Cachexia and head and neck squamous cell carcinoma: A scoping review.

OBJECTIVE: The objective of this paper was to provide an understanding of cachexia in relation to oral squamous cell carcinoma relevant to oral health care. The paper is a scoping review of aspects of the clinical presentation, aetiology and management of cachexia in relation to oral health and oral health care. METHODS: A combined search of MEDLINE and EMBASE databases (via OVID) was conducted using the terms ([Head and Neck] OR [Oral Squamous Cell Carcinoma]) AND (Cachexia). Duplicates were removed and results were subsequently limited to studies published between 2000 and 2023, humans and English language. After screening and full-text assessment a total number of 87 studies were included in the review. RESULTS: It is evident that cachexia is a not uncommon feature of patients with advanced malignancy of the head and neck driven by a multitude of mechanisms, induced by the tumour itself, that lead to reduced nutritional intake, increased metabolism and loss of adipose and skeletal tissue. CONCLUSION: While a variety of nutritional, physical, psychological and pharmacological interventions may improve quality and duration of life, ultimately the diagnosis of cachexia in relation to head and neck cancer remains an indicator of poor life expectancy.

Embedding patient and public involvement into a doctoral study: developing a point-of-care HIV testing intervention for dental settings.

Introduction: Patient and Public Involvement (PPI) can have a positive impact on research. PPI can make research more meaningful and appropriate as well as preventing research waste. For decades, patient advocates with HIV have played a key part in public health and research. This article presents the PPI activity undertaken during a doctoral study. The aim of this article is to demonstrate how PPI was embedded into a doctoral study that explored the feasibility of HIV testing in dental settings. Methods: Patients and the public were invited to be involved with the feasibility study through various organisations and charities. A comprehensive PPI activity strategy was devised, and appropriate funding was obtained. Patients and the public were predominantly consulted or collaboratively involved with several aspects of the study. Findings: Patients and the public positively contributed to the intervention development and the resources supporting its implementation. As a result, the study resources (i.e., questionnaire and information leaflets) were easier to read, and the intervention was more appropriate to the needs of patients. Furthermore, the training and focus groups conducted with dental patients and people with HIV benefitted from input of people with lived experience. Conclusions: PPI can be embedded within doctoral studies provided there is sufficient funding, flexibility, and supervisory support. However, PPI activity may be impacted by limited resource and a priori research protocol and funding agreements.

The use of modelling to evaluate and adapt strategies for animal disease control.

Disease is often associated with debilitating clinical signs, disorders or production losses in animals and/or humans, leading to severe socio-economic repercussions. This explains the high priority that national health authorities and international organisations give to selecting control strategies for and the eradication of specific diseases. When a control strategy is selected and implemented, an effective method of evaluating its efficacy is through modelling. To illustrate the usefulness of models in evaluating control strategies, the authors describe several examples in detail, including three examples of classification and regression tree modelling to evaluate and improve the early detection of disease: West Nile fever in equids, bovine spongiform encephalopathy (BSE) and multifactorial diseases, such as colony collapse disorder (CCD) in the United States. Also examined are regression modelling to evaluate skin test practices and the efficacy of an awareness campaign for bovine tuberculosis (bTB); mechanistic modelling to monitor the progress of a control strategy for BSE; and statistical nationwide modelling to analyse the spatio-temporal dynamics of bTB and search for potential risk factors that could be used to target surveillance measures more effectively. In the accurate application of models, an interdisciplinary rather than a multidisciplinary approach is required, with the fewest assumptions possible.

Potential oral manifestations of cardiovascular drugs.

OBJECTIVE: The aim of this work was to determine the frequency and nature of oral manifestations secondary to use of cardiovascular drugs. METHODS: Five hundred and thirty one patients attending an adult cardiology clinic in Saudi Arabia were questioned about the occurrence of oral dryness, dysgeusia, or burning sensation and were clinically evaluated for the presence of oral mucosal or gingival disease. Data were statistically analyzed with chi-squared tests, odds ratios and Student's t-test. RESULTS: Oral symptoms and/or signs were recorded in 75 (14.1%) patients with xerostomia being the most common (7.5%), followed by lichenoid (lichen planus-like) lesions (3.6%) and dysgeusia (1.9%). Xerostomia was significantly more frequent in patients with a history of diabetes mellitus and in female patients (P < 0.05). There were no statistically significant differences (P > 0.05) between patients with or without oral manifestations when age, gender, cardiovascular risk factor, cardiac disease, type of cardiac drug used or the number of medications were assessed. There was a trend for xerostomia to be less frequent in patients receiving therapy with angiotensin converting enzyme inhibitors and a slight trend of xerostomia to be more likely with increased number of non-cardiac and total number of agents per subject. The number of non-cardiac and total medications taken by patients with potential oral manifestations tended to be greater than that of patients without oral manifestations. CONCLUSIONS: The frequency of potential oral manifestations in patients receiving cardiovascular agents was 14.1%. The occurrence and character of the oral manifestations had no significant relation with individual cardiac drugs, although there was a trend for oral manifestations to be likely with increasing number of drugs.

Extensive oral shedding of human herpesvirus 8 in a renal allograft recipient.

INTRODUCTION: Studies were conducted to investigate changes in the extent of human herpesvirus 8 (HHV-8) shedding and diversity of HHV-8 strains in the mouth of a renal allograft recipient who developed cutaneous post-transplantation Kaposi's sarcoma. METHODS: Matched oral and blood samples were obtained from a Saudi Arabian renal allograft recipient from 3 days before to 38 weeks after transplantation, and from his kidney donor. Polymerase chain reaction (PCR) protocols to amplify selected HHV-8 sub-genomic regions were applied to detect and quantify HHV-8 DNA. Sequence diversity was determined by cloning the PCR products and subjecting them to denaturing gradient gel electrophoresis and to nucleotide sequencing. RESULTS: Before transplantation, the recipient was seropositive for anti-HHV-8 immunoglobulin G, but the donor was seronegative; HHV-8 DNA could be detected in the recipient's blood, whole-mouth saliva (WMS) and buccal exfoliates, and the salivary viral load was estimated as 2.6 million genome-copies/ml. Post-transplantation, the recipient's salivary viral load initially increased to 4.1 million genome-copies/ml, and thereafter declined precipitously, coinciding with an increase in the dosage of valaciclovir given; HHV-8 DNA was detected most often in WMS compared with parotid saliva, and buccal and palatal exfoliates. Carriage of multiple HHV-8 strains was evident in blood and oral samples; whereas before transplantation strains belonging to genotypes A1 and A5 were observed, after transplantation genotype A5 strains became dominant and A2 strains emerged. CONCLUSION: Immunosuppression and antiviral prophylaxis may interact to influence the spectrum of oral HHV-8 strains and the extent of post-transplantation HHV-8 shedding into the mouth.

Varicella zoster virus: review of its management.

Varicella zoster virus (VZV) is one of eight herpes viruses known to infect humans. Primary infection causes varicella (chickenpox), after which virus becomes latent. Years later, VZV reactivates and causes a wide range of neurological diseases. The aim of the present report was to critically examine the published literature to evaluate advantages and limitations of therapy of VZV infection in both immunocompetent and immunocompromised patients. Aciclovir (ACV) has been the drug of choice for many years for the treatment of VZV infections. Recently, other antiviral agents have been developed to overcome the low oral bioavailability of ACV, as well as to provide a more flattering dosage regime. Chickenpox is a benign self-limiting disease in the majority of cases and usually no specific treatment is required. Treatment of shingles is indicated to reduce the acute symptoms of pain and malaise, to limit the spread and duration of the skin lesions and to prevent the development of post-herpetic neuralgia. Different classes of drugs have been used for the treatment of post-herpetic neuralgia. The first choice of any of these medications should be guided by the patient's medical health, the likely adverse effects of the drug and the patient's preference.

Randomised controlled trial of the efficacy of HybenX in the symptomatic treatment of recurrent aphthous stomatitis.

BACKGROUND: The treatment of recurrent aphthous stomatitis (RAS) is principally directed towards reducing the pain and duration of each episode of ulceration; however, there remain few agents for which there is definitive evidence of benefit. AIMS: The aims of the present study were to determine the efficacy of HybenX (Epien Medical Inc., Minneapolis, MN, USA), vs another device used for the treatment of RAS (Salicept; Carrington Laboratories Inc., lrving, TX, USA) to reduce the symptoms and duration of RAS and determine the safety of HybenX for this clinical application. MATERIAL AND METHODS: Sixty-three individuals (36 male, 27 female, group median age 25 years, range 17.8-57.9 years) were entered into a prospective randomised controlled trial of HybenX vs an occlusive covering device (Salicept oral patches; Carrington Laboratories). RESULTS: Painful symptoms over a 5-day posttreatment period were reduced by both agents although HybenX was statistically more effective at day 2 than Salicept, and there was a trend for HybenX to cause greater pain reduction than Salicept over this 5-day period. Both agents gave rise to few adverse side effects--a total of nine adverse events in eight patients were recorded. All were unlikely to be related to the treatment devices. HybenX was only applied on one occasion to the HybenX group, while individuals in the Salicept group were able to self medicate as required. The mean number of Salicept patches used per day per subject was three (s.d. 3.3) on day 1 posttreatment, 3.4 (s.d. 3.1) on day 2 and 2.7 (s.d. 1.9) on day 3. Thereafter, the number of applications fell to a mean of 0.8 on day 7. CONCLUSION: It is concluded that HybenX safely and effectively reduces the painful symptoms of RAS.

Onset and progression of clinical manifestations of orofacial granulomatosis.

BACKGROUND: There remain few studies describing in detail the early occurrence and long-term progression of clinical manifestations of orofacial granulomatosis (OFG) in a substantial number of patients. OBJECTIVES: The aim of this study was to determine the early and late clinical manifestations of a large case series of patients with OFG. PATIENTS/METHODS: Clinically relevant data of 49 patients with OFG who attended an Oral Medicine unit in the UK were examined retrospectively. The analyzed parameters included occurrence and typology of initial manifestations at onset and with respect to long-term follow-up. RESULTS: Five major patterns of disease onset were observed. Recurrent facial swelling with/without intra-oral manifestations was the single most common presentation at onset followed by intra-oral ulcers, and other intra-oral and neurological manifestations. The majority of patients later developed a spectrum of additional features. CONCLUSIONS: OFG results in multiple manifestations at different time points. The disease onset is characterized by manifestations other than facial swelling in about half of affected individuals. However, patients can develop cosmetically unacceptable lip/facial swelling at a later stage. Nearly all affected individuals ultimately develop lip/facial swelling while about half of all patients develop oral ulceration.

Molecular markers relevant to myocardial injury following dental extraction in patients with or without coronary artery disease.

OBJECTIVES: The aim of this study was to characterize biological changes following dental extractions in patients with or without coronary artery disease (CAD). MATERIALS AND METHODS: Forty-five patients (36 males and 9 females) referred for dental extraction underwent treatment and provided blood samples before, immediately after, and 24 h after the procedure. A broad array of biomarkers was employed to assess myocardial injury (highly sensitive troponin T, hs-TnT), bacterial burden (LPS endotoxin activity), and systemic inflammation (CRP, fibrinogen, IFN-γ, IL-1ß, IL-6, IL-8, IL-10, IL-12, and TNF-α). RESULTS: Dental extraction in patients with or without CAD was associated with rises in hs-TnT (p = 0.013), hs-CRP (p < 0.001), fibrinogen (p = 0.005), endotoxin activity (p < 0.001), IFN-γ (p < 0.001), IL-6 (p < 0.001), IL-8 (p = 0.011), and IL-12 (p < 0.001) at 24 h compared with immediately post procedure. Changes in systemic inflammation and endotoxin activity were more evident in those with hs-TnT rise. CONCLUSIONS: Simple dental extractions may cause mild increase in hs-TnT, indicating minor myocardial injury in both patients with or without CAD. Acute systemic inflammation and endotoxemia could represent a possible link between invasive dental treatment and increased risk of acute cardiovascular events. These findings indicate that invasive dental treatment (as simple as a single dental extraction) may impact negatively on clinical outcomes in dental patients, especially those with CAD.

Vegetative endocarditis in equids (1994--2006).

BACKGROUND: Endocarditis is a rare heart condition with variable clinical expressions in equids. Risk factors for this disease are incompletely understood. OBJECTIVE: Describe risk factors for endocarditis in equids. ANIMALS: One hundred and fifty-three equids admitted to Liège University, 9 diagnosed with endocarditis and 144 free from endocarditis but admitted to the hospital with a differential diagnosis including this disease. METHODS: Retrospective case-control study. RESULTS: Equids with endocarditis were significantly younger (mean age = 4.84 +/- 5.74 years) than control equids (mean age = 10.8 +/- 7.73 years) (P = .01). No sex or breed predisposition was observed. Animals with hyperthermia (odds ratio [OR] = 24.4; confidence interval [CI] = 1.40-428), synovial distension (OR = 13.4; CI = 3.00-59.8), lameness (OR = 6.52; CI = 1.63-26.1), hyperglobulinemia (OR = 26.4; CI = 3.03-229), hypoalbuminemia (OR = 11.4; CI = 1.34-96.8), hyperfibrinogenemia (OR = 9.81; CI = 1.16-82.7), or leukocytosis (OR = 7.12; CI = 1.40-36.4) presented a significantly higher risk of having endocarditis than control horses. The presence of two of the clinical signs mentioned above significantly increased the probability of a diagnosis of endocarditis (P< or = .05). CONCLUSIONS AND CLINICAL IMPORTANCE: Age is associated with equine endocarditis. The diagnostic value of certain clinical signs and abnormalities in blood parameters in this disease are described.

Little clinical benefit of early systemic aciclovir for treatment of primary herpetic stomatitis.

DATA SOURCES: The Cochrane Oral Health Group's Trials Register; Cochrane Central Register of Controlled Trials, Medline and Embase were used to search for studies. There were no language restrictions. STUDY SELECTION: Randomised controlled trials (RCT) were eligible if they compared aciclovir to placebo in children and young adults (<25 years of age) who had been diagnosed with primary herpetic gingivostomatitis with or without herpes labialis. DATA EXTRACTION AND SYNTHESIS: Two review authors independently and in duplicate screened and extracted information from, and assessed the risk of bias, in the included clinical trials. The Cochrane Collaboration statistical guidelines were followed for data synthesis. RESULTS: Only two clinical trials, one with 72 participants and the other with 20, were included in this review. The second study failed to report details of its methods and was inconsistent in its reporting of the outcomes measured. The first trial, which had a moderate risk of bias, showed that aciclovir was better than placebo at reducing, in children of <6 years, the number with oral lesions [risk ratio (RR), 0.10; 95% confidence interval (CI), 0.02-0.38)], with new extra-oral lesions (RR; 0.04; 95% CI, 0.00-0.65), and difficulty in eating (RR, 0.14; 95% CI, 0.03-0.58) and drinking (RR, 0.11; 95% CI, 0.01-0.83) after 8 days of treatment. Following onset of treatment, three patients from the placebo group were admitted to hospital for rehydration (P 0.11 ). Four children (two from the aciclovir group and two taking placebo) suffered mild gastrointestinal symptoms that resolved spontaneously after 24-48 h without a change in the study regimen. CONCLUSIONS: Two relevant trials were found for this systematic review. Only one was able to provide weak evidence that aciclovir is an effective treatment in children under 6 years of age with primary herpetic gingivostomatitis, in reducing the number of oral lesions, preventing the development of new extra-oral lesions, decreasing the number of individuals experiencing difficulties eating and drinking and reducing hospital admission.

Field cancerization in oral lichen planus.

BACKGROUND: The concept of field cancerization describes the tendency of patients with premalignant and malignant lesions of head and neck mucosal sites to develop multiple carcinomas of the upper aerodigestive tract. Here we address whether this concept should be extended also to patients affected by oral lichen planus (OLP), an inflammatory disorder associated with an increased risk of cancer development. METHODS: Data from a cohort of 45 patients with OLP who subsequently developed severe dysplastic changes and/or oral squamous cell carcinoma were retrospectively reviewed. Patients who presented more than one oral neoplastic event were considered for further data analysis as regards incidence, localization, management and prognosis. RESULTS: Twenty (44.4%) patients were affected by one single neoplastic event while 25 (55.6%) developed multiple and often multifocal oral dysplastic and/or malignant events. In most cases, a careful surveillance programme led to diagnosis and effective treatment of oral neoplasias at an early intraepithelial and microinvasive stage, leading to long-term survival. In some patients, however, additional primary tumours occurred suddenly with rapid invasion, leading to advanced stage diagnosis and poor prognosis. Overall, three patients (12%) died due to malignant oral disease. CONCLUSIONS: Patients with OLP and subsequent development of dysplasia/ oral squamous cell carcinoma are at risk of having multiple and multifocal neoplastic events of the oral cavity, a phenomenon which parallels the concept of field cancerization of traditional head and neck cancers. If detected at an early stage, these neoplasias can be managed with superficial and complete resection. However a small number of patients have loco-regional tumour spread despite a standard surveillance protocol.

Oral manifestations of systemic disease.

While the majority of disorders of the mouth are centred upon the direct action of plaque, the oral tissues can be subject to change or damage as a consequence of disease that predominantly affects other body systems. Such oral manifestations of systemic disease can be highly variable in both frequency and presentation. As lifespan increases and medical care becomes ever more complex and effective it is likely that the numbers of individuals with oral manifestations of systemic disease will continue to rise. The present article provides a succinct review of oral manifestations of systemic disease. In view of this article being part of a wider BDJ themed issue on the subject of oral medicine, this review focuses upon oral mucosal and salivary gland disorders that may arise as a consequence of systemic disease.

Early Adhesion of Candida albicans onto Dental Acrylic Surfaces.

Denture-associated stomatitis is a common candidal infection that may give rise to painful oral symptoms, as well as be a reservoir for infection at other sites of the body. As poly (methyl methacrylate) (PMMA) remains the main material employed in the fabrication of dentures, the aim of this research was to evaluate the adhesion of Candida albicans cells onto PMMA surfaces by employing an atomic force microscopy (AFM) single-cell force spectroscopy (SCFS) technique. For experiments, tipless AFM cantilevers were functionalized with PMMA microspheres and probed against C. albicans cells immobilized onto biopolymer-coated substrates. Both a laboratory strain and a clinical isolate of C. albicans were used for SCFS experiments. Scanning electron microscopy (SEM) and AFM imaging of C. albicans confirmed the polymorphic behavior of both strains, which was dependent on growth culture conditions. AFM force-spectroscopy results showed that the adhesion of C. albicans to PMMA is morphology dependent, as hyphal tubes had increased adhesion compared with yeast cells ( P < 0.05). C. albicans budding mother cells were found to be nonadherent, which contrasts with the increased adhesion observed in the tube region. Comparison between strains demonstrated increased adhesion forces for a clinical isolate compared with the lab strain. The clinical isolate also had increased survival in blood and reduced sensitivity to complement opsonization, providing additional evidence of strain-dependent differences in Candida-host interactions that may affect virulence. In conclusion, PMMA-modified AFM probes have shown to be a reliable technique to characterize the adhesion of C. albicans to acrylic surfaces.

HCV infection: aspects of epidemiology and transmission relevant to oral health care workers.

Hepatitis C virus (HCV) infection is a common worldwide problem, giving rise to long-term viral carriage and risk of chronic hepatic disease, hepatic malignancy and a wide spectrum of immunologically mediated disorders. The present report describes relevant data suggesting that nosocomial transmission to oral health care workers is unlikely, but in view of medical and occupational consequences of such infection, and the absence of long-term effective treatment or vaccine, the oral surgery profession must continue to be vigilant and to maintain the highest standards of infection control procedures to minimize the possible acquisition of HCV during dental treatment.

An audit of the UK national cancer referral guidelines for suspected oral mucosal malignancy.

BACKGROUND AND OBJECTIVE: This study aimed to assess the effectiveness of the UK National Guidelines for identifying patients with potentially malignant oral disease which were introduced in 2000. DESIGN: Retrospective audit. SETTING: The oral medicine unit in a university teaching hospital in London. METHODS: All new referrals over a one year period were retrospectively reviewed in a departmental audit to evaluate guideline effectiveness. Reasons for referral and final diagnosis were compared in a randomly selected sub-population. RESULTS: Four hundred and eighty-seven of 901 new patients referred were classified as having potentially malignant disease from the referral letter. In a randomly selected subgroup of 241 patients, 18 actually had malignant (8) or dysplastic lesions (10). Of 75 patients referred with a persistent oral ulcer, only nine were actually malignant or dysplastic. Eight of 116 patients referred with a white patch and none with red patches were found to have dysplastic or malignant lesions. The criteria failed to identify three carcinomas and two severely dysplastic lesions (15% of the malignant or dysplastic lesions). All of the latter had been referred by primary care physicians with orofacial pain of unknown cause. CONCLUSIONS: UK National Guidelines discriminate poorly between potentially malignant and other oral mucosal disease.

Review article: oral ulcers and its relevance to systemic disorders.

Oral ulceration is a common problem, and is sometimes a marker of gastroenterological disease. Patients with signs or symptoms of oral ulcers are sometimes referred to gastroenterology clinics, however, in most instances the ulcers does not reflect gastrointestinal disease. Indeed, a spectrum of disorders other than those of the gut can give rise to oral mucosal ulcers ranging from minor local trauma to significant local disease such as malignancy or systemic illness. This present article reviews aspects of the aetiology, diagnosis and management of common ulcerative disorders of the oral mucosa.

Behçet's disease: a review.

Behçet's disease (BD) is a multi-system inflammatory disorder dominated clinically by recurrent oral and genital ulceration, uveitis, and erythema nodosum. Behçet's disease runs a chronic course, with unpredictable exacerbations and remissions whose frequency and severity may diminish with time. Behçet's disease typically arises in young adults, although childhood-onset BD has also been reported. The disease can affect both genders and has a worldwide distribution, although it is more prevalent in countries of the ancient Silk Route. The cause of BD remains unknown, although an autoimmune reaction triggered by an infectious agent in a genetically predisposed individual has been suggested. The treatment of BD is symptomatic and empirical, but generally specific to the clinical features of each patient. The majority of affected individuals do not have life-threatening disease, although mortality can be associated with vascular-thrombotic and neurological disease.

Tissue distribution of Epstein-Barr virus genotypes in hosts coinfected by HIV.

BACKGROUND: In healthy people, oral and pharyngeal epithelium preferentially carries Epstein-Barr virus (EBV) belonging to a genotype that possesses three copies of a 29 base-pair repeat in the first intron of the BZLF-1 gene, while peripheral blood mostly carries a genotype that bears two copies. Whether EBV shows differential tropism in HIV-1-coinfected hosts, who are prone to develop oral hairy leukoplakia, has not been studied. METHODS: Tongue scrapings and CD45+-enriched peripheral blood cells of 20 HIV1-infected patients and 40 healthy controls were examined. EBV-specific DNA was amplified from segments in the first intron of the BZLF-1 gene, in exon C of the LMP-1 gene, and the type A/B-specifying domain of the EBNA-3C gene. Size polymorphisms of these amplicons were assessed by agarose gel electrophoresis, and DNA sequence differences among BZLF-1 gene amplicons by single-strand conformation polymorphism analysis. RESULTS: The predominant EBV genotype in peripheral blood as well as tongue carried two copies of the BZLF-1 repeat. In controls, although the BZLF-1 genotype with two copies was exclusively detected in the blood, the genotype with three copies predominated in the tongue. The findings could not be correlated with EBV genotyped according size polymorphisms in the EBNA-3C or LMP-1 genes. DNA sequences of a proportion or all of the clones derived from the BZLF-1 amplicons in the tongues of HIV-1-infected patients were identical to those in the blood. CONCLUSIONS: These findings are consistent with EBV haematogenous superinfection of the tongue of HIV-positive individuals. Such superinfection may precede or lead to the development of oral hairy leukoplakia.

Tracking familial transmission of Kaposi's sarcoma-associated herpesvirus using restriction fragment length polymorphism analysis of latent nuclear antigen.

Intra-familial transmission of Kaposi's sarcoma associated herpesvirus (KSHV) is likely to occur in geographical regions where KSHV infection is highly endemic. Transmission has been studied previously indirectly using serological techniques, however direct documentation of specific transmission routes has yet to be reported. The internal repeat domain (IRD) of the KSHV opening reading frame (ORF) 73 was shown previously to exhibit restriction-fragment length polymorphism (RFLP). Analysis of such polymorphism was undertaken using nested ORF 73 IRD PCR products derived from the blood and mouth rinse samples of individuals in Malawian family groups. The resulting RFLP patterns were unique to an individual and could be compared between family members. In three of eight families studied, identical RFLP patterns were recovered from family members; in the remaining five families, dissimilar RFLP patterns were revealed. Results from RFLP analysis were compared to sequencing data recovered from family members for the first variable region of the hypervariable KSHV ORF K1. Patterns of intra- and extra-familial transmission inferred from ORF K1 sequencing data were corroborated mainly using ORF 73 IRD RFLP analysis.