an AT1 antagonist) shifted the Ang II curve to the right, whereas PD123319 (an AT2 antagonist) had no significant effect. L-NA, indomethacin, and des-Arg9-[Leu8]-BK (a B1 antagonist) did not significantly affect BK-induced contractions. HOE140 (a B2 antagonist) shifted the BK concentration-response curve to the right. The His curve was shifted to the right weakly by diphenhydramine (an H1 antagonist) and strongly by cimetidine (a H2 antagonist). ACh-induced relaxation was significantly inhibited by L-NA, atropine, and pFHHSiD (a muscarinic M3 antagonist), whereas pirenzepine and methoctramine (muscarinic M1 and M2 antagonists, respectively) showed no significant effects. At a resting vascular tone, L-NA-induced contraction and indomethacin induced relaxation. These results suggest that α-adrenergic, 5-HT1, 5-HT2, AT1, and B2 receptors might be important in arterial contraction, whereas M3 and H2 (>H1) receptors might modify these contractions, inducing relaxation.

Salmonella isolated from the feces of migrating cranes at the Izumi Plain (2002-2008): serotype, antibiotic sensitivity and PFGE type.

From November 2002 to February 2008, 2,251 crane feces were collected at the Izumi Plain in Kagoshima Prefecture. Salmonella enterica was isolated from 359 feces (15.9%), of which 332 (92.5%) were Salmonella Typhimurium (ST), 9 were S. Hvittingfoss/II, 4 were S. Abaetetuba, 3 were S. Enteritidis, 2 were S. Konstanz, 1 was S. Pakistan and 8 were untyped isolates, respectively. Against 12 antimicrobial agents, no resistant strains were found in 154 isolates examined, but one was found to be resistant to ampicillin. By pulsed-field gel electrophoresis (PFGE), all but one of the 68 ST isolates tested showed indistinguishable banding patterns; one had a different pattern. The results suggest that ST strains from the same origin would spread in crane flocks during their stay at Izumi Plain every winter.

Fracture status of wild cranes (Grus monacha and G. vipio) found dead or in a weak condition at Izumi Plain in Japan.

The Izumi Plain in Kagoshima Prefecture, Japan, is a major wintering ground for wild cranes. Between October 2009 and March 2016, a total of 204 Hooded Cranes Grus monacha and White-naped Cranes G. vipio were found dead or in a weak condition in the plane. Of these, 56 cranes had bone fractures. The rate of incidence of fracture was considered to be higher in White-naped than in Hooded Cranes. Tibia fractures were the most common. The leg and wing fracture numbers were almost equal. Forty six percent of the fracture cases were believed to be caused by collisions with the power line.

Vasomotor effects of 5-hydroxytryptamine, histamine, angiotensin II, acetylcholine, noradrenaline, and bradykinin on the cerebral artery of bottlenose dolphin (Tursiops truncatus).

From an evolutionary aspect, dolphins share a very close phylogenetic relationship with pigs. Previously, we characterized porcine cerebral artery responsiveness to intrinsic vasoactive substances. Therefore, here, we investigated dolphin (Tursiops truncatus) cerebral artery responsiveness to 5-hydroxytryptamine (5-HT), histamine (His), angiotensin (Ang) II, acetylcholine (ACh), noradrenaline (NA), and bradykinin (BK) to characterize their related receptor subtypes. We also compared dolphin cerebral artery responsiveness with porcine cerebral artery responsiveness. We found that 5-HT and His induced concentration-dependent contraction of the dolphin cerebral artery. Ketanserin (a 5-HT2 antagonist) and methiothepin (a 5-HT1 and 5-HT2 antagonist) shifted the concentration-response curve for 5-HT to the right. Although diphenhydramine (an H1 antagonist) shifted the concentration-response curve for His to the right, cimetidine (an H2 antagonist) had no such effect. Ang II and ACh did not produce any vasomotor actions. NA induced concentration-dependent relaxation. Propranolol (a ß antagonist) shifted the concentration-response curve for NA to the right, whereas phentolamine (an α antagonist) had no significant effect. BK induced relaxation followed by contraction in pre-contracted arteries with intact endothelium. HOE140 (a B2 antagonist) shifted the concentration-response curve for BK to the right, whereas des-Arg9-[Leu8]-BK (a B1 antagonist) had no significant effect. These results suggest that 5-HT1, 5-HT2, and H1 receptor subtypes are important in arterial contraction and that ß and B2 receptor subtypes modify these contractions to relaxations. The responsiveness of the dolphin cerebral artery is very similar to that of porcine cerebral artery, supporting their evolutionary linkage.

Increased Salmonella Schwarzengrund prevalence and antimicrobial susceptibility of Salmonella enterica isolated from broiler chickens in Kagoshima Prefecture in Japan between 2013 and 2016.

This study aimed to analyze the Salmonella serovars, measure the minimum inhibitory concentration of antimicrobials, and examine the antimicrobial resistance genes of Salmonella isolated from 192 broiler flocks in Kagoshima Prefecture in Japan, from 2013 to 2016. We found that all Salmonella isolates belonged to three serovars: Salmonella Manhattan, S. Infantis, and S. Schwarzengrund. Among them, S. Schwarzengrund prevalence has recently increased annually making the main serovar. Most recovered isolates were highly resistant to streptomycin, sulfamethoxazole, and oxytetracycline. We saw the reduction of third-generation cephalosporin resistance and identified the reason of increased kanamycin resistance to be the increased number of S. Schwazengrund isolates. Among the kanamycin-resistant Salmonella isolates, aphA1 constituted the main resistance gene detected.

Effects of dietary feed supplementation of heat-treated Lactobacillus sakei HS-1 on the health status, blood parameters, and fecal microbes of Japanese Black calves.

This study investigated the effect of heat-killed Lactobacillus sakei HS-1 (HK-LS HS-1) on the health and fecal bacteriological change of suckling Japanese Black calves as a supplement in milk replacers. Twelve calves were separated from dams to calf-hatch after calving for milk replacers feeding. They were randomly assigned to an HK-LS HS-1 supplement or a control without HK-LS HS-1 group in milk replacers. HK-LS HS-1 was administered from separation day to 3 weeks. Blood and fecal samples were examined. Two calves with a haptoglobin concentration of >500 µg/ml on day 0 were excluded from the experiment, and 10 calves were finally included. Glucose and vitamin A levels on day 7 were significantly higher (P<0.05) in the supplement group than in the control group. No significant differences were observed in haptoglobin or serum amyloid A between the groups. The number of Escherichia coli in feces was lower in the control group than in the supplement group on day 21 (P=0.06). No difference was observed in the number of bifidobacteria, but that of lactic acid bacteria was significantly higher (P<0.05) in the supplement group on day 21. The number of medications administered was significantly lower (P<0.05) in the supplement group (5.2 ± 3.9) than in the control group (10.6 ± 5.9) during the experimental period. The results indicated that HK-LS HS-1 is potentially beneficial for improving intestinal microbes and reducing the number of medical treatments.

Surveillance of antibiotic resistance in Escherichia coli isolated from wild cranes on the Izumi plain in Kagoshima prefecture, Japan.

The prevalence of antibiotic resistance in 376 Escherichia coli (E. coli) isolates from fecal samples of Hooded and White-naped cranes was investigated on the Izumi plain in Kagoshima prefecture, Japan, during winter 2016 and 2017. Resistance to oxytetracycline, ampicillin, and nalidixic acid were observed in 10.9%, 3.1-4.4%, and 2.1-7.7% of isolates, respectively. Since the previous surveillance in 2007, isolation rates of antibiotic-resistant E. coli recovered from wild cranes have remained at significantly low levels compared with those in Japanese livestock. Our results indicate that surveillance of antibiotic-resistant E. coli from wild cranes wintering in the Izumi plain could be a useful strategy to indicate natural environmental pollution by antibiotic-resistant bacteria in the environment.

Characterization of gizzards and grits of wild cranes found dead at Izumi Plain in Japan.

We analyzed the gizzards, and grits retained in the gizzards of 41 cranes that migrated to the Izumi Plain during the winter of 2015/2016 and died there, either due to accident or disease. These included 31 Hooded Cranes (Grus monacha) and 10 White-naped Cranes (G. vipio). We determined body weight, gizzard weight, total grit weight and number per gizzard, and size, shape, and surface roundness of the grits. Average gizzard weights were 92.4 g for Hooded Cranes and 97.1 g for White-naped Cranes, and gizzard weight positively correlated with body weight in both species. Average total grit weights per gizzard were 19.7 g in Hooded Cranes and 25.7 g in White-naped Cranes, and were significantly higher in the latter. Average percentages of body weight to grit weight were 0.8% in Hooded Cranes and 0.5% in White-naped Cranes. Average grit number per gizzard was 693.5 in Hooded Cranes and 924.2 in White-naped Cranes, and were significantly higher in the latter. The average grit size was 2.8 mm in both species. No differences were found in the shape and surface roundness of grits between the two species. To the best of our knowledge, this is the first study on the grits retained in the gizzards of Hooded and White-naped Cranes.

Ibuprofen or ozagrel increases NO release and l-nitro arginine induces TXA(2) release from cultured porcine basilar arterial endothelial cells.

The vascular resting tone of the porcine basilar artery appears to be mostly maintained by a balance between spontaneously released nitric oxide (NO) from endothelial cells and thromboxane (TX) A(2) from endothelial and smooth muscle cells. However the precise role of the interaction between the above two substances in the control of vascular tone is unclear. We attempted to clarify the interaction between NO and TXA(2) using cultured porcine basilar arterial endothelial cells. The cultured endothelial cells produced NO spontaneously, while TXB(2) (a stable metabolite of TXA(2)) production remained below the detection limit. Ibuprofen (a COX inhibitor) and ozagrel (a TXA(2) synthetase inhibitor) significantly increased the spontaneous production of NO, which was not affected by 1400W (an iNOS inhibitor). l-Nitro arginine (a NOS inhibitor) significantly induced TXB(2) production. These results suggest that NO may inhibit COX or TXA(2) synthetase, and that therefore inhibition of NOS might disinhibit COX or TXA(2) synthetase, subsequently inducing TXA(2) production. On the other hand, as TXA(2) and other contractility-related prostaglandin(s) may inhibit NOS, therefore the inhibition of COX or TXA(2) synthetase might disinhibit NOS, and then increase the spontaneous production of NO in porcine basilar arterial endothelial cells.

Morphologically abnormal beaks observed in chickens that were beak-trimmed at young ages.

A survey of beak morphological abnormalities was performed on 6,201 chickens (egg-laying hens and chickens for meat production belonging to 25 flocks) brought to a poultry processing plant. The observed abnormalities varied among flocks with occurrence rates ranging from 0.48 to 46.67%. The occurrence was high in flocks subjected to beak trimming and varied significantly according to chicken breed, with the highest rates of abnormalities in a certain chicken breed. The most widely observed abnormalities were: 1) uneven growth of the upper and lower mandibles, mostly with elongation of the lower mandible (accounting for 64.8% of all abnormalities); 2) misalignment of the upper and lower mandibles, causing lateral deviation or crossing (16.3%); 3) sharp or jagged deformities of the mandible tips (10.1%); 4) permanent open beak, a deformity in which the beak did not close completely even when closed (5.8%); and 5) formation of tubercular swellings at the tips of the upper or lower mandibles (3.1%). This is the first report on the occurrence of beak abnormalities in beak-trimmed poultry in Japan.

Characterization of variant infectious bursal disease virus from a broiler farm in Japan using immunized sentinel chickens.

We attempted the isolation of variant infectious bursal disease (IBD) viruses by using sentinel chickens immunized with inactivated classical-type IBD vaccine. Immunized sentinel chickens with high levels of neutralizing antibodies and non-immunized sentinel chickens were raised together with broiler chickens in a commercial farm. Severe atrophy of the bursa of Fabricius was observed from the second week after cohabitation in non-immunized sentinel chickens. However, in immunized sentinel chickens and broiler chickens, atrophy was observed from the third week after cohabitation. The IBD virus (IBDV) isolated from the bursa of Fabricius of immunized sentinel chickens, designated as strain IBDV TY2, showed severe atrophy of the bursa in infected SPF chickens. Antiserum to the IBDV TY2 strain showed higher neutralizing activity to heterologous IBDV strains than did antiserum to the K strain vaccine virus. Phylogenetic analysis revealed that the nucleotide sequences encoding the hypervariable region of virus protein 2 of the IBDV TY2 strain did not cluster with the classical, variant or very virulent IBDV groups. Based on these results, we suggest that the IBDV TY2 strain may constitute a novel variant type of IBDV.

Increases in pig major acute-phase protein in wasting pigs brought to the abattoir.

Plasma proteins of wasting pigs were quantitatively and qualitatively compared with those of normal fattening pigs. Higher expression of a 120 kDa protein was observed in the plasma of wasting pigs by SDS-PAGE. This protein was identified as pig major acute-phase protein (Pig-MAP) by proteomic analysis using two-dimensional electrophoresis and matrix-assisted laser desorption/ionization time-of-flight mass spectrometry. The plasma concentration of Pig-MAP in wasting pigs was 7-fold higher than that of normal ones.

Antagonistic Effects of Gingko biloba and Sophora japonica on Cerebral Vasoconstriction in Response to Histamine, 5-Hydroxytryptamine, U46619 and Bradykinin.

The aim of this study was to evaluate, for the first time, the antagonistic effects of Gingko biloba leaf (GB) and Sophora japonica L. flower bud (SJ) extracts on cerebral vasoconstriction in response to KCl, extracellular Ca[Formula: see text], histamine, 5-hydroxytryptamine (5-HT), 9,11-dideoxy-9[Formula: see text],11[Formula: see text]-methanoepoxy prostaglandin (PG) F[Formula: see text](U46619) and bradykinin (BK), in order to explain their traditional application for diseases associated with cerebral vasospasm. Isolated porcine basilar arteries (PBA) and endothelial cells from them were used as the study materials. Neither SJ nor GB had any effect on the contractions induced by KCl and extracellular Ca[Formula: see text]. SJ significantly inhibited the contraction induced by histamine, 5-HT, U46619 and BK, whereas GB inhibited histamine-induced contraction, but had no effects on the contractions induced by 5-HT, U46619 and BK. In the presence of diphenhydramine (a H1 receptor antagonist), ketanserin (a 5-HT2 receptor antagonist) and ONO-3708 (a thromboxane (TX) A2/PG receptor antagonist), the inhibitory effects of these extracts on the contractions induced by histamine, 5-HT and U46619 were abolished. SJ significantly inhibited the contractions induced by BK and PGF[Formula: see text], but in the presence of ONO-3708 (10[Formula: see text] M) had no effect on them. BK enhanced the production of PGF[Formula: see text] from cultured PBA endothelium cells, and SJ significantly attenuated this enhancement. These results suggest that SJ and GB have a H1-antagonistic effect, and that SJ also attenuates cerebral vasoconstriction mediated via 5-HT2 and TXA2/PG receptors. These findings appear to explain why SJ has been used traditionally as a therapeutic medication for cerebral vasospasm after cerebral hemorrhage.