Importation trends in antibiotics for veterinary use in Rwanda: A retrospective study between 2019 and 2021.

Estimating antibiotic consumption in animals is fundamental to guiding decision-making and research on controlling the emergence and spread of antibiotic-resistant bacteria in humans, animals, and the environment. This study aimed to establish importation trends of antibiotics for veterinary use in Rwanda between 2019 and 2021. Data was collected from the Rwanda Food and Drugs Authority's database. Quantities of imported antibiotic active ingredients were computed using the information extracted from the issued import licenses. These quantities were subsequently adjusted per animal biomass. In total, 35,291.4 kg of antibiotics were imported into Rwanda between 2019 and 2021, with an annual mean of 11,763.8 ± 1,486.9 kg. The adjustment of imported quantities of antibiotics per animal biomass revealed that 29.1 mg/kg, 24.3 mg/kg, and 30.3 mg/kg were imported in 2019, 2020, and 2021 respectively. A slight but not statistically significant decline in antibiotic importation was noted in 2020 (p-value = 0.547). Most of the imported antibiotics were indicated to be used in food-producing animals (35,253.8 kg or 99.9% of the imported antibiotics). Tetracyclines (17,768.6 kg or 50.3%), followed by sulfonamides (7,865.0 kg or 22.3%) and aminoglycosides (4,071.1 kg or 11.5%), were the most imported antibiotics over the studied period. It was noted that 78.9% of the imported antibiotics were categorized as highly important antimicrobials for human medicine. This study established a generalized overview of the importation of antibiotics for veterinary use in Rwanda. These results can serve as guidance for the control of antibiotic misuse. They can be used to make a correlation between antibiotic importation, antibiotic consumption, and the occurrence of antibiotic resistance in the country.

Trypanosomes infection, endosymbionts, and host preferences in tsetse flies (Glossina spp.) collected from Akagera park region, Rwanda: A correlational xenomonitoring study.

Akagera National Park and its surroundings are home to tsetse flies and a number of their mammalian hosts in Rwanda. A One-health approach is being used in the control and surveillance of both animal and human trypanosomosis in Rwanda. Determination of the infection level in tsetse flies, species of trypanosomes circulating in vectors, the source of tsetse blood meal and endosymbionts is crucial in understanding the epidemiology of the disease in animals and humans in the region. Tsetse flies (n = 1101), comprising Glossina pallidipes (n = 771) and Glossina morsitans centralis (n = 330) were collected from Akagera park and surrounding areas between May 2018 and June 2019. The flies were screened for trypanosomes, vertebrate host DNA to identify sources of blood meal, and endosymbionts by PCR - High Resolution Melting analysis and amplicon sequencing. The feeding frequency and the feeding indices (selection index - W) were calculated to identify the preferred hosts. An overall trypanosome infection rate of 13.9% in the fly's Head and Proboscis (HP) and 24.3% in the Thorax and Abdomen (TA) were found. Eight trypanosome species were identified in the tsetse fly HP and TA, namely: Trypanosoma (T.) brucei brucei, T. congolense Kilifi, T. congolense savannah, T. vivax, T. simiae, T. evansi, T. godfreyi, T. grayi and T. theileri. We found no evidence of human-infective T. brucei rhodesiense. We also identified eighteen species of vertebrate hosts that tsetse flies fed on, and the most frequent one was the buffalo (Syncerus caffer) (36.5%). The frequently detected host by selection index was the rhinoceros (Diceros bicornis) (W = 16.2). Most trypanosome infections in tsetse flies were associated with the buffalo blood meal. The prevalence of tsetse endosymbionts Sodalis and Wolbachia was 2.8% and 4.8%, respectively. No Spiroplasma and Salivary Gland Hypertrophy Virus were detected. These findings implicate the buffaloes as the important reservoirs of tsetse-transmitted trypanosomes in the area. This contributes to predicting the main cryptic reservoirs and therefore guiding the effective control of the disease. The study findings provide the key scientific information that supports the current One Health collaboration in the control and surveillance of tsetse-transmitted trypanosomosis in Rwanda.

Preliminary sampling of aflatoxin M1 contamination in raw milk from dairy farms using feed ingredients from Rwanda.

Milk is susceptible to aflatoxin M1 (AFM1) contamination when dairy cattle consume feed contaminated with aflatoxins and is considered as a public health concern. This pilot study assessed the prevalence and amount of total aflatoxin contamination in commercially available dairy feed and the corresponding AFM1 contamination in raw milk from samples collected at farms using local, commercially available dairy feed across Rwanda's five provinces. The inclusion criteria to select dairy farm participants were (1) to have at least two cows and (2) use of commercially prepared dairy feeds. Importantly, the majority of cattle rearing households in Rwanda rely principally on grazing or other freely available feedstock, rather than on commercially prepared feeds. In total, 170 raw milk samples were collected during one sampling period from dairy farms using commercially prepared dairy feeds. In addition, 154 dairy feed samples were collected simultaneously with the milk samples. These farms were previously targeted in a larger study measuring aflatoxin contamination of Rwandan feeds and feed ingredients. The mean AFM1 concentration in these samples was 0.89 ± 1.64 µg/l (median: 0.33 µg/l) with a maximum of 14.5 µg/l. Maize bran was the principal dairy feed ingredient used by farmers in the sampling, representing more than 65% of the total feed samples collected, with mean aflatoxin concentration of 90.5 µg/kg (median 32.3 µg/kg). The authors note that this preliminary sampling is not generalizable across Rwandan milk production and consumption; the limited pilot study presented here was not designed with the robustness necessary for broad-scale generalization. Thus, the data presented should not be broadly applied outside of the context of the study.

An environmental scan of one health preparedness and response: the case of the Covid-19 pandemic in Rwanda.

BACKGROUND: Over the past decade, 70% of new and re-emerging infectious disease outbreaks in East Africa have originated from the Congo Basin where Rwanda is located. To respond to these increasing risks of disastrous outbreaks, the government began integrating One Health (OH) into its infectious disease response systems in 2011 to strengthen its preparedness and contain outbreaks. The strong performance of Rwanda in responding to the on-going COVID-19 pandemic makes it an excellent example to understand how the structure and principles of OH were applied during this unprecedented situation. METHODS: A rapid environmental scan of published and grey literature was conducted between August and December 2020, to assess Rwanda's OH structure and its response to the COVID-19 pandemic. In total, 132 documents including official government documents, published research, newspaper articles, and policies were analysed using thematic analysis. RESULTS: Rwanda's OH structure consists of multidisciplinary teams from sectors responsible for human, animal, and environmental health. The country has developed OH strategic plans and policies outlining its response to zoonotic infections, integrated OH into university curricula to develop a OH workforce, developed multidisciplinary rapid response teams, and created decentralized laboratories in the animal and human health sectors to strengthen surveillance. To address COVID-19, the country created a preparedness and response plan before its onset, and a multisectoral joint task force was set up to coordinate the response to the pandemic. By leveraging its OH structure, Rwanda was able to rapidly implement a OH-informed response to COVID-19. CONCLUSION: Rwanda's integration of OH into its response systems to infectious diseases and to COVID-19 demonstrates the importance of applying OH principles into the governance of infectious diseases at all levels. Rwanda exemplifies how preparedness and response to outbreaks and pandemics can be strengthened through multisectoral collaboration mechanisms. We do expect limitations in our findings due to the rapid nature of our environmental scan meant to inform the COVID-19 policy response and would encourage a full situational analysis of OH in Rwanda's Coronavirus response.

Occurrence, diversity and distribution of Trypanosoma infections in cattle around the Akagera National Park, Rwanda.

BACKGROUND: African Trypanosomiases threaten the life of both humans and animals. Trypanosomes are transmitted by tsetse and other biting flies. In Rwanda, the African Animal Trypanosomiasis (AAT) endemic area is mainly around the tsetse-infested Akagera National Park (NP). The study aimed to identify Trypanosoma species circulating in cattle, their genetic diversity and distribution around the Akagera NP. METHODOLOGY: A cross-sectional study was carried out in four districts, where 1,037 cattle blood samples were collected. The presence of trypanosomes was determined by microscopy, immunological rapid test VerY Diag and PCR coupled with High-Resolution Melt (HRM) analysis. A parametric test (ANOVA) was used to compare the mean Packed cell Volume (PCV) and trypanosomes occurrence. The Cohen Kappa test was used to compare the level of agreement between the diagnostic methods. FINDINGS: The overall prevalence of trypanosome infections was 5.6%, 7.1% and 18.7% by thin smear, Buffy coat technique and PCR/HRM respectively. Microscopy showed a low sensitivity while a low specificity was shown by the rapid test (VerY Diag). Trypanosoma (T.) congolense was found at a prevalence of 10.7%, T. vivax 5.2%, T. brucei brucei 2% and T. evansi 0.7% by PCR/HRM. This is the first report of T.evansi in cattle in Rwanda. The non-pathogenic T. theileri was also detected. Lower trypanosome infections were observed in Ankole x Friesian breeds than indigenous Ankole. No human-infective T. brucei rhodesiense was detected. There was no significant difference between the mean PCV of infected and non-infected animals (p>0.162). CONCLUSIONS: Our study sheds light on the species of animal infective trypanosomes around the Akagera NP, including both pathogenic and non-pathogenic trypanosomes. The PCV estimation is not always an indication of trypanosome infection and the mechanical transmission should not be overlooked. The study confirms that the area around the Akagera NP is affected by AAT, and should, therefore, be targeted by the control activities. AAT impact assessment on cattle production and information on the use of trypanocides are needed to help policymakers prioritise target areas and optimize intervention strategies. Ultimately, these studies will allow Rwanda to advance in the Progressive Control Pathway (PCP) to reduce or eliminate the burden of AAT.

Genome-Wide Association Study of Growth Performance and Immune Response to Newcastle Disease Virus of Indigenous Chicken in Rwanda.

A chicken genome has several regions with quantitative trait loci (QTLs). However, replication and confirmation of QTL effects are required particularly in African chicken populations. This study identified single nucleotide polymorphisms (SNPs) and putative genes responsible for body weight (BW) and antibody response (AbR) to Newcastle disease (ND) in Rwanda indigenous chicken (IC) using genome-wide association studies (GWAS). Multiple testing was corrected using chromosomal false detection rates of 5 and 10% for significant and suggestive thresholds, respectively. BioMart data mining and variant effect predictor tools were used to annotate SNPs and candidate genes, respectively. A total of four significant SNPs (rs74098018, rs13792572, rs314702374, and rs14123335) significantly (p ≤ 7.6E-5) associated with BW were identified on chromosomes (CHRs) 8, 11, and 19. In the vicinity of these SNPs, four genes such as pre-B-cell leukaemia homeobox 1 (PBX1), GPATCH1, MPHOSPH6, and MRM1 were identified. Four other significant SNPs (rs314787954, rs13623466, rs13910430, and rs737507850) all located on chromosome 1 were strongly (p ≤ 7.6E-5) associated with chicken antibody response to ND. The closest genes to these four SNPs were cell division cycle 16 (CDC16), zinc finger, BED-type containing 1 (ZBED1), myxovirus (influenza virus) resistance 1 (MX1), and growth factor receptor bound protein 2 (GRB2) related adaptor protein 2 (GRAP2). Besides, other SNPs and genes suggestively (p ≤ 1.5E-5) associated with BW and antibody response to ND were reported. This work offers a useful entry point for the discovery of causative genes accountable for essential QTLs regulating BW and antibody response to ND traits. Results provide auspicious genes and SNP-based markers that can be used in the improvement of growth performance and ND resistance in IC populations based on gene-based and/or marker-assisted breeding selection.

An update on the distribution of Glossina (tsetse flies) at the wildlife-human-livestock interface of Akagera National Park, Rwanda.

BACKGROUND: Glossina (tsetse flies) biologically transmit trypanosomes that infect both humans and animals. Knowledge of their distribution patterns is a key element to better understand the transmission dynamics of trypanosomosis. Tsetse distribution in Rwanda has not been well enough documented, and little is known on their current distribution. This study determined the current spatial distribution, abundance, diversity, and seasonal variations of tsetse flies in and around the Akagera National Park. METHODS: A longitudinal stratified sampling following the seasons was used. Biconical traps were deployed in 55 sites for 6 consecutive days of each study month from May 2018 to June 2019 and emptied every 48 h. Flies were identified using FAO keys, and the number of flies per trap day (FTD) was used to determine the apparent density. Pearson chi-square (χ2) and parametrical tests (t-test and ANOVA) were used to determine the variations between the variables. The significance (p < 0.05) at 95% confidence interval was considered. Logistic regression was used to determine the association between tsetse occurrence and the associated predictors. RESULTS: A total of 39,516 tsetse flies were collected, of which 73.4 and 26.6% were from inside Akagera NP and the interface area, respectively. Female flies accounted for 61.3 while 38.7% were males. Two species were identified, i.e. G. pallidipes [n = 29,121, 7.4 flies/trap/day (FTD)] and G. morsitans centralis (n = 10,395; 2.6 FTD). The statistical difference in numbers was significant between the two species (p = 0.000). The flies were more abundant during the wet season (15.8 FTD) than the dry season (4.2 FTD). Large numbers of flies were trapped around the swamp areas (69.1 FTD) inside the park and in Nyagatare District (11.2 FTD) at the interface. Glossina morsitans was 0.218 times less likely to occur outside the park. The chance of co-existing between the two species reduced outside the protected area (0.021 times). CONCLUSIONS: The occurrence of Glossina seems to be limited to the protected Akagera NP and a narrow band of its surrounding areas. This finding will be crucial to design appropriate control strategies. Glossina pallidipes was found in higher numbers and therefore is conceivably the most important vector of trypanosomosis. Regional coordinated control and regular monitoring of Glossina distribution are recommended.

Correction: Genetic diversity and population structure of indigenous chicken in Rwanda using microsatellite markers.

[This corrects the article DOI: 10.1371/journal.pone.0225084.].

Genetic diversity and population structure of indigenous chicken in Rwanda using microsatellite markers.

Rwanda has about 4.5 million of indigenous chicken (IC) that are very low in productivity. To initiate any genetic improvement programme, IC needs to be accurately characterized. The key purpose of this study was to ascertain the genetic diversity of IC in Rwanda using microsatellite markers. Blood samples of IC sampled from 5 agro-ecological zones were collected from which DNA was extracted, amplified by PCR and genotyped using 28 microsatellite markers. A total of 325 (313 indigenous and 12 exotic) chickens were genotyped and revealed a total number of 305 alleles varying between 2 and 22 with a mean of 10.89 per locus. One hundred eighty-six (186) distinct alleles and 60 private alleles were also observed. The frequency of private alleles was highest in samples from the Eastern region, whereas those from the North West had the lowest. The influx of genes was lower in the Eastern agro-ecological zone than the North West. The mean observed heterozygosity was 0.6155, whereas the average expected heterozygosity was 0.688. The overall inbreeding coefficient among the population was 0.040. Divergence from the Hardy-Weinberg equilibrium was significant (p<0.05) in 90% of loci in all the populations. The analysis of molecular variance revealed that about 92% of the total variation originated from variation within populations. Additionally, the study demonstrated that IC in Rwanda could be clustered into four gene groups. In conclusion, there was considerable genetic diversity in IC in Rwanda, which represents a crucial genetic resource that can be conserved or optimized through genetic improvement.

Assessment of Aflatoxin and Fumonisin Contamination and Associated Risk Factors in Feed and Feed Ingredients in Rwanda.

Mycotoxins are fungal metabolites that contaminate crops, food, and animal feeds. Aflatoxins and fumonisins are among the mycotoxins that have been increasingly reported to affect health and productivity of livestock globally. Given that the health and productivity of livestock can directly influence human food safety and security, a study was conducted to assess the levels and factors for aflatoxin and fumonisin contamination in feed and feed ingredients in Rwanda. Aflatoxins and fumonisins were analyzed in 3328 feed and feed ingredient samples collected at six time points between March and October 2017 in all 30 districts of Rwanda. Of the 612 participants providing samples, there were 10 feed processors, 68 feed vendors, 225 dairy farmers, and 309 poultry farmers. Enzyme-Linked Immunosorbent Assay (ELISA) was used for aflatoxin and fumonisin analyses. Mean aflatoxin levels of 108.83 µg/kg (Median (MD): 43.65 µg/kg), 103.81µg/kg (MD: 48.4 µg/kg), 88.64 µg/kg (MD: 30.90 µg/kg), and 94.95 µg/kg (MD: 70.45 µg/kg) were determined for dairy farmers, poultry farmers, feed vendors, and feed processors, respectively. Mean fumonisin levels were 1.52 mg/kg (MD: 0.71 mg/kg), 1.21 mg/kg (MD: 0.56 mg/kg), 1.48 mg/kg (MD: 0.76 mg/kg), and 1.03 mg/kg (MD: 0.47 mg/kg) for dairy farmers, poultry farmers, feed vendors, and feed processors, respectively. Aflatoxin contamination was significantly affected by time of sampling and district from which feed samples originated (p < 0.05). Fumonisins did not show any correlation trends. Ninety-two percent of survey participants were unaware of aflatoxins and fumonisins and their adverse effects. This study has provided the basic understanding of the extent of feed contamination across the country and has established a baseline for future interventions in Rwanda. Further studies are needed to explore strategies for mitigating mycotoxins in the feed value chain in Rwanda.