Antimicrobial Resistance Genes in Pigeons from Public Parks in Costa Rica

Antimicrobial resistance is known to be an emerging problem, but the extent of the issue remains incomplete. The aim of this study was to determine the presence or absence of nine resistance genes (bla_TEM, catI, mecA, qnrS, sulI, sulII, tet(A), tet(Q), vanA) in the faeces of 141 pigeons from four urban parks in Alajuela, Guadalupe, Tres Ríos and San José in Costa Rica. The genes were identified by real‐time PCR directly from enema samples. About 30% of the samples were positive for genes catI and sulI; between 13% and 17% were positive for qnrS, sulII, tet(A) and tet(Q); and 4% were positive for bla_TEM. The mecA and vanA genes were not detected. The average of antimicrobial resistance genes detected per pigeon was 2. Eight different patterns of resistance were identified, without differences in the sampling areas, being the most common pattern 2 (sulII positive samples). During rainy season, the genes more frequently found were sulI and tet(A). In conclusion, the urban inhabiting pigeons tested are currently carrying antimicrobial resistance genes, potentially acting as reservoirs of resistant bacteria and vectors to humans. To the authors’ knowledge, this is the first study carried out on direct detection of resistance genes in the digestive metagenomes of pigeons.     

The effect of induced forelimb lameness on thoracolumbar kinematics during treadmill locomotion

REASONS FOR PERFORMING STUDY: Lameness has often been suggested to result in altered movement of the back, but there are no detailed studies describing such a relationship in quantitative terms. OBJECTIVES: To quantify the effect of induced subtle forelimb lameness on thoracolumbar kinematics in the horse. METHODS: Kinematics of 6 riding horses was measured at walk and at trot on a treadmill before and after the induction of reversible forelimb lameness grade 2 (AAEP scale 1-5). Ground reaction forces (GRF) for individual limbs were calculated from kinematics. RESULTS: The horses significantly unloaded the painful limb by 11.5% at trot, while unloading at walk was not significant. The overall flexion-extension range of back motion decreased on average by 0.2 degrees at walk and increased by 3.3 degrees at trot (P<0.05). Changes in angular motion patterns of vertebral joints were noted only at trot, with an increase in flexion of 0.9 degrees at T10 (i.e. angle between T6, T10 and T13) during the stance phase of the sound diagonal and an increase in extension of the thoracolumbar area during stance of the lame diagonal (0.7degrees at T13, 0.8 degres at T17, 0.5 degres at L1, 0.4 degrees at L3 and 0.3 degrees at L5) (P<0.05). Lameness further caused a lateral bending of the cranial thoracic vertebral column towards the lame side (1.3 degrees at T10 and 0.9 degrees at T13) (P<0.05) during stance of the lame diagonal. CONCLUSIONS: Both range of motion and vertebral angular motion patterns are affected by subtle forelimb lameness. At walk, the effect is minimal, at trot the horses increased the vertebral range of motion and changed the pattern of thoracolumbar motion in the sagittal and horizontal planes, presumably in an attempt to move the centre of gravity away from the lame side and reduce the force on the affected limb. POTENTIAL RELEVANCE: Subtle forelimb lameness affects thoracolumbar kinematics. Future studies should aim at elucidating whether the altered movement patterns lead to back and/or neck dysfunction in the case of chronic lameness.     

Humoral immune response in pregnant heifers inoculated with Neospora caninum tachyzoites by conjunctival route

The aim of this study was to compare systemic humoral immune responses in pregnant heifers inoculated with Neospora caninum tachyzoites by conjunctival and intravenous routes. Twenty nine heifers separated in three experimental groups were studied: Group 1 (n=10 animals) and Group 2 (n=9 animals) were inoculated with 10(8) of N. caninum tachyzoites by conjunctival and intravenous routes at 5th month of gestation, respectively; Group 3 (n=10 animals) were non-inoculated control animals. An indirect fluorescent antibody test (IFAT) and western immunoblotting (IB) were used to analyze the humoral immune response. All animals from Group 1 developed N. caninum specific antibody responses after conjunctival inoculation recording the highest antibody titer (mean+/-SE: 160+/-49.9) at 6th month of gestation. There were statistical differences between humoral immune responses found in Group 1 and 2 being higher in the second one at 6.5th, 8.5th and 9th months of gestation (P<0.05). Interestingly, all heifers from Group 1 reverted to seronegative status at the end of gestation. No increase in antibody was detected in the uninfected control group. Same pattern of N. caninum antigens was recognized by sera from heifers inoculated by conjunctival route and heifers inoculated by intravenous route. Recognized antigens were 116, 92, 84, 77, 45, 40, 25-26 and 17-18 kDa. The conjunctival instillation of N. caninum tachyzoites in pregnant heifers induces specific systemic antibodies. Further work is needed in order to clarify the consequences of this novel experimental route of infection not only on the fetus but also on the dam.     

An immunohistochemical study of the tonsils in pigs with acute African swine fever virus infection

An immunohistochemical study of the tonsils was carried out to gain further insight in the pathogenesis of acute African swine fever (ASF). Twenty-one pigs were inoculated by intramuscular route with a highly virulent isolate of ASF virus and painlessly killed at 1-7dpi. Viral antigen was highly distributed in the tonsil from 3 to 4dpi and an increase in the number of monocyte-macrophages was very evident at the same days post inoculation. This phenomenon was observed together with an increase of the expression of proinflammatory cytokines (Tumour necrosis factor alpha and Interleukin-1 alpha) and the apoptosis of lymphocytes studied by the terminal deoxynucleotidyltransferase-mediated dUTP nick end labelling (TUNEL) technique and haemorrhages. With these results, we can conclude that the tonsil is suffering similar lesions than those observed in other lymphoid organs in acute African swine fever, even when the route of inoculation is the intramuscular and not oral-nasal.     

Leishmania-specific isotype levels and their relationship with specific cell-mediated immunity parameters in canine leishmaniasis

In the current retrospective study, Leishmania infantum-specific IgG, IgA and IgM levels were determined by ELISA in 106 untreated dogs with clinically-patent leishmaniasis (Sx) and in 171 clinically healthy dogs (Asx) from Spain to investigate the relationship between these Ig isotypes and clinical status. In addition, we studied if different Leishmania-specific humoral pattern exists between Asx dogs with and without cellular mediated immunity (CMI). Fifty-six dogs were assessed by means of lymphoproliferation assay (LPA), interferon production (IFN) and leishmanin skin test (LST), 71 dogs by means of both LPA and IFN and 44 only by LST. Both Sx and Asx dogs produced Leishmania-specific IgG, IgA and IgM antibodies, however the levels and proportion of positive dogs for each Ig isotype were significantly higher in Sx than in Asx ones (P<0.001). Analysis of immunological profiles determined for each cellular technique (positive and negative cellular response for each technique combined with positive or negative specific humoral response) showed that Asx dogs constituted a high heterogeneous group. No correlations were observed between CMI tests and specific IgG or IgM levels. However, a significant inverse correlation was demonstrated between specific IgA levels and LPA response (Spearman's r=-0.220; P=0.035). In general, the low correlation detected between CMI tests and isotype levels might indicate that the immune response is not strongly polarized in the majority of Asx dogs. Additionally, this study suggests that dogs developing T-cell response are probably able to avoid the dissemination of the parasite at least to mucosal surfaces and, as a consequence, to produce low or background specific IgA levels. Further studies are needed to investigate the relationship between specific IgA and parasite load, especially at mucosal site.     

Gastrointestinal parasitism reduces the plasma availability of doramectin in lambs

A study was undertaken to investigate the effect of parasitism on plasma availability and pharmacokinetic behaviour of doramectin (DRM) in lambs. Fourteen parasitised grey face Suffolk lambs (26.9 ± 1.5 kg bodyweight) were selected for the study. Seven pairs of lambs were allocated to two groups to obtain an approximately even weight distribution. Group I (non-parasitised) was pre-treated with three repeated administrations of 5 mg/kg fenbendazole to maintain a parasite free condition. In group II (parasitised), the lambs did not receive any anthelmintic treatment. After the 85-day pre-treatment period, both groups of animals were treated with DRM by subcutaneous (SC) injection in the shoulder area at 200 μg/kg. Throughout the experimental period, both groups were maintained together under similar feeding and management conditions. Blood samples were collected by jugular venepuncture at different set times between 0.5 h and 60 days post-treatment. After plasma extraction and derivatisation, samples were analysed by high performance liquid chromatography (HPLC) with fluorescence detection. A computerised kinetic analysis was performed and the data were compared using the Student’s paired t test.The parent molecule was detected in plasma between 30 min and either day 20 (parasitised) or day 35 (non-parasitised) post-DRM treatment. The AUC values of the parasitised group (143.0 ± 18 ng d/mL) were significantly lower (P < 0.05) than those observed in the parasitically naïve animals (229.6 ± 21.7 ng d/mL). The mean residence time (MRT) in the parasitised group (3.4 ± 0.3 days) was significantly shorter (P < 0.05) than in the healthy group (6.6 ± 0.6 days). Study results have shown that parasitic disease, through alteration in the body condition, can produce significant changes in the plasma disposition of DRM when administered SC to parasitised lambs.     

Efficiency of Duddingtonia flagrans against Trichostrongyle infections of sheep on mountain pastures

The control of sheep nematode parasites in extensive mountain/transhumant management systems using the nematophagous fungus Duddingtonia flagrans was assessed in this study. Two groups of Churra Tensina ewes were allowed to graze for 8 weeks in autumn on two separate paddocks of infected pasture near their winter sheds in the valley. At lambing, ewes and their twin lambs were turned out into the same paddocks for the following 12 weeks. One group of ewes received a daily dose of 5 x 10(5) chlamydospores of Duddingtonia flagrans/kg live weight per day both in autumn and in spring, while the other group was used as a non-treated control. Daily dosing of grazing ewes with the fungus D. flagrans had a clear effect on reducing autumn pasture contamination. This had a subsequent effect on the over-wintering larvae population that was confirmed by a 20% lower worm burden of tracer lambs kept in early spring on the paddock previously grazed by fungus treated ewes. In spring, pasture contamination was also significantly reduced in the paddock grazed by fungi-treated ewes and their lambs showed a 61% lower worm burden and a better performance than the control lambs. Results herein show that fungal spores fed to sheep at critical times with regard to the epidemiology of parasite infection, can have a significant effect on the infective larvae present on pasture, which could further improve lambs performance. This novel approach to parasite control would be of interest amongst both organic and conventional sheep farmers operating in mountain regions.     

OC4Influence of Seminal Plasma Added to Highly Diluted Semen on Sperm Motility of Young Boar Feed with Selenium and Vitamin E

High dilution rates have been documented as detrimental for boar spermatozoa, shortening their lifespan (Centurion et al. 2003, Biol Reprod 69: 640–646). Addition of seminal plasma (SP) to semen extenders, or selenium (Se) and vitamin E (VE) in diet of boars could increase motility of highly diluted spermatozoa (HDS). The aim of this work was to evaluate the effect of seminal plasma on sperm motility of HDS from boars feed with Se and VE. Sixteen 12 month-old boars were designed to one of four dietary treatments: (i) control, Se 0 ppm–VE 0 IU/kg; (ii) Se 0–VE 250; (iii) Se 0.5–VE 250 and (iv) Se 0.5–VE 0. Boars were treated for 8 weeks before semen collection. Sperm rich fractions from each boar were diluted to 5 × 10⁶ sperm/ml in PBS medium and incubated at 37°C with or without 10% SP. The measurements were done at 0, 2 or 5 h. Data were analyzed as a mixed model for a factorial design [2 (Se) × 2 (VE) × 2 (SP) × 3 (h)]. Percentage of sperm motility (PSM) increases significantly (p < 0.001) with addition of Se (81.3 ± 1.52), VE (81.0 ± 1.62) and SP (81.5 ± 1.57) vs control (73.4 ± 1.61). There was significant interaction Se × VE (p < 0.001) and Se × VE × SP (p < 0.05) in PSM. However, PSM was affected significantly by time (0 h 83.4 ± 1.92; 2 h 80.7 ± 1.92 and 5 h 67.9 ± 1.92; p < 0.001). There was significant interaction SP × Time (p < 0.05) in PSM. These results indicate that Se, VE and SP improve seminal viability. Addition of 10% of SP maintains PSM at least during 5 hours.