National Veterinary Laboratory Research Articles

Rabies incidence and burden in three cities of Cameroon (2004–2013)

Rabies is a fatal disease occurring worldwide and especially in almost all the countries in Asia and Africa including Cameroon. Though animal and human rabies is prevalent in Cameroon, the epidemiology and socio-economic burden of the disease in the country is not known. Therefore, a 10-year (October 2004–April 2013) retrospective study on the incidence of animal and human rabies and its burden in Garoua, Ngaoundéré and Yaoundé in Cameroon was carried out. Records of human cases were extracted from the database of the regional hospitals, and animal cases from the databases of Centre Pasteur and National Veterinary Laboratory. The burden of the disease was assessed through the estimation of costs linked to preventive measures (vaccination), corrective procedures (Post Exposure Treatment), Disability-Adjusted Life Year (DALY) and overall societal cost of the disease. Overall, 56 rabies-suspected human deaths, corresponding to an incidence of 0.02 ± 0.00 ‱ and Animal Rabies Incidence (ARI) of 0.37 ± 0.00 % among 1844 suspected animal cases were recorded. The economic loss due to preventive measures of 326,046 ± 28,130.85 USD, related to corrective procedures of 806,741.25 ± 2,466.08 USD, and DALY of 1690.28 ± 4.76 years were estimated. This is the first study that highlights the enormous socio-economic burden associated with animal and human rabies in endemic parts of Cameroon and emphasizes on enhancing rabies eradication strategy focusing on the One Health approach.

Ionic liquid-based dispersive liquid-liquid microextraction of anthelmintic drug residues in small-stock meat followed by LC-ESI-MS/MS detection.

An ionic liquid-based dispersive liquid-liquid microextraction (IL-DLLME) of 20 anthelmintic drugs followed and detected by liquid chromatography-tandem mass spectrometry (LC-MS/MS) has been developed, optimized, and validated. The parameters affecting the anthelmintic extraction efficiencies such as selection of extraction solvent (ionic liquids), selection of disperser solvent, volume of extraction solvent, volume of disperser solvent, pH of the aqueous phase, extraction time, salt addition, and centrifugation time were optimized. Validation was conducted according to ISO/IEC 17025:2017 and Commission Implementing Regulation (EU) 2021/808 of 22 March 2021. Validation parameters such as calibration function, matrix effect, limit of detection (LOD), limit of quantification (LOQ), decision limit (CCα), accuracy, and precision were established. Coefficient of determination (R 2) values ranging from .99938 to .99995 were obtained using the matrix calibration curve spiked at 0, 0.25, 1.0, 1.5, and 2.0 times MRL. The LODs and LOQs were calculated using the standard deviation of the response and the slopes of the calibration curves ranged from 0.35 to 26.1 μg/kg and from 1.2 to 87.0 μg/kg, respectively, and were dependent on calibration range. The CCα values ranged from 23 to 1022.0 μg/kg and are also dependent on the MRL concentration levels. The coefficient of variation (CV) values calculated are within the reproducibility range of 16%-30% adapted from the Horwitz Equation CV = 2(1-0.5 log C) and ranged from 1.7% to 16.9%. The developed and validated and the standard QuEChERS method were compared. The IL-DLLME LC-MS/MS method was applied to 32 small stock (18 caprine [goat] and 14 ovine [sheep]) liver samples received from municipal abattoirs at Botswana National Veterinary Laboratory for the analysis of anthelmintic drug residues. The results obtained indicated that the anthelmintic drug residues were all below the detection capability, and therefore, the samples were passed as fit for human consumption.

PENICILLIN: A JOURNEY FROM A MAGIC BULLET TO A RESISTANT DRUG

PENICILLIN: A JOURNEY FROM A MAGIC BULLET TO A RESISTANT DRUG

TNF and HNRNPL Related Immunoregulatory Long non-coding RNA (THRIL) and long intergenic noncoding RNA-p21 (lincRNA-p21) as potential useful biomarkers for the diagnosis of tuberculosis

Early diagnosis is crucial in controlling tuberculosis globally and in developing countries with the emergence of drug-resistant Mycobacterium tuberculosis strains. Long non-coding RNAs (lncRNAs) are promising tuberculosis diagnostic biomarkers. Two lncRNA diagnostic markers, lncRNA THRIL and lincRNA-p21, were studied as tuberculosis diagnostic biomarkers. This cross-sectional study was conducted at the Center of Respiratory Diseases of LAQUINTINIE hospital and the National Veterinary Laboratory of Douala from December 2020 to August 2021. The ability of lncRNAs to distinguish between 19 healthy controls, 15 latent tuberculosis, and 21 active tuberculosis was estimated using quantitative polymerase chain reaction and Receiver Operating Characteristic curve analysis. Our analysis showed that lncRNA THRIL and lincRNA-p21 were significantly upregulated (P <0.05) in active and latent tuberculosis compared with healthy controls. LincRNA-p21 expression was significantly increased (P <0.05) in active tuberculosis compared with latent tuberculosis, whereas lncRNA THRIL was not significantly affected (P ≥0.05). Both lncRNA THRIL and lincRNA-p21 showed excellent performance in classifying latent tuberculosis and healthy controls (AUC = 92.86%). Furthermore, lncRNA THRIL was good at discriminating active tuberculosis from healthy controls (AUC = 89.79%), while lincRNA-p21 showed excellent discriminating performance (AUC = 100%). LncRNA THRIL was identified as a poor discriminator of latent tuberculosis from active tuberculosis (AUC = 64.28%), while lincRNA-p21 showed excellent diagnostic performance in this distinction (AUC = 92.86%). Our cross-sectional study suggests that lncRNA THRIL and lincRNA-p21 are promising tuberculosis diagnostic biomarkers that can differentiate between latent and active infection.

Seroprevalence and Risk Factors Associated with Toxoplasma gondii and Chlamydophila abortus Infection in Domestic Small Ruminants in Cameroon

Toxoplasmosis and chlamydiosis remain among the primary causes of abortion and fetal loss in small ruminants. Consequently, they are a source of considerable economic losses for farmers. The objectives of this study were to determine the seroprevalence and highlight the risk factors associated with Toxoplasma gondii and Chlamydophila abortus infection in small ruminants in Cameroon. A cross-sectional study was conducted in 200 small ruminant farms during a period from April to October 2021. A total of 1061 small ruminants were sampled, and the sera obtained were analyzed using the indirect ruminant multi-species ELISA test for specific IgG antibody detection for T. gondii and C. abortus at the National Veterinary Laboratory. A questionnaire was constructed to collect information about flock management and risk factors possibly associated with T. gondii and C. abortus infection in goats and sheep. Overall, 329 small ruminants tested positive for T. gondii-specific IgG antibodies and 45 for C. abortus-specific IgG antibodies with a seroprevalence of 31.1% (95% CI: 28.2–33.8) for T. gondii and 4.2% (95% CI: 3.0–5.5) for C. abortus, respectively. However, a significant co-infection of 1.8% (95% CI: 0.37–3.3) was obtained between T. gondii and C. abortus (p = 0.02). Linear regression model analysis revealed that, the seroprevalence of T. gondii at the flock level was significantly correlated with the region (0.22 CI: 0.17; 0.26) the breeding objectives (0.36 CI: 0.17; 0.56) the level of hygiene (0.11 CI: 0.01; 0.21) and presence abortions (0.55 CI: 0.46; 0.64) on the farms and that of C. abortus was significantly correlated with the regions (0.01 CI: 0.00–0.02) and degree of abortion on the farms (−0.26 CI: −039; −0.14). At the individual level, a significant correlation was observed between the seroprevalence of T. gondii and region (−0.09 CI: −0.14; −0.04), species (0.07 CI: 0.01–0.14), sex (−0.11 CI: −0.18; −0.03), age (0.08 CI: 0.04–0.12) and physiological status (0.21 CI: 0.18; 0.24). For C. abortus seroprevalence, a significant correlation was observed with the regions (0.02 CI: 0.00; 0.03) and breed (0.01 CI: 0.01; 0.02) of small ruminants reared in the northern regions of Cameroon. These results could be used to implement efficient management measures to prevent and control T. gondii and C. abortus infection in goats and sheep in Cameroon.

Isolation and Identification of Endo-Parasites from Fecal Ellets of Avian Species in Captivity

Introduction: Birds are an integral part of ecosystem, they are commonly found all over the world. They can be parasitized by a wide variety of endo-parasites. Objectives/Aim: The current study conducted to identify and investigate the prevalence of endo-parasites from various captive bird species at main campus of PMAS-Arid Agriculture University, Rawalpindi. Methodology: Different standardized methods were used to collect fecal pellets from each bird species in early morning in a plastic bag and transferred to the National Veterinary laboratory (NVL), Islamabad for parasitological examination, eggs, oocytes, cysts and larva of pathogens. Prevalence of endo-parasites was estimated through geometric mean at confidence interval of 95%. Results: Total 403 fecal samples were collected from 13 bird species, 10 out of 13 avian species were found infected with endoparasites. The data on percentage occurrence of infection in avian species viz; pigeons (41.9%) were the most infected with endo-parasites as compared to golden pheasant (19.3%), pea cock (16.1%), cochin hen (12.9%), ring neck pheasant (9.6%), aseel hen (6.45%), polish hen (6.4%), brown hen (3.2%), red jungle fowl (3.2%) and bentum hen (3.2%). However, all of Budgerigar parrots, ducks and guinea fowl were not infected. It was found that Eimeriasp was more prevalent and isolated from Aseel hen (38.7%) and Golden pheasant (29.03%). Trichuris spp. was also more prevalent and isolated from golden pheasant (38.7 %). and other pathogens Ascaris spp. isolated from golden pheasant (16.1%) and peafowl (6.4 %), followed by Ascaridia spp. isolated from polish hen (16.1 %). It is concluded that Pigeons were most infected bird species and Eimeria spp, (38.7 %) and Trichuris spp. (38.7 %) were the most prevalent pathogens. Conclusion: Parasitic load can be managed by providing better healthcare facilities to birds, proper cleaning of waterers, vaccination of birds, proper nutrition and management of environmental conditions.

Assessment of the National Veterinary Laboratory System Capacity for the Detection of Infectious Threats to Global Health Security in Benin

Introduction: The International Health Regulations 2005 require countries to establish laboratory systems for rapid and safe confirmation of public health emergencies including zoonoses. Objectives: This study assessed the national veterinary laboratory system capacity for the confirmation of infectious threats to global health security in Benin. Method: The study was descriptive, cross-sectional and evaluative. The non-probability sampling method with selection of the two existing veterinary laboratories were used. Two questionnaires, one observation grid and the Food and Agriculture Organization laboratory mapping tool-core were used. The system capacity was rated good if each laboratory met at least 80% of the assessed criteria. Otherwise, the capacity was rated insufficient. Results: The national veterinary laboratory system capacity in Benin was insufficient. Bohicon laboratory satisfied 27.2% of assessed criteria; With the laboratory mapping tool-core, Parakou laboratory had an average indicator of 50.7%. Laboratory collaboration and networking was the best performing domain with 66.7%. Only basic supplies category reached 80%. Laboratory budget, sample accession and availaible technology had the lowest scores. The veterinary laboratory system is facing shortage of staffs. Avian influenza virus is confirmed by the two laboratories; Bacillus anthracis and rift valley fever virus are only confirmed by Parakou laboratory. Ebola and Lassa fever are not confirmed. There is no collaboration between the laboratories in the country. Conclusion: The veterinary laboratories are not able to ensure timely detection of zoonoses and inform health system for preparedness. The development and implementation of the veterinary laboratory system improvement plan is essential for the global health security in Benin.

A virtual biobank for companion animals: A parvovirus pilot study.

There is a lack of national population data concerning infectious disease in companion animals. Here, we piloted the feasibility of linking diagnostic laboratories, population surveillance and modern sequencing approaches to extract targeted diagnostic samples from laboratories before they were discarded, as a novel route to better understand national epidemiology of major small animal pathogens. Samples tested for canine or feline parvovirus were requested from a national veterinary diagnostic laboratory and analysed by Sanger or next generation sequencing. Samples were linked to electronic health data held in the SAVSNET database. Sequences obtained from positive samples, together with associated metadata, provided new insights into the recent geographical distribution of parvovirus strains in circulation in the United Kingdom (UK). This collaboration with industry represents a 'National Virtual Biobank' that can rapidly be called on, to efficiently add new layers of epidemiological information of relevance to animal, and potentially human, population health.

The role of national and international veterinary laboratories.

In the field of diagnostic test validation, World Organisation for Animal Health (OIE) Reference Laboratories (RLs) have a pivotal role and provide the international community with impartial advice and support in the selection, development and validation of diagnostic tests, which can be applied to the specialist diseases for which they are designated. National RLs provide an invaluable function in supporting the introduction, ongoing validation and application of validated diagnostic tests in line with international standards. Experienced staff with extensive knowledge of such systems and access to specialist facilities for conducting work are available to monitor changes or advancements in technology. They consider their relevance and value to evolving diagnostic test requirements. Reference Laboratories often have a broad mandate of activity linking research or development programmes and surveillance activities to benefit the continual assessment and, if necessary, improvement of diagnostic tools. Reference Laboratories maintain or have access to unique biological archives (known positive and negative sample populations) and produce international reference standards, both of which are vital in establishing the necessary and detailed validation of any diagnostic test. Reference Laboratories act either singularly or in collaborative partnerships with other RLs or science institutes, but also, when required, and with impartiality, with the commercial sector, to ensure new tests are validated according to OIE standards. They promote and apply formal programmes of quality assurance (including proficiency testing programmes) for newly validated tests, ensuring ongoing monitoring and compliance with standards, or as required set out any limitations or uncertainties. Reference Laboratories publish information on test validation in the scientific literature and on relevant websites, as well as disseminating information at workshops and international conferences. Furthermore, they can offer training in the processes and systems underpinning test validation.

Epidemiology of African Swine Fever in Piggeries in the Center, South and South-West of Cameroon.

African Swine Fever (ASF) is enzootic in Cameroon. A cross-sectional study was conducted in the center, south and south-west regions of Cameroon in order to determine: the knowledge, skills and practices at risk of pig breeders; the prevalence of the disease in piggeries; the genome of the circulating virus. A total of 684 blood samples were collected in 209 farms for RT-PCR and ELISA analyses at the National Veterinary Laboratory (LANAVET) annex in Yaoundé. Prevalences of 15.2% (95CI: 12.5–17.9%) by ELISA, 23.8% (95CI: 20.6–27.0%) by RT-PCR, and 15.2% (95CI: 12.5–17.9%) by ELISA-PCR, were recorded. Of the farmers surveyed, 90% knew about the ASF and 55.3% have already experienced it. The 47.4% of them would not be able to recognize ASF if it occurred and, according to them, the risk of the disease introduction in farms would be 32% linked to the animal health personnel who work on farms. Molecular characterization revealed that only ASF genotype-I variable 19T-RSs is circulating. ASF is still hovering at a risky rate over the pig sector of Cameroon. The control of ASF needs an epidemiological surveillance, a better involvement of all stakeholders, sensitization of breeders and an effective State support for producers.

Detection of highly pathogenic avian influenza in Sekong Province Lao PDR 2018—Potential for improved surveillance and management in endemic regions

Significant global efforts have been directed towards understanding the epidemiology of highly pathogenic avian influenza (HPAI) across poultry production systems and in wild-bird reservoirs, yet understanding of disease dynamics in the village poultry setting remains limited. This article provides a detailed account of the first laboratory-confirmed outbreak of HPAI in the south-eastern provinces of Lao PDR, which occurred in a village in Sekong Province in October 2018. Perspectives from an anthropologist conducting fieldwork at the time of the outbreak, clinical and epidemiological observations by an Australian veterinarian are combined with laboratory characterization and sequencing of the virus to provide insights about disease dynamics, biosecurity, outbreak response and impediments to disease surveillance. Market-purchased chickens were considered the likely source of the outbreak. Observations highlighted the significance of a-lack-of pathognomonic clinical signs and commonness of high-mortality poultry disease with consequent importance of laboratory diagnosis. Sample submission and testing was found to be efficient, despite the village being far from the national veterinary diagnostic laboratory. Extensively raised poultry play key roles in ritual, livelihoods and nutrition of rural Lao PDR people. Unfortunately, mass mortality of chickens due to diseases such as HPAI and Newcastle disease (ND) imposes a significant burden on smallholders in Lao PDR, as in most other SE Asian countries. We observed that high mortality of chickens is perceived by locals as a new 'normal' in raising poultry; this sense of it being 'normal' is a disincentive to reporting of mortality events. Establishing effective people-centred disease-surveillance approaches with local benefit, improving market-biosecurity and veterinary-service support to control vaccine-preventable poultry diseases could all reduce mass-mortality event frequency, improve veterinary-producer relationships and increase the likelihood that mortality events are reported. Priority in each of these aspects should be on working with smallholders and local traders, appreciating and respecting their perspectives and local knowledge.

Development and Evaluation of Epitope-Blocking ELISA for Detection of Antibodies against Contagious Caprine Pleuropneumonia in Goat Sera.

Enzyme linked immunosorbent assays (ELISAs) have been developed for the detection of antibodies against contagious caprine pleuropneumonia (CCPP), the causative agent of which is Mycoplasma capricolum subsp. Capripneumoniae (Mccp). The currently available commercial CCPP competitive ELISA (CCPP cELISA) kit produced and supplied by IDEXX Company (Westbrook, Maine, United States) is relatively expensive for most African laboratories. To address this issue and provide a variety of choices, a sensitive and specific blocking-ELISA (b-ELISA) test to detect antibodies against CCPP was developed. We describe the newly developed CCPP blocking-ELISA based on the blocking of an epitope of a monoclonal antibody (Mccp-25) by a positive serum sample against the Mccp protein coated on a plate. The Percentage Inhibition (PI) cut-off value for the CCPP b-ELISA was set at 50 using 466 CCPP negative and 84 CCPP positive small ruminant sera. Of the negative sera, 307 were obtained from the Botswana National Veterinary Laboratory (BNVL) and 159 from the Friedrich-Loeffler-Institute (FLI) Germany. The 84 positive sera samples came from experimentally vaccinated goats at the AU-PANVAC facility in Debre-Zeit, Ethiopia. The relative diagnostic sensitivity and specificity of the CCPP b-ELISA was 93% and 88%, respectively. This test result indicated good correlation with that of the commercial CCPP cELISA by IDEXX Company (Westbrook, Maine, United States) with a Cohen’s κ agreement of κ agreement of 0.85. The newly developed CCPP b-ELISA will be useful in the detection of antibodies for the diagnosis CCPP and for sero-surveillance during vaccination campaigns.

The experience of contagious bovine pleuropneumonia ring trials in Botswana.

The Botswana National Veterinary Laboratory (BNVL) has conducted ring trials (proficiency testing) on an annual basis since 2010. Proficiency testing is carried out to evaluate the ability of veterinary laboratories to perform serological complement fixation tests (CFTs) and molecular polymerase chain reaction (PCR) tests for the diagnosis of contagious bovine pleuropneumonia (CBPP). In this paper, the authors discuss the experience gained and the lessons learned in coordinating these ring trials over a period of six years, from 2010 to 2015.The number of participating laboratories increased from five in 2010 to 11 in2015. Their performance also improved over this period. The proportion of unsatisfactory results decreased from 40% to 10% for serological testing, while questionable results decreased from 60% to 10%. The proportion of unsatisfactory results for the molecular test decreased from 33% to 0%. Systematic errors (i.e. technical errors or imperfect experimental design) were the principal causes of questionable and unsatisfactory results. An analysis of responses from customer satisfaction surveys conducted annually since 2013 provided valuable information that enabled BNVL to redesign the programme in 2014 and 2015 to improve the overall quality of the proficiency testing programme. Among the changes made were sending freeze-dried sera for CFTs and DNA for PCR instead of sera and liquid cultures.

A Review of Eight High-Priority, Economically Important Viral Pathogens of Poultry within the Caribbean Region.

Viral pathogens cause devastating economic losses in poultry industries worldwide. The Caribbean region, which boasts some of the highest rates of poultry consumption in the world, is no exception. This review summarizes evidence for the circulation and spread of eight high-priority, economically important poultry viruses across the Caribbean region. Avian influenza virus (AIV), infectious bronchitis virus (IBV), Newcastle disease virus (NDV), infectious laryngotracheitis virus (ILTV), avian metapneumovirus (aMPV), infectious bursal disease virus (IBDV), fowl adenovirus group 1 (FADV Gp1), and egg drop syndrome virus (EDSV) were selected for review. This review of serological, molecular, and phylogenetic studies across Caribbean countries reveals evidence for sporadic outbreaks of respiratory disease caused by notifiable viral pathogens (AIV, IBV, NDV, and ILTV), as well as outbreaks of diseases caused by immunosuppressive viral pathogens (IBDV and FADV Gp1). This review highlights the need to strengthen current levels of surveillance and reporting for poultry diseases in domestic and wild bird populations across the Caribbean, as well as the need to strengthen the diagnostic capacity and capability of Caribbean national veterinary diagnostic laboratories.

Epidemiological and histopathological analysis of 40 apocrine sweat gland carcinomas in dogs: a retrospective study

Abstract Introduction: Apocrine sweat gland carcinomas (ASGCs) are malignant neoplasms of dogs and other animals, rarely reported worldwide. The aim of this study was to summarise the occurrence of this cancer in a population of dogs in Poland between 2009 and 2014 with regards to histological features and body location of the tumours, as well as age, sex and breed of the cancer-affected dogs. Material and Methods: The study involved 40 canine ASGC cases diagnosed in five national veterinary pathology laboratories. The material was processed according to routine histological methods. Results: Histological types of the tumours involved simple and complex apocrine carcinoma of cystic/papillary (62.5%), solid (15%), and tubular type (12.5%), as well as apocrine ductal carcinoma (10%). The epidemiological analysis revealed peak incidence of the cancer in dogs between 8 and 14 years of age, with the most commonly affected sites being forelimbs and thorax. The highest number of the cancer cases was diagnosed in mixed breed dogs and German Shepherds; no sex predilection was noted. Conclusion: To the authors’ knowledge, this is the first report recounting the study on canine malignant apocrine sweat gland tumours in Poland providing detailed phenotypical and histological data, which are otherwise rarely described in veterinary literature. This type of cancer appears to be diagnosed more frequently in dogs than in humans. Being an easily accessible material for research, canine ASGCs might serve as a relevant animal model for studies related to pathogenesis of sweat gland tumours.

An appraisal of the Maltese national livestock database with regard to bovines.

The creation of a centralised national livestock database for the islands of Malta and Gozo is of crucial importance for the identification and traceability of bovines. It is also important for compliance with the legal obligations that followed Malta's accession to the European Union in May 2004. This paper describes how the processes of identification, registration and traceability of bovines have changed since Malta's accession. The validation and integration of data originating from different departmental sections (such as the identification and registration section), the slaughterhouse and the National Veterinary Laboratory, ensures that any discrepancies are highlighted and can be investigated. Events recorded in the database enable the compliance and eligibility of bovine producers to be cross-checked when applications for European Union benefits are made. The main drawbacks and weak points of the system include financial costs for the government department, potentially late notification of the births and deaths of newborn calves, and insufficient uptake among bovine producers of the latest technology for notification of events such as births, deaths and movement of bovines.

An appraisal on the occurrence of foot-and-mouth disease virus serotypes in cattle and buffaloes, Pakistan.

Foot-and-mouth disease (FMD) is a livestock disease that has serious economic repercussions. Here, we report the laboratory results for samples from suspected outbreaks that were sent for confirmation of FMD in Pakistan. From January 2010 to December 2011, 132 suspected outbreaks were investigated, and samples from 58 out of the 76 outbreaks sent to the National Veterinary Laboratory (NVL) were positive. The highest proportion of positives were of serotype O (65.52 %), followed by serotype A (24.14 %) and serotype Asia-1 (10.35 %), whereas amongst the samples sent to the World Reference Laboratory, Pirbright, UK (WRL), samples from 48 out of 56 outbreaks were confirmed to be FMD positive, with the following serotypes identified: O (56.25 %), Asia-1 (37.50 %) and A (6.25 %). The outbreaks affected cattle, buffalo and mixed (cattle and buffalo) herds at rates of 91, 70 and 76 %, respectively. The trend of positive outbreaks was higher in the months of winter and late spring (November to April). Although the serotype O isolates and some of the serotype A isolates from the field samples resembled the vaccine strains (r-value ≥ 0.3), this was not the case for the Asia-1 isolates. These results help to improve our understanding of the occurrence and distribution of FMD in cattle and buffaloes in Pakistan and to provide baseline information for the FMD progressive control program in the country.

A study on the sterility, safety, potency and purity of hydropericardium hepatitis syndrome (Angara disease) vaccines

The present study was conducted in order to determine the quality of commercially available Angara disease vaccines. For the experiment, 200 broiler chicks of one day old were reared at National Veterinary Laboratories, Islamabad in isolation units. Chicks were divided into three groups that is, 1, 2, 3 and having 25, 25 and 150 chicks, respectively. Group 1 was used to determine LD50 of the virus in chicks, whereas the chicks of group 2 and 3 were used to assess the safety and potency of the vaccines. The LD50 (lethal dose50) of the virus was determined in 26 days old broiler chickens divided into sub-groups a, b, c, d and e. The LD50 titre of the (10%) viral suspension was prepared from liver extract and determined as 10-2.4 per ml. During investigation before use, the sterility of the vaccines was carried out by culturing on microbiological media. All four vaccines were found free from any contamination. The safety standard of the vaccines was also tested by inoculating in 25 chicks of sub-groups F, G, H, I and J, each contained 5 chicks. The chicks were kept under observation for five days post-vaccination. All four vaccines used in chicks were found safe. During study, the potency of the vaccines was determined by vaccinating 150 chicks of sub-group K, L, M, N and O with dose of 0.2 ml. The vaccinated and non-vaccinated chicks of sub-groups were challenged with viral dose of 2 ml at day 17 post-vaccination to know the protection potency of the vaccines. No any chick showed clinical manifestation of disease up to five days post challenge. On post-mortem examination, lesions of hydropericardium syndrome such as straw colour like fluid was observed around the heart. Other lesions recorded were hydronephrosis and hepatitis. Sera of different groups were checked for demonstration of antibody titration. A higher antibody titre was observed at day 21 as compared to 7 and 14 days post vaccination. Purity of vaccines was also determined by sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE). Multiple proteins bands were seen however, a band at the position of 18.6 Kda was observed in all four vaccines and positive control as well, which was detected as viral protein. All four vaccines showed similar viral protein profile by SDS-PAGE. Key words: Quality, antibody, potency, Angara disease, titre.

Environmental Factors Associated with Nitrate Poisoning in Livestock in Botswana

Some cases of sudden deaths in cattle, sheep and goats reported to the National Veterinary Laboratory, Gaborone were investigated. The clinical signs included: stupor, weakness, staggering, trembling, dyspnoea (difficulty in breathing), salivation, diarrhoea, and convulsions for those in extremis. The most consistent sign was the muddy coloured mucous membranes. Necropsy findings included: marked gastroenteritis and generalized congestion and petichiation in all internal body organs and chocolate brown coloured blood that clotted poorly. The presence of nitrate/nitrite in the blood of live animals, amniotic fluid, urine, ocular fluid, urine, drinking water and consumed forage were demonstrated by the diphenylamine test and subsequently confirmed by Gries test. Further confirmation of nitrate/ nitrite poisoning was by response to therapeutic intervention with methylene blue using the intravenous route of inoculation. Environmental factors that may have contributed to the poisoning episodes included, drought, high ambient temperatures, low cloud cover, soil leaching after flooding, soil moisture content and forage nitrogen content and natural geoformations associated with high nitrate content in the water. This is the first account of confirmed cases of nitrate/ nitrite poisoning in livestock in Botswana.

Parasites of animals in Papua New Guinea recorded at the National Veterinary Laboratory: a catalogue, historical review and zoogeographical affiliations

The catalogue includes more than 700 parasites of domestic and wild animals recorded at the National Veterinary Laboratory, Papua New Guinea, since data began to be gathered at the end of World War 2. It incorporates some information already published and data on parasites, particularly of indigenous fauna, not recorded previously in the country. Wildlife host species include wild pig, deer, bats, murine rodents, marsupials, monotremes, birds, reptiles, amphibians, fishes and invertebrates. The range of parasites in domestic and many wild animals shows great affinity with that found in Australia. Some notable exceptions amongst domestic animal parasites are the endoparasites Trichinella papuae, Capillaria papuensis and Mammomonogamus laryngeus and the economically significant ectoparasites Chrysomya bezziana, Tropilaelaps mercedesae and Varroa jacobsoni that are not recorded in Australia. Unusual host-parasite associations include the larvae of the insects Chrysomya spp. and Lucilia sp., parasites of warm-blooded animals, infesting, respectively, cold-blooded crocodiles and cane toads, and the mammalian mite, Sarcoptes scabiei, on an avian host, cassowaries. No host switching of helminths was seen between domestic and wild animals, or between populations of deer, wild pigs and wallabies when grazing together. The economic importance of certain parasites for domestic animals, the potential threats from introduced or newly-discovered parasites, and the relationship between some parasites and their wildlife hosts, are discussed. Information is presented in two tables: a parasite–host list that includes the location of a parasite in or on a host as well as a list of references of relevance to the country, and a host–parasite list that contains the distribution of the parasites according to province or locality.