Abeir Bashir Hasan Mohamed 1

Abeir Bashir Hasan Mohamed 1. Anwaar A. Y. Kordofani 2. Huda H. Satti3 1. Assistant Professor of Pathology, Sudan. 2. Department of pathology,Faculty of Medicine, Khartoum University, Sudan. 3. Department of pathology,Faculty of Medicine, Khartoum University, Sudan. Abstract Background Visceral leishmaniasis (VL) is a worldwide disease. As WHO reported In 1997, the number of confirmed cases of VL in Sudan exploded, showing a 400 increase over the previous year. Bleeding tendency is known to occur in VL can be very severe. The pathogenesis of this bleeding tendency is still controversial. Objectives to assess the haemostatic changes in patients with VL by assessing PT,APTT,D-dimer and platelet count. Patients and Methods This study was conducted in the period from January through May 2007 at both Elazzaza village in Sinnar state and Tabarakalla village in Gedarif State in Sudan. Thirty five patients diagnosed as having VL either by lymph node or bone marrow aspiration and 70 apparently healthy controls were included in this study. Haemoglobin (Hb), Total white cells count (TWCC), Platelets count (Plt), PT and APTT and D-dimer were assessed in all patients and controls. Results D-dimer levels were elevated in (94.3) of patients. Thrombocytopenia occurred in (89) of patients. A positive correlation was found between the level of D-dimer and the clinical severity of the disease (p). PT and APTT were prolonged in only 9 of patients. D-dimer levels were elevated in 14 of normal controls. Conclusion D-dimer levels were high in most patients with VL (94.3). This may reflect an ongoing process of DIC. It is also concluded that D-dimer test is more sensitive than PT PTT tests in assessing haemostatic disturbances as only (9) of patients showed prolonged PT APTT while (94.3) had elevated levels of D-dimer. High levels of D-dimer may affect the prognosis in patients with visceral leishmaniasis. Elevated D-dimer in some controls could be due to previous or subclinical infections with leishmaniasis. Key words D-Dimer, Haemostatic changes, Disseminated intravascular coagulation. Corresponding Author Dr. Abeir Bashir Hasan Mohamed E mail [email protected] introduction v isceral leishmaniasis is a worldwide disease with an annual incidence of approximately 500 000 cases and overall prevalence of 12 million people. The population at risk is 350 million(1). VL is endemic in the Sudan and has been reported since 1904(2). The main endemic area is in the eastern part of the country and stretches from the western bank of the White Nile to the Sudanese-Ethiopian border in the east and in the north from the upper reaches of Atbara River to an area around Malakal and Sobat River in Upper Nile state in the south. Another focus is around Kapoeta in eastern Equatoria.(3) Other areas where VL has been reported are Darfur and Kordofan. As WHO reported In 1997, the number of confirmed cases of VL in Sudan exploded, showing a 400 increase over the previous year . VL is caused by protozoa of the genus Leishmania, transmitted to humans by sandflies.36 In Sudan, the parasite was isolated from humans and sandflies, and was found to belong to the L.donovani sensus lato cluster. There are few reports of isolation of L.archibaldi and L.infantum from humans and dogs in Gedaref state, Eastern Sudan.(4) The incubation period is usually about 1 or 2 months but may be up to 10 years. Multiplication of leishmaniae takes place in macrophages, especially in the liver and spleen, the bone marrow and lymphoid tissue. The disease is accompanied by malnutrition and immunosuppression. Acute intercurrent pneumococcal infection or tuberculosis is a common complication. Granulocytopenia and thrombocytopenia occur. Anaemia is due to haemolysis, hypersplenism and ineffective erythropoiesis. Serum albumin is low and globulin, mainly IgG, high. Hepatocellular damage and bleeding are late complications.(5) The onset is usually insidious with a low grade fever, the patient remains ambulant, or it may be abrupt with sweating and high intermittent fever, sometimes showing a double rise of temperature in 24 hours. The spleen soon becomes enlarged, often massively hepatomegaly is less marked. If not treated, the patient will become anemic and wasted, frequently with increased pigmentation, especially on the face. Cough and diarrhea develop. In Africa lymphadenopathy is common, and is rarely the only feature. Many subclinical cases occur and go unrecognized for each clinically recognized case.(6) Bleeding tendency is known to occur in VL patients can be very severe. It is usually in the form of epistaxis. The pathogenesis of this bleeding tendency is still controversial. Patients Methods This is a cross-sectional comparative case-control study, conducted at both Elazzaza village in Sinnar state and Tabarakalla village in Gedarif state, both areas recognized as endemic for VL. Data were collected by using a VL suspect form and eligibility form. In the period from 25th of January till 23rd of May 2007. The study involved 35 patients diagnosed as having VL either by lymph node or bone marrow aspiration, not on treatment and 70 apparently healthy controls. Written consent was obtained from patients and controls. For the target population and controls the following tests were done 1. Haemoglobin (Hb)done using cyanmethaemoglobin method 2. Total white cells count (TWCC)done manually using counting chamber. 3. Platelets count (Plt)done manually using well made blood film. 4. PT and APTT 5. D-Dimerdone by using DiaDimer LX-2 test kit (turbidimetric method). Results – All patients were febrile, 23 of them had positive blood film for malaria while 77 were negative.(figure1) -All patients had lymphadenopathy. – 9 of patients were jaundiced. .(figure2) -86 of patients had enlarged spleen and 29 of them had hepatomegalyas well. .(figure34) -17 of patients had Epistaxis at presentation. .(figure5) – All male and female patients had low hemoglobin levels. (table1) -75 of male controls showed normal hemoglobin levels while 25 of them had low hemoglobin levels . .(figure6) -56 of female controls had normal hemoglobin levels while 44 of them had low hemoglobin levels. .(figure7) – 80 of patients had low WBC while 20 had normal WBC.(figure8) -67 of controls had normal WBC while 33 had low WBC.(figure9) – 89 of patients had low platelet counts while 11 had normal platelet counts .(figure10, table 2) -All controls had normal platelets count .(table 3) – 9 of patients had prolonged PT and PTT . .(figure11, 12) – All controls showed normal PT and PTT values.(table 4, 5) .(figure5) -94.3 of patients showed elevated levels of D-Dimer while only 5.7 of them had normal levels . .(figure13), (table 7) – 86 of control had normal levels of D-Dimer while 14 of them showed elevated levels.(table6) SHAPE MERGEFORMAT SHAPE MERGEFORMAT SHAPE MERGEFORMAT SHAPE MERGEFORMAT Table 1 Hb level for patients HbFrequencyPercentValid percentCumulative percentValid 3.5-5.512.92.92.9 5.6-7.51234.334.337.1 7.6-9.51748.648.685.7 9.6-10.5514.314.3100.0 Total35100.0100.0 Table 2 Platelets count for patients PlateletsFrequencyPercentValid percentCumulative percentValid 60,000-90,0001645.845.845.891,000-120,0001131.431.477.1121,000-150,000411.411.488.6151,000411.411.4100.0Total35100.0100.0 Table 3 Platelets count for controls PlateletsFrequencyPercentValid percentCumulative percentValid 150,000-250,0005071.471.471.4251,000-350,0001927.127.198.6351,000-450,00011.41.4100.0Total70100.0100.0 Table 4 Prothrombin Time in controls PT FrequencyPercentValid percentCumulative percentValid 11-132535.735.735.7 13.1-153042.942.978.6 15.1-161521.421.4100.0 Total70100100 Table 5 Partial Thromboplastin Time in controls PTTFrequencyPercentValid percentCumulative percentValid 30-332130.030.030.0 33.1-352332.932.962.9 35.1-371521.421.484.3 37.1-401115.715.7100.0 Total70100100 Table 6 D-Dimer range in controls D-Dimer (g/L)FrequencyPercentValid percentCumulative percentHigh (221-750)1014.314.314.3Normal (27-150)6085.685.6100.0Total70100.0100.0 Table 7 D-Dimer range in patients group D-DimerFrequencyPercentValid percentCumulative percentValid 110-1000720.020.020.01001-2000720.020.040.02001-30001440.040.080.03001-500038.68.688.6 5000411.411.4100.0Total35100.0100.0 Table 8 The relation between D-Dimer, CBC, PT, PTT and Jaundice in patients with D-Dimer 5000 g/L PatientsD-Dimer (g/L)Hb (g/dl)WBC (L)Platelet (L)PT/secPTT/secJaundiceValid 1Valid 54405.6120080,0001340-ve 256273.51800110,0002351ve 362716.080060,0002148ve 471626.51800100,0001436-ve Mean61255.4140087,00017.843.8 Discussion VL is a worldwide disease the incidence of which has significantly increased during the last decades. It is a progressive disease, mortality rates in untreated cases range from 75 to 95. As mentioned in literature and noticed during this study, patients with VL are predisposed to bleeding. In this study platelet counts, activated partial thromboplastin time (APTT), prothrombin time (PT) and D-Dimer level were measured in order to assess the functional status of hemostasis in untreated patients with VL. All studied patients were febrile and anaemic, 80 were leucopenic, and 86 had splenomegaly, while hepatomegaly was found in 29 of them. The prevalence of thrombocytopenia in the studied patients was found to be 89, as 31 patients had low platelet count, 12 of these patients had epistaxis either before or at the time of presentation. The rest did not show any bleeding tendency. On the other hand two patients with a normal platelet counts had epistaxis. This suggests a different mechanism other than thrombocytopenia for the bleeding tendency in VL. Our findings are slightly different from those obtained by Cascio et-al who identified and analyzed the clinical and epidemiological characteristics of 111 consecutive cases of VL from 1980 to 2000 in a Sicilian pediatric hospital retrospectively. In their study fever and splenomegaly were present in all cases and hepatomegaly in 101 (90.1) cases. Thrombocytopenia and anemia were both observed in 78 (70.2) cases and leucopenia in 47 (42.3) cases.(7) One the other hand our findings are completely different from those obtained by with HYPERLINK http//www.ncbi.nlm.nih.gov/sites/entrezDbpubmedCmdSearchTerm22JC3BCttner20C225BAuthor5DitoolEntrezSystem2.PEntrez.Pubmed.Pubmed_ResultsPanel.Pubmed_DiscoveryPanel.Pubmed_RVAbstractPlusJttner C et al who evaluated haemostasis in 19 dogs with natural Leishmania infection, six of them with a history of epistaxis. The results were compared with the results from 24 healthy dogs. Buccal mucosa bleeding time was prolonged in the dogs with leishmaniasis (P 0.002) and most significantly in those with epistaxis (P 0.005). None of the Leishmania-infected dogs had thrombocytopenia, low levels of plasma von Willebrand factor antigen, a prolonged prothrombin time or activated partial thromboplastin time, a low plasma fibrinogen concentration or high serum fibrin degradation products. These results rule out defects of secondary haemostasis or disseminated intravascular coagulation as significant causes of epistaxis in non-complicated leishmaniasis. Histopathology of the nasal mucosa of 10 of the affected dogs, three of them with epistaxis, revealed ulcerative and inflammatory lesions in all of them.(8) 9 of our patients were severely ill and deeply jaundiced with episodes of epistaxis. They had prolongation of both Pt PTT. This might be duo to liver dysfunction, yet DIC could not be excluded. Elevated levels of D-Dimer were found in 33 patients (94.3), one patient (2.9) had D-Dimer level 1000 g/L, 28 patients (80) had D-Dimer levels 1000 g/L while 4 patients (11.4) had D-Dimer levels 5000g/L. These 4 patients, with D-Dimer levels 5000g/L were severely ill, emaciated, severely pancytopenic and two of them were jaundiced with prolonged PT and PTT. So in this study there is a positive correlation between the level of D-Dimer and the clinical severity of the disease(p0.003). Elevation in D-Dimer levels indicates an ongoing state of thrombus formation and fibrinolysis. These findings may indicate the presence of such a process in VL which becomes worse with increasing severity of the disease. Similar findings were reported by Lomtadze et al who studied 45 patients with visceral leishmaniasis in Georgia thrombocytopenia was observed in 95 of his patients. Prolonged PT and PTT were found in severe forms of the disease, D-Dimer level elevated in 95.6. They concluded that leishmania infection affects primary haemostasis and coagulation and these alterations are related to the severity of clinical symptoms. They thus suggested that in visceral leishmaniasis activation of intravascular coagulation takes place, particularly during the severe forms of the disease.(9) Of interest is the finding of mild to moderate elevation (up to 750 g/L) of D-Dimer in 10 of the controls (14). However two of them had past history of VL and 4 were close contacts to patients with VL but the remaining 4 gave no similar history. This raises a very important question as to whether increased D-Dimer levels can indicate previous or subclinical infections. Conclusion Despite limitations of this study, it showed a significant relationship between the degree of elevation in D-Dimer and the clinical severity of the disease. D-Dimer test is more sensitive than PT PTT tests in assessing the haemostatic derangement such as DIC and excessive fibrinolysis and it may be possible to use D-Dimer levels in the follow up of patients to assess response to treatment. Also VL patients with a bleeding tendency may benefit from early and proper treatment of DIC. Acknowledgement Thanks are due to Prof. Sayda H. Elsafi, Faculty of Laboratory Sciences, University of Khartoum and her field team of Visceral leishmaniasis studies, Ministry of Health staff in Elazzaza Tabarakalla villages for their cooperation help. References Arias, J. R. Monteiro, P. S. and Zicker, F. (1996). The Reemergence of Visceral Leishmaniasis in Brazil. Emerging Infectious Diseases 2(2)145146. Sheffield Neave, M.R.C.P Leishmania donovani In Soudan, available at Br Med J. 1904 May 28 1(2265) 1252. Lloyd C. Rohrs, United States Naval Medical Research Unit No. 3, Leishmaniasis in the Sudan Republic, Am. J. Trop. Med. Hyg., 13(2), 1964, pp. 265-271. Kirk, Studies in leishmaniasis in the Anglo-Egyptian Sudan. XII. Attempts to find a reservoir host,available at HYPERLINK javascriptAL_get(this,20jour,20Trans20R20Soc20Trop20Med20Hyg.) Trans R Soc Trop Med Hyg. 1956 Mar50(2)169-77. Yezid Gutierrez. Diagnostic Pathology of Parasitic Infections with Clinical Correlations With Clinical Correlations.Oxford University Press US, 2000 79-81. 6- Deniau, M., Canavate, C., Faraut-Gambarelli, F. and Marty.The biological diagnosis of leishmaniasis in HIV-infected patients. Annals of Tropical Medicine and Parasitology,vol 97 (supplement 1), (2003) 115133(19). Cascio1, C.Colomba1, S.Antinori1, M.Orobello1, D.Paterson2 and L.Titone1, Pediatric Visceral Leishmaniasis in Western Sicily, Italy A Retrospective Analysis of 111 Cases, HYPERLINK http//www.springerlink.com/content/101941/pe35c983f9a77442eac01c1551e70c1c7pi0 European Journal of Clinical Microbiology Infectious Diseases, HYPERLINK http//www.springerlink.com/content/rd0gund0nq1t/pe35c983f9a77442eac01c1551e70c1c7pi0 Volume 21, Number 4 / April, 2002, 277-282 HYPERLINK http//www.ncbi.nlm.nih.gov/sites/entrezDbpubmedCmdSearchTerm22JC3BCttner20C225BAuthor5DitoolEntrezSystem2.PEntrez.Pubmed.Pubmed_ResultsPanel.Pubmed_DiscoveryPanel.Pubmed_RVAbstractPlus Jttner C, HYPERLINK http//www.ncbi.nlm.nih.gov/sites/entrezDbpubmedCmdSearchTerm22RodrC3ADguez20SC3A1nchez20M225BAuthor5DitoolEntrezSystem2.PEntrez.Pubmed.Pubmed_ResultsPanel.Pubmed_DiscoveryPanel.Pubmed_RVAbstractPlus Rodrguez Snchez M, HYPERLINK http//www.ncbi.nlm.nih.gov/sites/entrezDbpubmedCmdSearchTerm22RollC3A1n20Landeras20E225BAuthor5DitoolEntrezSystem2.PEntrez.Pubmed.Pubmed_ResultsPanel.Pubmed_DiscoveryPanel.Pubmed_RVAbstractPlus Rolln Landeras E, HYPERLINK http//www.ncbi.nlm.nih.gov/sites/entrezDbpubmedCmdSearchTerm22Slappendel20RJ225BAuthor5DitoolEntrezSystem2.PEntrez.Pubmed.Pubmed_ResultsPanel.Pubmed_DiscoveryPanel.Pubmed_RVAbstractPlus Slappendel RJ, HYPERLINK http//www.ncbi.nlm.nih.gov/sites/entrezDbpubmedCmdSearchTerm22FragC3ADo20Arnold20C225BAuthor5DitoolEntrezSystem2.PEntrez.Pubmed.Pubmed_ResultsPanel.Pubmed_DiscoveryPanel.Pubmed_RVAbstractPlus Frago Arnold CEvaluation of the potential causes of epistaxis in dogs with natural visceral leishmaniasis, HYPERLINK javascriptAL_get(this,20jour,20Vet20Rec.) Vet Rec. 2001 Aug 11149(6)176-9. HYPERLINK http//www.ncbi.nlm.nih.gov/sites/entrezDbpubmedCmdSearchTerm22Lomtadze20ML225BAuthor5DitoolEntrezSystem2.PEntrez.Pubmed.Pubmed_ResultsPanel.Pubmed_DiscoveryPanel.Pubmed_RVAbstractPlus Lomtadze ML, HYPERLINK http//www.ncbi.nlm.nih.gov/sites/entrezDbpubmedCmdSearchTerm22Khochava20MA225BAuthor5DitoolEntrezSystem2.PEntrez.Pubmed.Pubmed_ResultsPanel.Pubmed_DiscoveryPanel.Pubmed_RVAbstractPlus Khochava MA, HYPERLINK http//www.ncbi.nlm.nih.gov/sites/entrezDbpubmedCmdSearchTerm22Shalamberidze20IA225BAuthor5DitoolEntrezSystem2.PEntrez.Pubmed.Pubmed_ResultsPanel.Pubmed_DiscoveryPanel.Pubmed_RVAbstractPlus Shalamberidze IA, HYPERLINK http//www.ncbi.nlm.nih.gov/sites/entrezDbpubmedCmdSearchTerm22Shilakadze20MA225BAuthor5DitoolEntrezSystem2.PEntrez.Pubmed.Pubmed_ResultsPanel.Pubmed_DiscoveryPanel.Pubmed_RVAbstractPlus Shilakadze MA, HYPERLINK http//www.ncbi.nlm.nih.gov/sites/entrezDbpubmedCmdSearchTerm22Dzhokhtaberidze20TG225BAuthor5DitoolEntrezSystem2.PEntrez.Pubmed.Pubmed_ResultsPanel.Pubmed_DiscoveryPanel.Pubmed_RVAbstractPlus Dzhokhtaberidze TG Functional status of haemostasis system in patients with visceral leishmaniasis, HYPERLINK javascriptAL_get(this,20jour,20Georgian20Med20News.) Georgian Med News. 2005 Nov(128)59-62. PAGE PAGE MERGEFORMAT 1 Low Normal 25 75 Fig(6) Hb in Male Controls Low Normal 44 56 Fig(7) Hb in Female Controls Low Normal 80 20 Fig(8) WBC in Patients group Low Normal 33 67 Fig (9) WBC in Controls group Normal Low 11 89 Fig (10) Platelet Count in Patients group prolonged Normal 9 91 Fig (11) PT for Patients group prolonged Normal 9 91 Fig (12) PTT for Patients group HIGH, 94.30 LOW, 5.70 Fig (13) D-dimer in Patients group Fig(1) Blood Films for Malaria in Patients group POSITIVE 23 NEGATIVE 77 0 Fig(2) Jaundice in patients group 91 9 Normal Jaundiced Fig (3 ) Splenomegaly in Patients group POSITIVE 86 NEGATIVE 14 Fig (4 )Hepatomegaly in Patients group POSITIVE 29 NEGATIVE 71 Low Normal 80 20 Fig(8) WBC in Patients group ) [email protected]_t)[email protected]/lV/ 3hm5fumjU5c_MMTUvcn0-RQafjeZcw.DJ9hKsb3C3h1O31 tKos @Yos ./A 6x2LKx9/b @YSIJBiRKw/mgCmc9xfcmnALop [email protected]/UB41kf Jky2cZM gI-PNCSlTlyL9Kjxvj6o9 Y, dXiJ(x(I_TS1EZBmU/xYy5g/GMGeD3Vqq8K)fw9
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