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ORIGINAL ARTICLE
Year : 2016  |  Volume : 29  |  Issue : 2  |  Page : 464-467

Study of the role of ascitic fluid lactoferrin levels in the diagnosis of spontaneous bacterial peritonitis


1 Department of Tropical Medicine, Faculty of Medicine, Menoufia University, Al Minufya, Egypt
2 Department of Clinical Pathology, Faculty of Medicine, Menoufia University, Al Minufya, Egypt

Date of Submission15-May-2013
Date of Acceptance12-May-2016
Date of Web Publication18-Oct-2016

Correspondence Address:
Ahmed Elmaaz
Department of Tropical Medicine, Faculty of Medicine, Menoufia University, Al Minufya, 32511
Egypt
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Source of Support: None, Conflict of Interest: None


DOI: 10.4103/1110-2098.192416

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  Abstract 

Objectives:
To study the role of ascitic fluid (AF) lactoferrin as a surrogate marker for the diagnosis of spontaneous bacterial peritonitis (SBP).
Background:
SBP is one of the most dangerous and life-threatening complications of liver cirrhosis and ascites. Its diagnosis is difficult, operator dependent, and time consuming.
Materials and methods:
This study was conducted on 60 patients with decompensated chronic liver disease and ascites admitted to the Tropical Medicine Department at Menoufia University Hospital. These patients were classified into two groups: group I (non-SBP group) and group II (SBP group). AF samples from both groups were examined for polymorphonuclear leukocyte cell count, AF culture, and lactoferrin levels.
Results:
The results showed a highly significant increase in AF lactoferrin in the SBP group and at a cut-off level of 255 ng/ml; the sensitivity and specificity of AF lactoferrin in the diagnosis of SBP were 100 and 88.9%, respectively. AF culture revealed the presence of organisms in 43.4% of patients in the SBP group; all organisms were gram negative.
Conclusion:
Our results support that AF lactoferrin could be used as a marker for screening and diagnosing SBP in patients with cirrhosis and ascites.

Keywords: ascites, cirrhosis, lactoferrin, spontaneous bacterial peritonitis


How to cite this article:
Abulseoud A, Ibrahim H, Essa A, Essa E, Elmaaz A. Study of the role of ascitic fluid lactoferrin levels in the diagnosis of spontaneous bacterial peritonitis. Menoufia Med J 2016;29:464-7

How to cite this URL:
Abulseoud A, Ibrahim H, Essa A, Essa E, Elmaaz A. Study of the role of ascitic fluid lactoferrin levels in the diagnosis of spontaneous bacterial peritonitis. Menoufia Med J [serial online] 2016 [cited 2019 Nov 15];29:464-7. Available from: http://www.mmj.eg.net/text.asp?2016/29/2/464/192416


  Introduction Top


Cirrhosis represents the final common histological pathway for a wide variety of chronic liver diseases. Cirrhosis is defined histologically as a diffuse hepatic process characterized by fibrosis and the conversion of normal liver architecture into structurally abnormal nodules. Some patients with cirrhosis are completely asymptomatic and have a reasonably normal life expectancy[1].

Occurrence of ascites is the most common presentation of liver cirrhosis, and is defined as an accumulation of excessive fluid within the peritoneal cavity and may be a complication of both hepatic and nonhepatic diseases [2].

Spontaneous bacterial peritonitis (SBP) is observed in 15–26% of patients hospitalized with ascites. The syndrome arises most commonly in patients whose low-protein ascites (<1 g/dl) contain low levels of complement, resulting in decreased opsonic activity. SBP appears to be caused by the translocation of gut bacteria across the gut wall and also by the hematogenous spread of bacteria. The most common causative organisms are  Escherichia More Details coli, Streptococcus pneumoniae, Klebsiella spp., and other gram-negative enteric organisms [3]. SBP is defined as the infection of ascitic fluid (AF) in the absence of a contiguous source of infection and/or an intra-abdominal inflammatory focus. An AF-polymorphonuclear (PMN) leukocyte count more than or equal to 250/mm irrespective of the AF culture result is universally accepted nowadays as the best surrogate marker for diagnosing SBP. Frequently, the results of the manual or automated PMN count do not reach the hands of the responsible medical personnel in a timely manner [4].

As a result of the potential for delays in the diagnosis of SBP, there has been interest in developing a surrogate test to identify an elevated AF-PMN cell count. Leukocyte reagent strips were evaluated in an initial attempt to develop a rapid screening test for the diagnosis of SBP. Even when lowering the threshold for positivity, the sensitivity of the leukocyte reagent strip test only improved to 79% [5].

Lactoferrin is an iron-binding protein that is found mainly in external secretions such as breast milk and in PMN leukocytes. Lactoferrin is released from PMN leukocytes on activation of these cells, and its presence in body fluids is proportional to the flux of neutrophils. It is hypothesized that measurement of AF lactoferrin could be clinically useful for the detection of SBP in patients with cirrhosis [6].


  Aim of the Study Top


The aims of this study were to evaluate the usefulness of AF lactoferrin for the diagnosis of SBP and to identify a clinically useful cut-off level that can be used for future development of an important clinical, economic, and time-saving rapid bedside test for the diagnosis of SBP in cirrhotic ascites.


  Materials and Methods Top


This study was conducted on 60 patients with decompensated chronic liver disease and ascites admitted to the Tropical Medicine Department at Menoufia University Hospital. These patients were selected according to the following inclusion and exclusion criteria.

Inclusion criteria

Patients with decompensated chronic liver diseases (Child B and C cirrhosis with ascites). They were classified into two groups.

Group I (non-SBP group): included 30 patients with cirrhotic ascites without SBP.

Group II (SBP group): included 30 patients with cirrhotic ascites with SBP.

Bacterial peritonitis was diagnosed when AF-PMN was more than 250/mm 3 with positive AF culture (SBP) or with negative AF culture (culture-negative neutrocytic ascites), as well as positive AF culture with PMN less than 250/mm 3 (non-neutrocytic bacterascites), in the absence of data compatible with secondary peritonitis.

Exclusion criteria

Patients with ascites due to any cause other than liver cirrhosis, evidence of active infection other than AF infection, prehospitalization antibiotic administration, any other cause of neutrocytic ascites such as pancreatitis, appendicitis, tuberculosis, peritoneal carcinomatosis, or hemorrhagic ascites, as well as patients with history of abdominal surgery within 3 months of the study were excluded from the present study.

All patients were subjected to thorough history, complete clinical examination, laboratory investigations (including complete blood count, erythrocyte sedimentation rate, liver and renal function tests, and viral markers), abdominal ultrasonography, and AF analysis (including total and differential cell count, total proteins, albumin, aerobic and anaerobic bacteria culture, and AF lactoferrin levels) using enzyme-linked immunosorbent assay (Assay Max Human Lactoferrin ELISA Kit, Catalog No. EL211; ASSYPRO, Saint Charles, Missouri, USA). This assay employs a quantitative sandwich enzyme immunoassay technique.

The procedures followed were in accordance with the ethical standards of the responsible institutional committee on human experimentation.


  Results Top


Out of the 60 patients included in the present study, 35 of them (58.3%) were males and 25 (41.7%) were females. Their ages ranged from 42 to 70 years, with a mean age of 56.17 ± 7.90.

Hepatitis C virus infection was the main etiology of liver cirrhosis in both groups, whereas hepatitis B virus infection was much less common. Hepatitis C virus and hepatitis B virus coinfection was present in 20% of the SBP group.

AF culture revealed the presence of organisms in 43.4% of patients in the SBP group (all organisms were gram negative) and absence of organisms in all non-SBP patients ([Table 1]).
Table 1: Ascitic fluid bacterial culture in non-SBP and SBP groups

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Cellular examination of AF revealed a highly significant increase in AF total leukocytic and PMN cell counts in the SBP group when compared with the non-SBP group ([Table 2]).
Table 2: AF mean total leukocytic and PMN cell counts in the studied groups

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Our results also revealed a highly significant increase in AF lactoferrin in the SBP group when compared with the non-SBP group with mean values of 3434.8 and 140.7 ng/ml, respectively ([Table 3]).
Table 3: AF lactoferrin levels in the studied groups

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A receiver operating characteristic curve was generated and showed that AF lactoferrin, at the cut-off level of 255 ng/ml, can distinguish patients with SBP from those without SBP with sensitivity (100%) and specificity (88.9%). The area under the receiver operating characteristic curve was 0.98 ([Figure 1]).
Figure 1: Receiver operating characteristic curve of ascitic fluid lactoferrin

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  Discussion Top


SBP is a common and serious infection occurring in 8–27% of hospitalized patients with cirrhosis and ascites [7]. It is also one of the potential life-threatening complications in cirrhotic patients with ascites, with mortality ranging between 30 and 50% [8], and therefore it requires a rapid and accurate diagnosis in addition to prompt effective therapy [9].

Lactoferrin is one of the main immune proteins of the neutrophil granulocytes, stored in secondary granules where it resides until cellular activation. On contact with a foreign antigen, these cells degranulate and release a number of antibacterial and antiviral peptides, including lactoferrin. The major function of lactoferrin probably resides in its bactericidal effects, either by sequestering free iron or by the effects of lactoferricin, an antibacterial peptide generated by proteolytic cleavage of lactoferrin [0].

Regarding this study, AF bacterial cultures were positive in 13 cases (43.4%) of the SBP group and negative in all cases of the non-SBP group. This is in agreement with Llovet et al. [1] who detected positive AF bacterial cultures in 41.7% of cases of SBP. However, culture-negative neutrocytic ascites, which are considered as a variant of SBP, were detected in 56.6% of SBP cases, consistent with a study conducted by Navasa and colleagues, who reported that, despite using good culture techniques, cultures are still negative in ∼30–50% of patients with an increased AF-PMN cell count. Culture negativity was explained by low concentrations of bacteria in AF [2]. Furthermore, AF culture is insensitive and leads to delay in diagnosis for several days. A delay in antibiotic therapy entails high mortality [6].

AF total leukocytic and PMN cell count were significantly higher in the SBP group than in the non-SBP group. Jansen stated that, although ascitic total white blood cell (WBC) count increases in SBP cases, it has low specificity because a large proportion of patients with sterile ascites have increased WBC counts. Moreover, diuretic therapy has been shown to increase the total WBC count but does not alter the PMN count [3]. In clinical practice, the diagnosis is based on a PMN cell count that must be greater than or equal to 250 cell/mm 3 in AF in the absence of an intra-abdominal cause of infection [7]. Total leukocytic and PMN cell counts in AF, however, are not always readily available [4]. The count of AF PMNs is operator dependent, and lysis of PMN cells during transport to the laboratory may lead to false-negative results [6].

On the basis of these limitations, we tried to study AF lactoferrin as a surrogate and alternative marker in early diagnosis of SBP. Our study revealed a highly statistical significant increase in SBP cases when compared with the non-SBP group. At a cut-off level of 255, the sensitivity of the test was 100% and the specificity was 88.9%. The results of this study were close to that reported by Parsi et al. [6] who stated that the sensitivity and specificity of the test were 95.5% with the cut-off level 242 ng/ml. However, it is important to underline that AF lactoferrin detection is a qualitative method and needs, therefore, to be further confirmed by slandered cytology of the AF. This test could merely be a screening modality providing only a presumptive diagnosis, but it does not have the potential to replace the manual counting method. Moreover, a qualitative test does not have applicability in the clinical management of SBP, as a reduction of more than 25% of the initial PMN cell count value is, according to the current guidelines, the criterion to establish the efficacy of the ongoing antibiotic treatment [5].


  Conclusion Top


AF lactoferrin at a cut-off level greater than 255 ng/ml can be used as an initial, sensitive (100%), and specific (88.9%) screening test for the detection of SBP in cirrhotic patients with ascites. Nevertheless, further confirmatory studies including evaluation of post-treatment AF lactoferrin are needed.

Financial support and sponsorship

Nil.

Conflicts of interest

There are no conflicts of interest.[15]

 
  References Top

1.
Friedman SL. Alchoholic liver disease, cirrhosis and its major sequele. In: Goldman L, Bennett JC editors. Cecil text book of medicine. 23rd ed. Philadelphia, PA: WB Sounders Company; 2008: 804–812.  Back to cited text no. 1
    
2.
Caldwell SH, Battle EH. Ascites and spontaneous bacterial peritonitis. In: Schiff ER, Sorrell MF, Maddrey WG editors. Schiff's disease of the liver. 10th ed. Philadelphia, PA: Lippincott Williams and Wilkins, Wolters Kluwer Company; 2007: 503–544.  Back to cited text no. 2
    
3.
Garcia-Tsao G. Spontaneous bacterial peritonitis. Gastroenterol Clin North Am 1992; 21:257–275.  Back to cited text no. 3
    
4.
Koulaouzidis A, Bhat S, Karagiannidis A, Tan WC, Linaker BD. Spontanous bacterial peritonitis. Postgrad Med J 2007; 83:379–383.  Back to cited text no. 4
    
5.
Nousbaum JB, Cadranel JF, Nahon P, Khac EN, Moreau R, Thévenot T, et al. Diagnostic accuracy of the Multistix 8 SG reagent strip in diagnosis of spontaneous bacterial peritonitis. Hepatology 2007; 45:1275–1281.  Back to cited text no. 5
    
6.
Parsi MA, Saadeh SN, Zein NN, Davis GL, Lopez R, Boone J, et al. Ascitic fluid lactoferrin for diagnosis of spontaneous bacterial peritonitis. Gastroenterology 2008; 135:803–807.  Back to cited text no. 6
    
7.
Thuluvath PJ, Morss S, Thompson R. Spontaneous bacterial peritonitis – In-hospital mortality, predictors of survival, and health care costs from 1988 to 1998. Am J Gastroenterol 2001; 96:1232–1236.  Back to cited text no. 7
    
8.
Thévenot T, Cadranel JF, Nguyen-Khac E, Tilmant L, Tiry C, Welty S, Merzoug N Diagnosis of spontaneous bacterial peritonitis in cirrhotic patients by use of two reagent strips. Eur J Gastroenterol Hepatol 2004; 16:579–583.  Back to cited text no. 8
    
9.
Guarner C, Runyon BA. Spontaneous bacterial peritonitis: pathogenesis, diagnosis and management. Gastroenterology 1995; 3:311–328.  Back to cited text no. 9
    
10.
Bellamy W, Takase M, Wakabayashi H, Kawase K, Tomita M Antibacterial spectrum of lactoferricin B, a potent bactericidal peptide derived from the N-terminal region of bovine lactoferrin. J Appl Bacteriol 1992; 73:472–479.  Back to cited text no. 10
    
11.
Llovet JM, Moitinho E, Sala M, Bataller R, Rodríguez-Iglesias P, Castells A, et al. Prevalence and prognostic value of hepatocellular carcinoma in cirrhotic patients presenting with spontaneous bacterial peritonitis. J Hepatol 2000; 33:423–429.  Back to cited text no. 11
    
12.
Navasa M, Follo A, Llovet JM, Clemente G, Vargas V, Rimola A, et al. Randomized, comparative study of oral ofloxacin versus intravenous cefotaxime in spontaneous bacterial peritonitis. Gastroenterology 1996; 111:1011–1017.  Back to cited text no. 12
    
13.
Jansen PL. Spontaneous bacterial peritonitis. Detection, treatment and prophylaxis in patients with liver cirrhosis. Neth J Med 1997; 51:123–128.  Back to cited text no. 13
    
14.
Runyon BA. Strips and tubes: refining the diagnosis of spontaneous bacterial peritonitis. Hepatology 2003; 37:745–747.  Back to cited text no. 14
    
15.
Riggio O, Angeloni S. Ascitic fluid analysis for diagnosis and monitoring of spontaneous bacterial peritonitis. World J Gastroenterol 2009; 15:3845–3850.  Back to cited text no. 15
    


    Figures

  [Figure 1]
 
 
    Tables

  [Table 1], [Table 2], [Table 3]



 

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