Pedro Fernández-Llario

Reproductive performance of the wild boar in a Mediterranean ecosystem under drought conditions

The reproductive biology of the wild boar was analyzed during a period of drought in a Mediterranean area, Doñana National Park (Spain), during 1993 and 1994. Births were highly synchronized, most of them occuring between February and April. The average litter size for litters of about 1 week of age was 3.05 piglets per birth. This figure is below most reported for other populations of the species which are mostly intrauterine litters, hence without the effect of perinatal mortality. The percentage of breeding adult females was below 17%, much smaller than that found for most Mediterranean areas. These results stress the breeding decision of females, rather than the timing of reproduction or the size of litters, as the main adjustment of reproductive effort in the wild boar under drought conditions, which may be common in fluctuating Mediterranean climates.

Environmental correlates of nest site selection by wild boarSus scrofa

Location of nests, built by free-living wild boarsSus scrofa Linnaeus, 1758, was analysed in order to assess the importance of environmental and social conditions in piglets’ survival during the first days after birth. The research was carried in a Mediterranean forest with different habitats and high density of natural predators. The results indicate that most nests were already constructed before the beginning of March, being located preferentially in areas with: (1) abundant plant cover, (2) water nearby and (3) a warmer temperature than in other places. These results suggest that female wild boar try to diminish mortality caused by natural predators by constructing their nests in places with dense cover. Water is very important because of the females’ new necessities derived from milk production. Warm nesting places might diminish deaths after birth in a species with important thermoregulation deficiencies. The distance between nests is also important because in this period the wild boar develops a territorial behaviour, possibly with the objective of establishing bonds between the mother and her offspring before returning to the familiar units made up of several females and their piglets, all of them of a similar age.

Methicillin resistant Staphylococcus aureus (MRSA) carriage in different free-living wild animal species in Spain

Methicillin-resistant Staphylococcus aureus (MRSA) is a life-threatening pathogen in humans and its presence in animals is a public health concern. The aim of this study was to measure the prevalence of MRSA in free-living wild animals. Samples from red deer (n=273), Iberian ibex (n=212), Eurasian Griffon vulture (n=40) and wild boar (n=817) taken from different areas in Spain between June 2008 and November 2011 were analyzed. Characterization of the isolates was performed by spa typing, multi-locus sequence typing (MLST) and antimicrobial susceptibility testing. A low prevalence of MRSA was found with 13 isolates obtained from 12 animals (0.89%; 95% CI: 0.46-1.56). All MRSA sequence types belonged to ST398 (t011 and t1451) and ST1 (t127). Genotypes and antimicrobial susceptibility patterns (tetracycline resistance in ST398 and clindamycin-erythromycin-tetracycline resistance in ST1) suggest that the MRSA found probably originated in livestock (ST398) or humans (ST1). This is the first report of MRSA carriers in free-living wild animals in Europe. Although our data showed that MRSA prevalence is currently low, free-living wild animals might act as reservoir and represent a potential risk for human health.

Staphylococcus aureus Carrying mecC Gene in Animals and Urban Wastewater, Spain

To the Editor: A new methicillin resistance mechanism gene, a divergent mecA homologue named mecC (formerly mecALGA251), was recently described in Staphylococcus aureus (1). Methicillin-resistant S. aureus (MRSA) isolates carrying mecC have been recovered from humans, ruminants, pets, and other animals such as rats, seals, and guinea pigs (1–3). It has been suggested that mecC-carrying MRSA isolates might not be detected by using MRSA selective media (4). For mecC-carrying S. aureus isolates, cefoxitin MICs of 4–64 mg/L have been demonstrated (1–2,4), values that would normally include susceptible isolates, according to the epidemiologic cutoff value established by the European Committee on Antibiotic Susceptibility Testing (EUCAST; www.eucast.org). mecC-carrying S. aureus isolates have been classified as heteroresistant (5), and MICs can be affected by the drug-susceptibility testing method used (1,5). These observations led us to retrospectively investigate the presence of mecC gene in a set of 361 mecA-negative S. aureus isolates collected during 2009–2012 (Table), independently of their susceptibility to cefoxitin. Isolates were recovered from healthy carriers in livestock (n = 39), from wild animals (n = 254), and from wastewater (effluents) from an urban sewage plant (n = 68). Specific amplification of the mecC gene was performed as described (6). The mecC-carrying S. aureus isolates were tested by broth microdilution using Microtiter EUST plates (Trek Diagnostic Systems, East Grinstead, UK) for susceptibility to benzylpenicillin, cefoxitin, chloramphenicol, ciprofloxacin, clindamycin, erythromycin, florfenicol, fusidic acid, gentamicin, kanamycin, linezolid, mupirocin, rifampin, sulfamethoxazole, streptomycin, quinupristin-dalfopristin, tetracycline, thiamulin, trimethoprim, and vancomycin. Additionally, susceptibility to oxacillin was determined by using microScan Gram Positive Combo panel 37 (Siemens, Erlangen, Germany). MICs were interpreted according to EUCAST epidemiologic cutoff values. Table Testing of Staphylococcus aureus isolates for presence of methicillin resistance mechanism gene mecC, Spain* mecC was detected in a total of 4 isolates from wild boar (n = 1), fallow deer (n = 2), and urban wastewater (n = 1); these isolates represent 1% of the 361 tested isolates. The 3 isolates recovered from animals were susceptible to all antimicrobial drugs tested other than β-lactams and to oxacillin (MICs 0.5–1 mg/L) but were resistant to penicillin (MICs 0.5–2 mg/L). Two of the isolates were resistant to cefoxitin (MICs 8 and 16 mg/L) and the third was susceptible (MIC 4 mg/L). The wastewater isolate was resistant to penicillin (MIC 2 mg/L) and erythromycin (MIC 16 mg/L) and susceptible to all other antimicrobial drugs tested, including cefoxitin (MIC 4 mg/L) and oxacillin (MIC ≤0.25 mg/L). Previous studies have described mecC-positive isolates as susceptible to all antimicrobial drugs tested except β-lactams (2,3), although sporadic resistance to fluoroquinolones has been found (4,7). We additionally found erythromycin resistance in 1 mecC-carrying S. aureus isolate. For the 4 mecC-carrying S. aureus isolates we detected, MICs of oxacillin were interpreted as susceptible, and 2 isolates were susceptible to cefoxitin according to EUCAST guidelines, findings that agree with previous reports (1–2,4). Thus, mecC presence is not always linked to resistance phenotypes for cefoxitin or oxacillin; such unclear findings could hinder the detection of mecC-carrying isolates. We further characterized the 4 mecC-carrying S. aureus isolates by spa typing and detection of Panton-Valentin leukocidin (PVL) toxin genes (6,8). Multilocus sequence typing (MLST) was performed according to Enright et al. (9) by using self-designed primers arc (down 5′-CGATTTGTTGTTGATTAGGTTC-3′), tpi (up 5′-CATTAGCAGATTTAGGCGTTA-3′), and yqiL (down 5′-GATTGGYTCACCTTTRCGTTG-3′). All 4 isolates were PVL negative. The 3 animal isolates were assigned to a new spa type (t11212) and to clonal complex (CC) 425 and sequence type (ST) 425 (Table). ST425 has been previously associated with mecC-carrying S. aureus isolates in cattle and humans (1–2); the animals we sampled were from a game estate and may have had contact with cattle and with urban wastewater. The wastewater isolate was assigned to spa type t843 and to a new allelic profile, ST2676, in CC130 (Table). ST2676 represents a single-locus variant of ST130 carrying a different allele for the gene aroE. MRSA isolates of CC130 have been associated with humans and animals (1–4,6). This result indicates that mecC-carrying S. aureus isolates can be found in urban wastewater, which may act as an environmental reservoir, as has been demonstrated for mecA-carrying S. aureus (10). In conclusion, we detected the methicillin resistance mechanism gene mecC in nonclinical S. aureus isolates from animals and urban wastewater in Spain. Although our data indicate that the frequency of this resistance mechanism is low, this gene appears to be expanding to new areas. Prospective studies should be performed to evaluate epidemiologic changes and to analyze the genetic lineages that carry this resistance mechanism.

Comparative pathology of the natural infections by Mycobacterium bovis and by Mycobacterium caprae in wild boar (Sus scrofa).

The potential role of wild animals in the maintenance and spread of tuberculosis (TB) infection in domestic livestock is of particular importance in countries where eradication programs have substantially reduced the incidence of bovine tuberculosis but sporadic outbreaks still occur. Mycobacterium bovis is the agent mainly isolated in wildlife in Spain, but recently, infections by Mycobacterium caprae have increased substantially. In this study, we have analysed 43 mandibular lymph nodes samples containing TB-like lesions from 43 hunted wild boar from Madrid and Extremadura (central and south-western regions of Spain). After isolation, identification and typing of Mycobacterium tuberculosis complex isolates, we found that 23 mandibular lymph nodes involved M. caprae infections and 20 M. bovis. The lesions were compared for histopathology (different granuloma stage and number of multinucleated giant cells (MNGCs)), and acid-fast bacilli (AFBs) were quantified in the Ziehl-Neelsen-stained slides. Granulomas produced by M. caprae showed more stage IV granulomas, more MNGCs and higher AFBs counts than those induced by M. bovis. In conclusion, lesions caused by M. caprae would be more prone to the excretion of bacilli, and infected animals result as a high-risk source of infection for other animals.

Spleen size variations and reproduction in a Mediterranean population of wild boar (Sus scrofa)

We analysed spleen size variations of free-ranging wild boars from the west-central Iberian Peninsula during the hunting season (autumn and winter) in relation to the rut, the gestation effort and the attainment of sexual maturity by males and females. Females had larger spleens than males once they reached their sexual maturity. Individuals shot in winter had larger spleens than those shot in autumn, the start of the rutting period. In contrast to other reports, we found no influence of the reproductive status of adult females on their spleen sizes. Our findings may point to an influence of sex hormones, and possibly also stress hormones, and environmental factors on spleen development, mainly in adult and subadult males.

Wild boar (Sus scrofa) harvesting using the espera hunting method: side effects and management implications

Harvesting of wildlife by man has been linked to demographic and evolutionary impacts in many populations. We investigated the sex ratio and age class structure in hunting bags of wild boar harvested by espera—nocturnal single hunt at bait—during four hunting seasons in Alentejo (Portugal). In addition, we assessed whether the hunting method is a significant predictor of the probability of harvesting an animal of a particular gender, of particular age class or of a particular combination of these two attributes. We found that the espera hunting method allows very selective harvesting regimes, and thus, it seems a highly effective population management tool. Removing a large proportion of adult males, however, may bias the population sex ratio towards females, reduce male life expectancy and raise the degree of polygyny. Our results suggest that recruitment rates are resilient to this skewed sex ratio, and possibly the higher proportion of females in the adult population may even increase productivity.

Management practices and their association with Mycobacterium tuberculosis complex prevalence in red deer populations in Southwestern Spain

Intensification of game management may increase the prevalence of tuberculosis (TB) in wildlife despite eradication programs implemented in cattle herds in the same areas. In this cross-sectional study, we investigated the association between wild game management practices and the presence of tuberculosis in red deer populations in Southwestern Spain. Five hundred and fifty-one animals were examined by necropsy to detect tuberculosis-like lesions in the main lymph nodes. Prevalence, as determined by TB-like lesions, was estimated to be 5.1% of animals, with 77% of TB-like lesions confirmed by PCR. Our results suggest that population density, in addition to factors which promote the local aggregation of animals, is factors associated with increased prevalence of TB in red deer populations. We suggest that management practices including supplementary feeding, fencing, water ponds and interaction with domestic livestock should be revised in order to prevent TB in wild deer both.

Supplemental feeding drives endoparasite infection in wild boar in Western Spain.

Wildlife population management is thought to destabilize existing host-parasite equilibriums in opposing directions, that is, it may increase parasite success or host resilience once infection takes place. This process is of special importance for species such as the wild boar (Sus scrofa) that are managed for game purposes throughout much of Europe. However, little is known about how this practices influcences either gastrointestinal or pulmonary parasitism in the wild boar. Twelve hunting estates were chosen in order to study the relationship of management measures (feeder density, wild boar abundance, the ratio of wild boar per feeder and the percentage of sclerophyllous vegetation) and host factors (age and sex) with gastrointestinal and pulmonary parasite aggregation, richness, infection probability and intensity of infection. Parasitological analyses from 300 wild boar gastrointestinal and 269 respiratory tracts were performed for this purpose. A set of general linear models with combinations of the explanatory variables was built and the model with the smallest Akaike Information Criterion was selected as the best. The feeder density increased gastrointestinal parasite traits (richness, infection probability and intensity of infection), probably due to the contamination of feeding sites with infective parasite forms. Pulmonary parasite traits, on the other hand, were only influenced by host sex and age class, and parasite aggregation was as expected for a wild population. Managers should be aware of the consequences on parasitism when implementing supplemental feeding in hunting estates.

Carriage of Staphylococcus aureus by Free-Living Wild Animals in Spain

ABSTRACT The presence of methicillin-susceptible Staphylococcus aureus (MSSA) was analyzed in different free-living wild animals to assess the genetic diversity and predominant genotypes on each animal species. Samples were taken from the skin and/or nares, and isolates were characterized by spa typing, multilocus sequence typing (MLST) and antimicrobial susceptibility testing. The proportion of MSSA carriers were 5.00, 22.93, 19.78, and 17.67% in Eurasian griffon vulture, Iberian ibex, red deer, and wild boar, respectively (P = 0.057). A higher proportion of isolates (P = 0.000) were recovered from nasal samples (78.51%) than skin samples (21.49%), but the 9.26% of red deer and 18.25% of wild boar would have been undetected if only nasal samples had been tested. Sixty-three different spa types were identified, including 25 new spa types. The most common were t528 (43.59%) in Iberian ibex, t548 and t11212 (15.79% and 14.04%) in red deer, and t3750 (36.11%) in wild boar. By MLST, 27 STs were detected, of which 12 had not been described previously. The most frequent were ST581 for Iberian ibex (48.72%), ST425 for red deer (29.82%), and ST2328 for wild boar (42.36%). Isolates from Eurasian griffon vulture belong to ST133. Host specificity has been observed for the most frequent spa types and STs (P = 0.000). The highest resistance percentage was found against benzylpenicillin (average, 22.2%), although most of the S. aureus isolates were susceptible to all antimicrobial tested. Basically, MSSA isolates were different from those MRSA isolates previously detected in the same animal species.