Listeria monocytogenes in Dairy Products of the Middle East Region: A Systematic Review, Meta-Analysis, and Meta-Regression Study.

Background: The contamination of food products by Listeria monocytogenes as a pathogen bacterium, threatening public health and raised a global concern for a long time. Dairy and meat products and ready-to-eat foods are recognized as the most common carriers for L. monocytogenes.
Methods: The related reports of the prevalence of L. monocytogenes in dairy products in Middle East countries from 2009 to 2020 were screened through some of the international databases such as Science Direct, Web of Science, Scopus, PubMed, and Google Scholar. While a random effect model was applied to estimate pooled or overall prevalence, 95% confidence intervals (95%CI) were used.
Results: Results showed severe heterogeneity (84.2%) in studies and estimated the overall prevalence of L. monocytogenes dairy food products from the Middle East region of 3.5% (CI: 2.2-5). The highest and lowest prevalence was associated with Jordan (17.6% CI: 9.8-26.9) and Iraq (1.6% CI: 0.3-3.7), respectively. Based on the type of product, the highest and lowest prevalence of L. monocytogenes was recognized for raw cow milk (5.8% CI: 2.7-9.7) and pasteurized cow milk (1.1% CI: 0-8), respectively.
Conclusion: There is no justification for severe heterogeneity (I2) of subgroups as prevalence is heterogenic innately, but Jordan and row cow milk subgroups were found to have a considerable effect on overall pooled prevalence. Thus, they were the reason for prevalence changes.

Introduction

Listeria is a genus from the Listeriaceae family that includes more than 20 species categorized into three main clades (1). It is a facultatively anaerobic, gram-positive, non-sporulating, and psychrotrophic bacterium (2). Some Listeria species are recognized as pathogenic such as L. monocytogenes, known as pathogens for both humans and animals, and L. ivanovii is considered pathogenic for animals (3). L. monocytogenes are highly important in terms of public health and grow easily at temperatures ranging from −0.4 °C to 45°C and tolerate osmotic stress up to 14% salt concentration. It also can undergo a broad zone of pH 4.0 to 9.6 (optimum pH 6–8). The water activity of 0.90 is distinguished as the lowest level of water activity that L. monocytogenes can survive (4, 5).

Listeriosis resulted from the consumption of contaminated food products by L. monocytogenes such as cheese, non-pasteurized milk, unwashed raw vegetables, and undercooked meat, and it is identified as one of the primary foodborne diseases for humans (6–10). The contamination of milk by L. monocytogenes is attributed to both infected and healthy animals besides poor hygiene conditions or practices (11, 12). Therefore, among different foods, milk, mainly raw milk, and other kinds of dairy products are particular sources for L. monocytogenes.

L. monocytogenes threaten human health by causing listeriosis. At the same time, diarrhea, vomiting, fever, headache, gastroenteritis, and myalgia are mild symptoms. Moreover, septicemia and meningitis that commonly resulted in stillbirth and abortion in pregnant women are the common invasive symptoms (8, 10, 13). According to the Center for Diseases Control (CDC), 467 listeriosis cases and 83 deaths were reported between 2011 and 2017 in the different states of the USA (14), while the number of confirmed cases of listeriosis reported by EU/EEA countries in 2016 was 2555 (15). Available data about the listeriosis epidemiology in the Middle East, especially Iran, are rare. Because in the Iranian health system, listeriosis cases are not reported precisely (16).

Studies for investigating the prevalence of Listeria spp. have been conducted based on different groups of foods, including meat, seafood, milk, dairy products, ready-to-eat foods, and eggs (4, 17). Contamination is different in various regions due to differences in livestock procedures conditions, storage and hygiene conditions, and technological practices (17). Several studies were carried out on the prevalence of L. monocytogenes. These studies do not have a specific pattern, and researchers have been reported inconsistencies in different regions like Iran, Egypt, and Italy. For instance, an Iranian research group represented 37% of raw milk samples contaminated with L. monocytogenes (18). However, some other researchers (19–21) described no contamination for L. monocytogenes in raw milk. Moreover, L. monocytogenes was detected in fifty percent of cheese samples (22), while other authors (20, 23) could not find L. monocytogenes in Iranian cheese samples. Besides, Kabuki et al isolated L. monocytogenes in 6.3% of Iranian cheese samples (24). Inconsistent statistics in L. monocytogenes prevalence in dairy products are also reported in other countries. For instance, in Egypt, L. monocytogenes range from 3.3 to 6.6% in milk products (25), and researchers in other countries like Syria reported that Listeria spp. exists in 10.96% of raw milk samples (26), or L. monocytogenes was positive in 4.8% of cheese samples in Turkey (27). Dairy product contamination with L. monocytogenes has been reported for European countries as well. For example, contamination of 2.2% was reported for Italian raw milk with L. monocytogenes (28). Furthermore, meta-analysis research conducted in Europe using EFSA reports and prior studies about the prevalence, contamination of L. monocytogenes in different cheese types showed the prevalence from 0.8 to 11.8% in European countries (29).

Although many studies have estimated the prevalence value of Listeria spp. in different food samples, no systematic review or meta-analysis regarding L. monocytogenes contamination of milk products in the Asian or the Middle East countries was reported cohesively. Hence, summarizing research from different databases that studied L. monocytogenes contamination in the Middle East countries in dairy products and considering the risk of bias assessment was the objective of this work.

Methods

Search scheme

According to the Cochrane protocol (PRISMA guideline), this systematic review and meta-analysis study was carried out (30). In the first step, Science Direct, Web of Science, Scopus, PubMed, and Google Scholar databases were used to retrieve published original studies in the Middle East countries between 2009 and 2020. The following search keywords and MeSH terms were used: (Cheese OR Cream OR Kashk OR Milk OR “Dairy Products” OR “Ice cream” OR Butter OR Yoghurt OR Ghee OR Buttermilk OR Kefir OR Koumiss OR Whey) AND (“Listeria monocytogenes” OR “L.monocytogenes” ) AND (“The Middle East” OR Afghanistan OR Bahrain OR Iran OR Iraq OR Israel OR Jordan OR Kuwait OR Lebanon OR Oman OR Qatar OR Saudi Arabia OR Syria OR Turkey OR United Arab Emirates OR Yemen) AND (Prevalence OR Outbreak OR Incident OR Contamination). Searching in the mentioned databases and screening was conducted. The title, abstract, and full text of retrieved articles were screened for eligibility. Moreover, to cover almost all papers, the references list of reviewed articles was considered.

Inclusion/exclusion criteria

The articles’ selection was based on our research criteria by two reviewers separately after screening the databases with the same keywords and procedures. In this step, each author was responsible for reviewing the papers in all parts (titles, abstracts, and full texts) to select the articles having the inclusion criteria; any differences were supposed to be resolved by consensus. Articles included in this study based on inclusion criteria (Eligibility criteria) including only dairy product samples, researches were done and reported in the middle east countries, available full-text in the English language; cross-sectional descriptive studies; L. monocytogene detection done with high accuracy technique (culture and PCR techniques); and prevalence data was presented clearly in the results of papers. Moreover, any other food samples like meat products or ready-to-eat-foods, reported prevalence of L. monocytogene in other countries, genetic, ecological, animal studies, case reports, thesis, books, non-English articles, and review articles were excluded (Fig. 1).

Fig. 1:

The PRISMA flowchart diagram of the study.

Data extraction and risk of bias assessment

Required data from selected articles including first author, publication year, method of L. monocytogenes detecting, type of dairy products, country, study area (province), sample size, and positive samples were extracted by two authors separately. Additionally, assessing the risk of bias of included studies was judged independently by the first author based on the JBI checklist with a minor modification (31).

Meta-analysis of data

In this meta-analysis, we applied random effect models for estimating pooled or overall prevalence and 95% confidence intervals (95%CI). The positive samples (pi) divided into the total sample (ni) indicates the prevalence of L. monocytogene in the dairy products (I=pi /ni). Overall prevalence was estimated using a Freeman-Tukey double arcsine transformation. Estimation of prevalence L. monocytogene was performed using the Meta-prop command (32). I statistics and Cochran’s Q Heterogeneity were used for exploring heterogeneity among studies. I2 is a variance variable (prevalence) divided by Q value. I ranges between 0–100 percent, and I >70% were considered heterogeneous (33, 34). A Forest plot was used to show L. monocytogene prevalence. In this study, subgroup analysis and meta-regression were performed to explore heterogeneity sources (35). Publication bias was not tested in this study because its aim is not to estimate the association between exposures and outcomes (35). Meta-analysis was done by STATA 14.0 (2015; STATA 14.0 Statistical Software, College Station, TX, USA). Statistical significance was selected less than 0.05 (P<0.05).

Results and discussion

This systematic review and meta-analysis study was carried out on 76 key measures calculated from 32 papers related to dairy products. Articles included in this systematic review and meta-analysis from each country (the Middle East) were summarized in Table 1. Iran (n=13) and Turkey (n=10) had the most considerable number of articles on the prevalence of L. monocytogenes. However, some other countries like Kuwait, Bahrain, and Yemen found no related publication in this field. In this research, dairy products were divided into eight main subgroups, in which most studies included cheese and raw cow milk, as well as the overall prevalence for the presence of L. monocytogenes in dairy products, is illustrated in Fig. 2, which was 3.5 % (CI: 2.2%–5.0%).

Table 1:

Summary of the studies reporting the Prevalence of Listeria monocytogenes in dairy products in the Middle East countries

Id	Assessment technique	Food groups	Country	Region	Year	Sample size	Positive samples	Quality assessment	Ref.
1	PCR	Cheese	Iran	Tehran	2012–2015	70	5	7	(46)
		Cream				20	2	7
		Kashk				17	2	6
2	PCR	Raw cow milk	Turkey	Samsun	2011–2012	100	5	7	(47)
		Cheese				70	9	7
		Ice cream				20	0	7
		Cream				20	0	7
3	PCR	Raw cow milk	Iran	-	2013–2015	37	1	7	(48)
		Cheese		-		130	4	7
4	PCR	Raw cow milk	Turkey	Southern Marmara	2009–2010	42	0	7	(49)
		Cheese				140	0	7
		Cream				12	0	6
		Yogurt				2	0	6
5	PCR	Raw cow milk	Iran	Tehran	2008–2010	240	13	7	(41)
		Raw sheep Milk				206	5	7
6	Culture	Cheese	Turkey	Van otlu	2011	120	34	7	(50)
7	Culture	Raw cow milk	Turkey	Sakarya	2008–2010	8	0	8	(51)
		Cheese				8	0	8
8	Culture	Raw cow milk	Turke	Aydin	2013	20	5	7	(52)
		Pasteurized cow milk	y			20	1	7
		Yogurt				40	2	7
		Cheese				50	5	7
		Cream				10	1	7
9	Culture	Raw cow milk	Iran	Tabriz	2014	18	9	7	(22)
10	Culture	Raw cow milk	Jordan	Karak	2011	20	9	7	(36)
		Cheese				20	7	7
		Yogurt				20	8	7
11	PCR and	Raw cow milk	Iran	Fars and Khuzestan	2010–2011	156	3	8	(10)
	Culture	Raw sheep Milk				104	2	8
12	PCR and Culture	Raw cow milk	Egypt	Sharkia Governorate	2015–2016	100	25	8	(22)
13	PCR	Kashk	Lebanon	Bekaa Valley	2009	83	6	8	(43)
		Cheese				81	17	8
14	Culture	Raw cow milk	Iran	Sari	2013	100	0	8	(19)
		Pasteurized cow milk				100	0	8
15	PCR	Pasteurized cow milk	Iran	-	2011	596	61	8	(54)
16	PCR	Raw cow milk	Egypt	-	2013	100	1	5	(55)
17	PCR	Cheese	Turkey	Erzurum	2015	104	17	6	(56)
	Culture	Pasteurized cow milk	Turkey	-	2012	10	0	7	(57)
18		Cheese		-		60	1	7
		Ice cream		-		20	0	7
		Cream		-		10	0	7
19	PCR	Raw cow milk	Iran	-	2015	60	0	7	(58)
20	Culture	Cheese	Jordan	Amman	2017	150	8	7	(59)
21	PCR and	Raw cow milk	Iran	Yazd	2016	140	11	8	(8)
	Culture	Cheese				220	10	8
		Cream				100	1	8
		Ice cream				85	0	8
22	Culture	Ice cream	Iran	Kermans	2013	67	0	8	(21)
		Pasteurized cow milk		hah		59	0	8
		Cheese				59	0	8
23	PCR and Culture	Raw cow milk	Iran	Kerman	2011	100	5	6	(60)
24	PCR and	Raw cow milk	Iran	Isfahan	2007–2009	138	1	8	(10)
	Culture	Raw sheep milk				122	6	8
		Cheese				90	9	8
		Ice cream				136	2	8
		Yogurt				130	0	8
		Cream				40	1	8
		Kashk				71	0	8
25	Culture	Yogurt	Turkey	Balikesir	2010–2011	100	3	7	(61)
		Cheese				100	5	7
26	PCR	Cheese	Egypt	El Giza Governorate	2013–2014	50	0	8	(62)
		Raw cow milk				50	4	8
27	Culture	Raw cow milk	Iran	Nooraba	2010	120	3	7	(63)
		Cheese		d		60	3	7
		Yogurt				180	0	7
28	PCR	Cheese	Jordan		2011	350	39	7	(64)
29	Culture	Pasteurized cow	Turkey	Bursa	2012	10	0	6	(38)
		milk
		Cheese				60	1	7
		Ice cream				20	0	7
		Cream				10	0	6
30	Culture	Row cow milk	Jordan	-	2016	305	32	8	(65)
31	PCR	Cheese	Turkey	Bolu	2016	40	0	8	(66)
		Ice cream				27	0	8
32	Culture	Cheese	Turkey	Hatay	2019	20	0	8	(67)
33	PCR	Cheese	Iraq	Dehok	2016–2017	121	1	8	(68)
		Raw sheep milk				118	3	8

Fig. 2:

The forest plot of prevalence of Listeria monocytogenes in dairy products based on food type/year of publication/country

Ice cream had the lowest mean prevalence of 0.0% (CI: 0.0–1.1%), followed by cream 0.6% (CI: 0.0–2.9%), pasteurized cow milk 1.1% (CI: 0.0.–8.0%), yogurt 1.6% (CI: 0.0–10.4%), raw ship milk 2.8% (CI: 1.5–4.4%), Kashk 3.8% (CI: 0.0–14.8%), cheese 5.5% (CI: 2.8–8.9%) and raw cow milk had the highest L. monocytogenes prevalence of 5.8% (CI: 2.7–9.7%). On the other hand, the prevalence rank order in terms of country showed the following order: Iraq 1.6% (CI: 0.3–3.7%), Turkey 2.2% (CI: 0.3–5.1%), Iran 2.5% (CI: 1.2–4%), Egypt 5.8% (CI: 0–20.6%), Lebanon 13.3% (CI: 8.4–19%) and Jordon 17.6% (CI: 9.8–26.9%).

The highest level of L. monocytogenes contamination in cheese (Fig. 3) was observed in Jordan with 35% contamination (CI: 15.4–59.2%) (36).

Fig. 3:

The forest plot of prevalence of Listeria monocytogenes in dairy products based on country/year of publication/food type

Moreover, the highest and lowest L. monocytogenes incidence in raw ship milk was observed in two studies carried out (10, 37) in Iran, which found values of 4.9 and 1.9%. The main reason cheese faced a high prevalence of L. monocytogenes is probably related to raw milk origin and survival ability of L. monocytogenes in high salt concentration and resistance to different pH (38). L. monocytegenes can survive in some cheeses like white cheese after several weeks, dangerous for public health. The estimated mean prevalence of L. monocytogenes in cheese products in the Middle East (5.5%) was more than results from European reports (2.3%) (3). The only study on L. monocytogenes contamination in Iraq reported a mean prevalence of 1.6% (CI: 0.3%–3.7%) in cheese and raw ship milk (39). Milk contamination of L. monocytogenes is not specific to raw cow milk, but also pasteurized cow milk could carry L. monocytogenes in case of inadequate pasteurization or post-pasteurization contamination. Furthermore, the high prevalence of L. monocytogenes in raw milk could be associated with animal infection or contamination of livestock feed, and contamination of raw milk during breastfeeding and milk storage could be another reason (40, 41). Similarly, in a study conducted by the EU’s Rapid Alert System for Food and Feedstuff between 2004 and 2009, 97 dairy products were positive for Listeria and 34 of them were pasteurized, and 22 samples were raw milk products (42). Moreover, the highest occurrence rate of L. monocytogenes in yogurt was found in Jordan (36), that 40% of samples were contaminated with L. monocytogenes. Moreover, in the only study on L. monocytogenes contamination in Lebanon, they reported a mean prevalence of 13.3% (CI: 8.4–19%) in cheese and Kashk (43). According to the results of this meta-analysis, contamination of cream and ice cream by L. monocytogenes was low.

Generally, occurrence evaluation of L. monocytogenes is necessary for dairy products as they are vehicles for this pathogen transmission that led to the highest death rate among infectious diseases (44). Preventing foods, especially dairy products, from being contaminated by L. monocytogenes is highly recommended. Additionally, adequate pasteurizing, forage quality, housing facilities, and milking hygienic conditions can affect milk’s microbial quality (45).

The meta-regression test investigates potential contributing parameters that affect study heterogeneity. This test was used to examine the year of publication and the quality score for each included study in our study. The year of publication’s meta-regression plot versus L. monocytogenes prevalence in dairy products showed no significant relationship (C=0.0006; P=0.88). As shown in Fig. 4, regarding the effect sizes of the research studies; there is a tendency to decrease in L. monocytogenes prevalence over the years between 2010 and 2020, which seems that during these years, various factors, including hygienic-sanitary quality of milk, storage conditions, and pasteurization method has improved as well as post-pasteurization contamination reduced. Generally, hygienic procedures in the Middle East have been introduced to some extent for quality improvement over the last decade. In this study, the quality assessment of research studies was conducted. We evaluated the methodological quality of the 32 research reports, using nine quality criteria relating to the prevalence studies. The items were determined to consist of sampling method, sample size adequacy, subject description in detail, data coverage analysis, applying valid method, response rate adequacy, using the standard method, and using appropriate statistical analysis. A meta-regression analysis was used to find the relationship between L. monocytogenes occurrence rate and the quality score of the included study. Figure 5 indicated that included papers’ quality scores have no considerable effects on the prevalence of L. monocytogenes in dairy products.

Fig. 4:

The relationship between the prevalence Listeria monocytogenes with year of study

Fig. 5:

The relationship between the prevalence of Listeria monocytogenes with quality score for included study

There are some limitations to this study. Although our search was comprehensive, there were limited studies in some countries or finding no studies on L. monocytogenes contamination. Moreover, no study was found about Listeria contamination in some samples, such as Kefir or Komis. Another limitation was that some of the available studies had limited sample sizes or a lack of data on Listeria contamination, or no full-text access. Despite these limitations, the authors tried to cover all the studies done in the Middle East region and showed a general view of the prevalence of L. monocytogenes. Moreover, researchers suggest that researchers aim to carry out secondary review articles (systematic review and meta-analysis study) to limit and make narrower the type of reviewed food samples and possibly widen reviewed regions. It helps to draw a much better image of the prevalence of L. monocytogenes in the world and finds critical and threatening points in L. monocytogenes contaminations affecting public health. Similarly, risk assessment studies for this pathogen in different dairy products in different regions and countries and even worldwide are suggested for future original research.

Conclusion

The prevalence of L. monocytogenes in dairy products in the Middle East based on defined subgroups was systematically reviewed and meta-analyzed. The order of contamination by L. monocytogenes among the investigated dairy products based on prevalence was raw cow milk> cheese> Kashk> raw ship milk> yogurt > pasteurized cow milk > cream > ice cream, demonstrating the required further attention in the production process of some products such as cheese and Kashk. The high prevalence of L. monocytogenes in some dairy products is probably related to the unhealthy production method and lack of adequate pasteurization temperature in these products, as well as environmental contamination by animal wastes, which can be a reason for this contamination. Consumption of raw milk and its products, especially cheese produced with insufficient heat treatment and the lack of appropriate control measures, might cause serious health problems.

Ethical considerations

Ethical issues (Including plagiarism, informed consent, misconduct, data fabrication and/or falsification, double publication and/or submission, redundancy, etc.) have been completely observed by the authors.