Evolution and spread of a highly drug resistant strain of Mycobacterium tuberculosis in Papua New Guinea.

BACKGROUND: Molecular mechanisms determining the transmission and prevalence of drug resistant tuberculosis (DR-TB) in Papua New Guinea (PNG) are poorly understood. We used genomic and drug susceptibility data to explore the evolutionary history, temporal acquisition of resistance and transmission dynamics of DR-TB across PNG. METHODS: We performed whole genome sequencing on isolates from Central Public Health Laboratory, PNG, collected 2017-2019. Data analysis was done on a composite dataset that also included 100 genomes previously sequenced from Daru, PNG (2012-2015). RESULTS: Sampled isolates represented 14 of the 22 PNG provinces, the majority (66/94; 70%) came from the National Capital District (NCD). In the composite dataset, 91% of strains were Beijing 2.2.1.1, identified in 13 provinces. Phylogenetic tree of Beijing strains revealed two clades, Daru dominant clade (A) and NCD dominant clade (B). Multi-drug resistance (MDR) was repeatedly and independently acquired, with the first MDR cases in both clades noted to have emerged in the early 1990s, while fluoroquinolone resistance emerged in 2009 (95% highest posterior density 2000-2016). We identified the presence of a frameshift mutation within Rv0678 (p.Asp47fs) which has been suggested to confer resistance to bedaquiline, despite no known exposure to the drug. Overall genomic clustering was significantly associated with rpoC compensatory and inhA promoter mutations (p < 0.001), with high percentage of most genomic clusters (12/14) identified in NCD, reflecting its role as a potential national amplifier. CONCLUSIONS: The acquisition and evolution of drug resistance among the major clades of Beijing strain threaten the success of DR-TB treatment in PNG. With continued transmission of this strain in PNG, genotypic drug resistance surveillance using whole genome sequencing is essential for improved public health response to outbreaks. With occurrence of resistance to newer drugs such as bedaquiline, knowledge of full drug resistance profiles will be important for optimal treatment selection.

STRIVE PNG: using a partnership-based approach in implementation research to strengthen surveillance and health systems in Papua New Guinea.

Successful implementation research requires effective and equitable relationships between policy-makers, researchers and implementers to effect evidence-based systems change. However, mainstream research grant models between Global North and Global South institutions often (unintentionally) reinforce power imbalances between partners, which result in missed opportunities for knowledge and learning exchange between policy-makers, researchers and implementers.This case study, centred on the STRIVE PNG project, describes how a partnership-based approach has been used to establish, maintain and review effective and equitable relationships between 13 partner organizations (independent research institutes, government health agencies and public health laboratories) to strengthen surveillance and health systems in Papua New Guinea (PNG). We provide an overview of key terms (with supporting conceptual frameworks), describe selected partnership processes and tools used within the project, and share observations regarding early outcomes achieved through this approach.

Surveillance of molecular markers of Plasmodium falciparum artemisinin resistance (kelch13 mutations) in Papua New Guinea between 2016 and 2018.

Plasmodium falciparum resistance to artemisinin-based combination therapy (ACT) is a global threat to malaria control and elimination efforts. Mutations in the P. falciparum kelch13 gene (Pfk13) that are associated with delayed parasite clearance have emerged on the Thai-Cambodian border since 2008. There is growing evidence of widespread Pfk13 mutations throughout South-East Asia and they have independently emerged in other endemic regions. In Papua New Guinea (PNG), Pfk13 "C580Y" mutant parasites with reduced in vitro sensitivity to artemisinin have been isolated in Wewak, a port town in East Sepik Province. However, the extent of any local spread of these mutant parasites in other parts of PNG is unknown. We investigated the prevalence of Pfk13 mutations in multiple malaria-endemic regions of PNG. P. falciparum isolates (n = 1152) collected between 2016 and 2018 and assessed for Pfk13 variation by sequencing. Of 663 high quality Pfk13 sequences a total of five variants were identified. They included C580Y, a mutation at a previously documented polymorphic locus: N499K, and three previously undescribed mutations: R471C, K586E and Y635C. All variants were found in single isolates, indicating that these Pfk13 mutations were rare in the areas surveyed. Notably, C580Y was absent from Maprik district, which neighbours Wewak where C580Y mutant parasites were previously identified. The single C580Y isolate was found in the port town of Lae, Morobe Province, a potential entry site for the importation of drug resistant parasites into PNG. Although sample size in this location was small (n = 5), our identification of a C580Y mutant in this second location is concerning, highlighting the urgent need for further surveillance in Lae. Other Pfk13 mutants were rare in PNG between 2016 and 2018. Continued surveillance for molecular markers of drug resistance is critically important to inform malaria control in PNG.

A prospective multicentre diagnostic accuracy study for the Truenat tuberculosis assays.

BACKGROUND: Bringing reliable and accurate tuberculosis (TB) diagnosis closer to patients is a key priority for global TB control. Molbio Diagnostics have developed the Truenat point-of-care molecular assays for detection of TB and rifampicin (RIF) resistance. METHODS: We conducted a prospective multicentre diagnostic accuracy study at 19 primary healthcare centres and seven reference laboratories in Peru, India, Ethiopia and Papua New Guinea to estimate the diagnostic accuracy of the point-of-care Truenat MTB, MTB Plus and MTB-RIF Dx assays for pulmonary TB using culture and phenotypic drug susceptibility testing as the reference standard, compared with Xpert MTB/RIF or Ultra. RESULTS: Of 1807 enrolled participants with TB signs/symptoms, 24% were culture-positive for Mycobacterium tuberculosis, of which 15% were RIF-resistant. In microscopy centres, the pooled sensitivity of Truenat MTB and Truenat MTB Plus was 73% (95% CI 67-78%) and 80% (95% CI 75-84%), respectively. Among smear-negative specimens, sensitivities were 36% (95% CI 27-47%) and 47% (95% CI 37-58%), respectively. Sensitivity of Truenat MTB-RIF was 84% (95% CI 62-95%). Truenat assays showed high specificity. Head-to-head comparison in the central reference laboratories suggested that the Truenat assays have similar performance to Xpert MTB/RIF. CONCLUSION: We found the performance of Molbio's Truenat MTB, MTB Plus and MTB-RIF Dx assays to be comparable to that of the Xpert MTB/RIF assay. Performing the Truenat tests in primary healthcare centres with very limited infrastructure was feasible. These data supported the development of a World Health Organization policy recommendation of the Molbio assays.

Quality microbiological diagnostics and antimicrobial susceptibility testing, an essential component of antimicrobial resistance surveillance and control efforts in Pacific island nations.

PROBLEM: Emerging bacterial antimicrobial (antibiotic) resistance (AMR) is a global threat to human health. However, most lower income countries do not have microbiological diagnostic testing for prompt, reliable confirmation of bloodstream infection and identification of AMR. CONTEXT: Clinicians in Pacific island nations are increasingly challenged by patients who have infection due to antimicrobial-resistant bacteria. Treatment of infection remains empirical because of a lack of diagnostic testing capacity and may follow guidelines that were formulated without reference to local measures of AMR prevalence. There is limited understanding among clinicians of microbiology testing and test interpretation. ACTION: Examine the lessons learnt from pilot laboratory development programmes in two Pacific island nations that focused on establishing standard procedures for micrological diagnostics and antimicrobial susceptibility testing (AST) and on improving the training of clinicians to increase their use of laboratory services. OUTCOME: The pilot programmes addressed a range of logistical difficulties and evaluated two blood culture systems. They also examined and improved internal QC implementation and evaluated the prevalence of AMR. DISCUSSION: Continued development of microbiological diagnostic capability in the Pacific region is paramount. Pacific Island nations need to develop the capability of at least one central laboratory to culture AMR pathogens and subject them to quality-controlled AST or arrange for suitable referral to a nearby country. DISCUSSION: This study demonstrated a persistently high prevalence of three major bacterial STIs across four countries in WHO's Western Pacific Region during nearly two decades. Further strengthening of strategies to control and prevent STIs is warranted.

Antimicrobial resistance in the Pacific Island countries and territories.

Antimicrobial resistance (AMR) is a critical global health threat with a disproportionate impact on low-income and middle-income countries (LMICs) due to their higher burden of infections, reduced laboratory surveillance infrastructure and fewer regulations governing antimicrobial use among humans or animals. While there have been increasing descriptions of AMR within many LMICs in WHO's Western Pacific and South East Asian regions, there remains a paucity of data from Pacific Island countries and territories (PICTs). The PICTs represent 22 predominantly middle-income countries and territories with a combined population of 12 million people and 20 official languages, spread over hundreds of separate islands spanning an area corresponding to more than 15% of the earth's surface. Our paper outlines the present state of the evidence regarding AMR in PICTs-discussing the present estimates of AMR and their accompanying limitations, important drivers of AMR, as well as outlining key priorities and potential solutions for tackling AMR in this region. Significant areas for action include developing National Action Plans, strengthening laboratory surveillance systems and educational activities targeted at both healthcare workers and the wider community. Ensuring adequate funding for AMR activities in PICTs is challenging given competing health and environmental priorities, in this context global or regional funding initiatives such as the Fleming Fund can play a key role.

Cross-Border Movement of Highly Drug-Resistant Mycobacterium tuberculosis from Papua New Guinea to Australia through Torres Strait Protected Zone, 2010-2015.

In this retrospective study, we used whole-genome sequencing (WGS) to delineate transmission dynamics, characterize drug-resistance markers, and identify risk factors of transmission among Papua New Guinea residents of the Torres Strait Protected Zone (TSPZ) who had tuberculosis diagnoses during 2010-2015. Of 117 isolates collected, we could acquire WGS data for 100; 79 were Beijing sublineage 2.2.1.1, which was associated with active transmission (odds ratio 6.190, 95% CI 2.221-18.077). Strains were distributed widely throughout the TSPZ. Clustering occurred more often within than between villages (p = 0.0013). Including 4 multidrug-resistant tuberculosis isolates from Australia citizens epidemiologically linked to the TSPZ into the transmission network analysis revealed 2 probable cross-border transmission events. All multidrug-resistant isolates (33/104) belonged to Beijing sublineage 2.2.1.1 and had high-level isoniazid and ethionamide co-resistance; 2 isolates were extensively drug resistant. Including WGS in regional surveillance could improve tuberculosis transmission tracking and control strategies within the TSPZ.

Characterization of pncA mutations in multi-drug and pyrazinamide resistant Mycobacterium tuberculosis isolates cultured from Queensland migrants and Papua New Guinea residents.

Outbreak of drug resistant tuberculosis in the Western province, Papua New Guinea is a concern to Queensland, Australia due to migration. We performed pncA mutation analysis and genotyping of multi-drug/pyrazinamide (MDR/PZA) resistant isolates from 18 Queensland (Qld) migrants and 81 Papua New Guinea (PNG) residents, to compare with phenotypic evidence of PZA resistance and to evaluate the genotypes obtained from the two countries. Seven different mutations were seen from Qld isolates of which 2 have not been described previously. A cluster of mutations were found between amino acids L35 and S65. Amongst the PNG isolates, 10 mutations were identified, of which 6 were unique and have not been described previously. Majority of the mutations formed 2 clusters, between amino acids Q10 to A20 and W68 to W119. Mutations identified at nucleotide (nt) position 202 and 307 were found to be the most common types, occurring in 25% and 51% of the PNG isolates respectively. The majority of the mutations were seen in MDR/PZA resistant isolates. These mutations could be utilized for direct screening of PZA resistance from PNG patient samples. Genotypic analysis of the isolates showed strong clustering amongst the PNG isolates as opposed to Qld isolates. A diversity of mutations and genotypes were seen amongst the Qld migrant isolates. Majority of PNG isolates had one genotype with two distinct pncA mutation patterns (T202C and T307G) which highlight on-going transmission. pncA mutation analysis provided a satisfactory alternative to PZA culture DST with high positive predictive value and an improved result turnaround time.

A complete high-quality MinION nanopore assembly of an extensively drug-resistant Mycobacterium tuberculosis Beijing lineage strain identifies novel variation in repetitive PE/PPE gene regions.

A better understanding of the genomic changes that facilitate the emergence and spread of drug-resistant Mycobacterium tuberculosis strains is currently required. Here, we report the use of the MinION nanopore sequencer (Oxford Nanopore Technologies) to sequence and assemble an extensively drug-resistant (XDR) isolate, which is part of a modern Beijing sub-lineage strain, prevalent in Western Province, Papua New Guinea. Using 238-fold coverage obtained from a single flow-cell, de novo assembly of nanopore reads resulted into one contiguous assembly with 99.92 % assembly accuracy. Incorporation of complementary short read sequences (Illumina) as part of consensus error correction resulted in a 4 404 064 bp genome with 99.98 % assembly accuracy. This assembly had an average nucleotide identity of 99.7 % relative to the reference genome, H37Rv. We assembled nearly all GC-rich repetitive PE/PPE family genes (166/168) and identified variants within these genes. With an estimated genotypic error rate of 5.3 % from MinION data, we demonstrated identification of variants to include the conventional drug resistance mutations, and those that contribute to the resistance phenotype (efflux pumps/transporter) and virulence. Reference-based alignment of the assembly allowed detection of deletions and insertions. MinION sequencing provided a fully annotated assembly of a transmissible XDR strain from an endemic setting and showed its utility to provide further understanding of genomic processes within Mycobacterium tuberculosis.

Multi-clonal evolution of multi-drug-resistant/extensively drug-resistant Mycobacterium tuberculosis in a high-prevalence setting of Papua New Guinea for over three decades.

An outbreak of multi-drug resistant (MDR) tuberculosis (TB) has been reported on Daru Island, Papua New Guinea. Mycobacterium tuberculosis strains driving this outbreak and the temporal accrual of drug resistance mutations have not been described. Whole genome sequencing of 100 of 165 clinical isolates referred from Daru General Hospital to the Supranational reference laboratory, Brisbane, during 2012-2015 revealed that 95 belonged to a single modern Beijing sub-lineage strain. Molecular dating suggested acquisition of streptomycin and isoniazid resistance in the 1960s, with potentially enhanced virulence mediated by an mycP1 mutation. The Beijing sub-lineage strain demonstrated a high degree of co-resistance between isoniazid and ethionamide (80/95; 84.2 %) attributed to an inhA promoter mutation combined with inhA and ndh coding mutations. Multi-drug resistance, observed in 78/95 samples, emerged with the acquisition of a typical rpoB mutation together with a compensatory rpoC mutation in the 1980s. There was independent acquisition of fluoroquinolone and aminoglycoside resistance, and evidence of local transmission of extensively drug resistant (XDR) strains from 2009. These findings underline the importance of whole genome sequencing in informing an effective public health response to MDR/XDR TB.

Evaluation of Xpert MTB/RIF assay in children with presumed pulmonary tuberculosis in Papua New Guinea.

BACKGROUND: The Gene Xpert MTB/ RIF assay (Xpert) is used for rapid, simultaneous detection of Mycobacterium tuberculosis (MTB) and rifampicin resistance. This study examined the accuracy of Xpert in children with suspected pulmonary tuberculosis (PTB). METHODS: Children admitted to Port Moresby General Hospital with suspected PTB were prospectively enrolled between September 2014 and March 2015. They were classified into probable, possible and TB-unlikely groups. Sputum or gastric aspirates were tested by Xpert and smear microscopy; mycobacterial culture was undertaken on a subset. Children were diagnosed with TB on the basis of standard criteria which were used as the primary reference standard. Xpert, smear for acid-fast bacilli (AFB) and the Edwards TB score were compared with the primary reference standard. RESULTS: A total of 93 children ≤14 years with suspected PTB were enrolled; 67 (72%) were classified as probable, 21 (22%) possible and 5 (5.4%) TB-unlikely. Eighty were treated for TB based on the primary reference standard. Xpert was positive in 26/93 (28%) MTB cases overall, including 22/67 (33%) with probable TB and 4/21 (19%) with possible TB. Three (13%) samples identified rifampicin resistance. Xpert confirmed more cases of TB than AFB smear (26 vs 13, p = 0.019). The sensitivity of Xpert, AFB smear and an Edwards TB score of ≥7 was 31% (25/80), 16% (13/80) and 90% (72/80), respectively, and the specificity was 92% (12/13), 100% (13/13) and 31% (4/13), respectively, when compared with the primary reference standard. CONCLUSION: Xpert sensitivity is sub-optimal and cannot be relied upon for diagnosing TB, although a positive result is confirmatory. A detailed history and examination, standardised clinical criteria, radiographs and available tests remain the most appropriate way of diagnosing TB in children in resource-limited countries. Xpert helps confirm PTB better than AFB smear, and identifies rifampicin resistance. Practical guidelines should be used to identify children who will benefit from an Xpert assay.

High Levels of Transmitted HIV Drug Resistance in a Study in Papua New Guinea.

INTRODUCTION: Papua New Guinea is a Pacific Island nation of 7.3 million people with an estimated HIV prevalence of 0.8%. ART initiation and monitoring are guided by clinical staging and CD4 cell counts, when available. Little is known about levels of transmitted HIV drug resistance in recently infected individuals in Papua New Guinea. METHODS: Surveillance of transmitted HIV drug resistance in a total of 123 individuals recently infected with HIV and aged less than 30 years was implemented in Port Moresby (n = 62) and Mount Hagen (n = 61) during the period May 2013-April 2014. HIV drug resistance testing was performed using dried blood spots. Transmitted HIV drug resistance was defined by the presence of one or more drug resistance mutations as defined by the World Health Organization surveillance drug resistance mutations list. RESULTS: The prevalence of non-nucleoside reverse transcriptase inhibitor transmitted HIV drug resistance was 16.1% (95% CI 8.8%-27.4%) and 8.2% (95% CI 3.2%-18.2%) in Port Moresby and Mount Hagen, respectively. The prevalence of nucleoside reverse transcriptase inhibitor transmitted HIV drug resistance was 3.2% (95% CI 0.2%-11.7%) and 3.3% (95% CI 0.2%-11.8%) in Port Moresby and Mount Hagen, respectively. No protease inhibitor transmitted HIV drug resistance was observed. CONCLUSIONS: The level of non-nucleoside reverse transcriptase inhibitor drug resistance in antiretroviral drug naïve individuals recently infected with HIV in Port Moresby is amongst the highest reported globally. This alarming level of transmitted HIV drug resistance in a young sexually active population threatens to limit the on-going effective use of NNRTIs as a component of first-line ART in Papua New Guinea. To support the choice of nationally recommended first-line antiretroviral therapy, representative surveillance of HIV drug resistance among antiretroviral therapy initiators in Papua New Guinea should be urgently implemented.

The Burden of Drug-Resistant Tuberculosis in Papua New Guinea: Results of a Large Population-Based Survey.

BACKGROUND: Reliable estimates of the burden of multidrug-resistant tuberculosis (MDR-TB) are crucial for effective control and prevention of tuberculosis (TB). Papua New Guinea (PNG) is a high TB burden country with limited information on the magnitude of the MDR-TB problem. METHODS: A cross-sectional study was conducted in four PNG provinces: Madang, Morobe, National Capital District and Western Province. Patient sputum samples were tested for rifampicin resistance by the Xpert MTB/RIF assay and those showing the presence of resistance underwent phenotypic susceptibility testing to first- and second-line anti-TB drugs including streptomycin, isoniazid, rifampicin, ethambutol, pyrazinamide, ofloxacin, amikacin, kanamycin and capreomycin. RESULTS: Among 1,182 TB patients enrolled in the study, MDR-TB was detected in 20 new (2.7%; 95% confidence intervals [CI] 1.1-4.3%) and 24 previously treated (19.1%; 95%CI: 8.5-29.8%) TB cases. No case of extensively drug-resistant TB (XDR-TB) was detected. Thirty percent (6/20) of new and 33.3% (8/24) of previously treated cases with MDR-TB were detected in a single cluster in Western Province. CONCLUSION: In PNG the proportion of MDR-TB in new cases is slightly lower than the regional average of 4.4% (95%CI: 2.6-6.3%). A large proportion of MDR-TB cases were identified from a single hospital in Western Province, suggesting that the prevalence of MDR-TB across the country is heterogeneous. Future surveys should further explore this finding. The survey also helped strengthening the use of smear microscopy and Xpert MTB/RIF testing as diagnostic tools for TB in the country.

Prevalence of patients with acute febrile illnesses and positive dengue NS1 tests in a tertiary hospital in Papua New Guinea.

Because the prevalence of dengue fever in urban settings in Papua New Guinea is unknown, we investigated the presence of dengue using the NS1 antigen test in an outpatient-based prospective observational study at Port Moresby General Hospital. Of 140 patients with acute febrile illnesses, dengue fever was diagnosed in 14.9% (20 of 134; 95% confidence interval [95% CI] = 9.6-22.4). Malaria (2 of 137; 1.5%; 95% CI = 0.3-5.7), chikungunya (3 of 140; 2.1%; 95% CI = 0.6-6.6), and bacterial bloodstream infections (0 of 80; 0%; 95% CI = 0-5.7) were uncommon. Dengue fever should no longer be considered rare in Papua New Guinea.

Multisite evaluation of point of care CD4 testing in Papua New Guinea.

Laboratory-based CD4 monitoring of HIV patients presents challenges in resource limited settings (RLS) including frequent machine breakdown, poor engineering support and limited cold chain and specimen transport logistics. This study assessed the performance of two CD4 tests designed for use in RLS; the Dynal assay and the Alere PIMA test (PIMA). Accuracy of Dynal and PIMA using venous blood was assessed in a centralised laboratory by comparison to BD FACSCount (BD FACS). Dynal had a mean bias of -50.35 cells/µl (r(2) = 0.973, p<0.0001, n = 101) and PIMA -22.43 cells/µl (r(2)= 0.964, p<0.0001, n = 139) compared to BD FACS. Similar results were observed for PIMA operated by clinicians in one urban (n = 117) and two rural clinics (n = 98). Using internal control beads, PIMA precision was 10.34% CV (low bead mean 214.24 cells/µl) and 8.29% (high bead mean 920.73 cells/µl) and similar %CV results were observed external quality assurance (EQA) and replicate patient samples. Dynal did not perform using EQA and no internal controls are supplied by the manufacturer, however duplicate testing of samples resulted in r(2) = 0.961, p<0.0001, mean bias =  -1.44 cells/µl. Using the cut-off of 350 cells/µl compared to BD FACS, PIMA had a sensitivity of 88.85% and specificity of 98.71% and Dynal 88.61% and 100%. A total of 0.44% (2/452) of patient samples were misclassified as "no treat" and 7.30% (33/452) "treat" using PIMA whereas with Dynal 8.91% (9/101) as "treat" and 0% as "no treat". In our setting PIMA was found to be accurate, precise and user-friendly in both laboratory and clinic settings. Dynal performed well in initial centralized laboratory evaluation, however lacks requisite quality control measures, and was technically more difficult to use, making it less suitable for use at lower tiered laboratories.

Quality of antimalarial drugs and antibiotics in Papua New Guinea: a survey of the health facility supply chain.

BACKGROUND: Poor-quality life-saving medicines are a major public health threat, particularly in settings with a weak regulatory environment. Insufficient amounts of active pharmaceutical ingredients (API) endanger patient safety and may contribute to the development of drug resistance. In the case of malaria, concerns relate to implications for the efficacy of artemisinin-based combination therapies (ACT). In Papua New Guinea (PNG), Plasmodium falciparum and P. vivax are both endemic and health facilities are the main source of treatment. ACT has been introduced as first-line treatment but other drugs, such as primaquine for the treatment of P. vivax hypnozoites, are widely available. This study investigated the quality of antimalarial drugs and selected antibiotics at all levels of the health facility supply chain in PNG. METHODS AND FINDINGS: Medicines were obtained from randomly sampled health facilities and selected warehouses and hospitals across PNG and analysed for API content using validated high performance liquid chromatography (HPLC). Of 360 tablet/capsule samples from 60 providers, 9.7% (95% CI 6.9, 13.3) contained less, and 0.6% more, API than pharmacopoeial reference ranges, including 29/37 (78.4%) primaquine, 3/70 (4.3%) amodiaquine, and one sample each of quinine, artemether, sulphadoxine-pyrimethamine and amoxicillin. According to the package label, 86.5% of poor-quality samples originated from India. Poor-quality medicines were found in 48.3% of providers at all levels of the supply chain. Drug quality was unrelated to storage conditions. CONCLUSIONS: This study documents the presence of poor-quality medicines, particularly primaquine, throughout PNG. Primaquine is the only available transmission-blocking antimalarial, likely to become important to prevent the spread of artemisinin-resistant P. falciparum and eliminating P. vivax hypnozoites. The availability of poor-quality medicines reflects the lack of adequate quality control and regulatory mechanisms. Measures to stop the availability of poor-quality medicines should include limiting procurement to WHO prequalified products and implementing routine quality testing.

Histological characteristics of 21 Papua New Guinean children with high-grade B-cell lymphoma, which is frequently associated with EBV infection.

The aim of the present study was to confirm the histopathological features of aggressive B-cell lymphoma in Papua New Guinea (PNG)-an EBV endemic region. The immunophenotypic features and expression of EBV-encoded proteins and RNA in B-cell lymphomas were analyzed in 21 PNG children, and compared to the corresponding features of 17 Japanese children with Burkitt lymphoma (BL). Histological diagnosis of the lymphomas from the PNG children was BL in nine patients; atypical Burkitt/Burkitt-like variant of BL (BLL) in three; diffuse large B-cell lymphoma (DLBCL) in four; and B-lymphoblastic lymphoma (B-LBL) in five. The lymphomas from the PNG children had a high positive rate on EBV-RNA in situ hybridization (EBV-ISH; 66.7%). With regard to the histological typing, 10 of 12 patients (83%) with BL/BLL, one of four (25%) with DLBCL, and three of five (60%) with B-LBL were positive for EBV-ISH. The findings of EBV-positive B-LBL were surprising because it is commonly considered that lymphoblastic lymphoma is not associated with EBV. EBV positivity was not detected in the 12 Japanese patients who were available for the EBV-ISH evaluation. It is concluded that it is possible that a proportion of DLBCL and B-LBL besides BL/BLL are associated with EBV in endemic region.

Burkitt lymphoma in Papua New Guinea--40 years on.

BACKGROUND: Burkitt lymphoma is common in tropical Africa and Papua New Guinea, where it has been reported to account for 16% of all childhood malignancies. AIM: This study aimed to compare the geographical distribution of recent cases and their anatomical site of presentation with findings from previous studies, and to determine survival using the current treatment protocol. METHODS: The study included all cases of Burkitt and Burkitt-like lymphoma in children up to 14 years of age diagnosed between January 1998 and December 2003. RESULTS: Thirty-six children were diagnosed with Burkitt lymphoma, accounting for 50% of all lymphomas and 13% of all childhood malignancies. The median age was 6 years (interquartile range 4-8 years) and the male:female ratio was 8:1. Facial structures were the most commonly affected sites, accounting for 21 (58%) cases, followed by spinal involvement in three. The majority (89%) of patients came from malaria-holo-endemic, coastal PNG and three were from the highland region. The national incidence was 1.7/100,000 but provincial rates varied, the highest of 13.4/100,000 being in Gulf province. Only two of the 16 patients who received chemotherapy were known to be in remission at 12 months. CONCLUSIONS: Burkitt tumour remains a common childhood malignancy in PNG. There is a need to improve diagnosis and reporting so that treatment can be started early. The most appropriate treatment regimen for use in PNG and other resource-poor countries remains to be determined.

The aetiology of severe anaemia among antenatal patients of the Port Moresby General Hospital.

Between October 1998 and September 2000, 111 consecutive pregnant patients admitted to the Port Moresby General Hospital antenatal ward with a haemoglobin level of 6 g/dl or less were studied. The main causes of the severe anaemia were as follows: iron deficiency on its own or in combination with another factor 66%--iron deficiency on its own 43% and combined folate and iron deficiency 23%--and folate deficiency 18%. Malaria was a contributory factor in 13 patients (12%). A combination of blood film, bone marrow study, serum assays of ferritin, folate and vitamin B12, and mean corpuscular volume (MCV) was used to determine the cause of the anaemia. Ferritin levels on their own poorly correlated with the presence of iron in the bone marrow. A low MCV correlated well with iron deficiency anaemia while a high MCV was associated with folic acid deficiency. It would seem therefore that while a bone marrow study is mandatory to reach a definitive diagnosis of severe anaemia, MCV, in conjunction with the red cell morphology on blood film, would be a good marker for iron and folic acid deficiency anaemia, especially as we do not have serum assays readily available for folate, ferritin and vitamin B12 in Papua New Guinea.