Blood Parasites PDF - Fifth Year Pure Science

# Fifth year Pure Science ## Infectious Diseases ### Blood Parasites **D/ Ahmed Magdy** * شرح الاونلاين + شرح المحاضرة * DR. PETER # I. Bovine Babesiosis ## Synonyms * Texas fever (first recorded in Texas) * Red water fever (red urine) * Cattle tick fever * Enzootic hemoglobinuria (Hb.uria) *...

# Fifth year Pure Science ## Infectious Diseases ### Blood Parasites **D/ Ahmed Magdy** * شرح الاونلاين + شرح المحاضرة * DR. PETER # I. Bovine Babesiosis ## Synonyms * Texas fever (first recorded in Texas) * Red water fever (red urine) * Cattle tick fever * Enzootic hemoglobinuria (Hb.uria) * Splenic fever (splenomegaly) ## Def. * Group of tick borne seasonal diseases of domestic, wild, lab animal & man caused by protozoa (genus babesia) * Acute, subacute or chronic tick borne protozoal disease of cattle & buffalo caused by babesia spp. * More than 71 species of babesia are known, only 18 causes diseases in domestic animals * Characterized by * High fever (41-42°C) * Hemolytic anemia * Hemoglobinuria & hemoglobinemia * Coffee black urine * Death due to anemic anoxia * Chronic form → cannot be defined clinically & associated with anemia & weight loss. ## Host & Etiology | Species | Shape | Susceptible Host | |---|---|---| | B. bigemina | Large in size | | | B. major | Pairs of babesia forming → Acute angle | | | B. Bovis | Small in size Pairs of babesia forming → Wide angle | Cattle & buffalo | | B. divergen | Small in size Pairs of babesia forming → Acute angle | | * Host specific * Intra-erythrocyte * Pear shape & pyriform bodies * Arranged in pairs forming acute or wide angle **DR. PETER** * Cattle more susceptible than buffalo * All ages are susceptible, but clinical signs & pathological changes are mild & short lived in animal under 6 months old while severity of signs increases with age. * Common age of infection 6-12 months of age. Why? **Endemic stability (innate resistance( هااااام** * Calf from previously immune (infected) dam takes maternal immunity from colostrum (passive immunity) → give protection for 3 months of age → followed by age resistance to 9 months old. * When calf exposed to infection during this time Rarely showed clinical signs & get long last immunity (endemic stability) ## Life cycle * 1st phase → multiply sexually by binary fusion in RBCs of vertebrate host * 2nd phase → in female tick by sporogony → Sporozoites (infective stage) in salivary gland of female ticks * Hard ticks have 4 life stages * Adult → egg → 6 legged larva → 8 legged nymph * Most ticks prefer to feed on different hosts at each life stage. * Ticks can feed on mammals, birds, reptiles, amphibians ## Notes * Typical egg masses range from 2000 to 10000 (recorded of 22000) ## Transmission **Infected Ticks (mainly)** * Biologically by * Transovarian * Trans-stadial (Stage to stage) **DR. PETER** * Mechanical transmission → contaminated needles, surgical instruments & Blood transfusion **عشان كده لازم تغير من الابرة بين الحيوانات زى BEF** * Biologically → Multiply and growth in I.H * Mechanical → No multiplication or growth on in I.H * Intra-uterine infection → not common **لكن ممكن بتحصل** **Ticks can transmit babesia by: هااااام** * Stage to stage (trans-stadial) transmission * Adult stage transmitting infection which they acquired as nymph * Trans-ovarian transmission (TOT) * Adult female engorged with infected blood → protozoa pass to ovary → all eggs ## Pathogenesis * Protozoa multiply in peripheral or visceral circulation by binary fusion * Destruction RBCs → * Hemolysis, anemia (hemolytic anemia) * ↓ Ph. → Pica (soil eating( بتاكل الطينه هااااام * Liver → jaundice * Kidney → hemoglobinuria * Some metabolized in liver → producing unspecific toxic substance → affect thermoregulatory center in brain → fever (41-42°C) * Death occurs due to anemic anoxic (destruction of RBCS ↓ O2 infusion for tissues( امتحااااان * If animal survive → become carrier & resist re-infection with the same species. * **DR. PETER** ## Signs * Depend on amount of parasite in blood) * IP 7-20 days (2-3 weeks) * Morbidity 40% * Mortality 60% * **تانى يوم الاكل يقل اكتر** * **الاعراض في اول يوم سخنيه لكن الحيوان بياكل (بيلقط) > وهنا سهل علاجه** * **تالت يوم bloody urine وهنا صعب العلاج، لو اديت علاج والبول رجع طبيعي تانى يوم ← يبقى فيه امل تستجيب وهتخف** * **لكن لو عدى تانى يوم العلاج والبول مدمم زى ما هو يبقى الحاله hopeless** * **Sudden onset of high Fever (41-42°C) + inappetence "in complete off food" (in early stage)** * **complete off food - BEF لكن في حالات الـ وده اللى بيفرقها عن مرض الـ BEF لان الاتنين بينتشروا في الصيف وحراره عاليه** * **كل ما ↑ حراره الحيوان ← تطمن ان الحاله هتستجيب مع العلاج؛ لكن لو الحراره قلت ← تقلق ان الحاله مش هتستجيب (حلها الوحيد نقل دم)** * **NB: Cases of High fever + normal appetite → babesia & pneumonia** * **Late stage → anorexia, depression, weakness, emaciation, cessation of rumination & ↓ milk yield** * **↑ Respiratory & heart rates** * **Congested M.M with petechial hemorrhage characteristic for babesia (due to prothrombin) → pale (due to anemia) → jaundice** * **Foamy bloody, dark red to brown urine (coffee colored urine) with very stable froth due to high content of protein (characteristic for babesia)** ## Present of babesia & ticks on animals without animals' losses or clinical signs → Endemic stability **الكلام ده كان زمان في مصر، دلوقتي ممكن عند عمر ٣-٤ ايام يجيله بابيزيا او ثيليريا عادي** ## i.e. Nymph (feed on infected animal and got infection) → molt to infected adult (transmit the babesia) ## 2 or 3 host ticks as Hyalomma, Rhypicephelus & Ixiodes spp. ## become infected → larva harbor infection but not transmit infection until it becomes nymph. ## So Nymph plays a principal role in disease transmission ## One host ticks as Boophilus spp. ## PM * Presence of ticks on carcass skin * Marked Jaundice * Liver → hepatomegaly & dark brown * Gall bladder → distended & contain thick granular bile * Splenomegaly (can reach one meter in length) → spleen has soft & pulpy consistency * Kidney → Dark & enlarged * Red urine in urinary bladder * Pericardium & endocardium → ecchymotic hemorrhage ## Diagnosis * **Field diagnosis** * History → Ticks (in summer season) & coffee colored urine * **حتى لو ملقتش الـ ticks لا تستبعد الـ blood parasite لان ممكن يكون الأونر استخدم دوا للقراض من اسبوع ۱۰ ايام** * **لازم تعرف الاونر لو الحاله عشر عشان ميقولش العلاج بتاعك هو اللي عملها abortion، وممكن الـ abortion يحصل بعد اسبوعين تلاته من العلاج** * **Clinical signs & PM** ## ## Signs indicate death (cases of bad prognosis( هاااااام **حالات ملهاش علاج (متدخلش فيها) حلها الوحيد نقل دم** * Foreign breeds (as it take sever infestation form) * Subnormal temperature (hypothermia) + cold extremities * Recumbency * **وصلت للمرحلة الاخيرة Complete off food .** * Panting & difficult breathing بينهج وبتأن → due to destruction of RBCs & ↓ 02 * One year old infection * Late pregnancy * Nervous signs **DR. PETER** * **between 2 thigh or udder & ،اكثر اماكن فيها القراض، تحت الذيل thigh** ## Lab diagnosis * **Samples** * Thin blood film * Peripheral blood smear from ear vein puncture (during febrile stage) * Using Giemsa stain * Pyriform in shape * Hemolymph smear & Egg smear from ticks ## متروحش لحاله وتديها علاج الا لما تشوف الـ urine ## في الـ male بتعمل مساج للـ prepuce، في الـ female بتركب قسطرة ## Nervous signs (Cerebral Babesiosis( هاااام * Due to clotting in brain capillaries (hopeless) * Incoordination, head pressing, mania, convulsion & coma * Using heparin might help * **95% mortality** * **استخدام الهيبارين ممكن يساعد لكن غالبا مش بتستجيب** ## In case of B. divergen → spasm in anal sphincter → defecation with great force in long tin stream → Pipe-stem feces ## ## In severe cases → death after 24 hr. ## ## Non-specific abortion (due to high fever) ## ## # II. Bovine Theileriosis ## ## # II. Bovine Theileriosis ## ## # II. Bovine Theileriosis ## ## # II. Bovine Theileriosis ## ## # II. Bovine Theileriosis ## ## # II. Bovine Theileriosis ## ## # II. Bovine Theileriosis ## ## # II. Bovine Theileriosis ## Synonyms * Tropical Theileriosis * Egyptian fever * Mediterranean coast fever ## Definition * Peracute, acute, subacute or chronic tick-borne protozoal disease of cattle & buffalo, caused by Theileria annulata * Characterized by fever, progressive anemia, wasting, enlargement of L.N & corneal opacity, abomasal ulcer ## Etiology * Theileria species * Host specific (No cross immunity occurs between different spp. of the parasite) | Species | Disease | Distribution | |---|---|---| | T. annulata | Tropical Theileriosis Egyptian fever | Northern, Mediterranean & subtropical regions of the world | | T. parva | East coast Fever | East & central Africa | | T. orientalis | Oriental theileriosis | Korea, Japan, Egypt | | T. mutans | Benign bovine theileriosis | Africa & south Sahara | ## The parasite occurs in two forms: * Erythrocytic (Piroplasm) Form * Parasite found inside RBCs during febrile period → * Lymphocytic form (Koch's blue bodies) * Schizont phase of parasite occur inside lymphocyte ## Dormant (without multiplication) & don't undergo further development until ingested by ticks in which they multiply sexually ## Presence of several form in RBCs represent multiple invasions as signet ring, oval, dot, coma shape) ## Susceptibility * Cattle & water buffaloes * Highly virulent for European dairy cattle & may be subclinical in local cattle * Water buffaloes → can be infected and be carrier. * **الجاموس بياخد العدوی chronic form ، لكن لو حصل stress بتقلب acute** ## Transmission * Mainly through tick (Hyalomma spp.) by * Stage to stage (trans-stadial) transmission only (3 Host ticks or 2 host ticks) * Contaminated needle or syringe * Transplacental transmission هاااااام **بتلاقي عجل عمر ٣ ايام مصاب بالثيليريا** ## Pathogenesis * Ticks must feed for 2-4 days before sporozoites (infective to cattle) become mature (infective to cattle) * Appear as chromatin dots seen in mass of bluish stain cytoplasm with Giemsa stained in aspirated smear of lymph node (Koch's blue bodies) * Sporozoites → enter to blood during blood meal then go to lymphocytes (develop Schizonts) in L.N in area of tick attachment → lymphocytes transformed to lymphoblast → infected lymphoblast disseminated through lymphoid system * Some schizonts differentiate to merozoites → invade RBCs * In RBCs, parasite transforms to piroplasm which are infective to ticks * Parasite inside tick undergoes developmental changes → sporozoites * Main pathologic changes caused by Schizonts in lymphocytes & merozoites in RBCs * Lymphoadenopathy due to * Division of schizonts in lymphocytes * Phagocytosis & destruction of piroplasm infected RBCs → hemolytic anemia with icterus * Anemia, bilirubinemia, hemoglobinuria & bilirubinuria ## Clinical signs * IP → 6-21 days * Morbidity → 60% * Mortality → 60% * A) Peracute form * Foreign Susceptible animals enter endemic areas * Fever, anorexia, weakness & die within 2-3 days ## B) Acute form هااااام * Fever persists for several days, inappetence & drop in milk production. * Enlargement of L.Ns (parotid, pre-scapular & precrural "pre-femoral") → due to rapid division of infected lymphoblast & accumulation of infected fluid * Skin lesion (reported in Tunisia) found schizonts in skin nodule S/C هاااااام ## LSD * Nodules → large & containing all skin layers. * Edema in limb & brisket. * Outbreak ## Theileria * Smaller * Corneal opacity. * No outbreak ## Dermatological signs in bovine tropical theileriosis (Theileria annulata infection). * Nasal & ocular discharge & swollen eyes * Corneal opacity within 2-3 days after fever (central to peripheral( هااااام * **DR. PETER** ## امتحااااان Eye Proptosis * Massively protruding of eye balls out of their sockets * Due to pressure of accumulating fluid in the membrane of the orbit (and may combined with Pink eye disease) * May progressed to complete removal of eye ball ## Eye Proptosis * Complete removal of eye ball ## Difficult breathing & cough (resp. signs هااااام * Due to pulmonary edema and 2ry bacterial infection by pasteurella → BRD * Die due to asphyxia ## After few days → anemia & pale M.M ## Later → hemoglobinuria, bilirubinemia & Jaundice ## Constipation (due to fever) then bloody diarrhea (1) * Due to ulceration of internal mucosa “Payer's patches") → bad prognosis ## Abomasal ulcer → Melena (2) (black terry diarrhea) * Animals become rapidly emaciated & about 40-80%. die over a period 8-18 days ## C) Subacute form * Intermittent fever for 2-4 weeks * Moderate progressive anemia & Jaundice * Animal may recover or develop acute form after stress & die ## D) Chronic form * More prolonged form of subacute disease. * Recovery is rare & some cases suddenly develop acute form & die * In Egyptian buffalo → chronic diarrhea (3), emaciation with normal temp. & appetite * **anthelmintic او antibiotics مش بتستحيب لاى diarrhea حالات** * **There are 3 types of diarrhea Bloody, Melena & Chronic. هاااام** ## PM * Pin-headed infarct (foci) in kidneys' cortex → due to aggregation of lymphocytes * Abomasal & intestinal mucosa → ulcerated (at Payer's patches) 2-12 mm surrounded by zone of inflammation * black diarrhea (bad prognosis) * Emaciation, icterus, Infested with ticks & anemia (pale M.M & yellowish discoloration of tissues) * Lung → pulmonary edema, hyperemia & congestion * Ln → enlarged & edematous & hyperemic * Liver & spleen → enlarged & friable & pat brown or yellow. * Heart → epicardial & endocardial hemorrhages. ## Diagnosis * **Field diagnosis** * History → suspected from area * Clinical signs & PM * Presence of ticks * **Lab diagnosis**  * Thin blood film (during febrile) from ear vein puncture → examine oil immersion lens found: * Ecinocytes هااااام * **كرات الدم الحمرا بيتكون على شكل النجمه وليها شوك** * Containing oval, signet ring, dots or coma shape piroplasm surrounded by hallow zone stain deposits والـ piroplasm هي اللى بتفرق بين الـ Hallow zone الـ * Smears from Enlarged L.N, liver, kidney & spleen → Koch's blue bodies "diagnostic" هااااام * Blood for hematology (CBC & PCV) → anemia, leukopenia * Liver function test → ↑ ALT + ↑ AST + Gamma-glutamyl Transferase (GGT) Test * Kidney Function test → ↑ creatinine + ↑ BUN (blood urea nitrogen) * Serum for serology → IFAT, CFT, ELISA * PCR for carrier animals * **Differential diagnosis** * Babesia, anaplasma ## MCF هاااااام * Corneal opacity from peripheral to central * Ulceration & necrosis in cheek papillae * Crusts & ulceration on muzzles & nostril * Not respond to antipyretics ## Trypanosomiasis  * **In camel & not common in Egypt** * **By blood smear → intercellular blood parasite** ## Treatment هاااااام * **Specific Treatment** * Buparavaquine (Butalex, Buta-ject, Theilcure( غالی * 1 ml/20 kg I/M 2 dose 48 hr. interval * وفي حالات ممكن تحتاج ٣ جرعات * Oxytetracycline L.A (Almycin LA) * 1 ml/10 kg I.M * Not recommended as it has load effect on liver) * **Marbofloxacine + Butalex combination better than Oxy + Butalex combination** * Alexoquine 250 mg tablets (chloroquine phosphate) * بشرى يستخدم لعلاج المالاريا * باستخدم شريط كامل باطحنه واديه للبقره يوميا لمده ٣ ايام ← بیدی نتایج كويسه جدا * Paraquone (cloxon) * 1 ml/15 kg I/M (not common) * (resistant for Butalex وخلها تبقى في حالات الـ theileria cytochrome B gene في جين mutation لان حصلها ## Treatment of 2ry disease امتحااااان * For BRD and pulmonary edema * Diuretic → Furosemide (Lasix) ampules بشری or Hydrochlorothiazide (Diurizone) * Diurizone is better than Furosemide * O as Furosemide cause loss of mineral with water) ## Abdominal ulcer treatment * Antacid as aluminum hydroxide “Mucogel” or Gaviscon * H2 blocker as famotidine "antodine or Gastrotidine" ampule / 40 kg every 12 hrs. * PPI "proton pump inhibitors" as Pantoprazole ## Marbofloxacine for 2ry bacterial infection * Acetylcysteine 600 mg sachets (mucolytic) → 2 sachets/ daily * Careful not to ↑ mucolytic dose as it↓ cilia movement ## Control * Tick control & Treatment * Chemo-immunization by injection of L.A Oxytetracycline ## Vaccination * A) Sporozoites (stabilate) vaccine (vaccination then treatment) * Vaccination (0.5 ml, S/C, near to pre-scapular L.N) then treatment with tetracycline (20 mg/kg) or Butalex at day 8 post-vaccination (to suppress activity of parasite at its peak of multiplication) * B) Schizont vaccine (live vaccine) * Whole blood vaccine (not common) → blood from infected cow with mild strain (as Kouba strain of T. annulata) * Cell culture vaccine (Rakshavac-T) * Prepared from live schizonts of T. annulata grown in lymphoid cell culture & attenuated by prolonged in vitro passage * 3 ml S/C, immunity for 1 year * Preserved in liquid nitrogen * Before using, must be put in water bath of 37°C * Not used for pregnant or weak animal **DR. PETER** ## (Controloc) ampoule / 100 kg every 24 hrs. ## III. Anaplasmosis ## Synonyms * Gall sickness * Yellow cow disease (all M.M become jaundice) ## Definition * Tick borne disease of ruminants, caused by rickettsia (anaplasma) * Characterized by → fever, anemia, jaundice & enlarged superficial L.N * Subclinical in sheep & goat. ## Etiology * Anaplasma spp. (rickettsia) * A. marginale Cause severe disease & Locates on margins of RBCs * A. centrale Cause mild disease & Locates on central of RBCS ## Obligate intracellular & multiply by binary fission of RBCs. ## Susceptibility * Cattle & buffalo * Sheep & goat (subclinical) * Young age usually resistant to disease (cattle over 3 years are commonly affected) * Any immune depressing as Live attenuated vaccine of LSD ## Transmission * Ticks * (mainly) * Stage to stage & Transovarian * Ex: Boophilus spp., Hyalomma spp. * Mechanical transmission * Contaminated needles, syringe or hemophilic diptera flies as Haematobia irritans, Tabanus Stomoxys & blood transfusion. * Anaplasma transmitted biologically by ticks, however different hemophilic diptera as Haematobia irritans & Stomoxys can mechanically spread rickettsia امتحااااااااااان * **تلاقی مزرعه مفيش فيها ticks لكن في نفس الوقت فيها anaplasma** * Transplacental | not common ## Pathogenesis * Infective initial bodies enter host along with saliva of vector, to capillaries of visceral organs then peripheral circulation → ↑ in size & multiply by binary fission in RBCs into 2- 8 new initial bodies * Eventually, initial bodies are liberated from RBCs & process repeated * Anemia result from → phagocytosis of RBCs in spleen (not hemolytic) * So anima & jaundice without Hb.uria (no bloody urine) * عكس البابيزيا والثيليريا، هنا تكسير الـ RBCs بيحصل في الـ spleen * Animal recovered become carrier for life. هاااام ## # # Clinical signs * IP 3-4 weeks * Peracute * In Cattle over 3 years → often fatal * High fever (40.5°C - 41.5°C) * Anemia & pale M.M → rapid breathing, excessive salivation * Occasionally nervous manifestations * Death * Acute form * Animal develop fever over few days (reaching 41°C – 41.5°C) * Enlargement of L.N * Anemia, sever jaundice without Hb.uria, icterus (yellow cow disease) * Petechial hemorrhage → indicate liver dysfunction (↓ prothrombin), so use Vit. K in t.t.t هاااااام * **لاحظ ان Vit. K ملوش لزمه في حالات البابيزيا (لانه تكسير في RBCs)** * Emaciation, weakness. * Abortion (nonspecific) & temporally infertility in bulls. * Affected animals are usually has dark brown (bloody) feces containing mucus with offensive odor (due to ↑ unconjugated bilirubin( هاااااام * However, the urine is not usually red as in babesiosis, but urine will be a dark yellow color هاااااام * In severe cases هاااام * By auscultation → you can hear heart sound from rumen (flank region) * Indicate → severe anemia with tachycardia **DR. PETER** ## Jugular vein pulsation * Puncture of jugular vein with needle → watery blood (bad prognosis) ## NB: Cases of hearing heart sound from rumen * Babesia, * Hepatization pneumonia, * Hemo-concentration (dehydration) * Severe anemia ## PM * Emaciated carcasses, pale & icteric tissues * Thin & watery blood * Enlargement of L.N (congested & edematous), * Hepato-splenomegaly * Heart → flabby & petechial hemorrhage on endocardium & pericardium. ## Diagnosis * **Field diagnosis** * History of endemic Anaplasmosis, signs & PM / anima & jaundice without Hb.uria * **Lab diagnosis** * Samples * Blood → Giemsa stain blood smears * CBC * Serum → Serum biochemistry * Liver function tests → ALT, AST, GGT ## Kidney function tests → urea, creatinine. ## Serology CFT, IFAT ## Carriers detected by → PCR & transmission test. ## Treatment * **A) Specific treatment** * Imidocarb (Imizol) → 2.5 ml / 100 k S/C. * Long-acting tetracycline → 1 ml/10 kg IM. * Marbocyl (marbofloxacine) * To prevent latent of carrier (carrier elimination( هاااام * 1) Imizol → 2 doses of 2 weeks apart or daily for 3 days. * 2) Oxytetracycline (S.A) → Daily dose 10 mg/kg for 10 days. * 3) Oxytetracycline (L.A) → 20 mg/kg 4 times at 3-7 days interval. **DR. PETER** ## عرف الاونر ان العلاج هيطول، ومش هترجع تاكل الا بعد ٣-٤ ايام، فلازم تدى مقويات للكبد بالذات ## Supportive treatment for blood parasite هاااام * 1. Freshness of the kidney اغسل الكلى * Fluid therapy with Duphalyte, phosphorus & Vit B. * Give animal boiled barley water (diuretic) * **نقطة المحاليل دى عليها خلاف لان المحاليل هتعمل dilution for blood وهو اصلا خفيف، لكن اتستخدمها ومتزودش عن ازازه واحده (٥٠٠ مل) ويفضل تشرب الحيوان شعير مغلى او بيريل** * 2. Vitamins & Electrolytes as * Iron dextran 7 ml / 100 kg I/M * Fercobsang for anemia (Cobalt + vitamins 20 ml/cow I/M * Duphalyte or Ornipural (sorbitol + 100 ml/cow I/M. ## A.A) as hepatic protective which convert Ammonia to urea * Jecuplex, Bovi Care (250 ml for one shot orally) or Bovi Pak "L-carnitine" as liver support * Tonophosphan 20% (phosphorus) * Epinosine B forte ampules * Vit. K (Amri K) ampules ## Prevention & control * 1) Tick control (as babesia). * 2) Detection of carriers by PCR. * 3) Restriction of Cattle movement. * 4) Vaccination * Live anaplasma marginale → creation of large number of Carrier animals * Attenuated vaccines (frozen anaplasma marginale) * Killed anaplasma marginale vaccine * 2 injection 4-6 weeks apart & the last dose given 2 weeks before vector season (summer) * Booster dose 2 weeks before next vector season. ## divided for 3 days ## 60 ml daily for 3 days given orally ## To treat pica ## Source for ATP & Vit. B ## For bleeding (in Anaplasmosis) ## Has no role in case of babesiosis ## من الاخر كده...... ازاى تتعامل مع حاله blood parasites في الفيلد؟؟! ## لو الحاله سخنه والبول لسه طبيعي (ممكن تكون بابيزيا او انابلازما) > فبدی Imizol بجرعه ٢٫٥ سم / ١٠٠ كيلو وبقسمهم على مرتين مره ## ## # ## ## # ## ## # ## ## # ## ## # ## ## # ## # # ## # # ## # # ## # # ## # # ## # # ## # # ## # # ## # # ## # # ## # # # ## # # # # ## # # # ## # # # ## # # ## # ## # # # ## # # # # # # ## # # # # # ## # # # # ## # * # * # ## # # # ## # # # # # # ## # # # # # ## # # # # ## # * # * # ## # # # ## # # # # # # ## # # # # # ## # # # # ## # * # * # ## # # # ## # # # # # # ## # # # # # ## # # # # ## # * # * # ## # # # ## # # # # # # ## # # # # # ## # # # # ## # * # * # ## # # # ## # # # # # # ## # # # # # ## # # # # ## # * # * # ## # # # ## # # # # # # ## # # # # # ## # # # # ## # * # * # ## # # # ## # # # # # # ## # # # # # ## # # # # ## # * # * # ## # # # ## # # # # # # ## # # # # # ## # # # # ## # * # * # ## # # # ## # # # # # # ## # # # # # ## # # # # ## # * # * # ## # # # ## # # # # # # ## # # # # # ## # # # # ## # * # * # ## # # # ## # # # # # # ## # # # # # ## # # # # ## # * # * # ## # # # ## # # # # # # ## # # # # # ## # # # # ## # * # * # ## # # # ## # # # # # # ## # # # # # ## # # # # ## # * # * # ## # # # # # ## # # # # # ## # # # # * # ## # # # # # ## # # # # # ## # # # # * # ## # # # # # ## # # # # # ## # # # # * # ## # # # ## # # # # # ## # # # # * # * # ## # # # ## # # # # # ## # # # # * # * # ## # # # ## # # # # # ## # # # # * # * # ## # # # ## # # # # # ## # # # # * # * # ## # # # ## # # # # # ## # # # # * # * # ## # # # ## # # # # # ## # # # # * # * # ## # # # ## # # # ## # # # ## # # # ## # # # # # ## # # # # * # * # ## # # # ## # # # # # ## # # # # * # * # ## # # # ## # # # # # ## # # # # * # * # ## # # # ## # # # # # ## # # # # * # * # ## # # # ## # # # ## # # # # ## # # # # # # ## # # # # # ## # # # # * # ## # # # # # ## # # # # # ## # # # # * # ## # # # # # ## # # # # # ## # # # # * # ## # # # ## # # # # # ## # # # # * # * # ## # # # ## # # # # # ## # # # #