Middle East respiratory syndrome: making the case for surveillance of transboundary coronaviruses in the Middle East.

Middle East respiratory syndrome (MERS) is a zoonotic viral disease identified in both animals and human beings. More than 2,200 laboratory-confirmed cases have been reported in humans from 27 countries, with a crude case fatality rate of 35% since the disease's emergence in the Middle East in 2012. In the coming years, MERS will continue to pose a severe threat to economic development as well as to the elimination of poverty and advances in food security. An important gap in the effort to keep MERS at bay is the lack of surveillance of animals in the Middle East. The authors identify the need for international collaboration to conduct MERS coronavirus (CoV) surveillance in animals in the Middle East, since the emergence of new MERS-CoV variants with the ability to sustain efficient person-to-person transmission is a genuine threat. However, effective surveillance will be very difficult, if not impossible, to achieve. There are multiple obstacles in the region to overcome, including a lack of transparency as governments in the Middle East generally do not disclose detailed information on animal diseases. In addition, there is minimal collaboration between local and international agencies in both the human and animal health sectors and a limited number of readily available qualified laboratories to screen animals for MERS- CoV. Last, but not least, there is a lack of adequate active communication between all relevant laboratories, local and abroad. However, with the support of the Food and Agriculture Organization of the United Nations (FAO), the World Organisation for Animal Health (OIE), and other partners, the responsibility of the Mediterranean Zoonosis Control Centre in Athens, Greece, could be widened to include the countries of the Middle East. This would foster a stronger alliance and far more effective collaboration in the spirit of One Health.

Le syndrome respiratoire du Moyen-Orient (MERS) est une maladie virale zoonotique qui affecte à la fois l'homme et les animaux. Plus de 2 200 cas humains confirmés au laboratoire ont été notifiés dans 27 pays depuis l'apparition de la maladie au Moyen-Orient en 2012, avec un taux brut de létalité de 35 %. Dans les années à venir, le MERS continuera à représenter une menace aussi bien pour le développement économique que pour la réussite des objectifs d'élimination de la pauvreté et de sécurisation de l'approvisionnement alimentaire. L'un des principaux obstacles empêchant de tenir le MERS en échec est l'absence de surveillance sanitaire exercée sur les populations animales au Moyen-Orient. Les auteurs soulignent la nécessité d'une collaboration internationale en matière de surveillance du coronavirus responsable du MERS (MERS-Cov) chez les animaux au Moyen-Orient, d'autant que l'émergence de nouveaux variants du MERS-CoV qui entretiennent l'infection en favorisant la transmission de personne à personne constitue un véritable danger. Toutefois, il sera extrêmement difficile, voire impossible de mettre en place une surveillance efficace. En effet les obstacles sont nombreux dans la région, en particulier l'absence de transparence puisque les gouvernements du Moyen-Orient ne publient généralement pas d'informations détaillées sur les maladies animales présentes sur leur territoire. En outre, la collaboration entre les agences locales et internationales des secteurs de la santé publique et animale est réduite au minimum et rares sont à ce jour les laboratoires possédant les compétences requises pour procéder au dépistage de l'infection par le MERS-CoV chez les animaux. Dernière difficulté mais non la moindre, les laboratoires compétents dans les pays et à l'étranger ne communiquent pas entre eux de manière proactive. Dans ce contexte, il est envisagé d'élargir la portée du Centre méditerranéen de lutte contre les zoonoses, situé à Athènes (Grèce) afin d'y intégrer les pays du Moyen-Orient, avec le soutien de l'Organisation des Nations Unies pour l'alimentation et l'agriculture (FAO), de l'Organisation mondiale de la santé animale (OIE) et d'autres partenaires. Cette initiative permettrait de renforcer les alliances et de déployer une collaboration bien plus efficace, dans une perspective Une seule santé.

El síndrome respiratorio de Oriente Medio (MERS, por su acrónimo inglés) es una enfermedad viral zoonótica que se ha descrito tanto en animales como en personas. Desde que en 2012 surgió en el Oriente Medio, se han notificado más de 2 200 casos confirmados en laboratorio que afectan a personas de 27 países, con una tasa bruta de letalidad del 35%. En los próximos años, el MERS seguirá constituyendo una grave amenaza para el desarrollo económico y también para el avance hacia la eliminación de la pobreza y la seguridad alimentaria. A la hora de poner coto a la enfermedad, un importante problema es la deficiente vigilancia zoosanitaria en el Oriente Medio. Los autores señalan la necesidad de colaboración internacional para hacer efectiva en la región la vigilancia del coronavirus (CoV) del MERS en los animales, pues la aparición de nuevas variantes de este virus capaces de transmitirse eficaz y sostenidamente entre las personas constituye un verdadero peligro. Sin embargo, resultará difícil, si no imposible, efectuar una vigilancia eficaz, habida cuenta de la multitud de obstáculos que hay que superar en el Oriente Medio, incluida la falta de transparencia de los gobiernos, que no acostumbran a revelar información detallada sobre las enfermedades animales. Además, la colaboración entre instancias locales y organismos internacionales en los sectores de la salud humana y la sanidad animal es mínima, y hay contados laboratorios cualificados para la detección del MERS­CoV en animales que estén en condiciones de intervenir con presteza. Por último, pero no menos importante, no hay una adecuada comunicación activa entre todos los laboratorios competentes, ya sean de los propios países o del extranjero. No obstante, con apoyo de la Organización de las Naciones Unidas para la Alimentación y la Agricultura (FAO), la Organización Mundial de Sanidad Animal (OIE) y otros colaboradores, sería posible extender a los países del Oriente Medio el ámbito de responsabilidad y actuación del Centro de Control de Zoonosis del Mediterráneo, sito en Atenas (Grecia), cosa que favorecería alianzas más sólidas y mucho más eficaces, conforme al espíritu de Una sola salud.

Genomics and zoonotic infections: Middle East respiratory syndrome.

The emergence of Middle East respiratory syndrome (MERS) and the discovery of MERS coronavirus (MERS-CoV) in 2012 suggests that another SARS-like epidemic is occurring. Unlike the severe acute respiratory syndrome (SARS) epidemic, which rapidly disappeared in less than one year, MERS has persisted for over three years. More than 1,600 cases of MERS have been reported worldwide, and the disease carries a worryingly high fatality rate of >30%. A total of 182 MERS-CoV genomes have been sequenced, including 94 from humans and 88 from dromedary camels. The 182 genomes all share >99% identity, indicating minimal variation among MERS-CoV genomes. MERS-CoV is a lineage C Betacoronavirus (ßCoV). MERS-CoV genomes can be roughly divided into two clades: clade A, which contains only a few strains, and clade B, to which most strains belong. In contrast to ORF1ab and structural proteins, the putative proteins encoded by ORF3, ORF4a, ORF4b, ORF5 and ORF8b in the MERS-CoV genome do not share homology with any known host or virus protein, other than those of its closely related lineage C ßCoVs. Human and dromedary viral genomes have intermingled, indicating that multiple camel-to-human transmission events have occurred. The multiple origins of MERS-CoV suggest that the virus has been resident in dromedaries for many years. This is consistent with the detection of anti-MERS-CoV antibodies in dromedary camels as early as the 1980s.

L'émergence du syndrome respiratoire du Moyen-Orient (SRMO, ou MERS d'après son sigle anglais) et l'identification en 2012 du coronavirus responsable de cette maladie (MERS-CoV) indiquent que nous sommes en présence d'une épidémie semblable à celle du syndrome respiratoire aigu sévère (SRAS). Toutefois, contrairement à l'épidémie du SRAS qui avait rapidement disparu en moins d'un an, le MERS persiste depuis plus de trois ans. Plus de 1 600 cas de MERS ont été notifiés dans le monde ; la maladie présente un taux de létalité particulièrement préoccupant, s'élevant à plus de 30 %. Au total, 182 génomes du MERS-CoV ont été séquencés jusqu'à présent, dont 94 provenaient de virus isolés chez l'homme et 88 chez des dromadaires. Ces 182 génomes ont en commun un pourcentage d'identité de 99 %, dénotant une très faible variabilité des génomes viraux. Le MERS-CoV appartient à la lignée C du genre Betacoronavirus (ßCoV). Les génomes du MERSCoV se répartissent, dans leurs grandes lignes, en deux clades : le clade A, qui ne contient que quelques souches, et le clade B regroupant l'immense majorité des souches. Contrairement à ce qui se produit avec la protéine ORF1ab et les protéines structurales, les protéines potentiellement codées par les gènes ORF3, ORF4a, ORF4b, ORF5 et ORF8b du génome du MERS-CoV ne présentent aucune homologie avec des protéines virales ou de l'hôte autres que celles d'autres bêtacoronavirus de la lignée C, qui lui sont étroitement apparentés. Les génomes des virus affectant l'homme et le dromadaire se sont entremêlés, ce qui montre que le virus a connu de multiples épisodes de transmission des camélidés à l'homme. Les origines multiples du MERS-CoV témoignent d'une présence prolongée du virus (plusieurs années) chez les dromadaires. Ce constat est corroboré par le fait que des anticorps anti-MERS-CoV ont été détectés chez des dromadaires dès le début des années 80.

La aparición del síndrome respiratorio de Oriente Medio (MERS, por sus siglas en inglés) y el descubrimiento del coronavirus que lo causa (MERS-CoV) en 2012 parecen apuntar al advenimiento de una nueva epidemia análoga a la del síndrome respiratorio agudo severo (SRAS). Pero a diferencia de lo ocurrido con la epidemia de SRAS, que en menos de un año había desaparecido, el MERS lleva más de tres años presente. En el mundo se han notificado más de 1.600 casos de MERS, y la enfermedad presenta una tasa de letalidad muy alta y preocupante, superior al 30%. Hasta ahora se han secuenciado un total de 182 genomas del MERS-CoV, 94 de ellos obtenidos a partir de personas y 88 a partir de dromedarios. Estos 182 genomas comparten identidad en más de un 99%, lo que pone de manifiesto un nivel mínimo de variación entre los genomas coronavíricos. El coronavirus del MERS pertenece al linaje C del género Betacoronavirus (ßCoV). Los genomas de este virus pueden ser divididos, a grandes rasgos, en dos clados: el clado A, que agrupa unas pocas cepas; y el clado B, al que pertenecen la gran mayoría de las cepas. A diferencia de lo que ocurre con la proteína ORF1ab y las proteínas estructurales, las proteínas que supuestamente codifican los genes ORF3, ORF4a, ORF4b, ORF5 y ORF8b del genoma del MERS-CoV no comparten homología con ninguna proteína conocida de otros virus o anfitriones, salvo con proteínas de otros betacoronavirus del linaje C estrechamente emparentados con él. Los genomas de los virus que afectan a personas y dromedarios se han entremezclado, lo que indica que se han producido numerosos episodios de transmisión de camélidos a humanos. De los múltiples orígenes del MERS-CoV se deduce que el virus lleva muchos años siendo residente en dromedarios, lo que concuerda con el hecho de que ya en los años ochenta se detectaran anticuerpos anti-MERS-CoV en dromedarios.

Camelid brucellosis: a review.

Camel brucellosis has been diagnosed in all camel-rearing countries except Australia. In many countries the infection is on the rise in Old World camels (OWCs) due to the uncontrolled trade of live animals. Knowledge of camelid brucellosis has increased over the last decade through field investigations, experimental infection trials and comprehensive laboratory testing. Infection with Brucella melitensis is frequent in OWCs and rare with B. abortus. New World Camels rarely contract brucellosis. In East African countries the seroprevalence of brucellosis can reach 40% (herd level) and depends on the management system. The highest incidence is found when camels are kept together with infected small ruminants. Only a combination of serological methods can detect all serological reactors. Culturing the pathogen is still the preferred test method, although several assays based on polymerase chain reaction have been developed.

Detection of a novel genotype of Chlamydia buteonis in falcons from the Emirates.

Chlamydiaceae are a family of obligate intracellular bacterial pathogens that affect both humans and animals. Recently, a new species named Chlamydia (C.) buteonis was isolated from hawks. In this study, we aimed to investigate the prevalence of Chlamydiaceae in 60 falcons that underwent a routine health check at a specialized clinic in Dubai, United Arab Emirates. Using real-time PCR, we analyzed cloacal and tracheal swabs from these birds and found that 39 of them tested positive for Chlamydiaceae. Subsequent real-time PCR assays specific for C. psittaci, C. abortus, C. avium, and C. gallinacea yielded negative results, while testing positive for C. buteonis. Analysis of ompA and MLST sequences indicated a highly conserved group of strains within this set of samples, but with sequences distinct from the C. buteonis RSHA reference strains and other C. buteonis strains isolated from hawks in the United States. Two strains were further isolated by cell culture and sequenced using whole-genome sequencing, confirming the clustering of these falcon strains within the C. buteonis species, but in a separate clade from the previously identified hawk strains. We also developed a SNP-based PCR-HRM assay to distinguish between these different genotypes. Overall, our findings suggest a high prevalence of C. buteonis in falcons in Dubai and highlight the importance of monitoring this pathogen in birds of prey.

Microbiological quality and somatic cell count in bulk milk of dromedary camels (Camelus dromedarius): descriptive statistics, correlations, and factors of variation.

The objectives of the present study were to monitor the microbiological quality and somatic cell count (SCC) of bulk tank milk at the world's first large-scale camel dairy farm for a 2-yr period, to compare the results of 2 methods for the enumeration of SCC, to evaluate correlation among milk quality indicators, and to determine the effect of specific factors (year, season, stage of lactation, and level of production) on milk quality indicators. The study was conducted from January 2008 to January 2010. Total viable count (TVC), coliform count (CC), California Mastitis Test (CMT) score, and SCC were determined from daily bulk milk samples. Somatic cell count was measured by using a direct microscopic method and with an automatic cell counter. In addition, production parameters [total daily milk production (TDM, kg), number of milking camels (NMC), average milk per camel (AMC, kg)] and stage of lactation (average postpartum days, PPD) were recorded for each test day. A strong correlation (r=0.33) was found between the 2 methods for SCC enumeration; however, values derived using the microscopic method were higher. The geometric means of SCC and TVC were 394×10(3) cells/mL and 5,157 cfu/mL during the observation period, respectively. Somatic cell count was >500×10(3) cells/mL on 14.6% (106/725) and TVC was >10×10(3) cfu/mL on 4.0% (30/742) of the test days. Both milk quality indicators had a distinct seasonal pattern. For log SCC, the mean was lowest in summer and highest in autumn. The seasonal pattern of log TVC was slightly different, with the lowest values being recorded during the spring. The monthly mean TVC pattern showed a clear difference between years. Coliform count was <10 cfu/mL in most of the samples (709/742, 95.6%). A positive correlation was found between log SCC and log TVC (r=0.32), between log SCC and CMT score (r=0.26), and between log TVC and CC in yr 1 (r=0.30). All production parameters and stage of lactation showed strong seasonal variation. Log SCC was negatively correlated with TDM (r=-0.35), AMC (r=-0.37), and NMC (r=-0.15) and positively correlated with PPD (r=0.40). Log TVC had a negative correlation with AMC (r=-0.40) but a positive correlation with NMC (r=0.32), TDM (r=0.16), and PPD (r=0.45). The linear mixed model with stepwise variable selection showed that the main sources of log SCC variation were PPD, TDM, PPD × season, and season. For log TVC, the same factors and year contributed to the variation.

Tuberculosis in camelids: a review.

Tuberculosis is a chronic, contagious, granulomatous disease caused by mycobacterial species belonging to the Mycobacterium tuberculosis complex. Camelids were not considered highly susceptible to tuberculosis, but in recent years increased numbers of cases have been experienced in some countries. In most of the cases, transmission probably occurs through contact with infected cattle or wildlife. None of the ante-mortem tests currently available can consistently provide accurate diagnosis of the infection in live camelids. Recently developed serological assays have the potential for rapid and accurate diagnosis of tuberculosis but still need to be validated.

Foot and mouth disease and similar virus infections in camelids: a review.

Foot and mouth disease (FMD) remains the most important animal disease. The FMD virus is highly contagious and occurs almost exclusively among cloven-hoofed animals such as cattle, sheep, goats, Bactrian camels and swine. Old World camels (OWCs) and New World camels (NWCs) inhabit FMD-endemic countries in South America, North and East Africa, and the Middle and Far East. Results of experimental infection of OWCs with the virus, and several clinical observations from the field over a century, confirm that the two closely related camel species of Bactrian and dromedary camels possess noticeably different susceptibilities to FMD. It is now certain that Bactrian camels can contract the disease. In contrast, dromedaries are not susceptible to FMD and do not transmit infection, even when in close contact with susceptible animals. The susceptibility of NWCs to the FMD virus has been demonstrated in the field and in experimental infection trials. However, these animals are not very susceptible and do not represent a serious risk in transmitting FMD to susceptible animal species.

Culture of chicken embryo in interspecific surrogate egg white.

The effect of interspecific egg white on the development of chicken embryos was investigated in a surrogate eggshell culture system. Egg yolks were separated from fertile White Leghorn chicken eggs and cultured in different egg whites from turkey (group TK), guineafowl (group GF), and duck (group DK), and chicken (group CK) was used as control. The viability of chicken embryos in groups CK, TK, GF, and DK after 3 d culture in system II was 98.3, 90.2, 96.1, and 91.1%. The whole contents (egg yolk and surrogate egg white) were further transferred into an eggshell from a 1.5 times heavier chicken egg with air space (system III), and incubated for further 16 d, before moving them to a hatcher. No significant difference between the 4 groups was found in their viabilities, which ranged between 72.9 and 81.3%, until 14 d postincubation (P > 0.05). After 21 d, the viability decreased to 60.4, 57.4, 50.0, and 27.7% in groups CK, TK, GF, and DK. The viability in group DK was significantly lower than in the other groups (P < 0.05). Weight loss in system III was approximately 12% in all the 4 groups without significant difference (P > 0.05). Hatchability of the chicken embryo was 60.4, 55.3, 47.9, and 19.1% in groups CK, TK, GF, and DK, respectively, and that in group DK was significantly lower than in the other groups (P < 0.05). There was no difference between the other groups (P > 0.05). These results show that chicken embryos can develop to hatch in duck, guineafowl, and turkey egg whites. However, the hatchability decreases according to the phylogenetic distance. The present study will provide a tool for manipulation of avian embryos and eventual conservation of endangered wild birds.

Production of the first cloned camel by somatic cell nuclear transfer.

In this study, we demonstrate the use of somatic cell nuclear transfer to produce the first cloned camelid, a dromedary camel (Camelus dromedarius) belonging to the family Camelidae. Donor karyoplasts were obtained from adult skin fibroblasts, cumulus cells, or fetal fibroblasts, and in vivo-matured oocytes, obtained from preovulatory follicles of superstimulated female camels by transvaginal ultrasound guided ovum pick-up, were used as cytoplasts. Reconstructed embryos were cultured in vitro for 7 days up to the hatching/hatched blastocyst stage before they were transferred to synchronized recipients on Day 6 after ovulation. Pregnancies were achieved from the embryos reconstructed from all cell types, and a healthy calf, named Injaz, was born from the pregnancy by an embryo reconstructed with cumulus cells. Genotype analyses, using 25 dromedary camel microsatellite markers, confirmed that the cloned calf was derived from the donor cell line and the ovarian tissue. In conclusion, the present study reports, for the first time, establishment of pregnancies and birth of the first cloned camelid, a dromedary camel (C. dromedarius), by use of somatic cell nuclear transfer. This has opened doors for the amelioration and preservation of genetically valuable animals like high milk producers, racing champions, and males of high genetic merit in camelids. We also demonstrated, for the first time, that adult and fetal fibroblasts can be cultured, expanded, and frozen without losing their ability to support the development of nuclear transfer embryos, a technology that may potentially be used to modify fibroblast genome by homologous recombination so as to generate genetically altered cloned animals.

Camelimonas lactis gen. nov., sp. nov., isolated from the milk of camels.

Three strains of Gram-negative, rod-shaped, non-spore-forming bacteria (M 2040(T), M 1973 and M 1878-SK2), isolated from milk of camels at a camel-milk production farm in the United Arab Emirates, were investigated for their taxonomic allocation. On the basis of 16S rRNA gene sequence similarities, all three strains were shown to belong to the Alphaproteobacteria and were most closely related to Chelatococcus asaccharovorans and Chelatococcus daeguensis (95.1 and 95.2 % sequence similarity to the respective type strains). meso-Diaminopimelic acid was detected as the characteristic peptidoglycan diamino acid. The predominant compound in the polyamine pattern was spermidine, and sym-homospermidine was not detectable. The quinone system was ubiquinone Q-10. The polar lipid profile included the major compounds phosphatidylcholine and diphosphatidylglycerol and moderate amounts of phosphatidylethanolamine, phosphatidylglycerol, an unidentified glycolipid and two unidentified aminolipids. Minor lipids were also detected. The major fatty acid profile, consisting of C19 :0 cyclo ω8c and C18:1 ω7c, with C18 :03-OH as the major hydroxylated fatty acid, was similar to those of the genus Chelatococcus. The results of DNA-DNA hybridization experiments and physiological and biochemical tests allowed both genotypic and phenotypic differentiation of the isolates from described Chelatococcus species. Isolates M 2040(T), M 1973 and M 1878-SK2 were closely related on the basis of DNA-DNA reassociation and therefore represent a single novel species. In summary, low 16S rRNA gene sequence similarities of 95 % with Chelatococcus asaccharovorans and marked differences in polar lipid profiles as well as in polyamine patterns support the description of a novel genus and species to accommodate these strains, for which the name Camelimonas lactis gen. nov., sp. nov. is proposed. The type strain of Camelimonas lactis is M 2040(T) (=CCUG 58638(T) =CCM 7696(T)).

Peste des petits ruminants in Arabian wildlife.

Recurrence of peste des petits ruminants (PPR) was diagnosed in the United Arabian Emirates in several wild ruminants confirmed by morphological, immunohistochemical, serological and molecular findings. Phylogenetic analysis revealed that the virus strain belongs to lineage IV, which is different to some previously isolated PPR strains from the Arabian Peninsula. This study shows that wild ruminants may play an important epidemiological role as virus source for domestic small ruminants.

Prevalence of Clostridium perfringens in faeces and ileal contents from grass sickness affected horses: comparisons with 3 control populations.

REASONS FOR PERFORMING STUDY: While previous studies have demonstrated an association between equine grass sickness (EGS) and the presence of Clostridium botulinum within ileal contents and faeces, no such associations with other intestinal-derived anaerobic bacteria have been extensively investigated. HYPOTHESIS: The prevalence of C. perfringens in the ileal contents and faeces of EGS horses is greater than control horses; the detection of C. perfringens in faeces by ELISA could be diagnostically beneficial in a clinical setting. METHODS: The prevalence of C. perfringens in faeces from EGS horses and healthy grazing control horses was determined by both selective culture and ELISA to permit both validation of the ELISA and inter-group comparisons. Additionally, the prevalence of C. perfringens (ELISA) in ileal contents from EGS horses was compared with that for control horses with nongastrointestinal disease. Finally, the prevalence of C. perfringens (ELISA) in faeces from EGS cases was compared with that from both horses with which they shared pasture at the time of disease onset and non-EGS colic horses. RESULTS: When compared with culture, the ELISA had a sensitivity and specificity of 86 and 98%, respectively. The prevalence of C. perfringens in faeces as determined by both culture and ELISA was significantly higher (P<0.001) for EGS horses (7/9 and 15/37, respectively) than for healthy grazing controls (0/60 and 1/74, respectively). The prevalence of C. perfringens in ileal contents from EGS horses (5/10) was greater than that for horses with nongastrointestinal disease (1/12) at a level that approached significance (P = 0.056). EGS cases had a significantly greater prevalence of C. perfringens in faeces (15/37) than co-grazing horses (1/18) and colic (1/16) horses. The specificity (93%) and PPV (94%) of the detection of C. perfringens by ELISA on faecal samples in relation to disease status (EGS compared with colic horses) was good. Sensitivity (41%) and NPV (39%) were poor. CONCLUSIONS AND POTENTIAL RELEVANCE: The use of a commercial ELISA to detect faecal C. perfringens may be diagnostically beneficial when differentiating EGS cases from colic cases, although further work is required to fully evaluate its potential.

In vitro maturation of dromedary (Camelus dromedarius) oocytes: effect of different protein supplementations and epidermal growth factor*.

The present experiment was aimed to compare the effect of different protein supplementation sources, foetal calf serum (FCS), oestrous dromedary serum (EDS) and BSA, in experiment 1, and the effect of different concentrations of epidermal growth factor (EGF), in experiment 2, on in vitro nuclear maturation of the dromedary oocytes. Cumulus oocyte complexes (COCs) were harvested from the ovaries collected from a local slaughterhouse by aspirating the visible follicles in PBS supplemented with 5% FCS. Pooled COCs were randomly distributed to 4-well culture plates containing 500 µl of the maturation medium and cultured at 38.5 °C in an atmosphere of 5% CO(2) in air for 32-36 h. The basic maturation medium consisted of TCM-199 supplemented with 0.1 mg/ml L-glutamine, 0.8 mg/ml sodium bicarbonate, 0.25 mg/ml pyruvate, 50 µg/ml gentamicin, 10 µg/ml bFSH, 10 µg/ml bLH and 1 µg/ml estradiol. In experiment 1, this medium was supplemented with 10% FCS, 10% EDS or 0.4% BSA, whereas in experiment 2, it was supplemented with 0.4% BSA and 0, 10, 20 or 50 ng/ml of EGF. The oocytes were fixed, stained with 1% aceto-orcein stain and their nuclear status was evaluated. Oocytes were classified as germinal vesicle, diakinesis, metaphase-I, anaphase-I (A-I), metaphase-II (M-II) and those with degenerated, fragmented, scattered, activated or without visible chromatin as others. There was no difference (p > 0.05) observed in the proportion of oocytes reaching M-II stage between the media supplemented with FCS (71.5 ± 4.8), EDS (72.8 ± 2.9) and BSA (72.7 ± 6.2). In experiment 2, a higher proportion (p < 0.05) of oocytes reached M-II stage when the medium was supplemented with 20 ng/ml of EGF (81.4 ± 3.2) when compared with the media supplemented with 10 ng/ml (66.9 ± 4.1) and control (67.2 ± 7.1) groups. It may be concluded that the maturation media for dromedary camel oocytes can be supplemented with any of the three protein sources, i.e. FCS, EDS and BSA without any significant differences on the maturation rates. Also, a supplementation of 20 ng/ml of EGF in the maturation medium seems to be optimal and improves the nuclear maturation of dromedary camel oocytes.

Development of a microsphere-based immunoassay for the serological detection of glanders in equids.

Burkholderia mallei is the etiologic agent of glanders, an infectious disease of solipeds, with renewed scientific interest due to its increasing incidence in different parts of the world. More rapid, sensitive and specific assays are required by laboratories for confirmatory testing of this disease. A microsphere-based immunoassay consisting of beads coated with B. mallei recombinant proteins (BimA, GroEL, Hcp1, and TssB) has been developed for the serological diagnosis of glanders. The proteins' performance was compared with the OIE reference complement fixation test (CFT) and an indirect enzyme-linked immunosorbent assay (iELISA) on a large panel of sera comprised of uninfected horses (n=198) and clinically confirmed cases of glanders from India and Pakistan (n=99). Using Receiver Operating Characteristics (ROC) analysis and adjusting the cutoff levels, Hcp1 (Se=100%, Sp=99.5%) and GroEL (Se= 97%, Sp=99.5%) antigens exhibited the best specificity and sensitivity. Neither Hcp1 and GroEL proteins, nor iELISA reacted with doubtful and positive CFT samples from glanders free countries which further confirmed the false positive reactions seen in CFT.

Camelid immunoglobulins and nanobody technology.

It is well established that all camelids have unique antibodies circulating in their blood. Unlike antibodies from other species, these special antibodies are devoid of light chains and are composed of a heavy-chain homodimer. These so-called heavy-chain antibodies (HCAbs) are expressed after a V-D-J rearrangement and require dedicated constant gamma-genes. An immune response is raised in these so-called heavy-chain antibodies following classical immunization protocols. These HCAbs are easily purified from serum, and the antigen-binding fragment interacts with parts of the target that are less antigenic to conventional antibodies. Since the antigen-binding site of the dromedary HCAb is comprised in one single domain, referred to as variable domain of heavy chain of HCAb (VHH) or nanobody (Nb), we designed a strategy to clone the Nb repertoire of an immunized dromedary and to select the Nbs with specificity for our target antigens. The monoclonal Nbs are well produced in bacteria, are very stable and highly soluble, and bind their cognate antigen with high affinity and specificity. We have successfully developed recombinant Nbs for research purposes, as probe in biosensors, to diagnose infections, and to treat diseases like cancer or trypanosomosis.

Corrigendum to "Incidence and detection of Beak and Feather disease virus in psittacine birds in the UAE" [Biomol. Detect. Quantif. 6 (January) (2016) 27-32].

[This corrects the article DOI: 10.1016/j.bdq.2015.10.001.].

Dromedaries (Camelus dromedarius) are of low susceptibility to inoculation with foot-and-mouth disease virus serotype O.

Two sheep and five dromedaries were inoculated with a highdose of a cattle-passaged type O strain of foot-and-mouth disease virus (FMDV). The sheep developed typical FMD. The inoculated camels, which were placed in contact with five further dromedaries and four sheep, showed no visible signs of illness or vesicular lesions. However, one of them had a raised body temperature at 3 days post-inoculation (pi) and a viraemia from days 2 to 10; probang samples from this animal were negative for infectious virus, but a low level of FMDV RNA was detected in a sample taken on day 6 pi, five other samples taken from days 3 to 28 being negative. Examination of mouth swabs indicated a low level of FMDV RNA at days 1-5 pi in four of the five inoculated camels, but no infectious FMDV or FMDV RNA was detected in serum, probang or mouth swab samples from contact-exposed animals (camels and sheep). All the contact-exposed camels and sheep and two of the inoculated camels were serologically negative for FMD when tested up to day 28. In contrast, the camel with viraemia became serologically positive from day 14, and the other two inoculated camels (which had been exposed to FMDV in an earlier experiment) became serologically positive from day 10. The experiment suggested that dromedaries (1) are of low susceptibility to FMDV serotype O, (2) do not transmit infection, even by close contact, and (3) are unlikely to play a significant epidemiological role in FMD.

Temporal assessment of seroconversion in response to inactivated foot-and-mouth disease vaccine in Arabian oryx (Oryx leucoryx).

Ten male Arabian oryx (Oryx leucoryx) were vaccinated with a commercially available standard aqueous foot-and-mouth-disease vaccine containing aluminium hydroxide as an adjuvant, and their antibody titres against serotypes O and A were measured using solid-phase blocking elisa and the virus neutralisation test. Mean elisa antibody titres greater than 1.45 log(10) were recorded for serotype A, but low elisa titres were recorded for serotype 0; low titres were recorded by VNT for both serotypes.

First molecular characterisation of a Brazilian Burkholderia mallei strain isolated from a mule in 2016.

We present the first molecular characterisation based on MLVA and SNP analysis of a strain of Burkholderia mallei isolated from a mule found dead in Brazil in 2016.

Tuberculosis outbreak in a dromedary racing herd and rapid serological detection of infected camels.

A recent outbreak of tuberculosis (TB) in a dromedary racing herd of 58 animals involved 3 infected animals. Disease was confirmed at necropsy by finding gross lesions from which Mycobacterium bovis (antelope type) was isolated. Sera collected from the camels in this herd were used to evaluate two new serological methods, Multiantigen Print Immunoassay (MAPIA) and rapid test (RT) developed using the lateral-flow technology, in comparison with the intradermal tuberculin tests. Antibodies were found in all three infected dromedaries by both RT and MAPIA, but not in the remaining 55 animals in the herd. With the limited number of animals tested in this study, the serological assays showed the potential for convenient, rapid, and accurate diagnosis of TB in live camels.