LEPTOSPIROSIS 1 Summary

Leptospirosis: bacterial zoonosis Transmission via contact with contaminated fresh water Fever, muscle pain, cough, red eyes, Hepatomegaly, icterus, haemorrhagic tendency, meningitis, nephritis Difficult clinical diagnosis: water contact, leukocytosis, urine analysis, lumbar puncture Serology and direct detection of bacteria are difficult to carry out Treatment tetracyclines, penicillin

2 General
Leptospirosis is a bacterial disease which is distributed world-wide. There are even occasional cases in Belgium and the Netherlands. Leptospires are the only pathogenic spirochaetes which are free-living in the environment. In comparison, Treponema pallidum is only found in humans and Borrelia spirochaetes are only found in arthropods and mammals. The severe form of leptospirosis was described in 1886 by the German Adolf Weil, Professor of Medicine at the University of Heidelberg. It is therefore still called Weils disease. In 1907 Stimson discovered the organism in kidney tissue from a patient who died during a yellow fever epidemic (see Clinical aspects). In 1914 Japanese researchers under the direction of Ryokichi Inada discovered spirochaetes in the liver of guinea pigs injected with blood from patients with Weils disease. The conditions at the front during the First World War (mud, water, trenches and rats) meant that leptospirosis became a significant disease with military implications. The generic name Leptospira originates from the Japanese Hideyo Noguchi, at that time associated with the Rockefeller Institute. In 1925 the West African Yellow Fever Commission was set up with the support of the Rockefeller Foundation. A laboratory was built near Lagos, Nigeria. For two years attempts were made to demonstrate that leptospires were the cause of yellow fever. In retrospect the confusion is easily understable. Clinically it is indeed very difficult to differentiate between yellow fever and leptospirosis. In regions where scrub typhus and hantavirosis are endemic, differentiation between Orientia tsutsugamushi, hantavirus infection and leptospirosis on clinical criteria alone, is just not possible.

3 Taxonomy
The genus Leptospira contains 2 species: L. biflexa which is apathogenic for man and L. interrogans which is pathogenic. The micro-organisms are very delicate and spiral-shaped, hence the name. They have a typical terminal hook (Gr. leptos = delicate, slender, speira = spiral, interrogans = question mark). Leptospires have a characteristic double membrane architecture with features of both Gram-positive and Gram-negative bacteria. There are many variants (240 serotypes or serovars, grouped into 23 serogroups), of which L. interrogans serovar icterohaemorrhagiae is the most important. Others include canicola, tarassovi, pyrogenes, bataviae, javanica, ballum, cynopteri, celledoni, panama, shermani, ranarum, bufonis, pomona, hebdomadis, autumnalis and grippotyphosa. The serovars are defined by agglutination after cross-absorption with homologous antigen. The agglutinating antigen is a lipopolysaccharide. Classification into 17 different genome species based on the percentage of DNA: DNA hybridisation is possible, but such a classification is as yet of no clinical significance. The subspecies are all morphologically identical. The bacteria are so thin that they cannot be detected with normal light microscopy. They can be detected using phase contrast or dark-field microscopy (urine) and by means of silver staining of tissue sections. There is a poor correlation between clinical severity and the precise serotype of the bacteria.

4 Transmission
The pathogenic bacteria can survive in fresh water, but die in seawater. Infected animals retain bacteria in their kidneys for a long time and eliminate them in the urine. Transmission follows contact with fresh water contaminated with the urine of infected animals. Rats form the main reservoir, but other animals such as cattle, dogs, cats and pigs may also become infected. It is an important animal disease. * Leptospires are killed by gastric acid and bile salts. They penetrate the body via wounds and via the mucosa of the mouth, nose and eyes (conjunctivae). Water is the most important route of transmission, but direct contact with infected animals may also be significant (slaughterhouse workers, veterinary surgeons). It is a disease associated with certain occupations, e.g. workers in paddy fields or on sugar cane plantations, farmers, workers in sewers and canals, gold prospectors (gold dust obtained from water courses). People who bathe or swim in infected surface water are at increased risk of this zoonosis. Now that rafting, kayaking and adventure sports in tropical regions have become popular, there is an increase in leptospirosis in tourists. Ideal conditions for transmission are produced when dirty streets with large rat populations are flooded. In 1995 there was an important epidemic of leptospirosis in

Central America, caused by chronic failure to collect household waste and rubbish leading to an increased rodent population. Heavy rains followedy. The presence of leptospires in water can be detected experimentally by the "immersion test". The abdomen of a non-infected guinea pig is shaved and the skin scarified. The damaged skin is brought into contact with water for one hour and not dried off. After an incubation period, it is possible to detect leptospires in the animal.

5 Course of the disease

In view of the many sub-species of leptospires, a wide spectrum of diseases is possible. Symptoms range from mild fever with a flu-like syndrome to atypical pneumonia, myocarditis, aseptic meningitis or the severe Weils disease with liver and kidney failure, meningitis and haemorrhaging. The disease course has three phases: the first septicaemic, the second with leptospiruria (leptospires in the urine) and the third, convalescence phase. During the first phase the leptospires are present in the blood in low numbers (too low to be detected in a blood smear using phase contrast microscopy). Subsequently the organisms disappear from the blood due to the formation of antibodies. Cellular defence also clears the bacteria from the various tissues. Leptospires persist in the kidney. In the renal tubuli the organisms can multiply and cause renal damage. Bacteria are eliminated with the urine, although the concentration is quite low: < 104/ml urine. The bacteria may remain in the kidneys for months, even after clinical recovery. Leptospires might also persist in the choroid plexus of the brain.

6 Clinical aspects
Leptospirosis exhibits a wide spectrum of symptoms. Mild forms are often atypical and are generally missed unless they are specifically sought. Severe forms exhibit a more dramatic symptomatology and course. In severe leptospirosis, after an incubation of 2 to 20 days there is sudden high fever. This is accompanied by pronounced headache (95%) and general malaise. Patients can sometimes pinpoint to within the hour when the illness began. The fever lasts approximately 7 days. The fever may then subside for a few days and then increase once more (biphasic fever). Absence of this fever pattern does not rule out the disease, however. Significant muscle pain is almost always present. If it is absent the diagnosis is improbable. The eyes are bloodshot (25%), but there is no pus, unlike in purulent conjunctivitis. The bacteria may be present for months in the aqueous humour of the anterior chamber of the eye, and in 2% of patients chronic inflammation (uveitis) may occur. There are two patterns: (1) self-limiting anterior uveitis with photophobia, blurred vision and pain, and (2) posterior or

panuveitis

with

cotton-wool

spots,

choroiditis,

retinal

haemorrhage,

vitreous

humour

membranes and papillitis. There is sometimes a sore throat and a dry cough, later possibly haemoptysis. In 10 to 30% of patients there is a spotty skin rash of the lower legs. [This was initially described as "Fort Bragg Fever", caused by L. interrogans autumnalis]. After a few days there are organ symptoms. Meningism may occur early, but is more frequent in the second phase. Neck stiffness is not always present. Two thirds of patients suffer nausea and/or vomiting. Swollen lymph nodes are only present in a minority of patients. The spleen is swollen in 20% of cases. Cd_1002_019c.jpg * Involvement of the liver is characterised by hepatomegaly, jaundice and a haemorrhagic tendency. The gall bladder may become inflamed (acute cholecystitis) as may the heart (myocarditis). The liver enzyme levels (transaminases) are only slightly elevated (no liver cell necrosis). Typically the icterus is described as more orange compared to that of viral hepatitis, but in practice this is not a very reliable sign. Leptospirosis may also exist without icterus. * Kidney damage leads to proteinuria, haematuria and uraemia. Hypovolaemia and poor renal circulation may further exacerbate the renal damage. Hypovolaemia is characterised by oliguria, low blood pressure, diminished skin turgor, flat neck veins. If it is not corrected by giving fluids tubular necrosis will follow. Myocarditis occurs and often leads to congestive heart failure and cardiogenic shock. Electrocardiographic abnormalities are common. During the second phase there is sometimes meningeal irritation with neck stiffness and a picture of "viral" meningitis. However, this usually only lasts for a few days. Organ complications such as kidney failure and haemorrhages also occur in this phase. The total period of illness is approximately 3 weeks to one month. The mortality is between 5 and 30 %, severe icterus having a poor prognosis. If the patient survives, there is usually no residual damage. A long convalescent period is typical. * In 2001 a new bacterium was discovered in Denmark, Leptospira faini serovar Hurstbridge. In two infected patients there was an unusual presentation: chronic infection characterised by subconjunctival bleeding (6 months), intermittent abdominal pain (5 months), severe headache for 2 months accompanied by dizziness and one month of jaundice. Further research needs to be carried out, however, to describe the full pathogenic spectrum of this bacterium.

7 Diagnosis
Diagnosis is quite difficult. This disease is often missed. The disease may be clinically

suspected. Exposure to potentially contaminated water (occupation, accident, swimming, etc.) should be enquired about. In a severe infection there is an influenzalike syndrome with sudden onset, followed by hepatitis with conjunctivitis, and then meningitis, haemorrhages and renal involvement. Milder infections generally exhibit a biphasic fever with muscle pain, red eyes, jaundice and meningism. All the signs are not always present. Cd_1056_014c.jpg * There is proteinuria, pyuria and microscopic haematuria. The cerebrospinal fluid initially contains neutrophils. Later lymphocytes predominate, together with elevated protein and normal glucose. In general, there is significant leukocytosis, but this too is not constant. Thrombocytopenia is common. Early in the course of the disease leptospires can rarely be found in the blood, urine or cerebrospinal fluid (the tests are not very sensitive). Subsequently the bacteria are only found in the urine. Since these are very thin organisms (0.1m diameter) a dark-field microscope is needed to detect them. Indirect illumination is used in this method instead of direct illumination, so that fine structures can be detected which are not visible with the traditional microscope. This method is not very sensitive, however, and has been responsible for many errors (many false positives and false negatives). Culture of the bacteria is the gold standard but is not practical in most developing countries, since special media are required. Serology can be performed. Since the traditional serology using micro-agglutination test or MAT, requires a well-functioning laboratory, this too will not be available in practice in third world countries. For this reason, in 1997 a simple dipstick method (the LEPTO Dipstick) was developed to detect anti-leptospire IgM in serum. The test is based on a broadly reactive leptospire antigen fixed to a solid strip, together with stabilised monoclonal anti-IgM which has been conjugated with a dye as a signal system. The sensitivity and specificity are quite good (both approximately 90%). The test is not available everywhere and still has to be validated in various geographical regions. Interpretation of MAT serology results to identify the responsible serovars is rather difficult, because the highest titre does not necessarily correlate with the actual serovar which is responsable the infection.

8 Differential diagnosis
This is very broad in view of the variable symptoms. It includes influenza, gastro-enteritis, meningitis, malaria, hepatitis, cholangitis, rickettsiosis (e.g. scrub typhus), borreliosis, typhoid fever, Reyes syndrome, arboviroses such as yellow fever, Rift valley Fever, Crimean-Congo haemorrhagic fever and West Nile fever as well as arenaviroses. In the case of haemorrhagic tendency, Gram-negative septicaemia and the various viral haemorrhagic fevers should be considered.

9 Treatment
The earlier treatment is started, the better the results. Antibiotics such as tetracyclines within the first 4 days are effective in shortening the illness. Sometimes leptospires persist in urine for a long time, even with correct treatment. It is preferable to give doxycycline 200 mg per day for 1 week. If there is vomiting IV penicillin is used. Ampicillin is also active against leptospires. Chloramphenicol, on the other hand, is not. Symptomatic and supportive therapy is very important.

10 Prevention
Cd_1079_057c.jpg Since rats form the main reservoir and contaminate surface water and drains, their control is important for prevention. Nevertheless it should not be forgotten that the animal reservoir is much broader (e.g. dogs etc.) and cannot be eradicated completely. Wearing boots when working in infected water is advisable, but sometimes utopian. Chemoprophylaxis of 200 mg doxycycline per week may be taken as a preventative in high-risk situations. After infection there is protection against the infecting serovar (the serological variety) but there is no crossimmunity. * A vaccine has been developed by the Pasteur Institute. It is aimed at L. icterohaemorrhagicae. This vaccine can be used for persons in high-risk occupations. Two injections are given SC with a 15-day interval, followed by a repeat after 6 months and then every 2 years. The vaccine is difficult to obtain and is of limited benefit. Many other vaccine candidates have unacceptable side effects and give brief protection without cross-immunity. There are vaccines for animals.

11 Exercises
.1 A labourer in Santos, a port in Brazil, suddenly gets a high fever. His eyes are bloodshot, but there is no pus. He has a terrible headache and pronounced muscle pains. He is coughing. The diagnosis of influenza is made. A few days later his urine is dark and hepatitis is suspected. Another few days later there is still severe headache and neck stiffness occurs. Meningitis is considered. Lumbar puncture shows an increased lymphocyte count, but no organisms. The man remarks that his stools are black and smell bad. Is the fact that 10 days

ago he fell into a sewer relevant? Is the detail of black stools relevant? If it is wished to rule out the initial diagnosis of fulminant hepatitis A, and serology will take too long, what could liver tests contribute? .2 A cyclone has led to heavy rains and floods in Myanmar (Burma). You find a man with a dry cough, fever, jaundice and various haemorrhages (nose, gums, melena). Your colleague thinks this is a case of yellow fever. What do you think? What about the possibility of scrub typhus? .3 Ethiopia. A large family of refugees arrives in a camp. One man has had a high fever, headache and muscle pain for three days. There is pronounced cough and dyspnoea. His eyes are bloodshot. This morning the fever has subsided but his general condition has worsened. He is bleeding from nose and mouth. You notice red spots on his arms, torso and legs which do not blanch upon pressure. The liver and spleen are swollen. You are told that the previous week the brother of this patient died with identical symptoms. His wife has also had a similar illness, but has survived. She has lost her unborn child, however, and now has paralysis of the left side of her face. Do you expect anything from a thick smear? What do you do? To avoid further cases, someone suggests finding and destroying all rats and rodents in the camp. What do you think? .4 Eritrea. A 16-year-old girl has had an ulcer on her right leg for two months. The edges are slightly elevated. The lesion is not painful, but is very gradually widening. Your colleague thinks it is a tropical ulcer. What do you think? Do you begin treatment with penicillin? .5 A white woman is visiting a number of remote villages in Senegal as a tourist. She is sleeping under primitive conditions "like the local people". A little while later she is febrile and there is pronounced malaise. After a few days she feels better. Two weeks later she develops the same symptoms. Malaria is suspected, but no trophozoites are found in a thick smear. It is reported that the quality of the smear and the thick smear are not so good, because some microscopic threads of cotton wool seem to have adhered to the slide. What do you think? Possible treatment? .6 Cuba. A man falls overboard at sea from a sailing boat when he is 5 km out from the coast. He is rescued and resuscitated. If he develops a cough or fever, should leptospirosis be included in the differential diagnosis? .7 Sumatra. You are visiting a village in swampy surroundings. You are told that there has never been a case of leptospirosis in the whole district. The argument is made that leptospires

have never been found in blood or urine and that they therefore no longer need to be tested for. What do you say? .8 Name three different spirochaetes which are neurotropic. .9 Timor. Three children were recently admitted with leptospirosis. They waded recently through a local lake. A Swiss fish biologist is doing research and has to go into the water daily for the next 4 weeks. Which advice on prevention do you give him? .10 Why do you think that historically leptospires were discovered during a yellow fever epidemic? .11Thailand. At the Cambodian border, a professional photographer is planning to take pictures of water beetles in their natural biotope every day over the next 4 weeks. Someone advises him to take one tablet of vibramycin daily. Any idea why? .12Nicaragua. In 1995 in the region of Achuapa, after excessive rainfall with floods there was an epidemic of an acute febrile illness, accompanied by headache, muscle pain, dyspnoea and pulmonary haemorrhages. In the end this proved to be a new variant of leptospirosis. What diseases should you include in the initial differential diagnosis: dengue, yellow fever, arenaviruses, filoviruses, rickettsioses, hantaviruses?