Genetic characterization of orf viruses isolated from various ruminant species of a zoo

In the present study, an outbreak of proliferative dermatitis in musk ox (Ovibos moschatus), Sichuan takin (Budorcas taxicolor tibetana) and domestic Shetland sheep (Ovis aries) in a zoo is described. Skin lesions consisted of severe, persistent, multifocal, proliferative dermatitis in musk ox, and mild, transient, focal, dermatitis in the Sichuan takin and Shetland sheep. Parapoxviruses were isolated from skin lesions, and characterized by restriction enzyme analysis and partial gene sequencing. The results of this investigation indicate that the outbreak of proliferative dermatitis was due to infection by a single parapoxvirus, which is genetically closely related to other orf virus (ORFV) strains but distant to bovine papular stomatitis virus (BPSV) and pseudocowpox virus (PCPV).     

Detection and molecular characterization of calf diarrhoea bovine coronaviruses circulating in South Korea during 2004-2005

Although the widespread occurrence of calf diarrhoea (CD) bovine coronavirus (BCoV) infections have been reported in most cattle producing countries, only the genetic differences in the BCoVs from American and Canadian isolates and/or strains have been identified and compared. Hence, it is unclear if the BCoVs circulating in the other countries have distinct genetic characteristics. The aim of this study was to determine the prevalence and genetic diversity of CD BCoVs based on the deduced amino acid (aa) sequences of the spike (S) and haemagglutinin/esterase (HE) proteins in South Korea. RT-PCR and nested PCR using the primer pairs specific to the nucleocapsid gene, BCoVs detected the BCoVs in 56 (15.6%) of 359 diarrhoeic faecal samples. Phylogenetic analysis of the entire S gene indicated that 10 Korean CD BCoV strains clustered with other Korean BCoV strains with different clinical forms but were different from the American and Canadian BCoV strains. Moreover, the phylogenetic data of the aa sequences of the HE gene revealed all the Korean CD strains to be distinct from the other Korean BCoV strains with different clinical forms. These results suggest that the Korean BCoVs cause endemic infections in diarrhoeic calves in Jeonnam province and have taken a different evolutionary pathway from the BCoVs in other countries. Moreover, the different BCoV strains are circulating in the different clinical forms in South Korea. These results also suggest that vaccines against the BCoVs can be developed with each Korean BCoV in different clinical forms.     

Detection and control of rotavirus infections in zoo animals

Fecal specimens from 15 exotic animal species, with and without diarrhea, were examined for the presence of rotavirus, bacterial enteropathogens, and intestinal parasites. A commercial enzyme-linked immunosorbent assay was used to detect antigens of rotavirus. Rotavirus was detected in the feces of 20 (57%) of 35 of the animals, which included addax (Addax nasomaculatus), nyala (Tragelaphus angasi), saiga (Saiga tatarica), white-tailed gnu (Connochaetus gnou), greater kudu (Tragelaphus strepsiceros), sitatunga (Tragelaphus spekei), Grant's gazelle (Gazella granti roosevelti), sable antelope (Hippotragus niger niger), kob (Kobus kob leucotis), pygmy marmoset (Callithrix pygmaea), bush dog (Speothos venaticus), grizzly bear (Ursus arctos horribilis), and red kangaroo (Megaleia rufa). Bacterial pathogens were found in 8 animals, 5 of which had concurrent rotavirus infections. Most (60%) of the animals with rotavirus infection were less than 2 weeks old; however, rotavirus also was detected in feces from adult animals. Although most of the cases of rotavirus infection were detected in nursery-reared animals, exhibit-reared animals also were infected with rotavirus.     

Detection of turkey enteric coronavirus by enzyme-linked immunosorbent assay and differentiation from other coronaviruses

A double-antibody ELISA for the detection of coronaviruses in intestinal contents from turkey poults with diarrhea was developed. Antibodies were raised in rabbits and guinea pigs against a Minnesota isolate of turkey enteric coronavirus (TCV) propagated in embryonating turkey eggs and were purified by density-gradient centrifugation. The specificity of antisera was confirmed by hemagglutination-inhibition and immunoelectron microscopy. Absorption of anti-TCV hyperimmune sera with egg extracts or egg ovalbumin and the use of different dilution and blocking buffers influenced the sensitivity and specificity of the ELISA. Reciprocal cross-reactivity was detected among turkey, chicken, bovine, and murine coronaviruses. Antisera to the transmissible gastroenteritis virus of swine, the rabbit enteric coronavirus, or the human coronavirus strain 299E failed to react with TCV. The TCV cross-reacted only moderately with the avian infectious bronchitis virus and the hemagglutinating encephalomyelitis virus of swine. Investigations with samples from 47 commercial turkey flocks in Quebec with episodes of transmissible enteritis revealed that the ELISA was more sensitive than was electron microscopy for detection of TCV.     

Bacterial studies on the occurrence of Mycobacterium paratuberculosis in fecal samples of zoo ruminants]

Mycobacterium (M.) paratuberculosis was isolated from fecal samples of 3 (21.4%) from 14 mouflons, of 10 (20.4%) from 49 dwarf goats, of 5 (14.3%) from 35 Cameroon sheep and of 1 (9.1%) from 11 alpine ibex. M. paratuberculosis could not found by cultural method in fecal samples of 22 Pinzgauer goats, of 15 bantengs, of 9 wild goats, of 9 skuddens, of 6 four-horned sheep, of 3 red-head sheep, and of 1 chamois. From all 19 animals with cultural positive fecal samples complement binding antibodies against M. paratuberculosis could not be found in the corresponding serum samples. The results confirm that M. paratuberculosis is more frequently in small zoo ruminants than up to now was suspected. The cultural examination of fecal samples has been proved to be a better method for detecting animal excretors than serological investigations by means of the complement fixation test.     

Detection and characterization of Clostridium species in soil of Zambia

In the retrospective study of soil-borne diseases of cattle in Zambia, malignant edema and blackquarter were widespread. One hundred and sixty-five cases with malignant edema and 103 cases with blackquarter were reported between 1985 and 1997. It was found that specific soil-conditions associate the emergence of the soil-borne diseases. Soil samples from five areas in Zambia were examined for the presence of genus Clostridium. Direct immuno-fluorescent assay (IFA) examination showed that C. septicum, C. novyi and C. chauvoei were detected in the soil of specific areas in Zambia, respectively. Causal organisms such as C. perfringens were isolated from the soil samples. The information of area-specific distribution of Clositridium species may give an efficient program in protecting cattle and man.     

The antagonism of ketamine/xylazine anesthesia ("Hellabrunn mixture") in wild zoo ruminants

The ability of the antagonists tolazoline, yohimbine and the combination of yohimbine with 4-aminopyridine to reverse the effects of the xylazine-component of the "Hellabrunn mixture" (125 mg/ml xylazine and 100 mg/ml ketamine) on nondomestic zoo ruminants is discussed. Arousal time, recovery time and changes in the parameter of circulatory and respiratory functions after antagonization are shown. Tolazoline is able to antagonize the xylazine effect completely within a short time. Using a dosage of 3-5 mg/kg there is a marked negative effect on the cardio-vascular system. Yohimbine in the used dosage of 0.25-0.3 mg/kg in non-domestic ruminants did not approve in its effects. Combining yohimbine (0.25-0.3 mg) with 4-aminopyridine (0.5 mg/kg) recovery time is about 30 minutes. The negative effect on the cardio-vascular system is less pronounced compared with tolazoline.     

Isolation of Campylobacter species from zoo animals and polymerase chain reaction-based random amplified polymorphism DNA analysis

A total of 104 fecal specimens from 30 mammals, 12 birds, and 3 reptiles at the Phoenix Zoological Gardens, Miyazaki City, Japan, were examined for the presence of Campylobacter species. All the animals examined were healthy with no fecal abnormality. Twenty-three (22.1%) thermophilic campylobacters, (9 C. jejuni, 11 C. hyointestinalis, 2 C. coli, and 1 C. lari), were isolated from 11 animals (7 mammals and 4 birds). C. jejuni and C. hyointestinalis were the predominant species isolated from these zoo animals and C. hyointestinalis was isolated frequently from simians. As selective media influence the numbers and species of campylobacters isolated, the agar medium was not supplemented with cephalothin. Campylobacters were isolated most frequently when a combination of enrichment culture and selective agar plating was performed at 42 degrees C. For the epidemiological study, a polymerase chain reaction (PCR)-based randomly amplified polymorphic DNA (RAPD) method was used as a tool to detect the heterogeneity of amplified DNAs of Campylobacter spp. isolated from zoo animals. The two arbitrary primers used in this study enabled even closely related strains of the same Campylobacter spp. to be differentiated. RAPD analysis revealed considerable diversity among the strains, suggesting that the transmission of Campylobacter spp. among animals in a defined area occurred through different mechanisms.Examination of the genotypic diversity among the multiple clones from the same host also revealed differences between clones. These results demonstrate that campylobacter populations in zoo animals are highly divergent.     

Detection of the saa gene in verotoxin-producing Escherichia coli from ruminants.

A total of 163 verotoxin-producing Escherichia coli (VTEC) strains isolated from diarrheic and healthy cattle, sheep, and goats were analyzed for the presence of the saa gene by polymerase chain reaction. Seventeen (45.9%) and 5 (29.4%) of the VTEC isolated from healthy cattle and diarrheic calves, respectively, had the saa gene. None of the saa-positive strains carried the eae gene, but 20 of the 22 saa positive were ehxA positive. In contrast with cattle VTEC, none of the VTEC isolated from small ruminants were saa positive. These results show that the saa gene is commonly associated with bovine eae-negative VTEC strains but not with ovine or caprine VTEC strains.     

Mycoplasma species and related organisms isolated from ruminants in Britain between 1990 and 2000

Between 1990 and 2000, more than 1600 mycoplasmas and the related acholeplasmas were identified from ruminant animals by the Mycoplasma Group at the Veterinary Laboratories Agency--Weybridge. Mycoplasma bovis was the most commonly identified pathogen, mostly from pneumonic calves but occasionally from cattle with mastitis and arthritis. Mycoplasma canis was first isolated in Britain in 1995 from pneumonic calves and the number of isolates increased to 18 per cent of the total mycoplasmas isolated from cattle in 1999. The ELISA for antibodies to M. bovis detected 1971 positive samples (22 per cent) among 8959 serum samples, mainly from pneumonic cattle. Other mycoplasmas identified included Mycoplasma dispar from the lungs of cattle with respiratory disease, and Mycoplasma bovigenitalium from the reproductive tract of cows with vulvovaginitis and infertility. Mycoplasma bovirhinis and Acholeplasma species were found commonly but are thought to be more opportunistic than pathogenic. In sheep and goats, the majority of Mycoplasma species isolated were identified as Mycoplasma ovipneumoniae from pneumonic sheep, Mycoplasma conjunctivae from sheep with keratoconjunctivitis, and the ubiquitous Mycoplasma arginini.     

Emergence and genetic variability of Anaplasma species in small ruminants and ticks from Central Europe

Anaplasmoses are common tick-borne zoonotic bacterial diseases of livestock and free-living ungulates from the genus Anaplasma that are recently emerging in Central Europe. The main aim of this study was to analyze the prevalence and genetic variability of Anaplasma phagocytophilum and Anaplasma ovis in small ruminants and questing ticks from six different sites in Slovakia and the Czech Republic using the PCR of the msp4 gene followed by the sequence analysis. At two farms from southeastern Slovakia, 66.1% small ruminants were infected with A. ovis in contrast to one positive animal from both sites in northern Slovakia. It was represented by two different genotypes. A. phagocytophilum was present in all tested flocks with the infection prevalence ranging from 0.9% to 5.7%. None of the tested questing ticks carried A. ovis. A. phagocytophilum was detected in 1.1% and 7.8% of questing Ixodes ricinus ticks collected around the farms located in southeastern and northern Slovakia, respectively. A. phagocytophilum revealed higher intraspecific diversity than A. ovis.     

Molecular detection of zoonotic tick-borne pathogens from ticks collected from ruminants in four South African provinces

Ticks carry and transmit a remarkable array of pathogens including bacteria, protozoa and viruses, which may be of veterinary and/or of medical significance. With little to no information regarding the presence of tick-borne zoonotic pathogens or their known vectors in southern Africa, the aim of our study was to screen for Anaplasma phagocytophilum, Borrelia burgdorferi, Coxiella burnetii, Rickettsia species and Ehrlichia ruminantium in ticks collected and identified from ruminants in the Eastern Cape, Free State, KwaZulu-Natal and Mpumalanga Provinces of South Africa. The most abundant tick species identified in this study were Rhipicephalus evertsi evertsi (40%), Rhipicephalus species (35%), Amblyomma hebraeum (10%) and Rhipicephalus decoloratus (14%). A total of 1634 ticks were collected. DNA was extracted, and samples were subjected to PCR amplification and sequencing. The overall infection rates of ticks with the target pathogens in the four Provinces were as follows: A. phagocytophilum, 7%; C. burnetii, 7%; E. ruminantium, 28%; and Rickettsia spp., 27%. The presence of B. burgdorferi could not be confirmed. The findings of this study show that zoonotic pathogens are present in ticks in the studied South African provinces. This information will aid in the epidemiology of tick-borne zoonotic diseases in the country as well as in raising awareness about such diseases in the veterinary, medical and tourism sectors, as they may be the most affected.     

Detection of Coxiella burnetii in placenta and abortion samples from British ruminants using real-time PCR

A real-time PCR was developed to detect Coxiella burnetii (the cause of Q fever) in ruminant placentas and aborted fetuses. Primer and probe sets previously developed for human tissue studies were used to target the insertion sequence IS1111 gene for C burnetii. The assay was highly sensitive, with a limit of detection of 10 copies of template, theoretically equating to a single bacterium, and did not cross-react with a panel of other bacteria. To determine sensitivity on field samples submitted for the diagnosis of abortion, results using the IS1111 PCR assay were compared with a com1 PCR assay. When applied to ruminant abortion material, including placental cotyledons and fetal samples, the IS1111 and com1 assays yielded positive results in 23 (25 per cent) of 93 and 19 (20 per cent) of 93 samples, respectively. One infected goat herd was monitored for 31 months: 57 (92 per cent) of 62 placental cotyledon samples from aborting and non-aborting goats, and 10 (30 per cent) of 33 fetal samples were positive by the IS1111 PCR assay.     

Characteristics of mature oocytes from four species of marine teleosts

The morphology and histology of the digestive tract of the branchiuran crustacean, Chonopeltis australis Box shall, 1976 are described from serial sections. The foregut is differentiated into a preoral cavity, containing the mandibles and tongue, an ascending oesophagus, with an H-shaped lumen invested with longitudinal, circular and dilator muscles, a horizontal oesophagus with a star-shaped lumen and lacking longitudinal and dilator muscles, and an oesophageal funnel consisting of inner and recurrent walls. The midgut is differentiated into anterior and posterior chambers, separated by an S-shaped muscular tube. The arborescent midgut glands open laterally into the anterior part of the anterior midgut. Columnar epithelial cells line the anterior midgut whereas tall, papilliform cells are present in the epithelium of the posterior midgut. The transition from posterior midgut to hindgut is marked by the presence of very tall epithelial cells. The terminology describing the various parts of the digestive tract of branchiurans is discussed.     

Detection and molecular characterization of a novel large Babesia species in a dog

Babesia canis has generally been considered the only large Babesia to infect dogs. Here we describe the molecular characterization of a large Babesia species that was detected in the blood and bone marrow of a dog with clinical and hematological abnormalities consistent with babesiosis. Analysis of the 18S rRNA genes revealed a unique sequence that shared 93.9% sequence identity with B. bigemina and 93.5% sequence identity with B. caballi, compared to 91.2-91.6% identity with B. canis canis, B. c. vogeli, and B. c. rossi. Cross-reactive antibodies against B. canis, B. gibsoni (Asian genotype), or B. gibsoni (California genotype) antigens were not detected in acute or convalescent serum samples. The dog was treated with imidocarb diproprionate, which resulted in the resolution of clinical signs, and subsequently Babesia DNA was not detectable by PCR in post-treatment samples. The organism described in this report represents a genetically unique large Babesia sp. and is the eighth genetically distinct piroplasm capable of infecting the domestic dog.     

Detection of respiratory and enteric shedding of bovine coronaviruses in cattle in Northwestern Turkey

Bovine coronavirus (BCoV) is an important cause of diarrhoea in calves, winter dysentery in adult cattle and respiratory tract disease in feedlot cattle. Serum, faecal and nasal swab samples were collected from a total of 96 cattle with clinical signs in 29 barns of 23 villages in Northwestern Turkey. The cattle were subdivided into 3 distinct age groups (0-30 days old, 4-12 months old and 2-7 years old). An indirect antigen-capture ELISA and an antibody-detection ELISA as well as geometric mean BCoV antibody titres were used to detect BoCV shed in the faeces and in the nasal secretions, respectively. Relationships between BCoV shedding and age group, seroconversion and clinical signs in cattle were also analysed. The rate of faecal shedding of BoCV was 37.1% (13/35) in 0-30 days old calves, 25.6% (10/39) in 4-12 months old feedlot cattle and 18.2% (4/22) in 2-7 years old cows. The overall rate of BCoV faecal shedding was 28.1% (27/96) in the cattle examined. Only one animal in the 4-12 months old age group was found to shed BoCV nasally. The analysis showed that there was a significant difference (P < 0.0001) with respect to faecal shedding between the clinical signs and the age groups. BCoV antibody titre in 50% of all cattle was < or =100 as detected by ELISA while 27.1% of the cattle had high titres ranging between 1,600 and 25,600. The seroconversion rate was 7.3% (7/96) in animals shedding BoCV in the faeces and 42.7% (41/96) in cattle negative for faecal shedding as detected by ELISA, and 20.8% of cattle with no seroconversion shed BCoV in the faeces. There was no statistically significant association between seroconversion and nasal or faecal BCoV shedding. These findings confirm the presence of BCoV infections in Turkey. Further studies are needed to isolate BCoV strains in Turkey and to investigate their antigenic and genetic properties.     

Serum biochemical and electrophoretic values from four deer species and from pronghorn antelope.

Serums from 4 species of deer and 1 species of antelope were analyzed for various components in order to define an animal disease model for sickle cell disease in people. Animal species included black-tailed deer (Odocoileus hemionus columbianus), mule deer (Odocoileus hemionus), sika deer (Cervus nippon nippon), fallow deer (Dama dama), and pronghorn antelope (Antilocapra americana). The mean serum values for total bilirubin, total protein, albumin, creatinine, urea nitrogen, and electrolytes were similar in all species and were in the normal range for human beings. Cholesterol and uric acid values for all animals were lower than those for people. Alkaline phosphatase values in the 4 cervid species were higher than in the pronghorn antelope. Values for glutamic oxalacetic transaminase were lower in the cervids than in the pronghorn antelope. Lactic dehydrogenase values were similar in the 5 species. High activities for glutamic oxalacetic transaminase and lactic dehydrogenase in the 5 species probably related to muscle mass and great muscular activity.