Foot-and-mouth Disease Serotype Research Articles

Seroprevalence and Associated Risk Factors of Foot and Mouth Disease in Cattle in West Shewa Zone, Ethiopia.

Foot and mouth disease (FMD) is a highly contagious viral disease of cloven-hoofed animals and one of the endemic diseases in Ethiopia. The study was aimed to estimate the seroprevalence and to assess associated risk factors of foot and mouth disease seroprevalence in West Shewa Zone. A total of 384 sera samples were collected from randomly selected cattle and tested using ELISA for antibodies against nonstructural proteins of foot and mouth disease viruses based on IDEXX FMD Multispecies Ab Test (IDEXX Laboratories Inc, USA). The seroprevalence of foot and mouth disease in West Shewa Zone was found to be 40.4% (95% CI: 35.46–45.27) at an animal and 74.7% (95% CI: 65.58–83.85) at the herd level. Multivariable logistic regression analysis indicated that districts, breed, and animal composition were the potential risk factors of FMD seropositivity. Accordingly, cattle found in Abuna Ginde Beret (odds ratio (OR): 9.1, 95% CI: 2.4–34.1, p=0.001), Cheliya (OR: 8.8, 95% CI: 2.5–31.3, p=0.001), Bako Tibe (OR: 7.6, 95% CI: 2.1–28.3, p=0.002), Tokekutaye (OR: 5.8, 95% CI: 1.7–19.5, p=0.004), and Jeldu (OR: 5.3, 95% CI: 1.3–21.5, p=0.020) districts were more at risk to be infected with FMD than cattle from Ambo. The odds of FMD seropositivity was significantly higher in cattle kept with small ruminants (OR: 2.1, 95% CI: 1.3–3.3, p=0.003) than cattle alone. The analysis also revealed that the odds of seropositivity were 6 times higher in crossbred compared with local cattle (p=0.003). The current study found high seroprevalence of FMD in West Shewa Zone. Therefore, cattle should be vaccinated regularly after the identification of specific FMD serotypes circulating in the study area.

The Development and Validation of a Novel Nanobody-Based Competitive ELISA for the Detection of Foot and Mouth Disease 3ABC Antibodies in Cattle.

Effective management of foot and mouth disease (FMD) requires diagnostic tests to distinguish between infected and vaccinated animals (DIVA). To address this need, several enzyme-linked immunosorbent assay (ELISA) platforms have been developed, however, these tests vary in their sensitivity and specificity and are very expensive for developing countries. Camelid-derived single-domain antibodies fragments so-called Nanobodies, have demonstrated great efficacy for the development of serological diagnostics. This study describes the development of a novel Nanobody-based FMD 3ABC competitive ELISA, for the serological detection of antibodies against FMD Non-Structural Proteins (NSP) in Uganda cattle herds. This in-house ELISA was validated using more than 600 sera from different Uganda districts, and virus serotype specificities. The evaluation of the performance of the assay demonstrated high diagnostic sensitivity and specificity of 94 % (95 % CI: 88.9–97.2), and 97.67 % (95 % CI: 94.15–99.36) respectively, as well as the capability to detect NSP-specific antibodies against multiple FMD serotype infections. In comparison with the commercial PrioCHECK FMDV NSP-FMD test, there was a strong concordance and high correlation and agreement in the performance of the two tests. This new developed Nanobody based FMD 3ABC competitive ELISA could clearly benefit routine disease diagnosis, the establishment of disease-free zones, and the improvement of FMD management and control in endemically complex environments, such as those found in Africa.

Serological Detection of FMD Serotypes by New Prepared Innovative Recombinant Hepta-Epitopic Peptide

Background: Foot-and-mouth disease (FMD) is a highly contagious and economically important disease that affects cloven-hoofed animals worldwide. In recent years, a series of outbreaks of FMD have occurred in many countries. Recombinant protein synthesis incorporating protective B- and T-cell epitopes are candidates for new safer and more effective (FMD) vaccines that have potential to provide protective immunity against diverse FMDV strains and to protect against future epidemics. Objectives: The aim of this study was to produce Recombinant Hepta-epitope peptide for detecting FMD O, A , Asia1 serotypes ,which can be used as a tool for diagnostic kits. Methods: In the present study, we designed and produced a Recombinant Hepta-epitopic peptide from FMDV epitopes of the viral proteins VP1, VP2, VP3, 3C (seven epitopic regions, amino acid residues VP1 140-160, Vp1 200-213, VP2 68-75 ,VP2 179-198 ,VP3129-148 ,VP3 190-199 , 3C121-135). Results: It was shown that the mentioned recombinant peptide could recognize the serum collected from cattle infected with FMD serotype A and O. Conclusion: Therefore we believe that this recombinant Hepta-epitopic peptide can be used for diagnostic serological assays. Furthermore this Recombinant peptide may be a potential candidate as an alternative vaccine against FMDV epidemic variants.

Spatial pattern of foot-and-mouth disease in animals in China, 2010-2016.

Foot-and-mouth disease (FMD) is a highly contagious disease of cloven-hoofed animals. An outbreak of FMD can produce devastating economic losses for a considerable length of time. In order to investigate the distribution characteristics of FMD in China, data from 2010 to 2016 were collected, including information on 65 outbreaks of FMD (25 by serotype A and 40 by serotype O), and 5,937 diseased animals (1,691 serotype A and 4,284 serotype O cases). Spatial autocorrelation, including global spatial autocorrelation and local spatial autocorrelation, as well as directional distribution analysis, were performed. Global spatial autocorrelation analysis of FMD cases from 2010 to 2016 did not show clustering (P > 0.05). In 2013 and 2014, the FMD serotype A hotspots areas were Tibet (Z = 3.3236, P < 0.001 in 2013; Z = 3.2001, P < 0.001 in 2014) and Xinjiang provinces (Z = 4.2113, P < 0.001 in 2013; Z = 3.9888, P < 0.001 in 2014). The FMD serotype O hotspots areas were: Xinjiang (Z = 2.5832, P = 0.0098) province in 2010; Tibet (Z = 3.8814, P < 0.001) and Xinjiang (Z = 4.9128, P < 0.001) provinces in 2011; and Tibet (Z = 3.0838, P = 0.0020), Xinjiang (Z = 3.8705, P < 0.001) and Qinghai (Z = 2.8875, P = 0.0039) provinces in 2013. The distribution of FMD cases from 2010 to 2016 showed a significant directional trend (northwest-southeast). In conclusion, our findings revealed the spatial patterns of FMD cases, which may provide beneficial information for the prevention and control of FMD.

Serological, hematological, Biochemical and Oxidative Markers During Foot and Mouth Disease Serotype ‘O’ Infection, Egypt

Foot and mouth disease (FMD) is an extremely grave communicable disease of livestock. It affects all wild and domestic animals with cloven hoof. It is caused by Aphtho virus (Apthous fever) or (FMDV) foot and mouth disease virus which is originated from family Picornaviridae. 30 adult female water buffaloes, 3-5 years old infected with FMD serotypes, O. These animals were located at Sharkia governorate, Egypt during the period beetwen December 2014 to March 2015. Hematological findings showed no significant change in erythrogram and reduction in total leukocytes in the early stage of FMDV infection. Moreover development of macrocytic normochromic anemia and increase in total leukocytes and lymphocytic counts was reported in the late stage of infection. A significant decrease in cholesterol , progesterone , total proteins, albumin , globulins, calcium and sodium levels in infected groups, while a significant increase in serum activities of ALT ,AST, glucose, total, direct ,indirect bilirubine, phosphorous potassium, NO. MDA, CK-MB, LDH and CTNI. Without alterations in creatinine level.

Selection of vaccine strains for serotype O foot-and-mouth disease viruses (2007-2012) circulating in Southeast Asia, East Asia and Far East.

Foot-and-mouth disease (FMD) is endemic in Southeast Asia (SEA) and East Asia with circulation of multiple serotypes and multiple genotypes within each serotype of the virus. Although countries like Japan and South Korea in the Far East were free of FMD, in 2010 FMD serotype O (O/Mya-98) outbreaks were recorded and since then South Korea has experienced several FMD outbreaks despite regular vaccination. In this study a total of 85 serotype O FMD viruses (FMDV) isolated from 2007 to 2012 from SEA, East Asia and Far East were characterized by virus neutralisation tests using antisera to four existing (O/HKN/6/83, O/IND/R2/75, O/SKR/2010 and O/PanAsia-2) and one putative (O/MYA/2009) vaccine strains, and by full capsid sequencing. Serological studies revealed broad cross-reactivity with the vaccine strains; O/PanAsia-2 exhibited a good match with 95.3%, O/HKN/6/83 with 91.8%, O/IND/R2/75 with 80%, and the putative strain O/MYA/2009 with 89.4% isolates employed in this study. Similarly O/PanAsia-2 and O/IND/R2/75 vaccines showed a good match with all eight viruses belonging to O-Ind-2001d sublineage whereas the vaccines of O/Mya-98 lineage, O/MYA/2009 and O/SKR/2010 exhibited the lowest match indicating their unsuitability to protect infections from O-Ind-2001d viruses. A Bayesian analysis of the capsid sequence data indicated these circulating viruses (n = 85) to be of either SEA or Middle East-South Asian (ME-SA) topotype. The ME-SA topotype viruses were mainly detected in Lao PDR, Vietnam, Myanmar and Thailand reflecting the trade links with the Indian subcontinent, and also within the SEA countries. Implications of these results in the context of FMD control in SEA and East Asian countries are discussed.

Evaluation of foot and mouth vaccination for yak (Bos grunniens) in Pakistan.

In northern Pakistan, many farming communities rely on domestic yak (Bos grunniens) as a principle source of income. A 2006 participatory disease surveillance report from this region indicated that foot-and-mouth disease (FMD) is the most prevalent annual disease of yak. Our objectives of this study were to determine exposure levels of yak to FMD virus; implement a vaccination program based on current, regional FMD virus serotypes and subtypes; and quantify immune responses following vaccination. Blood samples were used to determine pre-vaccination exposure of animals to FMD virus by antibody presence to non-structural proteins of FMD virus using a 3-ABC trapping indirect ELISA. Vaccine used consisted of FMD serotypes 'O' (PanAsia-2), 'A' (Iran-05), and 'Asia-1' (Shamir), but changed later during the study to match newly circulating viruses in the country ('O'-PanAsia-2; 'A'-Turk-06 and Asia-1-Sindh-08). Three hundred sixty-three blood samples were tested from selected villages to determine pre-vaccination FMD virus exposure in yak with an average of 37.7%. Immune responses from initial vaccination and booster dose 30days later showed clear protective levels (as mean percent inhibition) of antibodies against structural proteins of serotypes 'O,' 'A,' and 'Asia-1.' These responses remained above threshold positive level even at day 210 following initial vaccination. Results of sero-surveillance and anecdotal information of repeated FMD outbreaks demonstrate the persistence of FMD virus of yak in northern Pakistan. Laboratory results and field observations clearly indicated that yak can be protected against FMD with a good quality vaccine with FMD serotype(s) matching current, regionally circulating FMD virus.

Tracing the antibody mediated acquired immunity by Foot and Mouth disease and Rift Valley Fever combined vaccine in pregnant ewes and their lambs

Aim: The aim of this study is to provide adequate protection to ewes and their lambs against Foot and Mouth disease (FMD) and Rift Valley Fever (RVF). Materials and Methods: A combined inactivated FMD and RVF oil vaccine was prepared successfully. Such vaccine was found to be free from foreign contaminants, safe and potent as determined by quality control tests such as challenge protection percentage for FMD and mice ED 50 for RVF. Vaccination of pregnant ewes with the prepared combined vaccine and determination of the antibody level via serum neutralization test (SNT) and Enzyme Linked immune sorbent assay (ELISA) in the vaccinated pregnant ewes and their lambs. Results: Vaccination of pregnant ewes revealed that these ewes exhibited high levels of specific antibodies against the included vaccine antigens (Foot and Mouth disease virus type A Iran O5, O PanAsia and SAT2/EGY/2012 and RVFV-ZH501). FMD antibodies recorded their peaks by the 10 th week post vaccination while those of RVF recorded their peaks by the 12 th week post vaccination then all antibodies began to decrease gradually to reach their lowest protective titers for FMD by the 32 nd week post vaccination and those for RVF by the 34 th week post vaccination. Potency test of the prepared combined vaccine expressed as protection percentage of vaccinated sheep against target virulent FMD virus serotypes reflected a protection percentage of 80% against type O and SAT2 and 100% against A while for RVF, the mice ED 50 was found to be 0.009 indicating the potency of the prepared vaccine. The antibody titer in serum and colostrum of vaccinated pregnant ewes at day of parturition (10-12 week post vaccination) recorded a high titer against FMD serotype (O), serotype (A), serotype (SAT2) and against RVF. It was noticed that the colostrum antibody titers were slightly higher than those in the sera of vaccinated ewes at time of parturition. The newly born lambs from vaccinated ewes, exhibited good levels of maternal immunity against the included antigens through suckling their mother colostrum by the 24 hrs. post parturition. The newly born lambs recorded their highest level by 1 week of birth and lasted within a protective level up to 11-12 week of birth. Conclusion: Using of FMD/RVF combined vaccine provided a sufficient immune status for pregnant ewes as well as for their lambs. Vaccination of newly born lambs should be carried out by the 4 th month of birth.

Antigenic variation of foot-and-mouth disease virus serotype A

The current measures to control foot-and-mouth disease (FMD) include vaccination, movement control and slaughter of infected or susceptible animals. One of the difficulties in controlling FMD by vaccination arises due to the substantial diversity found among the seven serotypes of FMD virus (FMDV) and the strains within these serotypes. Therefore, vaccination using a single vaccine strain may not fully cross-protect against all strains within that serotype, and therefore selection of appropriate vaccines requires serological comparison of the field virus and potential vaccine viruses using relationship coefficients (r1 values). Limitations of this approach are that antigenic relationships among field viruses are not addressed, as comparisons are only with potential vaccine virus. Furthermore, inherent variation among vaccine sera may impair reproducibility of one-way relationship scores. Here, we used antigenic cartography to quantify and visualize the antigenic relationships among FMD serotype A viruses, aiming to improve the understanding of FMDV antigenic evolution and the scope and reliability of vaccine matching. Our results suggest that predicting antigenic difference using genetic sequence alone or by geographical location is not currently reliable. We found co-circulating lineages in one region that were genetically similar but antigenically distinct. Nevertheless, by comparing antigenic distances measured from the antigenic maps with the full capsid (P1) sequence, we identified a specific amino acid substitution associated with an antigenic mismatch among field viruses and a commonly used prototype vaccine strain, A22/IRQ/24/64.

Geo-spatial distribution of serologically detected bovine Foot and Mouth Disease (FMD) serotype outbreaks in Ilesha Baruba, Kwara State-Nigeria

In this study, geo-spatial distribution of bovine Foot and Mouth Disease (FMD) outbreaks were investigated to establish the prevalence and distribution of FMD serotypes in Ilesha Baruba, Kwara state-Nigeria. This purposive study involved 15 FMD outbreaks occurred during December 2010 to March 2011. For geo-spatial analysis, the outbreak locations were indexed using Global Positioning Service (GPS) device (EtrexR). Randomly sampled bovine sera (n=64) from herd representatives were subjected to FMD 3ABC enzyme-linked immunosorbent assay (FMD 3ABC ELISA) and solid-phase competitive ELISA (SP-cELISA), for the screening and serotyping of FMD virus, respectively. The serotypes were indexed with the specific outbreak locations. Through ELISA, the FMD serotypes detected in this study were- serotype O (83%; n=53/64), serotype A (7.8%; n=5/64), serotype vaccine O (1.6%; n=1/64)), and serotype vaccine SAT2 (1.6%; n=1/64). Multiple serotypes were observed in two different combinations; these were O and A (4.7%; n=3/64), and O and SAT2 (1.6%; n=1/64). FMD multiple serotype infections were associated with absence of cross-immunity between serotypes and cross reactivity enhanced by clustered herds, highland study area topography, road and river interconnectivity, possible human settlements, activities and traffic. This study provides baseline information on geo-spatial distribution, and identification of prevalent FMD serotypes in Ilesha Baruba, Kwara state-Nigeria.

Proper Quality Control of Formulated Foot-and-Mouth Disease Vaccines in Countries with Prophylactic Vaccination is Necessary

Vaccination is considered as an important tool to control foot-and-mouth disease (FMD). A good quality vaccine containing relevant serotypes and matching strains is a pre-requisite for vaccination to be effective. The present study investigated the quality of different brands of FMD vaccine available in Pakistan, including three locally produced and two imported products. All the vaccines were found free of bacterial or fungal contamination. No adverse effects were noted in suckling mice and buffalo calves inoculated with the vaccines, showing that the vaccines were sterile and safe. The humoral immune response to the FMD vaccines was determined in buffalo calves for 234days post-vaccination. Very low humoral immune responses against FMD serotypes O, A and Asia 1 viruses were detected to the locally produced vaccines. The imported vaccines, however, elicited a higher antibody response which persisted for a long period in one of the 2 vaccines. The present study highlights the need of assessing an independent vaccine quality control of finished FMD vaccine products.

Spatial trend of Foot and Mouth Disease virus (FMDV) serotypes in cattle and buffaloes, Pakistan

The present study describes the frequency of Foot and Mouth Disease (FMD) virus serotypes (O, A and Asia-1) in major regions (all provinces) of Pakistan using Indirect Sandwich ELISA. Also, spatial distribution of various FMD serotypes and their comparison is discussed. A total of 590 samples (Epithelial tissue) have been analyzed during a period of five years (2005-2009). Out of 590 samples, 180 were found positive, giving an overall confirmation of FMDV about 33.2 %. Of the prevalent serotypes, FMDV 'O' serotype caused most outbreaks (20.7 %), followed by serotype A (6.6 %) and serotype Asia-1 (4.6 %) while there was no positive case of type 'C'. The study clearly showed that the disease was more frequent in the agro-climatic zones than in hilly areas. Based on the data of 590 samples (>50 outbreaks), the overall prevalence of FMDV in cattle and buffaloes in Pakistan was 33.2 %, while in cattle alone, it was 37.1 %, higher than in buffalo (28.7 %). There were eight cases of mixed serotypes infection, indicating the presence of endemic state of disease. Another significant feature was the change over time. In phase-I (2005-2007), there was an overall prevalence of 29.4 %, while the occurrence of the serotype O, A and Asia-1 was 20.4 %, 2.9 % and 4.7 %, respectively. During phase-II (2008-2009), the overall prevalence was 59.21 %, while those of serotype O, A and Asia-1 were 22.4 %, 31.6 % and 4.0 %, respectively. This clearly indicated a shift from serotype O to A, which may help to explain the occurrence of more severe outbreaks, despite vaccination.

Southeast Asian Foot-and-Mouth Disease Viruses in Eastern Asia

Foot-and-mouth disease (FMD) outbreaks recently affected 2 countries (Japan and South Korea) in eastern Asia that were free of FMD without vaccination. Analysis of viral protein 1 nucleotide sequences indicated that FMD serotype A and O viruses that caused these outbreaks originated in mainland Southeast Asia to which these viruses are endemic.

Detection of Foot-and- Mouth Disease Sub-clinical infection in sheep imported from free zones of Georgia during Hajj season 2009 in Kingdom of Saudi Arabia

Foot and mouth disease (FMD) sub-clinically infected animals, are always a threat to susceptible herds. During Hajj season 2009 (1431 Hijri) the kingdom of Saudi Arabia (KSA) imported about 204,583 sheep from FMD free areas from Republic of Georgia through Jeddah Islamic seaport. The animals were clinically free from FMD and authorized as not been previously vaccinated. However, but during the routine laboratory examination of serum samples using FMD-3ABC ELISA some sheep consignments exhibited positivness for FMD anti-bodies. The liquid phase blocking ELISA (LPBE) was performed as a confirmatory test which revealed antibodies against FMD serotype O, the suggesting that animals may be susceptible to FMD infection from any endemic countries passed through during overseas transportation. This study will contribute towards the development of an appropriate strategy for FMD control, including the choice of countries of the animal importation, as well as assist to improve our understanding of the epidemiology of FMD.

Phylogenetic analyses of the polyprotein coding sequences of serotype O foot-and-mouth disease viruses in East Africa: evidence for interserotypic recombination

BackgroundFoot-and-mouth disease (FMD) is endemic in East Africa with the majority of the reported outbreaks attributed to serotype O virus. In this study, phylogenetic analyses of the polyprotein coding region of serotype O FMD viruses from Kenya and Uganda has been undertaken to infer evolutionary relationships and processes responsible for the generation and maintenance of diversity within this serotype. FMD virus RNA was obtained from six samples following virus isolation in cell culture and in one case by direct extraction from an oropharyngeal sample. Following RT-PCR, the single long open reading frame, encoding the polyprotein, was sequenced.ResultsPhylogenetic comparisons of the VP1 coding region showed that the recent East African viruses belong to one lineage within the EA-2 topotype while an older Kenyan strain, K/52/1992 is a representative of the topotype EA-1. Evolutionary relationships between the coding regions for the leader protease (L), the capsid region and almost the entire coding region are monophyletic except for the K/52/1992 which is distinct. Furthermore, phylogenetic relationships for the P2 and P3 regions suggest that the K/52/1992 is a probable recombinant between serotypes A and O. A bootscan analysis of K/52/1992 with East African FMD serotype A viruses (A21/KEN/1964 and A23/KEN/1965) and serotype O viral isolate (K/117/1999) revealed that the P2 region is probably derived from a serotype A strain while the P3 region appears to be a mosaic derived from both serotypes A and O.ConclusionsSequences of the VP1 coding region from recent serotype O FMDVs from Kenya and Uganda are all representatives of a specific East African lineage (topotype EA-2), a probable indication that hardly any FMD introductions of this serotype have occurred from outside the region in the recent past. Furthermore, evidence for interserotypic recombination, within the non-structural protein coding regions, between FMDVs of serotypes A and O has been obtained. In addition to characterization using the VP1 coding region, analyses involving the non-structural protein coding regions should be performed in order to identify evolutionary processes shaping FMD viral populations.

Epidemiological Patterns of Foot-and-Mouth Disease Worldwide

Foot-and-Mouth Disease (FMD) is a clinical syndrome in animals due to FMD virus that exists in seven serotypes, whereby recovery from one sero-type does not confer immunity against the other six. So when considering intervention strategies in endemic settings, it is important to take account of the characteristics of the different serotypes in different ecological systems. FMD serotypes are not uniformly distributed in the regions of the world where the disease still occurs. For example, the cumulative incidence of FMD serotypes show that six of the seven serotypes of FMD (O, A, C, SAT-1, SAT-2, SAT-3) have occurred in Africa, while Asia contends with four sero-types (O, A, C, Asia-1), and South America with only three (O, A, C). Periodically there have been incursions of Types SAT-1 and SAT-2 from Africa into the Middle East. This paper describes the global dynamics for the seven sero-types and attempts to define FMD epidemiological clusters in the different regions of the world. These have been described on a continent by continent basis. The review has reaffirmed that the movement of infected animals is the most important factor in the spread of FMD within the endemically infected regions. It also shows that the eco-system based approach for defining the epidemiological patterns of FMD in endemic, which was originally described in South America, can apply readily to other parts of the world. It is proposed that any coordinated regional or global strategy for FMD control should be based on a sound epidemiological assessment of the incidence and distribution of FMD, identifying risk sources as either primary or secondary endemic eco-systems.

Foot-and-mouth disease in camelids: a review

Foot-and-mouth disease (FMD) in South American camelids, in dromedaries and Bactrians is reviewed. Recent well-executed experimental studies in New World camels indicate that, although the llama and alpaca can be infected with FMD virus (FMDV) by direct contact, they are not very susceptible and do not pose a risk in transmitting FMD to susceptible animal species. They do not become FMDV carriers. Reports on FMD in dromedaries are, however, conflicting. Serological investigations in Africa and the United Arab Emirates (UAE) on thousands of camel sera were negative and experimental infections have been conducted on only a few dromedaries with one serotype and in one country. The design and execution of most of these experiments were poor and therefore the conclusions are questionable. From these investigations, it seems that dromedaries can contract the disease after experimental infection and through close contact with FMD diseased livestock, but do not present a risk in transmitting FMD to susceptible animals. They do not become FMDV carriers. Recent reports from Mongolia describe similar FMD lesions in Bactrian camels. However, so far no samples have tested positive for FMD. To clarify the situation in Bactrians, samples from suspected clinical cases should be tested because other viral vesicular diseases cannot be distinguished from FMD. Thus, further research on the epidemiology of FMD in camelids is necessary. This would include large-scale serological investigations and experimental infections with different FMD serotypes in connection with susceptible contact animals. The Office International des Épizooties (OIE) Code chapter on FMD includes camelids as being susceptible species to FMD, giving the impression that they are similar to cattle, sheep, goats and pigs in their potential involvement in the epidemiology of FMD. This is clearly not the case, and this issue should be re-addressed by the relevant authorities.