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More important is the increase in resistance to some antimicrobial agents, particularly the resistance to trimethoprimsulfamethoxazole seen in E. coli. Physicians distinguish UTIs from other diseases that have similar clinical presentations with use

For urine specimens obtained from outpatients, it is not necessary to routinely inoculate a medium that is selective for grampositive bacteria, because nearly all UTIs in outpatients are caused by aerobic and facultative gram-negative bacteria.

Even in patient populations in which Staphylococcus saprophyticus is a common cause of UTIs, it is not necessary to use selective

media and urine specimens which obtained from hospitalized patients are likely to contain Enterococci, which have emerged as

the second most common cause of nosocomial infections.

Urine cultures should be incubated overnight at 35°C-37°C in ambient air before being read. There is no added benefit to

incubating routine urine cultures for 48 h, provided that specimens are incubated for a full 24 h and that urine specimens

containing <104

uropathogens or specimens from patients with suspected funguria are incubated for 48 hours.

Most pathogenic yeasts grow well on blood agar plates, so it is unnecessary to use selective fungal media for urine cultures, even

for samples obtained from patients with suspected funguria. Selective fungal media can be used in those rare instances in which

there is a high clinical probability that a UTI is caused by a more fastidious yeast or mold. Urine specimens obtained from patients

with suspected mycobacterial UTIs should be processed and plated to the appropriate mycobacterial media

Detection of bacteriuria by urine microscopy

Bacteriuria can be detected microscopically using Gram staining of uncentrifuged urine specimens, Gram staining of centrifuged

specimens, or direct observation of bacteria in urine specimens. Gram stain of uncentrifuged urine specimens is a simple method.

A volume of urine is applied to a glass microscope slide, allowed to air dry, stained with Gram stain, and examined

microscopically.

The performance characteristics of the test are not well-defined, because different criteria have been used to define a positive test

result. In one study, the test was found to be sensitive for the detection of ⩾105 CFU/ml but insensitive for the detection of lower

numbers of bacteria.

The urine Gram stain test has the important advantage of providing immediate information as to the nature of the infecting

bacterium or yeast and thereby guiding the physician in selecting empiric antimicrobial therapy. This is of importance in some

settings, but the Gram stain test has 3 disadvantages that limit its usefulness in most clinical settings:

1- It is an insensitive test, being reliably positive only if the concentration of bacteria in the urine is ⩾105 CFU/ml; infections

with bacterial concentrations of 102

-103 CFU/ml may not be detected by this test.

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2- The test is too labor intensive for it to be practical for most clinical microbiology laboratories to offer it on more than a select

basis.

3- Because it may not detect bacteria at concentrations of 102

-103 CFU/ml, it should not be used in the outpatient setting for

patients with uncomplicated UTIs.

Because of these limitations, its use should be limited to patients with cases of acute pyelonephritis, patients with invasive UTIs,

or other patients for whom it is important to have immediate information as to the nature of the infecting pathogen.

Detection of bacteriuria by nitrite test

Bacteriuria can be detected chemically when bacteria produce nitrite from nitrate. The biochemical reaction that is detected by the

nitrite test is associated with members of the family Enterobacteriaceae (the pathogens most commonly responsible for UTIs),

but the usefulness of the test is limited because nitrite production is not associated with urinary-tract pathogens such as S.

saprophyticus, Pseudomonas species, or Enterococci.

Another limitation to the test is that it requires testing a specimen of the first urine produced in the morning, as ⩾4 h are required

for bacteria to convert nitrate to nitrite at levels that are reliably detectable.

Detection of pyuria by urine microscopy

Pyuria can be detected and quantified microscopically by measuring the urinary leukocyte excretion rate, counting leukocytes

with a hemocytometer, counting leukocytes in urine specimens using Gram staining, or counting leukocytes in a centrifuged

specimen.

The advantages to urine microscopy are that leukocytes, leukocyte casts, and other cellular elements are observed directly. One

disadvantage to urine microscopy is that leukocytes deteriorate quickly in urine that is not fresh or that has not been adequately

preserved. Because of these disadvantages, urine microscopy should be limited to patients in whom pyelonephritis or other more

serious infections are suspected.

The most accurate microscopic method for quantitating pyuria is to measure the urinary leukocyte excretion rate. Patients with

symptomatic UTIs have urinary leukocyte excretion rates of ⩾400,000 leukocytes/hour. The test is impractical for clinical use,

however, making it necessary for laboratories to use other methods. A simple and inexpensive alternative is to count urine

leukocytes with a hemocytometer. Comparison of hemocytometer counts with urinary leukocyte excretion rates has shown that a

hemocytometer count of ⩾10 leukocytes/mm3

correlates with a urinary leukocyte excretion rate of ⩾400,000 leukocytes/h.

Moreover, the correlation of hemocytometer counts with urine colony counts has shown that patients with symptomatic UTIs and

bacterial concentrations of >105 CFU/ml have urine leukocyte counts of ⩾10 leukocytes/mm3

. Although using a hemocytometer

to count leukocytes is easier than measuring urinary leukocyte excretion rates, it is impractical for clinical laboratories to use a

hemocytometer to count leukocytes on a routine basis. The most practical microscopic method involves counting the number of

leukocytes in the sediment of centrifuged urine specimens; this method is inaccurate because of inadequate standardization of the

method. For these reasons, and to facilitate the processing of large numbers of specimens, most laboratories use rapid tests for

leukocyte esterase as a surrogate for microscopic leukocyte counts.

Detection of pyuria by leukocyte esterase tests

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Leukocyte esterase tests are based on the hydrolysis of ester substrates by proteins with esterolytic activity; human neutrophils

produce as many as 10 proteins with esterolytic activity. These proteins react with ester substrates to produce alcohols and acids

that then react with other chemicals to produce a color change that is proportional to the amount of esterase in the specimen.

These tests have the advantage of detecting both esterases in intact leukocytes and esterases released after cell lysis; therefore,

even specimens that have not been preserved properly may yield a positive test result.

Leukocyte esterase tests can yield false-positive test results when the urine is contaminated with bacteria present in vaginal fluid;

when the specimen contains eosinophils or Trichomonas species, both of which can act as sources of esterases; and when

oxidizing agents or formalin react with the test strips to generate false-positive test results.

Leukocyte esterase tests may show a decrease in positive test results when the specimen has an elevated specific gravity and/or

elevated levels of protein and glucose; when boric acid preservatives are present; when large amounts of ascorbic or oxalic acid

are present; and when the patient has received antimicrobial agents, such as Cephalothin, Cephalexin, or Tetracycline.

Routine bacterial urine cultures

Urine culture may not be necessary as part of the evaluation of outpatients with uncomplicated UTIs. However, urine cultures are

necessary for outpatients who have recurrent UTIs, experience treatment failures, or have complicated UTIs.

Urine cultures are also necessary for inpatients that develop UTIs. The bacterial culture remains an important test in the diagnosis

of UTI, not only because it helps to document infection, but also because it is necessary for determination of the identity of the

infecting microorganism(s) and for antimicrobial susceptibility testing. This is particularly true because of the increased incidence

of antimicrobial resistance.

The most commonly used criterion for defining significant bacteriuria is the presence of ⩾105 CFU per milliliter of urine. This

criterion was established only for women with acute pyelonephritis or women who were asymptomatic but had multiple urine

cultures that yielded this number of bacteria; however, the criterion is often applied to other patient populations. Most patients

with UTIs, however, do not fall into either category, and 30%-50% of patients with acute urethral syndrome will have colony

counts of <105 CFU/ml. For this reason, many laboratories have opted to use lower colony counts as a criterion for interpreting

and reporting results. One common criterion is a colony count of 104 CFU/ml, which would be expected to increase the sensitivity

of the test without making the test impractical for clinicians and laboratories to use.

Catheterized patients (who may have low concentrations of bacteria that can progress to higher concentrations) and many patients

with infections of the lower urinary tract have colony counts much lower than 105 CFU/ml if the specimens are obtained via

suprapubic aspirate or catheterization. Accordingly, the most appropriate diagnostic criterion for urine culture specimens obtained

via suprapubic aspirate or catheterization is a bacterial concentration of ⩾102 CFU/ml.

Follow-up cultures are recommended for patients with infections that do not respond to therapy, patients who have recurrent

UTIs, patients who have anatomic or functional abnormalities of the urinary tract, or patients who continue to have unexplained

abnormal urinalysis findings.

Interpretation of urine culture results

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Microbiologists need to interpret the microbiologic relevance of growth on culture plates to determine whether further

identification and antimicrobial susceptibility testing are necessary. Most culture results can be interpreted readily; no growth and

gross contamination are both unambiguous results, as are pure cultures of common pathogens growing in a quantity of >105 CFU

per milliliter of urine.

The interpretation of cultures that yield pure growth in lower quantities is also clear for specimens obtained via suprapubic

aspiration or straight catheterization. On the other hand, interpretation of urine cultures that yield mixed flora in varying quantities

can be difficult. Test reports for cultures that yield mixed flora in varying quantities should specify the microorganisms that were

recovered, the quantity of each microorganism, and the probable clinical importance of each isolate.

Antimicrobial susceptibility testing

Each laboratory should have guidelines by which pathogens are tested for antimicrobial susceptibility. These guidelines should be

developed and antimicrobial susceptibility tests should be performed and reported according to the most recent version of the

NCCLS guidelines. Bacterial or fungal isolates of uncertain clinical importance should not be tested for antimicrobial

susceptibility for purposes of routine patient care

Most patients with uncomplicated acute cystitis have cases that are clinically straightforward, and they may not require any

laboratory testing beyond urinalysis. For a significant number of patients, however, the clinical history and physical findings

alone may be insufficient to make a definitive diagnosis of UTI.

For those patients and for patients with complicated UTIs, laboratory tests are necessary to make the diagnosis and to provide

specific information regarding the identity and the antimicrobial susceptibility pattern of pathogens. Both the laboratory diagnosis

and the clinical diagnosis of laboratory test results must be made in light of the method of collection used; clinicians should

specify the method of collection on test requisition forms.

Of the available laboratory tests, urinalysis is helpful primarily as a means of excluding bacteriuria, but it is not a surrogate for

culture. Although cultures identify pathogens, the accurate interpretation of culture results requires clinical information that is

usually available only to the clinician.

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Bloodstream Infections :

Invasion of the bloodstream by microorganisms constitutes one of the most serious situations in

infectious disease. Microorganisms present in the circulating blood—whether continuously, intermittently, or

transiently—are a threat to every organ in the body.

The suffix emia is derived from the Greek word meaning “blood” and refers to the presence of a substance in

the blood; bacteremia refers to the presence of bacteria in the blood, fungemia refers to the presence of fungi

in the bloodstream, and septicemia indicates bacteria are present in the blood, producing an infection and

reproducing within the bloodstream.

Microbial invasion of the bloodstream resulting from any organism can have serious immediate

consequences, including shock, multiple organ failure, disseminated intravascular coagulation (DIC), and death.

Approximately 200,000 cases of bacteremia and fungemia occur annually, with mortality

rates ranging from 20% to 50%. Timely detection and identification of blood-borne pathogens are two of the

most important functions of the microbiology laboratory.

Pathogens of all four major groups of microbes—bacteria, fungi, viruses, and parasites—may be found

circulating in blood during the course of many diseases. Positive blood cultures may help provide a clinical

diagnosis, as well as a specific etiologic diagnosis.

General considerations:

The successful recovery of microorganisms from blood by the laboratory depends on many, often complex,

factors: the type of bacteremia, the specimen collection method, the blood volume, the number and timing of

blood cultures, the interpretation of results, and the type of patient population being served by the laboratory.

All of these parameters must be considered in the development of the blood culture protocol within the

laboratory in order to maximize the detection and recovery of microorganisms and ensure quality patient care.

Etiology:

As previously mentioned, all major groups of microbes can be present in the bloodstream during the course of

many diseases.

Bacteria:

The organisms most commonly isolated from blood are gram-positive cocci, including coagulase-negative

staphylococci, Staphylococcus aureus, and Enterococcus spp., and other organisms likely to be inhabitants of

the hospital environment that colonize the skin, oropharynx, and gastrointestinal tract of patients. Some of the

most common, clinically significant bacteria isolated from

blood cultures are listed in Table 1. (Table 1 Organisms Commonly Isolated from Blood Cultures)

Organisms Commonly Isolated from Blood Cultures:

Staphylococcus aureus

Escherichia coli

Coagulase-negative staphylococci

Enterococcus spp.

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Candida albicans

Pseudomonas aeruginosa

Klebsiella pneumoniae

Viridans streptococci

Streptococcus pneumoniae

Enterobacter cloacae

Proteus spp.

Beta-hemolytic streptococci

Anaerobic bacteria: Bacteroides and Clostridium spp.

In general, the number of fungi and coagulase-negative staphylococc has increased, whereas the number of

clinically significant anaerobic isolates has decreased since the early 2000s.

Of importance, the laboratory isolation of certain bacterial species from blood can indicate the presence of an

underlying, occult, or undiagnosed neoplasm. Alterations in local conditions at the site of the neoplasm

allowing bacteria to proliferate and seed the bloodstream have been suggested as a potential mechanism for the

association between bacteremia and cancer. Another possible mechanism is reduced killing of bacterial cells by

the host phagocytes. Organisms associated with neoplastic disease include Clostridium septicum and other

uncommonly isolated clostridial species, Streptococcus galldyticus, Aeromonas hydrophila, Plesiomonas

shigelloides, and Campylobacter spp. Finally, if Streptococcus anginosis group bacteria are isolated from

blood, the possibility of an abscess should be considered.

Fungi:

Fungemia (the presence of fungi in blood) is usually a serious condition, occurring primarily in

immunosuppressed patients and in those with serious or terminal illness. Candida albicans is by far the most

common species, but Malassezia furfur can often be isolated in patients, particularly neonates, receiving lipidsupplemented parenteral nutrition. Candida spp. account for approximately 8% to 10% of all nosocomial

bloodstream infections. Except for Histoplasma, which multiply in leukocytes

(white blood cells), fungi do not invade blood cells, but their presence in the blood usually indicates a focus of

infection elsewhere in the body.

Fungi in the bloodstream can disseminate (be carried) to all organs of the host, where they may grow, invade

normal tissue, and produce toxic products. Fungi gain entrance to the circulatory system via loss of integrity of

the gastrointestinal or other mucosa; through damaged skin; from primary sites of infection, such as the lung or

other organs; or by means of intravascular catheters. Systemic fungal infections begin as pneumonia and may

disseminate from the lungs, which serve as the portal of entry. Arthroconidia of Coccidioides immitis and

microconidia of Histoplasma capsulatum and Blastomyces dermatitidis are ingested by alveolar macrophages

in the lung.

These macrophages carry the fungi to nearby lymph nodes, usually the hilar nodes. The fungi multiply within

the node tissue and ultimately are released into the circulating blood, from which they are capable of seeding

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other organs or are destroyed by the body’s defenses. Molds are particularly insensitive to host defenses such as

antibody and phagocytic cells because of their large size and their sterol containing cell wall structure.

Parasites:

Eukaryotic parasites may be found transiently in the bloodstream as they migrate to other tissues or organs.

Their presence, however, cannot be considered consistent with a state of good health. For example, tachyzoites

of the parasite Toxoplasma gondii may be found in circulating blood. They invade cells within lymph nodes

and other organs, including the lungs, liver, heart, brain, and eyes. The resulting cellular destruction accounts

for the manifestations of toxoplasmosis. Also, microfilariae are

seen in peripheral blood during infection with Dipetalonema, Mansonella, Loa loa, Wuchereria, or Brugia.

Malarial parasites invade host erythrocytes and hepatic parenchymal cells. The significant anemia and

subsequent tissue hypoxia (reduction in oxygen levels) may result from destruction of red blood cells by the

parasite. Vascular trapping of normal erythrocytes by the infected red blood cells, which are less flexible and

tend to clog small capillaries, is a major cause of morbidity. The host’s immunologic response is to remove the

parasites and damaged red blood cells; the immune response may also

have deleterious effects. Parasites in the bloodstream are usually detected by direct visualization. Those

parasites for which traditional diagnosis is dependent on observation of the organism

in peripheral blood smears include Plasmodium, Trypanosoma, and Babesia. Patients with malaria or filariasis

may display a periodicity in their episodes of fever that allows the physician to time the collection of blood for

microscopic examination intended for optimal detection. Rapid serological methods and molecular methods are

currently used to detect malaria, babesiosis, and trypanosomiasis.

Viruses:

Although many viruses do circulate in the peripheral blood at some stage of disease, the primary pathology

relates to infection of the target organ or cells. Those viruses that preferentially infect blood cells are EpsteinBarr virus (invades lymphocytes), cytomegalovirus (invades monocytes, polymorphonuclear cells, and

lymphocytes), and human immunodeficiency virus (HIV)

(involves only certain T lymphocytes and perhaps macrophages) and other human retroviruses that attack

lymphocytes. The pathogenesis of viral diseases of the blood is the same as that for viral diseases of any organ;

by diverting the cellular machinery to create new viral components

or by other means, the virus may prevent the host cell from performing its normal function. The cell

may be destroyed or damaged by viral replication, and immunologic responses of the host may also contribute

to the pathogenesis. Although many viral diseases have a viremic stage,

recovery of virus particles or detection of circulating viruses is used in the diagnosis of only a few diseases.

Types of bacteremia:

Bacteremia may be transient, continuous, or intermittent. Most people have experienced transient bacteremia;

teething infants and people having dental procedures have had oral flora gain entry to the bloodstream through

breaks in the gums. Other conditions in which bacteria are only transiently present in the bloodstream include

manipulation of infected tissues, devices or instrumentation inserted through contaminated mucosal surfaces,

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and surgery involving nonsterile sites. These circumstances may also lead to significant septicemia, although

normally the bacteria are cleared from the blood by scavenging

leukocytes, resulting in no infection. Septicemia can occur when the bacteria multiply more rapidly than the

immune system is capable of killing and removing the organism. In septic shock, bacterial endocarditis, and

other endovascular infections, organisms are released into the bloodstream at a fairly constant rate (continuous

bacteremia). Also, during the early stages of specific infections, including typhoid fever, brucellosis, and

leptospirosis, bacteria are continuously present in the bloodstream.

In most other infections, such as in patients with undrained abscesses, bacteria can be found intermittently in

the bloodstream. Of note, the causative agents of meningitis, pneumonia, pyogenic arthritis, and osteomyelitis

are often recovered from blood during the early course of these diseases. In the case of transient seeding of the

blood from a sequestered focus of infection, such as an

abscess, bacteria are released into the blood approximately 45 minutes before a febrile episode.

The symptoms of septicemia are fever, chills, and malaise; these are caused by the presence of the invading

microorganism and the toxins produced by these microorganisms. The older the patient is, the greater the risk

and the rate of mortality as a result of septicemia.

Types of blood stream infections:

The two major categories of bloodstream infections are intravascular (those that originate within the

cardiovascular system) and extravascular (those that result from bacteria entering the blood circulation through

the lymphatic system from another site of infection). Of note, other organisms, such as fungi, may also cause

intravascular or extravascular infections. However, because bacteria account for the majority of significant

vascular infections, these types of bloodstream infections are discussed in more detail. Factors contributing to

the initiation of bloodstream infections are immunosuppressive agents, widespread use of broad-spectrum

antibiotics that suppress the normal flora and allow the emergence

of resistant strains of bacteria, invasive procedures allowing bacteria access to the interior of the host, more

extensive surgical procedures, and prolonged survival of debilitated and seriously ill patients.

1) Intravascular Infections:

Intravascular infections include infective endocarditis, mycotic aneurysm, suppurative thrombophlebitis, and

intravenous (IV), catheter-associated bacteremia. Because these infections are within the vascular system,

organisms are present in the bloodstream at a fairly constant rate (i.e., a continuous bacteremia). These

infections in the cardiovascular system are extremely serious and considered life threatening.

Infective Endocarditis. The development of infective endocarditis (infection of the endocardium most

commonly caused by bacteria) is believed to involve several independent events. Cardiac abnormalities, such as

congenital valvular diseases that lead to turbulence in blood flow or direct trauma from IV catheters, can

damage cardiac endothelium. This damage to the endothelial

surface results in the deposition of platelets and fibrin. If bacteria transiently gain access to the bloodstream

(this can occur after an innocuous procedure such as brushing the teeth) after alteration of the capillary

endothelial cells, the organisms may stick to and then colonize the damaged cardiac endothelial cell surface.

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After colonization, the surface will rapidly be covered with a protective layer of fibrin and platelets. This

protective environment is favorable to further bacterial multiplication. This web of platelets, fibrin,

inflammatory cells, and entrapped organisms is called a vegetation (Figure -1).

The resulting vegetations ultimately seed bacteria into the blood at a slow but constant rate. The primary

causes of infective endocarditis are the viridans streptococci, comprising several species (Box 68-2). These

organisms are normal inhabitants of the oral cavity, often gaining entrance to the bloodstream as a result of

gingivitis, periodontitis, or dental manipulation. Heart valves, especially those previously damaged, present

convenient surfaces for attachment of these bacteria. Streptococcus sanguis and Streptococcus mutans are

frequently isolated in streptococcal endocarditis.

Gram-negative bacilli, known as the AACEK group, Aggregatibacter aphrophilus, Actinobacillus

actinomycetemcomitans Cardiobacterium hominis, Eikenella corrodens, and Kingellakingae, can also be

associated with endocarditis.

With the ever-increasing use of IV catheters, arterial lines, and vascular prostheses, organisms considered

normal or hospital-acquired inhabitants of the human skin are able to gain access to the bloodstream and attach

to various surfaces, including heart valves and vascular endothelium. It has been estimated that more than

200,000 nosocomial infections (bloodstream) occur annually in the United States in adults and children. The

majority of these infections are caused by the use of intravascular catheters. Staphylococcus epidermidis and

other coagulase-negative staphylococci have been increasingly implicated as the cause of infection associated

with intravascular catheters. S. epidermidis is the most common etiologic agent identified in prosthetic valve

endocarditis, with S. aureus being the second most common. S. aureus is an important cause of septicemia

without endocarditis and is found in association with other foci, such as abscesses, wound infections, and

pneumonia, as well as sepsis related to indwelling intravascular catheters.

Figure 1 Vegetations of bacterial endocarditis. Arrow indicates the vegetations.

(Courtesy Celeste N. Powers, MD, PhD, Virginia Commonwealth

University Medical Center, Medical College of Virginia Campus, Richmond, Va.

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Table 2 Agents of Infective Endocarditis

Agents of Infective Endocarditis:

Aggregatibacter aprophilus

Viridans streptococci*

Nutritionally deficient streptococci (Abiotrophia spp. and

Granulicatella spp.)

Enterococci*

Streptococcus bovis

Staphylococcus aureus*

Staphylococci (coagulase-negative)

Enterobacteriaceae

Pseudomonas spp. (usually in drug users)

Haemophilus spp.

Unusual gram-negative bacilli (e.g., Actinobacillus,

Cardiobacterium, Eikenella, Coxiella burnetii)

Yeast

Other (including polymicrobial infectious endocarditis)

*Most common organisms associated with native valve endocarditis in non-drug-using

adults.

Table -3 Common Agents of IV Catheter–Associated Bacteremia:

Staphylococcus epidermidis

Other coagulase-negative staphylococci

Staphylococcus aureus

Enterobacteriaceae

Pseudomonas aeruginosa

Figure 2 Short-term, triple-lumen central venous catheter. The ends from which the catheter is accessed

are usually referred to as the hubs. After the catheter is inserted, the tip resides within the bloodstream.

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