Molecular characterisation of Cryptosporidium isolates from pet reptiles

In this study, we investigated the presence of Cryptosporidium in 171 faecal samples from reptiles commonly used as pet animals. These include lizards belonging to the genera Eublepharis, Pogona, Chlamydosaurus, Hemiteconyx, Teratoscincus, Tiliqua, Iguana, and Chamaeleo, snakes of the genera Lampropeltis, Elaphe, Python, Boa and Corallus, and tortoises belonging to the genera Testudo and Kinixys. Cryptosporidium oocysts were detected by immunofluorescence using a commercially available kit and cryptosporidial DNA by amplification of a polymorphic fragment of the 18S rDNA and the HSP70 locus.Cryptosporidium was detected in 38.6% and 25.1% of the samples analysed by immunofluorescence and PCR, respectively. Molecular characterisation of the isolates confirmed that C. serpentis and C. varanii (syn. C. saurophilum) are the main species involved in infection in pet reptiles but also showed the presence of C. parvum and C. muris, as well as other species or genotypes of this parasite including the Cryptosporidium mouse genotype and Cryptosporidium tortoise genotype previously described in reptiles. In addition, a Cryptosporidium sp. was isolated from a chameleon and a python.     

Hematology and serum biochemistry comparison in wild and captive Central American river turtles (Dermatemys mawii) in Tabasco, Mexico

Hematological and serum biochemistry analyses were determined on 51 Central American river turtles (Dermatemys mawii) during the dry and rainy seasons of 2006. Turtles came from two sites: Pantanos de Centla Biosphere Reserve and a turtle breeding farm, both located in Tabasco State, Mexico. Physical examination and body measures of animals were performed. Incidence and prevalence of hemoparasites were explored.Captive organisms were in poor physical condition while wild turtles were apparently healthy. There were differences in several hematological parameters related with the condition and the season. During the dry season captive turtles exhibited higher levels of uric acid and urea, as well as lower levels of glucose. Haemogregarina sp. was detected in 100% of the wild individuals, but not in captive individuals. Its incidence was greater during the rainy season. This is the first health assessment and hematology study of this critically endangered species.     

Contamination of Salmonella spp. in a pig finishing herd, from the arrival of the animals to the slaughterhouse

The aim of the present study was to investigate the contamination sources and the transmission of Salmonella within a pig finishing herd in Italy. Nine sets of samples were collected during the fattening period from cleaned and disinfected pens, animals at different ages (4 days after arrival, 90, 150, 170 and 240 days of age) and at slaughter. Salmonella was isolated from cleaned pens, individual faecal samples, the truck used to transport the pigs to the abattoir and after slaughter (cecal contents, mesenteric lymph nodes and carcasses). Several serovars were isolated: Salmonella typhimurium and Salmonella derby on farm; Salmonella bovismorbificans, Salmonella bredeney, Salmonella blockley, Salmonella hadar and Salmonella corvallis from the truck; S. derby, S. hadar, S. bredeney, S. bovismorbificans and Salmonella infantis at slaughter. Antibiotic resistance of the strains was tested and PFGE was carried out to investigate the on-farm epidemiology of Salmonella. The results showed that the environmental contamination may have represented a major source of infection for the pigs both on farm and during transport to the abattoir.     

Identification of Salmonella spp. isolates from chicken abattoirs by multiplex-PCR

The present study was carried out to report the occurrence Salmonella spp., Salmonella Enteritidis, and Salmonella Typhimurium in chicken abattoirs. Samples of feces; feathers; scald, evisceration, and chiller water; and rinse water of non-eviscerated, eviscerated, and chilled carcass were collected from six chicken abattoirs. Salmonella isolates were identified by a multiplex-PCR using three sets of primers targeting the invA, pefA, and sefA gene sequences from Salmonella spp., S. Typhimurium and S. Enteritidis, respectively. Salmonella spp. was detected in 10% (29/288) of the samples, whereas serovars Enteritidis and Typhimurium were identified in 62% (7/288), respectively. The results indicate the need to improve hygiene and sanitary standards in poultry slaughter lines, besides the education of food handlers and information to consumers.     

Interaction between the effects of evaporation rate and amount of simulated rainfall on development of the free-living stages of Haemonchus contortus

A factorial experiment (3 x 4 x 2 x 3) was conducted in programmable incubators to investigate interaction between the effects of rainfall amount, rainfall distribution and evaporation rate on development of Haemonchus contortus to L3. Sheep faeces containing H. contortus eggs were incubated on sterilised soil under variable temperatures typical of summer in the Northern Tablelands of NSW, Australia. Simulated rainfall was applied in 1 of 3 amounts (12, 24 or 32 mm) and 4 distributions (a single event on the day after deposition, or the same total amount split in 2, 3 or 4 equal events over 2, 3 or 4 days, respectively). Samples were incubated at either a Low or High rate of evaporation (Low: 2.1-3.4 mm/day and High: 3.8-6.1 mm/day), and faeces and soil were destructively sampled at 4, 7 and 14 days post-deposition. Recovery of L3 from the soil (extra-pellet L3) increased over time (up to 0.52% at day 14) and with each increment of rainfall (12 mm: <0.01%; 24 mm: 0.10%; 32 mm: 0.45%) but was reduced under the High evaporation rate (0.01%) compared with the Low evaporation rate (0.31%). All rainfall amounts yielded significantly different recoveries of L3 under Low evaporation rates but there was no difference between the 12 and 24 mm treatments under the High evaporation rate. The distribution of simulated rainfall did not significantly affect recovery of infective larvae. Faecal moisture content was positively associated with L3 recovery, as was the ratio of cumulative precipitation and cumulative evaporation (P/E), particularly when measured in the first 4 days post-deposition. The results show that evaporation rate plays a significant role in regulating the influence of rainfall amount on the success of L3 transmission.     

Simulation model estimates of test accuracy and predictive values for the Danish Salmonella surveillance program in dairy herds

The Danish government and cattle industry instituted a Salmonella surveillance program in October 2002 to help reduce Salmonella enterica subsp. enterica serotype Dublin (S. Dublin) infections. All dairy herds are tested by measuring antibodies in bulk tank milk at 3-month intervals. The program is based on a well-established ELISA, but the overall test program accuracy and misclassification was not previously investigated. We developed a model to simulate repeated bulk tank milk antibody measurements for dairy herds conditional on true infection status. The distributions of bulk tank milk antibody measurements for infected and noninfected herds were determined from field study data. Herd infection was defined as having either >or=1 Salmonella culture-positive fecal sample or >or=5% within-herd prevalence based on antibody measurements in serum or milk from individual animals. No distinction was made between Dublin and other Salmonella serotypes which cross-react in the ELISA. The simulation model was used to estimate the accuracy of herd classification for true herd-level prevalence values ranging from 0.02 to 0.5. Test program sensitivity was 0.95 across the range of prevalence values evaluated. Specificity was inversely related to prevalence and ranged from 0.83 to 0.98. For a true herd-level infection prevalence of 15%, the estimate for specificity (Sp) was 0.96. Also at the 15% herd-level prevalence, approximately 99% of herds classified as negative in the program would be truly noninfected and 80% of herds classified as positive would be infected. The predictive values were consistent with the primary goal of the surveillance program which was to have confidence that herds classified negative would be free of Salmonella infection.     

Salmonella Prevalence Among Reptiles in a Zoo Education Setting

Clinically healthy reptiles may shed Salmonella and therefore act as a potential zoonotic threat. Most people in Northern European countries are rarely exposed to reptiles, but many zoos have education departments where children have direct contact with this group of animals. The objectives of this study were to determine the prevalence and serotype distribution of Salmonella among reptiles in the Education Department (n = 55) at Copenhagen Zoo and compare it to the Zoo’s main reptile collection (n = 145) to evaluate the zoonotic risk. Salmonella was isolated from cloacal swabs by selective enrichment, and a single isolate from each positive sample was further identified by biochemical tests and serotyped. The overall prevalence was 35% (69/200) with significant difference between the Education Department (64%, 35/55) and the main reptile collection (23%, 34/145). A total of 28 serotypes were detected. Ten serotypes were isolated from more than one specimen and four from more than one species. Salmonella enterica subsp. enterica serovar Eastbourne was the predominant serotype (32%, 22/69) and was also the serotype isolated from most reptile species (n = 7). Transmission of serotypes from one department to another was very limited indicated by the serotype distribution. Despite the relative high prevalence observed among the reptiles in the Zoo’s Education Department compared to the reptiles in the Zoo’s main reptile collection, no Salmonella cases have been linked to the Zoo, and Salmonella ser. Eastbourne is very rarely isolated from humans in Denmark. Simple hygienic procedures such as hand washing which is consistently carried out following handling of reptiles at the Education Department may reduce the risk and therefore contribute to this low prevalence.     

四川省动物性食品源沙门氏菌的耐药性监测与分析

近年食源致病菌耐药性较严重,为了解四川省动物性食品源沙门氏菌（Salmonella spp.）耐药情况,本试验采用肉汤微量稀释法,对源自四川省动物性食品的76株沙门氏菌进行了25种抗生素(组合)的敏感性试验。结果表明76株沙门氏菌对利福平（96.05%）、黏菌素（78.95%）、磺胺异噁唑（64.47%）、先锋唑啉（60.53%）、氨苄西林（60.53%）、阿莫西林—克拉维酸（56.58%）、甲氧苄啶（53.95%）及强力霉素（52.63%）耐药率较高。76株沙门氏菌对25种抗生素（组合）共产生了66种耐药谱,多重耐药率达97.37%（74/76）,主要对青霉素类、先锋唑啉、链霉素、四环素类、磺胺类药物及黏菌素表现出多重耐药。结果表明,四川地区食源沙门氏菌耐药性已较为严重,应加强连续监测。     

Phenotypic and genetic characterization of Salmonella enterica subsp. enterica serovar Typhimurium isolated from pigs in Rio Grande do Sul, Brazil

This study aimed to investigate the relatedness of porcine Salmonella enterica subsp. enterica (S.) serovar Typhimurium strains isolated in Southern Brazil. Sixty-six isolates from pigs belonging to three commercial companies were submitted to phage typing, XbaI-macrorestriction (PFGE), IS200 hybridization, rep-PCR, antimicrobial susceptibility testing, and PCR assay targeting the spvR region. All strains presented a unique rep-PCR pattern and 63 strains had a common IS200 profile. One pulse-type (XA) was the most prevalent (39/66 strains) and included strains of phage types DT177, DT192, DT194 and RDNC. The spvR region was detected in three strains, which harboured plasmids of 90 kb. High rates of tetracycline, sulfonamide and streptomycin resistance were found. Isolates from farms located in different geographic regions but associated to the same commercial companies clustered together and presented a common resistance profile. Results suggested that clonal groups of S. Typhimurium are present in pig commercial companies in Southern Brazil.     

Culling decisions of dairy farmers during a 3-year Salmonella control study

Salmonella enterica subsp. enterica-serotypes lead to periodically increased morbidity and mortality in cattle herds. The bacteria can also lead to serious infections in humans. Consequently, Denmark has started a surveillance and control programme in 2002. The programme focuses on Salmonella Dublin which is the most prevalent and most persistent serotype in the Danish cattle population. A field study in 10 dairy herds with persistent Salmonella infections was carried out over three years to gain experience with control procedures including risk assessment, targeted control actions and test-and-cull procedures. From autumn 2003 until end of 2006 quarterly milk quality control samples from all lactating cows and biannual blood samples from all young stock above the age of three months were tested using an indirect antibody ELISA. The most recent and previous test results were used to categorise all animals into risk groups. These risk groups and all individual ELISA-results were communicated to the farmers as colour-coded lists four to six times per year. Farmers were advised to manage the risk of Salmonella transmission from cattle with repeatedly high ELISA results (flagged as "red") or cows with at least one recent moderately high ELISA result (flagged as "yellow") on the lists. Risk management included, e.g. culling or separation of the cows at calving. We analysed culling decisions using two models. For heifers a hierarchical multivariable logistic model with herd as random effect evaluated if animals with red and yellow flags had higher probability of being slaughtered or sold before first calving than animals without any risk flags. For adult cows a semi-parametric proportional hazard survival model was used to test the effect of number of red and yellow flags on hazards of culling at different time points and interactions with prevalence in the herd while accounting for parity, stage of lactation, milk yield, somatic cell count and the hierarchical structure of the data with animals clustered at herd level. This study illustrates how investigation of culling decisions made by herd managers when they have access to test-status of individual animals and overall apparent prevalence during control of an infection can lead to useful new knowledge. Overall herd managers were more likely to cull cattle with increasing number of yellow and red flags than animals with no flags. However, cattle were more likely to be culled with yellow and red flags during times with low or medium high within-herd seroprevalence than at times with high seroprevalence. These results are valuable knowledge for modelling and planning of control strategies and for making recommendations to farmers about control options.     

Prevalence of intestinal parasites and genotyping of Giardia intestinalis in pet shop puppies in east Japan

The current study examined the prevalence of intestinal parasites and genotypes of Giardia intestinalis in puppies from nine pet shops in east Japan. Fresh fecal samples from 1794 puppies (≦3 months old) were collected on one occasion. Giardia spp. was examined for specific coproantigen using ELISA kit (SNAP^®Giardia, IDEXX Laboratories, Inc., USA). Other intestinal parasites were detected microscopically using the formalin-ethyl acetate sedimentation technique. Genotyping was determined for the random 29 stool samples identified as Giardia spp. positive using PCR and direct sequencing of the glutamate dehydrogenase (gdh) gene. Overall prevalence of protozoan Giardia spp. and Cystoisospora spp. revealed 23.4% and 11.3%, respectively. Prevalence of ascarids, Strongyloides spp. and hookworms were recorded 1.8%, 1.1% and 0.1%, respectively. Protozoan Giardia spp. and Cystoisospora spp., thus, represent important pathogens among pet shop puppies. All genotyped G. intestinalis isolates were belonged to assemblage C or D, identified as dog-specific genotypes. Zoonotic assemblage A and B were not demonstrated. The result suggests that the risk of zoonotic transmission of G. intestinalis from pet shops puppies to humans may be quite low in Japan.     

Antimicrobial resistance of Salmonella enterica isolates from apparently healthy and clinically ill finishing pigs in Spain

This study was the first conducted in Spain to evaluate the occurrence of antimicrobial resistance and multi-resistance in Salmonella isolates recovered from finishing pigs from Spanish swine farms distributed over the whole country. For this purpose, 290 Salmonella isolates recovered from apparently healthy finishing pigs in a farm-based cross-sectional study and 192 Salmonella isolates recovered from faecal samples of finishing pigs suffering from diarrhoea were investigated. Resistance to a panel of 17 antimicrobials was determined using a broth microdilution technique. Resistance was a common finding and was detected in 90.3% of the Salmonella isolates from apparently healthy finishing pigs and 95.3% of the Salmonella isolates from clinically diseased finishing pigs. Resistance was particularly high among isolates of serogroup B and serovars Typhimurium and its monophasic variant S. 4,5,12:i:-. Higher frequencies of resistance were found to tetracycline, sulphamethoxazole, streptomycin, spectinomycin, ampicillin, chloramphenicol and trimethoprim-sulphamethoxazole. Less than 10% of the isolates were resistant to amoxicillin/clavulanic acid, neomycin, cephalotin, apramycin and gentamicin. Resistance to ciprofloxacin, colistin and ceftiofur was rare (under 1%). Multi-resistance, defined as resistance to four or more drugs, was detected in more than 50% of the isolates. Although multi-resistance was particularly frequent among isolates of S. Typhimurium, it was also high among other serovars as Bredeney and the S. Typhimurium monophasic variant. 4,5,12:i:-.     

Salmonella enterica isolates from faeces of domestic reptiles and a study of their antimicrobial in vitro sensitivity

From October 2001 to February 2002, the faecal samples of 305 reptiles (165 saurians, 99 ophidians and 41 chelonians) were bacteriologically examined to detect Salmonella enterica. S. enterica was isolated from 73 (23.93%) faecal samples including 44 (60.27%) samples collected from saurians, 15 (20.55%) from chelonians and 14 (19.18%) from ophidians; considering the number of samples taken for each reptile group, S. enterica was isolated from the 36.58% of chelonians, 26.66% of saurians and 14.14% of ophidians. The isolates were distributed among 38 serotypes. Sixty-nine (94.52%) isolates were resistant to erythromycin. About one-third of the isolates was resistant to sulfisoxazole (35.61%), gentamycin (32.88%), amoxycillin (31.51%) and ampicillin (27.40%). All but one of the isolates were sensitive to chloramphenicol. A high percentages of isolates were sensitive to enrofloxacin (84.93%), nitrofurantoin (80.82%), trimethoprim (76.71%) and tetracycline (75.34%).     

Prevalence and Antimicrobial Resistance Pattern of Salmonella Isolates from Apparently Healthy Slaughtered Cattle in Ethiopia

The prevalence and antimicrobial resistance pattern of Salmonella isolates was determined from apparently healthy slaughtered cattle at Debre Zeit (Ethiopia). A total of 323 cattle were examined for the presence of Salmonella in faeces, mesenteric lymph nodes, abdominal and diaphragmatic muscles. Salmonellae were cultured from 23 (7.1%) of the animals. Salmonellae were isolated from 2 (3.1%) and 3 (4.5%) of 65 pooled faecal and mesenteric lymph node samples, respectively. Nine (2.8%) abdominal muscle and 10 (3.1%) diaphragmatic muscle samples (n = 323 of each) were contaminated by Salmonella. About 60% of the serovars identified in the abdominal and diaphragmatic muscles were also detected from faeces and mesenteric lymph node samples. The five different serovars isolated were Salmonella mishmarhaemek (48%), S. typhimurium (20%), S. enteritidis (12%), S. guildford (12%) and S. dublin (48%). The antimicrobial resistance profiles of 25 of the Salmonella isolates with 17 antimicrobials showed that 52% (13/25) of them were resistant to three or more antimicrobials. Both strains of Salmonella (S. mishmarhaemek and S. typhimurium) showed multiple resistance to ampicillin, sulfamethoxazole and ticarcillin. This revised version was published online in July 2006 with corrections to the Cover Date.     

Antimicrobial resistance pattern of Salmonella serotypes isolated from food items and personnel in Addis Ababa, Ethiopia

Antimicrobial susceptibility test of 98 isolates of Salmonella was assayed from September 2003 to February 2004 using the guidelines of the National Committee for Clinical Laboratory Standards (NCCLS).The result revealed that 32.7% of Salmonella isolates were resistant to one or more of the 24 antimicrobials tested. Generally resistance for 13 different antimicrobial drugs was recognized. The most common resistance was to streptomycin (24/32, 75%), ampicillin (19/32, 59.4%), tetracycline (15/32, 46.9%), spectinomycin (13/32, 40.6%) and sulfisoxazole (13/32, 40.6%). All the three Salmonella Kentucky isolates showed resistance to at least 8 antimicrobials. Out of the 12 Salmonella Braenderup isolates, 10 (83.3%) showed multidrug resistance to ampicillin, spectinomycin, streptomycin, sulfisoxazole, sulfamethoxazole/trimethoprim, amoxicillin/clavulanic acid and trimethoprim. Among the 8 S. Hadar isolates 7 (86.5%) showed antimicrobial resistance. All the 6 S. Dublin isolates were resistant to carbadox (100%). All the 6 S. Haifa isolates were resistant for at least ampicillin, streptomycin and tetracycline. Up to ten different antimicrobial resistances pattern was observed. Multiple antimicrobial drug resistance was observed in 23 Salmonella isolates (23.5%). The level of antimicrobial resistance was significantly higher for isolates from chicken carcass (18/29, 62.1%) and pork isolates (5/22, 22.7%) (p = 0.003). The findings of the present study ascertain that significant proportion Salmonella isolates have developed resistance for routinely prescribed antimicrobial drugs and poses considerable health hazards to the consumers unless prudent control measures are instituted.     

A multiplex real-time PCR for differential detection and quantification of Salmonella spp., Salmonella enterica serovar Typhimurium and Enteritidis in meats

Salmonella (S.) Typhimurium and S. Enteritidis are the major causative agents of food-borne illnesses worldwide. Currently, a rapid detection system using multiplex real-time polymerase chain reaction (PCR) has been applied for other food-borne pathogens such as Escherichia coli, Staphylococcus aureus and Streptococcus spp. A multiplex real-time PCR was developed for the simultaneous detection of Salmonella spp., especially S. Typhimurium and S. Enteritidis, in beef and pork. For the specific and sensitive multiplex real-time PCR, three representative primers and probes were designed based on sequence data from Genbank. Among the three DNA extraction methods (boiling, alkaline lysis, and QIAamp DNA Mini Kit), the QIAamp DNA Mini Kit was the most sensitive in this study. The optimized multiplex real-time PCR was applied to artificially inoculated beef or pork. The detection sensitivity of the multiplex real-time PCR was increased. The specificity of the multiplex real-time PCR assay, using 128 pure-cultured bacteria including 110 Salmonella isolates and 18 non-Salmonella isolates, was 100%, 100% and 99.1% for Salmonella spp., S. Typhimurium and S. Enteritidis, respectively. The sensitivity was 100%, 100% and 91.7% for Salmonella spp., S. Typhimurium and S. Enteritidis, respectively. The multiplex real-time PCR assay developed in this study could detect up to 0.54 ± 0.09 and 0.65 ± 0.07 log₁₀ CFU/ml for S. Typhimurium and S. Enteritidis for beef, 1.45 ± 0.21 and 1.65 ± 0.07 log₁₀ CFU/ml for S. Typhimurium and S. Enteritidis for pork, respectively, with all conditions optimized. Our results indicated that the multiplex real-time PCR assay developed in this study could sensitively detect Salmonella spp. and specifically differentiate S. Typhimurium from S. Enteritidis in meats.     

A survey of Salmonella spp and Campylobacter spp in dairy goat faeces and bulk tank milk in the Murcia region of Spain

This study was designed to investigate the occurrence of Salmonella spp and Campylobacter spp in faeces samples from 222 healthy Murciano-Granadina dairy goats reared on 12 farms in Spain and in samples of bulk tank milk from 11 of those herds. Neither Salmonella spp nor Campylobacter spp were isolated from any of the samples. Our results suggest that, under the management practices applied to this breed in Spain, Murciano-Granadina goats are not likely to be a significant reservoir for these food-borne pathogens.     

动物源性沙门氏菌的耐药性分析及氟苯尼考类耐药基因的鉴定

为了解动物沙门氏菌的流行情况和药物敏感性及氟苯尼考耐药株的耐药基因分布,本试验对临床上疑似患沙门氏菌病的病料进行病原分离和细菌的多重PCR鉴定;采用K-B法测定分离株对23种抗菌药物的敏感性;选择氟苯尼考耐药菌株扩增floR、fexA、fexB、cfr和pexA基因。结果显示,共鉴定出61株沙门氏菌,其中肠炎沙门氏菌10株,鸡白痢沙门氏菌12株,鼠伤寒沙门氏菌39株。所有菌株对青霉素、红霉素、万古霉素耐药,90.16%对6种及6种以上抗菌药耐药。floR基因广泛存在于鼠伤寒沙门氏菌氟苯尼考耐药菌株中(8/12,66.67%),未发现其他耐药基因。研究结果表明鼠伤寒沙门氏菌是鹅源分离株中的优势血清型;floR基因主要介导沙门氏菌对氟苯尼考耐药性,但可能还存在其他机制。     

Seroprevalence of Salmonella spp. in Wild Reindeer (Rangifer tarandus tarandus) in Iceland

Veterinary Research Communications -